Microorganism

Appl Environ Microbiol, 1997 Mar, 63(3), 1131 - 8
Characterization of the genes encoding the three-component membrane-bound alcohol dehydrogenase from Gluconobacter suboxydans and their expression in Acetobacter pasteurianus; Kondo K et al.; The three-component membrane-bound alcohol dehydrogenase (ADH) of Gluconobacter suboxydans IFO12528 was purified, and the NH2-terminal amino acid sequence of each subunit was determined . On the basis of the amino acid sequences, the genes adhA, encoding the 72-kDa dehydrogenase, adhB, encoding the 44-kDa cytochrome c-553 (a CO-binding cytochrome c), and adhS, encoding a 15-kDa protein, were cloned and the amino acid sequences of their products were deduced from the nucleotide sequences . The dehydrogenase and cytochrome genes were clustered with the same transcription polarity, as is the case in species of Acetobacter, another genus of acetic acid bacteria . These AdhA and AdhB subunits showed similarity in amino acid sequence to those from Acetobacter spp., whereas AdhS showed no similarity to the corresponding subunit of the ADH complex of Acetobacter pasteurianus . Consistent with this, adhS of G . suboxydans could not complement a defect in the corresponding subunit of A . pasteurianus . When the adhA-adhB gene cluster of G . suboxydans was expressed in an ADH-deficient mutant of A . pasteurianus, the transformant showed distinct ADH activity . The ADH complex was purified to near homogeneity and consisted of two subunits, the dehydrogenase and the cytochrome c subunits derived from G . suboxydans, without any other subunit . These data suggested that AdhS, the smallest subunit of ADH, from G . suboxydans is not essential for ADH activity in A . pasteurianus, in contrast to the essential role of A . pasteurianus AdhS, which is required for correct assembly of the dehydrogenase and cytochrome c subunits on the membrane.

J Gen Virol, 1997 Mar, 78 ( Pt 3), 535 - 42
The beet yellows closterovirus p65 homologue of HSP70 chaperones has ATPase activity associated with its conserved N-terminal domain but does not interact with unfolded protein chains; Agranovsky AA et al.; The positive-strand RNA genome of beet yellows closterovirus (BYV) encodes a 65 kDa protein (p65) related to the HSP70 family of cell chaperones . The full-sized BYV p65, and N- and C-terminal fragments, with (His)6 tails, were overexpressed in bacteria and purified by metal-chelate chromatography . Using a polyclonal antiserum raised against the C-terminal fragment of p65, evidence was obtained for expression of the viral protein in planta . Purified recombinant p65 and its N-terminal 40 kDa fragment exhibited Mg2+-dependent ATPase activity in vitro . However, unlike its cellular HSP70 homologues, p65 was unable to bind to denatured protein and its ATPase activity was not stimulated by synthetic peptides which are known to stimulate HSP70 ATPases . Hence, the BYV p65, although being a chaperone-type ATPase, may have a distinct substrate specificity and function in BYV-infected cells.

J Bacteriol, 1997 Mar, 179(5), 1628 - 35
Relationship of Treponema denticola periplasmic flagella to irregular cell morphology; Ruby JD et al.; Treponema denticola is an anaerobic, motile, oral spirochete associated with periodontal disease . We found that the periplasmic flagella (PFs), which are located between the outer membrane sheath and cell cylinder, influence its morphology in a unique manner . In addition, the protein composition of the PFs was found to be quite complex and similar to those of other spirochetes . Dark-field microscopy revealed that most wild-type cells had an irregular twisted morphology, with both planar and helical regions, and a minority of cells had a regular right-handed helical shape . High-voltage electron microscopy indicated that the PFs, especially in those regions of the cell which were planar, wrapped around the cell body axis in a right-handed sense . In those regions of the cell which were helical or irregular, the PFs tended to lie along the cell axis . The PFs caused the cell to form the irregular shape, as two nonmotile, PF-deficient mutants (JR1 and HL51) were no longer irregular but were right-handed helices . JR1 was isolated as a spontaneously occurring nonmotile mutant, and HL51 was isolated as a site-directed mutant in the flagellar hook gene flgE . Consistent with these results is the finding that wild-type cells with their outer membrane sheath removed were also right-handed helices similar in shape to JR1 and HL51 . Purified PFs were analyzed by two-dimensional gel electrophoresis, and several protein species were identified . Western blot analysis using antisera to Treponema pallidum PF proteins along with N-terminal amino acid sequence analysis indicated T . denticola PFs are composed of one class A sheath protein of 38 kDa (FlaA) and three class B proteins of 35 kDa (FlaB1 and FlaB2) and one of 34 kDa (FlaB3) . The N-terminal amino acid sequences of the FlaA and FlaB proteins of T . denticola were most similar to those of T . pallidum and Treponema phagedenis . Because these proteins were present in markedly reduced amounts or were absent in HL51, PF synthesis is likely to be regulated in a hierarchy similar to that found for flagellar . synthesis in other bacteria.

J Clin Microbiol, 1997 Mar, 35(3), 691 - 6
Aseptic meningitis and encephalitis: the role of PCR in the diagnostic laboratory; Read SJ et al.; In this study we have devised a simple and robust PCR strategy to detect a wide range of viruses, bacteria, and parasites, all of which are capable of causing aseptic meningitis and encephalitis . The techniques developed have been used in a routine diagnostic virology laboratory to test prospectively 2,233 cerebrospinal fluid specimens . A virus was detected in 147 specimens of cerebrospinal fluid from 143 patients . Four sets of primers were sufficient to detect the virus in 135 (94%) of the PCR-positive patients . We conclude that with appropriate primers, PCR can be systematically and economically applied to test for a range of organisms in a routine diagnostic laboratory . In our opinion, PCR will soon become the "gold standard" test for viral infections of the central nervous system.

Cytometry, 1997 Mar 1, 27(3), 269 - 74
Flow cytometric determination of endocytosis of viable labelled Legionella pneumophila by Acanthamoeba palestinensis; Harf C et al.; Endocytosis of fluorescently labelled cells of Legionella pneumophila (L . pneumophila) by free-living Acanthamoeba palestinensis (A . palestinensis) has been studied using flow cytometry . L . pneumophila cells were labelled with CM-DiI, a lipophilic fluorescent probe under conditions that did not modify viability . Coculturing the bacteria with amoebae was accompanied by rapid endocytosis; after 5 min, 90% of the amoebae had internalized bacteria . This percentage remained unchanged during further coculture, but the number of bacteria ingested per amoeba increased . Moreover, the number of ingested bacteria was found to be dependent on the size of the amoeba . The validity of the internalization analyzed by flow cytometry was confirmed by observation using epifluorescence and phase contrast microscopy . CM-DiI labelling associated with flow cytometry provides a very valuable technique for the determination of bacteria endocytosis by free-living amoeba.

Infect Immun, 1997 Mar, 65(3), 1088 - 94
The Chlamydia trachomatis parasitophorous vacuolar membrane is not passively permeable to low-molecular-weight compounds; Heinzen RA et al.; Chlamydia trachomatis is an obligately intracellular bacterial parasite of eucaryotic cells that undergoes a biphasic life cycle within a parasitophorous vacuole (PV) called an inclusion . The parasitophorous vacuolar membrane (PVM) constitutes a barrier between the replicating bacteria and the nutrient-rich environment of the host cytoplasm . To determine whether the chlamydial PVM contains pores that allow passive diffusion of metabolites between the host cytoplasm and the PV, fluorescent tracer molecules were introduced directly into the cytoplasm of infected cells by transfection or microinjection . Fluorescence microscopy and laser scanning confocal microscopy were subsequently employed to determine whether equilibration of the fluorescent tracers between the cytoplasm and the PV occurred . No movement of tracer molecules as small as 520 Da from the cytoplasm to the PV was observed . These data suggest that the chlamydial PV is not passively permeable to small molecules through open channels in the PVM.

J Immunol, 1997 Mar 1, 158(5), 2218 - 27
A three-domain T cell receptor is biologically active and specifically stains cell surface MHC/peptide complexes; Plaksin D et al.; We have expressed in bacteria a single-chain T cell receptor (scTCR) with specificity for an HIV gp120-derived peptide bound to the murine MHC-I molecule, H-2Dd . This scTCR consists of V alpha covalently linked to the VbetaCbeta domains that was solubilized, refolded, and purified in high yield . Specific binding of the scTCR to MHC/peptide complexes was demonstrated by surface plasmon resonance, with a Kd of 2 to 8 x 10(-6) M . This scTCR specifically inhibited T cell activation, and stained cell surface MHC/peptide complexes as measured by cytofluorimetry . The preservation of binding specificity by such a three-domain scTCR suggests that this structure is sufficient for specific MHC/peptide recognition and that this strategy will be of general use as applied to other TCR.

Sci Total Environ, 1997 Feb 24, 194-195, 247 - 62
Biology of the Humber rivers; Whitton BA et al.; An overview of the literature is presented on the biology of the rivers entering the Humber, eastern England, together with some of their tributaries . Particular emphasis is given to dynamic aspects, including transport and movement within rivers, movement between rivers, processes within rivers and long-term changes.

Presse Med, 1997 Feb 22, 26(5), 248 - 54
{Search for a HIV vaccine}; Sicard D et al.; A DIFFICULT SITUATION FOR A VACCINE: The human immunodeficiency virus has an exceptional capacity to mutate and macrophage reservoirs where it is harbored after penetration are highly inaccessible to antibodies . Use of a live vaccine would increase the risk of recurrent virulence and integration into the genome . UNKNOWN IMMUNOLOGY: Induction of cytotoxic cells appears to be essential, but investigations into this type of response are not well standardized and not easily quantifiable . Neutralizing antibodies would have a protective effect, but facilitating antibodies would have the opposite effect . SEVERAL POSSIBILITIES UNDER STUDY: These vaccine use live attenuated viruses with a more or less deleted genome, recombinant live viruses constructed from other viruses or bacteria, pseudo-particles, or recombinant proteins as well as vaccines synthetized from peptides or lipopeptides . An association of different vaccines would appear to be the best solution as live vectors usually induce cytotoxic cells while proteins or peptides induce production of neutralizing antibodies . RESPONSE TO VACCINES IS USUALLY WEAK: Both in animal models and in human volunteers, immune responses obtained to date have been quite variable and of short duration . Canarypox type recombinant vaccines followed by recombinant protein vaccines appear to give the best results at the present time . SEVERAL PROBLEMS: Anti-HIV vaccination protocols raise major ethical problems for the participating volunteers (ELISA test becomes positive, lack personal protection) and for the population involved in phase II/III trials . In addition, the virus rapidly penetrates via the mucosa without yielding sufficient mucosal immune response.

J Theor Biol, 1997 Feb 21, 184(4), 451 - 69
A study of oligonucleotide occurrence distributions in DNA coding segments; Castrignano T et al.; In this paper we present a general strategy designed to study the occurrence frequency distributions of oligonucleotides in DNA coding segments and to deal with the problem of detecting possible patterns of genomic compositional inhomogeneities and disuniformities . Identifying specific tendencies or peculiar deviations in the distributions of the effective occurrence frequencies of oligonucleotides, with respect to what can be a priori expected, is of the greatest importance in biology . Differences between expected and actual distributions may in fact suggest or confirm the existence of specific biological mechanisms related to them . Similarly, a marked deviation in the occurrence frequency of an oligonucleotide may suggest that it belongs to the class of so-called "DNA signal (target) sequences" . The approach we have elaborated is innovative in various aspects . Firstly, the analysis of the genomic data is carried out in the light of the observation that the distribution of the four nucleotides along the coding regions of the genoma is biased by the existence of a well-defined "reading frame" . Secondly, the "experimental" numbers found by counting the occurrences of the various oligonucleotide sequences are appropriately corrected for the many kinds of mistakes and redundancies present in the available genetic Data Bases . A methodologically significant further improvement of our approach over the existing searching strategies is represented by the fact that, in order to decide whether or not the (corrected) "experimental" value of the occurrence frequency of a given oligonucleotide is within statistical expectations, a measure of the strength of the selective pressure, having acted on it in the course of the evolution, is assigned to the sequence, in a way that takes into account both the value of the "experimental" occurrence frequency of the sequence and the magnitude of the probability that this number might be the result of statistical fluctuations . If the strength of the selective pressure evaluated in this fashion turns out to be sufficiently large, the corresponding sequence will be considered to have an occurrence frequency beyond expectations and, hence, to be statistically and biologically interesting.

J Mol Biol, 1997 Feb 21, 266(2), 306 - 16
Mutant RecA proteins which form hexamer-sized oligomers; Logan KM et al.; We have analyzed the oligomeric properties of a number of mutant RecA proteins containing single amino acid substitutions within one region of the subunit interface . In contrast to wild-type RecA, which forms a heterogeneous population of different-sized oligomers, we find that many of these mutant proteins exist in a more homogeneous oligomeric form, which approximates to the size of a RecA hexamer . Some of these mutants have a significant level of activity in vivo for recombinational DNA repair and thus represent the first mutant RecA proteins identified which retain activity yet can exist in a discrete oligomeric state as free protein.

Circulation, 1997 Feb 18, 95(4), 814 - 7
Left ventricular assist device infection is associated with increased mortality but is not a contraindication to transplantation; Herrmann M et al.; BACKGROUND: Left ventricular assist devices (LVADs) are increasingly used as a bridge to transplantation . Infection is a frequent and major complication associated with the use of these devices, however, the correlation of infection and outcome has not yet been evaluated in a prospective fashion . METHODS AND RESULTS: Twenty-five patients (24 male, 1 female) with end-stage cardiac failure and resulting organ dysfunction were included . Patients were bridged with the Novacor N100 portable LVAD (median duration of support, 55 days) and were evaluated prospectively by device surface cultures on explantation, molecular typing of isolates, and correlation of infection with survival to transplant . Twelve (48%) of 25 patients had LVAD infection as defined by recovery of multiple isolates of identical genotype from the device surface . Whereas only 5 (42%) of 12 patients with LVAD infection survived until transplantation, 11 (85%) of 13 patients without infection were successfully transplanted (P < .05) . Death of the 7 patients with proven LVAD infection was associated with multiple organ failure or other signs of acute infection . CONCLUSIONS: LVAD infection is associated with a significantly decreased survival probability . It does not preclude successful bridging but rather may pose an indication for urgent transplantation.

Biochem J, 1997 Feb 15, 322 ( Pt 1), 207 - 11
Difference in hepatic metallothionein content in Antarctic red-blooded and haemoglobinless fish: undetectable metallothionein levels in haemoglobinless fish is accompanied by accumulation of untranslated metallothionein mRNA; Scudiero R et al.; Icefish (family Channichthyidae, suborder Nothothenioidei) are a group of Antarctic fish that have evolved unique phenotypes in order to adapt to the environment in which they live . Besides the lack of haemoglobin and the drastic reduction in the number of erythrocyte-like cells, another striking feature of the icefish is that their liver is devoid of metallothionein . These cysteine-rich heavy-metal-binding proteins are usually present in large amounts in a large variety of organisms, from bacteria to mammals . Despite the failure to detect appreciable levels of metallothionein in icefish liver, a cDNA encoding metallothionein was produced from total RNA by reverse transcriptase PCR . The icefish metallothionein showed high percentage identity with metallothionein from Trematomus bernachii, a red-blooded Antarctic fish in which a normal content of hepatic metallothionein was found . Steady-state mRNA levels were assessed in fish liver by high-stringency hybridization of the metallothionein probe with total RNA . The results showed that icefish livers retain large amounts of untranslated metallothionein mRNA . The stability of the icefish transcript might be correlated with the lack of specific motifs in the untranslated 3' ends of mRNA.

Anal Biochem, 1997 Feb 15, 245(2), 115 - 22
Recombinant strategies for rapid purification of catalytic subunits of cAMP-dependent protein kinase; Hemmer W et al.; Knowledge of the crystal structure of the catalytic subunit (C) of cAMP-dependent protein kinase provided for the first time a molecular basis for probing function by site-directed mutagenesis . The purification of mutant C-subunits, however, presented new and unanticipated challenges due to instability, insolubility, and underphosphorylation of the altered proteins . To overcome these barriers, a rapid and efficient method for purifying recombinantly expressed C-subunits was developed . Purification to near homogeneity is achieved in less than 5 h . The procedure is based on colysis of bacteria that overexpress the C-subunit with bacteria that overexpress a poly-His-tagged mutant of the type II regulatory subunit H6RII (R213K) . This mutant R-subunit with an altered cAMP binding site A forms holoenzyme rapidly in bacterial extracts, and the Ka (cAMP) for the resulting holoenzyme, 27-37 microM, is nearly 50-fold increased compared to holoenzyme formed with wild-type RII . Thus, after batchwise immobilizing the holoenzyme on Ni(2+)-resin, the free C-subunit can be directly eluted batchwise with high concentrations of cAMP . The method is described for the purification of wild-type C, with yields of approximately 5 mg/liter . In addition, a mutant subunit, C{G52S}, which is defective in ATP binding and could not be isolated using previously described methods, was purified with equal efficiency.

Transplantation, 1997 Feb 15, 63(3), 407 - 14
Effects of hypoxemia on early postoperative course of liver transplantation in pediatric patients with intrapulmonary shunting; Uemoto S et al.; Nine pediatric patients (mean age, 10 years) with biliary atresia, who had hypoxemia related to intrapulmonary shunting, underwent living related liver transplantation . The effects of hypoxemia during the early postoperative period after liver transplantation on cardiopulmonary and renal function, as well as on transplanted liver, were analyzed . Based on the degree of shunt ratio calculated by technetium-99m macroaggregated albumin scintigraphy, the nine patients were included in the moderate group (shunt ratio under 40%, n=4) or the severe group (shunt ratio over 40%, n=5) . Partial pressure of arterial oxygen was maintained at normal range in the moderate group, while that in the severe group persistently had very low values (<50 mmHg), in spite of a high degree of oxygen supply . However, all patients in the severe group maintained stable cardiopulmonary vital signs, including systemic blood pressure, heart rate, respiratory rate, and cardiac index . They also demonstrated stable renal function . None of the patients died of cardiopulmonary or renal insufficiency after transplantation, but three patients died of portal vein thrombosis, sepsis, and intracranial hemorrhage (one each) . The minimal adverse effect of hypoxemia on the transplanted liver was confirmed by a rapid increase of arterial ketone body ratio, low peak values (under 200 IU/L) of aspartate aminotransferase, and a steady decrease of serum total bilirubin . Four patients encountered surgical complications, including two bile leaks from the cut liver surface, two leaks from bilioenteric anastomosis, and one intestinal perforation . Six patients suffered from bacterial infections, including four wound infections, three right subphrenic abscesses, one cholangitis, and two systemic sepses . All patients in the moderate group recovered from hypoxemia, but four of five patients in the severe group have not recovered during the follow-up period between 4 and 9 months . It was concluded that the adverse effects of hypoxemia on cardiopulmonary and renal function and transplanted liver were minimal, so that patients with severe hypoxemia could tolerate the stress of liver transplantation without special management . However, the high incidence of surgical complication and infection suggested the adverse effects of hypoxemia on wound healing and resistance to bacteria infection.

Structure, 1997 Feb 15, 5(2), 153 - 7
A good turn for DNA: the structure of integration host factor bound to DNA; Ellenberger T et al.; The crystal structure of integration host factor (IHF) complexed with DNA shows how a small heterodimeric protein can induce a big bend in DNA . IHF exerts leverage in the minor groove and wraps DNA around the body of the protein, providing another example of sequence-specific recognition of the minor groove.

J Biol Chem, 1997 Feb 14, 272(7), 4058 - 64
Molecular mechanism of polyamine stimulation of the synthesis of oligopeptide-binding protein; Igarashi K et al.; Polyamine stimulation of the synthesis of oligopeptide-binding protein (OppA) was shown to occur mainly at the level of translation by measuring OppA synthesis and its mRNA level . Several artificial oppA genes were constructed by site-directed mutagenesis . These synthesize different kinds of OppA mRNAs: mRNAs differing in the size of 5'-untranslated region; mRNAs having the Shine-Dalgarno (SD) sequence in a different position; mRNAs having different secondary structure in the region of the SD sequence; and fusion mRNAs consisting of the 5'-untranslated region of OppA mRNA and the open reading frame of beta-galactosidase . By measuring the synthesis of OppA or beta-galactosidase from these mRNAs, we found that the 171-nucleotide 5'-untranslated region and 145 nucleotides of the ORF of OppA mRNA are involved in the polyamine stimulation of OppA synthesis . When the secondary structure of the above region of OppA mRNA was analyzed by optimal computer folding, it was shown that the degree of polyamine stimulation of OppA protein synthesis was dependent on the structure of the SD sequence in addition to its position . Loose base pairing of the SD sequence with other regions of the mRNA caused strong polyamine stimulation, while intense base pairing of the SD sequence with other regions of the mRNA resulted in insignificant or weak polyamine stimulation.

Proc Natl Acad Sci U S A, 1997 Feb 4, 94(3), 802 - 7
Binding of SecB to ribosome-bound polypeptides has the same characteristics as binding to full-length, denatured proteins; Randall LL et al.; The interaction of the chaperone SecB with ribosome-bound polypeptides that are in the process of elongation has been studied using an in vitro protein synthesis system . The binding is characterized by the same properties as those demonstrated for the binding of SecB to full-length proteins that are in nonnative conformation: it is readily reversible and has no specificity for the leader peptide . In addition, it is shown that the growing polypeptide chains must achieve a critical length to bind tightly enough to allow their isolation in complex with SecB . This explains the longstanding observation that, even when export is cotranslational, it begins late in synthesis . Furthermore, the required length is approximately the same as the length that defines the binding frame within denatured, full-length proteins bound to SecB.

Am J Perinatol, 1997 Feb, 14(2), 103 - 6
A pilot study identifying type V collagenolytic activity in human amniotic fluid; Polzin WJ et al.; Amniorrhexis complicates pregnancies if it occurs in a preterm pregnancy or remote from the onset of labor in a term pregnancy . There are different collagen types (I-V) that create the extracellular matrix of the amnion . Collagenases specific to these collagen types, with the exception of type V collagen, are found in human amniotic fluid, fibroblasts, polymorphonuclear leukocytes and bacteria . Type V collagen is a major component of the amniotic basement membrane and is responsible for maintaining a barrier to bacteria and to the loss of amniotic fluid . We sought to find evidence of type V collagenolytic activity in human amniotic fluid obtained from pregnancies in different clinical states.

Endod Dent Traumatol, 1997 Feb, 13(1), 31 - 5
Experimental apexigenesis in baboons; Das S et al.; Apexification with calcium hydroxide is a routine procedure . However, some clinical reports suggest that root completion can occur by controlling the infection without use of a catalyst . The present study investigated the use of tetracycline treatment (in root canals) on root growth in immature teeth, rendered non-vital experimentally . Incisors in 3 young baboons were exposed and canals were left open . After 2 months all canals were cleaned and treated with either tetracycline or formocresol . Some canals in each group were filed . Animals were sacrificed after 6 months . Bacterial evaluations were done before placing medications, one week later and six months after that . The number of bacteria were reduced in all treatment groups . Root growth almost near completion was observed in more teeth treated with tetracycline than in the formocresol group.

J Infect Dis, 1997 Feb, 175(2), 474 - 7
Interleukin-8 and chemotactic activity of middle ear effusions; Storgaard M et al.; The importance of interleukin (IL)-8 in the chemotactic activity of middle ear effusions (MEEs) was evaluated . There was a significantly higher IL-8 concentration in MEEs of children with acute otitis media (AOM) (n = 17; 136 ng/mL) than in children with otitis media with effusion (OME) (n = 28; 65 ng/mL) . The IL-8 concentration in MEEs with bacteria (149 ng/mL) was significantly higher than in MEEs without bacteria (66 ng/mL) . MEEs from children with AOM and OME had equally higher chemotactic activity than the diluent alone (23.3% and 24.8% vs . 9.2%) . The chemotactic activity was not altered by the presence of bacteria nor did it correlate with IL-8 concentration . Fractionation of MEEs by gel chromatography demonstrated that the main chemotactic activity could clearly be separated from the IL-8 activity, thus excluding IL-8 as a main chemotactic component in MEEs.

Eur J Cancer, 1997 Feb, 33(2), 209 - 13
Bile duct stents: is there an increased rate of complications in patients receiving chemotherapy?
Lofts FJ, Evans TR, Mansi JL, Glees JP, Knight MJ.
The aim of this study was to determine whether palliative chemotherapy accelerates the rate of biliary stent occlusion, in patients with a malignant biliary obstruction . Such treatment can induce neutropenia and increase the risk of bacterial sepsis . Overgrowth of bacteria within the bile of patients receiving chemotherapy could accelerate the rate of stent occlusion . Retrospective analysis of treatment records for 80 consecutive patients with a diagnosis of adenocarcinoma arising from the pancreas, bile ducts or gall bladder was conducted . Two groups were identified, those with a biliary stent in situ (primary stent group: 47/80; 59%) at the time of referral and those without (no stent group: 33/80; 41%) . The majority of patients went on to receive chemotherapy, 64% and 70% in the primary stent group and no stent group, respectively . The rate of febrile neutropenia was similar in the two groups (5% versus 7% of all chemotherapy cycles in the primary stent group and no stent group, respectively) . The rate of stent occlusion was not significantly different between those exposed to chemotherapy (37%; 95% CI 20-54%) and those unexposed (39%; 95% CI 19-59%) . Similarly, the mean duration of patency was not shortened by chemotherapy (105 days in the chemotherapy group versus 119 days in the non-chemotherapy group; P = 0.97, Mann-Whitney U-test) . We conclude that there is no evidence of increased rate of bile duct-related complications in patients receiving chemotherapy . In particular, we find no indication for the use of prophylactic antibiotics.

J Comp Pathol, 1997 Feb, 116(2), 203 - 16
An immunohistochemical study of the lesions of demodicosis in the dog; Day MJ; Histopathological and immunohistochemical examination of skin biopsies from 32 dogs with demodicosis is reported . There was no association between the different clinical presentations of the disease and the histopathological character of the biopsies, which included absence of inflammation (n = 2), dominant perifolliculitis (n = 11), interface mural folliculitis (n = 7), mural folliculitis (n = 1), furunculosis (n = 10) and nodular dermatitis (n = 1) . In eight of 32 biopsies colonies of coccoid bacteria or Malassezia pachydermatis-like yeasts were observed . IgG-bearing plasma cells were found in similar numbers in the inflammatory infiltrates of all types of histological lesion, and were invariably more numerous than IgM or IgA plasma cells . The IgG plasma cells were largely IgG4 in lesions of perifolliculitis, but consisted of a mixture of IgG2 and IgG4 where folliculitis or furunculosis was present . CD3+T lymphocytes were prominent within the interface infiltrates of follicular epithelium and also within the lesions of furunculosis . Dermal inflammatory cells and epidermal Langerhans cells expressing MHC Class II were observed in similar number in all types of lesion . The study demonstrated an active local cutaneous immune response in canine demodicosis, which increased as the dermal pathology progressed from perifolliculitis to furunculosis.

Acta Paediatr Jpn, 1997 Feb, 39(1), 105 - 13
Trends in development of anti-infective agents in Japan; Yagisawa M et al.; Newly developed anti-infective agents have been continuously supplied in the clinics of Japan over the past 50 years, and were beneficial in saving patients from life-threatening infections . However, the emergence of resistant bacteria and uncommon pathogens has caused complications in chemotherapy . Further efforts have been made to develop newer and more effective agents to minimize such complications . The newest and the most effective agents are available even at primary-care clinics, as the health insurance system allows the use of any agents approved by the government . Despite potential risks of the emergence and spread of resistant bacteria and the occurrence of adverse drug reactions, current usage of such effective agents shows more successful results, particularly in prognosis, than that of less effective ones . Because of the extreme changes in infectious diseases over the last 10-15 years, guidelines for the evaluation of anti-infective agents have been proposed . It is essential to harmonize with the USA and the European guidelines for mutual acceptance of clinical data . However, it is true that practices of anti-infective therapy in Japan, in the USA and in Europe are different both in dose regimen and assessments of efficacy and safety . As the leading country in the development of newer anti-infective agents, Japan proposes the most sophisticated way to evaluate those agents to update knowledge in medical science.

Nutrition, 1997 Feb, 13(2), 128 - 32
Effect of total parenteral nutrition with different lipid emulsions of human monocyte and neutrophil functions; Waitzberg DL et al.; Parenteral nutrition (TPN) with lipid emulsions is claimed to be associated with impaired monocyte (M) and neutrophil (N) functions . Long-chain triglycerides (LCT) and a mixture containing 50% medium-chain triglycerides (MCT) and 50% LCT, currently used in nutritional therapy with TPN, were evaluated for their ex vivo effects on human N and M chemotaxis, phagocytosis, bacterial killing, and oxidative metabolism by nitroblue tetrazolium reduction test . Cell functions were examined in a randomized, crossover, blind trial in 10 malnourished patients with gastric cancer . Prior to the operation (2 wk), central TPN (40 kcal/kg) with 25% of caloric energy provided as LCT or MCT/LCT emulsion was infused over 48 h . After the crossover period fat-free TPN was given over 48 h . Function tests were done for N and M before and after each lipid emulsion infusion . Every cell function test performed for each patient was controlled by another test done in healthy adult volunteers and the results were compared with the normal range of values previously established for a healthy adult population . All the patients completed the studies without complications . Crossover validity was statistically established . Bacterial killing was the only function reduced in neutrophils after LCT emulsion (% killed bacteria = 79.0 +/- 8.5 versus 67.4 +/- 19.2; P < 0.05), although this function remained within the normal range values in 80% of the patients . In conclusion, the lipid emulsions did not affect any monocyte functions and only moderately decreased neutrophil bacterial killing.

J R Army Med Corps, 1997 Feb, 143(1), 31 - 4
Dermatological conditions in winter in primary health care on Operation Resolute (Bosnia); Winfield DA; The distribution of dermatological conditions has been studied in a total of 1822 consultations with British troops in a primary health care setting on Operation Resolute (Bosnia) between 1 January and 4 March 1996 . Approximately one in eight (12%) of the consultations were for skin conditions; eczema was the most common complaint, but, taken as a whole, infections due to virus (excluding warts), fungus and bacteria made up 30% . The overall distribution of diseases was similar to that seen in British general practice.

Int J Parasitol, 1997 Feb, 27(2), 173 - 80
Endoparasite control strategies: implications for biodiversity of native fauna; Spratt DM; Efforts to control the spectrum of diseases that affect humans, our crops and our animals pose problems which need to be debated openly . Widespread use of chemicals in such a broad sphere raises important concerns not only about safety for the users, consumers and target species, but especially about the not so obvious effects upon the ecosystems in which they are used . Some undetermined level of biological diversity is necessary to maintain ecological function and resilience . These, in turn, are necessary for generating the biological resources (trees, fish, wildlife, crops) and ecological services (watershed protection, air cleansing, climate stabilisation, erosion control) on which economic activity and human welfare depend . The driving forces behind decline of biodiversity stem entirely from human activities . Underlying causes are those resulting from the cultural and social factors associated with economic activities and lead to direct depletion of species, and degradation or destruction of habitats . The broad spectrum and high efficacy of the macrocyclic lactones against nematode and arthropod parasites of livestock and companion animals are unprecedented . Cattle, horses, sheep, swine, dogs--to varying degrees all are utilised by humans for economic gain . Detrimental impact upon non-target animals is considered acceptable in eradicating parasites because of their economic importance to commercial livestock production . Production will increase when these parasites are eliminated, but we remain oblivious to the long-term consequences of our actions . What are the ecological limits to rural economic activities? Decomposing animal faeces help to maintain our ecosystem by returning valuable nutrients to the soil . Dung fauna-fungi, yeast, bacteria, nematodes, insects and earthworms--play a non-conspicuous but important and varied role in this decomposition process, a role dependent upon many factors, especially environmental ones . Anthelmintics and pesticides are of considerable value in agriculture, but largely at an unevaluated cost to the greater environment . We have insufficient knowledge of the extent to which a spectrum of anthelmintics and pesticides affect ecological function and ecosystem resilience in our commercial plant and animal production systems . It is time we developed a genuine interest in avoiding "the dialogue of the deal" that in the past has minimised interdisciplinary research between environmental ecology and commercial plant and animal production.

Hybridoma, 1997 Feb, 16(1), 11 - 6
Combinatorial antibodies against human malignant melanoma; Pereira S et al.; The general responsiveness of human melanoma to immunotherapy has been well established, but active immunotherapy of melanoma has been hampered by insufficient information on the immunogenicity of melanoma antigens in patients . We have attempted to identify melanoma-associated antigens recognized by patients' B cells using an antibody phage display approach . Antibody display on filamentous phages allows direct screening of cDNA libraries for expression of cell-surface-reactive antibodies, without the need for antibody production and purification using bacteria or eukaryotic cell systems . This approach was used to identify melanoma-associated cell-surface antigens recognized by patients' B cells . Antibodies produced by the B cells of a melanoma patient (in remission for > 7 years following periodic vaccination with allogeneic melanoma cell vaccine) were displayed as Fabs on the surfaces of filamentous phages . A library of 10(8) phages was absorbed to normal melanocytes, followed by phage binding to and elution from melanoma cells (human lymphocyte antigen nonmatched and vaccine melanoma cells) . Phages were further selected for reactivities with tunicamycin-treated melanoma cells . These procedures resulted in a > 10(6)-fold enrichment of tumor-specific phages from the original phage library . One phage-Fab bound to melanoma cells, other tumor cells, and a few normal cells in cultured cell lines and in tissue sections.

Curr Biol, 1997 Feb 1, 7(2), R108 - 11
Cloning in silico; Cutler S et al.; Cellulose is a major component of plant cell walls, but identification of the enzymes that synthesize it has proven difficult . Now, however, several candidate proteins with sequence homology to bacterial cellulose synthases have been identified by partial sequencing of anonymous cDNA clones from cotton fibers.

Gut, 1997 Feb, 40(2), 211 - 4
Chemical synthesis of nitric oxide in the stomach from dietary nitrate in humans; McKnight GM et al.; BACKGROUND/AIMS: It has been suggested that dietary nitrate, after concentration in the saliva and reduction to nitrite by tongue surface bacteria, is chemically reduced to nitric oxide (NO) in the acidic conditions of the stomach . This study aimed to quantify this in humans . METHODS: Ten healthy fasting volunteers were studied twice, after oral administration of 2 mmol of potassium nitrate or potassium chloride . Plasma, salivary and gastric nitrate, salivary and gastric nitrite, and gastric headspace NO concentrations were measured over six hours . RESULTS: On the control day the parameters measured varied little from basal values . Gastric nitrate concentration was 105.3 (13) mumol/l (mean (SEM), plasma nitrate concentration was 17.9 (2.4) mumol/l, salivary nitrate concentration 92.6 (31.6) mumol/l, and nitrite concentration 53.9 (22.8) mumol/l . Gastric nitrite concentrations were minimal (< 1 mumol/l) . Gastric headspace gas NO concentration was 16.4 (5.8) parts per million (ppm) . After nitrate ingestion, gastric nitrate peaked at 20 minutes at 3430 (832) mumol/l, plasma nitrate at 134 (7.2) mumol/l, salivary nitrate at 1516.7 (280.5) mumol/l, and salivary nitrite at 761.5 (187.7) mumol/l after 20-40 minutes . Gastric nitrite concentrations tended to be low, variable, and any rise was non-sustained . Gastric NO concentrations rose considerably from 14.8 (3.1) ppm to 89.4 (28.6) ppm (p < 0.0001) after 60 minutes . All parameters remained increased significantly for the duration of the study . CONCLUSIONS: A very large and sustained increase in chemically derived gastric NO concentrations after an oral nitrate load was shown, which may be important both in host defence against swallowed pathogens and in gastric physiology.

Genetics, 1997 Feb, 145(2), 467 - 78
Transposon-disruption of a maize nuclear gene, tha1, encoding a chloroplast SecA homologue: in vivo role of cp-SecA in thylakoid protein targeting; Voelker R et al.; A nuclear mutant of maize, tha1, which exhibited defects in the translocation of proteins across the thylakoid membrane, was described previously . A transposon insertion at the tha1 locus facilitated the cloning of portions of the tha1 gene . Strong sequence similarity with secA genes from bacteria, pea and spinach indicates that tha1 encodes a SecA homologue (cp-SecA) . The tha1-ref allele is either null or nearly so, in that tha1 mRNA is undetectable in mutant leaves and cp-SecA accumulation is reduced > or = 40-fold . These results, in conjunction with the mutant phenotype described previously, demonstrate that cp-SecA functions in vivo to facilitate the translocation of OEC33, PSI-F and plastocyanin but does not function in the translocation of OEC23 and OEC16 . Our results confirm predictions for cp-SecA function made from the results of in vitro experiments and establish several new functions for cp-SecA, including roles in the targeting of a chloroplast-encoded protein, cytochrome f, and in protein targeting in the etioplast, a nonphotosynthetic plastid type . Our finding that the accumulation of properly targeted plastocyanin and cytochrome f in tha1-ref thylakoid membranes is reduced only a few-fold despite the near or complete absence of cp-SecA suggests that cp-SecA facilitates but is not essential in vivo for their translocation across the membrane.

Hum Reprod, 1997 Feb, 12(2), 286 - 91
Fertilization with human testicular spermatids: four successful pregnancies; Antinori S et al.; Between July 1995 and May 1996, 36 patients with non-obstructive azoospermia of secretory origin underwent intracytoplasmic injection of spermatids . A previous histological biopsy was performed on all patients: 15 had spermatogenic arrest, a further 13 had Sertoli cell-only syndrome, and the remaining eight had post-cryptorchidism tubal atrophy . The ejaculate was duly examined and a complete absence of spermatozoa and spermatids was confirmed, with only bacteria and debris being found . Testicular sperm extraction (TESE) was then performed . In 19 out of 36 cases round spermatids only were found, while elongated spermatids were found in the remaining 17 . Both round and elongated spermatids were isolated and used for injection . A total of 135 oocytes at metaphase II were recovered from 19 partners and injected with round spermatids, while 123 mature oocytes from 17 partners were injected with elongated spermatids . The number of oocytes fertilized, as judged by the presence of two pronuclei, was 75 (55.5%) and 71 (57.7%) respectively . By 34 h after injection, the number of embryos which had cleaved to the 2-cell stage was 56 (74.6%) with round spermatids and 55 (77.4%) with elongated spermatids . All cleaved embryos were transferred into the uterus of the partners . Clinical pregnancies were established in two cases of round spermatid cycles (10.5%) (both are still ongoing), and three cases of elongated spermatid cycles (17.6%) (two are still ongoing; one was lost after 8 weeks of gestation) . Chromosomal analysis showed that all fetuses had a normal karyotype (three male and one female) with no chromosomal abnormalities.

Surg Endosc, 1997 Feb, 11(2), 93 - 4
Thoracoscopic decortication in infants and children; Rothenberg SS et al.; BACKGROUND: The treatment of pediatric empyemas remains controversial . While thoracentesis and tube thoracostomy appear adequate for relatively benign organisms, virulent bacteria cause thick fibrinous pleural peels entrapping the lung . Open thoracotomies have been effectively used for decortication but are painful . METHODS: We report the use of minimally invasive thoracoscopic decortication in 12 patients (mean age 5 years) . All failed conventional management with persistent fever, increasing oxygen requirement, recurrent effusion, and pleural consolidation; 5- and 10-mm trocars were used and complete decortication was accomplished . RESULTS: Ten of 12 patients were afebrile by 72 h and discharged 4-12 days after surgery . Eleven of 12 patients had clear chest x-rays by 1 month . CONCLUSION: Thoracoscopic decortication is a safe and effective means of treating pediatric empyemas.

Br J Dermatol, 1997 Feb, 136(2), 260 - 3
Skin infection caused by Mycobacterium avium; Ichiki Y et al.; A patient with skin infection due to Mycobacterium avium is reported . A 9-year-old female had 10 subcutaneous nodules and two ulcers on the abdomen and legs . She had no medical history of systemic disease, skin disease or immunosuppressive therapy . Cultures of a biopsy specimen and of aspirated seropurulent fluid in nodules showed acid-fast bacteria, identified as M . avium by the DNA-DNA hybridization method . We treated her with a combination of surgery and the antibiotics, cycloserine, isoniazid and clarithromycin.

Biochem Mol Biol Int, 1997 Feb, 41(2), 243 - 55
The growth kinetics of intracellular bacille Calmette-Guerin (BCG) within human monocyte-derived macrophages of different ethnic populations; Fazal N; In this paper we have examined intracellular mycobacterial growth in human monocyte-derived macrophage cell cultures . Cells from three population groups were selected to compare and contrast the kinetics of intracellular BCG growth both by determining changes in metabolic activity of dividing bacteria by radio-isotope incorporation and growth/survival by estimating colony forming unit by Fazal's Microcolony Counting Method . These data indicate that BCG infects and multiplies intracellularly in human macrophages and may serve as an in vitro model for studying inherent capability of mycobacteria to grow and infect macrophage populations . There is however, no inhibition or reduction of intracellular mycobacterial growth from a Caucasian male and female, and an Afrocarribean female donor macrophages . On the contrary, there is a ten-fold increase in the replication of mycobacteria over a 7-day infection period . Patterns of BCG growth within macrophage cultures from different donors were determined and variation calculated at different days post-infection . Experiment to experiment coefficient of variation of intracellular BCG growth estimates for a single donor is between 4-12% . In contrast, the coefficient of variation of intracellular BCG growth between three different macrophage donors studied is < 4%.

Mol Med Today, 1997 Feb, 3(2), 61 - 8
Microsatellite instability in human solid tumors; Lothe RA; A genome-wide instability has been found in almost all analyzed malignant tumors from patients with hereditary non-polyposis colorectal cancer (HNPCC), and in a subgroup of sporadic (non-inherited) cancers of the same type . This mutator phenotype was initially seen as novel alleles at microsatellite loci (a family of repetitive DNA sequences) and was shown to be caused by mutations in the highly conserved mismatch repair genes . Mutations have been found in each of four of these human genes: hMSH2, hMLH1, hPMS1 and hPMS2, in the germline of HNPCC patients and in their tumors, as well as in sporadic tumors . These recent discoveries provide new molecular diagnostic tools for the detection of patients at high risk of developing carcinomas of the large bowel and other HNPCC-related tumors . Ongoing international research is progressively solving many of the unanswered questions at the genotypic and phenotypic levels of this newly identified mechanism in carcinogenesis.

J Interferon Cytokine Res, 1997 Feb, 17(2), 69 - 75
Treatment of an infected murine macrophage cell line (J774A.1) with interferon-gamma but not tumor necrosis factor-alpha or live Mycobacterium intracellulare alone modulates the expression of adhesion molecules; Pourshafie MR et al.; In the present study, we evaluated whether the activation of a murine macrophage cell line (J774.1A) by treatment with recombinant murine tumor necrosis factor-alpha (rTNF-alpha) or recombinant murine interferon-gamma (rIFN-gamma) before or simultaneous with infection with Mycobacterium intracellulare would affect their ability to express lymphocyte function-associated antigen-1 (LFA-1) and to restrict growth and kill the ingested M . intracellulare . The data showed that the effect of lipopolysaccharide (LPS) in increasing the level of LFA-1 was the same in the presence or absence of M . intracellulare . The inability of M . intracellulare to affect the level of expression of LFA-1 was irrespective of the M . intracellulare to J774A.1 ratio . A significant increase in the expression of LFA-1 was observed when J774A.1 cells were prestimulated with IFN-gamma 1 day before the addition of the bacteria . The addition of IFN-gamma with M . intracellulare simultaneously, however, did not affect the expression of the adhesion molecules as compared with the IFN-gamma alone . Our results indicated no change in the level of LFA-1 on J774A.1 following exposure with TNF-alpha . We observed that preexposure with 10-10(4) IU/ml of TNF-alpha can significantly decrease the number of ingested M . intracellulare . Simultaneous addition of 10(3) and 10(4) IU/ml of TNF-alpha, however, did not have any mycobactericidal effect . This indicates that the TNF-alpha-induced killing by J774A.1 cells was relatively selective, depending on the concentration and the time of presence of TNF-alpha . The data may suggest that the uptake of M . intracellulare is carried out via other adhesion receptors when M . intracellulare and IFN-alpha are present simultaneously and that in the presence of TNF-alpha other surface receptors are involved in the uptake of M . intracellulare . Flow cytometry analysis of the spleen cells removed at various times from M . intracellulare-infected mice also indicated no change in the level of LFA-1 beta or MAC-1, a finding comparable with that of the J774A.1 cells.

Biol Pharm Bull, 1997 Feb, 20(2), 127 - 33
Characterization of lactoferrin-binding proteins of human macrophage membrane: multiple species of lactoferrin-binding proteins with polylactosamine-binding ability; Eda S et al.; Human lactoferrin (LF) specifically binds to human monocytic leukemia cell line THP-1 cells differentiated into macrophages, and it has been suggested that the poly-N-acetyllactosaminyl saccharide chains of LF are involved . We partially purified and characterized LF-binding proteins with affinity for polylactosamines from THP-1 cells . LF-binding activity was solubilized by nonionic detergent Triton X-100 from THP-1 cell membrane, and subjected to affinity chromatography using an LF-Sepharose column . LF-binding activity, detected by ligand blotting assay, was eluted and further fractionated by affinity chromatography using a Sepharose column coupled with band 3, a polylactosaminyl chain-containing glycoprotein of human erythrocyte membrane . LF-binding activity was separated into three fractions (frs . B1, B2, and B3) . These fractions exhibited band 3-binding activity as detected by ligand blotting assay . Dodecylsulfate-polyacrylamide gel electrophoresis of frs . B1, B2, and B3, followed by detection of LF-binding activity on Western blots, indicated that frs . B1, B2, and B3 contained LF-binding proteins with a molecular mass of 35, 50 and 80, and 35-37 kDa, respectively . Binding of LF to each of the fractions on the dot blots was partially inhibited by LF oligosaccharides, band 3 oligosaccharides and lacto-N-neotetraose, each containing di-N-acetyllactosaminyl or analogous structure, Gal beta 1-->4GlcNAc beta 1-->3Gal beta 1-->4GlcNAc (or Glc) . These results suggest that the 35, 50 and/or 80, and 35-37 kDa proteins on THP-1 cells are LF-binding proteins with polylactosamine-binding ability.

Biol Pharm Bull, 1997 Feb, 20(2), 122 - 6
Localization of enzymes involved in metabolism of glycyrrhizin in contents of rat gastrointestinal tract; Akao T; Most digested food 2 h after overnight feeding in rat remained in the stomach, duodenum, upper small intestine, lower small intestine, cecum and colon, all of which indicated pH between 4 and 7 and had glycyrrhizin (GL) hydrolyzing activity . This enzyme activity was highest in the cecal and colonic contents among all gastrointestinal contents . Also, 3 alpha-hydroxyglycyrrhetic acid (3 alpha-hydroxyGA) and 3 beta-hydroxyglycyrrhetic acid (3 beta-hydroxyGA) oxidizing enzymes were localized in the same cecal content . Namely, rat gastrointestinal bacteria had the ability to hydrolyze GL to 3 beta-hydroxyGA by glycyrrizin beta-D-glucuronidase and to oxide 3 beta-hydroxyGA and 3 alpha-hydroxyGA to 3-oxoGA by 3 beta-hydroxyglycyrrhetinate dehydrogenase and 3 alpha-hydroxyglycyrrhetinate dehydrogenase, respectively . In medium of pH 1 to pH 10, metabolites 3 beta-hydroxyGA, 3-oxoGA and 3 alpha-hydroxyGA obtained from the metabolism of GL were the highest in pH 8 . The intestinal contents of pH 6 or pH 7 were able to produce metabolites 3 beta-hydroxyGA in the metabolism of GL . However, the stomach content at pH 4.2 was lowest in metabolite 3 beta-hydroxyGA . It is unknown whether or not GL is metabolized to 3 beta-hydroxyGA by the stomach content in vivo.

Am J Infect Control, 1997 Feb, 25(1), 11 - 5
A randomized trial of surgical scrubbing with a brush compared to antiseptic soap alone; Loeb MB et al.; BACKGROUND: The difference between use of a scrub brush versus soap alone in reducing hand bacterial counts has never been established by a prospective, comparative study . METHODS: Fifteen volunteers were taught the 5-minute surgical scrub . Baseline specimens were obtained by the glove fluid sampling procedure . Subjects were randomized to (1) scrub with an inert scrub brush and 4% chlorhexidine soap with isopropyl alcohol or (2) wash with 4% chlorhexidine soap with isopropyl alcohol alone . Specimens were obtained immediately after the scrub was completed and 45 minutes later . The experiment was repeated by use of a cross-over design after a 1-week washout period . The data were analyzed by three methods that took into account the broad range of baseline hand counts (5 x 10(1) to 11.2 x 10(4): method 1, the discordance between presence/absence of hand bacterial counts within individuals at 45 minutes for soap versus soap and brush; method 2, the absolute reduction of bacteria (baseline vs 45 min.) for soap versus soap and brush; and method 3, the proportional change in bacterial counts at 45 minutes from baseline for soap versus soap and brush . RESULTS: Although there was no statistically significant difference for any method, the point estimates for the odds ratio (OR) showed that up to twice the number of subjects had a greater reduction in bacterial counts when they washed with soap than when they scrubbed with a brush, as evidenced by the following data: method 1, OR 2.3 (95% confidence interval {CI} 0.53, 13.99) for soap alone; method 2, OR 1.0 (CI 0.23, 4.35); and method 3, OR 2.0 (CI 0.54, 9.10) for soap alone . CONCLUSIONS: The effect of use of soap alone in reducing hand bacterial counts at 45 minutes was similar to use of soap and brush . Soap can be used alone and the surgical infection rate prospectively monitored.

Poult Sci, 1997 Feb, 76(2), 248 - 55
The applicability of particleboard residue as a litter material for male turkeys; Hester PY et al.; Particleboard residue is a by-product of the secondary wood products manufacturing industries . Large quantities of this product are landfilled for lack of better use . The objective of the current study was to investigate the possibility of using particleboard residue as a litter source for male turkeys . Two sizes of particleboard residue, fine and coarse, were compared to hardwood shavings . Compared to hardwood shavings, fine and coarse particleboard was a drier, cleaner product initially, as indicated by lower moisture content as well as bacteria and mold counts at Day 0 . Turkeys reared for 123 d on fine particleboard had several advantages over those reared on either the coarse particleboard or hardwood shavings, which included significantly lowered incidences of breast buttons and leg abnormalities . Perhaps due to the jagged edges and coarser texture, coarse particleboard increased the incidence of foot pad dermatitis when compared to the other two litter sources . Turkeys reared on fine particle-board had a 0.16 kg reduction (P < 0.01) in live market body weights compared to the toms reared on hardwood shavings, but this was offset by a 0.22 kg gain in muscle deposition (P < 0.05) . Mortality, breast weights and yields, and feed efficiency were unaffected by litter source . Based on the variables studied, it was concluded that fine particleboard residue could be used as an alternative bedding material for male turkeys.

J Orthop Trauma, 1997 Feb-Mar, 11(2), 121 - 5
Benzalkonium chloride: a potential disinfecting irrigation solution; Gainor BJ et al.; OBJECTIVE: To determine the disinfecting properties of benzalkonium chloride as an irrigation agent . DESIGN: Comparison was made between irrigation of contaminated muscle strips with benzalkonium chloride and normal saline (control) . SUMMARY OF BACKGROUND DATA: Benzalkonium chloride is a cationic disinfectant, which has questionable efficacy in an organic environment . However, no previous study has attempted to use high volumes of this cationic solution to overcome the neutralizing effect of organic tissue and thus maintain this detergent's germicidal properties . METHODS: 2.5 cm x 0.5 cm x 0.5 cm pieces of bovine muscle were aseptically cut from the center of freshly harvested beef muscle and incubated with 1.0 x 10(7) colony forming units of bacteria for 15 minutes . The muscle strips were then irrigated with either 100 mL, 1 L, or 10 L of benzalkonium chloride at a 1:2000 concentration in normal saline . Normal saline was used as the control . The muscle strips were sonicated to remove adherent bacteria; the number of living organisms was determined by quantitatively culturing the sonicate . RESULTS: In vitro, benzalkonium chloride was superior to normal saline at disinfecting bovine muscle (p < or = 0.001) . When 10 L of benzalkonium chloride irrigation was used, no living bacteria could be recovered (p < or = 0.012) . CONCLUSION: In this experimental setting benzalkonium chloride was an effective disinfection agent, with enhanced activity at large volumes.

J Anim Sci, 1997 Feb, 75(2), 556 - 65
Cellular defense mechanisms in the udder and lactation of goats; Paape MJ et al.; Migration of neutrophils into mammary tissue provides the first immunological line of defense against bacteria that penetrate the physical barrier of the teat canal . Evasion of neutrophil defenses provides an opportunity for invading bacteria to become established . Depletion of neutrophils results in a dramatic increase in susceptibility to intramammary infection . Numerous cytoplasmic particles are shed from the apical surface of mammary secretory cells during milk secretion in goats . Only those counting methods that are specific for deoxyribonucleic acid can distinguish cell-like particles from somatic cells and thereby give reliable estimates of somatic cell numbers in goat milk . Unlike in milk from dairy cows, the somatic cell count in goat milk is influenced by the presence of nucleated cytoplasmic particles, stage of lactation, parity, and caprine arthritis-encephalitis . Investigations indicate that a dry period is necessary for optimal milk production in dairy cows but may not be necessary in goats . However, in many other respects regulation of bovine and caprine lactation seems to be quite similar . Studies have demonstrated additive galactopoietic effects of growth hormone and frequent milking in both species and a recently isolated chemical feedback inhibitor of lactation seems effective across both species . Increasing lactational performance has the potential for decreasing milk somatic cell counts in late lactation.

Naunyn Schmiedebergs Arch Pharmacol, 1997 Feb, 355(2), 190 - 7
In U-937 promonocytes, misteltoe lectin I increases basal {Ca2+}i, enhances histamine H1- and complement C5a-receptor-mediated rises in {Ca2+}i, and induces cell death; Wenzel-Seifert K et al.; Mistletoe lectin I (ML I) from Viscum album inhibits cell growth and induces apoptosis (programmed cell death) in several cell types . Because increases in cytosolic Ca2+ concentration ({Ca2+}i) constitute a signal for the induction of apoptosis, we studied the effects of ML I on basal {Ca2+}i, receptor-mediated rises in {Ca2+}i and cell viability, using human U-937 promonocytes as model system . Treatment of U-937 cells with ML I (30-100 ng/ml) significantly increased basal {Ca2+}i . ML I (10-30 ng/ml) enhanced histamine-induced rises in {Ca2+}i up to five-fold . The effect of histamine was inhibited by clemastine but not by famotidine, indicative for its mediation via H1-receptors . ML I additionally enhanced the stimulatory effect of complement C5a on {Ca2+}i, whereas the effect of ATP was unaffected . ML I did not induce responsiveness of U-937 cells towards a bacteria-derived chemotactic peptide . ML I up to 10 ng/ml did not affect cell viability and growth of U-937 cells . ML I at 30 ng/ml moderately inhibited cell growth and reduced cell viability . At 100 ng/ml, ML I was strongly cytotoxic . Our data support the view that Ca2+ plays a role as intracellular signal molecule in the induction of apoptosis and point to an accelerating role of H1- and C5a-receptors in the regulation of this process.

Plant Mol Biol, 1997 Feb, 33(3), 381 - 92
Light induces accumulation of isocitrate lyase mRNA in a carotenoid-deficient mutant of Chlamydomonas reinhardtii; Petridou S et al.; A cDNA with sequence similarity to isocitrate lyase (ICL) genes was isolated from the unicellular eukaryotic green alga Chlamydomonas reinhardtii as a light-induced mRNA in the carotenoid biosynthetic mutant strain FN68 . The 416 amino acid open reading frame shows significant sequence similarity to isocitrate lyases of bacteria (70%), molds (48%), yeasts (45%), and plants (47%) . Expression of the Chlamydomonas ICL gene was tested in the mutant strain FN68, which when grown in the dark fails to accumulate carotenoids and is deficient in chlorophyll, and in CC400G, a strain that accumulates wild-type levels of carotenoids and chlorophyll . In vegetative CC400G cells, ICL mRNA accumulated to a high level in the dark and declined to a barely detectable level within 30 min of exposure to light . This response was more sensitive to white (tungsten filament) or red light than green or blue light, excluding cryptochrome and rhodopsin as the photoreceptor . These results are consistent with excitation by chlorophyll and/or a phytochrome-related photoreceptor . In vegetative FN68 cells, ICL mRNA abundance was very low in the dark, but increased dramatically in response to light . At intensities above threshold, excitation by far-red or red light-induced ICL mRNA accumulation to the highest levels . The threshold of the response was lowest for far-red and blue light . These results are consistent with excitation of a photochromic far-red-responsive pigment.

Antonie Van Leeuwenhoek, 1997 Feb, 71(1-2), 159 - 78
Evolution of novel metabolic pathways for the degradation of chloroaromatic compounds; van der Meer JR; Chlorobenzenes are substrates not easily metabolized by existing bacteria in the environment . Specific strains, however, have been isolated from polluted environments or in laboratory selection procedures that use chlorobenzenes as their sole carbon and energy source . Genetic analysis indicated that these bacteria have acquired a novel combination of previously existing genes . One of these gene clusters contains the genes for an aromatic ring dioxygenase and a dihydrodiol dehydrogenase . The other contains the genes for a chlorocatechol oxidative pathway . Comparison of such gene clusters with those from other aromatics degrading bacteria reveals that this process of recombining or assembly of existing genetic material must have occurred in many of them . Similarities of gene functions between pathways suggest the incorporation of existing genetic material has been the most important mechanism of expanding a metabolic pathway . Only in a few cases a horizontal expansion, that is acquisition of gene functions to accommodate a wider range of substrates which are then all transformed in one central pathway, is observed on the genetic level . Evidence is presented indicating that the assembly process may trigger a faster divergence of nearby gene sequences . Further 'fine-tuning', for example by developing a proper regulation, is then the next step in the adaptation.

Plant Physiol, 1997 Feb, 113(2), 463 - 8
Origin of the thiazole ring of camalexin, a phytoalexin from Arabidopsis thaliana; Zook M et al.; The principal phytoalexin that accumulates in Arabidopsis thaliana after infection by fungi or bacteria is 3-thiazol-2'-yl-indole (camalexin) . Detached noninoculated leaves of Arabidopsis and leaves inoculated with the fungus Cochliobolus carbonum were fed {35S}cysteine (Cys) and {35S}methionine . Inoculated leaves incorporated more than a 200-fold greater amount of radioactivity from {35S}Cys into camalexin, as compared with noninoculated leaves . The amount of radioactivity from {35S}Cys that was incorporated into camalexin from inoculated Arabidopsis leaves was 10-fold greater than the amount of radioactivity that was incorporated into camalexin from {35S}methionine . Additional labeling experiments were performed to determine whether other atoms of Cys are incorporated into camalexin . {14C}Cys and {35S}Cys were incorporated into camalexin with approximately the same efficiency . Cys labeled either with deuterium (D3-Cys{2,3,3}) or 13C and 15N ({U-13C,15N}Cys) was also fed to inoculated leaves of Arabidopsis; camalexin was analyzed by mass spectroscopic analysis . The average ratio of molecular ion intensities of 203/200 for {U-13C,15N}Cys-labeled camalexin was 4.22, as compared with 0.607 for the average 203/200 ratio for unlabeled camalexin . The mass fragment-ion intensity ratios of 60/58 (thiazole ring ion fragment) and 143/142 were also higher for {U-13C,15N}Cys-labeled camalexin, as compared with unlabeled camalexin . The 59/58 and 201/200 ratios were higher for D3-Cys-labeled camalexin as compared with unlabeled camalexin . These data are consistent with the predicted formation of the thiazole ring of camalexin from Cys.

Bioessays, 1997 Feb, 19(2), 117 - 26
Chloride channels: an emerging molecular picture; Jentsch TJ et al.; Chloride channels are probably found in every cell, from bacteria to mammals . Their physiological tasks range from cell volume regulation to stabilization of the membrane potential, signal transduction, transepithelial transport and acidification of intracellular organelles . These different functions require the presence of many distinct chloride channels, which are differentially expressed and regulated by various stimuli . These include various intracellular messengers (like calcium and cyclic AMP), pH, extracellular ligands and transmembrane voltage . Three major structural classes of chloride channels are known to date, but there may be others not yet identified . After an overview of the general functions of chloride channels, this review will focus on these cloned chloride channels: the CLC chloride channel family, which includes voltage-gated chloride channels, and the cystic fibrosis transmembrane regulator (CFTR), which performs other functions in addition to being a chloride channel . Finally, a short section deals with GABA and glycine receptors . Diseases resulting from chloride channel defects will be specially emphasized, together with the somewhat limited information about how these proteins work at the molecular level.

Mol Microbiol, 1997 Feb, 23(3), 537 - 49
Substrate induction and glucose repression of maltose utilization by Streptomyces coelicolor A3(2) is controlled by malR, a member of the lacl-galR family of regulatory genes; van Wezel GP et al.; malR of Streptomyces coelicolor A3(2) encodes a homologue of the Lacl/GalR family of repressor proteins, and is divergently transcribed from the malEFG gene cluster, which encodes components of an ATP-dependent transport system that is required for maltose utilization . Transcription of malE was induced by maltose and repressed by glucose . Disruption or deletion of malR resulted in constitutive, glucose-insensitive malE transcription at a level markedly above that observed in the parental malR+ strain, and overproduction of MalR prevented growth on maltose as carbon source . Consequently, MalR plays a crucial role in both substrate induction and glucose repression of maltose utilization . malR is expressed from a single promoter with transcription initiating at the first G of the predicted GTG translation start codon.

Microbiology, 1997 Feb, 143 ( Pt 2), 539 - 46
A new insertion sequence IS1452 from Acetobacter pasteurianus; Kondo K et al.; A new insertion sequence element, IS1452, was found to be associated with inactivation of the alcohol dehydrogenase by insertion in the adhS gene encoding subunit III of the three-component membrane-bound alcohol dehydrogenase complex in Acetobacter pasteurianus . Cloning and sequencing analyses of the mutated subunit III gene locus revealed that IS1452 was inserted at or near the ribosome-binding sequence of adhS . Analysis of transcription using the chloramphenicol acetyltransferase gene as the reporter indicated that IS1452 abolished transcription of adhS by separating its promoter from the subunit III structural gene . IS1452 was 1411 bp in length and had a terminal inverted repeat of 21 bp . IS1452 contained one long ORF of 416 amino acids rich in basic amino acids . This protein showed homology with a putative transposes, Tra1, of IS701 isolated from the cyanobacterium Calothrix species PCC 7601 . Like IS701, IS1452 was found to generate a 4 bp direct repeat at the site of insertion upon transposition . The target site specificity was rather strict, and a CTA(A or G) sequence appeared to be preferentially recognized . Transposition of IS1452 was replicative, since it was accompanied by an increase in the copy number of IS1452 . Several strains belonging to the genus Acetobacter also contained IS1452 at varying copy numbers from one to more than ten . These observations suggest that IS1452 is one of the insertion sequences that are responsible for genetic instability leading to deficiencies in various physiological properties in acetic acid bacteria.

Scand J Immunol, 1997 Feb, 45(2), 115 - 31
Host responses and antigens involved in protective immunity to Mycobacterium tuberculosis; Andersen P; Tuberculosis (TB) is the largest single infectious cause of human mortality . The incidence of TB has remained high in most of the developing world and the disease has recently re-emerged as a public health problem in industrialized countries . The development of a new improved TB vaccine is a highly prioritized international research area, which has been further stimulated by the appearance of multi-drug resistant strains of Mycobacterium tuberculosis . The present status of the attempts to characterize the protective immune response to TB will be reviewed with special emphasis on recent progress in the identification and characterization of target molecules recognized by protective cells . This paper will focus on proteins released from live bacteria and discuss their role in the host-pathogen interaction and the ongoing attempts to use these molecules in TB subunit vaccines.

Protein Sci, 1997 Feb, 6(2), 450 - 8
Thermoanaerobacter brockii alcohol dehydrogenase: characterization of the active site metal and its ligand amino acids; Bogin O et al.; The active-site metal ion and the associated ligand amino acids in the NADP-linked, tetrameric enzyme Thermoanaerobacter brockii alcohol dehydrogenase (TBADH) were characterized by atomic absorption spectroscopy analysis and site-directed mutagenesis . Our preliminary results indicating the presence of a catalytic zinc and the absence of a structural metal ion in TBADH (Peretz & Burstein . 1989 . Biochemistry 28:6549-6555) were verified . To determine the role of the putative active-site zinc, we investigated whether exchanging the zinc for other metal ions would affect the structural and/or the enzymatic properties of the enzyme . Substituting various metal ions for zinc either enhanced or diminished enzymatic activity, as follows: Mn2+ (240%); Co2+ (130%); Cd2+ (20%); Cu2+ or V3+ (< 5%) . Site-directed mutagenesis to replace any one of the three putative zinc ligands of TBADH, Cys 37, His 59, or Asp 150, with the non-chelating residue, alanine, abolished not only the metal-binding capacity of the enzyme but also its catalytic activity, without affecting the overall secondary structure of the enzyme . Replacing the three putative catalytic zinc ligands of TBADH with the respective chelating residues serine, glutamine, or cysteine damaged the zinc-binding capacity of the mutated enzyme and resulted in a loss of catalytic activity that was partially restored by adding excess zinc to the reaction . The results imply that the zinc atom in TBADH is catalytic rather than structural and verify the involvement of Cys 37, His 59, and Asp 150 of TBADH in zinc coordination.

J Bone Miner Res, 1997 Feb, 12(2), 165 - 71
Human stanniocalcin inhibits renal phosphate excretion in the rat; Wagner GF et al.; Stanniocalcin (STC) is a glycoprotein hormone first identified in bony fishes where it counteracts hypercalcemia by inhibiting gill calcium uptake and stimulating renal inorganic phosphate (Pi) reabsorption . Human STC (hSTC) has recently been cloned and sequenced and is highly homologous to the fish hormone at the amino acid level . The objective of this study was to examine the possible effects of hSTC on electrolyte homeostasis and renal function in the rat . Recombinant hSTC was expressed in bacteria and purified by metal-ion affinity chromatography and reverse-phase high performance liquid chromatography . Anesthetized animals were given bolus infusions of 1, 5, or 10 nmol hSTC per kilogram of body weight . Control animals received solvent alone . The most effective dosage was 5 nmol/kg, which caused significant reductions in both absolute and fractional phosphate excretion in comparison with control rats . The hSTC had no effect on the renal excretion of other ions, the glomerular filtration rate, renal blood flow, blood pressure, or plasma electrolytes (Na+, K+, Ca2+, Pi, Mg/+) . The maximum effect of hSTC on phosphate excretion was observed 60-80 minutes postinjection . Lesser effects were obtained with higher and lower dosages of hormone . When renal cortical brush-border membrane vesicles were isolated from control and hormone-treated animals 80 minutes postinjection, the rate of Na+/Pi cotransport was found to be 40% higher in vesicles from hormone-treated animals (p < 0.01; 5 nmol hSTC/kg) . Together, the renal clearance and membrane vesicle data indicate that hSTC participates in the renal regulation of Pi homeostasis in mammals.

Curr Opin Immunol, 1997 Feb, 9(1), 17 - 23
Cytokines acting on or secreted by macrophages during intracellular infection (IL-10, IL-12, IFN-gamma); Trinchieri G; The three cytokines IL-12, IL-10, and IFN-gamma have important and cross-regulatory roles in infection . In the past year, much progress has been made in the understanding of the cellular and molecular mechanisms involved in the regulation (and cross-regulation) of these three cytokines and their role in pathology . IL-12 is rapidly produced after infection and acts as a proinflammatory cytokine eliciting the production, by T cells and natural killer cells, of IFN-gamma which activates phagocytic cells . The production of IL-12 is strictly regulated by negative and positive feedback mechanisms . If IL-12 and IL-12-induced IFN-gamma are present during early T cell expansion in response to antigen, Th1 cell generation is favored and the generation of Th2 cells is inhibited . Thus, IL-12 is also a potent immunoregulatory cytokine which promotes Th1 differentiation and is instrumental in the Th1-dependent resistance to infections by bacteria, intracellular parasites, fungi, and certain viruses . Viruses inducing a permanent or transient immunodepression, such as HIV and measles, may act, in part, by suppressing IL-12 production.

Nat Struct Biol, 1997 Feb, 4(2), 140 - 6
The structure of the cytochrome p450BM-3 haem domain complexed with the fatty acid substrate, palmitoleic acid; Li H et al.; The substrate-bound structures of two cytochrome P450s, P450cam and P450eryF, are known . While these structures reveal important features that control substrate specificity, the problem of how conformational changes allow for substrate entry and product release remains unsolved . The structure of the haem domain of the bacterial fatty acid hydroxylase, P450BM-3, previously was solved in the substrate-free form . Unlike the substrate-bound P450cam and P450eryF structures, the substrate access channel is open in substrate-free P450BM-3 . Here we present the X-ray structure of P450BM-3 at 2.7 A bound with a fatty acid substrate, palmitoleic acid . A comparison of the substrate-bound and -free forms reveals major conformational differences and provides the first detailed picture of substrate-induced conformational changes in a P450.

Appl Environ Microbiol, 1997 Feb, 63(2), 687 - 93
Transformation yields of chlorinated ethenes by a methanotrophic mixed culture expressing particulate methane monooxygenase; Anderson JE et al.; Transformation yields for the aerobic cometabolic degradation of five chlorinated ethenes were determined by using a methanotrophic mixed culture expressing particulate methane monooxygenase (pMMO) . Transformation yields (expressed as moles of chlorinated ethene degraded per mole of methane consumed) were 0.57, 0.25, 0.058, 0.0019, and 0.00022 for trans-1,2-dichloroethylene (t-DCE), vinyl chloride (VC), cis-1,2-dichloroethylene (c-DCE), trichloroethylene (TCE), and 1,1-dichloroethylene (1,1-DCE), respectively . Degradation of t-DCE and VC was observed only in the presence of formate or methane, sources of reducing energy necessary for cometabolism . The t-DCE and VC transformation yields represented 35 and 15%, respectively, of the theoretical maximum yields, based on reducing-energy availability from methane dissimilation to carbon dioxide, exclusive of all other processes that require reducing energy . The yields for t-DCE and VC were 20 times greater than the yields reported by others for cells expressing soluble methane monooxygenase (sMMO) . Transformation yields for c-DCE, TCE, and 1,1-DCE were similar to or less than those for cultures expressing sMMO . Although methanotrophic biotreatment systems have typically been designed to incorporate cultures expressing sMMO, these results suggest that pMMO expression may be highly advantageous for degradation of t-DCE or VC . It may also be much easier to maintain pMMO expression in treatment systems, because pMMO is expressed by all methanotrophs whereas sMMO is expressed only by type II methanotrophs under copper-limited conditions.

J Bacteriol, 1997 Feb, 179(4), 1143 - 52
Cloning and sequence of cymA, a gene encoding a tetraheme cytochrome c required for reduction of iron(III), fumarate, and nitrate by Shewanella putrefaciens MR-1; Myers CR et al.; The cymA gene, which encodes a tetraheme cytochrome c, was cloned from Shewanella putrefaciens MR-1 . This gene complemented a mutant which had a TnphoA insertion in cymA and which was deficient in the respiratory reduction of iron(III), nitrate, fumarate, and manganese(IV) . The 561-bp nucleotide sequence of cymA encodes a protein of 187 amino acids with a predicted molecular mass of 20.8 kDa . No N-terminal signal sequence was readily apparent; consistent with this, a cytochrome with a size of 21 kDa was detected in the wild type but was absent in the insertional mutant . The cymA gene is transcribed into an mRNA; the major transcript was approximately 790 bases, suggesting that it is not part of a multicistronic operon . This RNA transcript was not detected in the cymA mutant . The CymA protein was found in the cytoplasmic membrane and soluble fraction of MR-1, and it shares partial amino acid sequence homology with multiheme c-type cytochromes from other bacteria . These cytochromes are ostensibly involved in the transfer of electrons from the cytoplasmic membrane to acceptors in the periplasm . The localization of the fumarate and iron(III) reductases to the periplasm and outer membrane of MR-1, respectively, suggests the possibility of a similar electron transfer role for CymA.

J Leukoc Biol, 1997 Feb, 61(2), 133 - 40
Marked inhibition of the intracellular multiplication of Legionella pneumophila in monocytes isolated from carriers of human T lymphotrophic virus type I; Funai N et al.; Intracellular growth patterns of Legionella pneumophila were examined in monocytes obtained from carriers of human T-lymphotropic virus type I (HTLV-1) and controls who were HTLV-1 seronegative . All subjects were seronegative for antibodies against L . pneumophila . Bacterial growth was determined 0, 1, 2, and/or 3 days after infecting peripheral blood mononuclear cells (PBMCs) with the bacteria . The intracellular growth of L . pneumophila was markedly inhibited in HTLV-1 carriers compared with normal controls . When the lymphocytes were depleted from the HTLV-1 carrier PBMC cultures before infection, this inhibition was abolished . Inhibition reappeared, however, when the 72-h culture supernatants of PBMCs from HTLV-1 carriers were added to the lymphocyte-depleted cultures . Culture supernatants of infected and uninfected PBMCs from HTLV-1 carriers exhibited markedly increased levels of interferon-gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) compared with the HTLV-1 seronegative controls . In the HTLV-1 carriers, IFN-gamma was produced by the CD4+ lymphocytes, whereas TNF-alpha was secreted by the monocytes . Addition of anti-IFN-gamma or anti-TNF-alpha antibodies to the HTLV-1 carrier PBMC cultures diminished the inhibition of intracellular growth of L . pneumophila . Results suggest that the monocytes are activated in HTLV-1 carriers . These findings may explain why an opportunistic infection by certain intracellular pathogens is rarely seen among HTLV-1 carriers.

Antimicrob Agents Chemother, 1997 Feb, 41(2), 475 - 7
Comparative evaluation of the inhibitory activities of the novel penicillanic acid sulfone Ro 48-1220 against beta-lactamases that belong to groups 1, 2b, and 2be; Tzouvelekis LS et al.; The inhibitory activity of the penicillanic acid sulfone Ro 48-1220 against group 1, 2b, and 2be beta-lactamases was evaluated . Ro 48-1220 inhibited TEM and SHV as effectively as clavulanate and tazobactam . It also inhibited group 1 beta-lactamases at lower concentrations than tazobactam . Ro 48-1220, at a concentration of 4 micrograms/ml, protected ceftriaxone and ceftazidime against strains producing group 1 and 2be beta-lactamases.

Antimicrob Agents Chemother, 1997 Feb, 41(2), 379 - 84
A cytotoxicity assay for evaluation of candidate anti-Pneumocystis carinii agents; Cushion MT et al.; A series of over 60 agents representing several different classes of compounds were evaluated for their effects on the ATP pools of Pneumocystis carinii populations derived from immunosuppressed rats . A cytotoxicity assay based on an ATP-driven bioluminescent reaction was used to determine the concentration of agent which decreased the P . carinii ATP pools by 50% versus untreated controls (IC50) . A ranking system based on the IC50 value was devised for comparison of relative responses among the compounds evaluated in the cytotoxic assay and for comparison to in vivo efficacy . With few exceptions, there was a strong correlation between results from the ATP assay and the performance of the compound in vivo . Antibiotics, with the exception of trimethoprim-sulfamethoxazole (TMP-SMX), were ineffective at reducing the ATP pools and were not active clinically or in the rat model of P . carinii pneumonia . Likewise, other agents not expected to be effective, e.g., antiviral compounds, did not show activity . Standard anti-P . carinii compounds, e.g., TMP-SMX, pentamidine, and dapsone, dramatically reduced ATP levels . Analogs of the quinone and topoisomerase inhibitor groups were shown to reduce ATP concentrations and hold promise for further in vivo investigation . The cytotoxicity assay provides a rapid assessment of response, does not rely on replicating organisms, and should be useful for assessment of structure-function relationships.

Am J Clin Oncol, 1997 Feb, 20(1), 24 - 30
Phase I trial of etoposide, carboplatin, and GM-CSF in extensive small-cell lung cancer: a Cancer and Leukemia Group B study (CALGB 8832); Luikart SD et al.; The maximum tolerated dose (MTD) of etoposide and carboplatin without growth factor support was previously defined by Cancer and Leukemia Group B (CALGB) as 200 and 125 mg/m2/day x 3, respectively, given every 28 days to previously untreated patients who have extensive, small-cell lung cancer (SCLC) . Myelosuppression was dose-limiting . The purpose of this phase I trial was to determine if granulocyte macrophage colony-stimulating factor (GM-CSF) support allows the dosage of the combination of etoposide and carboplatin to be increased above the previously determined MTD . In this CALGB study of 44 evaluable patients with performance status 0-2, cohorts were treated with etoposide and carboplatin given intravenously on days 1-3 followed by GM-CSF (molgramostim) given subcutaneously on days 4-18 . Four dose levels of bacteria-derived recombinant GM-CSF (5, 10, 20 microg/kg/day and 5 microg/kg every 12 h), three dose levels of etoposide (200, 250, and 300 mg/m2/day x 3), and two dose levels of carboplatin (125 and 150 mg/m2/day x 3) were evaluated . There was no chemotherapy dose escalation in individual patients . With 5 microg/kg/d GM-CSF, the first etoposide and carboplatin cycle of 300 and 150 mg/m2/day x 3, respectively, could be administered with acceptable toxicity . However, GM-CSF did not allow repeated administration of this dose-escalated regimen every 21 days, since delayed platelet and/or neutrophil recovery was dose limiting in later cycles . These results demonstrate that GM-CSF alone has limited capability to support the repeated administration of high doses of etoposide and carboplatin . CALGB currently is testing the ability of interleukin (IL)-6 given with GM-CSF to ameliorate the cumulative myelosuppression of this intense regimen.

Infect Immun, 1997 Feb, 65(2), 750 - 7
Inhibition of Helicobacter pylori binding to gastrointestinal epithelial cells by sialic acid-containing oligosaccharides; Simon PM et al.; Helicobacterpylori, the ulcer pathogen residing in the human stomach, binds to epithelial cells of the gastric antrum . We have examined binding of 13 bacterial isolates to epithelial cell lines by use of a sensitive microtiter plate method in which measurement of bacterial urease activity provides the means for quantitation of bound organisms . Several established human gastrointestinal carcinoma cell lines grown as monolayers were compared for suitability in these assays, and the duodenum-derived cell line HuTu-80 was selected for testing bacterial binding inhibitors . When bacteria are pretreated with oligosaccharides, glycoproteins, and glycolipids, a complex picture of bacterial-epithelial adherence specificities emerges . Among the monovalent inhibitors tested, 3'-sialyllactose (NeuAc alpha2-3Gal beta1-4Glc; 3'SL) was the most active oligosaccharide, inhibiting adherence for recent clinical isolates of H . pylori with a millimolar 50% inhibitory concentration (IC50) . Its alpha2-6 isomer (6'SL) was less active . Most of the recent clinical isolates examined were inhibited by sialyllactose, whereas long-passaged isolates were insensitive . Among the long-passaged bacterial strains whose binding was not inhibited by 3'SL was the strain ATCC 43504, also known as NCTC 11637 and CCUG 17874, in which the proposed sialyllactose adhesin was recently reported to lack surface expression (P . G . O'Toole, L . Janzon, P . Doig, J . Huang, M . Kostrzynska, and T . H . Trust, J . Bacteriol . 177:6049-6057, 1995) . Pretreatment of the epithelial monolayer with neuraminidase reduced the extent of binding by those bacteria that are sensitive to inhibition by 3'SL . Other potent inhibitors of bacterial binding are the glycoproteins alpha1-acid glycoprotein, fetuin, porcine gastric and bovine submaxillary mucins, and the glycolipid sulfatide, all of which present multivalent sialylated and/or sulfated galactosyl residues under the conditions of the binding assay . Consistent with this pattern, a multivalent neoglycoconjugate containing 20 mol of 3'SL per mol of human serum albumin inhibited bacterial binding with micromolar IC50 . The H . pylori isolate most sensitive to inhibition by 3'SL was least sensitive to inhibition by sulfatide, gastric mucin, and other sulfated oligosaccharides . Bacteria that have been allowed to bind epithelial cells are also effectively detached by 3'SL . These results describe a heterogeneous adherence repertoire for these bacteria, but they also confirm the critical role of the 3'SL structure on human gastric epithelial cells as an adherence ligand for recent isolates of H . pylori.

Infect Immun, 1997 Feb, 65(2), 422 - 7
Role of the carboxyl-terminal region of Porphyromonas gingivalis fimbrillin in binding to salivary proteins; Nagata H et al.; Porphyromonas gingivalis fimbriae are considered to play an important role in the adherence and colonization of the bacteria in the oral cavity . In this study, we generated and purified three carboxyl-terminal variants of recombinant fimbrillin (r-FimA 224-337, r-FimA 266-337, and r-FimA 287-337, corresponding to amino acid residues 224 to 337, 266 to 337, and 287 to 337, respectively, of the 43-kDa fimbrillin of P . gingivalis 2561) . They were used as inhibitors of P . gingivalis cell binding to human salivary protein-coated hydroxyapatite (HAP) beads . All of the carboxyl-terminal region polypeptides inhibited binding in a dose-dependent manner; however, the inhibitory effect of r-FimA 287-337 was less than that of the other two polypeptides when HAP beads were coated with whole saliva or purified salivary proline-rich protein 1 (PRP1) . Assays of binding of a synthetic peptide corresponding to amino acid residues 266 to 286 of P . gingivalis 2561 fimbrillin to salivary proteins showed that this peptide bound strongly to whole saliva or PRP1 but only weakly to statherin . These results suggest that the carboxyl-terminal region corresponding to amino acid residues 266 to 337 of P . gingivalis fimbrillin plays an important role in binding to salivary proteins and that the domain corresponding to amino acids 266 to 286 is likely a major binding site for PRP1s and the domain corresponding to amino acids 287 to 337 is likely a major binding site for statherin.

J Bacteriol, 1997 Feb, 179(3), 663 - 9
Analysis of the cbbXYZ operon in Rhodobacter sphaeroides; Gibson JL et al.; Three genes, cbbX, cbbY, and cbbZ were found downstream from the form I ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes of Rhodobacter sphaeroides . As in chemoautotrophic bacteria, cbbZ was shown to encode phosphoglycolate phosphatase (PGP), whereas the identities of cbbX and cbbY are not known . To determine the physiological function of the cbbXYZ gene products, we constructed R . sphaeroides strains in which the genes were inactivated and characterized the resultant mutant strains according to growth phenotype and levels of RubisCO and PGP . Only a mutation in cbbX resulted in a discernible phenotype, namely, impaired photoautotrophic growth . No PGP activity was observed in any of the mutants, suggesting that the three genes are transcriptionally linked . Studies with a spontaneous chemoautotrophic competent derivative of the CBBX mutant suggested that the cbbXYZ gene products are not essential for chemoautotrophic growth . PGP activity determined in the wild-type strain grown under a variety of growth conditions, and in various strains containing mutations in Calvin-Benson-Bassham cycle structural and regulatory genes, indicated that transcription of the cbb(I) operon influenced expression of the downstream cbbXYZ operon.

J Clin Microbiol, 1997 Feb, 35(2), 455 - 61
Concordance of Porphyromonas gingivalis colonization in families; Tuite-McDonnell M et al.; Periodontitis is a widespread disease that appears to be due to a specific bacterial infection . Several species of bacteria have been investigated as potential pathogens, and particularly strong evidence links the presence of Porphyromonas gingivalis with indicators of periodontitis . Information concerning the transmission of P . gingivalis between human contacts may be important in determining risk factors for disease and developing preventive strategies . A few small studies have provided some evidence of transmission between related individuals, but no large-scale study of families that would reflect the typical transmission of this pathogen in the population has been reported . The purpose of this study was to investigate the transmission of P . gingivalis within randomly selected, extended families . The colonization status of 564 members of multigeneration families was determined, and the degree of concordance observed among members of these families was then compared to that expected to occur based on the prevalence of colonization in the population studied . A PCR assay was used for detection of P . gingivalis . Concordance in colonization was more frequently observed within entire families (P = 0.0000) and for spouses (P < 0.001), children and their mothers (P < 0.001), children and their fathers (P < 0.01), adults and their mothers (P < 0.005), and siblings (P < 0.05) than would be expected if P . gingivalis were randomly distributed in the population studied . Results showed that contact with an infected family member substantially increased the relative risk of colonization in these intrafamilial pairs . This indicates that P . gingivalis is commonly transmitted by contact with an infected family member.

Endocrinology, 1997 Feb, 138(2), 602 - 6
Thyroid-stimulating hormone-induced down-regulation of thyroid transcription factor 1 in rat thyroid FRTL-5 cells; Saito T et al.; Thyroid transcription factor 1 (TTF-1) is thought to play an important role in the expression of genes that encode thyroid-specific proteins such as thyroglobulin, thyroid peroxidase, and TSH-receptor . The role of TSH in the regulation of TTF-1 messenger RNA (mRNA) and protein abundance was investigated in rat thyroid FRTL-5 cells . Northern blot analysis revealed that TSH reduced TTF-1 mRNA abundance in a dose- and time-dependent manner . Immunoblot analysis with rabbit antibodies prepared against a recombinant fragment of TTF-1 expressed in bacteria showed that TSH also reduced the amount of TTF-1 protein in FRTL-5 cells . Whereas the effect of TSH on TTF-1 mRNA was apparent after 3 h, the effect on TTF-1 protein was not apparent until 12 h after TSH addition to the cells . Both TTF-1 mRNA and protein were significantly decreased after the addition of (Bu)2 cAMP or forskolin for 24 h, whereas they were not decreased by 12-O-tetradecanoyl-phorbol-13-acetate . These results indicate that TSH down-regulates TTF-1 expression in FRTL-5 cells via the cAMP pathway.

J Virol, 1997 Feb, 71(2), 1547 - 57
Physical and functional interactions between herpes simplex virus immediate-early proteins ICP4 and ICP27; Panagiotidis CA et al.; The ordered expression of herpes simplex virus type 1 (HSV-1) genes, during the course of a productive infection, requires the action of the virus immediate-early regulatory proteins . Using a protein interaction assay, we demonstrate specific in vitro protein-protein interactions between ICP4 and ICP27, two immediate-early proteins of HSV-1 that are essential for virus replication . We map multiple points of contact between these proteins . Furthermore, by coimmunoprecipitation experiments, we demonstrate the following . (i) ICP4-ICP27 complexes are present in extracts from HSV-1 infected cells . (ii) ICP27 binds preferentially to less modified forms of ICP4, a protein that is extensively modified posttranslationally . We also demonstrate, by performing electrophoretic mobility shift assays and supershifts with monoclonal antibodies to ICP4 or ICP27, that both proteins are present in a DNA-protein complex with a noncanonical ICP4 binding site present in the HSV thymidine kinase (TK) gene . ICP4, in extracts from cells infected with ICP27-deficient viruses, is impaired in its ability to form complexes with the TK site but not with the canonical site from the alpha4 gene . However, ICP4 is able to form complexes with the TK probe, in the absence of ICP27, when overproduced in mammalian cells or expressed in bacteria . These data suggest that the inability of ICP4 from infected cell extracts to bind the TK probe in the absence of ICP27 does not reflect a requirement for the physical presence of ICP27 in the complex . Rather, they imply that ICP27 is likely to modulate the DNA binding activity of ICP4 by affecting its posttranslational modification status . Therefore, we propose that ICP27, in addition to its established role as a posttranscriptional regulator of virus gene expression, may also modulate transcription either through direct or indirect interactions with HSV regulatory regions, or through its ability to modulate the DNA binding activity of ICP4.

J Biol Chem, 1997 Jan 31, 272(5), 2936 - 41
Cloning and expression of a novel mammalian thioredoxin; Spyrou G et al.; We have isolated a 1276-base pair cDNA from a rat heart cDNA library that encodes a novel thioredoxin (Trx2) of 166 amino acid residues with a calculated molecular mass of 18.2 kDa . Trx2 possesses the conserved thioredoxin-active site, Trp-Cys-Gly-Pro-Cys, but lacks structural cysteines present in all mammalian thioredoxins . Trx2 also differs from the previously described rat thioredoxin (Trx1) by the presence of a 60-amino acid extension at the N terminus . This extension has properties characteristic for a mitochondrial translocation signal, and the cleavage at a putative mitochondrial peptidase cleavage site would give a mature protein of 12.2 kDa . Western blot analysis from cytosolic, peroxisomal, and mitochondrial rat liver cell fractions confirmed mitochondrial localization of Trx2 . Northern blot and reverse transcriptase-polymerase chain reaction analyses revealed that Trx2 hybridized to a 1.3-kilobase message, and it was expressed in several tissues with the highest expression levels in heart, muscle, kidney, and adrenal gland . N-terminally truncated recombinant protein was expressed in bacteria and characterized biochemically . Trx2 possessed a dithiol-reducing enzymatic activity and, with mammalian thioredoxin reductase and NADPH, was able to reduce the interchain disulfide bridges of insulin . Furthermore, Trx2 was more resistant to oxidation than Trx1.

J Biol Chem, 1997 Jan 31, 272(5), 2914 - 9
Identification of the region in actin-binding protein that binds to the cytoplasmic domain of glycoprotein IBalpha; Meyer SC et al.; Actin-binding protein (ABP-280) is a component of the submembranous cytoskeleton and interacts with the glycoprotein (GP) Ibalpha subunit of the GP Ib-IX complex in platelets . In the present studies, we have identified the binding site for GP Ibalpha in ABP-280 . A melanoma cell line lacking ABP-280 was stably transfected with the cDNAs coding for GP Ib-IX, then transiently transfected with cDNA coding for various carboxyl-truncates of ABP-280 . Immunocapture assays and co-immunoprecipitation experiments from detergent-lysed cells showed that deletion of the carboxyl-terminal repeats 20-24 of ABP-280 had no effect on GP Ib-IX binding, but deletion of residues 2099 through 2136 within repeat 19 abolished binding . In the yeast two-hybrid system, an ABP-280 fragment comprising repeats 17-19 bound GP Ibalpha . Deletion from either end abolished binding . Individual or multiple repeats of ABP-280 were expressed as fusion protein in bacteria and purified; structural folding was evaluated, and binding to GP Ib-IX was assessed . Binding depended on the presence of repeats 17-19 . None of the individual repeats were able to bind to GP Ib-IX . These findings demonstrate that residues 1850-2136 comprising repeats 17-19 contain the binding site for GP Ib-IX.

Fortschr Med, 1997 Jan 30, 115(3), 35 - 6
{Flies--reservoirs and vectors of Helicobacter pylori}; Grubel P et al.; The route by which humans get infected with H.p . is unknown . Viable bacteria have been shown to be excreted in human feces from infected individuals . We have recently shown that flies are able to carry and disseminate viable H . p . with their excreta, when they have fed on H.p . We propose a hypothesis that H . p . is acquired from human feces by flies, which then, while crawling on human food, contaminate it with their intestinal excreta . The food, swallowed by a susceptible individual then leads to a new infection . Flies should be considered prime suspects for the transmission of H . p., particularly in developing countries, where sanitary and domestic facilities are poor.

FEBS Lett, 1997 Jan 27, 402(1), 50 - 2
Expression and activity of a recombinant chimeric protein composed of pokeweed antiviral protein and of human interleukin-2; Dore JM et al.; The pokeweed antiviral protein (PAP) has already been used to chemically construct immunotoxins . Here we tested the recombinant approach for the production of PAP-containing cytotoxic fusion-proteins . A cDNA encoding a mutated PAP (PAP9), which is expressed at high levels in bacteria, was fused to human interleukin-2 (IL-2) cDNA . The resulting PAP9-IL-2 protein was as active as the free PAP9 in inhibiting an eukaryotic cell-free translation system . Only the chimeric protein desaminated the 28S rRNA and inhibited translation of the CTLL-2 cell line which expresses the IL-2 receptor . These results show that PAP is a suitable toxin for the production of recombinant immunotoxins.

J Biol Chem, 1997 Jan 24, 272(4), 2129 - 35
Evaluation of secondary structure of OxlT, the oxalate transporter of Oxalobacter formigenes, by circular dichroism spectroscopy; Fu D et al.; OxlT, the oxalate/formate exchange transporter of Oxalobacter formigenes, was purified as a histidine-tagged variant, OxlTHis, using Ni2+-linked affinity chromatography . OxlTHis was readily obtained in high purity (>/=95%) and reasonable yield (>/=60%), and showed kinetic and biochemical features characteristic of its parent, OxlT, including an unusually high maximal velocity (60 micromol/min per mg of protein at 4 degrees C) . Circular dichroism spectroscopy of purified OxlTHis identified the alpha-helix as its dominant secondary structural unit, encompassing 60-70% of OxlTHis residues and consistent with a model suggesting 60% of OxlT (OxlTHis) residues are involved in the construction of 12 transmembrane alpha-helices (Abe, K., Ruan, Z.-S., and Maloney, P . C . (1996) J . Biol . Chem . 271, 6789-6793) . In either octyl glucoside/lipid or dodecylmaltoside/lipid micelles, solubilized OxlTHis showed a striking substrate-induced stabilization of function, and at saturating levels of substrate (1000 x KD) activity recoverable by reconstitution disappeared with a half-life of 7 days at 23 degrees C . Measurement of changes of ellipticity at 222 nm as a function of time and substrate concentration showed that maintenance of function was attributable to a substrate-induced stabilization of the alpha-helical ensemble with a KD of 10 microM for the 1:1 binding of oxalate to OxlTHis.

Science, 1997 Jan 24, 275(5299), 515 - 8
An Fe2IVO2 diamond core structure for the key intermediate Q of methane monooxygenase; Shu L et al.; A new paradigm for oxygen activation is required for enzymes such as methane monooxygenase (MMO), for which catalysis depends on a nonheme diiron center instead of the more familiar Fe-porphyrin cofactor . On the basis of precedents from synthetic diiron complexes, a high-valent Fe2(micro-O)2 diamond core has been proposed as the key oxidizing species for MMO and other nonheme diiron enzymes such as ribonucleotide reductase and fatty acid desaturase . The presence of a single short Fe-O bond (1.77 angstroms) per Fe atom and an Fe-Fe distance of 2.46 angstroms in MMO reaction intermediate Q, obtained from extended x-ray absorption fine structure and Mossbauer analysis, provides spectroscopic evidence that the diiron center in Q has an Fe2IVO2 diamond core.

Biochemistry, 1997 Jan 21, 36(3), 489 - 94
Three-dimensional structure of tetrahydrodipicolinate N-succinyltransferase; Beaman TW et al.; The conversion of tetrahydrodipicolinate and succinyl-CoA to N-succinyltetrahydrodipicolinate and CoA is catalyzed by tetrahydrodipicolinate N-succinyltransferase and is the committed step in the succinylase pathway by which bacteria synthesize L-lysine and meso-diaminopimelate, a component of peptidoglycan . The X-ray crystal structure of THDP succinyltransferase has been determined to 2.2 A resolution and has been refined to a crystallographic R-factor of 17.0% . The enzyme is trimeric and displays the left-handed parallel beta-helix (L beta H) structural motif encoded by the "hexapeptide repeat" amino acid sequence motif {Raetz, C.R.H., & Roderick, S.L . (1995) Science 270, 997-1000} . The approximate location of the active site of THDP succinyltransferase is suggested by the proximity of binding sites for two inhibitors: p-(chloromercuri)benzenesulfonic acid and cobalt ion, both of which bind to the L beta H domain.

J Exp Med, 1997 Jan 20, 185(2), 317 - 28
Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures; Winzler C et al.; The signals controlling the checkpoints of dendritic cells (DC) maturation and the correlation between phenotypical and functional maturational stages were investigated in a defined model system of growth factor-dependent immature mouse DC . Three sequential stages of DC maturation (immature, mature, and apoptotic) were defined and characterized . Immature DC (stage 1) had low expression of costimulatory molecules, highly organized cytoskeleton, focal adhesion plaques, and slow motility; accordingly, they were very efficient in antigen uptake and processing of soluble proteins . Further, at this stage most of major histocompatibility complex class II molecules were within cytoplasmic compartments consistent with a poor allostimulatory capacity . Bacteria or cytokines were very efficient in inducing progression from stage 1 towards stage 2 (mature) . Morphological changes were observed by confocal analysis including depolymerization of F-actin and loss of vinculin containing adhesive structures which correlates with acquisition of high motility . Antigen uptake and presentation of native protein antigen was reduced . In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation . This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed.

Gene, 1997 Jan 15, 184(2), 291 - 8
Physical and genetic map of the chromosome of Dichelobacter nodosus strain A198; La Fontaine S et al.; A physical map of the chromosome of Dichelobacter nodosus strain A198 was constructed using the restriction endonucleases EagI and StuI . Mapping data indicated the presence of a single, circular chromosome of 1.54 Mb . The three rRNA operons and the virulence related locus (vrl) were precisely positioned at the junctions of EagI and StuI fragments, and their transcriptional orientations were also determined . Other D . nodosus genes were assigned to specific EagI and StuI fragments . Analysis of the resultant map revealed that the putative virulence genes were not clustered on the chromosome which suggests that the D . nodosus virulence determinants have been acquired gradually and that virulence in D . nodosus is an evolving trait.

Eur J Biochem, 1997 Jan 15, 243(1-2), 474 - 81
Assignment of the ligand geometry and redox potentials of the trihaem ferricytochrome c3 from Desulfuromonas acetoxidans; Turner DL et al.; Cytochrome c551.5 is a trihaem cytochrome of the cytochrome c3 family isolated from Desulfuromonas acetoxidans . Although several X-ray structures are available for tetrahaem cytochromes of this family, there is no X-ray structure for trihaem cytochromes . Cytochrome C551.5 was studied in the oxidized form by means of two-dimensional NMR . The pattern of observed interhaem NOESY connectivities is in agreement with the haem core structure previously determined by NMR for the reduced protein {Coutinho, I . B., Turner, D . L., Liu, M . Y., LeGall, J . & Xavier, A . V . (1996) J . Biol . Inorg . Chem . 1, 305-311} . The similarities found between the haem core structure and the amino acid sequence of cytochrome c551.5 and those of tetrahaem cytochromes c3 allows each of the haems to be specifically assigned in the polypeptide sequence, and the attribution of the midpoint redox potentials to the individual haems . This also allows individual redox potentials to be assigned to each haem in the NMR spectrum . The paramagnetic shifts of the 13C resonances of the haem substituents were analyzed in terms of pi molecular orbitals with perturbed D4h symmetry . The parameters of this analysis have been shown to be controlled by the orientation of the axial ligands in several other bis-His-coordinated haems and hence the ligand geometry was deduced for cytochrome C551.5 . The structural analogy between the relative haem plane orientations in cytochrome c551.5 and the tetrahaem cytochromes c3 is found to extend to the axial ligands with the largest differences being in the vicinity of the deleted fourth haem, using the numbering of cytochrome c3 haems.

Eur J Biochem, 1997 Jan 15, 243(1-2), 393 - 9
An ultracentrifugal approach to quantitative characterization of the molecular assembly of a physiological electron-transfer complex: the interaction of electron-transferring flavoprotein with trimethylamine dehydrogenase; Wilson EK et al.; The interaction between two physiological redox partners, trimethylamine dehydrogenase and electron-transferring flavoprotein, has been characterized quantitatively by analytical ultracentrifugation at 4 degrees C . Analysis of sedimentation-equilibrium distributions obtained at 15 000 rpm for mixtures in 10 mM potassium phosphate, pH 7.5, by means of the psi function {Wills, P . R., Jacobsen, M . P . & Winzor, D . J . (1996) Biopolymers 38, 119-130} has yielded an intrinsic dissociation constant of 3-7 microM for the interaction of electron-transferring flavoprotein with two equivalent and independent sites on the homodimeric enzyme . This investigation indicates the potential of sedimentation equilibrium for the quantitative characterization of interactions between dissimilar macromolecules.

Eur J Biochem, 1997 Jan 15, 243(1-2), 209 - 18
Identification and structure of activated-platelet protein-1, a protein with RNA-binding domain motifs that is expressed by activated platelets; Houng AK et al.; Beyond their critical role in thrombosis, platelets perform important functions in vascular remodeling, inflammation, and wound repair . Many of these functions are executed by molecules expressed by activated platelets . A novel molecule, activated-platelet protein-1 (APP-1), was identified by a monoclonal antibody against activated rabbit platelets . When platelets were stimulated by thrombin, A23187 or ADP, APP-I was expressed on the platelet surface . APP-1 was also detected in whole cell lysates of platelets, but not on the external surfaces of resting platelets . With maximal activation by thrombin, 15 900 +/- 2800 molecules APP-1 were expressed/platelet . A 2.3-kb cDNA fragment containing a partial coding sequence for APP-1 was isolated from a rabbit bone marrow library by expression cloning with the anti-APP-1 monoclonal antibody . When expressed as a recombinant fusion protein in bacteria, APP-1 bound specifically to poly(A)-Sepharose . The full-length cDNA coding for human APP-1, obtained by DNA hybridization techniques, showed 98.7% amino acid sequence identity with the rabbit protein . Northern analysis with human APP-1 identified a 3.7-kb mRNA transcript in megakaryocytic lines that express transcripts for platelet proteins . Human APP-1 has four ribonucleotide binding domains with ribonucleoprotein 1 and 2 motifs . By virtue of its ribonucleotide binding domains, APP-1 is structurally related to polyadenylate-binding protein, which regulates translation initiation and polyadenylate shortening, and to nucleolysin, a specific effector molecule found in the granules of cytotoxic T lymphocytes.

EMBO J, 1997 Jan 15, 16(2), 417 - 29
Pop3p is essential for the activity of the RNase MRP and RNase P ribonucleoproteins in vivo; Dichtl B et al.; RNase MRP is a ribonucleoprotein (RNP) particle which is involved in the processing of pre-rRNA at site A3 in internal transcribed spacer 1 . Although RNase MRP has been analysed functionally, the structure and composition of the particle are not well characterized . A genetic screen for mutants which are synthetically lethal (sl) with a temperature-sensitive (ts) mutation in the RNA component of RNase MRP (rrp2-1) identified an essential gene, POP3, which encodes a basic protein of 22.6 kDa predicted molecular weight . Over-expression of Pop3p fully suppresses the ts growth phenotype of the rrp2-1 allele at 34 degrees C and gives partial suppression at 37 degrees C . Depletion of Pop3p in vivo results in a phenotype characteristic of the loss of RNase MRP activity; A3 cleavage is inhibited, leading to under-accumulation of the short form of the 5.8S rRNA (5.8S(S)) and formation of an aberrant 5.8S rRNA precursor which is 5'-extended to site A2 . Pop3p depletion also inhibits pre-tRNA processing; tRNA primary transcripts accumulate, as well as spliced but 5'- and 3'-unprocessed pre-tRNAs . The Pop3p depletion phenotype resembles those previously described for mutations in components of RNase MRP and RNase P (rrp2-1, rpr1-1 and pop1-1) . Immunoprecipitation of epitope-tagged Pop3p co-precipitates the RNA components of both RNase MRP and RNase P . Pop3p is, therefore, a common component of both RNPs and is required for their enzymatic functions in vivo . The ubiquitous RNase P RNP, which has a single protein component in Bacteria and Archaea, requires at least two protein subunits for its function in eukaryotic cells.

J Am Vet Med Assoc, 1997 Jan 15, 210(2), 235 - 9
Pulmonary lobectomy in the management of pneumonia in dogs: 59 cases (1972-1994); Murphy ST et al.; OBJECTIVE: To evaluate the risk and efficacy of pulmonary lobectomy in dogs with pneumonia . DESIGN: Retrospective study . ANIMALS: 59 dogs with pneumonia . PROCEDURE: Review of medical records and telephone conversations . RESULTS: 54.2% of dogs had resolution of pneumonia after lobectomy, 20.3% died in the perioperative period, and 25.4% survived the perioperative period but pneumonia did not resolve . Pneumonia was caused by bacteria (25 dogs), fungi (12), foreign bodies (8), parasites (1), viruses (1), and allergies (1) . In 11 dogs, the etiologic agent was not isolated . Bacterial or fungal pneumonias were significantly less likely to resolve compared with foreign body pneumonia or when an etiologic agent was not isolated . Perioperative mortality rate increased significantly with an increase in number of pulmonary lobes removed . Complications during surgery significantly increased perioperative mortality rate . Surgical era (1972 to 1983 vs 1984 to 1994) was a significant predictor of mortality, with the odds of dying in the perioperative period being 11 times greater between 1972 to 1983 . The odds of failure to resolve pneumonia was 3 times greater during 1972 to 1983 . CLINICAL IMPLICATIONS: Number of pulmonary lobes removed and complications during surgery significantly affect perioperative mortality rate . Identification of etiologic agents may help in predicting dogs likely to resolve pneumonia after surgery.

Biochem Biophys Res Commun, 1997 Jan 13, 230(2), 242 - 6
Characterization of scFv-421, a single-chain antibody targeted to p53; Jannot CB et al.; A gene encoding a single-chain antibody (scFv) which specifically binds the tumor suppressor protein p53 has been constructed from RNA of hybridoma cells producing Pab 421 . scFv-421 which was expressed and purified from bacteria specifically binds p53 . scFv-421, as well as the previously described scFv-FRP5 and -R1R (1), were expressed intracellularly in mammalian cells and targeted to different subcellular locations, including the nucleus, cytoplasm, and endoplasmic reticulum (ER) . High levels of all ER targeted scFv proteins, but not nuclear or cytoplasmic targeted proteins, were found in transfected COS-1 cells . In an attempt to stabilize the proteins, sequences encoding the mouse immunoglobin CK constant domain were added to each scFv construct . This led to a moderate increase in the cytoplasmic expression of scFv-FRP5.

Biochim Biophys Acta, 1997 Jan 10, 1355(1), 50 - 60
Re-investigation of glucose metabolism in Fibrobacter succinogenes, using NMR spectroscopy and enzymatic assays . Evidence for pentose phosphates phosphoketolase and pyruvate formate lyase activities; Matheron C et al.; The glucose metabolism of Fibrobacter succinogenes S85 was studied in detail; key intermediates and alternative pathways were evidenced by NMR and/or enzymatic assays . A high phosphoketolase activity was detected in four strains of Fibrobacter under strictly anaerobic conditions, with ribose-5-phosphate as substrate, no activity was evidenced with fructose-6-phosphate . This is the first report of a pentose phosphates phosphoketolase in bacteria unable to use pentoses . In contrast, the Entner-Doudoroff pathway and the oxidative branch of the pentose phosphate pathway could not be evidenced . Incubation of living cells of F . succinogenes with Na2(13)CO3 confirmed the incorporation of 13CO2 in the carboxylic group of succinate . The presence of fumarase was evidenced by in vivo 4C-NMR using 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO): the enzyme showed a high reversibility under physiological conditions . The production of formate from glucose catabolism was evidenced by enzymatic assay and by NMR and a pyruvate formate lyase activity was detected using strictly anaerobic conditions.

Biochemistry, 1997 Jan 7, 36(1), 41 - 8
An exposed tyrosine on the surface of trimethylamine dehydrogenase facilitates electron transfer to electron transferring flavoprotein: kinetics of transfer in wild-type and mutant complexes; Wilson EK et al.; In wild-type trimethylamine dehydrogenase, tyrosine-442 is located at the center of a concave region on the surface of the enzyme that is proposed to form the docking site for the physiological redox acceptor, electron transferring flavoprotein . The intrinsic rate constant for electron transfer in the reoxidation of one-electron dithionite-reduced wild-type trimethylamine dehydrogenase (modified with phenylhydrazine) by electron transferring flavoprotein was investigated by stopped-flow spectroscopy . Analysis of the temperature dependence of the reaction rate by electron transfer theory yielded values for the reorganizational energy of 1.4 eV and the electronic coupling matrix element of 0.82 cm-1 . The role played by residue Tyr-442 in facilitating reduction of ETF by TMADH was investigated by isolating three mutant forms of the enzyme in which Tyr-442 was exchanged for a phenylalanine, leucine, or glycine residue . Rates of electron transfer from these mutants of TMADH to ETF were investigated by stopped-flow spectroscopy . At 25 degrees C, modest reductions in rate were observed for the Y442F (1.4-fold) and Y442L (2.2-fold) mutant complexes, but a substantial decrease in rate (30.5-fold) and an elevated dissociation constant for the complex were seen for the Y442G mutant enzyme . Inspection of the crystal structure of wild-type TMADH reveals that Tyr-442 is positioned along one side of a small cavity on the surface of the enzyme: Val 344, located at the bottom of this cavity, is the closest surface residue to the 4Fe-4S center of TMADH and is likely to be positioned on a major electron transfer pathway to ETF . The reduced electron transfer rates in the mutant complexes are probably brought about by decreases in electronic coupling between the electron transfer donor and acceptor within the complex, either directly or indirectly due to unfavorable change in the orientation of the two proteins with respect to one another.

Biochemistry, 1997 Jan 7, 36(1), 8 - 14
Active site mutants implicate key residues for control of color and light cycle kinetics of photoactive yellow protein; Genick UK et al.; To understand how the protein and chromophore components of a light-sensing protein interact to create a light cycle, we performed time-resolved spectroscopy on site-directed mutants of photoactive yellow protein (PYP) . Recently determined crystallographic structures of PYP in the ground and colorless I2 states allowed us to design mutants and to study their photosensing properties at the atomic level . We developed a system for rapid mutagenesis and heterologous bacterial expression for PYP apoprotein and generated holoprotein through formation of a covalent thioester linkage with the p-hydroxycinnamic acid chromophore as found in the native protein . Glu46, replaced by Gln, is buried in the active site and hydrogen bonds to the chromophore's phenolate oxygen in the ground state . The Glu46Gln mutation shifted the ground state absorption maximum from 446 to 462 nm, indicating that the color of PYP can be fine-tuned by the alteration of hydrogen bonds . Arg52, which separates the active site from solvent in the ground state, was substituted by Ala . The smaller red shift (to 452 nm) of the Arg52Ala mutant suggests that electrostatic interactions with Arg52 are not important for charge stabilization on the chromophore . Both mutations cause interesting changes in light cycle kinetics . The most dramatic effect is a 700-fold increase in the rate of recovery to the ground state of Glu46Gln PYP in response to a change in pH from pH 5 to 10 (pKa = 8) . Prompted by this large effect, we conducted a careful reexamination of pH effects on the wild-type PYP light cycle . The rate of color loss decreased about 3-fold with increasing pH from pH 5 to 10 . The rate of recovery to the colored ground state showed a bell-shaped pH dependence, controlled by two pKa values (6.4 and 9.4) . The maximum recovery rate at pH 7.9 is about 16 times faster than at pH 5 . The effect of pH on Arg52Ala is like that on wild type except for faster loss of color and slower recovery . These kinetic effects of the mutations and the changes with pH demonstrate that both phases in PYP's light cycle are actively controlled by the protein component.

Biochem Biophys Res Commun, 1997 Jan 3, 230(1), 30 - 4
The formate dehydrogenase isolated from the aerobe Methylobacterium sp . RXM is a molybdenum-containing protein; Duarte RO et al.; The formate dehydrogenase (FDH) isolated from cells of Methylobacterium sp . RXM grown on molybdenum-containing mineral medium using methanol as carbon source, was partially purified (at least 90% pure as revealed by SDS-PAGE) . The enzyme is unstable under oxygen and all the purification steps were conducted under strict anaerobic conditions . The molecular mass is 75 kDa (gel exclusion 300 kDa) . The enzyme was characterized in terms of the kinetic parameters towards different substrates and electron acceptors, pH and temperature dependence and the effect of a wide range of compounds in the enzymatic activity . The EPR spectra of the dithionite reduced sample show, at low temperature (below 20 K), two rhombic EPR signals due to two distinct {Fe-S} centres (centre I at g-values 2.023, 1.951 and 1.933, and centre II at g-values 2.054 and 1.913) . At high temperature (around 100 K) another rhombic EPR signal is optimally observed at g-values 2.002, 1.987 and 1.959 and attributed to the molybdenum site . The EPR signals assigned to the iron-sulfur centres show a strong analogy with the aldehyde oxido-reductase from Desulfovibrio gigas known to contain a Mo-pterin and two {2Fe-2S} centres and whose crystallographic structure was recently resolved.

Gene, 1997 Jan 3, 184(1), 121 - 3
A natural longer glycine-rich region in IKe filamentous phage confers no selective advantage; Bruno R et al.; Filamentous phage infect bacteria bearing pili . The phage protein involved in the recognition of pili and subsequent penetration of the phage into bacteria is the gene 3 protein (g3p) . This is a multi-domain protein with glycine-rich regions separating some of the domains . Here we have found an insertion within the glycine-rich domain of the g3p of IKe, a filamentous phage which infects bacteria bearing N pili . Although this insertion considerably increases the length of the glycine-rich domain it has no selective advantage or disadvantage in infection or production of phage, and can therefore be considered a neutral mutation.

Biochim Biophys Acta, 1997 Jan 3, 1350(1), 75 - 9
Compilation of ribosomal 5S ribonucleic acid nucleotide sequences: eukaryotic 5S rRNAs; Szymanski M et al.; 5S Ribosomal RNA is the smallest RNA component of the ribosomes . Due to relatively simple isolation and sequencing procedures as well as a potential use of the sequence data in evolutionary analyses, the amount of known nucleotide sequences on both RNA and DNA levels was rapidly growing . In this paper we present the updated (March 1996) compilation of eukaryotic 5D rRNA and 5S rDNA sequences.

FEBS Lett, 1997 Jan 3, 400(2), 243 - 6
Complementation of a yeast delta pkc1 mutant by the Arabidopsis proteinANT; Vergani P et al.; The Saccharomyces cerevisiae protein kinase C homologue, PKC1, is involved in maintenance of cell integrity during polarized growth . We have used a mutant complementation approach to investigate related signal transduction pathways in higher plants . Here we report the isolation of a cDNA from Arabidopsis thaliana which partially suppresses the lytic defect of a delta pkc1 yeast strain . The encoded protein, ANT, belongs to the AP2-related gene family and is essential for ovule development . Expression in yeast of a LexA-ANT fusion protein activates transcription of a reporter gene from promoters containing lexA operators . Our results support the idea that ANT acts as transcriptional activator in planta.

J Biol Chem, 1997 Jan 3, 272(1), 580 - 5
Membrane topology of the transposon 10-encoded metal-tetracycline/H+ antiporter as studied by site-directed chemical labeling; Kimura T et al.; The transposon (Tn) 10-encoded metal-tetracycline/H+ antiporter (Tn10-TetA) is predicted to have a membrane topology involving 12 transmembrane domains on the basis of the hydropathy profile of its sequence and the results of limited proteolysis; however, the experimental results of limited proteolysis are not enough to confirm the topology because proteases cannot gain access from the periplasmic side (Eckert, B., and Beck, C . F . (1989) J . Biol . Chem . 264, 11663-11670) . One or two cysteine residues were introduced into each predicted hydrophilic loop or the N-terminal segment of Tn10-TetA by site-directed mutagenesis, and then the topology of the protein was determined by examining whether labeling of the introduced Cys residue by membrane-permeant {14C}N-ethylmaleimide ({14C}NEM) was prevented by preincubation of intact cells with the membrane-impermeant maleimide, 4-acetamido-4'-maleimidylstilbene-2,2'-disulfonic acid (AMS) . The binding of {14C}NEM to the S36C (loop 1-2), L97C (loop 3-4), S156C (loop 5-6), R238C (loop 7-8), S296C (loop 9-10), Y357C, and D365C (loop 10-11) mutants was completely blocked by pretreatment with AMS, indicating that these residues are located on the periplasmic surface . In contrast, {14C}NEM binding to the S4C (N-terminal segment), S65C (loop 2-3), D120C (loop 4-5), S199C and S201C (loop 6-7), T270C (loop 8-9), and S328C (loop 10-11) mutants was not affected by pretreatment with AMS, indicating that these residues are on the cytoplasmic surface . These results for the first time thoroughly confirm the 12-transmembrane topology of the metal-tetracycline/H+ antiporter.

J Biol Chem, 1997 Jan 3, 272(1), 146 - 53
Effect of dimerization on signal transduction and biological function of oncogenic Ros, insulin, and insulin-like growth factor I receptors; Chan JL et al.; The avian sarcoma virus UR2 codes for an oncogenic Gag-Ros fusion protein-tyrosine kinase (PTK) . We have previously derived two retroviruses, T6 and NM1, coding for oncogenic Gag-insulin receptor and Gag-insulin-like growth factor I receptor (IGFR) fusion proteins, respectively . The Gag-IGFR fusion protein dimerizes, whereas Gag-Ros does not . To identify sequences affecting dimerization and the effect of dimerization on signaling and biological functions, we generated recombinants exchanging the extracellular and transmembrane sequences among the three fusion receptors . The presence of multiple cysteines in the Gag sequence appears to preclude dimerization, since deletion of the 3' cysteine residue allows for dimerization . Most of the chimeric receptors retain high PTK activity and induce transformation regardless of their configuration on the cell surface . UT, a UR2/T6 chimera, retained mitogenic activity but has a markedly reduced transforming ability, while UN7, a UR2/NM1 recombinant, which also harbors Y950F and F951S mutations in IGFR, exhibits dramatic reductions in both activities . All of the fusion receptors can phosphorylate insulin receptor substrate 1 and activate PI 3-kinase . UT protein induces Shc phosphorylation, whereas UN7 protein does not, but both are unable to activate mitogen-activated protein kinase . Our results show that overexpressed oncogenic Gag-fusion receptors do not require dimerization for their signaling and transforming functions and that the extracellular and transmembrane sequences of a receptor PTK can affect its specific substrate interactions.

EMBO J, 1997 Jan 2, 16(1), 35 - 43
Ezrin is a cyclic AMP-dependent protein kinase anchoring protein; Dransfield DT et al.; cAMP-dependent protein kinase (A-kinase) anchoring proteins (AKAPs) are responsible for the subcellular sequestration of the type II A-kinase . Previously, we identified a 78 kDa AKAP which was enriched in gastric parietal cells . We have now purified the 78 kDa AKAP to homogeneity from gastric fundic mucosal supernates using type II A-kinase regulatory subunit (RII) affinity chromatography . The purified 78 kDa AKAP was recognized by monoclonal antibodies against ezrin, the canalicular actin-associated protein . Recombinant ezrin produced in either Sf9 cells or bacteria also bound RII . Recombinant radixin and moesin, ezrin-related proteins, also bound RII in blot overlay . Analysis of recombinant truncations of ezrin mapped the RII binding site to a region between amino acids 373 and 439 . This region contained a 14-amino-acid amphipathic alpha-helical putative RII binding region . A synthetic peptide containing the amphipathic helical region (ezrin409-438) blocked RII binding to ezrin, but a peptide with a leucine to proline substitution at amino acid 421 failed to inhibit RII binding . In mouse fundic mucosa, RII immunoreactivity redistributed from a predominantly cytosolic location in resting parietal cells, to a canalicular pattern in mucosa from animals stimulated with gastrin . These results demonstrate that ezrin is a major AKAP in gastric parietal cells and may function to tether type II A-kinase to a region near the secretory canaliculus.

J Exp Biol, 1997, 200(Pt 21), 2797 - 805
The effects of exposure to ammonia on ammonia and taurine pools of the symbiotic clam
Lee R, Childress J, Desaulniers N.
The nutrition of the gutless clam Solemyareidi is supported by the activity of intracellular chemoautotrophic bacteria housed in its gill filaments . Ammonia (the sum of NH3 and NH4+) is utilized as a nitrogen source by the association and is abundant in the clam's environment . In the present study, clams were exposed to 0.011.3mmoll-1 ammonia for 2223h in the presence of thiosulfate as a sulfur substrate . Ammonia exposure increased the ammonia concentration in the tissue pools of the gill, foot and visceral mass from 0.5 to 2µmolg-1wetmass, without added ammonia, to as much as 12µmolg-1wetmass in the presence of 0.7 and 1.3mmoll-1 external ammonia . Gill tissue ammonia concentrations were consistently higher than those in the foot and visceral mass . The elevation of tissue ammonia concentration compared with the medium may be due in part to an ammonia trapping mechanism resulting from a lower intracellular pH compared with sea water and greater permeability to NH3 compared with NH4+ . Rates of ammonia incorporation into organic matter (assimilation) were determined using 15N as a tracer . 15N-labeled ammonia assimilation was higher in gill than in foot and increased as a function of 15N-labeled ammonia concentration in the medium . The size of the free amino acid (FAA) pool in the gill also increased as a function of ammonia concentration in the medium . This entire increase was accounted for by a single amino acid, taurine, which was the predominant FAA in both gill and foot tissue . Aspartate, glutamate, arginine and alanine were also abundant but their levels were not influenced by external ammonia concentration . Ammonia assimilation appeared to occur at rates sufficient to account for the observed increase in taurine level . These findings suggest that taurine is a major product of ammonia assimilation.

J Exp Biol, 1997, 200(Pt 5), 883 - 96
Inorganic carbon acquisition by the hydrothermal vent tubeworm Riftia pachyptila depends upon high external PCO2 and upon proton-equivalent ion transport by the worm
Goffredi S, Childress J, Desaulniers N, Lee R, Lallier F, Hammond D.
Riftia pachyptila is the most conspicuous organism living at deep sea hydrothermal vents along the East Pacific Rise . To support its large size and high growth rates, this invertebrate relies exclusively upon internal chemosynthetic bacterial symbionts . The animal must supply inorganic carbon at high rates to the bacteria, which are far removed from the external medium . We found substantial differences in body fluid total inorganic carbon (CO2) both within and between vent sites when comparing freshly captured worms from a variety of places . However, the primary influence on body fluid CO2 was the chemical characteristics of the site from which the worms were collected . Studies on tubeworms, both freshly captured and maintained in captivity, demonstrate that the acquisition of inorganic carbon is apparently limited by the availability of CO2, as opposed to bicarbonate, and thus appears to be accomplished via diffusion of CO2 into the plume, rather than by mediated transport of bicarbonate . The greatly elevated PCO2 measured at the vent sites (up to 12.6 kPa around the tubeworms), which is a result of low environmental pH (as low as 5.6 around the tubeworms), and elevated CO2 (as high as 7.1 mmol l-1 around the tubes) speeds this diffusion . Moreover, despite large and variable amounts of internal CO2, these worms maintain their extracellular fluid pH stable, and alkaline, in comparison with the environment . The maintenance of this alkaline pH acts to concentrate inorganic carbon into extracellular fluids . Exposure to N-ethylmaleimide, a non-specific H+-ATPase inhibitor, appeared to stop this process, resulting in a decline in extracellular pH and CO2 . We hypothesize that the worms maintain their extracellular pH by active proton-equivalent ion transport via high concentrations of H+-ATPases . Thus, Riftia pachyptila is able to support its symbionts' large demand for inorganic carbon owing to the elevated PCO2 in the vent environment and because of its ability to control its extracellular pH in the presence of large inward CO2 fluxes.

Med Trop (Mars), 1997, 57(1), 77 - 82
{Helicobacter pylori infection in developing countries}; Guisset M et al.; Helicobacter pylori infection is probably the most common bacterial infection in the world . The pathogenicity of this bacteria has been recognized since 1989 in the developed world where prevalence is 20 to 40% . Its role in gastric and duodenal disease is certain and its low recurrence rate justifies eradication . In the developing world prevalence of Helicobacter pylori infection is over 80% with contamination being maximal in children . Transmission is oro-oral and even feco-oral . Crowded living conditions is the determinant factor . Helicobacter pylori infection exhibits special features in developing world . The prevalence of gastroduodenal disease varies according to geographic area independently of the prevalence of Helicobacter pylori infection and stomach cancer is uncommon . In newborns Helicobacter pylori infection seems to be the primary event for chronic malnutrition and diarrhea syndrome with failure to thrive . In practice detection of Helicobacter pylori is difficult in the developing world and presumptive treatment is always followed by recurrence . In the future active or passive immunization will be the solution.

Acta Otolaryngol Suppl, 1997, 529, 30 - 3
Prognosis of acute otitis media . Factors associated with the development of recurrent acute otitis media; Jero J et al.; Clinical factors associated with the development of recurrent acute otitis media (RAOM) (> or = 3 recurrences during 6 months' follow-up period) after acute otitis media (AOM) were analysed in 121 children aged 3 months to 7 years (median 2 years 6 months) . After AOM, 19 (16%) children had primary recurrence (pre-treatment signs and symptoms firstly improved or cured, but worsened or recurred within 30 days' post-treatment) and 33 (27%) developed RAOM during 6 months' follow-up period . It seemed that children < 2 years of age (p = 0.04), children with bilateral disease (p = 0.007), strong infection status (p = 0.05), primary clinical failure (p = 0.04) and development of primary recurrence after AOM (p = 0.001) were significantly related to the development of RAOM in univariate analysis, but only children < 2 years of age (OR 1.5, 95% CI 1.0-5.7, p = 0.04) and the development of primary recurrence (OR 5.1, 95% CI 1.8-14.1, p = 0.002) related to the development of RAOM in multivariate analysis . None of the bacteria cultured from middle ear effusion were related to the development of RAOM.

Int J Aging Hum Dev, 1997, 44(4), 317 - 34
Olfactory acuity as a function of age and gender: a comparison of African and American samples; Barber CE; A frequently reported finding in age-related sensory impairment is that olfaction shows consistent and uniform decline with age . In most studies, discerning whether loss in olfaction is due to aging per se or to factors extrinsic to the aging process (e.g., smoking, chemical exposure, head injury) is difficult . Moreover, studies of olfaction have generally relied on data collected from samples drawn primarily from Western societies . As such, little is known regarding differences in olfaction involving non-Western cultures . Using international data from the 1986 National Geographic Smell Survey, responses of 19,219 American respondents and 3,204 respondents from Africa were analyzed . All respondents were screened for factors negatively affecting olfaction . Measures of olfactory acuity included odor detection, identification, intensity, and quality . The odor of interest was androstenone, a scent produced by bacteria on the human body and appearing in sweat . The results indicate that some measures of olfactory acuity tend to decline across age groups, but that this decline is less marked than reported in previous studies . The most important finding is that loss of olfaction is not consistent or uniform between geographic regions of America or Africa, between male vs . female respondents, or among the four measures of olfactory acuity . African respondents (both men and women) had significantly higher percentages of detection than did American respondents, women generally reported higher levels of olfactory functioning than did men, and some measures of olfaction were stable across age groups, or were higher among older respondents (e.g., odor identification).

Acta Vet Hung, 1997, 45(3), 373 - 86
DNA amplification methods for diagnosis and epidemiological investigations of avian mycoplasmosis; Kempf I; Rapid, sensitive and specific tests that detect nucleic acid from pathogenic mycoplasmas are very attractive for the laboratory detection of infected flocks, and methods for direct detection of the four main pathogenic mycoplasmas have been developed . Moreover, most avian mycoplasma species can be differentiated, according to their unique restriction fragment length polymorphism (RFLP) patterns generated with different restriction enzymes . However, this method is limited to the identification of pure cultures of avian mycoplasmas as other bacteria may be amplified by the set of primers chosen . Another application of PCR-RFLP is the ability to distinguish between very closely related species such as M . gallisepticum and M . imitans . In order to characterise isolates below the species level, PCR-based subtyping methods have been introduced . One of them, arbitrarily primed-PCR, results in strain-specific arrays of DNA fragments that can distinguish even closely related strains of a given species . This method was successfully used to investigate the molecular epidemiology of vaccine strains or of Mycoplasma gallisepticum conjunctivitis in songbirds . Major issues in the development of DNA amplification tests concern the selection of the appropriate target for amplification, specimen collection, DNA preparation and detection of amplification reaction inhibitors . Detection of amplified products is most commonly performed after gel electrophoresis or probe-based methods . Careful consideration to the design and work flow of the facility are necessary to avoid false-positive results.

Parasitol Res, 1997, 83(7), 731 - 3
Observations on the surface coat of Blastocystis hominis; Zaman V et al.; The surface coat of Blastocystis hominis was studied in the electron microscope . In some cells the surface coat was seen in two layers; the external layer was more electron-dense and fragmented than the internal layer . It appears that the surface coat is being continuously formed by the parasite and shed in the environment . The fibrillar material of the surface coat attaches to the bacteria, in some cases, completely surrounding them, possibly causing cytoplasmic damage to the bacterial cell as indicated by loss of electron density.

Dev Biol Stand, 1997, 90, 303 - 9
Strengths and weaknesses of different challenge methods; Nordmo R; The development of effective fish vaccines is greatly dependent on reproducible standardised challenge methods . Different test protocols involving bath exposure, intra-peritoneal and intra-muscular injection and co-habitation are all well described in the literature . Ideally, to represent the many facets of infectious diseases in fish, a test challenge method should closely mimic natural exposure to the pathogen and ensure that immune mechanisms located in the body surfaces play their role . Bath and co-habitation challenges best fulfil this requirement . On the other hand, these methods are more difficult to control and standardise than injection methods . The pathogenicity of some strains of bacteria may also be insufficient to induce a satisfactory outbreak of disease in the target species when using bath or co-habitation challenge . A common measure of efficacy of a vaccine is by Relative Percent Survival (RPS), which expresses the proportional relationship between mortalities in a vaccinated group compared to unvaccinated controls . It has been demonstrated that RPS may vary significantly between challenge methods in groups of Atlantic salmon vaccinated with the same batch of vaccine . Also, RPS may vary for the same batch of vaccine when used in identical test systems at different times . Great care should therefore be taken when competitive products evaluated in different tests are compared . As several vaccines have been marketed and shown to have very good efficacy under field conditions, such products could be introduced as reference standards when testing new candidates for marketing authorisation.

Microbios, 1997, 89(360-361), 151 - 6
Interaction of synthetic pyrethroids with micro-organisms: a review; Johri AK et al.; Pyrethroids are widely used insecticides in agriculture and public health . They are photostable with high insecticidal activity and low toxicity to birds and mammals . At lower concentrations, they are less toxic, but have significant effects on micro-organisms at high concentrations . Pyrethroids constitute about 25% of the total pesticides used in the world and due to the restricted use of organochlorine insecticides, the application of pyrethroids is expected to increase . The present review deals with the interaction of pyrethroids with micro-organisms.

Crit Rev Oral Biol Med, 1997, 8(3), 269 - 90
Short-chain carboxylic-acid-stimulated, PMN-mediated gingival inflammation; Niederman R et al.; This communication reviews the effects of short-chain carboxylic acids on human cells of importance to the periodontium . The central hypothesis is that these acids can alter both cell function and gene expression, and thus contribute to the initiation and prolongation of gingival inflammation . Short-chain carboxylic acids {CH3-(CH2)x-COOH, x < 3} are metabolic intermediates with a broad range of apparently paradoxical biological effects . For example, lactic acid (CH3-CHOH-COOH), a 3-carbon alpha-hydroxy-substituted acid, is widely recognized for its cariogenicity . Lactic acid, however, also occurs in tropical fruits, and is the active ingredient in a variety of anti-wrinkle creams developed by dermatologists . In marked contrast, the unsubstituted 3-carbon propionic acid (CH3-CH2-COOH) is used as a food preservative and is the active principle for one class of non-steroidal anti-inflammatory agents . Interestingly, the addition of one carbon to propionic acid dramatically changes the biological effects . The unsubstituted 4-carbon butyric acid (CH3-CH2-CH2-COOH) is used by hematologists as a de-differentiating agent for the treatment of sickle cell anemia, but by oncologists as a differentiating agent for cancer chemotherapy . Finally, acting either individually or in concert, these acids can increase vascular dilation . Clearly, these acids, while metabolically derived, have a number of very divergent activities which are cell-type-specific (Fig . 1) . It may be telling that periodontal bacteria produce these acids in millimolar concentrations, and that these bacteria can be characterized by their acid production profiles . It is no less interesting that these acids occur in the gingival crevices of human subjects with severe periodontal disease at millimolar levels which are > 10-fold higher than those found in mildly diseased subjects, and are undetectable in healthy subjects . Further, when applied directly to healthy human gingiva, short-chain carboxylic acids stimulate a gingival inflammatory response and inflammatory cytokine release . At the cellular level, these acids inhibit proliferation of gingival epithelial and endothelial cells, and inhibit leukocyte apoptosis and function, but can stimulate leukocyte cytokine release . At the molecular level, these acids can stimulate neutrophil gene transcription, translation, and protein expression . Thus, the likelihood is high that these acids, in addition to their cariogenic activity, can promote and prolong gingival inflammation . Our challenge will be to identify the cell or cells of the periodontium which respond to short-chain carboxylic acids, to delineate their responses and the molecular mechanism(s) of these effects, and to categorize the aspects of the inflammatory components which damage and those which protect the host . With this information, it may be possible to begin to rationally identify and test pharmaceutical agents which diminish the harmful aspects, while enhancing the beneficial components, of the inflammatory response.

DNA Seq, 1997, 7(5), 267 - 84
Identification, sequences, and expression of Treponema pallidum chemotaxis genes; Greene SR et al.; Treponema pallidum, the agent of syphilis, is a pathogenic spirochete that has no known mechanisms of genetic exchange and cannot be continuously cultivated in vitro . A probe based on the nucleotide sequence of the T . pallidum cheA gene was used to screen a T . pallidum genomic DNA library . A treponemal DNA region containing four open reading frames (orfs) was identified . The proteins encoded by these orfs have significant homology with proteins involved in bacterial chemotaxis . The orfs have been designated cheA, cheW, cheX, and cheY . The cheA, cheW, and cheY genes were individually-cloned and expressed in vitro . The observed molecular mass of each protein correlated well with its predicted molecular mass . Reverse transcriptase-PCR data indicate that cheA through cheY are co-transcribed . The organization of these genes suggests that they comprise an operon . We hypothesize that the ability to sense and respond to nutrient gradients is important for the survival and dissemination of T . pallidum in vivo . The presence of a putative che operon strongly suggests that T . pallidum has the potential for a chemotactic response.

Microbiol Immunol, 1997, 41(6), 487 - 94
Moraxella (Branhamella) catarrhalis adherence to human bronchial and oropharyngeal cells: the role of adherence in lower respiratory tract infections; Rikitomi N et al.; To study the role of Moraxella (subgenus Branhamella) catarrhalis (B . catarrhalis) adherence to airway cells in lower respiratory tract infections, the in vitro attachments of B . catarrhalis to upper airway (oropharyngeal) and lower airway (bronchial) epithelial cells were compared . The adherence of 4 strains (1 nonfimbriated and 3 fimbriated) of B . catarrhalis to respiratory tract epithelial cells collected from 11 patients with chronic pulmonary disease (CPD) and 11 healthy individuals was evaluated . Both the fimbriated and nonfimbriated strains showed increased attachment to oropharyngeal cells in the CPD patients (mean +/- SEM; 25.0 +/- 3.2/cell; P < 0.01) when compared to the control subjects (12.1 +/- 1.1/cell) . On the average, the attachment to bronchial cells was 6.1 to 13.6 times greater per surface area (bacteria/micron2) than the attachment to oropharyngeal cells . The fimbriated strains tended to adhere in higher numbers to bronchial cells (19.0 +/- 1.8/cell) than the nonfimbriated strain (8.7 +/- 1.2/cell), although there was no difference between the CPD and control groups . In conclusion, the attachment of B . catarrhalis to oropharyngeal cells may be an enhancing factor for colonization in the upper respiratory tract in patients with CPD, and elevated adherence of the bacteria to bronchial cells may suggest pathogenic importance when mucociliary function is impaired.

Probl Tuberk, 1997, (2), 32 - 3
{Immune responsive features in patients with pulmonary tuberculosis complicated by intestinal dysbacteriosis}; Lineva ZE; Clinical and immunological studies made in 130 patients with various types of pulmonary tuberculosis ascertain that the absolute majority (90%) patients who begin to develop intestinal dysbacteriosis due to long-term chemotherapy have T-cell immunity impairments, which appears as their quantitative shortage and functional insufficiency . There is a marked decrease in T helper levels with subpopulation imbalance and lower Tx/Tc ratios . The applied complex of corrective means produces stimulating and correcting effects on cellular and humoral immunological responses, promotes the increase of T helper counts, by eliminating the imbalance of regulatory subpopulations of T lymphocytes and having a beneficial effect on the clinical and X-ray changes in pulmonary tuberculosis.

Postepy Hig Med Dosw, 1997, 51(2), 115 - 38
{Immunologic reactions in infections caused by Helicobacter pylori}; Chmiela M et al.; Helicobacter pylori is recognized as an important cause of chronic antral gastritis and peptic ulceration . Moreover, H . pylori associated inflammatory process has been linked with gastric carcinoma . Many putative virulence factors of H . pylori have been suggested, including motility, urease and cytotoxins production and bacterial adhesins . An accessory function of CagA antigen and bacterial heat-shock proteins in the pathogenesis of H . pylori infections have been also considered . H . pylori-induced immunological response is discussed as regards local and general antibody production, the interaction of the bacteria with the phagocytes and still controversial involvement of T cells . Data on the importance of cytokines and inflammatory mediators in the disruption of the gastric mucosal barriers as well as the evidence to support a role for H . pylori as a risk factor for gastric carcinoma are also presented.

Prev Vet Med, 1997 Jan, 29(3), 161 - 77
Assessing infections at multiple levels of aggregation; Kadohira M et al.; The patterns of sero-prevalence of antibodies to four infectious diseases, representing a broad range of pathogens (bacteria: brucellosis; mycoplasma: contagious bovine pleuropneumonia; viruses: infectious bovine rhinotracheitis; protozoa: trypanosomosis) were investigated at three levels of organization (farm, area and district) . Three contrasting districts in Kenya were compared: an arid and pastoral area (Samburu); a tropical highland area (Kiambu), and a tropical coastal area (Kilifi) . Cattle in three districts were selected by two-stage cluster sampling between August 1991 and 1992 . Schall's algorithm, a generalized linear mixed model suitable for multi-level analysis, was compared to ordinary logistic regression (OLR), which ignores clustering of responses; generalized estimating equations (GEE) or Jacknife, to account for clustering at the farm level; SAS VARCOMP, which provides normal-theory random-effects models . Schall's algorithm provided similar estimates to GEE (regression effects) and Jackknife (standard errors) for farm-level clustered data . Extending Schall's procedure for additional district and area-within-district random effects usually provided additional information . In general, models that included only a farm-level random effect consistently provided larger estimates of farms' variance components than did models with additional district and area random effects . The four type diseases exhibited various amounts of clustering . Brucellosis had moderate farm clustering plus some area and district clustering . Contagious bovine pleuropneumonia had only a small amount of clustering, mostly by area . Infectious bovine rhinotracheitis exhibited a large amount of clustering, primarily at the farm level . Trypanosomiasis antibody prevalence varied by district, area and farm . We believe that patterns of disease clustering identified by multi-level analysis can be used to better target high-risk units for disease control and guide research to understand disease transmission factors.

Med Lav, 1997 Jan-Feb, 88(1), 13 - 23
{Genotoxicity of urban air particulate matter}; Clonfero E; This paper reviews the genetic toxicology of urban air particulate . Dusts present in the urban environment are composed of a special type of soot, currently mainly emitted by motor vehicles . This soot contains variable quantities of adsorbed highly active genotoxic compounds (polycyclic aromatic hydrocarbons and their nitro derivatives, i.e., nitroarenes) . Nitroarenes are among the compounds with the highest genotoxic activity according to the Ames test (genetic point mutation on bacteria), and are particularly abundant in ultrafine particulate matter (< 1.1 microns) emitted by diesel engines, mainly trucks . Diesel emissions have been considered as probable carcinogenic agents for man by the International Agency for Research on Cancer (IARC) . Highly mutagenic activity is present in the air of all the cities in the world, and there is increasing concern about possible carcinogenic effects on the general population as a result of exposure to urban particulate matter . Early epidemiological studies on city-dwellers exposed to heavy pollution indicate an excess of lung tumours . Lastly, there is a possible carcinogenic risk for occupationally exposed workers, such as traffic police and those working for city road cleaning services, etc., who are constantly obliged by their jobs to be exposed to polluted city air at work.

Int Rev Cytol, 1997, 175, 137 - 240
Sialic acids in molecular and cellular interactions; Kelm S et al.; Sialic acids (Sias) are terminal components of many glycoproteins and glycolipids especially of higher animals . In this exposed position they contribute significantly to the structural properties of these molecules, both in solution and on cell surfaces . Therefore, it is not surprising that Sias are important regulators of cellular and molecular interactions, in which they play a dual role . They can either mask recognition sites or serve as recognition determinants . Whereas the role of Sias in masking and in binding of pathogens to host cells has been documented over many years, their role in nonpathological cellular interaction has only been shown recently . The aim of this chapter is to summarize our knowledge about Sias in masking, for example, galactose residues, and to review the progress made during the past few years with respect to Sias as recognition determinants in the adhesion of pathogenic viruses, bacteria, and protozoa, and particularly as binding sites for endogenous cellular interaction molecules . Finally, perspectives for future research on these topics are discussed.

Parasitol Res, 1997, 83(5), 499 - 503
Electron microscopy studies on the a-bacteroids in the fat bodies of the lantern bug Pyrops candelaria Linn . (Homoptera: Fulgoridae); Wang JB et al.; The ultrastructure of alpha-bacteroids in relation to the fat-body cells of the lantern bug Pyrops candelaria was described . The fat-body-cell cytoplasm contained numerous mitochondria, rough endoplasmic reticula, vacuoles, and storage granules . Its nucleus had scattered chromatin materials . The alpha-bacteroid was enveloped by three membrane layers, namely, the plasma membrane, the cell wall, and the membrane envelope . Its cytoplasm contained amorphous dense bodies . The bacteroid reproduced by binary fission . Tracheoles were also found among fat-body cells.

Eur J Surg Suppl, 1997, (579), 31 - 3
Airborne particles from latex gloves in the hospital environment; Newsom SW et al.; Air sampling studies are described which show high levels of airborne starch powder contamination in areas where powdered latex gloves are used . Furthermore, culture of collected samples show a clear association between starch particles and bacterial colonies in an experimental system suggesting that airborne particles could act as a vector for pathogens in the hospital environment.

Adv Exp Med Biol, 1997, 419, 203 - 7
Effects of inhibitors of ADP-ribosylation on macrophage activation; Le Page C et al.; Nitric oxide (NO) is a potent mediator involved in many biological functions including macrophage cytotoxicity and non-specific immunity against parasites, bacteria and viruses . Murine macrophages possess the capacity to express the inducible NO synthase (iNOS) which is not constitutively expressed but induced at the transcriptional level by interferon gamma (IFN-gamma) alone or synergistically with LPS . We have investigated the possible role of ADP-ribosylation reactions in the signaling pathway involved in NO synthase induction, since ADP-ribosylation has been reported to be involved in the expression of certain IFN-gamma and LPS-inducible genes . We found that inhibitors of ADP-ribosylation inhibited nitrite synthesis in RAW 264.7 macrophages after stimulation by IFN-gamma and LPS . These ADP-ribosylation inhibitors acted by preventing NO synthase mRNA induction, without inhibiting NO synthase enzyme activity . IRF-1, a transcription factor induced and activated by IFN-gamma was recently shown to be involved in iNOS induction . We showed that inhibitors of ADP-ribosylation had no effect on IFN-gamma-mediated mRNA induction of IRF-1 nor on its activation and binding to its target sequence in the iNOS gene . In addition, the inhibitors failed to impair the IFN-gamma-mediated antiviral activity against VSV virus . Since induction by IFN-gamma of IRF-1 and induction of the antiviral state proceed through the JAK/STAT signalling pathway, our results imply that ADP-ribosylation reactions are not involved in triggering this pathway . Although the precise mechanism requires further investigation, our results indicate that ADP-ribosylation is a crucial step restricted to the signalling pathway which leads to iNOS mRNA induction, as well as TNF and MHC class II induction during macrophage activation.

Adv Exp Med Biol, 1997, 412, 21 - 9
Neuro-immune pathobiology of infectious enteric disease; Argenzio RA; Recent knowledge of neuro-endocrine-immune communication in the intestinal mucosa has provided a new paradigm for the pathophysiology of diarrheal disease that will significantly alter and advance therapeutic strategies . Mast cells, enteroendocrine cells and phagocytes are the proximate mediators of signalling cascades activated by parasitic nematodes and food allergens, enterotoxigenic bacteria, and at least some of the invasive pathogens, respectively . These proximate, trigger cells give rise to products that affect epithelial function directly, or indirectly through stimulation of prostaglandin production by mesenchymal cells, and enteric nerve stimulation, which can markedly amplify the initial stimulus . The enteric nervous system in fact may mediate the majority of the secretory response induced by enterotoxins or phagocytes . The signalling network mediated by cells in the lamina propria provides new points of control for pharmacological therapy.

Rev Gastroenterol Mex, 1997 Jan-Mar, 62(1), 22 - 8
{Helicobacter pylori infection and gastric cancer in Mexico . A challenge for prevention and population control}; Lopez Carrillo L et al.; The International Agency for Research on Cancer (IARC) of the WHO has recognized a cause-effect relationship between Helicobacter pylori (Hp) infection and gastric cancer of such magnitude that the presence of this infection increases the risk of gastric cancer approximately four times . Gastric cancer is currently the second cause of mortality due to malignant neoplasms in Mexico City . This article explores the association between Hp infection and gastric cancer incidence through an epidemiological study including 109 gastric cancer patients and 177 hospital controls in Mexico City . The study estimates that, in the population studied, Hp infection was present in 87.2% of the cases, compared with 82.5% of the controls . The odds ratio of having gastric cancer if infected with Hp was 1.44 IC95% 0.7-2.8 . In addition, it was calculated that with eradication of Hp infection in the general population, gastric cancer incidence would decrease by at least 26.6% . An improvement of the actual sanitary conditions along with the development of an effective vaccine for Hp infection and the existence of increasingly effective treatments to eradicate the bacteria are the necessary next step for populational prevention and control of gastric cancer.

Cell Motil Cytoskeleton, 1997, 37(2), 139 - 48
Reactivation of cell surface transport in Reticulomyxa; Orokos DD et al.; Granuloreticulosean protists transport particles (e.g., bacteria, algae, and sand grains) along the outer surfaces of their pseudopodia . This cell surface transport plays a vital role in feeding, reproduction, shell construction, and locomotion and can be visualized by the movements of extracellularly adherent polystyrene microspheres (i.e., latex beads) . Our videomicroscopic analyses of transport associated with the pseudopodia of Reticulomyxa filosa revealed two distinct types of both intracellular and cell surface transport: (1) saltatory, bidirectional transport of individual or clustered organelles and/or surface-attached particles, and (2) continuous, unidirectional bulk or "resolute" motion of aggregated organelles and/or surface-bound particles . Organelles and surface-attached polystyrene microspheres remained firmly attached to the microtubule cytoskeletons of detergent-extracted pseudopodia . Both saltatory and resolute organelle and surface transport reactivated upon the addition of 0.01-1.0 mM ATP . At 1 mM ATP, the velocities of reactivated saltatory transport were indistinguishable from those observed in vivo . The reactivated transport was microtubule-dependent and was not inhibited by incubation with Ca(2+)-gelsolin under conditions that abolish rhodamine-phalloidin detection of actin filaments . These findings provide further support that both intracellular organelle and membrane surface transport are mediated by a common mechanism, and establish Reticulomyxa as a unique model system to further study the mechanochemistry of cell surface transport in vitro.

HPB Surg, 1997, 10(3), 143 - 7
The internal biliary fistula--reappraisal of incidence, type, diagnosis and management of 33 consecutive cases; Yamashita H et al.; To reevaluate the current features of spontaneous internal biliary fistulas, we reviewed 1,929 consecutive patients who had been treated for biliary tract diseases during the recent 12-year period . Thirty-three patients had internal biliary fistulas and the incidence was 1.9% . Of 33 patients, 20 were women and 13 were men with the average age 63 years, and their mean duration of illness was 4 years . A total of 37 fistulas were found and the most common type was choledochoduodenal (62%), followed by cholecystoduodenal (19%), cholecystocholedochal (11%) and cholecystocolonic (8%) fistulas . Internal biliary fistulas of thirty-one patients were caused by biliary stones and those of two patients by malignant tumors . All of the 17 bile samples examined were bacteria positive and the majority of calculi were brown pigment stones . All of the choledochoduodenal fistulas were correctly diagnosed by endoscopic retrograde cholangiography . In 14 patients with cholecystoenteric or cholecystocholedochal fistulas, direct evidence of the internal fistula was obtained only in 7 patients (50%) preoperatively . Pneumobilia, a small atrophic gallbladder adherent to the neighboring organs and a history of spontaneous disappearance of jaundice in elderly patients may indicate the presence of a cholecystoentric fistula . Since the preoperative diagnostic rate for internal biliary fistula involving the gallbladder is still low, care is necessary before and at the time of surgery especially during laparoscopic cholecystectomy for elderly patients with cholelithiasis.

Vet Res, 1997, 28(1), 65 - 76
Production and characterization of monoclonal antibodies against Taylorella equigenitalis; Gradinaru DA et al.; Monoclonal antibodies were produced against Taylorella equigenitalis using two reference strains . Out of the 79 hybridoma clones shown to express antibodies to T equigenitalis by indirect immunofluorescence assay, 16 were selected for monoclonal antibody production and characterization . These clones recognized different field strains of T equigenitalis isolated in France . They showed no cross-reaction with bacterial strains with previously reported antigenic cross-reactivity, nor did they react with other bacteria commonly found in genital flora . The epitopes recognized by eight of the monoclonal antibodies were situated in proteins of 150, 120, 52.7 and 22 kDa . These epitopes were resistant to the extraction denaturing conditions . These monoclonal antibodies could be used as reagents for specific detection of T equigenitalis.

Bull Acad Natl Med, 1997 Jan, 181(1), 21 - 40; discussion 40-2
{Degradation of soil quality: health and environmental risks}; Robert M; This is a general survey of soil quality degradation and its consequences on human health and environment quality . The first part deals with the large complexity and reactivity of soil and its specific position as an environmental interface . The origin and behaviour of the main pollutant accumulated in soils are treated in the second and third parts . A special attention is paid to the main pollutants i.e., anions, cations (trace elements), pesticides, persistent organic pollutants (POP), bacteria, DNA.. . As they are closely linked to the soil constituents (clays, humus...), their real toxicity or effectiveness is examined . In the last part, the risks of soil pollutants for the environment and human health are shortly considered . Direct risks for health are few and mainly concern the ingestion of polluted soil by children; as regards the ecotoxicological and toxicological risks, the Critical pollutant loads have to be defined . The main risks are indirect and involve pollution of either the food chain or of the water and sediments by vertical or lateral (erosion) transfers.

Biol Trace Elem Res, 1997 Jan, 56(1), 63 - 91
Genomic structures of viral agents in relation to the biosynthesis of selenoproteins; Taylor EW et al.; The genomes of both bacteria and eukaryotic organisms are known to encode selenoproteins, using the UGA codon for seleno-cysteine (SeC), and a complex cotranslational mechanism for SeC incorporation into polypeptide chains, involving RNA stem-loop structures . These common features and similar codon usage strongly suggest that this is an ancient evolutionary development . However, the possibility that some viruses might also encode selenoproteins remained unexplored until recently . Based on an analysis of the genomic structure of the human immunodeficiency virus HIV-1, we demonstrated that several regions overlapping known HIV genes have the potential to encode selenoproteins (Taylor et al . {31}, J . Med . Chem . 37, 2637-2654 {1994}) . This is provocative in the light of overwhelming evidence of a role for oxidative stress in AIDS pathogenesis, and the fact that a number of viral diseases have been linked to selenium (Se) deficiency, either in humans or by in vitro and animal studies . These include HIV-AIDS, hepatitis B linked to liver disease and cancer, Coxsackie virus B3, Keshan disease, and the mouse mammary tumor virus (MMTV), against which Se is a potent chemoprotective agent . There are also established biochemical mechanisms whereby extreme Se deficiency can induce a proclotting or hemorrhagic effect, suggesting that hemorrhagic fever viruses should also be examined for potential virally encoded selenoproteins . In addition to the RNA stem-loop structures required for SeC insertion at UGA codons, genomic structural features that may be required for selenoprotein synthesis can also include ribosomal frameshift sites and RNA pseudoknots if the potential selenoprotein module overlaps with another gene, which may prove to be the rule rather than the exception in viruses . One such pseudoknot that we predicted in HIV-1 has now been verified experimentally; a similar structure can be demonstrated in precisely the same location in the reverse transcriptase coding region of hepatitis B virus . Significant new findings reported here include the existence of highly distinctive glutathione peroxidase (GSH-Px)-related sequences in Coxsackie B viruses, new theoretical data related to a previously proposed potential selenoprotein gene overlapping the HIV protease coding region, and further evidence in support of a novel frameshift site in the HIV nef gene associated with a well-conserved UGA codon in the 1-reading frame.

Vet Pathol, 1997 Jan, 34(1), 50 - 1
Gastric ulcer associated with a Helicobacter-like organism in a cougar (Felis concolor); Hill JE et al.; A 12-year-old captive female cougar (Felis concolor) died following perforation of a gastric ulcer . Histologically, erosions and ulcers were present in the antral area of the stomach . Fibroplasia and infiltrates of neutrophils bordered the ulcers . Modified Steiner silver stain revealed numerous tightly coiled helical bacteria . The bacteria stained positively with a rabbit polyclonal anti-Helicobacter pylori antibody . Morphology, location, and positive immunohistochemical staining suggests that the organism is a Helicobacter.

Rays, 1997 Jan-Mar, 22(1), 127 - 56
Idiopathic interstitial lung disease: anatomoradiologic pathogenesis; Chiodera P; Interstitial lung disease encompasses a large number of clinical disorders that affect the epithelium, the endothelium or both cell surfaces of alveolar wall and satellite structures including terminal and respiratory bronchioles . Causative factors are over 200 from bacteria, fungi, viruses, protozoans, to collagen disease, hypersensitivity and inorganic pneumoconiosis . Clinical and histological findings of open lung and transbronchial biopsies from 50 patients are reported, 18 patients were affected by diffuse alveolar damage (DAD), 15 patients by usual interstitial pneumonia (UIP), 8 patients by non specific interstitial pneumonia-fibrosis (NSIP-F), 9 patients by bronchiolitis obliterans organizing pneumonia (BOOP) correlated with conventional chest radiography in 30 patients and with HRCT in 31 patients . Interstitial lung disease other than histiocytosis X share anatomoradiologic features indicative for activity, chronic progression, chronic quiescence, chronic advanced and irreversible disease . In general, the histologic features correlate with radiographic patterns and even if radiologic findings do not always supply definitive diagnosis, some HRCT patterns are highly suggestive and usually classified into 4 categories: normal; with ground glass attenuation; linear, nodular or reticulo-nodular; honeycombing, suggestive for end-stage fibrosis . Correlation of HRCT with histologic findings in 31 patients with idiopathic interstitial fibrosis (IIF) allowed assessment of disease activity, follow-up and therapeutic result . HRCT definitely better than conventional radiology detects the presence, type and extent of parenchymal alterations, differentiating potential reversible lesions (inflammatory) from potentially irreversible (fibrotic) lesions . In IIF, for diagnostic accuracy the specimen of open lung (the gold standard), transbronchial or video-thoracoscopic biopsy must be preoperative, HRCT-assisted and centered on ground glass opacties (or nodules in suspected histiocytosis X) since a diagnostically reliable biopsy correlates with HRCT morphology of histologic specimen . Interstitial lung disease includes benign as well as malignant forms, thus only a multidisciplinary approach can prevent long term hazardous effects as severe cor pulmonale or a fatal outcome . The histologic HRCT-assisted assessment of "active" lesions is crucial for correct careful treatment of these patients.

Environ Mol Mutagen, 1997, 29(3), 296 - 302
Enhancement of the clastogenicity of N-nitrosodialkylamines plus near-ultraviolet irradiation by ethanol in Chinese hamster lung fibroblasts; Yamashita Y et al.; Early work from our laboratory showed a synergistic action of N-nitrosodialkylamines and near-ultraviolet light (UVA, 320-400 nm) to cause mutations in bacteria and phages . Recently we reported that N-nitrosodialkylamines + UVA induces chromosome aberrations in Chinese hamster lung cells in culture . We have now found that ethanol can potentiate this clastogenic action of N-nitrosodialkylamines + UVA . When the cells were treated with N-nitrosopyrrolidine (NPYR) or N-nitrosodiethylamine (NDEA) + UVA for 2 hours in the presence of 1% ethanol, approximately 2-fold increase in the numbers of cells with aberrant chromosomes was observed, compared to those found without the ethanol . NPYR/NDEA only ethanol only, or ethanol + UVA did not cause the aberrations . The enhancement was dependent on the concentration of ethanol . Treatment of cells with ethanol before the NPYR + UVA was ineffective . By contrast, treatment of cells with NPYR-UVA and then with ethanol was as effective as with the simultaneous treatment . Methanol showed synergistic effects similar to those of ethanol, but mannitol did not . Intracellular hydrogen peroxide was found to be increased twofold over that in the background by a treatment with ethanol + UVA . The alcohol-mediated enhancement of the clastogenic action of N-nitrosodialkylamines + UVA may be a consequence of an increase in intracellular oxidative stress, or simply due to increased membrane permeability.

Acta Vet Scand, 1997, 38(1), 9 - 16
Collagenolytic activity and its sensitivity to doxycycline inhibition in tracheal aspirates of horses with chronic obstructive pulmonary disease; Koivunen AL et al.; The collagenolytic activity and its sensitivity to doxycycline inhibition in tracheal aspirates (TA) of horses with chronic obstructive pulmonary disease (COPD) was analyzed with SDS-PA gel electrophoresis (SDS-PAGE), using Type 1 collagen as the substrate . Both autoactive and total collagenase activities were significantly higher in TAs of horses with symptomatic COPD than in TAs of healthy horses . Doxycycline inhibition studies suggest that most of the TA collagenase is of the neutrophil type (MMP-8), but some is derived from other cells such as fibroblasts and monocyte/macrophages (MMP-1) and bacteria (bacterial collagenases) . Drugs inhibiting collagenases in the respiratory tract might be worth a trial in the treatment of COPD in horses.

Emerg Infect Dis, 1997 Jan-Mar, 3(1), 21 - 32
Mycoplasmas: sophisticated, reemerging, and burdened by their notoriety; Baseman JB et al.; Mycoplasmas are most unusual self-replicating bacteria, possessing very small genomes, lacking cell wall components, requiring cholesterol for membrane function and growth, using UGA codon for tryptophan, passing through "bacterial-retaining" filters, and displaying genetic economy that requires a strict dependence on the host for nutrients and refuge . In addition, many of the mycoplasmas pathogenic for humans and animals possess extraordinary specialized tip organelles that mediate their intimate interaction with eucaryotic cells . This host-adapted survival is achieved through surface parasitism of target cells, acquisition of essential biosynthetic precursors, and in some cases, subsequent entry and survival intracellularly . Misconceptions concerning the role of mycoplasmas in disease pathogenesis can be directly attributed to their biological subtleties and to fundamental deficits in understanding their virulence capabilities . In this review, we highlight the biology and pathogenesis of these procaryotes and provide new evidence that may lead to increased appreciation of their role as human pathogens.

Arch Virol, 1997, 142(2), 305 - 21
Protein interaction domains of the measles virus nucleocapsid protein (NP); Liston P et al.; The currently accepted model for measles virus (MV) transcription and replication assumes the nucleocapsid (NP) protein to possess the ability to bind to RNA, to other NP molecules, and to the phosphoprotein (P) during ribonucleocapsid (RNP) assembly, as well as to the matrix protein (M) during virion assembly . We have cloned the MV NP open reading frame and have expressed the protein in bacteria as a fusion with glutathione-S-transferase (GST) . Affinity purified GST-NP fusion protein has been used as a probe to examine the interaction of NP with {35S} methionine labeled proteins from MV-infected cells . We have demonstrated definite and specific interactions between NP and itself and between NP and P, but have been unable to demonstrate any interaction between NP and M . We have been able to provide independent confirmation of this pattern of interaction using the yeast two-hybrid assay . We have, in addition, been able to map the domains of NP involved in these interactions by assays using sets of amino- and carboxy-terminal deletion mutants of GST-NP . The NP-NP interaction domain was found to reside in the highly conserved middle and amino-terminal domains of the protein . The hyper-variable carboxy-terminus and the conserved middle domain appear to constitute separate and independent sites for the binding of P to NP . The significance of these findings in regard to MV transcription and replication is discussed.

J Pharm Pharmacol, 1997 Jan, 49(1), 35 - 9
In-vivo assessment of extrahepatic metabolism of paeoniflorin in rats: relevance to intestinal floral metabolism; Takeda S et al.; The extraction ratios of paeoniflorin in gut wall (EG), liver (EH) and lung (EL) were assessed by comparing AUCs after various routes of its administration to estimate the first-pass effects and the metabolism by intestinal flora . Pulmonary extraction ratio of paeniflorin was assessed by comparing AUCs calculated from venous and arterial plasma concentrations after its intravenous administration (0.5 mg kg-1) . The mean pulmonary extraction ratio was estimated to be 0.06 . The hepatic extraction ratio (EH was assessed by comparing AUCs after intraportal and intravenous administrations (0.5 and 5 mg kg-1) . The plasma concentration profiles of paeoniflorin after intraportal administration were very close to those after intravenous administration, suggesting a negligible hepatic extraction ratio of paeoniflorin . The AUC value after intraperitoneal administration (0.5 mg kg-1) was greater than that after intraportal or intravenous administration . This finding suggests that paeoniflorin is not metabolized in the gut wall . The transference of paeoniflorin from the serosal side to the mucosal side was evaluated by the in-vitro everted sac method . The low intestinal permeability (19.4% at 60 min) was demonstrated by the comparison with phenobarbital (63.1% at 60 min) . We conclude that paeoniflorin is not metabolize by gut wall, liver and lung, its poor absorption from the intestine results in extremely low bioavailability and the unabsorbed fraction of paeoniflorin is degraded by the intestinal flora.

Free Radic Biol Med, 1997, 22(5), 775 - 85
Ex vivo induction of apoptosis in lymphocytes is mediated by oxidative stress: role for lymphocyte loss in HIV infection; Dobmeyer TS et al.; Programmed cell death (apoptosis) of T-lymphocytes observed in human immunodeficiency virus (HIV) infected individuals could be linked to oxidative stress . Therefore, we have investigated whether reactive oxygen species (ROS) induce apoptosis, which might contribute to the cell loss during progression of HIV-1 infection . ROS were generated in peripheral blood mononuclear cells (PBMC) obtained from HIV-1-positive patients and from healthy controls by stimulation with bacteria or by treatment with hypoxanthine/xanthine oxidase, which has been shown to generate ROS without direct involvement of cytokines . A dose-dependent inhibition of ROS formation correlated with the reduction of apoptosis induced by both bacterial and hypoxanthine/xanthine oxidase stimulation, suggesting that ROS generation was responsible for the induction of apoptosis . In addition, hydrogen peroxide (H2O2) rather than superoxide (O2.-) was observed to induce apoptosis . ROS-dependent apoptosis was shown to be independent of cytokines such as tumor necrosis factor-alpha (TNF-alpha) . ROS-induced apoptosis was significantly enhanced in HIV-infected subjects even in the very early stages after infection . Moreover, ROS-mediated apoptosis was not restricted to a particular lymphocyte subset . In view of the diminished oxidative resistance of HIV-infected individuals, our results suggest that ROS-mediated apoptosis might contribute to the deletion of lymphocytes and to the pathogenesis of the disease.

Environ Mol Mutagen, 1997, 29(2), 143 - 51
Mutations induced in a shuttle vector plasmid exposed to monofunctionally activated mitomycin C; Maccubbin AE et al.; Reductive activation of mitomycin C leads to its covalent binding to DNA, forming monoadducts and cross-links . The cytotoxicity of mitomycin C has been attributed to cross-link formation, whereas monoadducts are assumed to cause mutagenicity . We have developed a 32P-postlabeling technique to measure mitomycin C DNA adducts . Using this technique, we have measured monoadduct formation in the shuttle vector plasmid pSP189 and have determined mutations induced by monoadduct formation . The shuttle vector plasmid was incubated with mitomycin C under conditions favoring monofunctional activation of mitomycin C . The plasmid was then replicated in human Ad293 cells, rescued in bacteria, and analyzed for mutations in the supF tRNA gene sequence of pSP189 . One major mitomycin C/DNA adduct was observed by 32P-postlabeling and was characterized as a monoadduct of guanine . When pSP189 was exposed to monofunctionally activated mitomycin C, increases in adduct levels and mutation frequency were found to be related to mitomycin C concentration . The majority of the mutations involved single bases, with base substitutions making up 59.1% of the total mutations observed . Of the base substitutions, 67.2% were transversions and 32.8% were transitions, with nearly 80% of all base substitutions involving G:C base pairs . Deletions, either as single bases or large deletions, also involved G:C base pairs the majority of the time . The observed bias of mutations at G:C and the formation of a mitomycin C/DNA monoadduct involving guanine suggests that monoadduct formation may be responsible for the mutations.

Genes Cells, 1997 Jan, 2(1), 13 - 28
The N-end rule pathway of protein degradation; Varshavsky A; The N-end rule relates the in vivo half-life of a protein to the identity of its N-terminal residue . Similar but distinct versions of the N-end rule operate in all organisms examined, from mammals to fungi and bacteria . In eukaryotes, the N-end rule pathway is a part of the ubiquitin system . Ubiquitin is a 76-residue protein whose covalent conjugation to other proteins plays a role in many biological processes, including cell growth and differentiation . I discuss the current understanding of the N-end rule pathway.

Food Chem Toxicol, 1997 Jan, 35(1), 127 - 58
IRAG working group 5 . Other assays . Interagency Regulatory Alternatives Group; Curren RD et al.; As part of the Interagency Regulatory Alternatives Group (IRAG) program to evaluate the state of the art in the development of alternative (non-whole animal) eye irritation tests, academic and industrial organizations were invited to submit in vitro eye irritation data generated in their laboratories to one of several working groups for review . The assays reviewed in this report (from Working Group 5 . "Other Assays") were the EYTEX assay, tissue equivalent assay, a cytotoxicity assay using three-dimensional human fibroblast constructs, the Microtox assay, and other miscellaneous assays . Each submission consisted of raw data for chemicals and products tested, a description of the methodology, and an analysis (generally by regression analysis and Pearson's correlation coefficient) for the performance of the in vitro test relative to its ability to predict individual ocular tissue scores or total ocular score . In vivo data were generated according to the scoring methods proposed by Draize . Working Group 5 evaluated the submissions and commented on the utility of the assays . The variability of the in vivo data made conclusions difficult in many situations . Most of these assays were deemed useful (within limited chemical classes) for screening purposes or for use in conjunction with other toxicological information.

Exp Eye Res, 1997 Jan, 64(1), 37 - 43
Fibronectin concentration in tears of contact lens wearers; Baleriola-Lucas C et al.; The tears of soft contact lens wearers (wearing lenses on an extended were schedule) were analysed for the amount of fibronectin, albumin and total protein . Tears were collected during the day without stimulation and the levels were then compared to levels in non-contact lens wearers tears . The level of fibronectin in the tears of contact lens wearers (120 ng ml-1) was significantly higher than in the tears of non-contact lens wearers (21 ng ml-1; P < 0.01) . There were no differences in the amount of albumin or total protein . When contact lens wearers were divided into those subjects who carried significant levels of bacteria on their contact lenses and non-carriers, the amount of fibronectin and albumin was found to be higher in the tears of bacterial carriers (P < 0.03), whereas the amount of total protein remained the same . The fibronectin in the tears and on the contact lens may predispose subjects to contamination by bacteria and thus increase the risk of inflammatory or infectious diseases associated with bacterial contamination of contact lenses.

Subcell Biochem, 1997, 28, 57 - 87
Polyprenyl diphosphate synthases; Ogura K et al.; It is noteworthy that in spite of the similarity of the reactions catalyzed by these prenyltransferases, the modes of expression of catalytic function are surprisingly different, varying according to the chain length and stereochemistry of reaction products . These enzymes are summarized and classified into four groups, as shown in Figure 13 . Short-chain prenyl diphosphates synthases such as FPP and GGPP synthases require no cofactor except divalent metal ions, Mg2+ or Mn2+, which are commonly required by all prenyl diphosphate synthases . Medium-chain prenyl diphosphate synthases, including the enzymes for the synthesis of all-E-HexPP and all-E-HepPP, are unusual because they each consist of two dissociable dissimilar protein components, neither of which has catalytic activity . The enzymes for the synthesis of long-chain all-E-prenyl diphosphates, including octaprenyl (C40), nonaprenyl-(C45), and decaprenyl (C50) diphosphates, require polyprenyl carrier proteins that remove polyprenyl products from the active sites of the enzymes to maintain efficient turnovers of catalysis . The enzymes responsible for Z-chain elongation include Z,E-nonaprenyl-(C45) and Z,E-undecaprenyl (C55) diphosphate synthases, which require a phospholipid . The classification of mammalian synthases seems to be fundamentally similar to that of bacterial synthases except that no medium-chain prenyl diphosphate synthases are included . The Z-prenyl diphosphate synthase in mammalian cells is dehydrodolichyl PP synthase, which catalyzes much longer chain elongations than do bacterial enzymes . Dehydrodolichyl PP synthase will be a major target of future studies in this field in view of its involvement in glycoprotein biosynthesis.

J Basic Microbiol, 1997, 37(1), 11 - 21
Analysis of the PHA granule-associated proteins GA20 and GA11 in Methylobacterium extorquens and Methylobacterium rhodesianum; Follner CG et al.; Electrophoretic analysis of the proteins bound to poly(3-hydroxybutyric acid), PHB-, granules in Methylobacterium extorquens, M . rhodesianum as well as the PHB-leaky mutants Mu 1 and Mu 11, which were isolated from the latter, resulted in two dominant low-molecular weight proteins, which were referred to as GA11 and GA20 . After purification of these proteins antibodies against the GA11 and GA20 protein of M . extorquens were obtained . Both proteins bound to the surface of PHB granules as revealed by immunoelectron microscopy of whole cells of M . extorquens and M . rhodesianum . With cells of the PHB-leaky mutants Mu 1 and Mu 11 no specific labeling was observed . The N-terminal amino acid sequences of the GA11 and the GA20 protein were determined . We found significant homologies between the sequences of the investigated strains . The use of oligonucleotide probes based on the N-terminal sequences of the GA20 protein from M . rhodesianum to identify the corresponding structural genes in various genomic libraries failed.

J Med Entomol, 1997 Jan, 34(1), 56 - 63
Cellular hemolymph response of Simulium vittatum (Diptera:Simuliidae) to intrathoracic injection of Onchocerca lienalis (Filarioidea:Onchocercidae) microfilariae; Cupp MS et al.; Hemolymph cellular changes in Simulium vittatum Zetterstedt in response to intrathoracic injection with Onchocerca lienalis Stiles were characterized by increased numbers of tissue fragments that contained cells embedded within a noncellular matrix . Cell numbers within the matrix fragments increased both for sham and microfilariae-injected files, indicating a blastogenic response to injection alone . Morphological characteristics of fixed, Giemsa-stained cells within these tissue fragments were most similar to those previously described for prohemocytes . The total population of freely circulating hemocytes collected 24 h after injection also responded to injection alone . Differential cell counts showed a complex pattern of changes that was influenced strongly by increased numbers of prohemocytes at 24 h in microfilariae-injected flies . Fat body fragments collected in hemocoel perfusates at 24 h were fewer when flies were maintained at 27 degrees C than at 21 degrees C . More fat body fragments were collected from microfilariae-injected flies than from control and sham-injected flies held at 27 degrees C . Injection of 1.25-5 micrograms of lipopolysaccharide per fly did not elicit similar hemolymph-associated matrix tissue and fat body changes, indicating that S . vittatum response to microfilaria infection and bacteria infection are likely to involve different mechanisms.

J Periodontal Res, 1997 Jan, 32(1 Pt 2), 166 - 71
Underlying mechanisms at the bone-surface interface during regeneration; Schwartz Z et al.; The goal of regenerative therapy around teeth and implants is to create a suitable environment in which the natural biological potential for functional regeneration of periodontal ligament and/or bone can be maximized . In order for the regenerative process to be successful, the following factors must be addressed: prevention of acute inflammation from bacteria, mechanical stability of the wound, creation and maintenance of blood clot-filled space, isolation of the regenerative space from undesirable competing tissue types, and the creation of a desirable surface chemistry, energy, roughness and microtopography that can directly influence cellular response, ultimately affecting the rate and quality of new tissue formation and, therefore, the regeneration process . This paper will review how surface characteristics (chemistry and roughness) can affect cell response and local factor production . To evaluate the effect of surface chemistry on cell proliferation and differentiation costochondral chondrocytes were grown on standard tissue culture plastic dishes sputter-coated with different materials . The results indicate that surface materials can elicit differential responses in cell metabolism and phenotypic expression in vitro . In a second study, the effect of varying titanium surface roughnesses on osteoblast-like cell behavior was examined . Surface roughness was found to alter osteoblast proliferation, differentiation and matrix production in vitro . In addition, production of PGE2 and TGF beta by these cells was also shown to increase with increasing surface roughness, indicating that substrate surface roughness also affects cytokine and growth factor production . The role of surface roughness in determining cellular response was further explored by comparing the response of osteoblasts grown on new and previously used surfaces . The results of these latter studies showed that cell proliferation, expression of differentiation markers and overall matrix production are not altered when cells are grown on used vs . virgin surfaces . This suggests the possibility that implants may be re-used, especially in the same patient, if they are appropriately treated . In this context, it should also be noted that rougher titanium surfaces may require more extensive cleaning procedures . From a global perspective, these studies provide some insight into how bone regeneration can be optimized in the presence of an implant or tooth root residing at the site of a bony defect . Since the new bone being produced, during regeneration, grows from a distal area toward the implant or tooth root surface, it is hypothesized that the osteoblasts growing on the surface of the implant may produce local factors that can affect the bone healing process distally . In short, it appears that the surface characteristics of an implant, particularly roughness, may direct tissue healing and, therefore, subsequent implant success in sites of regeneration by modulating osteoblast phenotypic expression.

J Periodontal Res, 1997 Jan, 32(1 Pt 2), 104 - 9
Molecular factors associated with compartmentalization of gingival immune responses and transepithelial neutrophil migration; Tonetti MS; PMN migration into the gingival sulcus is a tightly regulated process aimed at selectively increasing leukocyte availability at the site of bacterial plaque aggression, i.e . the superficial portion of the junctional epithelium . The evidence reviewed in this paper indicates that, besides the action of complement fragments, arachidonic acid metabolites, formyl peptides and other bacterial products, the establishment of a gradient of ICAM-1 expression across the junctional epithelium and the expression of IL-8 in its superficial layers probably represent important regulatory mechanisms leading to PMN migration into the gingival sulcus . Such mechanisms can be regulated by the autocrine and paracrine action of some pro-inflammatory cytokines and could, possibly be initiated by specific bacteria-keratinocyte interactions . The advantage of such a redundant regulatory mechanism leading to PMN transepithelial migration is probably related to the key role of the neutrophil in the maintenance of a local host-parasite equilibrium on one side, and on the tissue injury associated with PMN persistence or random migration within periodontal tissues on the other . Several investigations are in progress aimed at identifying the initial environmental stimuli leading to PMN recruitment into the gingival sulcus and at further exploring the important regulatory events.

J Periodontal Res, 1997 Jan, 32(1 Pt 2), 81 - 9
Genetic studies of syndromes with severe periodontitis and palmoplantar hyperkeratosis; Hart TC et al.; The Papillon-Lefevre and Haim Munk syndromes are characterized by the presence of both palmoplantar hyperkeratosis (PPK) and severe early onset periodontitis . It is the early onset periodontal disease component that distinguishes these from other more common forms of PPK . It has been proposed that the periodontal disease component may be a casual association in individuals with PPK . Genetic syndromes with palmoplantar keratosis and severe ealry onset periodontitis may be due to specific bacterial infections in individuals with PPK . Recently, keratin gene mutations have been identified in several conditions typified by palmoplantar keratosis . The present study sought to test the hypothesis that a keratin gene defect similar to those previously identified in other PPK conditions is responsible for the Haim Munk and the Papillon . Lefevre syndromes . We have performed genetic linkage studies to test for linkage between polymorphic DNA loci within 2 cytokeratin gene families and the disease phenotype in Haim Munk syndrome and Papillon-Lefevre syndrome . Families with individuals segregating for the Haim Munk syndrome and the Papillon-Lefevre syndrome were examined to determine disease status, and genotyped for microsatellite DNA markers closely linked to the acidic (type I) and the basic (type II) cytokeratin genes on chromosomes 12 and 17 . Genotype data were evaluated for microsatellite allele homozygosity in affected individuals . Results of these preliminary genetic studies suggest that the gene defect in Haim Munk syndrome is not due to a gene defect in either the type I or the type II keratin gene clusters . These findings suggest that Haim Munk syndrome may be genetically distinct from other more common forms of PPK that have been linked to the cytokeratin gene families, and suggest that mutations in genes other than keratin genes are responsible . Additional family studies are needed to confirm these preliminary findings.

Arch Toxicol Suppl, 1997, 19, 377 - 86
Lymphoma induction by heterocyclic amines in E mu-pim-1 transgenic mice; Sorensen IK et al.; The usefulness of transgenic E mu-pim-1 mice bearing in their genome the pim-1 oncogene supplemented with an upstream immunoglobulin enhancer and a downstream murine leukaemia virus long terminal repeat, as sensitive test organisms was studied in two short-term carcinogenicity studies . The mice were fed standard diet Altromin 1314 supplemented either with 0.03% 2-amino-1-methyl-6-phenylimidazo{4,5-b}pyridine (PhIP) for 7 months or with 0.03% 2-amino-3-methylimidazo{4,5-f}quinoline (IQ) for 6 months . PhIP and IQ are heterocyclic amines formed during cooking of meat and fish and are mutagenic to bacteria and cultured mammalian cells . PhIP is a potent mouse lymphomagen, while IQ is a liver, lung and forestomach carcinogen in mice . We found that transgenic E mu-pim-1 mice are highly susceptible to PhIP induced lymphomagenesis but do not respond to IQ treatment . PhIP feeding of E mu-pim-1 mice not only increased the total number of T-cell lymphomas but also decreased the latency time compared to either transgenic or wild-type controls . The effect was most pronounced in the treated female E mu-pim-1 mice, which showed a higher incidence of PhIP induced T-cell lymphomas than transgenic males and a strongly reduced latency period after PhIP treatment compared to non-transgenic mice . Our results suggest that the transgenic E mu-pim-1 mouse may be a useful model for short-term carcinogenicity screening of potential genotoxic carcinogens having the lymphoid system as target tissue . Carcinogens that do not target this tissue, like IQ, however will not be recognised.

J Comp Pathol, 1997 Jan, 116(1), 21 - 33
Demonstration of Helicobacter pylori-like organisms in the gastric mucosa of captive exotic carnivores; Jakob W et al.; Samples of gastric tissue from the cardiac, fundic and pyloric region of 30 carnivores comprising 12 tigers (Panthera tigris), 10 lions (Panthera leo), three pumas (Felis concolor), two leopards (Panthera pardus), one serval (Felis serval), one wolf (Canis lupus) and one hyena (Crocuta crocuta) kept at German zoological gardens were subjected to histopathological and immunohistochemical examination . Selected tissue specimens of 12 animals were examined also by electron microscopy . The purpose of this study was to determine the prevalence of Helicobacter-like organisms in carnivores and to record infection rates, degree of colonization and associated histopathological changes . Three morphologically different types of spiral-shaped bacteria were demonstrated . A Helicobacter pylori-like organism (HPLO) was found in 42% of the tigers and 90% of the lions examined . Large Helicobacter-like organisms (HLOs) were identified in three pumas, one serval, one hyena and in three lions (in the latter, in coexistence with HPLOs) . A third organism with a spiral periplasmic fibril (Helicobacter felis-like) was demonstrated in a wolf . The most striking histopathological finding associated with HPLO and HLO colonization was the formation of lymphoid follicles in the mucosa . Additional lymphoplasmacytic and neutrophilic infiltrates in the gastric mucosa were found in a number of tigers and lions infected with HPLOs, but none in the other carnivores infected with HLOs . From these results it is concluded that gastric bacteria similar or identical with H . pylori may also be an important cause of chronic gastritis in tigers and lions.

J Pathol, 1997 Jan, 181(1), 31 - 8
Apoptosis of human monocytes/macrophages in Mycobacterium tuberculosis infection; Placido R et al.; Tuberculosis (TB) is still a major health problem, both as a single disease entity and as a cofactor in AIDS . The interaction between macrophage and Mycobacterium tuberculosis (MTB) is a critical step in the establishment of an early chronic infection . This study analyses the capacity of MTB to induce apoptosis in cells obtained by broncho-alveolar lavage (BAL) from patients with reactive pulmonary tuberculosis and from AIDS patients with disseminated pulmonary tuberculosis . Apoptosis was increased three-fold in BAL cells obtained from patients with pulmonary tuberculosis and even more markedly in alveolar macrophages of MTB-infected AIDS patients, compared with controls . Apoptosis was analysed and characterized by propidium iodide (PI) incorporation, terminal deoxy transferase (TDT)-mediated dUTP-biotin nick end labelling (TUNEL), and tissue transglutaminase (tTG) expression . The MTB-macrophage interaction was also investigated in vitro by infecting monocyte-derived macrophages (MDM) with MTB (virulent strain H37Rv) . The induction of apoptosis by MTB required viable bacteria, was dose-dependent, and was restricted to H37Rv . Infection with either Mycobacterium avium complex (MAC) or HIV-1 and treatment with heat-killed MTB failed to induce apoptosis.

Life Sci, 1997, 60(10), 715 - 24
Identification of a membrane-bound carboxypeptidase as the mammalian homolog of duck gp180, a hepatitis B virus-binding protein; McGwire GB et al.; A unique membrane-bound carboxypeptidase was discovered and characterized in membrane fractions of human skin fibroblasts and the mouse monocyte-macrophage cell line J774A.1 and was partially purified from human placenta . Enzymatic characterization identified it as a new member of the regulatory B-type metallocarboxypeptidases, different from carboxypeptidases B, E, M, N and U . It is, however, similar to the newly described bovine carboxypeptidase D, suggested to be a homolog of duck gp180, a 180 kDa hepatitis B virus-binding protein . To prove this, a partial cDNA encoding a 20 kDa fragment of the human homolog of duck gp180 was expressed in bacteria and the recombinant protein was purified . Antibodies raised to the protein immunoprecipitated 94% or 72% of the low pH carboxypeptidase activity in human skin fibroblasts or J774A.1 cells and gave a 175 kDa protein band in Western blots . Thus, carboxypeptidase D is the mammalian homolog of duck gp180 and is distributed in a variety of different cell types.

Crit Rev Oral Biol Med, 1997, 8(1), 76 - 89
Cell-mediated immune system regulation in periodontal diseases; Mathur A et al.; The adaptive immune system consists of humoral and cell-mediated immunity . T-lymphocytes are the key components of cell-mediated immunity . CD4+ helper T-lymphocytes facilitate B-cells to differentiate and produce specific antibodies, whereas CD8+ cytotoxic T-lymphocytes kill virally infected cells . Periodontal diseases have been associated with a variety of imbalances in the regulation of immune responses . Changes in the ratios of peripheral blood CD4+ and CD8+ T-lymphocytes, depressed proliferative responses of peripheral blood lymphocytes, and increased frequency of CD45RO+ memory T-lymphocytes in diseased tissues have been reported in individuals with various forms of periodontal disease . While some studies have shown an increased frequency of gamma delta + T-cells in periodontal lesions, the role of gamma delta + T-cells in periodontal disease remains controversial . The ability of putative periodontopathic bacteria selectively to stimulate certain V beta-expressing T-cells is intriguing and could determine whether a CD4+ Th1 or a CD4+ Th2 cell response is elicited . The prominence of a particular subset of helper T-cells within the periodontal lesion could be a reflection of the stage and activity of the disease, or the types of bacteria present . Regardless, longitudinal studies of the involvement of T-cell subsets and cytokines in periodontal disease are clearly needed.

Insect Biochem Mol Biol, 1997 Jan, 27(1), 49 - 54
Structural organization and developmental expression of the protein isoaspartyl methyltransferase gene from Drosophila melanogaster; O'Connor MB et al.; A protein carboxyl methyltransferase activity (PCMT) with a specificity for age-damaged protein D-aspartyl and L-isoaspartyl residues (E.C . 2.1.1.77) has been identified and cloned in Drosophila . The Drosophila gene was localized by chromosome in-situ hybridization to region 83AB of the third chromosome . The methyltransferase coding sequence is distributed among four exons within a 1.4-kb segment of the genome; it predicts a polypeptide of 226 amino acids that is 55% identical to the mouse enzyme . When expressed in bacteria, the Drosophila protein exhibits PCMT activity . A single 1.4-kb Pcmt transcript is detected in RNA preparations from embryos, larvae, pupae and adults . The abundance of the transcript, which is lowest in larvae and highest in adults, parallels the specific activity of the enzyme measured in extracts from the same developmental stages . It has been proposed that the PCMT initiates the repair of structurally damaged cellular proteins . The constitutive expression of PCMT and the relatively high level of expression in postmitotic adult cells suggest that PCMT activity is required through development, but acquires additional significance in aging tissues.

Zentralbl Bakteriol, 1997 Jan, 285(2), 182 - 94
Pyrolysis mass spectrometry: data processing in classification studies; Magee JT; A scheme for numerical processing of pyrolysis mass spectrometry (Py-MS) data is detailed, along with methods for combining the results of conventional phenotypic and Py-MS taxonomic surveys . The importance of this combined approach in polyphasic taxonomy is emphasised; it yields data on cell composition and the nutritional and physiological interactions of strains with their environment . Large collections of strains can be surveyed rapidly and economically, yielding presumptive classifications, which may then be confirmed with a few representative strains in more demanding, difficult and expensive approaches . An objective, non-arbitrary method of establishing suitable cut-off points to delineate group structures in dendrograms is also described.

Int J Sports Med, 1997 Jan, 18(1), 35 - 9
Lactate and ammonia concentration in blood and sweat during incremental cycle ergometer exercise; Ament W et al.; It is known that the concentrations of ammonia and lactate in blood increase during incremental exercise . Sweat also contains lactate and ammonia . The aim of the present study was to investigate the physiological response of lactate and ammonia in plasma and sweat during a stepwise incremental cycle ergometer exercise test in ten subjects . During this test lactate and ammonia were measured in blood obtained from the earlobe and in sweat collected in a bag attached to the back of the subject . At the end of each interval this bag was emptied for measuring lactate and ammonia . A disproportional increase in the concentration of lactate and ammonia in blood was found, in sweat a disproportional decrease . The lactate concentrations in sweat were higher than those in blood . We hypothesise that lactate in sweat is produced from glycogen granules of the clear cell of the eccrine gland . This lactate production results in acidification of sweat, which facilitates the diffusion of ammonia from eccrine duct cell to duct lumen . It is uncertain how far duct cell ammonia originates from plasma, the duct cell itself might produce ammonia . Part of the ammonia in sweat could come from the breakdown of urea by skin bacteria.

Curr Opin Hematol, 1997 Jan, 4(1), 59 - 66
Function and clinical use of interleukin-12; Trinchieri G; Interleukin-12 is a heterodimeric cytokine produced by phagocytic cells, professional antigen-presenting cells such as dendritic cells and skin Langerhans cells, and B cells . Interleukin-12 production is induced by bacteria, intracellular pathogens, fungi, viruses, or their products in a T-cell-independent pathway or a T-cell-dependent pathway, the latter mediated through CD40 ligand-CD40 interaction . Interleukin-12 is produced rapidly after infection and acts as a proinflammatory cytokine eliciting production of interferon gamma, by T and natural killer cells, which activates phagocytic cells . The production of interleukin-12 is strictly regulated by positive and negative feedback mechanisms . If interleukin-12 and interleukin-12-induced interferon gamma are present during early T-cell expansion in response to antigen, T-helper type-1 cell generation is favored and generation of T-helper type-2 cells is inhibited . Thus interleukin-12 is also a potent immunoregulatory cytokine that promotes T-helper type-1 differentiation and is instrumental in the T-helper type-1-dependent resistance to infections by bacteria, intracellular parasites, fungi, and certain viruses . By inhibiting T-helper type-2 cell response, interleukin-12 has a suppressive effect on allergic reactions; by promoting T-helper type-1 responses it participates in the immunopathology responsible for several organ-specific autoimmune diseases . Viruses inducing a permanent or transient immunodepression, such as HIV and measles, may act, in part, by suppressing interleukin-12 production . Because of its ability to enhance resistance to several infectious diseases and to act as an adjuvant in vaccination, and because of its powerful antitumor effect in vivo, interleukin-12 is currently in clinical trials in cancer patients and HIV-infected patients, and it is being considered for therapeutic use in other diseases.

Curr Opin Hematol, 1997 Jan, 4(1), 32 - 40
Killing mechanisms of cytotoxic lymphocytes; Berke G; Killer lymphocytes are primary immune effectors of virus, certain bacteria, and tumor immunity and play a role in autoimmunity and transplant rejection . This article reviews progress in deciphering the mechanisms by which they kill target cells through induction of apoptosis by either the secretory, perforin/granzyme-based pathway or the nonsecretory pathway, (i.e., by triggering the cell-surface death receptor Fas (CD95) by the membrane-bound Fas ligand of the killer).

J Clin Periodontol, 1997 Jan, 24(1), 57 - 64
The detection of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Prevotella intermedia using an ELISA in an adolescent population with early periodontitis; Clerehugh V et al.; The aim of the study was to compare the occurrence and levels of A . actinomycetemcomitans, P . gingivalis, and P . intermedia in the subgingival plaque from sites with and without early periodontitis in adolescents using an ELISA . 47, 15- to 16-year-old adolescents (39 Indo-Pakistani, 8 white Caucasian) were examined for clinical attachment level, probing depth, supragingival plaque, subgingival calculus and bleeding on probing on the mesio-buccal and disto-buccal aspects of the 1st molars and the incisors . Based on the clinical data, 2 sites per subject were selected for subgingival plaque sampling 3 weeks later: in 32 subjects with loss of attachment > or = 1 mm, a diseased site (D) and a healthy comparison control site (C) were sampled; in 15 subjects in whom loss of attachment had not yet developed, 1 of the upper molar sites was selected, called the at-risk site (R), together with a C site . The presence and levels of A . actinomycetemcomitans, P . gingivalis, and P . intermedia were determined using an ELISA . The loss of attachment subgroup had significantly more pockets > or = 4 mm, subgingival calculus and bleeding on probing (p < 0.05) . Significantly more of the D than C sites had P . gingivalis both at detectable and at measurable levels (p < 0.05) . In subjects who had no loss in clinical attachment levels, fewer sampled sites harboured any of the suspected periodontopathogens investigated, and no significant differences were found between the R or C sites (p > 0.05) . Although there was a significantly higher prevalence and extent of loss of attachment > or = 1 mm in the Indo-Pakistani subjects compared with the Caucasians (p < 0.05), no differences could be identified in the distribution of the bacteria . It is concluded that monitoring of the subgingival plaque may be useful in studies of early periodontitis in adolescents, and the role of P . gingivalis needs to be elucidated in prospective longitudinal investigations.

Br J Dermatol, 1997 Jan, 136(1), 60 - 5
Bacillary angiomatosis: presentation of six patients, some with unusual features; Schwartz RA et al.; Bacillary angiomatosis (BA) is an unusual systemic vascular proliferation seen predominantly in patients with the acquired immunodeficiency syndrome . These vascular lesions are probably due to infection with a Bartonella species, most often B . henselae and, in some patients, B . quintana . BA is treatable and often curable, but without therapy, may be life-threatening . Clinically, the lesions, when superficial, are said to often resemble pyogenic granulomas, appearing polypoid histologically with an epidermal collarette . We now report six patients, three of whom showed lesions of BA morphologically and histologically distinct from the other patients reported to date . Two patients lesions appeared clinically as violaceous plaques and tumours resembling Kaposi's sarcoma; one of them had lesions histologically reminiscent of a papular angiokeratoma; and the other had lesions histologically suggestive of a combination of Kaposi's sarcoma and BA . Another patient presented with soft subcutaneous nodules which histologically showed extensive acute inflammation characteristic of an acute abscess, but which also displayed proliferating dilated small blood vessels with bulbous endothelial cells adjacent to numerous bacteria and also containing them . The Grocott-methenamine silver stain and the Warthin-Starry stain showed the organisms to better advantage in lesions of all six patients, although bacteria were also evident with the haematoxylin and eosin, periodic acid-Schiff and alcian blue stains.

New Microbiol, 1997 Jan, 20(1), 63 - 8
In vitro virulence factors of Arcobacter butzleri strains isolated from superficial water samples; Musmanno RA et al.; Eighteen isolates of A . butzleri from river water samples were examined for their biotype, serogroup and putative virulence characteristics . Toxin profiles based on cytotonic, cytotoxic and cytolethal distending factors were determined after analysis responses in Vero and CHO cells Adhesivity and invasivity tests were performed on HeLa and Intestine 407 cells . Six biotypes and five serogroups were determined in our isolates . All strains but one induced cytotoxic effects on cells in culture . The cytotoxic negative strain caused elongation of CHO cells (a cytotonic-like effect) . This strain was the only one which adhered to cells in vitro . Invasiveness was never observed . Our results show a phenotypic heterogeneity of arcobacters isolated from environmental sources, and indicate that some strains could be potentially virulent.

Plant Mol Biol, 1997 Jan, 33(1), 181 - 5
Chloroplastic isoforms of DnaJ and GrpE in pea; Schlicher T et al.; The folding and assembly of proteins in cells often requires the assistance of molecular chaperones such as the Hsp70 and Hsp60 heat shock proteins . Hsp70 chaperones cooperate with DnaJ and GrpE homologues to ensure a productive folding cycle . In this study we describe the gene for the first chloroplast localized DnaJ homologue and present evidence that the gene product is at least partially associated with the inner envelope membrane . Immunoblot analysis also provides evidence for the presence of a GrpE homologue in plastids.

Aust Vet J, 1997 Jan, 75(1), 56 - 9
Failure of passive transfer in foals: incidence and outcome on four studs in New South Wales; Tyler-McGowan CM et al.; OBJECTIVE: To determine the regional incidence and effectiveness of treatment of failure of passive transfer (FPT) in foals . DESIGN: A study of disease incidence . ANIMALS: Eighty-eight foals and 57 mares from four studs in the practice area of the Rural Veterinary Centre were tested . PROCEDURE: Foals were tested for their serum IgG and total serum protein (TSP) concentration within the first 72 hours of life . Colostrum was collected from mares and specific gravity determined . FPT and partial failure of passive transfer (PFPT) of immunoglobulins was diagnosed when serum IgG concentrations were < 4 g/L and 4 to 8 g/L respectively . Owners of foals diagnosed with FPT were offered treatment with 1 to 2 L plasma (TSP > 70 g/L); 9 (64%) of the affected foals were treated . RESULTS: Fourteen foals (16%) had FPT whereas 15 (17%) had PFPT . There were significant differences between the mean TSP concentration in foals with FPT (42.6 +/- 4.2 g/L), PFPT (48.1 +/- 3.9 g/L) and those acquiring adequate passive immunity (58.9 +/- 5.5 g/l) (p < 0.01) . Sixteen (29%) mares had pre-suck colostral specific gravity < 1.060 and 12 (71%) foals raised by these mares had FPT or PFPT . The incidence of severe disease (categorised by a sepsis score > 11, positive culture of bacteria from blood or disease requiring hospitalisation) in all foals in the first 2 months of life was 10% . However, none of the nine foals with FPT that received plasma experienced severe disease . In contrast, foals with PFPT had an increased susceptibility to severe disease (p < 0.001) when compared with normal foals . CONCLUSION: Treatment of foals with FPT may reduce the subsequent incidence of severe disease . Pre-suck colostral specific gravity and foal TSP may be used to predict the likelihood of FPT and PFPT . Even though the number of foals studied is small the results highlight the importance of optimal management practices in reducing the incidence of FPT and disease associated with this process.

Aust Vet J, 1997 Jan, 75(1), 18 - 20
Atresia of the external acoustic meatus in a dog; Simpson D; A 3 year old, female great Dane with atresia of the right external ear canal had recurrent episodes of ear pain . Radiography revealed absence of air in the right external acoustic meatus, thickened bone of the right tympanic bulla and increased radiodensity of the chamber of the bulla . Total ear canal ablation and lateral bulla osteotomy were performed . The superficial portion of the external ear canal was absent and the deeper segment of the vertical ear canal began as a blunt ended cartilage tube . A patent lumen in the existent portion of the external ear canal and the tympanic bulla contained wax, hair and exfoliated squames . The tympanic membrane was not intact . No bacteria were cultured from the contents of the external and middle ear . The dog responded well to surgery and was free of pain 11 months later . Failure to surgically correct atresia of the ear canal in young dogs may allow the accumulation of cellular and sebaceous debris with subsequent involvement of the middle ear in an inflammatory response.

FASEB J, 1997 Jan, 11(1), 68 - 76
A superfamily of conserved domains in DNA damage-responsive cell cycle checkpoint proteins; Bork P et al.; Computer analysis of a conserved domain, BRCT, first described at the carboxyl terminus of the breast cancer protein BRCA1, a p53 binding protein (53BP1), and the yeast cell cycle checkpoint protein RAD9 revealed a large superfamily of domains that occur predominantly in proteins involved in cell cycle checkpoint functions responsive to DNA damage . The BRCT domain consists of approximately 95 amino acid residues and occurs as a tandem repeat at the carboxyl terminus of numerous proteins, but has been observed also as a tandem repeat at the amino terminus or as a single copy . The BRCT superfamily presently includes approximately 40 nonorthologous proteins, namely, BRCA1, 53BP1, and RAD9; a protein family that consists of the fission yeast replication checkpoint protein Rad4, the oncoprotein ECT2, the DNA repair protein XRCC1, and yeast DNA polymerase subunit DPB11; DNA binding enzymes such as terminal deoxynucleotidyltransferases, deoxycytidyl transferase involved in DNA repair, and DNA-ligases III and IV; yeast multifunctional transcription factor RAP1; and several uncharacterized gene products . Another previously described domain that is shared by bacterial NAD-dependent DNA-ligases, the large subunits of eukaryotic replication factor C, and poly(ADP-ribose) polymerases appears to be a distinct version of the BRCT domain . The retinoblastoma protein (a universal tumor suppressor) and related proteins may contain a distant relative of the BRCT domain . Despite the functional diversity of all these proteins, participation in DNA damage-responsive checkpoints appears to be a unifying theme . Thus, the BRCT domain is likely to perform critical, yet uncharacterized, functions in the cell cycle control of organisms from bacteria to humans . The carboxyterminal BRCT domain of BRCA1 corresponds precisely to the recently identified minimal transcription activation domain of this protein, indicating one such function.

Microb Pathog, 1997 Jan, 22(1), 47 - 57
Heparin-mediated inhibition of Chlamydia psittaci adherence to HeLa cells; Gutierrez-Martin CB et al.; The adherence of human strains of Chlamydia trachomatis has been recently shown to be inhibitable by heparin and heparitinase, leading to the proposal that Chlamydia binding to host cells may be mediated by a glycosaminoglycan (GAG)-dependent mechanism . We here describe the adherence of the guinea-pig pathogen, Chlamydia psittaci GPIC, to HeLa cells, which was measured by cytofluorometry with chlamydiae whose DNA was fluorescently labelled . Adherence could be inhibited by heat or trypsin pretreatment of the bacteria, and binding was much faster at 37 degrees C (reaching a plateau within 1 h) than 4 degrees C . Little binding remained when host cells were pre-fixed with paraformaldehyde, suggesting that host cell receptor mobility may be required for effective adherence . Visualization by confocal microscopy confirmed that the bacteria were at or near the host cell surface during the entire time-course of these experiments . Adherence increased as a function of pH between pH 6 and pH 8.0-8.5 . Both adherence and infection of HeLa cells could be inhibited with heparin when the adherence step was performed at 4 degrees C, but only infection was inhibited when the adherence step was performed at 37 degrees C, even though heparitinase could block adherence at either 4 degrees C or 37 degrees C . Even at 4 degrees C, heparin-mediated inhibition was significantly lower at pH 8 than pH 7.4, suggesting that GAG-independent mechanisms may play a role in the higher adherence observed at basic pH . These results therefore demonstrate that a GAG-dependent adherence step may be operative in C . psittaci, and raise the possibility that other adherence mechanisms may also contribute to binding by this chlamydial strain . Furthermore, they suggest that there may not be a strict correlation between C . psittaci adherence and the ability to cause productive infections.

Dermatology, 1997, 194(1), 32 - 5
Pityriasis rotunda: a survey of 42 cases observed in Sardinia, Italy; Aste N et al.; BACKGROUND: Pityriasis rotunda (PR) is an uncommon dermatosis characterized by multiple, round or oval, sharply demarcated scaling patches that are dyschromic and asymptomatic . It has been described in Japanese and in blacks, usually in association with certain infective or malignant systemic diseases . OBJECTIVE: The aim of this study is to further clarify this rare entity which in Italy seems to be confined to the island of Sardinia . METHODS: We studied 42 Sardinian patients, 22 males and 20 females, in an age range of 3-32 years . In 29 cases, the disease involved more than one family member . The patients were observed in Cagliari, the capital city of Sardinia . RESULTS: Bacterial, viral and fungal investigation yielded negative results . Haematochemical and immunological examination and thyroid, hypophyseal and adrenal hormones did not reveal any alterations . No systemic pathologies were found associated with the disease . CONCLUSIONS: The cases studied by us and those previously reported seem to indicate the presence of two distinct types of PR with significant prognostic differences.

J Periodontol, 1997 Jan, 68(1), 39 - 44
Fusobacterium nucleatum T18 aggregates human mononuclear cells and inhibits their PHA-stimulated proliferation; Kinder Haake S et al.; In previous studies Fusobacterium nucleatum has been shown to induce either stimulatory or inhibitory effects on human mononuclear cells . We examined the interaction of human mononuclear cells with human and cynomolgus monkey strains of F . nucleatum . Peripheral blood mononuclear cells (PBMCs) isolated from normal donors were aggregated in the presence of cells of F . nucleatum but not control bacteria . The aggregation of PBMCs and F . nucleatum T18 was inhibited by either L-arginine, L-lysine, or pretreatment of the bacterial cells with heat, but was unaffected by the presence of sugars or normal human serum . Strain T18 aggregated purified T-cells and monocytes at approximately equal concentrations . When F . nucleatum T18 was incubated with PHA-stimulated PBMCs, DNA synthesis in the PBMCs was significantly inhibited and detection of IL-2R alpha on the PBMCs was reduced . These studies indicate that F . nucleatum aggregates PBMCs, and that this interaction is associated with both an inhibition of PBMC proliferation and a decrease in IL-2 receptor expression . The ability of F . nucleatum to inhibit mononuclear cell proliferation may be significant in the pathogenesis of periodontal diseases.

J Periodontol, 1997 Jan, 68(1), 12 - 7
Alveolar bone loss in rats infected with a strain of Prevotella intermedia and Fusobacterium nucleatum isolated from a child with prepubertal periodontitis; Yoshida-Minami I et al.; Prevotella intermedia and fusobacterium nucleatum are associated with various forms of periodontal disease . The purpose of the present study was to infect the clinical isolates of these periodontopathic bacteria and to induce a significant loss of alveolar bone in specific pathogen-free (SPF) rats in the absence of ligatures . P . intermedia YKD8 and F . nucleatum YKZ5 were isolated from a prepubertal periodontitis patient, while P . gingivalis MWB13 was from a patient with juvenile periodontitis . At first, SPF Sprague-Dawley rats (70 days of age, male) were infected with A . viscosus Ny1R and subsequently superinfected with P . gingivalis MWB13, P . intermedia YKD8, or F . nucleatum YKZ5, respectively . The control group was infected with A . viscosus Ny1R alone . All rats were killed and periodontal bone levels were assessed morphometrically 135 days after the first infection with A . viscosus . P . intermedia YKD8 was recovered frequently from rats, with serum antibody levels remaining highly elevated throughout the experiment . Significant loss of alveolar bone was found in rats infected with P . intermedia YKD8, the virulence of which was equivalent to that of P . gingivalis MWB13 . F . nucleatum YKZ5 also induced alveolar bone loss, but not significantly when compared with rats infected with A . viscosus Ny1R alone.

J Anim Sci, 1997 Jan, 75(1), 159 - 69
Evaluation of the 15N-isotope dilution technique for determining the recovery of endogenous protein in ileal digesta of pigs: effect of the pattern of blood sampling, precursor pools, and isotope dilution technique; Lien KA et al.; The 15N-enrichments (atom percentage excess) were determined in the plasma free amino acids of blood samples taken at the time of feeding and in samples taken hourly and pooled over 12 h, as well as in ileal digesta, crude mucin, and bacteria collected at the distal ileum of pigs fed barley while continuously administered {15N}leucine intravenously . The branched-chain amino acids were the only indispensable amino acids to exhibit incorporation of 15N (P < .05) . All dispensable amino acids exhibited some incorporation . Enrichments in free leucine and alanine were higher (P < .02) in blood samples taken at the time of feeding, compared to those in pooled blood samples, resulting in an underestimation of the endogenous ileal recoveries of these amino acids . Enrichments in amino acids in crude mucin were usually similar to those in pooled plasma samples, providing some support for the use of plasma free amino acids to estimate enrichments in endogenous amino acids in ileal digesta . Enrichments in bacteria were not different (P > .05) from those in ileal digesta . The recoveries of endogenous amino acids in ileal digesta determined with the {15N}leucine and 15N-amino acid dilution techniques demonstrate the overestimation of these criteria with the 15N-isotope dilution technique, applied in its current form, and suggest that modifications in the composition of endogenous protein can occur when pigs are fed protein-containing diets . These study supports the use of 15N-isotope dilution techniques, with modifications, for determining the recovery of endogenous protein in ileal digesta of pigs fed protein-containing diets.

Exp Parasitol, 1997 Jan, 85(1), 55 - 62
Trichuris suis: thiol protease activity from adult worms; Hill DE et al.; Trichuris suis, the whipworm of swine, causes anemia, weight loss, anorexia, mucohemorrhagic diarrhea, and death in heavy infections . A zinc metalloprotease has been suggested to play a role in the severe enteric pathology associated with infection and the infiltration of opportunistic bacteria into deeper tissues in the swine colon . In this study, a thiol protease from gut extracts of adult T . suis and from excretory/secretory components (E/S) of adult worms was characterized using fluorogenic peptide substrates and protein substrate gels . The protease cleaved the fluorogenic substrate Z-Phe-Arg-AMC, and this cleavage was completely inhibited by the thiol protease inhibitors E-64, leupeptin, Z-Phe-Ala-CH2F, and Z-Phe-Arg-CH2F . Gelatin substrate gels and fluorescence assays using both the gut and the stichosome extracts and E/S revealed enhanced activity when 2 mM dithiothreitol or 5 mM cysteine was included in the incubation buffer, and optimal activity was seen over a pH range of 5.5 to 8.5 . Incubation of gut extracts or E/S material with inhibitors of aspartic, serine, or metalloproteases had no effect on the cleavage of Z-Phe-Arg-AMC . Thiol protease activity was found in extracts of gut tissue but not in the extracts of stichocytes of adult worms . N-terminal amino acid sequencing of the protease revealed sequence homologies with cathepsin B-like thiol protease identified from parasitic and free-living nematodes.

Lett Appl Microbiol, 1997 Jan, 24(1), 23 - 6
A rapid plate assay for screening L-asparaginase producing micro-organisms; Gulati R et al.; A pH and dye-based fast procedure for screening L-asparaginase producing micro-organisms is reported . The procedure is suitable for bacterial and fungal screening . The results are obtained within 24 and 48 h for bacteria and fungi, respectively . The results correlate with quantitative estimations in culture broths.

J Exp Biol, 1997 Jan, 200(Pt 1), 1 - 8
Bafilomycins and concanamycins as inhibitors of V-ATPases and P-ATPases; Drose S et al.; Bafilomycins and concanamycins, two groups of the plecomacrolide-defined class of macrolide antibiotics, have recently been recognized as important tools for studying the physiological role of vacuolar-type, proton-translocating ATPases (V-ATPases) and ATPases with phosphorylated states (P-ATPases) in animal and plant cells as well as in yeast, fungi and bacteria . The following review will give an account of the classification and function of these antibiotics.

Genetics, 1997 Jan, 145(1), 197 - 205
An allelic series of blue fluorescent trp1 mutants of Arabidopsis thaliana; Rose AB et al.; Nine blue fluorescent mutants of the flowering plant Arabidopsis thaliana were isolated by genetic selections and fluorescence screens . Each was shown to contain a recessive allele of trp1, a previously described locus that encodes the tryptophan biosynthetic enzyme phosphoribosylanthranilate transferase (PAT, called trpD in bacteria) . The trp1 mutants consist of two groups, tryptophan auxotrophs and prototrophs, that differ significantly in growth rate, morphology, and fertility . The trp1 alleles cause plants to accumulate varying amounts of blue fluorescent anthranilate compounds, and only the two least severely affected of the prototrophs have any detectable PAT enzyme activity . All four of the trp1 mutations that were sequenced are G to A or C to T transitions that cause an amino acid change, but in only three of these is the affected residue phylogenetically conserved . There is an unusually high degree of sequence divergence in the single-copy gene encoding PAT from the wild-type Columbia and Landsberg erecta ecotypes of Arabidopsis.

Nucleic Acids Res, 1997 Jan 1, 25(1), 263 - 4
The ribonuclease P database; Brown JW; Ribonuclease P is responsible for the removal of leader sequences from tRNA precursors . Ribonuclease P is a ribonucleoprotein, and in bacteria the RNA subunit alone is catalytically active in vitro, i.e . it is a ribozyme.The Ribonuclease P Database is a compilation of ribonuclease P sequences, sequence alignments, secondary structures, three-dimensional models, and accessory information, available via the World Wide Web.

J Clin Gastroenterol, 1997 Jan, 24(1), 18 - 20
Arthralgia as an early extraintestinal symptom of Whipple's disease . Report of five cases; Schilling D et al.; Five patients with Whipple's disease all suffered from arthralgia for a long time (15 years in one case) before developing gastrointestinal or other symptoms . In all patients, arthralgia was seronegative, and there was no evidence of joint destruction . Arthralgias were symmetric and migrating . Whipple's disease is part of the differential diagnosis of enteropathic arthralgia . Thereby, the polymerase chain reaction can be a helpful tool to prove Whipple's disease in difficult differential diagnosis.

Proc Soc Exp Biol Med, 1997 Jan, 214(1), 69 - 75
Psychoactive cannabinoids increase mortality and alter acute phase cytokine responses in mice sublethally infected with Legionella pneumophila; Smith MS et al.; Marijuana contains both psychoactive and nonpsychoactive cannabinoids which have varying effects on the immune response system . Previous studies with delta-9-tetrahydrocannabinol (THC), the major psychoactive component of marijuana, showed that this substance augmented the susceptibility of mice to infection with the opportunistic pathogen Legionella pneumophila . The present study compared the enhancement of Legionella-induced mortality in mice due to two other major of marijuana components, cannabinol and cannabidiol, as well as the synthetic psychoactive cannabinoid CP 55,940 . Inbred BALB/c mice, relatively resistant to infection with Legionella, were given the marijuana component 1 day before and 1 day after a sublethal intravenous infection with Legionella . Unlike the effect of THC, an 8 mg/kg dose of either cannabinol or cannabidiol did not affect mortality of the mice sublethally infected with Legionella . Mice given a 16 mg/kg dose of these components of marijuana, however, showed a slight to moderately increased mortality following the sublethal infection with Legionella . In contrast, a dose of 6 mg/kg of the synthetic psychoactive cannabinoid CP 55,940 given 1 day before and 1 day after infection with Legionella resulted in about 50% of the animals dying, the same level of augmentation of lethality induced by THC . Liver, spleen, and lung tissues were removed from the surviving mice 24 hr after the second THC injection and tested for the presence of viable Legionella using a standard CFU assay . The mice injected with THC before and after infection showed significantly higher levels of bacteria in their lung than mice that were not given any cannabinoid but were infected sublethally with the Legionella . Mice injected with the other cannabinoids, including the synthetic cannabinoid, showed a much smaller increase in the number of Legionella in their lung when infected with Legionella and treated with the drug . The number of bacteria recovered from the kidney and liver of the mice regardless of treatment with a cannabinoid, including with THC, did not show significant changes . RNA isolated from the spleen of the THC- and Legionella-treated animals was examined to determine at the molecular level whether acute phase pro-inflammatory cytokines (i.e., IL-1, IL-6 and TNF-alpha) were altered following drug treatment and infection, since previous studies had shown there were increased serum levels of these cytokines in the mice . It was found that the mRNA levels for IL-1 remained generally constant regardless of whether the infected animals were treated with a cannabinoid . However, the mRNA level for IL-6 was markedly increased following treatment of the infected animals with THC, suggesting the possible involvement of this pro-inflammatory cytokine in increased mortality . The mRNA level for TNF-alpha was generally low and not significantly altered in the drug treated animals . Mice given other cannabinoids, including cannabinol and cannabidiol, as well as the synthetic CP 55,940, showed no significant change in the level of mRNA for any of the cytokines tested.

J Clin Invest, 1997 Jan 1, 99(1), 77 - 87
Secretion of proinflammatory cytokines by epithelial cells in response to Chlamydia infection suggests a central role for epithelial cells in chlamydial pathogenesis; Rasmussen SJ et al.; Chlamydia species infect epithelial cells at mucosal surfaces, and are major causes of sexually transmitted diseases . Infection is characterized by inflammation which is exacerbated upon reinfection, ultimately leading to tissue damage and scarring . Although central for the development of disease manifestations, little is known about the mechanisms that initiate and sustain the inflammatory response to Chlamydia . Infection of cervical and colonic epithelial cells with Chlamydia trachomatis and Chlamydia psittaci is shown in the present studies to upregulate mRNA expression and secretion of the proinflammatory cytokines IL-8, GRO alpha, GM-CSF, and IL-6 . In contrast to the rapid, but transient, cytokine induction following infection with other invasive bacteria, the epithelial cytokine response to Chlamydia was delayed until 20-24 h after infection, persisted throughout the chlamydial growth cycle (2-4 d), and required bacterial protein synthesis . Moreover, epithelial cell lines and primary endocervical epithelial cells released IL-1alpha after Chlamydia infection, and increased secretion of the proinflammatory cytokines could be inhibited by anti-IL-1alpha . This suggests that IL-1alpha, released following lysis of infected epithelial cells, may amplify the inflammatory response by stimulating additional cytokine production by noninfected neighboring cells . These findings suggest a novel pathophysiologic concept wherein the acute host response to Chlamydia at mucosal surfaces is primarily initiated and sustained by epithelial cells, the first and major targets of chlamydial infection.

J Clin Invest, 1997 Jan 1, 99(1), 19 - 23
Mutation of human keratin 18 in association with cryptogenic cirrhosis; Ku NO et al.; Mutations in 11 of the more than 20 keratin intermediate filaments cause several epidermal and oral associated diseases . No disease-associated mutations have been described in keratin 8 or 18 (K8/18) which are the major keratin pair in simple-type epithelia, as found in the liver, pancreas, and intestine . However, transgenic mice that express mutant keratin 18 develop chronic hepatitis, and have an increased susceptibility to drug-induced hepatotoxicity . Also, ectopic expression of epidermal K14 in mouse liver results in chronic hepatitis, and disruption of mouse K8 leads to embryo lethality with extensive liver hemorrhage . We tested if patients with liver disease of unknown cause may harbor mutations in K18 . We describe a his127-->leu (H127L) K18 mutation in a patient with cryptogenic cirrhosis that is germline transmitted . The K18 H127L isolated from the liver explant, or after expression in bacteria, showed an altered migration on two-dimensional gel analysis as compared with normal human liver or bacterially expressed K18 . Electron microscopy of in vitro assembled K18 H127L and wild type K8 showed an assembly defect as compared with normal K8/18 assembly . Our results suggest that mutations in K18 may be predispose to, or result in cryptogenic cirrhosis in humans.

Clin Exp Immunol, 1997 Jan, 107(1), 158 - 65
IgG and IgA subclass mRNA-bearing plasma cells in periodontitis gingival tissue and immunoglobulin levels in the gingival crevicular fluid; Takahashi K et al.; The humoral immune response, especially the production of IgG and IgA, is considered to have a protective role in the pathogenesis of periodontal disease, but the precise mechanisms are still unknown . In order to determine local IgG and IgA production, we investigated the presence of human IgG and IgA subclass mRNA-bearing plasma cells within periodontal tissue by in situ hybridization using digoxigenin-labelled oligonucleotide probes in 24 gingival biopsy samples (pocket depth > or = 5 mm) which were obtained from eight patients with adult periodontitis . Furthermore, we examined IgG and IgA subclass proteins and digested IgA1 Fab portions in the gingival crevicular fluid (GCF), corresponding to the sites from which the tissues were taken, by ELISA . IgG and IgA subclass mRNA-expressing cells were detected in all serial formalin-fixed/paraffin-embedded gingival tissue sections sampled . Plasma cells showed strong cytoplasmic staining with a high contrast and a good retention of morphology with these probes . IgG1 mRNA-expressing cells were predominant (mean 63%) and IgG2 mRNA-expressing cells were present at around 23% of total IgG plasma cells, while IgG3 and IgG4 mRNA-expressing cells were present to a much lesser extent (3% and 10%, respectively) . Similar proportions of IgG subclass proteins in GCF were detected, which were also consistent with 'normal' serum levels . In terms of IgA subclass, IgA1 mRNA-positive cells were predominant (mean 65.1%, P < 0.001) . In contrast, IgA2 protein in the GCF samples were detected at higher concentrations than IgA1 (P < 0.001) . The ratio of total IgG to IgA mRNA-positive plasma cells was approximately 7.5:1 . There was a good correlation between the amounts of IgG subclass proteins in GCF and the number of IgG subclass mRNA-positive cells in the same sites, but not between IgA subclass proteins and the number of IgA subclass mRNA-positive cells . These data suggest that IgG and IgA subclass proteins can be locally produced in the periodontitis gingiva . In addition, as we detected IgA1 Fab fragments in GCF, this is further confirmation that secreted IgA1 protein in GCF may be digested by periodontal bacteria.

Clin Exp Immunol, 1997 Jan, 107(1), 76 - 82
Cytotoxic effect of autocrine and macrophage-derived nitric oxide on cultured rat mesangial cells; Hruby Z et al.; Expression of the inducible form of nitric oxide synthase (iNOS) has been found to be up-regulated in cytokine-stimulated mesangial cells (MC) and in experimental glomerulonephritis . Since direct toxicity of nitric oxide (NO) has been implicated in damage of bacteria, neoplastic and intact pancreatic cells, we investigated whether NO is cytotoxic to cultured MC, which may be relevant to pathogenesis of glomerular injury . MC isolated from rat glomeruli generated substantial amounts of nitrite, the stable NO end-product, when cells were stimulated with IL-1beta and tumour necrosis factor-alpha (TNF-alpha) . Total DNA synthesis was significantly reduced in the presence of IL-1beta and TNF-alpha, and this effect was completely reversed by N(G)-monomethyl-L-arginine (L-NMMA), an inhibitor of iNOS . Stimulation of MC with IL-1beta and TNF-alpha caused remarkable toxicity to these cells, measured by the MTT test (3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide cleavage, specific cytotoxicity 41.5 +/- 20.3%), and much less prominent MC lysis (3H-thymidine release, specific cytolysis 11.5 +/- 5.3%) . Toxic effects of cytokines were fully reversible by the iNOS inhibitor . Lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma), but not IL-1beta and TNF-alpha, induced rat peritoneal macrophages to produce large amounts of nitrite . In co-culture, such prestimulated macrophages had significantly cytotoxic (MTT test 62.9 +/- 19.9%) and cytolytic (3H-thymidine release 57.9 +/- 13.8%) effects on MC . Again, this toxicity was totally inhibited in the presence of L-NMMA . We conclude from these results that cytokine-stimulated generation of NO by MC or macrophages is directly toxic to MC, and may play a role in pathogenesis of glomerular injury involving mesangiolysis.

Can J Vet Res, 1997 Jan, 61(1), 1 - 7
Detection of antibodies against Actinobacillus pleuropneumoniae serotype 5 using an inhibition enzyme immunoassay; Stenbaek EI et al.; An inhibition enzyme immunoassay (EIA) for detection of antibodies against A . pleuropneumoniae serotype 5 (App-5) in pig sera, based on the inhibition of the binding of an App-5 specific monoclonal antibody was established . The monoclonal antibody (MAb 210-F11) was found to be directed against an epitope on the O-chain of App-5 LPS . In the inhibition EIA, highly purified App-5 LPS was used to coat microtitre plates . Serial dilutions of pig sera were added to the plates prior to the addition of the MAb 210-F11 . The degree of binding of App-5 antibodies from pig sera was determined as the percentage inhibition of the MAb 210-F11 . Pig serum from specific pathogen free (SPF) herds, from experimentally infected animals, and from acutely and chronically infected herds were tested . A serum dilution of 1/30 was found to be optimal, when using 50% inhibition as the discriminating inhibition percentage . No cross-reactivity was observed with serum from pigs infected with other App serotypes or bacteria isolated from the respiratory tract, such as A . suis and H . parasuis . The inhibition EIA will be used for surveillance of App-5 antibodies in SPF and conventional herds.

EXS, 1997, 81, 109 - 20
New biosensors for environmental analysis; Hansen PD et al.; Environmental analysis requires fast and reliable measurement results . Biosensors, which facilitate integral monitoring as well as single substance analysis, achieve high sensitivities in a minimum of measuring time . Four new on-line biosensors, which cover a wide range of environmentally relevant substances, are introduced: A water-quality monitoring bacteria electrode, whose gradual development is described as an example, a heavy-metal screening urease inhibition sensor, a genotoxic potential as well as a immunotoxic potential indicating sensor . Future prospects are given.

J Rheumatol, 1997 Jan, 24(1), 227 - 8
Influence of the QKRAA/QRRAA/RRRAA motifs of the third hypervariable region of HLA-DRB1 in the development of rheumatoid arthritis; Auger I et al.; Most patients with rheumatoid arthritis (RA) express HLA-DR4, HLA-DR1, or HLA-DR10 . The DRB1 chains of these alleles have highly homologous 3rd hypervariable regions, with the motifs QKRAA, QRRAA, or RRRAA . The role played by the QKRAA, QRRAA, and RRRAA motifs in the development of RA is unknown . It may involve interaction with a T cell, an antigenic peptide, or an unknown ligand . We investigated the role played by the QKRAA motif and observed that it was expressed on numerous proteins from bacteria and viruses . However, we detected no anti-QKRAA autoimmunization in patients with RA . We found that QKRAA is a binding motif for bacterial and human 70 kDa heat shock proteins, and suggest that this may explain both its being expressed on so many proteins and its role in the development of RA.

J Leukoc Biol, 1997 Jan, 61(1), 58 - 62
Chemotaxis by human neutrophils and their cytokineplasts treated with inhibitors of nitric oxide synthase: no suppression of orientation or trajectory; Malawista SE et al.; Inhibitors of nitric oxide (NO) synthase are reported to inhibit both the adherence of polymorphonuclear leukocytes (PMN) to substrate and chemotaxis (directed locomotion) of PMN as determined in Boyden chamber assays . In the current study, we examined both human blood PMN and granule-poor motile cytoplasts derived from them (cytokineplasts, CKP), under direct microscopic observation with concomitant time-lapse video recording, for their ability to respond chemotactically to an erythrocyte destroyed by laser microirradiation . In this system we can observe directly and continuously the orientation and trajectory of PMN before, during, and after establishment of a chemotactic gradient . For both PMN and CKP we employed three different inhibitors of NO synthase (N(omega)-methyl-L-arginine, N-iminoethyl-L-ornithine, and diphenyleneiodonium) in at least twice the concentrations employed to inhibit chemotaxis of PMN in Boyden chambers or killing of bacteria in CKP . Although small differences in adhesion might not have been appreciated, treated PMN and CKP were each indistinguishable from untreated controls in their ability to orient in a newly created chemotactic gradient and in their trajectories toward the chemotactic target.

Arch Microbiol, 1997 Jan, 167(1), 46 - 53
Methylosulfonomonas methylovora gen . nov., sp . nov., and Marinosulfonomonas methylotropha gen . nov., sp . nov.: novel methylotrophs able to grow on methanesulfonic acid; Holmes AJ et al.; Two novel genera of restricted facultative methylotrophs are described; both Methylosulfonomonas and Marinosulfonomonas are unique in being able to grow on methanesulfonic acid as their sole source of carbon and energy . Five identical strains of Methylosulfonomonas were isolated from diverse soil samples in England and were shown to differ in their morphology, physiology, DNA base composition, molecular genetics, and 16S rDNA sequences from the two marine strains of Marinosulfonomonas, which were isolated from British coastal waters . The marine strains were almost indistinguishable from each other and are considered to be strains of one species . Type species of each genus have been identified and named Methylosulfonomonas methylovora (strain M2) and Marinosulfonomonas methylotropha (strain PSCH4) . Phylogenetic analysis using 16S rDNA sequencing places both genera in the alpha-Proteobacteria . Methylosulfonomonas is a discrete lineage within the alpha-2 subgroup and is not related closely to any other known bacterial genus . The Marinosulfonomonas strains form a monophyletic cluster in the alpha-3 subgroup of the Proteobacteria with Roseobacter spp . and some other partially characterized marine bacteria, but they are distinct from these at the genus level . This work shows that the isolation of bacteria with a unique biochemical character, the ability to grow on methanesulfonic acid as energy and carbon substrate, has resulted in the identification of two novel genera of methylotrophs that are unrelated to any other extant methylotroph genera.

Clin Immunol Immunopathol, 1997 Jan, 82(1), 60 - 7
HLA-DR polymorphism modulates the cytokine profile of Mycobacterium leprae HSP-reactive CD4+ T cells; Mitra DK et al.; In the present study, in vitro attempts have been made to define the cytokine profile of CD4+ T cells from polar leprosy patients and healthy individuals against Mycobacterium leprae-derived heat shock proteins (HSPs), HSP65 and HSP18, and their trypsin-digested fragments, relating to HLA-DR polymorphism . While all tryptic fragments of optimal digestion and undigested HSPs could stimulate CD4+ T cells from tuberculoid (TT) leprosy patients and healthy contacts (stimulation index, SI > 2.0), only two fragments, TDB65-2 (18 kDa) and TDB18-3 (3 kDa) triggered CD4+ T cells of anergic lepromatous (LL) leprosy patients . Both of these HSPs and their tryptic fragments showed diverse HLA-DR restriction, with DR15 providing the strongest restriction . Cytokine analysis demonstrated that HSP65 and HSP18 induced Th1-like activity in the context of all the restricting HLA-DR alleles, except DR1 and DR7 which induced a Th2 type of response against HSP65 and HSP18, respectively . These Th2 inducer epitopes on HSP65 (DR1 restricted) and HSP18 (DR7 restricted) were absent from TDB65-2 and TDB18-3 which exclusively triggered Th1 cells in both TT and LL forms of leprosy in the context of multiple DR alleles, DR15 being the major antigen-presenting allele . These studies suggest that the major histocompatibility complex phenotype of the antigen-presenting cell can modulate Th1-like versus Th2-like activity against M . leprae pathogens in leprosy and healthy individuals.

FEMS Microbiol Lett, 1997 Jan 1, 146(1), 31 - 8
The methanol oxidation genes mxaFJGIR (S) ACKLD in Methylobacterium extorquens; Amaratunga K et al.; MxaJ is a protein of unknown function encoded by mxaJ in the mxaFJGI operon . We have constructed a mxaJ mutant of M . extorquens with a deletion which does not affect transcription of downstream genes . It contained cytochrome cL (MxaG), but neither subunit of methanol dehydrogenase (MxaF and MxaI) . MxaJ is probably involved in processing this enzyme . We have sequenced the region between mxaFJGI and five other methanol oxidation genes, mxaACKLD; it includes one open reading frame (mxaR) and a possible second open reading frame (mxaS), demonstrating the presence in M . extorquens of the following gene cluster: mxaFJGIR(S) ACKLD.

Am J Gastroenterol, 1997 Jan, 92(1), 84 - 8
Percentage changes in serum pepsinogens are useful as indices of eradication of Helicobacter pylori; Furuta T et al.; OBJECTIVE: Eradication of Helicobacter pylori has been gaining significance for the treatment of gastroduodenal diseases . Establishment of a precise diagnostic method for H . pylori is of great value . The aim of this study was to establish a new method for precisely judging the eradication of this bacteria . METHODS: We measured serum pepsinogen I (PG I) and pepsinogen II (PG II) levels in 105 cases of peptic ulcer with H . pylori infection before and after anti-H . pylori treatment, determined percentage changes in serum PG I:PG II ratios before and 1 month after the treatment, and established cut-off values for them to distinguish success from failure of H . pylori eradication . Cut-off values for percentage changes in serum PG I:PG II ratios were tentatively set as +40%, +25%, and +10% when the serum PG I:PG II ratios before treatment were less than 3.0, not less than 3.0 but less than 5.0, and not less than 5.0, respectively . RESULTS: With these cut-off values, the sensitivity, specificity, and validity for determination of eradication of H . pylori-on the basis of culture, histology, the rapid urease test, and a polymerase chain reaction method-were 100.0%, 93.1%, and 96.2%, respectively . These cut-off values could be applied to both gastric ulcer and duodenal ulcer . CONCLUSION: Our findings suggest that percentage changes in serum PG I:PG II ratios are useful as indices for distinguishing success from failure in eradication therapy for H . pylori.

Int J Syst Bacteriol, 1997 Jan, 47(1), 224 - 7
Comparison of the 16S/23S ribosomal intergenic regions of "Candidatus Liberobacter asiaticum" and "Candidatus Liberobacter africanum," the two species associated with citrus huanglongbing (greening) disease; Jagoueix S et al.; 16S/23S intergenic spacer regions from the rRNA operons of two strains of "Candidatus Liberobacter asiaticum" and one strain of "Candidatus Liberobacter africanum" were cloned and sequenced . The intergenic spacers of the two "Candidatus L . asiaticum" strains studied are identical and contain the genes for isoleucine tRNA (tRNA(Ile)) and alanine tRNA (tRNA(Ala)) separated by 11 nucleotides . The intergenic spacer of the "Candidatus L . africanum" strain contains only one tRNA gene (tRNA(Ala)) . The level of homology between the intergenic spacers of the two liberobacter species is 79.46% . Ribosomal operons with 16S/23S spacer regions other than those studied might be present in the two "Candidatus Liberobacter" species.

Int J Syst Bacteriol, 1997 Jan, 47(1), 46 - 53
Phylogenetic analysis of the genus Actinomyces based on 16S rRNA gene sequences: description of Arcanobacterium phocae sp . nov., Arcanobacterium bernardiae comb . nov., and Arcanobacterium pyogenes comb . nov; Ramos CP et al.; A systematic phylogenetic analysis of the genus Actinomyces was performed . The 16S rRNA gene sequences of 13 Actinomyces species, an unnamed Actinomyces strain (ATCC 49338), and an Actinomyces-like isolate from sea mammals were determined . Comparative sequence analysis with closely related taxa revealed phylogenetic diversity and internal structure within the genus Actinomyces . In addition, some members of other genera (viz., the genera Arcanobacterium, Mobiluncus, and Rothia) were shown to be phylogenetically intermixed with the Actinomyces species . It was evident from both distance and tree topology considerations that the genus Actinomyces is in urgent need of taxonomic revision and requires subdivision into several genera . Based on the results of the present study it is proposed that Actinomyces bernardiae and Actinomyces pyogenes be assigned to the genus Arcanobacterium as Arcanobacterium bernardiae comb . nov . and Arcanobacterium pyogenes comb . nov., respectively . In addition, a new species, Arcanobacterium phocae, is proposed for Actinomyces-like bacteria isolated from seals.

Methods, 1997 Jan, 11(1), 116 - 27
Interleukin-12: an immunoregulatory cytokine produced by B cells and antigen-presenting cells; Sartori A et al.; Interleukin 12 (IL-12) is a heterodimeric protein produced by B cells, phagocytic cells, and other antigen-presenting cells . IL-12 was originally purified from the supernatant fluids of human EBV-transformed cell lines and later observed to be produced by the large majority of such cell lines, especially and at high levels from those derived from AIDS-associated lymphomas . However, phagocytic cells rather than B cells appear to be the most important physiological producers of IL-12 . There are two pathways of IL-12 induction in phagocytic cells: a T-cell-independent one, induced primarily by bacteria, bacterial products, or intracellular parasites and important in the early inflammatory response of innate resistance; and a T-cell-dependent one, induced by the interaction of CD40L on activated T cells with CD40 receptor on IL-12-producing cells (phagocytic cells and antigen-presenting cells) and important in the regulation of adaptive immunity . IL-12 induces production of cytokines, especially interferon-gamma, from both T and NK cells, enhances the cytotoxic activity of NK cells and the generation of cytotoxic T cells, and has a proliferative activity on T and NK cells . Both in vivo and in vitro, IL-12 is a powerful inducer of T helper type 1 (Th1) response, whereas it inhibits Th2-type responses.

J Bacteriol, 1997 Jan, 179(1), 267 - 71
Anabaena sp . strain PCC 7120 responds to nitrogen deprivation with a cascade-like sequence of transcriptional activations; Cai Y et al.; Anabaena sp . strain PCC 7120 adapts to deprivation of fixed nitrogen by undergoing physiological and genetic changes that include formation of N2-fixing heterocysts . Whether or not certain of the genes involved are interdependently expressed has been studied.

J Bacteriol, 1997 Jan, 179(1), 148 - 56
Fine-structure evidence for cell membrane partitioning of the nucleoid and cytoplasm during bud formation in Hyphomonas species; Zerfas PM et al.; Hyphomonas spp . reproduce by budding from the tip of the prosthecum, distal to the main body of the reproductive cell; thus, the chromosome must travel through the prosthecum to enter the progeny, the swarm cell . When viewed by electron microscopy, negatively stained whole cells, ultrathin-sectioned cells, and freeze-etched and frozen hydrated cells all had marked swellings of the cytoplasmic membrane (CM) in the prosthecum which are termed pseudovesicles (PV) . PV were separated by constrictions in the contiguous CM . In replicating cells, PV housed ribosomes and DNA, which was identified by its fibrillar appearance and by lactoferrin-gold labeling . The micrographs also revealed that the CM bifurcates at the origin of the prosthecum so that one branch partitions the main body of the reproductive cell from the prosthecum and swarm cell . The results of this fine-structure analysis suggest models explaining DNA segregation and the marked asymmetric polarity of the budding reproductive cell.

Appl Environ Microbiol, 1997 Jan, 63(1), 335 - 7
Development of a species-specific gene probe for Hyphomicrobium facilis with the inverse PCR; Fesefeldt A et al.; A species-specific gene probe for Hyphomicrobium facilis was generated from a transposon Tn5-132 insertion mutant defective in methanol oxidation by the inverse PCR . With this probe, the abundance of H . facilis in a garden soil was determined as a subfraction of the total cultivable hyphomicrobia.

Pediatr Res, 1997 Jan, 41(1), 114 - 9
Modified natural porcine surfactant inhibits superoxide anions and proinflammatory mediators released by resting and stimulated human monocytes; Walti H et al.; Pulmonary surfactant has a potential role in modulating inflammation in normal and injured lungs . In lung injury, monocytes become activated and participate in lung inflammation . We therefore, investigated the proinflammatory functions of stimulated human blood monocytes after an overnight preincubation period with modified natural porcine surfactant (Curosurf) (500-1000 micrograms/mL) . Monocytes were stimulated either with phorbol myristate acetate (PMA), bacterial extract OM-85, lipopolysaccharide (LPS), or Ca2+ ionophore A23187 . The present study shows that Curosurf significantly inhibits: 1) the production of superoxide anions stimulated with OM-85 (1 mg/mL, 30 min), but not with PMA (100 ng/mL, 30 min); 2) the release of cyclooxygenase metabolites prostaglandin E2 and thromboxane B2 stimulated with OM-85 (1 mg/mL, overnight); 3) the release of lipoxygenase metabolite leukotriene C4 stimulated with A23187 (10 microM, 10 min); 4) the release of the cytokine TNF-alpha stimulated overnight with either OM-85 (1 mg/mL) or LPS (10 micrograms/mL)) in a dose-dependent fashion . In addition, Curosurf decreases the spontaneous adherence of monocytes to plastic culture wells in a dose-dependent fashion . Experiments performed with staurosporine, an inhibitor of protein kinase C (PKC) indicate that, in contrast with PMA, the production of superoxide anions stimulated by OM-85 is not related to PKC activation . Consequently, we propose that the mechanism involved in the suppressive effects of Curosurf is PKC-independent . In summary, the present study provides experimental evidence that favors the anti-inflammatory role of modified natural porcine surfactant (Curosurf) in human monocytes in vitro.

Infect Immun, 1997 Jan, 65(1), 227 - 35
Development of antibody-secreting cells and antigen-specific T cells in cervical lymph nodes after intranasal immunization; Wu HY et al.; Intranasal (i.n.) immunization with bacterial protein antigens coupled to cholera toxin B subunit (CTB) effectively induces mucosal, especially salivary immunoglobulin A (IgA), and nonmucosal antibody responses in mice . To examine the regional distribution of antigen-specific B and T cells after i.n . immunization, antibody-secreting cells and antigen-responsive T cells in cervical lymph nodes (CLN) were compared with those found after intraoral or subcutaneous (in the neck) administration of the same antigen and with T cells found in mesenteric lymph nodes (MLN) and spleen after intragastric immunization . The i.n . immunization induced predominantly IgA antibody-secreting cells in salivary glands and IgA and IgG antibody-secreting cells in the superficial and central CLN; these responses were quantitatively enhanced if the antigen was coupled to CTB . Intraoral immunization also induced IgA and IgG antibody-secreting cells in the superficial and central CLN, but only if intact cholera toxin was included as an adjuvant . In contrast, subcutaneous (neck) immunization induced IgG antibody-secreting cells mainly in the draining facial lymph nodes . CLN cell populations resembled those of MLN, except that CLN lymphocytes had higher proportions of T cells and lower proportions of B cells and a slightly higher CD4+/CD8+ ratio among T cells than the MLN lymphocytes did . T cells that proliferated in response to antigen in vitro were found especially in central CLN 2 days after i.n . immunization and persisted for up to 6 months, whereas after intragastric immunization, responsive T cells were not found in the MLN for up to 14 days . After culture with antigen in vitro, T cells from the superficial CLN of i.n . immunized mice secreted both gamma interferon and interleukin-4 . Therefore, after i.n . immunization, superficial and central CLN represent sites of regional lymphocyte development, and the central CLN in particular appear to be sites where memory T cells persist.

Infect Immun, 1997 Jan, 65(1), 122 - 6
Marked reduction of mouse peritoneal CD5+ B cells by intraperitoneal administration of lipopolysaccharide; Paeng N et al.; Intraperitoneal administration of lipopolysaccharide to mice induced a marked reduction of CD5+ B cells in the peritoneal cavity . The reduction was not induced by intravenous, subcutaneous, or oral administration of lipopolysaccharide . The reduction continued for about 10 days after the injection, and the CD5+ B-cell count recovered to the normal state about 14 days after the injection . The reduction of peritoneal CD5+ B cells might be caused by apoptotic cell death . Injection of lipopolysaccharide did not result in production of antibody to lipopolysaccharide . On the other hand, intraperitoneal injection of heat-killed bacteria did not induce a reduction of peritoneal CD5+ B cells and elicited the definite production of antibody to lipopolysaccharide.

Infect Immun, 1997 Jan, 65(1), 35 - 41
Butyric acid-induced apoptosis of murine thymocytes, splenic T cells, and human Jurkat T cells; Kurita-Ochiai T et al.; The purpose of this study was to examine the effect of butyric acid, an extracellular metabolite from periodontopathic bacteria, on apoptosis induction in murine thymocytes, splenic T cells, and human Jurkat T cells . Butyric acid significantly suppressed T-cell viability in both a concentration- and time-dependent fashion . The results of DNA fragmentation assay indicated that butyric acid rapidly induced apoptosis in thymocytes (with 1.25 mM butyric acid and 6 h after treatment) and in splenic T cells and Jurkat cells (with 2.5 mM butyric acid and 16 h after treatment) . Incubation of thymocytes or Jurkat cells with 5 mM butyric acid for 21 h resulted in the typical ladder pattern of DNA fragmentation . Furthermore, Jurkat cells treated with 5 mM butyric acid showed the characteristic pattern of apoptotic cells such as chromatin condensation and hypodiploid nuclei . Experiments with fractionated subpopulations of splenic T cells revealed that DNA fragmentation was predominantly observed in CD4+ T cells . Butyric acid-induced apoptosis of thymocytes was decreased by the protein kinase inhibitors H7 and staurosporine . These inhibitors were less effective with similarly treated splenic T cells and Jurkat cells . These data suggest that butyric acid, one of the volatile fatty acids produced by periodontopathic bacteria and one that easily penetrates the oral mucosa, can modulate the immunoregulatory cell population in periodontal tissue by inducing T-cell death through apoptosis.

Mol Cell Biol, 1997 Jan, 17(1), 482 - 94
Molecular cloning and characterization of a transcription factor for the copia retrotransposon with homology to the BTB-containing lola neurogenic factor; Cavarec L et al.; By transfection experiments, we previously identified a 72-bp enhancer sequence within the Drosophila copia retrotransposon which is involved in the control of the transcription level of this mobile element in cells in culture . Gel shift assays with nuclear extracts from Drosophila hydei-derived DH-33 cells further demonstrated specific interactions of at least two nuclear factors with this enhancer sequence . Using this sequence as a probe for the screening of an expression cDNA library that we constructed from DH-33 cells RNA, we have isolated a cDNA clone encoding a 110-kDa protein with features common to those of known transcription factors; these include a two-zinc-finger motif at the C terminus, three glutamine-rich domains in the presumptive activation domain of the protein, and an N-terminal domain which shares homology with the Bric-a-brac, Tramtrack, and Broad-Complex BTB boxes . The precise DNA recognition sequence for this transcription factor has been determined by both gel shift assays and footprinting experiments with a recombinant protein made in bacteria . The functionality of the cloned element was demonstrated upon transcriptional activation of copia reporter genes, as well as of a minimal promoter coupled with the identified target DNA sequence, in cotransfection assays in cells in culture with an expression vector for the cloned factor . Southern blot and nucleotide sequence analyses revealed a related gene in Drosophila melanogaster (the lola gene) previously identified by a genetic approach as involved in axon growth and guidance . Transfection assays in cells in culture with lola gene expression vectors and in situ hybridization experiments with lola gene mutants finally provided evidence that the copia retrotransposon is regulated by this neurogenic gene in D.melanogaster, with a repressor effect in the central nervous systems of the embryos.

Mol Cell Biol, 1997 Jan, 17(1), 61 - 8
Suppression of grp78 core promoter element-mediated stress induction by the dbpA and dbpB (YB-1) cold shock domain proteins; Li WW et al.; The highly conserved grp78 core promoter element plays an important role in the induction of grp78 under diverse stress signals . Previous studies have established a functional region in the 3' half of the core (stress-inducible change region {SICR}) which exhibits stress-inducible changes in stressed nuclei . The human transcription factor YY1 is shown to bind the SICR and transactivate the core element under stress conditions . Here we report that expression library screening with the core element has identified two new core binding proteins, YB-1 and dbpA . Both proteins belong to the Y-box family of proteins characterized by an evolutionarily conserved DNA binding motif, the cold shock domain (CSD) . In contrast to YY1, which binds only double-stranded SICR, the Y-box/CSD proteins much prefer the lower strand of the SICR . The Y-box proteins can repress the inducibility of the grp78 core element mediated by treatment of cells with A23187, thapsigargin, and tunicamycin . In gel shift assays, YY1 binding to the core element is inhibited by either YB-1 or dbpA . A yeast interaction trap screen using LexA-YY1 as a bait and a HeLa cell cDNA-acid patch fusion library identified YB-1 as a YY1-interacting protein . In cotransfection experiments, the Y-box proteins antagonize the YY1-mediated enhancement of transcription directed by the grp78 core in stressed cells . Thus, the CSD proteins may be part of the stress signal transduction mechanism in the mammalian system.

FEMS Immunol Med Microbiol, 1996 Dec 31, 16(3-4), 247 - 55
Cryptic domains of a 60 kDa heat shock protein of Helicobacter pylori bound to bovine lactoferrin; Amini HR et al.; Bovine lactoferrin binds to a 60 kDa heat shock protein of Helicobacter pylori . Binding ability was related to human immunoglobulin G because bovine lactoferrin binding proteins were isolated by extraction of cell surface associated proteins with distilled water, applied on IgG-Sepharose and nickel sulphate chelate affinity chromatography . Binding was demonstrated by Western blot after purified protein was digested with alpha-chymotrypsin and incubated with peroxidase-labeled bovine lactoferrin . Binding was inhibited by bovine lactoferrin, lactose, rhamnose, galactose, and two iron-containing proteins, ferritin and haptoglobin . Helicobacter pylori binds ferritin and haptoglobin via charge or hydrophobic interactions because this binding was not inhibited by specific and various glycoproteins or carbohydrates . Carbohydrate moieties of bovine lactoferrin molecules seem to be involved in binding because glycoproteins with similar carbohydrate structures strongly inhibited binding . Scatchard plot analysis of the binding of peroxidase-labeled bovine lactoferrin to H . pylori cells yielded a kd 2.88 x 10(-6) M . In addition, binding of H . pylori cells to bovine lactoferrin was enhanced when bacteria treated with pepsin or alpha-chymotrypsin after isolation from iron-restricted and iron-containing media.

Brain Res Mol Brain Res, 1996 Dec 31, 43(1-2), 267 - 78
Serotonin transporter antibodies: production, characterization, and localization in the brain; Zhou FC et al.; Serotonin (5-HT) transporter, the mechanism for 5-HT high affinity uptake, is the essential component for the termination of 5-HT transmission . In order to identify transporter sites on 5-HT neurons or on other 5-HT uptaking cells, three rabbit antisera against cocaine sensitive-serotonin transporter (5-HTT) were produced . Antisera 5-HTT55 (against amino acid sequence 55-68 in cytoplasmic N-terminal) and 5-HTT315 (against amino acid sequence 315-325, a 3rd external loop peptide) were produced against synthetic multiple-antigenic peptides (MAP) . Antiserum 5-HTTN was produced against a fusion protein of the first 71 amino acids of N-terminal peptide expressed in recombinant DNA transformed bacteria . SDS-PAGE/Western blots indicate that 5-HTT55 and 5-HTT315 recognized bands of 74 and 64 kDa in rat brains with densities in the order of cortex > or = hippocampus > cerebellum, but not in liver, or muscle . The 5-HTTN recognized the fusion protein expressed in the bacteria, and the 64 kDa band with a similar density profile in the rat brain regions, and negative in liver and muscle . The immunocytochemistry of all three antisera revealed 5-HTT-immunostaining (5-HTT-im) in a pattern similar to 5-HT fiber distribution . 5-HTT55 and 5-HTT315 stainings were punctate in appearance, while 5-HTTN outlined the fibers in the 5-HT fiber areas, and neurons in raphe but not in substantia nigra or locus ceruleus . The preimmune serum and immune serum preabsorbed with 5-HTTN showed negative or diminished staining . Specific neurotoxin, 5,7-dihydroxytryptamine lesion removed all of the 5-HTTN fibers from the injection site, indicating, that 5-HTTN-im fibers are 5-HT fibers in nature . Our study indicates that the three antibodies we produced recognize various domains of the 5-HTT . Our 5-HTT antibodies could be used as new markers of 5-HT fibers, and are particularly useful for the study of the plasticity of 5-HT fibers free of the complications involved with 5-HT content.

Biochemistry, 1996 Dec 24, 35(51), 16852 - 62
Kinetic mechanism of folding and unfolding of Rhodobacter capsulatus cytochrome c2; Sauder JM et al.; In spite of marginal sequence homology, cytochrome c2 from photosynthetic bacteria and the mitochondrial cytochromes c exhibit some striking structural similarities, including the tertiary arrangement of the three main helices . To compare the folding mechanisms for these two distantly related groups of proteins, equilibrium and kinetic measurements of the folding/unfolding reaction of cytochrome c2 from Rhodobacter capsulatus were performed as a function of guanidine hydrochloride (GuHCl) concentration in the absence and presence of a stabilizing salt, sodium sulfate . Quenching of the fluorescence of Trp67 by the heme was used as a conformational probe . Kinetic complexities due to non-native histidine ligation are avoided, since cytochrome c2 contains only one histidine, His17, which forms the axial heme ligand under native and denaturing conditions . Quantitative kinetic modeling showed that both equilibrium and kinetic results are consistent with a minimal four-state mechanism with two sequential intermediates . The observation of a large decrease in fluorescence during the 2-ms dead-time of the stopped-flow measurement (burst phase) at low GuHCl concentration, followed by a sigmoidal recovery of the initial amplitude toward the unfolding transition region, is attributed to a well-populated compact folding intermediate in rapid exchange with unfolded molecules . A nearly denaturant-independent process at low GuHCl concentrations reflects the rate-limiting conversion of a compact intermediate to the native state . At high GuHCl concentrations, a process with little denaturant dependence is attributed to the rate-limiting Met96-iron deligation process during unfolding, which is supported by the kinetics of imidazole binding . The strong GuHCl-dependence of folding and unfolding rates near the midpoint of the equilibrium transition is attributed to destabilization of each intermediate and their transition states in folding and unfolding . Addition of sodium sulfate shifts the rate profile to higher denaturant concentration, which can be understood in terms of the relative stabilizing effect of the salt on partially and fully folded states.

Eur J Biochem, 1996 Dec 15, 242(3), 695 - 702
Membrane-bound c-type cytochromes in Heliobacillus mobilis . Characterisation by EPR and optical spectroscopy in membranes and detergent-solubilised material; Nitschke W et al.; The spectral and electrochemical parameters, as well as the orientations of the heme plane with respect to the membrane plane, of the c-type hemes present in membrane fragments from Heliobacillus mobilis were characterised by optical and EPR spectroscopy . Cytochrome C53, was thereby shown to represent at least four and possibly five heme species with the following characteristics: Em = -60 mV +/- 10 mV, g, = 2.92, 60 degrees; Em = +90 mV +/- 10 mV, g, = 2.92, 90 degrees; Em = +120 mV +/- 20 mV, g, = 3.03; and Em = +170 mV +/- 20 mV, g, = 3.03 . The latter component may correspond to two hemes with redox midpoint potentials of Em = +160 mV +/- 20 mV and Em = +180 mV +/- 20 mV (all Em values at pH 7.0) . For the heme species having g, peaks at g approximately 3.03, determination of individual orientations was precluded due to the superposition of several differently oriented hemes . About one copy of each heme was found to be present per photosynthetic reaction centre, with the exception of the +120 mV component for which a stoichiometry of 2 hemes/reaction centre was obtained . The heme proteins were detergent-solubilised and partially purified . Three c-type cytochromes that migrated with apparent molecular masses of 18, 29 and 50 kDa were detected on SDS/PAGE . Optical redox titrations at pH 7.0 showed redox midpoint potentials of +160 mV +/- 10 mV for the 18-kDa cytochrome, and -60 mV +/- 10 mV, with possible contributions around +160 mV, for the 50-kDa cytochrome . A tentative attribution of heme species observed in membranes to the isolated heme proteins is presented . The results obtained on H . mobilis are compared with those reported for green sulphur bacteria.

J Clin Invest, 1996 Dec 15, 98(12), 2764 - 70
Differences in endogenous peptides presented by HLA-B*2705 and B*2703 allelic variants . Implications for susceptibility to spondylarthropathies; Boisgerault F et al.; The association between HLA-B27 and spondylarthropathies is currently being reinvestigated in the light of HLA-B27 subtyping . At least 11 different subtypes have been described among which B*2703, B*2706, and B*2709 could be less closely associated with disease at the population level . Differences in the presentation of antigenic peptides by these subtypes could be related to differences in disease susceptibility . We focused our work on the comparison of B*2705 and B*2703 which differ at a single position at residue 59 in pocket A of the peptide binding groove . Endogenous peptides from the human C1R line transfected by B*2705 or B*2703 were acid-eluted and separated by HPLC . Major individual fractions were sequenced by Edman NH2-terminal degradation . Differences observed between B*2705 versus B*2703 individual ligands were confirmed in an in vitro stabilization assay with T2-B*2705 or B*2703 transfected cells in the presence of synthetic peptides . One B*2705 associated peptide is derived from the sequence 169-179 in the second extracellular domain of several HLA class I molecules including HLA-B27 . This sequence (RRYLENGKETL) is highly homologous to a previously reported sequence (LRRYLENGK) sharing similarities with proteins from enteric bacteria . We show here that it is naturally presented as a major endogenous peptide by B*2705 and B*2702 disease-associated subtypes and not by B*2703.

FEMS Microbiol Lett, 1996 Dec 15, 145(3), 433 - 7
Anaerobic degradation of benzene in BTX mixtures dependent on sulfate reduction; Phelps CD et al.; Enrichment cultures from marine sediments mineralized benzene while using sulfate as the terminal electron acceptor . Parallel cultures using river marsh sediment displayed no activity . Mineralization was confirmed by release of 14CO2 from radiolabeled benzene . The dependence on sulfate reduction was demonstrated by stoichiometric balances and the use of specific inhibitors . This work supports recent observations that anaerobic benzene degradation takes place coupled to sulfate reduction.

J Immunol, 1996 Dec 15, 157(12), 5503 - 11
A nine-amino acid peptide from IL-1beta augments antitumor immune responses induced by protein and DNA vaccines; Hakim I et al.; The idiotypic determinants of B cell lymphoma provide a tumor-specific Ag and a target for immunotherapy . We have developed several generations of idiotype vaccines that were tested in an animal model, the 38C13 mouse B cell lymphoma . Initially we showed that effective tumor immunity was elicited by the syngeneic Id when it was conjugated to a carrier protein and mixed with an adjuvant . A subsequent generation of Id vaccines eliminated the need for a carrier protein and for an adjuvant by incorporating cytokines into fusion proteins containing the Id . A third generation of vaccines consisting of naked DNA encoding the Id-granulocyte-macrophage colony-stimulating factor (GM-CSF) fusion proteins was equally effective in inducing tumor immunity . To determine whether Ig variable regions, in the absence of constant regions, could be immunotherapeutic in this model, we tested the use of single-chain Fv (scFv) . scFv proteins, produced in bacteria, and naked DNA encoding scFv were used in this study . scFv was tested alone or fused to GM-CSF or an immunoenhancing peptide derived from IL-1beta . Here we demonstrate that scFv-GM-CSF was effective only when injected as a protein, not as a DNA vaccine . In contrast, both scFv-IL-1beta peptide fusion protein and naked DNA encoding it induced tumor immunity that protected mice from tumor challenge.

Presse Med, 1996 Dec 14, 25(39), 1909 - 15
{Tuberculosis . Current therapeutics}; Tattevin P et al.; Due to the constant decline in incidence up to the mid eighties, eradication of tuberculosis appeared to be an attainable objective in developed countries . Since then multiple factors (HIV epidemic, poor social conditions in certain unfavored areas, population migrations, urbanization) have led to an increased frequency, making an excellent knowledge of tuberculosis a priority for all physicians . Multi-resistant mycobacteria have also made their appearance leading to numerous clinical and experimental studies which provide new insights into the correct management of patients with tuberculosis . Despite these recent changes, the classical treatment for tuberculosis remains the same in most cases, allowing nearly-certain cure when applied correctly in patients infected with a susceptible bacteria, including those with HIV infection or extrapulmonary localizations . On the contrary, the spontaneous aggravation of multi-resistant tuberculosis, even in some cases being treated, emphasizes the need to test the strain's susceptibility to the antituberculous agents used . Certain new antibiotics, including fluoroquinolones, may play an important role in some cases . The contribution of surgery, isolation and strict compliance must also be emphasized . Resistant strains may also led to renewed indications for the Calmette-Guerin vaccine.

J Biol Chem, 1996 Dec 13, 271(50), 31787 - 90
Activation of myosin-I by members of the Ste20p protein kinase family; Wu C et al.; The heavy chain of myosin-ID isolated from Dictyostelium was identified as an in vitro substrate for members of the Ste20p family of serine/threonine protein kinases which are thought to regulate conserved mitogen-activated protein kinase pathways . Yeast Ste20p and Cla4p and mammalian p21-activated protein kinase (PAK) phosphorylated the heavy chain to 0.5-0.6 mol of Pi/mol and stimulated the actin-dependent Mg2+-ATPase activity to an extent equivalent to that of the Ste20p-like myosin-I heavy chain kinase isolated from Dictyostelium . PAK purified from rat brain required GTPgammaS-Cdc42 to express full activity, whereas recombinant mouse mPAK3 fused to glutathione S-transferase and purified from bacteria, and Ste20p and Cla4p purified from yeast extracts were fully active without GTPgammaS-Cdc42 . These results suggest, together with the high degree of structural and functional conservation of Ste20p family members and myosin-I isoforms, that myosin-I activation by Ste20p family protein kinases may contribute to the regulation of morphogenetic processes in organisms ranging from yeast to mammalian cells.

J Biotechnol, 1996 Dec 10, 52(2), 107 - 20
DNA recovery and PCR quantification of catechol 2,3-dioxygenase genes from different soil types; Wikstrom P et al.; With the objective of monitoring xenobiotic degrading bacteria in soil, a method for rapid extraction of DNA from soil, amenable to amplification by PCR, was developed . The method was based on lysis by freeze-thawing and subsequent addition of sodium dodecyl sulfate (SDS), hexadecyltrimethylammonium bromide and proteinase K . The extraction method required 2 h and was tested on six different soils differing in organic content, water holding capacity and pH, including ones from which DNA extraction is difficult . DNA yields from the soils ranged from 6.1 to 54.0 micrograms of DNA per g soil . The efficiency and reproducibility of the DNA extraction method were evaluated by competitive PCR . The organic content in the soils was a major factor affecting the amount of obtained DNA amenable for amplification by PCR . A PCR primer-pair was designed on the basis of the known nucleotide sequences of several catechol 2,3-dioxygenase genes . The specificity of the primer-pair was demonstrated on different sequenced catechol 2,3-dioxygenase genes and on site-specific bacterial isolates from polycyclic aromatic hydrocarbon (PAH)-contaminated soil . The concentration of catechol 2,3-dioxygenase DNA in PAH-contaminated sediment undergoing an ex situ compost process was quantified by competitive PCR over a period of 16 weeks . The concentration of PAHs and catechol 2,3-dioxygenase DNA in the soil samples, was found to correlate.

Proc Natl Acad Sci U S A, 1996 Dec 10, 93(25), 14614 - 7
Presence of a mitochondrial-type 70-kDa heat shock protein in Trichomonas vaginalis suggests a very early mitochondrial endosymbiosis in eukaryotes; Germot A et al.; Molecular phylogenetic analyses, based mainly on ribosomal RNA, show that three amitochondriate protist lineages, diplomonads, microsporidia, and trichomonads, emerge consistently at the base of the eukaryotic tree before groups having mitochondria . This suggests that these groups could have diverged before the mitochondrial endosymbiosis . Nevertheless, since all these organisms live in anaerobic environments, the absence of mitochondria might be due to secondary loss, as demonstrated for the later emerging eukaryote Entamoeba histolytica . We have now isolated from Trichomonas vaginalis a gene encoding a chaperone protein (HSP70) that in other lineages is addressed to the mitochondrial compartment . The phylogenetic reconstruction unambiguously located this HSP70 within a large set of mitochondrial sequences, itself a sister-group of alpha-purple bacteria . In addition, the T . vaginalis protein exhibits the GDAWV sequence signature, so far exclusively found in mitochondrial HSP70 and in proteobacterial dnaK . Thus mitochondrial endosymbiosis could have occurred earlier than previously assumed . The trichomonad double membrane-bounded organelles, the hydrogenosomes, could have evolved from mitochondria.

Proc Natl Acad Sci U S A, 1996 Dec 10, 93(25), 14515 - 20
A global two component signal transduction system that integrates the control of photosynthesis, carbon dioxide assimilation, and nitrogen fixation; Joshi HM et al.; Photosynthesis, biological nitrogen fixation, and carbon dioxide assimilation are three fundamental biological processes catalyzed by photosynthetic bacteria . In the present study, it is shown that mutant strains of the nonsulfur purple photosynthetic bacteria Rhodospirillum rubrum and Rhodobacter sphaeroides, containing a blockage in the primary CO2 assimilatory pathway, derepress the synthesis of components of the nitrogen fixation enzyme complex and abrogate normal control mechanisms . The absence of the Calvin-Benson-Bassham (CBB) reductive pentose phosphate CO2 fixation pathway removes an important route for the dissipation of excess reducing power . Thus the mutant strains develop alternative means to remove these reducing equivalents, resulting in the synthesis of large amounts of nitrogenase even in the presence of ammonia . This response is under the control of a global two-component signal transduction system previously found to regulate photosystem biosynthesis and the transcription of genes required for CO2 fixation through the CBB pathway and alternative routes . In addition, this two-component system directly controls the ability of these bacteria to grow under nitrogen-fixing conditions . These results indicate that there is a molecular link between the CBB and nitrogen fixation process, allowing the cell to overcome powerful control mechanisms to remove excess reducing power generated by photosynthesis and carbon metabolism . Furthermore, these results suggest that the two-component system integrates the expression of genes required for the three processes of photosynthesis, nitrogen fixation, and carbon dioxide fixation.

J Long Term Eff Med Implants, 1997, 7(3-4), 225 - 34
Endotoxin and particulate matter on surgical gloves; Holmdahl L et al.; Because of the continual need to protect of health care workers, gloves are increasingly used . Most gloves purchased are powdered . Besides having deleterious effects on wound healing, glove powder can act as a vehicle for latex allergens, endotoxin, and possibly bacteria . Few tested gloves had undetectable levels of endotoxin, and only one was free from particulate matter . Since powder particles can become airborne and are thus easily spread, concerns for health care professionals and patients arise . To reduce risks for exposure, glove-selection criteria must include low or undetectable levels of latex allergens, endotoxin, and particulate matter.

J Biol Chem, 1996 Dec 6, 271(49), 31502 - 7
Purification and identification of a 28-kDa calcium-regulated heat-stable protein . A novel secretagogue-regulated phosphoprotein in exocrine pancreas; Groblewski GE et al.; This study reports the purification and identification of a novel 28 kDa phosphoprotein from rat pancreatic acini, previously described as being highly regulated by calcium mobilizing secretagogues, which we have designated calcium-regulated heat-stable protein 28 (CRHSP-28) . Internal amino acid sequences of purified CRHSP-28 were obtained following trypsin digestion and found to match with >95% identity the predicted amino acid sequence of a novel cDNA recently identified as being highly expressed in human breast carcinomas . Verification that this cDNA codes for human CRHSP-28 was demonstrated by the ability of antiserum raised against purified rat CRHSP-28 to recognize the recombinant human protein when expressed in bacteria . Furthermore, this antibody was found to specifically react with CRHSP-28 in rat acini following one- and two-dimensional electrophoresis and underwent a marked acidic shift in mobility after cholecystokinin stimulation, a phenomenon indicative of an increase in its phosphorylation . CRHSP-28 is predicted to be extremely hydrophilic, is phosphorylated entirely on serine residues, and bears little homology to any known proteins . Finally, the distribution of the CRHSP-28 protein in various rat tissues revealed that although it was present at low levels in almost all tissues, it was most highly expressed in pancreas, followed by the gastric, intestinal, and colonic mucosa . In view of its relative abundance throughout the digestive system and its apparent regulation by calcium-mobilizing agents, this protein may provide valuable insight into the mechanism(s) of calcium signaling in these tissues.

J Biol Chem, 1996 Dec 6, 271(49), 31135 - 44
Modified ligands to FA and FB in photosystem I . Proposed chemical rescue of a {4Fe-4S} cluster with an external thiolate in alanine, glycine, and serine mutants of PsaC; Jung YS et al.; The FB and FA electron acceptors in Photosystem I (PS I) are {4Fe-4S} clusters ligated by cysteines provided by PsaC . In a previous study (Mehari, T., Qiao, F., Scott, M . P., Nellis, D., Zhao, J., Bryant, D., and Golbeck, J . H . (1995) J . Biol . Chem . 270, 28108-28117), we showed that when cysteines 14 and 51 were replaced with serine or alanine, the free proteins contained a S = 1/2, {4Fe-4S} cluster at the unmodified site and a mixed population of S = 1/2, {3Fe-4S} and S = 3/2, {4Fe-4S} clusters at the modified site . We show here that these mutant PsaC proteins can be rebound to P700-FX cores, resulting in fully functional PS I complexes . The low temperature EPR spectra of the C14XPsaC.PS I complexes (where X = S, A, or G) show the photoreduction of a wild-type FA cluster and a modified FB' cluster, the latter with g values of 2.115, 1.899, and 1.852 and linewidths of 110, 70, and 85 MHz . Since neither alanine nor glycine contains a suitable side group, an external thiolate provided by beta-mercaptoethanol has likely been recruited to supply the requisite ligand to the {4Fe-4S} cluster . The EPR spectrum of the C51SPsaC.PS I complex differs from that of the C51APsaC.PS I or C51GPsaC.PS I complexes by the presence of an additional set of resonances, which may be derived from the serine oxygen-ligated cluster . In all other mutant PS I complexes, a wild-type spin-coupled interaction spectrum appears when FA and FB are simultaneously reduced . Single turnover flash studies indicate approximately 50% efficient electron transfer to FA/FB in the C14SPsaC.PS I, C51SPsaC.PS I, C14GPsaC.PS I, and C51GPsaC.PS I mutants and less than 40% in the C14APsaC.PS I and C51APsaC.PS I mutants, compared with approximately 76% in the PS I core reconstructed with wild-type PsaC . These data are consistent with the measurements of the rates of cytochrome c6-NADP+ reductase activity, indicating lower rates in the alanine mutants . It is proposed that the chemical rescue of a {4Fe-4S} cluster with a recruited external thiolate at the modified site allows the mutant PsaC proteins to rebind to PS I and to function in forward electron transfer.

FEBS Lett, 1996 Dec 2, 398(2-3), 187 - 92
Primary structure of a putative serine protease specific for IGF-binding proteins; Zumbrunn J et al.; From a subtracted cDNA library we have isolated a cDNA clone coding for a novel transformation-sensitive protein which is expressed by human fibroblasts, but not by their matched SV40 transformed counterparts . This protein has a molecular mass of 51 kDa and is highly related to the HtrA family of serine proteases from bacteria . At the N-terminal end, it contains an IGF-binding domain which may modulate the activity of the associated serine protease . Our data are consistent with the assumption that the novel protein represents one of the proteases that regulate the availability of IGFs by cleaving IGF-binding proteins.

Braz J Med Biol Res, 1996 Dec, 29(12), 1599 - 602
Cloning and sequencing of the nitrogenase structural genes nifHDK of Herbaspirillum seropedicae; Machado IM et al.; The nitrogenase structural genes (nifHDK) of the endophytic diazotroph Herbaspirillum seropedicae were isolated from a genomic bank by plate hybridization . Sequence analysis of the DNA showed a consensus promoter region upstream for the nifH gene containing a -24/-12 type promoter together with NifA- and integration host factor (IHF)- binding sites . The derived protein sequences of NifH, NifD and NifK contained conserved cysteine residues for binding iron-sulfur clusters and the iron-molybdenum cofactor . These protein sequences showed the strongest similarities to the nifHDK gene products of the symbiotic diazotroph Bradyrhizobium japonicum (93.5%, 91.3% and 83.3%, respectively), the plant-associated diazotroph Azospirillum brasilense (90.0%, 83.7% and 75.1%, respectively) and to Thiobacillus ferrooxidans (91.0%, 83.4% and 81.1%, respectively) of the same phylogenetic group of the protobacteria.

J Endod, 1996 Dec, 22(12), 643 - 5
Removal of smear layer in the root canal using oxidative potential water; Hata G et al.; We investigated oxidative potential water (OPW) for its ability to remove the smear layer using a scanning electron microscope . OPW has been studied mainly in Japan and is known to suppress bacteria and viruses without harming living systems . We found that OPW used as in irrigant during and after root canal instrumentation is as effective as 5% NaOCl or 17% EDTA for opening and keeping patent the dentinal tubules.

Voen Med Zh, 1996 Dec, 317(12), 26 - 8, 80
{The efficacy of using ozone preparations in the combined treatment of paranasal sinusitis}; Petrov GM et al.; In controllable study efficiency of application of ozone preparations in 39 patients with acute and chronic purulent maxillary polysinusitis was studied . Purulent maxillary sinusitis, which in 44% was combined with frontitis and in 56%--with fronto-ethmoiditis was diagnosed in all patients . Besides traditional therapy all patients were washed out daily inflamed sinuses with 0.9% solution of sodium chloride, saturated of ozone . After washing out the sinuses were aerated during 15-20 minutes through paracentesis needles and catheters with ozone-oxygen mixture, containing 6 mg/l of ozone . Speed of mixture flow--1 1/min . To the half of patient with phenomena of intoxication were introduced I/V 400 ml of physiologic salt solution (ozone concentration--0.8 mg/l) . Treatment rate is 3-6 infusions . In patients pyorrhea and allocation of bacteria stopped 2-3 days earlier, than in control group, level of molecules of average weight was faster reduced to norm in the plasma of blood . Reduction of concentration of molecules of average weight corresponded to essential improvement of patient clinical condition (P < 0.01) . Experience of ozone application testified about its high efficiency in paranasal sinusitis treatment.

Ital J Gastroenterol, 1996 Dec, 28(9), 512 - 7
Helicobacter pylori infection in families of Helicobacter pylori-positive children; Bonamico M et al.; Relatives of Helicobacter pylori positive patients show a higher incidence of Helicobacter pylori infection than the general population, probably due to relapses and/or reinfections between members of the family . Aim of this study was to evaluate the prevalence of the infection in 121 relatives of 41 children with Helicobacter pylori positive gastritis . Specific IgG antibodies (ELISA) were evaluated, and bacteria on gastric biopsy specimens were investigated by urease-rapid test, culture test and GIEMSA or acridine orange staining . Of the eighty-two relatives, 68% were antibody positive . Thirty-five agreed to undergo endoscopy . With the exception of one brother, all subjects (97%) were found to be infected by Helicobacter pylori . Two symptomatic relatives, with normal antibody titres, were submitted to endoscopy and found to be colonized by Helicobacter pylori . The present data confirm the high prevalence of infection within families and appear to demonstrate the usefulness of endoscopy for all subjects showing positive antibody titres as well as for symptomatic relatives, even if serologically negative, to confirm the presence of any pathological conditions and reduce the risk of relapses within families.

Can Vet J, 1996 Dec, 37(12), 729 - 34
Effects of formalin, chloramine-T, and low salinity dip on the behavior and hemolymph biochemistry of the American lobster; Speare DJ et al.; The purpose of this research was to investigate the salinity and formalin sensitivity of a ciliate parasite (Anophryoides haemophila) of the American lobster (Homarus americanus), and to examine the target-animal (lobster) safety of chemical-bath treatments involving low salinity, formalin, or chloramine-T that could be used to control this parasite in lobster pounds . "Bumper car" disease, caused by An . haemophila, is an important concern to lobster pound operators in eastern North America, because of the implicated lobster mortality rate and the general lack of preventive and therapeutic intervention regimes . We determined, using an in vitro method, that formalin at 50 mg/L, or low salinity at 8.0 parts per thousand (ppt) for 1 hour killed 100% of the parasites . When healthy lobsters were exposed to formalin at 200 mg/L, there were no negative behavioral responses and no significant differences in a panel of hemolymph biochemical indices . Similar results occurred when lobsters were exposed to chloramine-T, a common finfish therapeutic agent for topical bacteria and protozoa, at 10 mg/L for 1 hour . The low salinity treatment (8.0 ppt) resulted in significant adverse changes in lobster behavior and biochemical indices; however, these changes did not persist for more than 1 week after treatment ended . Although these treatments are unlikely to kill parasites that have already invaded the lobster carapace, they should be effective in reducing parasite loads on the gill and carapace surface of the lobster and in the environment of the impoundment housing.

Recenti Prog Med, 1996 Dec, 87(12), 616 - 22
{Common variable immunodeficiency}; Silvestris N et al.; Common variable immunodeficiency (CVI) is a heterogeneous syndrome characterized by defective antibody formation, resulting in abnormally low serum immunoglobulin levels . Clinical presentation usually includes recurrent infections of the respiratory tract, mostly induced by capsular bacteria . Patients are also highly prone to Giardia lamblia infections and related gastrointestinal disorders, as well as to a variety of autoimmune diseases which appear in approximately 20% of them . In addition, CVI can be frequently associated with a non-Hodgkin lymphoma or gastric carcinoma . In spite of its relatively frequent occurrence, the pathogenesis of CVI still remains poorly defined . In this review the authors describe clinical features, immunological abnormalities and replacement treatment with intravenous immunoglobulins of this antibody deficiency syndrome.

Recenti Prog Med, 1996 Dec, 87(12), 594 - 6
{Common variable immunodeficiency . The authors' personal cases from the last 5 years}; Silvestris N et al.; Common variable immunodeficiency (CVI) is a heterogeneous syndrome characterized by defective antibody formation, resulting in abnormally low serum immunoglobulin levels . Clinical presentation usually includes recurrent infections of the respiratory tract, mostly induced by capsular bacteria . Patients are also highly prone to Giardia lamblia infections and related gastrointestinal disorders, as well as to a variety of autoimmune diseases which appear in approximately 20% of them . In addition, CVI can be frequently associated with a non-Hodgkin lymphoma or gastric carcinoma . In spite of its relatively frequent occurrence, the pathogenesis of CVI still remains poorly defined . An overview of the syndrome is published in this issue (pages 616-622) . The authors experience (18 cases during last five years) is here reported.

Arch Microbiol, 1996 Dec, 166(6), 405 - 10
Competition between beta-ketothiolase and citrate synthase during poly(beta-hydroxybutyrate) synthesis in Methylobacterium rhodesianum; Mothes G et al.; The enzymes beta-ketothiolase and citrate synthase from the facultatively methylotrophic Methylobacterium rhodesianum MB 126, which uses the serine pathway, were purified and characterized . The beta-ketothiolase had a relatively high Km for acetyl-CoA (0.5 mM) and was strongly inhibited by CoA (Ki 0.02 mM) . The citrate synthase had a much higher affinity for acetyl-CoA (Km 0.07 mM) and was significantly inhibited by NADH (Ki 0.15 mM) . The intracellular concentration of CoA metabolites and nucleotides was determined in M . rhodesianum MB 126 during growth on methanol . The level of CoA decreased from about 0.6 nmol (mg dry mass)-1 during growth to the detection limit when poly(beta-hydroxybutyrate) (PHB) accumulated . Nearly unchanged intracellular concentrations of NADH, NADPH, and acetyl-CoA of about 0.5, 0.6-0.7, and 1.0 nmol (mg dry mass)-1, respectively, were determined during growth and PHB synthesis . During growth, the beta-ketothiolase was almost completely inhibited by CoA, and acetyl-CoA was principally consumed by the citrate synthase . During PHB accumulation, the beta-ketothiolase had about 75% of its maximum activity and showed much higher activity than citrate synthase, which at the actual NADH concentration was about 75% inhibited . NADPH concentration was sufficiently high to allow the unlimited activity of acetoacetyl-CoA reductase (Km NADPH 18 microM) . PHB synthesis is probably mainly controlled by the CoA concentration in M . rhodesianum MB 126.

Br J Theatre Nurs, 1996 Dec, 6(9), 29 - 32
The role of air ventilation and air sampling in reducing the incidence of surgical wound infection rates; Madeo M; Air ventilation systems are a common feature in today's operating theatres . The main function is to decrease the bacterial load in the operating theatre, reducing the incidence of surgical wound infection rates . The ventilation system is only one of many factors which can contribute to a safe operating environment . Ever since the time of Lister the significance of airborne bacteria in the operating room has been a matter of interest and of dispute . The role of airborne bacteria acting as a source of infection for most types of operations continues to be a matter for debate . Despite the uncertainty, the development of laminar air flow and Plenum (conventional) ventilation systems has gained momentum, making it an essential standard of design and ventilation in the modern operating theatre.

Br J Surg, 1996 Dec, 83(12), 1779 - 81
High prevalence of Helicobacter pylori infection in duodenal ulcer perforations not caused by non-steroidal anti-inflammatory drugs; Ng EK et al.; There has been controversy regarding the relationship between Helicobacter pylori and perforated peptic ulcer, which is known to have a high recurrence rate if only simple patch repair is performed . The aim of this study was to evaluate the association between H . pylori infection and intake of non-steroidal anti-inflammatory drugs (NSAIDs) in patients with perforated duodenal ulcers . Of the 73 patients recruited over a 16-month period, 51 (70 per cent) had evidence of H . pylori infection by intraoperative gastroscopy and antral biopsies . The infection rate rose to 80 per cent if NSAID users were excluded . The H . pylori-infected group was significantly younger (mean 47.6 versus 62.5 years), with a male preponderance (49 of 51 versus 14 of 22 patients), and had significantly less NSAID consumption (three of 51 versus ten of 22) and more prolonged dyspepsia (40 of 51 versus ten of 22), compared with H . pylori-negative patients . H . pylori infection probably plays an important role in the causation of non-NSAID-induced duodenal ulcer perforation . Whether eradication of the bacteria can alleviate the strong ulcer diathesis in this subgroup of patients is unknown.

Plant Cell Physiol, 1996 Dec, 37(8), 1150 - 60
Disruption analysis of the gene for a cold-regulated RNA-binding protein, rbpA1, in Anabaena: cold-induced initiation of the heterocyst differentiation pathway; Sato N et al.; A cold-regulated operon, rbpA1-rpsU, encodes an RNA-binding protein and a ribosomal protein in Anabaena variabilis M3 . The level of expression of this gene cluster was about ten times higher at temperatures below 30 degrees C than at 38 degrees C . To study the role of the RbpA1 protein in vivo, we constructed insertional disruptants of rbpA1 . These strains were totally devoid of RbpA1 protein but contained a normal level of the ribosomal protein S21, a product of the rpsU gene . The disruptants were morphologically normal at 38 degrees C, but at 22 degrees C they produced unusual cells at a low frequency . These cells were probably at an initial stage of proheterocyst formation . Various molecular events that are related to heterocyst initiation, namely, excision of the 11-kbp DNA element in nifD and the accumulation of transcripts of xisA and hetR, also occurred in the disruptants at 22 degrees C in the presence of nitrate ions, but these events did not occur in the presence of ammonium ions or at 38 degrees C . The results suggest that RbpA1 is required for enhanced repression of heterocyst initiation at low temperatures in the presence of nitrate . Possible mechanisms of the action of RabA1 are discussed.

Plant Cell Physiol, 1996 Dec, 37(8), 1043 - 8
The excessive production of indole-3-acetic acid and its significance in studies of the biosynthesis of this regulator of plant growth and development; Kawaguchi M et al.; Because of the importance of indole-3-acetic acid (IAA) in the growth and development of plants, extensive studies of the biosynthesis of IAA have been performed during the four decades since the discovery of IAA as a plant hormone . The pathway for the biosynthesis of IAA in plants remains, however, to be unelucidated, even though studies within the past decade have revealed unexpected aspects of such biosynthesis . By contrast, two pathways to IAA have been characterized in bacteria at the molecular level: the indole-3-acetamide (IAM) pathway (L-tryptophan-->IAM-->IAA); the indole-3-pyruvic acid pathway (L-tryptophan-->indole-3-pyruvic acid-->indole-3-acetaldehyde-->IAA) (Fig . 1) . In both pathways, the details of the biosynthesis of IAA were clarified using IAA-overproducing bacteria . After a description of recent advances of the studies of the biosynthesis of IAA in plants, this review focuses on the excessive production of IAA in several organisms and its significance in the studies of the biosynthesis of IAA.

J Gastroenterol, 1996 Dec, 31(6), 907 - 16
Alterations of the mucosal immune system in inflammatory bowel disease; MacDermott RP; The normal intestinal immune system is under a balance in which proinflammatory and anti-inflammatory cells and molecules are carefully regulated to promote a normal host mucosal defense capability without destruction of intestinal tissue . Once this careful regulatory balance is disturbed, nonspecific stimulation and activation can lead to increased amounts of potent destructive immunologica and inflammatory molecules being produced and released . The concept of balance and regulation of normal mucosal immune and inflammatory events is indicative of how close the intestine is to developing severe inflammation . The normal intestinal mucosal immune system is constantly stimulated by lumenal contents and bacteria . The stimulatory molecules present in the intestinal lumen that activate and induce subsequent mucosal immunologic and inflammatory events include bacterial cell wall products, such as peptidoglycans and lipopolysaccharides, as well as other chemotactic and toxic bacterial products that are produced by the many different types of bacteria within the gastrointestinal tract . These highly stimulatory bacterial cell wall products are capable of activating macrophages and T lymphocytes to release potent proinflammatory cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-alpha) . IL-1, IL-6, and TNF-alpha increase the presence of human leukocyte antigen (HLA) class II antigen-presenting molecules on the surfaces of epithelial cells, endothelial cells, macrophages, and B cells, thus increasing their ability to present lumenal antigens and bacterial products . The proinflammatory cytokines IL-1 and TNF-alpha also increase the ability of epithelial cells, endothelial cells, macrophages, and fibroblasts to secrete potent chemotactic cytokines, such as interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1), which serve to increase the movement of macrophages and granulocytes from the circulation into the inflamed mucosa . Thus, through lumenal exposure to potent, nonspecific stimulatory bacterial products, the state of activation of the intestinal immune system and mucosal inflammatory pathways are markedly up-regulated . This raises the question of whether there is a deficiency in effective down-regulation through the absence of normally suppressive cytokines such as interleukin-10 (IL-10), transforming growth factor-beta (TGF-beta), interleukin-4 (IL-4), and IL-1 receptor antagonist . Normally, the turning off of the active and destructive immunologic and inflammatory events should occur following the resolution of a bacterial or viral infection that has been appropriately defended against and controlled by the mucosal immune system . In inflammatory bowel disease (IBD), however, the down-regulatory events and processes that should turn off the immunologic and inflammatory protective processes, once the pathogenic agent has been cleared, appear to be deficient or only partially effective . We may find that we ultimately are dealing with disease processes that have more than one genetic or cellular basis . The improved understanding of the immunopathophysiology of IBD will allow exploration of novel immunologic and genetic approaches, such as gene replacement therapy, administration of a suppressor cytokine or an altered cell surface antigen, the administration of humanized monoclonal antibodies directed against proinflammatory cytokines, or the development of newer strategies against fundamental cell biologic mechanisms such as adhesion molecules.

Nihon Kyobu Shikkan Gakkai Zasshi, 1996 Dec, 34(12), 1380 - 4
{Pulmonary actinomycosis--diagnosis and therapy based on past reports of actinomycosis}; Tanaka S et al.; A 57-year-old man who complained of bloody sputum was admitted to the hospital because of a spherical mass on a chest X-ray film . Despite a detailed examination, no definitive diagnosis was obtained, and a left lower lobectomy was done . Histopathologic examination of tissue from the resected lobe revealed sulfur granules and branching filamentous bacteria, and therefore pulmonary actinomycosis was diagnosed . Examination of reports published in Japan in the last five years suggested a slight increase in the frequency of diagnosis before treatment of actinomycosis and a slight decrease in the frequency of surgical treatment . However, these changes are not surprising because the frequency of surgical treatment is 58% . These findings suggest that pulmonary actinomycosis should be included in the differential diagnosis of a mass on a chest X-ray film.

Nutr Clin Pract, 1996 Dec, 11(6), 269 - 73
The effects of rinsing enteral delivery sets on formula contamination; Kohn-Keeth C et al.; This study investigated whether rinsing enteral delivery sets before formula addition affects formula contamination . Both a simulated and a clinical phase were conducted . In the simulated phase, Osmolite (Ross Laboratories, Columbus, OH) was infused continuously through 52 delivery sets into a flask via enteral infusion pumps for 24 hours . The delivery sets were randomly assigned to two groups of equal size . One group was rinsed with tap water before new formula was added at 8 and 16 hours, and the other group was not rinsed . At 8, 16, and 24 hours, samples of formula were collected from the delivery sets, and bacteria counts were obtained . In the clinical phase, 23 critically ill patients receiving Osmolite continuously were randomly assigned to a rinse or no-rinse group . The same formula addition and rinse protocol from phase I was used . Formula samples were obtained at 24 hours . In both phases, there were no significant differences between the rinse and no-rinse groups with respect to bacteria counts at any time period . The findings suggest that rinsing may be unnecessary if delivery sets are used continuously for 24 hours or less; however, the possibility of a type II error because of the small sample size of this study must be recognized.

Immunology, 1996 Dec, 89(4), 613 - 8
Lipids from Mycobacterium leprae cell wall are endowed with an anti-inflammatory property and inhibit macrophage function in vivo; Moura AC et al.; In general, the majority of bacteria are pre-inflammatory when injected in experimental animals . However, Mycobacterium leprae has no inflammatory effect when injected into mouse footpad, but using the delipidated mycobacteria we observed a mild significant increase in footpad oedema . Other mycobacteria, Mycobacterium bovis-BCG or M . tuberculosis induce a strong paw oedema . Furthermore, M . leprae reduced locally the BCG-induced inflammatory reaction in mouse footpad, whereas delipidated M . leprae did not influence this reaction . Both M . leprae and M . leprae cell wall lipids blocked immune phagocytosis in vivo by inflammatory macrophages (from an induced focus) . In contrast delipidated M . leprae stimulated the phagocytosis reaction . Neither intact M . leprae . delipidated M . leprae, nor its lipids had any toxic effect on macrophages or on cell migration . Although M . leprae did not interfere on cell influx and cell type in an induced-inflammatory site, this mycobacterium led to the appearance of a distinct cell population in vivo . The hypothesis is that M . leprae would transform macrophages in epithelioid cells, suggested by morphology analysis of cells by fluorescence-activated cell sorter and observed under optic microscopy.

Dig Dis Sci, 1996 Dec, 41(12), 2477 - 81
Antagonistic effects of sulfide and butyrate on proliferation of colonic mucosa: a potential role for these agents in the pathogenesis of ulcerative colitis; Christl SU et al.; It has been shown that feces of patients with ulcerative colitis uniformly contain sulfate reducing bacteria . Sulfide produced by these bacteria interferes with butyrate-dependent energy metabolism of cultured colonocytes and may be involved in the pathogenesis of ulcerative colitis . Mucosal biopsies from the sigmoid rectum of 10 patients (no caner, polyps, inflammatory bowel disease) were incubated with either NaCl, sodium hydrogen sulfide (1 mmol/L), a combination of both sodium hydrogen sulfide and butyrate (10 mmol/L), or butyrate . Mucosal proliferation was assessed by bromodeoxyuridine labeling of cells in S-phase . Compared to NaCl, sulfide increased the labeling of the entire crypt significantly, by 19% (p < 0.05) . This effect was due to an expansion of the proliferative zone to the upper crypt (compartments 3-5), where the increase in proliferation was 54% . Sulfide-induced hyperproliferation was reversed when samples were coincubated with sulfide and butyrate . The study shows that sodium hydrogen sulfide induces mucosal hyperproliferation . Our data support a possible role of sulfide in the pathogenesis of UC and confirm the role of butyrate in the regulation of colonic proliferation and in the treatment of UC.

J Biomol NMR, 1996 Dec, 8(4), 477 - 86
AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR; Laskowski RA et al.; The AQUA and PROCHECK-NMR programs provide a means of validating the geometry and restraint violations of an ensemble of protein structures solved by solution NMR . The outputs include a detailed breakdown of the restraint violations, a number of plots in PostScript format and summary statistics . These various analyses indicate both the degree of agreement of the model structures with the experimental dat, and the quality of their geometrical properties . They are intended to be of use both to support ongoing NMR structure determination and in the validation of the final results.

Ecotoxicol Environ Saf, 1996 Dec, 35(3), 231 - 5
Toxicity identification evaluation of Lake Orta (Northern Italy) sediments using the Microtox system; Guzzella L et al.; Pore waters extracted by centrifugation from Lake Orta (Northern Italy) sediments were studied with a modified Toxicity Identification Evaluation (TIE) procedure using the Microtox bacterial luminescence toxicity test system . The most toxic pore water samples were from stations near a rayon factory, known as a source of copper and ammonium discharges . The TIE manipulations used were filtration, EDTA chelation, and C18 solid-phase resin adsorption . The most effective treatments to remove toxicity were the EDTA and C18, indicating that both metals and nonpolar organic compounds contribute to the observed toxicity.

Microbiology, 1996 Dec, 142 ( Pt 12), 3525 - 30
Phylogenetic diversity of intra-amoebal legionellae as revealed by 16S rRNA gene sequence comparison; Birtles RJ et al.; Following a recent phylogenetic analysis of all 39 species of the genus Legionella and related organisms, the name Legionella lytica has been proposed for bacteria previously considered either as Sarcobium lyticum or as 'Legionella-like amoebal pathogens' (LLAPs) . To investigate the phylogenetic integrity of this newly proposed species, we determined the 16S rRNA gene sequences of 10 LLAPs isolated from various environmental sources . All 10 isolates clustered within a monophyletic group containing all other members of the genus Legionella . Eight of the 10 isolates formed a monophyletic subgroup within the genus which also included the two previously characterized L . lytica strains . Four of these 10 isolates shared a specific and very close relationship with the L . lytica type strain (> 99% sequence similarity) . However, although clearly legionellae, the remaining two LLAP strains bore no specific evolutionary relationship to either L . lytica or any other Legionella species (< 96% sequence similarity) . Both isolates lay on their own relatively deep-rooted branches within the radiation of the Legionella cluster . LLAPs do not, therefore, represent a unique species or even a single line of descent within the genus, and investigation of more isolates may reveal them to be as evolutionarily diverse as the other presently recognized Legionella species.

FASEB J, 1996 Dec, 10(14), 1598 - 606
Osmotic regulation of gene expression; Burg MB et al.; Cells react to increased osmolality with numerous changes in gene expression . The specific genes affected differ between species, but the known osmoprotective effects of the gene products are remarkably similar, particularly with regard to cellular accumulation of compatible organic osmolytes . Here we concentrate on the molecular basis for osmotic regulation of gene expression, emphasizing certain genes expressed in bacteria, yeast, and the mammalian renal medulla because their expression is best understood . Thus, we emphasize 1) bacterial and yeast two-component histidine kinase systems, each consisting of a membrane osmolality sensor and a separate cytoplasmic response regulator that, when phosphorylated, alters transcription, 2) volume regulatory increases in cellular K+ salts that can prompt increased gene transcription in bacteria through direct effects on DNA and that in mammalian renal cells increase transcription, seemingly via trans-activating proteins, 3) a yeast kinase cascade that transmits an osmotic signal to the gene regulating the level of glycerol, and 4) in mammalian cells, several homologous cascades that are activated by hypertonicity, but whose osmoregulatory targets are not yet known.

J Periodontol, 1996 Dec, 67(12), 1251 - 9
Full- versus partial-mouth disinfection in the treatment of periodontal infections . Long-term clinical observations of a pilot study; Vandekerckhove BN et al.; A classical treatment for chronic adult periodontitis consists of four to six consecutive sessions of scaling and root planing at a 1- to 2-week interval . Such a so-called "quadrant or sextant therapy" might result in a reinfection of a previously disinfected area by bacteria from an untreated region . The purpose of this study was to investigate, over an 8-month period, the clinical benefits of full-mouth disinfection within a 24-hour period in the control of chronic periodontitis . Ten adult patients with advanced chronic periodontitis were randomly assigned to a test and a control group . The control group received the standard scheme of initial periodontal therapy, consisting of scaling and root planing of the four quadrants was performed within 24 hours and immediately followed by a thorough supra- and subgingival chlorhexidine application to limit any transfer of bacteria . The latter involved tongue brushing with a 1% chlorhexidine gel for 60 seconds, mouthrinsing with a 0.2% chlorhexidine solution twice for 60 seconds, repeated subgingival irrigation of all pockets with a 1% chlorhexidine gel (3 times within 10 minutes), and mouthrinsing twice daily with a 0.2% chlorhexidine solution during 2 weeks . In addition, both groups received thorough oral hygiene instructions . The plaque index, gingival index, probing depth, gingival recession, and bleeding on probing were recorded prior to professional cleaning and at 1, 2, 4, and 8 months afterwards . Although the test group scored higher plaque indices than the control group, especially at months 2 and 4, the gingival index and bleeding tendency showed similar improvements with time . However, when the gingival/plaque ratio was considered, the latter was lower in the test group at all follow-up visits . For pockets > or = 7 mm, full-mouth disinfection showed a significantly (P = 0.01) higher reduction in probing depth at each follow-up visit with, at month 8, a reduction of 4 mm (from 8 mm to 4 mm), in comparison to 3 mm (from 8 mm to 5 mm) for the classical therapy . The increase in gingival recession in the full-mouth disinfection group remained below 0.7 mm, while in the control group it reached 1.9 mm after 8 months . This resulted in a gain of clinical attachment level of 3.7 mm for the test group versus 1.9 mm for the control group . A radiographical examination also indicated a superior improvement for the test group when compared to the control group . This pilot study suggests that a full-mouth disinfection in one day results in an improved clinical outcome in chronic periodontitis as compared to scalings per quadrant at 2-week intervals over several weeks.

Can J Microbiol, 1996 Dec, 42(12), 1248 - 51
Detection and identification of Entamoeba gingivalis by specific amplification of rRNA gene; Kikuta N et al.; A pair of oligonucleotide primers were designed from the nucleotide sequence of the gene encoding the small subunit ribosomal RNA (SrRNA) of the oral protozoan parasite Entamoeba gingivalis . The primers amplified a 1.4-kb DNA fragment by polymerase chain reaction and were specific for Entamoeba gingivalis but not for other protozoa, oral protists and bacteria, or human leukocytes . With this method, the DNA from as few as 30 cells of Entamoeba gingivalis could be detected . These results suggest that this approach is applicable to the detection and identification of Entamoeba gingivalis in the human oral cavity.

Mol Mar Biol Biotechnol, 1996 Dec, 5(4), 299 - 303
Isolation of cDNA encoding a putative chitinase precursor in the kuruma prawn Penaeus japonicus; Watanabe T et al.; Amino acid sequences of chitinases have been determined in insects, plants, yeast, and bacteria, but not in crustaceans . We searched for chitinase-encoding cDNA in the kuruma prawn Penaeus japonicus by polymerase chain reaction (PCR) amplification of hepatopancreas cDNA using degenerate oligonucleotide primers derived from the two conserved regions of known chitinases . Using a PCR product as a probe, a cDNA clone was isolated . This clone contains an open reading frame for a protein (named Pjchi-1) of 572 amino acids that exhibits sequence similarities to known chitinases, especially to a chitinase from the tobacco hornworm Manduca sexta . Transcription of the mRNA was detected in the hepatopancreas but not in epidermal tissues.

Photochem Photobiol, 1996 Dec, 64(6), 949 - 52
Heat shock transcription factor, HSF, is activated by ultraviolet irradiation; Ohnishi K et al.; We demonstrated previously that human glioblastoma cell lines accumulated heat shock protein (hsp)72, not only after heat shock, but also after, gamma-ray or UV irradiation . In the present study, we investigated whether the binding activity of heat shock transcription factor (HSF) to the heat shock element (HSE) of the hsp72 gene promoter increased after UV irradiation of human glioblastoma A-172 cells . A gel mobility-shift assay showed that the activated HSF level increased markedly after UV irradiation . Furthermore, UV irradiation of nuclear extracts in vitro did not activate HSF, whereas in vitro heat shock treatment did . These results suggest that HSF activation can be induced by UV irradiation at normal physiological temperature and hsp72 accumulation results from an increased activated HSF level, i.e . a transcriptional up-regulation of hsp72 . In addition, the mechanism responsible for UV-induced HSF activation may differ from the process that operates in heat-treated cells.

Mol Microbiol, 1996 Dec, 22(5), 919 - 28
Localized domains of DNA supercoiling: topological coupling between promoters; Mojica FJ et al.; According to the twin-supercoiled domain model, a local domain with a high level of DNA supercoiling can be generated between two divergently transcribed promoters . We have tested this model directly using the rate of DNA photo-cross-linking by 4,5', 8-trimethylpsoralen to measure local regions of unconstrained supercoiling in vivo . We demonstrate that, in plasmids, a localized domain of highly supercoiled DNA is generated upstream of the tetA promoter . The level of supercoiling of this domain depends on the activity of the tetA promoter and on the extent of translation of the tetA gene product . The highest level of supercoiling within this localized domain was observed in a topA strain, although the generation of a localized supercoiling domain could also be detected in a topA+ background, consistent with potential physiological significance . The level of supercoiling in this localized domain correlated with activation of the supercoiling-sensitive leu-500 promoter located upstream of tetA . These data provide a direct demonstration that localized domains of increased negative supercoiling can be generated upstream of an actively transcribed promoter, and that this can result in supercoiling-mediated transcriptional coupling between two promoters.

Mol Microbiol, 1996 Dec, 22(5), 895 - 908
Comparative analysis of the virulence control systems of Bordetella pertussis and Bordetella bronchiseptica; Martinez de Tejada G et al.; Bordetella pertussis and Bordetella bronchiseptica contain nearly identical BvgAS signal-transduction systems that mediate a biphasic transition between virulent (Bvg+) and avirulent (Bvg-) phases . In the Bvg+ phase, the two species express a similar set of adhesins and toxins, and in both organisms the transition to the Bvg- phase occurs in response to the same environmental signals (low temperature or the presence of nicotinic acid or sulphate anion) . These two species differ, however, with regard to Bvg(-)-phase phenotypes, host specificity, the severity and course of the diseases they cause, and also potentially in their routes of transmission . To investigate the contribution of the virulence-control system to these phenotypic differences, we constructed a chimeric B . bronchiseptica strain containing bvgAS from B . pertussis and compared it with wild-type B . bronchiseptica in vitro and in vivo . The chimeric strain was indistinguishable from the wild type in its ability to express Bvg(+)- and Bvg(-)- phase-specific factors . However, although the chimeric strain responded to the same signals as the wild type, it differed dramatically in sensitivity to these signals; significantly more nicotinic acid or MgSO4 was required to modulate the chimeric strain compared with the wild-type strain . Despite this difference in signal sensitivity, the chimeric strain was indistinguishable from the wild type in its ability to cause respiratory-tract infections in rats, indicating that the bvgAS loci of B . pertussis and B . bronchiseptica are functionally interchangeable in vivo . By exchanging discrete fragments of bvgAS, we found that the periplasmic region of BvgS determines signal sensitivity.

Mol Biol Cell, 1996 Dec, 7(12), 1895 - 907
Is outer arm dynein intermediate chain 1 multifunctional?
Ogawa K, Takai H, Ogiwara A, Yokota E, Shimizu T, Inaba K, Mohri H.
The outer arm dynein of sea urchin sperm axoneme contains three intermediate chains (IC1, IC2, and IC3; M(r) 128,000, 98,000, and 74,000, respectively) . IC2 and IC3 are members of the WD family; the WD motif is responsible for a protein-protein interaction . We describe here the molecular cloning of IC1 . IC1 has a unique primary structure, the N-terminal part is homologous to the sequence of thioredoxin, the middle part consists of three repetitive sequences homologous to the sequence of nucleoside diphosphate kinase, and the C-terminal part contains a high proportion of negatively charged glutamic acid residues . Thus, IC1 is a novel dynein intermediate chain distinct from IC2 and IC3 and may be a multifunctional protein . The thioredoxin-related part of IC1 is more closely related to those of two redox-active Chlamydomonas light chains than thioredoxin . Antibodies were prepared against the N-terminal and middle domains of IC1 expressed as His-tagged proteins in bacteria . These antibodies cross-reacted with some dynein polypeptides (potential homologues of IC1) from distantly related species . We propose here that the three intermediate chains are the basic core units of sperm outer arm dynein because of their ubiquitous existence . The recombinant thioredoxin-related part of IC1 and outer arm dyneins from sea urchin and distantly related species were specifically bound to and eluted from a phenylarsine oxide affinity column with 2-mercaptoethanol, indicating that they contain vicinal dithiols competent to undergo reversible oxidation/reduction.

Kyobu Geka, 1996 Dec, 49(13), 1119 - 21
{A case of endobronchial tuberculosis with peripheral mucoid impaction showing a solitary pulmonary nodular shadow}; Miyoshi K et al.; In a 70-year-old nonsmoking, asymptomatic man, chest radiography before a operation for early gastric cancer revealed a solitary nodular shadow in the right lower lobe . Laboratory studies for bacteria and mycobacteria were negative and histological diagnosis could not be obtained . Right lower lobectomy was performed because the mass could be malignant . Histologically the right B10 was almost completely obstructed by tuberculous granuloma and the distal bronchus were dilated containing inspissated mucus . The abnormal nodular shadow was considered the obstructive mucoid impaction secondary to tuberculous bronchitis . It is quite rare that solitary nodular shadow is found in such a case of endobronchial tuberculosis.

Am J Kidney Dis, 1996 Dec, 28(6), 943 - 50
Bowel as a kidney substitute in renal failure; Friedman EA; Maintenance peritoneal dialysis and hemodialysis sustain the lives of approximately 250,000 uremic patients in industrialized nations worldwide . The cost of uremia therapy, however, exceeds health care budgets in developing countries . As a consequence, most of those alive today have no chance of effective treatment should their kidneys fail . Extraction, modification, or recycling of nitrogenous wastes by the gastrointestinal tract is a potentially low-cost means of substituting for missing renal function . Multiple approaches to the bowel as a substitute kidney have been attempted . Direct removal of nitrogen-containing compounds by an external gut fistula, gastric, ileal, or colonic gavage (dialysis) or induced diarrhea extract water and urea but only minimal amounts of larger molecules such as creatinine . Binding of nitrogen compounds to inert orally administered sorbents such as charcoal or oxystarch has been pursued in advanced uremia . Modification of nitrogen compounds by ingesting enzymes derived from soil bacteria or packaged in artificial cells is an approach that, although exciting, is incompletely evaluated . Evidence indicates that strains of bacteria can be induced to synthesize enzymes that recycle urea and other nitrogen compounds retained by uremic individuals . Clinical trials to substantiate uncontrolled trials of bowel substitution in renal failure will, if positive, accelerate development of a practical regimen to extend life where no other means are possible.

J Bacteriol, 1996 Dec, 178(24), 7031 - 6
Heat induction of hsp18 gene expression in Streptomyces albus G: transcriptional and posttranscriptional regulation; Servant P et al.; In Streptomyces albus G, HSP18, a protein belonging to the small heat shock protein family, could be detected only at high temperature . The nucleotide sequence of the DNA region upstream from hsp18 contains an open reading frame (orfY) which is in the opposite orientation and 150 bp upstream . This open reading frame encodes a basic protein of 225 amino acids showing no significant similarity to any proteins found in data banks . Disruption of this gene in the S . albus chromosome generated mutants that synthesized hsp18 RNA at 30 degrees C, suggesting that orfY plays either a direct or indirect role in the transcriptional regulation of the hsp18 gene . In addition, thermally induced expression of the hsp18 gene is subject to posttranscriptional regulation . In the orfY mutant, although hsp18 RNA was synthesized at a high level at 30 degrees C, the HSP18 protein could not be detected except after heat shock . Synthesis of the HSP18 protein in the orfY mutant was also heat inducible when transcription was inhibited by rifampin . Furthermore, when wild-type cultures of S . albus were shifted from high temperature to 30 degrees C, synthesis of the gene product could no longer be detected, even though large amounts of hsp18 RNA were present.

Anal Biochem, 1996 Dec 1, 243(1), 1 - 7
BODIPY-alpha-casein, a pH-independent protein substrate for protease assays using fluorescence polarization; Schade SZ et al.; BODIPY-alpha-casein is a new fluorescent protein substrate designed for fluorescence polarization studies to measure proteolytic activity at any pH over the range from pH 2 to 11 . Kinetic protease assays in real-time were performed in 1 to 5 min using an FPM-1 fluorescence polarization instrument . A purified enzyme or bacterial culture was mixed with the BODIPY-alpha-casein in a buffer of an appropriate pH and the decrease in fluorescence polarization was automatically recorded at 0.5-min intervals . The initial decrease in fluorescence polarization with time was dependent on protease concentration . In 3-min assays at 37 degrees C, the sensitivity of detection was 8 mU for pepsin at pH 2.0, 1 mU for papain at pH 6.0, 0.6 mU for proteinase K at pH 7.4, and 2 mU for Streptomyces griseus alkaline protease at pH 11 . Only 1-10 microliters of a growing culture was necessary to assay the protease activity of Porphyromonas gingivalis or Treponema denticola, oral bacteria that possess certain proteases on their surfaces . These assays have clinical applications, since certain pathogens use proteolytic activity as a virulence mechanism and differ from their nonpathogenic counterparts in this characteristic . Fluorescence polarization assays are simple, rapid, and reproducible.

Magn Reson Med, 1996 Dec, 36(6), 829 - 33
Finger printing of Mycobacterium tuberculosis in patients with intracranial tuberculomas by using in vivo, ex vivo, and in vitro magnetic resonance spectroscopy; Gupta RK et al.; In vivo, ex vivo, and in vitro proton magnetic resonance spectroscopy was performed in 12 patients with intracranial tuberculomas with an aim of detecting the biochemical constituents of Mycobacterium tuberculosis in a granuloma . One dimensional (1D) single pulse and spin-echo sequences and 2D correlative spectroscopy were used for the ex vivo study to confirm the resonances seen on in vivo study . Spectroscopic studies of the perchloric acid and lipid extract of granuloma and M . tuberculosis were performed to look for similarity of resonance . In vivo study showed the presence of lipids at 0.9, 1.3, 2.0, 2.8 ppm, and phosphoserine at 3.7 ppm . All these resonances were confirmed on ex vivo study . In addition, distinct resonances of serine and phenolic lipids were seen on ex vivo and in vitro study of tuberculous granuloma, which have not been observed in other intracranial tumors . Lipid extract of granuloma and M . tuberculosis showed phenolic lipids at 7.1 and 7.4 ppm, a constituent of the cell wall of the bacteria in a tuberculoma . It appears that it may be possible to finger print the biochemicals of the cell wall of M . tuberculosis in a tuberculous granuloma and thus may help in detection and diagnosis of such lesions.

Infect Immun, 1996 Dec, 64(12), 5274 - 83
Identification and comparison of macrophage-induced proteins and proteins induced under various stress conditions in Brucella abortus; Rafie-Kolpin M et al.; Brucella abortus is a facultative intracellular pathogen of cattle and humans that is capable of survival inside macrophages . In order to understand how B . abortus copes with the conditions during intracellular growth in macrophages, the protein synthesis pattern of the bacteria grown inside bovine macrophages has been compared with that of bacteria grown in the cell culture medium by two-dimensional polyacrylamide gel electrophoresis . Approximately 24 new proteins that are not detected in the bacteria grown in the cell culture medium have been induced during intracellular growth in macrophages . In contrast, approximately 50 proteins that were expressed during growth in cell culture medium were completely repressed during intracellular growth . The level of expression of 19 proteins increases while that of 54 proteins decreases during intracellular growth . To understand these results, the protein synthesis pattern of B . abortus during intracellular growth was compared with those during other stress conditions . Under each stress condition studied, several new proteins were induced that were not present during regular growth conditions . Comparison of the protein synthesis pattern of B . abortus during intracellular growth with those obtained under various stress conditions has indicated that the response to intracellular growth was not just a simple sum of stress conditions studied so far.

Infect Immun, 1996 Dec, 64(12), 5151 - 60
Effector mechanisms responsible for gamma interferon-mediated host resistance to Legionella pneumophila lung infection: the role of endogenous nitric oxide differs in susceptible and resistant murine hosts; Heath L et al.; To facilitate identification of the effector mechanism(s) responsible for gamma interferon (IFN-gamma)-mediated host resistance to Legionella pneumophila, a murine model of legionellosis in BALB/c mice with a targeted disruption in the IFN-gamma gene (gamma knockout {GKO} mice) was developed . Immunocompetent BALB/c mice and GKO mice were inoculated intratracheally with virulent L . pneumophila (10(6) bacteria per mouse), and bacterial clearance and the pulmonary inflammatory response were assessed . L . pneumophila did not replicate in, and was rapidly cleared from, the lungs of immunocompetent BALB/c mice, demonstrating that immunocompetent BALB/c mice are resistant to replicative L . pneumophila pulmonary infections . In contrast, similarly infected GKO mice developed persistent, replicative intrapulmonary L . pneumophila infections with extrapulmonary dissemination of the bacteria to the spleen . Histopathologic and flow cytometric analysis of L . pneumophila-infected lung tissue demonstrated that while immunocompetent BALB/c mice develop multifocal pneumonitis which resolves, similarly infected GKO mice develop diffuse pneumonitis with persistent neutrophil recruitment into the lung . Intratracheal administration of exogenous IFN-gamma to L . pneumophila-infected GKO mice facilitated intrapulmonary clearance of the bacteria, confirming the pivotal role of IFN-gamma in innate host defenses to L . pneumophila lung infection in this murine host . The potential role of endogenous reactive nitrogen intermediates, including nitric oxide (NO), in IFN-gamma-mediated resistance to L . pneumophila pulmonary infections in immunocompetent BALB/c mice was subsequently assessed . Macrophage inducible nitric oxide synthetase (an enzyme responsible for the production of NO) was induced in alveolar cells from L . pneumophila-infected immunocompetent BALB/c mice (with maximal expression at 48 h postinfection) but was not induced in similarly infected GKO mice . However, administration of the NO synthetase inhibitor N-monomethyl-L-arginine did not significantly inhibit clearance of L . pneumophila from the lung of immunocompetent BALB/c mice (compared with that in similarly infected mice not administered N-monomethyl-L-arginine) . In contrast, we have previously demonstrated that IFN-gamma-induced host resistance to replicative L . pneumophila lung infections in a susceptible murine host (A/J mice) is mediated, in part, by endogenous NO . Taken together, these studies identify a differing role of endogenous NO in IFN-gamma-mediated resistance to L . pneumophila pulmonary infection in susceptible and resistant murine hosts.

Infect Immun, 1996 Dec, 64(12), 5098 - 105
Characterization of the fim2 and fim3 fimbrial subunit genes of Bordetella bronchiseptica: roles of Fim2 and Fim3 fimbriae and flagella in adhesion; Savelkoul PH et al.; With DNA probes derived from the fimbrial subunit genes fim2 and fim3 of Bordetella pertussis, two homologous subunit genes of Bordetella bronchiseptica were identified and cloned . The nucleotide sequences of these genes were determined . Comparison of these nucleotide sequences with the B . pertussis fimbrial fim2 and fim3 subunit genes showed a pronounced homology . Therefore, the B . bronchiseptica genes were also designated fim2 and fim3 . Expression of the two B . bronchiseptica genes was demonstrated by Western blotting (immunoblotting) with polyclonal antiserum directed against the Fim2 and Fim3 fimbrial subunit proteins of B . pertussis and by enzyme-linked immunosorbent assay with monoclonal antibodies . After growth of B . bronchiseptica in the presence of MgSO4, no expression of both fimbrial subunit genes was observed . While no fimbriae were expressed, expression of flagella was observed under these circumstances . A longer C-stretch (up to 19 cytosine residues) than the one in front of the fim2 and fim3 genes of B . pertussis is present in front of the B . bronchiseptica fimbrial genes . In adherence experiments, fimbriated (Bvg+) as well as flagellated (Bvg-) B . bronchiseptica bacteria were able to adhere to HeLa cells, whereas nonflagellated B . pertussis did not . This suggests that fimbriae as well as other factors (possibly flagella) contribute to adherence of B . bronchiseptica to eukaryotic cells.

Nat Genet, 1996 Dec, 14(4), 421 - 9
A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants; Leister D et al.; Plant genes for pathogen resistance can be used to engineer disease resistant crops . Oligonucleotides were designed from sequence motifs conserved between resistance genes of tobacco and Arabidopsis thaliana and used as PCR primers in potato DNA . Amplification products were obtained that were homologous to known resistance genes and linked without recombination with the nematode resistance locus Gro1 and the Phytophthora infestans resistance locus R7 of potato . Map positions of PCR-derived potato gene fragments were also correlated with resistance loci of the related tomato and tobacco genomes . Our results indicate that plant resistance genes that are effective against nematodes, fungi, viruses and bacteria may be isolated based on common sequence motifs and PCR methodology.

J Immunol, 1996 Dec 1, 157(11), 5002 - 8
Humoral immunity and regulation of intrapulmonary growth of Legionella pneumophila in the immunocompetent host; Brieland JK et al.; The potential role of humoral immunity in regulating intrapulmonary growth of Legionella pneumophila in the immunocompetent host was investigated using a murine model of Legionnaires' disease . Intratracheal inoculation of A/J mice with a virulent strain of L . pneumophila (10(6) bacteria per mouse) resulted in the recruitment of B lymphocytes into the lung and the development of anti-L . pneumophila Ab . Opsonization of L . pneumophila in vitro with anti-L . pneumophila-specific mAb resulted in a significant decrease in intrapulmonary growth of the bacteria at 24 to 72 h postinfection . Transmission electron microscopic analysis of lung tissue from L . pneumophila- infected mice demonstrated that while there was no significant difference between phagocytosis of the unopsonized and opsonized L . pneumophila by alveolar macrophages at 24 h postinfection, phagocytosis of opsonized bacteria by alveolar mononuclear phagocytic cells was significantly enhanced at 48 h postinfection . Depletion of A/J mice of complement before intratracheal inoculation of opsonized L . pneumophila (10(6) bacteria per mouse) did not significantly alter intrapulmonary growth of L . pneumophila . These results suggest that anti-L . pneumophila Ab, produced during replicative L . pneumophila lung infections, may regulate intrapulmonary growth of L . pneumophila in the immunocompetent host by decreasing the viability of extracellular L . pneumophila and by enhancing phagocytosis of the bacteria by alveolar mononuclear phagocytic cells by a complement-independent mechanism.

J Clin Microbiol, 1996 Dec, 34(12), 3165 - 70
Prevalence and varieties of Helicobacter species in dogs from random sources and pet dogs: animal and public health implications; Eaton KA et al.; Gastric bacteria of a variety of ultrastructural morphologies have been identified in or isolated from domestic carnivores, but their prevalence in different populations of animals and their clinical significance are still unknown . The purposes of this study were (i) to evaluate the prevalence and morphologic types of gastric bacterial in three different populations of dogs; (ii) to determine which of the organisms were culturable, and if the cultured organisms were morphologically similar to the organisms seen in situ; (iii) to identify the isolated organisms; and (iv) to determine if gastric bacteria were associated with gastritis . Three groups of dogs were examined: healthy laboratory dogs, healthy dogs from an animal shelter, and pet dogs with various nongastric illnesses . Of these, 100% of laboratory and shelter dogs and 67% of pet dogs were colonized by large, tightly coiled gastric spiral bacteria morphologically similar to Gastrospirillum hominis or Helicobacter felis (referred to as gastrospirilla) . Regardless of the presence or density of gastric bacteria, all of the dogs in the study except one had mild to moderate gastritis . Helicobacter spp . were isolated from only 6 of 39 stomachs cultured, and only three of the organisms isolated were morphologically similar to the bacteria seen in situ . Five helicobacters were identified by 16S rDNA (genes coding for rRNA) sequence analysis . Three were strains of H . felis, one was H . bilis, and one was a novel helicobacter morphologically similar to "Flexispira rappini." Gastrospirilla are almost universal in the stomachs of domestic dogs, and in most infected dogs, they do not appear to be associated with clinical signs or histologic lesions compared with uninfected dogs . Nongastrospirillum helicobacters are rare in dogs and are not histologically detectable . Helicobacter pylori was not isolated from domestic dogs.

Cell Tissue Res, 1996 Dec, 286(3), 493 - 505
Type XV collagen exhibits a widespread distribution in human tissues but a distinct localization in basement membrane zones; Myers JC et al.; The collagen family of proteins consists of 19 types encoded by 33 genes . One of the more recently discovered collagens is the alpha1 chain of type XV . Type XV collagen is comprised of a 577-amino-acid, highly interrupted, triple-helical region that is flanked by amino and carboxy noncollagenous domains of 555 and 256 residues, respectively . To address questions of where this collagen is localized and what its function may entail, we produced a bacteria-expressed recombinant protein representing the first half of the type XV collagen carboxy-terminal domain in order to generate highly specific polyclonal antisera . Immunoscreening of an expression library with the affinity-purified antibody revealed three clones coding for part of the type XV triple-helical region and the entire noncollagenous carboxy-terminus . Western blot analysis of human tissue homogenates identified a 116-kDa collagenase-sensitive protein and a 27-kDa collagenase-resistant fragment, whose electrophoretic mobilities were unchanged in the presence and absence of reductant . Northern blot hybridization to human tissue RNAs indicated that type XV has a prevalent and widespread distribution . To determine the precise localization of type XV collagen, immunohistochemical analyses at the light- and electron-microscopic levels were performed . Type XV exhibited a surprisingly restricted and uniform presence in many human tissues as evidenced by a strong association with vascular, neuronal, mesenchymal, and some epithelial basement membrane zones . These data suggest that type XV collagen may function in some manner to adhere basement membrane to the underlying connective tissue stroma.

Ned Tijdschr Geneeskd, 1996 Nov 30, 140(48), 2406 - 10
{Causative role of infections with viruses or atypical pathogens in acute exacerbations of chronic bronchitis less frequent than expected}; Roessingh PH et al.; OBJECTIVE: To determine the prevalence of viruses and atypical pathogens as causes of acute severe exacerbations of chronic bronchitis . DESIGN: Retrospective . SETTING: University Hospital Utrecht, the Netherlands . METHODS: In 4 studies with 305 patients with severe type-1 exacerbations (increase of dyspnoea, of sputum volume and of sputum purulence) serological tests for viral and atypical pathogens were performed . RESULTS: Positive serology for viral infections was seen in 18 (5.9%) patients and for the atypical pathogens in 12 (3.9%) patients . CONCLUSIONS: Since viruses and atypical pathogens are the cause of the exacerbation in only few cases, clinical signs can be used in decision-making whether or not to prescribe antibiotics in acute exacerbations of chronic bronchitis.

Gene, 1996 Nov 28, 181(1-2), 19 - 27
Differential expression of two sporulation specific sigma factors of Streptomyces aureofaciens correlates with the developmental stage; Kormanec J et al.; In previous experiments, the Streptomyces aureofaciens (Sa) rpoZ, and sigF genes have been shown to encode putative sigma factors essential in differentiation . In an attempt to investigate expression of these genes during differentiation, we have performed S1 nuclease mapping using RNA prepared from Sa in various developmental stages . Two putative promoters were identified upstream of the rpoZ coding region . The promoters significantly differed in their strength, and were active in distinct developmental stages; the weaker, rpoZ-P1, was active only in substrate mycelium, and the stronger, rpoZ-P2, was induced at the beginning of aerial mycelium formation . Transcriptional analysis of sigF revealed two apparent transcription start points, both being detectable only in the late phase of aerial mycelium formation . No sigF transcription was detected in an rpoZ-disrupted Sa strain . Promoter-bearing DNA fragments from rpoZ and sigF were inserted into several promoter-probe vectors, to give expression patterns consistent with the results of direct RNA analysis . The results implicate temporally different expression of rpoZ and sigF during the differentiation of Sa, and direct or indirect dependence of sigF expression on the rpoZ-encoded putative sigma factor, thus indicating a cascade of sigma factors in Streptomyces development.

Biochem Biophys Res Commun, 1996 Nov 21, 228(3), 695 - 703
Low temperature enhancement of reporter genes expression directed by human immunodeficiency virus type 1 long terminal repeat; Chevrier-Miller M et al.; Bacteria and eukaryotic cells respond to cold stress by inducing and enhancing the synthesis of specific arrays of proteins . We describe here cold-induced enhancement of expression for two reporter genes; luciferase and beta-galactosidase, both under the control of HIV-1 LTR sequences, observed in mouse fibroblasts and human HeLa cells respectively . Increased expression of luciferase in fibroblasts when shifted to 25 degrees C was detectable at 30 degrees C but was not observed following cold shock at 4 degrees C . To sustain the cold-induced effect, cells had to be kept at subphysiological temperature . The observed enhancement of luciferase activity did not result from a particular site of integration of the reporter gene and was evident whether cold-stressed cells were stationary or growing . Cold-induced expression of luciferase was evidenced at the protein level, enzymatic activity and RNA level, furthermore, active transcription and translation were required for overexpression . The cold effect which has been generalized with the reporter gene beta-galactosidase appears to be a process involving, at least in part, the HIV-1 LTR sequences and might correspond to an increase in the half-life of mRNA . The cold-dependent enhanced expression of luciferase and beta-galactosidase reported here, together with data describing the activation of HIV-1 LTR by hyperthermia, point out the particular temperature sensitivity of these regulatory sequences . This potential thermal modulation may be useful in the comprehension of regulatory processes in latency and reactivation of viral expression during HIV-1 infection.

Nature, 1996 Nov 21, 384(6606), 285 - 8
Structural basis for the binding of a globular antifreeze protein to ice; Jia Z et al.; Antifreeze proteins (AFPs) have the unique ability to adsorb to ice and inhibit its growth . Many organisms ranging from fish to bacteria use AFPs to retard freezing or lessen the damage incurred upon freezing and thawing . The ice-binding mechanism of the long linear alpha-helical type I AFPs has been attributed to their regularly spaced polar residues matching the ice lattice along a pyramidal plane . In contrast, it is not known how globular antifreeze proteins such as type III AFP that lack repeating ice-binding residues bind to ice . Here we report the 1.25 A crystal structure of recombinant type III AFP (QAE isoform) from eel pout (Macrozoarces americanus), which reveals a remarkably flat amphipathic ice-binding site where five hydrogen-bonding atoms match two ranks of oxygens on the {1010} ice prism plane in the <0001> direction, giving high ice-binding affinity and specificity . This binding site, substantiated by the structures and properties of several ice-binding site mutants, suggests that the AFP occupies a niche in the ice surface in which it covers the basal plane while binding to the prism face.

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