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J Bacteriol, 2003 Jan, 185(2), 461 - 5
Bioenergetic properties of the thermoalkaliphilic Bacillus sp . strain TA2.A1; Olsson K et al.; The thermoalkaliphilic Bacillus sp . strain TA2.A1 was able to grow in pH-controlled batch culture containing a nonfermentable growth substrate from pH 7.5 to 10.0 with no significant change in its specific growth rate, demonstrating that this bacterium is a facultative alkaliphile . Growth at pH 10.0 was sensitive to the protonophore carbonyl cyanide m-chlorophenylhydrazone, suggesting that a proton motive force (Deltap) generated via aerobic respiration was an obligate requirement for growth of strain TA2.A1 . Strain TA2.A1 exhibited intracellular pH homeostasis as the external pH increased from 7.5 to 10.0; however, the maximum DeltapH generated over this pH range was only 1.1 units at an external pH of 9.5 . The membrane potential (Deltapsi) was maintained between -114 mV and -150 mV, and little significant change was observed over the pH range for growth . In contrast, the Deltap declined from -164 mV at pH 7.5 to approximately -78 mV at pH 10.0 . An inwardly directed sodium motive force (DeltapNa(+)) of -100 mV at pH 10.0 indicated that cellular processes (i.e., solute transport) dependent on a sodium gradient would not be affected by the adverse Deltap . The phosphorylation potential of strain TA2.A1 was maintained between -300 mV and -418 mV, and the calculated H(+)/ATP stoichiometry of the ATP synthase increased from 2.0 at pH 7.5 to 5.7 at pH 10.0 . Based on these data, vigorous growth of strain TA2.A1 correlated well with the DeltapNa(+), phosphorylation potential, and the ATP/ADP ratio, but not with Deltap . This communication represents the first report on the bioenergetics of an extremely thermoalkaliphilic aerobic bacterium.

Int J Syst Evol Microbiol, 2002 Nov, 52(Pt 6), 2127 - 33
Two novel psychrotolerant species, Bacillus psychrotolerans sp . nov . and Bacillus psychrodurans sp . nov., which contain ornithine in their cell walls; Abd El-Rahman HA et al.; Eleven psychrotolerant Bacillus strains with ornithine as diamino acid in position 3 of the peptide side chain of the cell wall and a G+C range of 35.7-38.4 mol% were characterized taxonomically . DNA-DNA hybridization studies confirmed previously physiologically established groups . High DNA-binding values (> 70%) were found within groups I A (consisting of the type strain of Bacillus insolitus DSM 5(T) and Bacillus insolitus DSM 2272), I B (consisting of isolates 3H1(T0, 71H1, 84E1, 87H2 and 4H2) and I C (consisting of isolates 68E39T), 61E1, 4E3 and 67E1) . Low DNA-binding values (< 60%) were revealed between the three groups . Consequently, strains of groups I B and I C were considered as being representatives of new psychrotolerant species . For group I B strains the name Bacillus psychrotolerans sp . nov . is proposed with the type strain 3H1(T) (= DSM 11706(T) = NCIMB 13838(T)) and for group I C strains the name Bacillus psychrodurans sp . nov . is proposed with the type strain 68E3(T) (= DSM 11713(T) = NCIMB 13837(T)).

Int J Syst Evol Microbiol, 2002 Nov, 52(Pt 6), 1985 - 9
Bacillus luciferensis sp . nov., from volcanic soil on Candlemas Island, South Sandwich archipelago; Logan NA et al.; Aerobic, endospore-forming bacteria were found in soil taken from an active fumarole on Lucifer Hill, Candlemas Island, South Sandwich archipelago . Amplified rDNA restriction analysis, SDS-PAGE, repetitive element primed-PCR (rep-PCR) and routine phenotypic tests suggested that six of the isolates represent a novel taxon, and 16S rDNA sequence comparisons support the proposal of a novel species, Bacillus luciferensis sp . nov., the type strain of which is strain LMG 18422(T) (= CIP 107105(T)).

J Infect Dis, 2003 Jan 1, 187(1), 117 - 23 Epub 2002 Dec 02.
Deletion of RD1 from Mycobacterium tuberculosis mimics bacille Calmette-Guérin attenuation; Lewis KN et al.; The tuberculosis (TB) vaccine bacille Calmette-Guerin (BCG) is a live attenuated organism, but the mutation responsible for its attenuation has never been defined . Recent genetic studies identified a single DNA region of difference, RD1, which is absent in all BCG strains and present in all Mycobacterium tuberculosis (MTB) strains . The 9 open-reading frames predicted within this 9.5-kb region are of unknown function, although they include the TB-specific immunodominant antigens ESAT-6 and CFP-10 . In this study, RD1 was deleted from MTB strain H37Rv, and virulence of H37Rv:DeltaRD1 was assessed after infections of the human macrophage-like cell line THP-1, human peripheral blood monocyte-derived macrophages, and C57BL/6 mice . In each of these systems, the H37Rv:DeltaRD1 strain was strikingly less virulent than MTB and was very similar to BCG controls . Therefore, it was concluded that genes within or controlled by RD1 are essential for MTB virulence and that loss of RD1 was important in BCG attenuation.

J Mol Biol, 2003 Jan 24, 325(4), 677 - 95
Lysyl-tRNA synthetase from Bacillus stearothermophilus: the Trp314 residue is shielded in a non-polar environment and is responsible for the fluorescence changes observed in the amino acid activation reaction; Takita T et al.; Three Trp variants of lysyl-tRNA synthetase from Bacillus stearothermophilus, in which either one or both of the two Trp residues within the enzyme (Trp314 and Trp332) were substituted by a Phe residue, were produced by site-directed mutagenesis without appreciable loss of catalytic activity . The following two phenomena were observed with W332F and with the wild-type enzyme, but not with W314F: (1) the addition of L-lysine alone decreased the protein fluorescence of the enzyme, but the addition of ATP alone did not; (2) the subsequent addition of ATP after the addition of excess L-lysine restored the fluorescence to its original level . Fluorometry under various conditions and UV-absorption spectroscopy revealed that Trp314, which was about 20A away from the lysine binding site and was shielded in a non-polar environment, was solely responsible for the fluorescence changes of the enzyme in the L-lysine activation reaction . Furthermore, the microenvironmental conditions around the residue were made more polar upon the binding of L-lysine, though its contact with the solvent was still restricted . It was suggested that Trp314 was located in a less polar environment than was Trp332, after comparison of the wavelengths at the peaks of fluorescence emission and of the relative fluorescence quantum yields . Trp332 was thought, based on the fluorescence quenching by some perturbants and the chemical modification with N-bromosuccinimide, to be on the surface of the enzyme, whereas Trp314 was buried inside . The UV absorption difference spectra induced by the L-lysine binding indicated that the state of Trp314, including its electrostatic environment, changed during the process, but Trp332 did not change . The increased fluorescence from Trp314 at acidic pH compared with that at neutral pH suggests that carboxylate(s) are in close proximity to the Trp314 residue.

Int J Food Microbiol, 2003 Jan 26, 82(1), 71 - 9
Influence of pH and temperature on growth of Bacillus cereus in vegetable substrates; Valero M et al.; Bacillus cereus is a food-borne pathogen which most often contaminates foods of plant origin . Spores of psychrotrophic strains have the ability to germinate and grow at refrigeration temperatures in different vegetable substrates, such as carrot broth, zucchini broth, and cooked carrot puree . In some circumstances, factors such as pH, heat treatment, and storage temperature play a fundamental role in controlling the growth of these psychrotrophic strains and in extending the shelf life of refrigerated, minimally processed vegetable-based products in relation to pathogenic spore-forming bacteria . The combination of mild acidification (pH 5.0) and refrigeration (</=8 degrees C) inhibited B . cereus growth for at least 60 days in vegetable substrates similar to those mentioned above . This protection was maintained even when the temperature to which the food was exposed reached 12 degrees C . Psychrotrophic strains of B . cereus were inhibited in carrot broth by heating at 90 degrees C for 7.5 min, if the broth was refrigerated at a temperature of 8 degrees C or lower . If the vegetable product was exposed to temperatures of mild abuse (12 degrees C), it was necessary to implement a more drastic heat treatment (90 degrees C for 30 min).

Indian J Pediatr, 2002 Nov, 69(11), 1003 - 5
Cutaneous bacillary angiomatosis; Asharaf M et al.; Bacillary angiomatosis is characterized by unique vascular lesions caused by infection with a small Gram staining bacillus of the genus Bartonella . It usually occurs in immunocompromised persons but can also occur in immunocompetent persons . We report a case of cutaneous bacillary angiomatosis in a 5-year-old immunocompetent child . He had infected lesions on the lips, after an injury, which was followed by lesions over the knees, buttocks, near the ankles and the elbows . Diagnosis was proved on histology . The lesions cleared after administration of erythromycin for 3 months . It is well to be aware of this condition in the context of increasing prevalence of AIDS.

Expert Rev Anticancer Ther, 2002 Dec, 2(6), 667 - 70
Prognostic value of p53 overexpression in bladder tumors treated with Bacillus Calmette-Guerin; Peyromaure M et al.; Bacillus Calmette-Guerin intravesical therapy is the standard treatment of superficial bladder tumors at high risk of recurrence and progression to muscle-invasion disease . To date, there is no well established predictive factor of response to Bacillus Calmette-Guerin intravesical therapy . The prognostic value of p53 overexpression in bladder tumors is controversial . Most investigators have found no correlation between p53 status assessed before Bacillus Calmette-Guerin intravesical therapy and patient outcome . On the other hand, it is acknowledged that the persistence of p53 overexpression after Bacillus Calmette-Guerin intravesical therapy is predictive of progression in patients treated for carcinoma in situ . Since conflicting data have been reported, further evaluation of the impact of p53 overexpression in patients treated with Bacillus Calmette-Guerin intravesical therapy for bladder carcinoma is required.

J Agric Food Chem, 2003 Jan 1, 51(1), 100 - 5
Enterotoxigenicity and cytotoxicity of Bacillus thuringiensis strains and development of a process for Cry1Ac production; Yang CY et al.; Bacillus thuringiensis is indistinguishable from Bacillus cereus except for the production of insecticidal crystal proteins (ICPs) . B . thuringiensis strains may show enterotoxin profiles and toxin levels similar to those of B . cereus strains isolated from food-poisoning cases . It is important for the food industry and farmers to consider that with the application of B . thuringiensis strains to crops, their spores may be introduced into the human food chain . In this study, 59 B . thuringiensis strains were assayed for their hemolysin BL (HBL) using a BCET-RPLA kit and their cytotoxicity to Chinese hamster ovary (CHO) cells . The enterotoxin titer was as high as that of B . cereus diarrheal-type strain ATCC 49064 . In an attempt to obtain a food safety strain for bioinsecticide use, in this study, a 3.5-kb cry1Ac DNA fragment was amplified with PCR from the total DNA of B . thuringiensis subsp . kurstaki CCRC 11502 and cloned into the Bacillus expression vector pHY300PLK . The alpha-amylase promoter, amyE, was then introduced into the promoter region and, afterward, the recombinant plasmid pHYe1Ac35 was introduced into a non-enterotoxigenic and non-cytotoxic B . thuringiensis subsp . kurstaki Tt14 strain . The transformant, without any detectable enterotoxigenicity or cytotoxicity, produced Cry1Ac toxin properly, and its insecticidal activity against Trichoplusia ni larvae was found to be satisfactory.

J Gen Appl Microbiol, 2002 Oct, 48(5), 243 - 50
Application of the hypervariable region of the 16S rDNA sequence as an index for the rapid identification of species in the genus Alicyclobacillus; Goto K et al.; A comparison of the 16S rRNA gene (rDNA) sequences of seven type strains belonging to different Alicyclobacillus species (i.e., five validated species, one proposed species and one genomic species) suggested that the 5' end hypervariable region (259-273 bases in length) of 16S rDNA was specific for the respective type strains . Further phylogenetic analysis based on DNA sequences of the hypervariable region using 24 Alicyclobacillus strains revealed that the strains could be categorized into five species and the A . acidocaldarius-Alicyclobacillus genomic species 1 group . The hypervariable region was highly conserved among the five species: A . acidiphilus, A . acidoterrestris, A . cycloheptanicus, A . herbarius, and A . hesperidum . The strains in the A . acidocaldarius-Alicyclobacillus genomic species 1 group were subdivided into two clusters (Clusters I and II) based on DNA sequences in the hypervariable region . On the basis of phenotypic characteristics, chemotaxonomic and phylogenetic analyses, and DNA-DNA hybridization data, strains in Cluster I were grouped as Alicyclobacillus genomic species 1 and strains in Cluster II were re-identified as A . acidocaldarius, thereby demonstrating that the hypervariable regions were also highly conserved within these two species . These results suggest that as is the case with Bacillus, the hypervariable region is significantly species-specific in the genus Alicyclobacillus to distinguish Alicyclobacillus species by DNA sequence comparison of the hypervariable region.

J Gen Appl Microbiol, 1999 Aug, 45(4), 149 - 153
Production of 2-phenylethylamine by decarboxylation of L-phenylalanine in alkaliphilic Bacillus cohnii; Hamana K et al.; Cellular polyamine fraction of alkaliphilic Bacillus species was analyzed by HPLC . 2-Phenylethylamine was found selectively and ubiquitously in the five strains belonging to Bacillus cohnii within 27 alkaliphilic Bacillus strains . A large amount of this aromatic amine was produced by the decarboxylation of L-phenylalanine in the bacteria and secreted into the culture medium . The production of 2-phenylethylamine may serve for the chemotaxonomy of alkaliphilic Bacillus.

J Gen Appl Microbiol, 1999 Dec, 45(6), 283 - 287
In vitro degradation of keratin by two species of Bacillus; Lal S et al.; The ability of two species of Bacillus to degrade child's scalp hair, cow horn, cow hooves, and human nails in vitro under static conditions was studied by the determination of soluble sulphhydryl compounds as cysteine, disulphides as cystine, and release of extracellular keratinase along with changes in alkalinity of the culture filtrate . Child's scalp hair was found to be the most favored keratin substrate for Bacillus spp.

J Gen Appl Microbiol, 1997 Dec, 43(6), 341 - 347
Cloning of a chitinase gene into Bacillus thuringiensis subsp . aizawai for enhanced insecticidal activity; Tantimavanich S et al.; Chitinase from a high producing strain (TP-1) of Bacillus licheniformis was used with B . thuringiensis subsp . aizawai (B.t.a.) in a combined larvicidal assay against the pest, Spodoptera exigua . With 10 mU of this chitinase, the LD(50) of B.t.a . was reduced by 7.6, 13.8 and 15 times on days 3, 5 and 7, respectively when compared to use of B.t.a . alone . In addition, a combination of chitinase (10 mU) and B.t.a . at a sub-lethal dose retarded growth and development of S . exigua . In preparation for transformation of B.t.a., the TP-1 chitinase gene was cloned in E . coli DH5alpha and sequenced to reveal a single open reading frame of 1,815 bp . This open reading frame encoded for a protein of 604 amino acids and a characteristic signal peptide sequence of 35 amino acids . The gene was subsequently introduced into B.t.a . where it was expressed constitutively . The transformed strain showed slightly improved activity against S . exigua when compared to the non-transformed strain . This was probably due to the low chitinase activity (15 mU/ml) of the transformant, which might be improved by further gene manipulation to overexpress enzyme production.

J Gen Appl Microbiol, 1998 Apr, 44(2), 133 - 138
Seasonal distribution and characterization of Bacillus thuringiensis isolated from sericultural environments in Korea; Kim HS et al.; To isolate naturally occurring novel Bacillus thuringiensis strains, we investigated the distribution and characteristics of B . thuringiensis from samples of sericultural farms in various regions of Korea in the spring and fall . Fifty-four B . thuringiensis strains out of 164 samples and 34 B . thuringiensis strains out of 135 samples were isolated in the spring and fall, respectively . Seventy percent of the isolates in the spring and 15% in the fall were toxic to lepidopteran larvae . Dipteran-active isolates were rare (7% in spring and 3% in fall isolation) . Particularly, B . thuringiensis isolates, which are toxic to both Lepidoptera and Diptera, were widely distributed (19% in spring and 62% in fall isolation) . Non-toxic isolates were also found (4% in spring and 20% in fall isolation) . B . thuringiensis isolates in the sericultural farms represented 11 H serotypes; they were principally B . thuringiensis subsp . aizawai in the spring and kurstaki in the fall . B . thuringiensis isolates of serotypes 1, 3a, 3a3b, 4a4c, 6, 7 and 12 were toxic to Lepidoptera . Seventy isolates produced typical rhomboidal inclusions, and the remainder produced parasporal inclusions with various morphologies . PCR analysis using cryI gene type-specific primers showed that cryIAa and cryIC genes are frequently found in the spring and cryIAa gene is a predominant type in the fall . Toxicity, H serotype and the cryI gene contents of B . thuringiensis isolated from sericultural farms showed that distribution varied depending on the season.

Neurology, 2002 Dec 24, 59(12), 1837 - 43
Immunization and MS: a summary of published evidence and recommendations; Rutschmann OT et al.; OBJECTIVE: To review the risk of MS exacerbations after infectious episodes potentially preventable by vaccination, and the risks and benefits of immunizing patients with MS . METHODS: The authors searched MEDLINE (1966 to January 2001; U.S . National Library of Medicine, Bethesda, MD), HealthSTAR, and Cumulative Index to Nursing and Allied Health Literature (CINAHL) database (Cinahl Information Systems, Glendale, CA) for English-language articles . Each study was summarized and rated for quality of evidence . Then feasible data were pooled and analyzed in meta-analysis . RESULTS: The risk of contracting common infectious diseases in patients with MS is not well established . There is strong evidence for an increased risk of MS exacerbations during weeks around an infectious episode . There is strong evidence against an increased risk of MS exacerbation after influenza immunization . There is no evidence that hepatitis B, varicella, tetanus, or Bacille Calmette-Guerin vaccines increase the risk of MS exacerbations . Insufficient evidence was found for other vaccines . CONCLUSIONS: Evidence supports 1) strategies to minimize the risk of acquiring infectious diseases that may trigger exacerbations of MS; and 2) the safety of using influenza, hepatitis B, varicella, tetanus, and Bacille Calmette-Guerin (BCG) vaccines in patients with MS.

Antimicrob Agents Chemother, 2003 Jan, 47(1), 383 - 6
gyrA mutations in ciprofloxacin-resistant Bartonella bacilliformis strains obtained in vitro; Minnick MF et al.; We isolated and characterized mutants of Bartonella bacilliformis that are resistant to the fluoroquinolone antibiotic ciprofloxacin, which targets the A subunit of DNA gyrase . Mutants had single point mutations in the gyrA gene that changed either Asp-90 to Gly or Asp-95 to Asn and had 3- or 16-fold higher resistance, respectively, to ciprofloxacin than did wild-type B . bacilliformis . Asp-95 is homologous to Asp-87 of Escherichia coli GyrA and is a common residue mutated in fluoroquinolone-resistant strains of other bacteria . This is the first report of a mutation at an Asp-90 homologue, which corresponds to Asp-82 in E . coli GyrA.

Clin Immunol, 2002 Dec, 105(3), 326 - 31
Oral recombinant Mycobacterium bovis bacillus Calmette-Guérin expressing HIV-1 antigens as a freeze-dried vaccine induces long-term, HIV-specific mucosal and systemic immunity; Kawahara M et al.; Induction of HIV-1-specific immune responses was evaluated using a recombinant BCG (rBCG) vector-based vaccine expressing HIV-1 Env V3 peptide (rBCG-pSOV3J1) . rBCG-pSOV3J1 was manufactured as a freeze-dried preparation based on good laboratory practice guidelines . Guinea pigs were immunized with the freeze-dried rBCG vaccine by oral administration to test the effectiveness of what is generally considered the most convenient and practical route for vaccination . While delayed-type hypersensitivity (DTH) skin reactions to purified protein derivative were not detected in any of the animals receiving oral rBCG-pSOV3J1, HIV-1 V3J1 antigen-specific DTH responses were detected in all of the immunized guinea pigs 1.5 years after immunization . In addition, significant proliferative responses against HIV-1 V3J1 antigen were measured in peripheral blood mononuclear cells and splenocytes from all animals receiving oral rBCG . Interestingly, intestinal intraepithelial lymphocytes from the animals also exhibited high levels of proliferative activity against HIV-1 V3J1 antigen . These results suggest that oral vaccination of guinea pigs with freeze-dried rBCG-pSOV3J1 induces high levels of functional T cells specific for HIV-1 antigens in both mucosal and systemic compartments and suggest that this approach has potential for use as a vaccine against HIV-1.

Clin Immunol, 2002 Dec, 105(3), 296 - 303
Dendritic cells are decreased in blood and accumulated in granuloma in tuberculosis; Uehira K et al.; Immunity against tuberculosis consists of innate and adaptive immune responses . In this study, we investigated the dynamics of dendritic cells (DC), which are known to elicit a variety of immune responses, in patients with tuberculosis . CD11c(+) peripheral blood DC were decreased in patients with tuberculosis . Immunohistochemical analyses demonstrated that a number of fascin(+), CD11c(+), HLA-DR(+) DC were infiltrating the lymphocyte areas of the tuberculous granulomas (tubercles) . Immunohistochemical analyses also demonstrated that interferon-gamma-producing Th1 cells were increased in the tubercles of the patients, indicating the presence of Th1 polarization at least in the context of inflammatory tissues . In vitro coculture of autologous naive T cells with CD11c(+) or CD11c(-) DC pretreated with Bacillus Calmette Guerin augmented the production of Th1 cells . These findings suggested that the trafficking of DC from the peripheral blood into the tubercles causes a dominant Th1 balance and thus plays an essential role in the immunity against tuberculosis.

J Immunol, 2003 Jan 1, 170(1), 463 - 9
Characterization of lung gamma delta T cells following intranasal infection with Mycobacterium bovis bacillus Calmette-Guérin; Dieli F et al.; The lungs are considered to have an impaired capacity to contain infection by pathogenic mycobacteria, even in the presence of effective systemic immunity . In an attempt to understand the underlying cellular mechanisms, we characterized the gammadelta T cell population following intranasal infection with Mycobacterium bovis bacillus Calmette-Guerin (BCG) . The peak of gammadelta T cell expansion at 7 days postinfection preceded the 30 day peak of alphabeta T cell expansion and bacterial count . The expanded population of gammadelta T cells in the lungs of BCG-infected mice represents an expansion of the resident Vgamma2 T cell subset as well as an influx of Vgamma1 and of four different Vdelta gene-bearing T cell subsets . The gammadelta T cells in the lungs of BCG-infected mice secreted IFN-gamma following in vitro stimulation with ionomycin and PMA and were cytotoxic against BCG-infected peritoneal macrophages as well as against the uninfected J774 macrophage cell line . The cytotoxicity was selectively blocked by anti-gammadelta TCR mAb and strontium ions, suggesting a granule-exocytosis killing pathway . Depletion of gammadelta T cells by injection of specific mAb had no effect on the subsequent developing CD4 T cell response in the lungs of BCG-infected mice, but significantly reduced cytotoxic activity and IFN-gamma production by lung CD8 T cells . Thus, gammadelta T cells in the lungs might help to control mycobacterial infection in the period between innate and classical adaptive immunity and may also play an important regulatory role in the subsequent onset of alphabeta T lymphocytes.

Infect Immun, 2003 Jan, 71(1), 101 - 8
Oral delivery of Mycobacterium bovis BCG in a lipid formulation induces resistance to pulmonary tuberculosis in mice; Aldwell FE et al.; A lipid-based formulation has been developed for oral delivery of Mycobacterium bovis bacille Calmette-Guerin (BCG) vaccine . The formulated M . bovis BCG was fed to BALB/c mice to test for immune responses and protection against M . bovis infection . The immune responses included antigen-specific cytokine responses, spleen cell proliferation, and lymphocyte-mediated macrophage inhibition of M . bovis . Oral delivery of formulated M . bovis BCG to mice induced strong splenic gamma interferon levels and macrophage inhibition of virulent M . bovis compared with results with nonformulated M . bovis BCG . Formulated oral M . bovis BCG significantly reduced the bacterial burden in the spleen and lungs of mice following aerosol challenge with virulent M . bovis . Our data suggest that oral delivery of formulated M . bovis BCG is an effective means of inducing protective immune responses against tuberculosis . Lipid-based, orally delivered mycobacterial vaccines may be a safe and practical method of controlling tuberculosis in humans and animals.

Biophys J, 2002 Dec, 83(6), 3416 - 24
Bidirectional control of sphingomyelinase activity and surface topography in lipid monolayers; Fanani ML et al.; Lipid lateral organization is increasingly found to modulate membrane-bound enzymes . We followed in real time the reaction course of sphingomyelin (SM) degradation by Bacillus cereus sphingomyelinase (SMase) of lipid monolayers by epifluorescence microscopy . There is evidence that formation of ceramide (Cer), a lipid second messenger, drives structural reorganization of membrane lipids . Our results provide visual evidence that SMase activity initially alters surface topography by inducing phase separation into condensed (Cer-enriched) and expanded (SM-enriched) domains . The Cer-enriched phase grows steadily as the reaction proceeds at a constant rate . The surface topography derived from the SMase-driven reaction was compared with, and found to differ from, that of premixed SM/Cer monolayers of the same lipid composition, indicating that substantial information content is stored depending on the manner in which the surface was generated . The long-range topographic changes feed back on the kinetics of Smase, and the onset of condensed-phase percolation is temporally correlated with a rapid drop of reaction rate . These observations reveal a bidirectional influence and communication between effects taking place at the local molecular level and the supramolecular organization . The results suggest a novel biocatalytic-topographic mechanism in which a surface enzymatic activity can influence the function of amphitropic proteins important for cell function.

J Food Prot, 2002 Dec, 65(12), 1930 - 6
Altered ability of Bacillus cereus spores to grow under unfavorable conditions (presence of nisin, low temperature, acidic pH, presence of NaCl) following heat treatment during sporulation; Faille C et al.; The effect of thermal treatment on the heat resistance of Bacillus cereus spores and their ability to germinate and grow under more or less adverse conditions during sporulation was investigated . Spores produced by sporulating cells subjected to a mild heat treatment (at a temperature 15 degrees C higher than the growth temperature) were more resistant to heat than were spores produced by untreated cells . Spore germination and growth (the lag time, the maximal growth rate, and the occurrence of a decrease in population) may be greatly affected by adverse environmental conditions brought about by the addition of nisin, low temperatures, acidic pHs, and, to a lesser extent, the addition of NaCl . Furthermore, heat treatments applied to sporulating cells or to mature spores induced a modification of the lag time (interaction of both treatments) . Therefore, mild heat treatments applied during sporulation may affect the heat resistance of spores and the ability of these spores to germinate under adverse conditions and may thus increase the risk associated with the presence of spores in lightly processed foods.

J Antimicrob Chemother, 2003 Jan, 51(1), 107 - 12
Emergence of penicillin resistance among Fusobacterium nucleatum populations of commensal oral flora during early childhood; Nyfors S et al.; Penicillin resistance due to beta-lactamase production is surprisingly common among oral bacteria in childhood . Fusobacterium nucleatum, a Gram-negative anaerobic bacillus, is a member of the developing oral commensal flora . As part of the investigation on the emergence of oral bacterial resistance, the aim of the present study was to examine longitudinally the penicillin resistance among salivary F . nucleatum populations as related to age, day care attendance and sibling history, and exposure to antimicrobial agents . Altogether 1492 F . nucleatum isolates from saliva of 44 healthy infants followed at a study clinic at 2, 6, 12, 18 and 24 months of age were tested for beta-lactamase production . Furthermore, the 276 beta-lactamase-positive isolates were examined for their in vitro susceptibility to penicillin G by the NCCLS-approved agar dilution method . Statistical analysis of the associations between penicillin-resistant isolates and infants' age, day care attendance, number of siblings and their ear infections, and exposure to antimicrobial agents was performed by SPSS Windows Version 10 . The prevalence of infants harbouring beta-lactamase-producing F . nucleatum strains increased from 2% to 49% during the follow-up time . In nearly all cases beta-lactamase-producing F . nucleatum isolates were found simultaneously with beta-lactamase-negative isolates . Most beta-lactamase-producing isolates (80%) showed an MIC of > or =8 mg/L . In conclusion, the prevalence of infants harbouring penicillin-resistant F . nucleatum due to beta-lactamase production increased with age and usage of antimicrobial agents during the first year of life.

J Appl Microbiol, 2003, 94(1), 60 - 4
Properties of Bacillus thuringiensis isolated from bank voles; Swiecicka I et al.; AIMS: To assess the properties of B . thuringiensis naturally occurring in the intestines of bank voles . METHODS AND RESULTS: Seventeen Bacillus thuringiensis strains, exhibiting typical growth on selective medium for the B . cereus group and characterized by the ability to produce parasporal crystals, were isolated from bank voles trapped in the Lomza Landscape Park of the Narew River Valley (north-east Poland) . All isolates were characterized by pulsed field gel electrophoresis (PFGE) of chromosomal DNA and SDS polyacrylamide gel electrophoresis (SDS-PAGE) of whole-cell proteins . Six pulsotypes were found with PFGE typing, using SmaI or NotI as restriction enzymes . Significant differences in chromosome size, ranging from 2.4 to 4.2 Mb for the B . thuringiensis strains studied, were noted . Strain heterogeneity in pulsotypes was also reflected by the similarity of whole-cell protein profiles of the strains . Environmental isolates and reference strains grouped at 71% similarity according to SDS-PAGE data and at 84% on the basis of biochemical tests . CONCLUSIONS: B . thuringiensis from intestines of bank voles demonstrated an important level of heterogeneity . The comparison of PFGE profiles and SDS-PAGE of whole-cell protein patterns may be useful to evaluate the relationship between B . thuringiensis isolates . SIGNIFICANCE AND IMPACT OF THE STUDY: The results presented in this paper may help to explain the diversity of B . thuringiensis.

Zhonghua Nan Ke Xue, 2002, 8(4), 266 - 9
{The establishment of testicular fibrosis model in Wistar rats}; Wang T et al.; OBJECTIVES: To establish the testicular fibrosis model in rats . METHODS: Wistar rats were divided into control group(n = 12) and model group(n = 40) randomly . Testicular fibrosis model was built with the classical method of establishing experimental autoimmune orchitis (EAO) combined with injecting Bacille Calmette-Guerin (BCG) into left testis . RESULTS: The incidence rate of EAO and the rate of testicular fibrosis were 100%, 11.1% and 100%, 81.5% at 80, 140 days after the first infection, respectively . CONCLUSIONS: The model of rat testicular fibrosis was established successfully.

Clin Infect Dis, 2003 Jan 1, 36(1), 16 - 23 Epub 2002 Dec 13.
Improved detection of Mycobacterium tuberculosis in Peruvian children by use of a heminested IS6110 polymerase chain reaction assay; Montenegro SH et al.; A novel heminested IS6110 polymerase chain reaction (PCR) assay was evaluated as a tool for diagnosing tuberculosis in 222 children . In an analysis of 392 specimens (gastric aspirates, nasopharyngeal aspirates, and sputum samples), results of PCR were compared with those of 3 culture methods, acid-fast bacillus (AFB) staining, and clinical assessment by the Stegen-Toledo score . The sensitivity of PCR (67%) was comparable to that of the 3-culture method (71%) and was significantly higher than that of Lowenstein-Jensen culture (54%) or AFB stain (42%) for children with highly probable tuberculosis . PCR detection rates for culture-positive specimens were 100% for smear-positive samples and 76.7% for smear-negative samples . The specificity of PCR was 100% in control children . Compared with culture, PCR demonstrated a sensitivity of 90.4%, a positive predictive value of 89%, a specificity of 94%, and a negative predictive value of 95% (kappa=.85) . With clinical assessment as the standard, PCR had a sensitivity of 71%, a positive predictive value of 92%, a specificity of 95%, and a negative predictive value of 79% (kappa=.67) . PCR is a rapid and sensitive method for the early diagnosis of pediatric tuberculosis.

Arch Virol, 2002 Dec, 147(12), 2393 - 404
Sequence analysis of an Australian isolate of sugarcane bacilliform badnavirus; Geijskes RJ et al.; The genome of an Australian isolate of Sugarcane bacilliform virus (SCBV-IM) was cloned, sequenced and analysed . The genome consisted of 7687 nucleotides and contained three open reading frames which were similar in size and organisation to those of other badnaviruses . SCBV-IM was found to be most similar to the SCBV-Morocco isolate with amino acid sequence similarity of 91.4 %, 83.8 % and 85.3 % in the ORF I, II and III coding regions, respectively . Phylogenetic analysis of the SCBV-IM ORF III deduced amino acid sequence showed that SCBV isolates were more closely related to each other than to other badnaviruses . Amplification of SCBV sequences from three different sugarcane varieties revealed considerable variability in the viral populations, both within single infected plants as well as between infected plants, suggesting that the SCBV isolates sequenced to date may not be representative of the range of virus variability.

Gene, 2002 Nov 13, 301(1-2), 13 - 20
Characterization of two regulatory genes of the mercury resistance determinants from TnMERI1 by luciferase-based examination; Huang CC et al.; The broad-spectrum mercury resistance transposon, TnMERI1, of Bacillus megaterium strain MB1, contains three proposed operator/promoter (O/P) transcriptional start sites and two regulatory genes (merR1 and merR2) . A series of luciferase (lux)-based transcriptional fusion plasmids were studied in Escherichia coli to show that both merR1 and merR2 gene products repressed transcription from O/PmerB3, O/PmerR1, and O/PmerR2 under uninduced conditions . Derepression occurred when the merR1 gene was present and Hg(2+) functioned as an inducer . In the presence of organomercurial compounds, basal transcription of merB3 was needed to produce inorganic Hg(2+) as the inducer of expression regulated by MerR1 at O/PmerB3 . The presence of merR2 repressed transcription from all three O/Pmer sites under both non-induced conditions and when inorganic Hg(2+) or organomercurials were added . These results show that MerR1 functions as a repressor in the absence of Hg(2+) and as an activator in the presence of Hg(2+), while MerR2 functions as a repressor.

Zhonghua Jie He He Hu Xi Za Zhi, 2002 Oct, 25(10), 595 - 7
{A study on the use of mycobacterium vaccine in the treatment of tuberculous pleurisy}; Zhou X et al.; OBJECTIVE: To study the effects of immunotherapy with mycobacterium vaccine in patients with tuberculous pleurisy . METHODS: The time for effusion resolution and lymphocyte counts in the effusions were observed in patients with tuberculous pleurisy who received anti-tuberculous therapy and thoracentesis (group A) or anti-tuberculous therapy and thoracentesis plus intramuscularly administered polysaccharide nucleic acid fraction of Bacillus Calmette Guerin (PSN-BCG,) (group S) . In situ end-labeling technique of fragment DNA was used to detect the apoptosis of lymphocytes in the effusions . RESULTS: The average resolution times in group A and group S were 21.2 days and 28.7 days respectively (P < 0.01) . The lymphocyte counts in the two groups after 1 week, 2 week, and 3 week therapy were 1.6 x 10(9)/L (A) and 2.1 x 10(9)/L (S) (P < 0.01), 0.83 x 10(9)/L and 1.52 x 10(9)/L (P < 0.01), 0.55 x 10(9)/L and 1.16 x 10(9)/L (P < 0.01) respectively . The average apoptotic half-time was 94 h (A) and 124 h (S) (P < 0.01), 84 h and 123 h (P < 0.01), 79 h and 120 h (P < 0.01) respectively . CONCLUSIONS: Mycobacterium vaccine was found to prolong lymphocyte activation, inhibit lymphocyte apoptosis, and delay the resolution of pleural effusions . It was found not helpful as an adjunct therapy for tuberculous pleurisy.

Clin Lab Med, 2002 Dec, 22(4), 937 - 62
Laboratory diagnosis of Bartonella infections; Agan BK et al.; Bartonella species are pathogens of emerging and reemerging significance, causing a wide array of clinical syndromes . In North America and Europe, they are increasingly recognized as a cause of culture negative endocarditis, neuroretinitis, and disease among homeless, HIV-infected, and other immunosuppressed individuals . In South America, bartonellosis continues to plague those in endemic regions and poses a significant threat to travelers in these areas . As the clinician is increasingly faced with these illnesses, which may be difficult to diagnose, laboratory techniques to confirm or refute the diagnosis are becoming increasingly important . Culture methods have improved over the past decade demonstrating increased sensitivity, but still require prolonged periods before isolation of the organism . Specimen handling, media selection, and growth conditions all may affect results and must be optimized in order to provide the highest likelihood of recovering the organism . Pure culture of the bacteria not only provides morphologic information, but also provides material for further diagnostic testing . Work with liquid media, which may provide a more rapid means of cultivation has shown some promise and should continue to be pursued . Improved blood culture techniques were a primary factor in the discovery of Bartonella endocarditis and continued improvements will likely demonstrate further clinical insights . Serologic testing for B henselae infections has become the cornerstone of clinical diagnosis, replacing the skin test that was poorly standardized and posed a potential risk to the patient . Immunofluorescence assays have been well characterized and validated in clinical trials, however they are not universally available . Vero cell cocultivated antigens appear to provide higher sensitivity and specificity when compared with agar-derived antigens . IFA assays are inherently difficult to perform, requiring significant expertise to provide reproducible results . On the contrary, enzyme immunoassays offer ease of use and a high level of reproducibility, however ideal antigens for use in the diagnosis of Bartonella infections have not been clearly identified . Continued work to define antigenic targets of the human response to infection and incorporation of these into a widely available EIA will provide a cost-effective tool for the clinician and epidemiologist alike . Due to the close phylogenetic relationship of B henselae and B quintana, differentiation between these species by serologic means may prove difficult . Molecular techniques including PCR offer high sensitivity and specificity, rapid availability of information, and the ability to differentiate Bartonella organisms at the highest level . Results of studies to date are promising and as methods are refined it will be important to conduct clinical studies to define the role of these assays . In disseminated Bartonella infections such as bacillary angiomatosis, peliosis, endocarditis, and urban trench fever, PCR currently offers the ability to establish the diagnosis when other tests may be unrevealing . For CSD, this technique should be used as a confirmatory technique when the diagnosis is unclear by other means . PCR analysis of blood specimens offers a minimally invasive approach to diagnosis, but clinical data are scarce and further studies are needed . As DNA microarrays move into the clinical arena, specific hybridization probes may allow improved identification and differentiation of Bartonellae at the molecular level.

J Med Microbiol, 2003 Jan, 52(Pt 1), 69 - 74
Production by Bacillus pumilus (MSH) of an antifungal compound that is active against Mucoraceae and Aspergillus species: preliminary report; Bottone EJ et al.; A compound produced by Bacillus pumilus (MSH) that inhibits Mucoraceae and Aspergillus species is described . Fungicidal activity was demonstrated by lawn-spotting and by diffusion through 0.45 microm Millipore membranes placed on 5 % sheep-blood agar, nutrient agar, trypticase soy agar and Mueller-Hinton agar, followed by spore inoculation of the bacterium-free underlying agar surface . With either technique, zones of fungal inhibition correlated with the zone of haemolysis produced by B . pumilus (MSH) . The active compound inhibited Mucor and Aspergillus spore germination and aborted elongating hyphae, presumably by inducing a cell-wall lesion . Antifungal activity was stable in agar for a minimum of 8 days, resistant to Pronase degradation, and partially inactivated by chloroform exposure and at pH 5.6 . Its molecular mass was determined by diffusion through dialysis membrane to be 500-3000 Da . Attempts at further isolation of the compound have proven unsuccessful to date.

Biochim Biophys Acta, 2002 Dec 23, 1567(1-2), 113 - 22
Estimation of the radius of the pores formed by the Bacillus thuringiensis Cry1C delta-endotoxin in planar lipid bilayers; Peyronnet O et al.; Pore formation constitutes a key step in the mode of action of Bacillus thuringiensis delta-endotoxins and various activated Cry toxins have been shown to form ionic channels in receptor-free planar lipid bilayers at high concentrations . Multiple conductance levels have been observed with several toxins, suggesting that the channels result from the multimeric assembly of a variable number of toxin molecules . To test this possibility, the size of the channels formed by Cry1C was estimated with the non-electrolyte exclusion technique and polyethylene glycols of various molecular weights . In symmetrical 300 mM KCl solutions, Cry1C induced channel activity with 15 distinct conductance levels ranging from 21 to 246 pS and distributed in two main conductance populations . Both the smallest and largest conductance levels and the mean conductance values of both populations were systematically reduced in the presence of polyethylene glycols with hydrated radii of up to 1.05 nm, indicating that these solutes can penetrate the pores formed by the toxin . Larger polyethylene glycols had little effect on the conductance levels, indicating that they were excluded from the pores . Our results indicate that Cry1C forms clusters composed of a variable number of channels having a similar pore radius of between 1.0 and 1.3 nm and gating synchronously.

Intern Med, 2002 Nov, 41(11), 990 - 2
Pulmonary sequestration associated by Mycobacterium intracellulare infection; Shiota Y et al.; A case of a 29-year-old woman with intralobar pulmonary sequestration infected with Mycobacterium intracellulare is presented . A chest CT scan revealed a density in the posterior segment of the left lower lobe, and an acid-fast bacillus sputum culture yielded Mycobacterium intracellulare . After 3 months of treatment with clarithromycin, streptomycin, rifampicin and ethambutol, the patient underwent partial resection of the left lower lobe . At the 6-month follow-up the patient's clinical status is excellent . A review of the literature revealed only three case reports of pulmonary sequestration associated with Mycobacterium avium-intracellulare complex infection.

J Mol Evol, 2002 Dec, 55(6), 632 - 7
Simultaneous horizontal gene transfer of a gene coding for ribosomal protein l27 and operational genes in Arthrobacter sp; Garcia-Vallve S et al.; Phylogenetic analysis of bacterial L27 ribosomal proteins showed that, against taxonomy, the L27 protein from the Actinobacteria Arthrobacter sp . clusters with protein sequences from the Bacillus group . The L27 gene clusters in the Arthrobacter sp . genome with six genes responsible for creatinine and sarcosine degradation . Phylogenetic analyses of orthologue proteins encoded by three of these genes also showed a phylogenetic relationship with Bacillus species . Comparisons between the synonymous codon usage of the Arthrobacter sp . genes and those from complete genomes showed that Arthrobacter genes encoding the L27 ribosomal protein and the proteins responsible for the degradation of creatinine and sarcosine have a codon usage that is more similar to that of Bacillus species than that of Arthrobacter . We suggest that the Arthrobacter sp . genes encoding the L27 ribosomal protein and the proteins responsible for the degradation of creatinine and sarcosine were acquired simultaneously through horizontal gene transfer from an unknown Bacillus species.

Int J Lepr Other Mycobact Dis, 2002 Sep, 70(3), 174 - 81
Combined 12-month WHO/MDT MB regimen and Mycobacterium w . vaccine in multibacillary leprosy: a follow-up of 136 patients; Kaur I et al.; A total of 136 patients with BI > or = 2 having been followed up for at least 2 years or more were included in the analyses . Seventy-seven out of 136 patients had completed three years follow up . All patients were given WHO/MDT MB regimen for 12 months and additionally 4 doses of Mycobacterium w . vaccine at 3-month intervals . The age of the patients varied from 6 to 77 years (mean 34 +/- 11.3 years) and they had the disease varying from 3 months to 7 years (mean = 1.9 +/- 1.4 years) . The mean of the BI before starting treatment was 3.6 +/- 1.3 . At the end of 2 years follow-up, a total of 54 patients out of the 136 (39.7%) had become smear-negative . A larger proportion of patients, 39/46 (84.8%) with BI of < or = 3 had become smear-negative, whereas, only 10/32 (31.3%) patients with BI between 3.1 to 4 and 5/58 (8.6%) highly bacillated patients having initial BI > 4 had become smear-negative at the end of 2 years . Out of the 77 patients who were available for follow up at 3 years, 30/33 (90.9%) patients with BI of < or = 3, 15/20 (75%) patients with BI between 3.1 to 4 and 13/24 (54.2%) patients having initial BI > 4, respectively, had attained smear negativity . Reactions occurred more frequently after 6 months of therapy and over a period of time their frequency gradually decreased, however, they continued to occur even two years after RFT . During the course of MDT and thereafter in follow up 4.6% and 1.3% of the patients developed new deformities or an increase in the existing grade of deformities, respectively . Three relapses (2 in LL and 1 in BL) occurred in patients having initial BI of > 4 . One patient relapsed in the second year and the other two relapsed in the third year of follow up and were successfully treated with reintroduction of the same MDT MB regimen . Local ulceration healing with scar formation and regional lymphadenopathy were the only local reactions to the vaccine seen in 47/136 (34.5%) patients . All the patients showed histopathological improvement in the form of a gradual reduction of granuloma fraction . Although the results of this limited period follow up are satisfactory, a long-term follow-up in larger number of patients will settle the issue of safety and efficacy of shortened MDT MB regimen and the place of immunotherapy with M . w . vaccine in multibacillary patients.

J Gen Appl Microbiol, 2001 Oct, 47(5), 263 - 267
Production and characterization of tannase from Bacillus cereus KBR9; Mondal KC et al.; A tannase-producing soil bacteria has been isolated and identified as Bacillus cereus . It can degrade tannic acid and produce maximum tannase (0.22 U/ml) at stationary phases of growth (24 h) . Maximum growth and enzyme production occurred with initial medium pH of 4.5-5.0 . Partial purified tannase showed optimum activity at pH 4.5 and 40 degrees C . It remains stable up to 30 degrees C and pH 4.5 to 5.0 . The enzyme is salt tolerant, stable up to 2 m of NaCl and retains 82% original activity in 3 m.

J Gen Appl Microbiol, 2000 Feb, 46(1), 1 - 8
Application of the partial 16S rDNA sequence as an index for rapid identification of species in the genus Bacillus; Goto K et al.; A comparison of 16S rRNA gene (rDNA) sequences was made among type strains of 69 Bacillus species approved in the International Journal of Systematic Bacteriology (IJSB) until 1998 . The results suggested that 5' end region (approx . 275 bp) was the hypervariant region (HV region) in the gene and was highly specific for each type strain . Furthermore, a sequence analysis of the HV region of Bacillus strains revealed that this region was highly conserved within the species . These results indicate that the HV region is a useful index for the identification or grouping of Bacillus species.

J Eur Acad Dermatol Venereol, 2002 Nov, 16(6), 612 - 4
Bacillary angiomatosis of the scalp in a human immunodeficiency virus-negative patient; Kayaselcuk F et al.; Bacillary angiomatosis (BA) is a recently recognized bacterial infectious disease that is mainly seen in patients with the acquired immunodeficiency syndrome . BA has rarely been reported in immunocompetent individuals . A case of BA of the scalp in an immunocompetent patient, who was human immunodeficiency virus seronegative, is reported . The importance of differential diagnosis of this lesion is emphasized.

Parazitologiia, 2002 Sep-Oct, 36(5), 337 - 44
{Cytopathological effect of Bacillus thuringiensis israelensis endotoxins on the intestines of Aedes aegypti mosquito larvae}; Zalunin IA et al.; The dynamics of pathological changes in the intestine of Aedes aegypti larvae under the influence of toxins Cry11A and Cry4B produced by Bacillus thuringiensis israelensis was studied by means of electron microscope . Most significant ultrastructure changes in the intestine of the second instar larvae were observed in the midgut . The cytoplasm of cells disintegrated, and elongated lacunae appeared . The number of microvilli decreased, or they disappeared in the result of destruction . The peritrophic membrane displaced to the lumen of midgut . Any changes in epithelial cells and cuticle in time of foregut and hindgut were not observed in a comparison to control . The toxin Cry4B caused the most effective destruction of the midgut epithelium.

Biochem Biophys Res Commun, 2003 Jan 3, 300(1), 102 - 6
Differential expression of CYP102 in Bacillus megaterium by 17-beta-estradiol and 4-sec-butylphenol; Rowley CW et al.; Previously we have reported the induction of CYP102 in Bacillus megaterium by 17beta-estradiol (E2) and 4-sec-butylphenol (4-sBP) . Electrophoretic mobility shift assay analyses demonstrated that E2 and 4-sBP both cause a dose-dependent disassociation of the Bm3R1 repressor protein from its binding site on the operator sequence of the CYP102 gene . Equimolar combinations of E2 and 4-sBP demonstrated additive induction of CYP102 compared to equivalent samples of E2 and 4-sBP added alone . Two gene constructs were used in this investigation . One construct designated BMC143 contained the entire regulatory region of CYP102 . The other gene construct, designated BMA45, had the "Barbie box" sequence deleted . While the induction of CYP102 by 4-sBP was much higher in the BMC 143 construct, E2 induced CYP102 in both constructs to the same extent . This difference in induction of CYP102 by these two inducers indicates that they act at different sites, either on the Bm3R1 repressor protein or on positive regulatory sites, or that they act, in part, through different mechanisms.

FEMS Microbiol Lett, 2002 Dec 17, 217(2), 263 - 7
Analysis of Bacillus megaterium lipolytic system and cloning of LipA, a novel subfamily I.4 bacterial lipase; Ruiz C et al.; The lipolytic system of Bacillus megaterium 370 was investigated, showing the existence of at least two secreted lipases and a cell-bound esterase . A gene coding for an extracellular lipase was isolated and cloned in Escherichia coli . The cloned enzyme displayed high activity on short to medium chain length (C(4)-C(8)) substrates, and poor activity on C(18) substrates . On the basis of amino acid sequence homology, the cloned lipase was classified into subfamily I.4 of bacterial lipases .

Water Sci Technol, 2002, 46(10), 247 - 54
Simultaneous production of biopesticide and alkaline proteases by Bacillus thuringiensis using sewage sludge as a raw material; Tyagi RD et al.; The simultaneous production of Bacillus thuringiensis (Bt) based biopesticide and proteases was studied using synthetic medium and wastewater sludge as a raw material . The studies were conducted in shake flask and computer controlled 15-L capacity fermentors . Measuring viable cell and spore counts, entomotoxicity and protease activity monitored the progress of the biopesticide production process . A higher viable cell count and spore count was observed in synthetic Soya medium, however, higher entomotoxicity and protease activity were observed in wastewater sludge medium . Thus, the wastewater sludge is a better raw material than commercial Soya medium for the biopesticides and enzyme production . The maximum entomotoxicity and protease activity observed in the fermentor was 9,332 IU/microL and 4.58 IU/mL, respectively . The proteases produced by Bt were also characterised . Two types of proteases were detected; neutral proteases with pH optimum 7.0 and alkaline proteases with pH optimum 10-11 . Further, two types of alkaline proteases were detected; one having a pH and temperature optimum at 10 and 50 degrees C while the other at 11 and 70 degrees C . The protease thermal stability was found to increase in the presence of CaCl2, indicating the proteases were metalloproteases.

Chem Commun (Camb), 2002 Dec 7, (23), 2860 - 1
Biosynthesis of aminoglycoside antibiotics: cloning, expression and characterisation of an aminotransferase involved in the pathway to 2-deoxystreptamine; Huang F et al.; The gene btrR from Bacillus circulans has been cloned and expressed and shown to produce a protein which catalyses the transamination of 2-deoxy-scyllo-inosose to give 2-deoxy-scyllo-inosamine, an intermediate in the biosynthesis of 2-deoxystreptamine.

J Urol, 2003 Jan, 169(1), 357 - 60
A phase I study of intravesical suramin for the treatment of superficial transitional cell carcinoma of the bladder; Uchio EM et al.; PURPOSE: Suramin is a polysulfonated naphthylurea that inhibits proliferation and DNA synthesis of transitional cell carcinoma cell lines . Its large molecular size and negative charge inhibit bladder absorption, making suramin an excellent candidate for intravesical chemotherapy . Intravesical suramin was evaluated in a phase I study to define dose limiting toxicity and systemic absorption, determine a starting dose and regimen for phase II studies and provide a preliminary assessment of in vivo antitumor activity . MATERIALS AND METHODS: Intravesical suramin treatment was administered in 9 patients with histologically identified transitional cell carcinoma (Tcis, Ta or T1) in whom at least 1 course of standard intravesical chemotherapy (bacillus Calmette-Guerin, thiotepa or mitomycin C) had failed . Suramin was administered once weekly for 6 weeks . Patients were treated in groups of 3 using a 60 cc volume and intrapatient dose escalation schedule . Suramin doses of 0.3 to 614.4 mg./ml . were administered intravesically . The last group was treated with the same weekly dose for 6 weeks . RESULTS: The 9 patients underwent 54 treatments with suramin . Plasma suramin concentration after treatment was 1.9 to 38.0 microg./ml . and was not related to treatment dose . The dose escalation phase was limited by the solubility of suramin in solution . Complications included self-limited bladder spasms (less than 24 hours) in 4 of 54 treatments (7%) and new or worsening vesicoureteral reflux in 3 ureters (17%) . Another patient who was treated after the Foley balloon was inflated in the urethra experienced bladder spasms, skin flushing and fever (39C) . Mean bladder capacity before and after treatment was 600 and 540 ml., respectively . At followup 7 patients had stage Ta tumors and 2 had carcinoma in situ . CONCLUSIONS: An intravesical suramin dose of 153 mg./ml was defined as a safe treatment parameter with acceptable plasma concentrations and minimal side effects . Phase II studies are needed to assess the antitumor activity of suramin in patients with transitional cell carcinoma of the bladder.

J Urol, 2003 Jan, 169(1), 96 - 100; discussion 100
A retrospective analysis of 153 patients treated with or without intravesical bacillus Calmette-Guerin for primary stage T1 grade 3 bladder cancer: recurrence, progression and survival; Shahin O et al.; PURPOSE: We retrospectively evaluated the long-term outcome in patients with newly diagnosed stage T1 grade 3 bladder cancer treated with transurethral resection with or without intravesical bacillus Calmette-Guerin (BCG) . MATERIALS AND METHODS: Of 153 patients with a median age of 67 years (range 36 to 88) and a male-to-female ratio of 4:1 we treated 92 with transurethral bladder resection and additional BCG, and 61 with transurethral bladder resection alone . BCG was administered intravesically as 120 mg . BCG Pasteur F dissolved in 50 ml . saline, retained for up to 2 hours weekly for 6 weeks and repeated as necessary . RESULTS: Median followup was 5.3 years (range 0.4 to 18.2) . Disease recurred in 70% of the patients treated with BCG and in 75% treated with transurethral resection alone . Median time to recurrence was 38 and 22 months for BCG and resection alone (p = 0.19) . Tumor progressed in 33% of patients with BCG and in 36% with resection alone . Deferred cystectomy was performed in 29% of the patients with BCG and in 31% with resection alone . Overall and disease specific survival did not differ significantly . CONCLUSIONS: Our results suggest that intravesical BCG therapy after transurethral bladder resection for stage T1 grade 3 bladder cancer may delay the time to recurrence and cystectomy but it does not substantially alter the final outcome . Our findings reflect the rule of 30% for stage T1 grade 3 cancer, namely approximately 30% of patients never have recurrence, 30% ultimately die of metastatic disease and 30% require deferred cystectomy.

J Urol, 2003 Jan, 169(1), 90 - 5
Intravesical bacillus Calmette-Guerin versus mitomycin C for superficial bladder cancer: a formal meta-analysis of comparative studies on recurrence and toxicity; Bohle A et al.; PURPOSE: We compare the therapeutic efficacy and toxicity of intravesical bacillus Calmette-Guerin (BCG) with mitomycin C on recurrence of stages Ta and T1 bladder carcinoma . MATERIALS AND METHODS: Combined published and unpublished data from comparative studies on BCG versus mitomycin C for superficial bladder carcinoma considering possible confounding factors were analyzed . Odds ratio (OR) and its 95% CI were used as primary effect size estimate . Toxicity data were evaluated descriptively . RESULTS: In 11 eligible clinical trials 1,421 patients were treated with BCG and 1,328 were treated with mitomycin C . Within the overall median followup time of 26 months 38.6% of the patients in the BCG group and 46.4% of those in the mitomycin C group had tumor recurrence . In 7 of 11 studies BCG was significantly superior to mitomycin C, in 3 studies no significant difference was found, while in 1 study mitomycin C was significantly superior to BCG . An overall statistically significant superiority of BCG versus mitomycin C efficacy in reducing tumor recurrence was detected (OR 0.56, 95% CI 0.38 to 0.84, p = 0.005) . In the subgroup treated with BCG maintenance all 6 individual studies showed a significant superiority of BCG over mitomycin C (OR 0.43, 95% CI 0.35 to 0.53, p <0.001) . In 4 of the 5 studies with reported data on toxicity BCG associated cystitis was significantly more frequent than in the mitomycin C group (53.8% versus 39.2%) . The combined cystitis OR was 1.81 (95% CI 1.48 to 2.23, p <0.001) . The OR for cystitis in the BCG maintenance group did not significantly differ from that in the nonmaintenance therapy group . CONCLUSIONS: The results suggest superiority of BCG over mitomycin C for prevention of tumor recurrences in the combined data and particularly in the BCG maintenance treatment subgroup, irrespective of the actual (intermediate or high) tumor risk status . The toxicity with BCG is higher but does not differ between BCG maintenance and nonmaintenance groups.

Eur Urol, 2002 Dec, 42(6), 542 - 6
Beware the BCG failures: a review of one institution's results; Lockyer CR et al.; OBJECTIVE: To review the outcome of all superficial transitional cell (TCC) bladder cancer treated with intravesical Bacille Calmette-Guerin (BCG) at one institution and, in particular, the prognosis for those patients who gained little benefit from BCG therapy . PATIENTS AND METHODS: The notes of 122 patients treated with BCG over a nine-year period were reviewed . The following details were recorded: time of diagnosis; time of decision to treat with BCG; results of cystoscopies before and after BCG; duration of follow up; time of progression if occurred, mortality and cause of death . RESULTS: Complete follow up data was available for 112 patients . At a median follow up of 23 months (range 3-107) 57 patients (51%) remained free of tumour, 30 (27%) had progressed and 18 (16%) had died of transitional cell carcinoma . There was a significant association between a positive initial check cystoscopy and subsequent progression (p<0.001) and disease specific mortality (p<0.001) . Of the 35 patients who had a positive cystoscopy after BCG treatment 21 (60%) progressed and 14 (40%) died of transitional cell carcinoma compared with 9 (12%) and 4 (5%) of the 77 with a negative cystoscopy . Adjusted odds ratios for progression and death from TCC for patients with a positive initial check cystoscopy were 21 and 13, respectively . CONCLUSION: In our series the patients found to have tumour at the initial check cystoscopy following intravesical BCG had a poor prognosis . This should be remembered when considering treatment options and counselling patients . Follow up of all BCG patients need to be rigorous and protocols would help to unify the treatment patients receive.

J Steroid Biochem Mol Biol, 2002 Oct, 82(2-3), 257 - 61
Influence of substituents at C11 on hydroxylation of progesterone analogs by Bacillus sp; Das S et al.; Transformation of progesterone analogs viz., progesterone, 11 alpha-, 11 beta-hydroxyprogesterones and 11-ketoprogesterone by Bacillus sp . is reported . Both progesterone and 11-ketoprogesterone were hydroxylated while the C(11) epimeric alcohols of progesterone remained unaltered under the conditions used . The major bioconverted products obtained from progesterone and 11-ketoprogesterone were characterized as 6 beta- and 14 alpha-hydroxyprogesterones and 14 alpha-hydroxy-11-ketoprogesterone respectively by mass and NMR spectra . The conversion of 11-ketoprogesterone to its 14 alpha-hydroxy derivative by microbe is unprecedented and novel . Moreover, hydroxylation at 6 beta- and 14 alpha-positions of progesterone by Bacillus sp . is significant . In conclusion, the present data showed that the substituents at 11-position of steroid play important role on hydroxylation by microbe.

J Biol Chem, 2003 Feb 28, 278(9), 7663 - 73 Epub 2002 Dec 09.
Crystal structure of Bacillus sp . GL1 xanthan lyase, which acts on the side chains of xanthan; Hashimoto W et al.; Xanthan lyase, a member of polysaccharide lyase family 8, is a key enzyme for complete depolymerization of a bacterial heteropolysaccharide, xanthan, in Bacillus sp . GL1 . The enzyme acts exolytically on the side chains of the polysaccharide . The x-ray crystallographic structure of xanthan lyase was determined by the multiple isomorphous replacement method . The crystal structures of xanthan lyase and its complex with the product (pyruvylated mannose) were refined at 2.3 and 2.4 A resolution with final R-factors of 17.5 and 16.9%, respectively . The refined structure of the product-free enzyme comprises 752 amino acid residues, 248 water molecules, and one calcium ion . The enzyme consists of N-terminal alpha-helical and C-terminal beta-sheet domains, which constitute incomplete alpha(5)/alpha(5)-barrel and anti-parallel beta-sheet structures, respectively . A deep cleft is located in the N-terminal alpha-helical domain facing the interface between the two domains . Although the overall structure of the enzyme is basically the same as that of the family 8 lyases for hyaluronate and chondroitin AC, significant differences were observed in the loop structure over the cleft . The crystal structure of the xanthan lyase complexed with pyruvylated mannose indicates that the sugar-binding site is located in the deep cleft, where aromatic and positively charged amino acid residues are involved in the binding . The Arg(313) and Tyr(315) residues in the loop from the N-terminal domain and the Arg(612) residue in the loop from the C-terminal domain directly bind to the pyruvate moiety of the product through the formation of hydrogen bonds, thus determining the substrate specificity of the enzyme.

Urol Oncol, 2002 Mar-Apr, 7(2), 67 - 72
Differential C-erbB-2 and VEGF expression following BCG immunotherapy in superficial papillary transitional cell carcinoma of the bladder; Morgan BE et al.; Bacillus Calmette Guerin (BCG) is generally regarded as an effective immunotherapy for superficially invasive papillary transitional cell carcinoma of the bladder . The exact mechanism(s) which underlie its efficacy are unknown . As C-erbB-2 oncoprotein and vascular endothelial growth factor (VEGF) have been shown to be over-expressed in TCC of the bladder, it has been postulated that they may be important in its pathogenesis . The purpose of this study was to 1.) differentially evaluate the effect of BCG immunotherapy in treated and untreated cohorts on the immunohistochemical expression of C-erbB-2 and VEGF in formalin-fixed paraffin-embedded sections of superficial and superficially invasive (Stage Ta-T1) transitional cell carcinoma of the bladder . Immunolabeling intensity was assessed independently by two pathologists and reported as a mean labeling index . The results confirm previous studies that 1.) both c-erbB-2 and VEGF are over-expressed in these tumors MLI = 90.1 and 45.7 respectively, 2.) that VEGF is an early and sensitive indicator of TCC, and 3.) that BCG has a salutary effect on papillary TCC, 66% vs . 89% recurrence rate, P = .04 . Our findings show that 1.) C-erbB-2 expression is decreased in patients tumors which show response to BCG (45.7 to 38.5), P = 0.15, 2.) that BCG administration has no effect on the expression of VEGF . While the decrement in c-erbB-2 immunostaining observed in those patients who received BCG contrasts with the increase in c-erbB-2 immunolabeling observed in patients who did not receive BCG, the differences were not statistically significant and could reflect tumor grade or stage regression associated with BCG therapy . However, this study suggests that BCG differentially influences the expression of C-erbB-2 and VEGF.

Trans R Soc Trop Med Hyg, 2002 Sep-Oct, 96(5), 549 - 50
High rate of Bartonella henselae infection in HIV-positive outpatients in Johannesburg, South Africa; Frean J et al.; The emerging opportunistic pathogen Bartonella henselae has a wide range of clinical presentation, which includes, particularly, bacillary angiomatosis . This non-random pilot survey of outpatients attending HIV clinics in Johannesburg, South Africa, sampled 188 patients, in whom there was a 10% prevalence of Bartonella bacteraemia, as determined by nested polymerase chain reaction.

J Biol Chem, 2003 Feb 28, 278(9), 7310 - 9 Epub 2002 Dec 06.
Tracking the putative biosynthetic precursors of oxygenated mycolates of Mycobacterium tuberculosis . Structural analysis of fatty acids of a mutant strain deviod of methoxy- and ketomycolates; Dinadayala P et al.; Disruption of the mma4 gene (renamed hma) of Mycobacterium tuberculosis has yielded a mutant strain defective in the synthesis of both keto- and methoxymycolates, with an altered cell-wall permeability to small molecules and a decreased virulence in the mouse model of infection (Dubnau, E., Chan, J., Raynaud, C., Mohan, V . P., Laneelle, M . A., Yu, K., Quemard, A., Smith, I., and Daffe, M . (2000) Mol . Microbiol . 36, 630-637) . Assuming that the mutant would accumulate the putative precursors of the oxygenated mycolates of M . tuberculosis, a detailed structural analysis of mycolates from the hma-inactivated strain was performed using a combination of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, proton NMR spectroscopy, and chemical degradation techniques . These consisted most exclusively of alpha-mycolates, composed of equal amounts of C(76)-C(82) dicyclopropanated (alpha(1)) and of C(77)-C(79) monoethylenic monocyclopropanated (alpha(2)) mycolates, the double bond being located at the "distal" position . In addition, small amounts of cis-epoxymycolates, structurally related to alpha(2)-mycolates, was produced by the mutant strain . Complementation of the hma-inactivated mutant with the wild-type gene resulted in the disappearance of the newly identified mycolates and the production of keto- and methoxymycolates of M . tuberculosis . Introduction of the hma gene in Mycobacterium smegmatis led to the lowering of diethylenic alpha mycolates of the recipient strain and the production of keto- and hydroxymycolates . These data indicate that long-chain ethylenic compounds may be the precursors of the oxygenated mycolates of M . tuberculosis . Because the lack of production of several methyltransferases involved in the biosynthesis of mycolates is known to decrease the virulence of the tubercle bacillus, the identification of the substrates of these enzymes should help in the design of inhibitors of the growth of M . tuberculosis.

Mem Inst Oswaldo Cruz, 2002 Oct, 97(7), 1041 - 8
Study of the safety, immunogenicity and efficacy of attenuated and killed Leishmania (Leishmania) major vaccines in a rhesus monkey (Macaca mulatta) model of the human disease; Amaral VF et al.; We have compared the efficacy of two Leishmania (Leishmania) major vaccines, one genetically attenuated (DHFR-TS deficient organisms), the other inactivated {autoclaved promastigotes (ALM) with bacillus Calmete-Guerin (BCG)}, in protecting rhesus macaques (Macaca mulatta) against infection with virulent L . (L.) major . Positive antigen-specific recall proliferative response was observed in vaccinees (79% in attenuated parasite-vaccinated monkeys, versus 75% in ALM-plus-BCG-vaccinated animals), although none of these animals exhibited either augmented in vitro gamma interferon (IFN-gamma) production or positive delayed-type hypersensitivity (DTH) response to the leishmanin skin test prior to the challenge . Following challenge, there were significant differences in blastogenic responses (p < 0.05) between attenuated-vaccinated monkeys and naive controls . In both vaccinated groups very low levels of antibody were found before challenge, which increased after infective challenge . Protective immunity did not follow vaccination, in that monkeys exhibited skin lesion at the site of challenge in all the groups . The most striking result was the lack of pathogenicity of the attenuated parasite, which persisted in infected animals for up to three months, but were incapable of causing disease under the conditions employed . We concluded that both vaccine protocols used in this study are safe in primates, but require further improvement for vaccine application.

Anal Biochem, 2002 Dec 15, 311(2), 103 - 18
Radical-generating coordination complexes as tools for rapid and effective fragmentation and fluorescent labeling of nucleic acids for microchip hybridization; Kelly JJ et al.; DNA microchip technology is a rapid, high-throughput method for nucleic acid hybridization reactions . This technology requires random fragmentation and fluorescent labeling of target nucleic acids prior to hybridization . Radical-generating coordination complexes, such as 1,10-phenanthroline-Cu(II) (OP-Cu) and Fe(II)-EDTA (Fe-EDTA), have been commonly used as sequence nonspecific "chemical nucleases" to introduce single-strand breaks in nucleic acids . Here we describe a new method based on these radical-generating complexes for random fragmentation and labeling of both single- and double-stranded forms of RNA and DNA . Nucleic acids labeled with the OP-Cu and the Fe-EDTA protocols revealed high hybridization specificity in hybridization with DNA microchips containing oligonucleotide probes selected for identification of 16S rRNA sequences of the Bacillus group microorganisms.We also demonstrated cDNA- and cRNA-labeling and fragmentation with this method . Both the OP-Cu and Fe-EDTA fragmentation and labeling procedures are quick and inexpensive compared to other commonly used methods . A column-based version of the described method does not require centrifugation and therefore is promising for the automation of sample preparations in DNA microchip technology as well as in other nucleic acid hybridization studies.

Mycorrhiza, 2002 Dec, 12(6), 313 - 6 Epub 2002 Sep 24.
Inoculation of field-established mulberry and papaya with arbuscular mycorrhizal fungi and a mycorrhiza helper bacterium; Mamatha G et al.; The effects of soil inoculation with arbuscular mycorrhizal (AM) fungi and a mycorrhiza helper bacterium (MHB) were investigated on mulberry and papaya plants already established in the field . Ten-year-old mulberry plants (var . M5) were inoculated with Glomus fasciculatum and 1.5-year-old papaya plants (var . Solo) were inoculated with a mixed culture of G . mosseae and G . caledonium with or without Bacillus coagulans at two levels of P fertilizer . Growth, P uptake, yield and AM colonization levels were monitored . Leaf yield in mulberry and fruit yield in papaya were minimal in uninoculated plants given 50% recommended P . However, crop yields of both mulberry and papaya inoculated with AM fungi alone or together with the bacterium and given 50% recommended P were statistically on a par with that of uninoculated plants given 100% recommended P . As inoculation of B . coagulans increased mycorrhiza levels in AM fungal-inoculated plants, this may be included in the class of MHB . Thus, mulberry and papaya already established in the field may respond to AM inoculation and MHB may increase symbiosis development by efficient AM fungi.

Appl Microbiol Biotechnol, 2002 Dec, 60(4), 381 - 95 Epub 2002 Oct 12.
An overview on fermentation, downstream processing and properties of microbial alkaline proteases; Gupta R et al.; Microbial alkaline proteases dominate the worldwide enzyme market, accounting for a two-thirds share of the detergent industry . Although protease production is an inherent property of all organisms, only those microbes that produce a substantial amount of extracellular protease have been exploited commercially . Of these, strains of Bacillus sp . dominate the industrial sector . To develop an efficient enzyme-based process for the industry, prior knowledge of various fermentation parameters, purification strategies and properties of the biocatalyst is of utmost importance . Besides these, the method of measurement of proteolytic potential, the selection of the substrate and the assay protocol depends upon the ultimate industrial application . A large array of assay protocols are available in the literature; however, with the predominance of molecular approaches for the generation of better biocatalysts, the search for newer substrates and assay protocols that can be conducted at micro/nano-scale are becoming important . Fermentation of proteases is regulated by varying the C/N ratio and can be scaled-up using fed-batch, continuous or chemostat approaches by prolonging the stationary phase of the culture . The conventional purification strategy employed, involving e.g., concentration, chromatographic steps, or aqueous two-phase systems, depends on the properties of the protease in question . Alkaline proteases useful for detergent applications are mostly active in the pH range 8-12 and at temperatures between 50 and 70 degrees C, with a few exceptions of extreme pH optima up to pH 13 and activity at temperatures up to 80-90 degrees C . Alkaline proteases mostly have their isoelectric points near to their pH optimum in the range of 8-11 . Several industrially important proteases have been subjected to crystallization to extensively study their molecular homology and three-dimensional structures.

Proc Natl Acad Sci U S A, 2002 Dec 24, 99(26), 16581 - 6 Epub 2002 Dec 03.
Bacillus thuringiensis-toxin resistance management: stable isotope assessment of alternate host use by Helicoverpazea; Gould F et al.; Data have been lacking on the proportion of Helicovera zea larvae that develop on noncotton host plants that can serve as a refuge from selection pressure for adaptation to transgenic cotton varieties that produce a toxin from the bacterium Bacillus thuringiensis . We found that individual H . zea moths that develop as larvae on cotton and other plants with C3 physiology have a different ratio of 13C to 12C than moths that develop on plants with C4 physiology, such as corn . We used this finding in determining the minimum percentage of moths that developed on noncotton hosts in two cotton-growing areas . Our results indicate that local corn can serve as a refuge for H . zea in midsummer . Our results contrast dramatically with the prevailing hypothesis that the large majority of late-season moths are produced from larvae feeding on cotton, soybean, and other C3 plants . Typically, <50% of moths captured in August through October have isotope ratios indicative of larval feeding on C3 plants . In one October sample, 100% of the moths originated from C4 hosts even though C4 crops were harvested at least 1 mo earlier, and no common wild C4 hosts were available . These findings support other research indicating that many late-season H . zea moths captured in Louisiana and Texas are migrants whose larvae developed on corn in more northern locations . Our isotope data on moths collected in Texas early in the season indicate that the majority of overwintering H . zea do not originate from cotton-feeding larvae and may be migrants from Mexico . Non-Bt corn in Mexico and the U.S . corn belt appears to serve as an important refuge for H . zea.

Cell, 2002 Nov 27, 111(5), 747 - 56
Sensing small molecules by nascent RNA: a mechanism to control transcription in bacteria; Mironov AS et al.; Thiamin and riboflavin are precursors of essential coenzymes-thiamin pyrophosphate (TPP) and flavin mononucleotide (FMN)/flavin adenine dinucleotide (FAD), respectively . In Bacillus spp, genes responsible for thiamin and riboflavin biosynthesis are organized in tightly controllable operons . Here, we demonstrate that the feedback regulation of riboflavin and thiamin genes relies on a novel transcription attenuation mechanism . A unique feature of this mechanism is the formation of specific complexes between a conserved leader region of the cognate RNA and FMN or TPP . In each case, the complex allows the termination hairpin to form and interrupt transcription prematurely . Thus, sensing small molecules by nascent RNA controls transcription elongation of riboflavin and thiamin operons and possibly other bacterial operons as well.

Lett Appl Microbiol, 2002, 35(6), 523 - 7
Optimization of a fermentation medium for the production of Penicillin G acylase from Bacillus sp; Rajendhran J et al.; AIMS: Optimization of Penicillin G acylase (PAC) production from a novel isolate of Bacillus sp . METHODS: Fermentation medium for PAC production was optimized using a two-level fractional factorial design with seven components . RESULTS: A maximum production of 9.5 U ml(-1) of PAC was obtained in an optimized medium containing (g l(-1): K2HPO4, 1.0; MgSO4.7H2O, 0.1; CaCl2.2H2O, 0.1; PAA, 2.0; tryptone, 5.0; yeast extract, 3.0; and sucrose, 50.0 . SIGNIFICANCE AND IMPACT OF THE STUDY: The two-step medium optimization resulted in a twofold increase in PAC production . Since the strain Bacillus sp . PGS10 produces a high level of PAC, it could be a potential candidate for industrial production of PAC.

Immunology, 2002 Dec, 107(4), 500 - 6
Phagocytosis of bacille Calmette-Guérin-infected necrotic macrophages induces a maturation phenotype and evokes antigen-presentation functions in dendritic cells; Goldmann O et al.; The interaction of pathogens with dendritic cells (DCs) seems to play a critical role in the initiation of the immune response . Tissue damage and induction of an inflammatory reaction are events frequently associated with the progression of the infection . Although DCs are very efficient at phagocytosing pathogens, the capacity of these cells to uptake microbes from a necrotic environment has not yet been proven . Here we have investigated the ability of murine bone marrow-derived DCs to maturate and acquire antigen-presentation functions when cocultured with bacille Calmette-Guerin (BCG)-infected necrotic macrophages . Immature DCs exhibited a prominent capacity to ingest necrotic material as demonstrated by flow cytometry analysis and confocal microscopy . Furthermore, after exposure to BCG-infected necrotic macrophages, DCs underwent phenotypic changes, including the up-regulation of maturation specific markers (major histocompatibility complex class II, CD40, CD80, and CD86) and the capacity to stimulate antigen-specific CD4+ T cells with higher efficiency than when they were directly infected with a similar number of bacteria . Antigen presentation following phagocytosis of BCG-infected necrotic macrophages was demonstrated by their ability to stimulate in vitro proliferation and interferon-gamma production of antigen-specific CD4+ T cells . These results suggest that the functional changes occurring in DCs after interaction with a pathogen can be favoured when the encounter takes place in a necrotic environment and it may constitute an important mechanism for the amplification of class II-restricted immune responses induced during infection.

Virus Res, 2002 Dec, 90(1-2), 269 - 74
Genomic instability of prawn white spot bacilliform virus (WSBV) and its association to virus virulence; Lan Y et al.; Prawn White Spot Bacilliform Virus (WSBV) is a major pathogen that causes prawn diseases . In this study, we examined the sequence of WSBV genome DNA in the shrimp Penaeus japonicus, P . vannamei, P . Monodon, P . chinensis and Metapenaeus ensis through successive PCR amplification of the DNA fragments in the whole WSBV genome . We found a sequence deletion hotspot in the WSBV genome that is 305107 bp in length . The sizes of the deleted fragments were 4.6, 4.8 or 8.1 kbp depending on the species of prawn . Since the mortality of shrimp infected by the intact WSBV was always significantly higher than that of shrimp infected by DNA fragment-deleted WSBV, we suggest that this deletion be somehow linked to the virulence of the virus itself . This result may lead to the discovery of the molecular mechanism of the pathogenicity of WSBV in shrimps.

Res Microbiol, 2002 Nov, 153(9), 605 - 9
Regulation of crystal protein biosynthesis by Bacillus thuringiensis: II . Effects of carbon and nitrogen sources; Icgen Y et al.; The regulation of crystal protein production in Bacillus thuringiensis 81 by sources of carbon and nitrogen was investigated . The highest titers of toxin were obtained on sucrose, lactose and inulin which also supported sporulation . Whey and molasses were also potential carbon substrates for toxin production . Other carbohydrates including glucose, glycerol, maltose, starch and dextrin yielded lower amounts of toxin . Nitrogen sources were found to exert the most profound controls . Peptone was the best organic nitrogen source, supporting optimum production and sporulation as well as high cell density . The formation of CryI and CryII toxin proteins was found to be differentially regulated by the inorganic nitrogenous compounds incorporated into the medium.

Res Microbiol, 2002 Nov, 153(9), 599 - 604
Regulation of crystal protein biosynthesis by Bacillus thuringiensis: I . Effects of mineral elements and pH; Icgen Y et al.; Crystal protein synthesis by a local isolate of Bacillus thuringiensis was monitored and compared in association with growth and sporulation in media differing in mineral element content . Mg and Cu were the most important metals for the biosynthesis of 135 kDa and 65 kDa toxin components in that the former was essential and the latter was greatly stimulatory at 10(-6) to 10(-7) M concentration . Also the inclusion of Mn favored toxin production at concentrations ranging from 3 x 10(-4) to 10(-5) M . The omission of Zn and Ca had no effect on toxin formation . Crystal protein synthesis and sporulation did not generally seem to be co-regulated by the minerals as these processes responded differently to mineral levels . There was no evidence for suppression of biosynthesis by inorganic phosphate over a range of 3 to 100 mM . Crystal protein production was more efficient in buffered medium, especially when the initial pH was adjusted to 6.5.

Acta Crystallogr D Biol Crystallogr, 2002 Dec, 58(Pt 12), 2198 - 200 Epub 2002 Nov 23.
The 2.2 A resolution structure of thermolysin (TLN) crystallized in the presence of potassium thiocyanate; Gaucher JF et al.; A new crystallization protocol for thermolysin (EC 3.4.24.27) from Bacillus thermoproteolyticus is presented . After dissolving the protein in the presence of KSCN, which avoids the use of DMSO and CsCl, crystals were obtained following the salting-in method . Crystal cell parameters are isomorphous with those previously reported from DMSO/CsCl mixtures . The new SCN(-) crystal structure has been analyzed . It shows the presence of one thiocyanate ion in the catalytic site and several rearrangements in the S(1) and S(2) subsites . These results are in agreement with the measurements of Inouye et al . {(1998), J . Biochem . (Tokyo), 123, 847-852}, who observed in solution that the solubility of TLN, which is particularly poor in low ionic strength solutions, increases dramatically in the presence of several neutral salts . The results reported here suggest possible explanations for the solubility increase and for the inhibitory effects of high SCN(-) concentrations on thermolysin activity.

Acta Crystallogr D Biol Crystallogr, 2002 Dec, 58(Pt 12), 2175 - 6 Epub 2002 Nov 23.
Crystallization and preliminary X-ray diffraction analysis of a thermostable D-hydantoinase from the mesophilic Bacillus sp . AR9; Agrawal V et al.; D-hydantoinase catalyzes the conversion of DL-hydantoin derivatives to the corresponding optically pure N-carbamoyl amino acids, the first step in the industrial preparation of optically pure amino acids using biological catalysts . A thermostable D-hydantoinase from the mesophilic bacteria Bacillus sp . AR9 has been crystallized in three different crystal forms . The hexagonal faced crystals were the best looking, but did not diffract . One of the crystal forms is star-shaped and appeared to be twinned, but diffracted as a single crystal to a resolution of 2.3 A . These crystals belong to space group P6(4) and have unit-cell parameters a = b = 129.55, c = 102.86 A, alpha = beta = 90, gamma = 120 degrees.

J Clin Microbiol, 2002 Dec, 40(12), 4750 - 2
Performance of the microscopic observation drug susceptibility assay in drug susceptibility testing for Mycobacterium tuberculosis; Park WG et al.; The drug susceptibility testing performance of a broth-based method with microscopic reading of bacillary growth, the microscopic observation drug susceptibility (MODS) assay, was compared to that of the reference 7H10 agar method of proportion by using 53 isolates of Mycobacterium tuberculosis from persons at risk for multidrug-resistant TB . For isoniazid (0.1 micro g/ml) and rifampin (2.0 micro g/ml), there was 100% agreement between MODS results read at day 11 and the reference method . Levels of agreement for ethambutol tested at 2.5 and 7.5 micro g/ml were 70 and 58%, respectively . Levels of agreement for streptomycin tested at 2.0 and 6.0 micro g/ml were 77 and 51%, respectively . For isoniazid and rifampin drug susceptibility testing, MODS is as accurate as and more rapid than the reference method.

Emerg Infect Dis, 2002 Nov, 8(11), 1327 - 33
Tuberculosis-related deaths within a well-functioning DOTS control program; Garcia-Garcia Mde L et al.; To describe the molecular epidemiology of tuberculosis (TB)-related deaths in a well-managed program in a low-HIV area, we analyzed data from a cohort of 454 pulmonary TB patients recruited between March 1995 and October 2000 in southern Mexico . Patients who were sputum acid-fast bacillus smear positive underwent clinical and mycobacteriologic evaluation (isolation, identification, drug-susceptibility testing, and IS6110-based genotyping and spoligotyping) and received treatment from the local directly observed treatment strategy (DOTS) program . After an average of 2.3 years of follow-up, death was higher for clustered cases (28.6 vs . 7%, p=0.01) . Cox analysis revealed that TB-related mortality hazard ratios included treatment default (8.9), multidrug resistance (5.7), recently transmitted TB (4.1), weight loss (3.9), and having less than 6 years of formal education (2) . In this community, TB is associated with high mortality rates.

J Appl Microbiol, 2002, 93(6), 937 - 43
Sterilization of plastic containers using electron beam irradiation directed through the opening; Cleghorn DA et al.; AIMS: Studies were performed to demonstrate the efficacy of a novel electron beam irradiation system for the on-line sterilization of polymeric containers using energetic electrons directed through the container opening . METHODS AND RESULTS: The distribution of dosage delivered during conveyance beneath the electron beam treatment system was determined for two sizes (8 or 16 ounces) of high-density polyethylene (HDPE) blow-moulded bottles . The biological effects of treatment were then determined using Bacillus pumilus ATCC 27142 spores inoculated as 10 microl droplets dried (i) onto the surface of flat coupons of the high-density polyethylene bottle material, and (ii) onto the region of lowest delivered dose (the bottom side wall) within each bottle . The inactivation obtained was determined by examining the level of survival after swab recovery of the inoculated spores, with reference to the level recovered from untreated control samples . CONCLUSIONS: The inactivation of B . pumilus spores during treatment was reproducible and proportional to the applied dose . In each instance, the logarithm of the surviving fraction of spores was well fitted by linear regression models and in good general agreement with values reported for inactivation of the same organism using gamma irradiation . SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this work demonstrate the potential of a new physical method capable of high throughput for in-line sterilization of polymeric containers . The ability of the process to eliminate the washing step involved with traditional chemical sterilants, such as hydrogen peroxide, greatly reduces the size and environmental impact of these systems.

J Appl Microbiol, 1997 Feb, 82(2), 253 - 8
Natural occurrence of Bacillus thuringiensis on cabbage foliage and in insects associated with cabbage crops; Damgaard PH et al.; Bacillus thuringiensis was isolated from the phylloplane of organically grown cabbage in one field during two growth seasons (1992-93) . The frequency of B . thuringiensis varied between 0.02 and 0.67 of the total B . cereus/B . thuringiensis population, with an average of 0.11 . Characterization of the B . thuringiensis isolates from foliage showed that the majority (64% of 150 isolates) belonged to serovar kurstaki, had bipyramidal crystals and toxicity towards Pieris brassicae and/or Trichoplusia ni . Other serovars were also found on the foliage but occurred at very low frequencies (one to three isolates of each serovar) . Bacillus thuringiensis was also isolated from insects associated with the cabbage crop (Pieris rapae (Lep.), Delia radicum (Dip.), Syrphidae ribesii (Dip.) and Aleochara bilineata (Col.)), which were collected alive at different developmental stages in the same field . Serologically these isolates were assigned to the serovars kurstaki, aizawai, tochigiensis, colmeri and indiana/colmeri.

J Gen Physiol, 2002 Dec, 120(6), 845 - 53
The cation selectivity filter of the bacterial sodium channel, NaChBac; Yue L et al.; The Bacillus halodurans voltage-gated sodium-selective channel (NaChBac) (Ren, D., B . Navarro, H . Xu, L . Yue, Q . Shi, and D.E . Clapham . 2001b . SCIENCE: 294:2372-2375), is an ideal candidate for high resolution structural studies because it can be expressed in mammalian cells and its functional properties studied in detail . It has the added advantage of being a single six transmembrane (6TM) orthologue of a single repeat of mammalian voltage-gated Ca(2+) (Ca(V)) and Na(+) (Na(V)) channels . Here we report that six amino acids in the pore domain (LESWAS) participate in the selectivity filter . Replacing the amino acid residues adjacent to glutamatic acid (E) by a negatively charged aspartate (D; LEDWAS) converted the Na(+)-selective NaChBac to a Ca(2+)- and Na(+)-permeant channel . When additional aspartates were incorporated (LDDWAD), the mutant channel resulted in a highly expressing voltage-gated Ca(2+)-selective conductance.

Appl Environ Microbiol, 2002 Dec, 68(12), 6410 - 5
Existence of a true phosphofructokinase in Bacillus sphaericus: cloning and sequencing of the pfk gene; Alice AF et al.; Some strains of Bacillus sphaericus are entomopathogenic to mosquito larvae, which transmit diseases, such as filariasis and malaria, affecting millions of people worldwide . This species is unable to use hexoses and pentoses as unique carbon sources, which was proposed to be due to the lack of glycolytic enzymes, such as 6-phosphofructokinase (PFK) . In this study, PFK activity was detected and the pfk gene was cloned and sequenced . Furthermore, this gene was shown to be present in strains belonging to all the homology groups of this heterogeneous species, in which PFK activity was also detected . A careful sequence analysis revealed the conservation of different catalytic and regulatory residues, as well as the enzyme's phylogenetic affiliation with the family of allosteric ATP-PFK enzymes.

Vaccine, 2002 Dec 13, 21(3-4), 158 - 66
Combined intrarectal/intradermal inoculation of recombinant Mycobacterium bovis bacillus Calmette-Guérin (BCG) induces enhanced immune responses against the inserted HIV-1 V3 antigen; Kawahara M et al.; The development of a successful recombinant Mycobacterium bovis bacillus Calmette-Guerin (rBCG) vector-based vaccine for human immunodeficiency virus type 1 (HIV-1) requires the induction of high levels of HIV-1-specific immunity while at the same time maintaining immunity to tuberculosis . To examine a combined vaccination strategy for enhancement of immune responses specific for HIV-1, guinea pigs were inoculated with either a single or combination intradermal (i.d.), intrarectal (i.r.) and intranasal (i.n.) administration of rBCG-pSOV3J1 which secretes a chimeric protein of HIV-1 V3J1 peptide and alpha-antigen . Significant level of delayed-type hypersensitivity to both V3J1 peptide and tuberculin was induced in guinea pigs inoculated with human doses of rBCG-pSOV3J1 by a combination of intrarectal and intradermal routes . Guinea pigs inoculated by combined routes also had significantly higher titers of HIV-1-specific serum IgG and IgA compared with those animals immunized only intrarectally, which led to the enhanced neutralization activity against HIV-1(MN) . In addition, the induction of high levels of IFNgamma and interleukin-2 (IL-2) mRNA in PBMC, splenocytes, and intraepithelial lymphocytes from the immunized animals was detected until at least 110 weeks post-inoculation . These results suggest that enhanced immune responses specific for HIV-1 are efficiently induced by combined intrarectal and intradermal immunization with rBCG-HIV, and antigen-specific Th1-type memory cells are maintained for more than 2 years in the immunized animals . Thus, inoculation with rBCG-HIV by combined routes represents an effective vaccination strategy to elicit high levels of HIV-1-specific immune responses.

Lepr Rev, 2002 Sep, 73(3), 254 - 61
Anti-leprosy protective vaccination of rhesus monkeys with BCG or BCG plus heat-killed Mycobacterium leprae: lepromin skin test results; Gormus BJ et al.; Groups of rhesus monkeys (RM) were vaccinated and boosted with living Mycobacterium bovis Bacillus Calmette-Guerin (BCG) or BCG + low dose (LD) heat-killed Mycobacterium leprae (HKML) or high dose (HD) HKML or were unvaccinated . Animals vaccinated with BCG + LD and HD HKML were lepromin skin tested 2 weeks after boosting . All groups were lepromin tested 37 and 46 months after challenge with live M . leprae . Fernandez (72 h) and Mitsuda (28 day) responses were recorded . Ten of 10 rhesus monkeys in each of the two BCG + HKML-vaccinated groups significantly converted to strong positive Fernandez status within 2 weeks of boosting, compared to one of six positives in the unvaccinated unchallenged normal control group . Both BCG + HKML groups were significantly protected from clinical leprosy . Six of 10 in each of the two BCG + HKML groups significantly converted to Mitsuda positivity within 2 weeks of boosting compared to zero of six in the normal control group . The sizes of the Mitsuda responses were larger in the LD group than the HD HKML vaccinated/boosted group, suggesting suppression by vaccination with higher doses of HKML in combination with BCG . Fernandez responses were negative in normal RM as well as in the unvaccinated, ML-challenged group and the BCG-vaccinated, ML-challenged group at 37 or 46 months after ML inoculation, although the BCG-vaccinated group was significantly protected from leprosy and the unvaccinated group was not . In contrast, at 37 months the Fernandez reaction was positive in the BCG plus LD and the BCG plus HD HKML-vaccinated groups, both of which were significantly protected from clinical leprosy . By 46 months, the Fernandez responses were below significance in all groups . Thus, Fernandez reactivity is not a reliable correlate to protection from experimental leprosy in RM . Mitsuda responses became strongly positive in all four ML-challenged groups by 37 months and remained strongly positive at 46 months after ML inoculation, suggesting that strong Mitsuda reactivity reflects responses to living ML . BCG or BCG + LD or HD HKML vaccination/boosting of RM produced significant clinical protection from leprosy and there was a good correlation between protection from LL forms of leprosy and positive Mitsuda skin test responses after challenge with live ML . Positive Mitsuda responses were generated in essentially all individuals after challenge with live ML, and this response was primed by prior vaccination/boosting with BCG + HKML as shown by conversion to positivity 2 weeks after boosting . The data show that resistance to clinical leprosy is reflected by Mitsuda responses in ML-exposed RM, similar to results from human studies, and confirm the suitability of RM as a model for leprosy vaccine studies.

Prikl Biokhim Mikrobiol, 2002 Nov-Dec, 38(6), 669 - 76
{Development of the biological preparation enatin with broad-range antimicrobial action}; Romanovskaia TV et al.; Physiological and biochemical traits of epiphytic spore forming bacteria Bacillus pumilis BIM V-263 were examined . The nutrient medium and conditions for submerged cultivation of the strain were selected . The growth dynamics and antagonistic activity during cultivation in a laboratory fermenter ANKUM-2M were studied . The results provide grounds for development of the biological preparation Enatin with broad-range antimicrobial effect . The plant-protective and growth-stimulating effect of Enatin was examined in laboratory and field experiments . The preparation holds promise as means for biological control of crop pathogens.

Prikl Biokhim Mikrobiol, 2002 Nov-Dec, 38(6), 634 - 8
{Biosynthesis of inulinases by Bacillus bacteria}; Zherebtsov NA et al.; Biosynthesis of extracellular inulinase by bacteria Bacillus polymyxa 29, B . polymyxa 722, and B . subtilis 68 was studied . The optimal parameters for the producer growth were as follows: pH 7.0, 33-35 degrees C, the growth duration within 72 h . The presence of mineral reduced or of organic nitrogen was necessary for the enzyme biosynthesis . The inulinase biosynthesis was sharply activated in the presence of carbohydrates . B . polymyxa 722 and B . polymyxa 29 displayed the maximal activity on a starch-containing culture medium, the maximal activity of B . subtilis 68 was found in the presence of sucrose . Inulin did not induce the inulinase biosynthesis by the strains studied . The time course of bacteria growth and of the enzyme biosynthesis was studied.

Antonie Van Leeuwenhoek, 2002 Aug, 81(1-4), 565 - 74
Toward an understanding of microbial communities through analysis of communication networks; Dunn AK et al.; Bacteria receive signals from diverse members of their biotic environment . They sense their own species through the process of quorum sensing, which detects the density of bacterial cells and regulates functions such as bioluminescence, virulence, and competence . Bacteria also respond to the presence of other microorganisms and eukaryotic hosts . Most studies of microbial communication focus on signaling between the microbe and one other organism for empirical simplicity and because few experimental systems offer the opportunity to study communication among various types of organisms . But in the real biological world, microorganisms must carry on multiple molecular conversations simultaneously between diverse organisms, thereby constructing communication networks . We propose that biocontrol of plant disease, the process of suppressing disease through application of a microorganism, offers a model for the study of communication among multiple organisms . Successful biocontrol requires the sending and receiving of signals between the biocontrol agent and the pathogen, plant host, and microbial community surrounding the host . We are using Bacillus cereus, a biocontrol agent, and the organisms it must interact with, to dissect a communication network . This system offers an excellent starting point for study because its members are defined and well studied . An understanding of signaling in the B . cereus biocontrol system may provide a model for network communication among organisms that share a habitat and provide a new angle of analysis for understanding the interconnections that define communities.

Water Res, 2002 Nov, 36(19), 4850 - 60
Studies on the production of B . thuringiensis based biopesticides using wastewater sludge as a raw material; Vidyarthi AS et al.; Growth and delta-endotoxin yield of Bacillus thuringiensis (Bt) subsp kurstaki in tryptic soy yeast extract (TSY) medium, soybean meal based commercial medium and wastewater sludge medium were studied . The viable spores (VS) count in sludge medium was comparable to that obtained in laboratory and commercial media . The entomotoxicity of the fermentation liquid (Bt grown sludge) against Choristoneura fumiferana was comparable to the concentrated commercial Bt formulation available in the market (Foray 48B) . A higher entomotoxicity was observed in a sludge medium than in the TSY or soybean meal media . The secondary and mixed (mixture of primary and secondary) sludges from various wastewater treatment plants were also evaluated for spore formation and entomotoxicity yield . The VS count was higher in a mixed sludge compared to the secondary sludge at a similar sludge solids concentration . Both VS count and entomotoxicity yield was found to be a function of sludge solids concentration in the medium . The optimum value of solids concentration for Bt production was found to be 25 g (-1) (dry weight basis) . Beyond this concentration, a drop in VS count and entomotoxicity yield was observed . A low C:N ratio in the secondary sludge and a high C:N ratio in the mixed sludge resulted in a higher entomotoxicity . The optimum value of C:N ratio in combined sludge for Bt production was found to be 7.9-9.9 . Relationships between entomotoxicity and maximum specific growth as well as with specific sporulation rate were developed.

J Infect Dis, 2002 Dec 15, 186(12), 1797 - 807 Epub 2002 Nov 19.
Tuberculin skin testing and in vitro T cell responses to ESAT-6 and culture filtrate protein 10 after infection with Mycobacterium marinum or M . kansasii; Arend SM et al.; T cell responses to ESAT-6 and culture filtrate protein 10 (CFP-10), antigens expressed by Mycobacterium tuberculosis but not by M . bovis bacille Calmette-Guerin (BCG), were found to discriminate reliably between infection with M . tuberculosis and BCG vaccination . Because the esat-6 and cfp-10 genes occur in M . kansasii and M . marinum, T cell responses to ESAT-6 and CFP-10 were investigated in patients infected with M . kansasii or M . marinum, persons intensively exposed to environmental mycobacteria, and unexposed control subjects . Tuberculin skin tests were performed, and peripheral blood mononuclear cells were cocultured with ESAT-6, CFP-10, peptide mixtures of ESAT-6 and CFP-10, and control antigens . When enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunospot assay (ELISPOT) were used to measure interferon-gamma production, most M . kansasii- or M . marinum-infected patients and several persons exposed to environmental mycobacteria were found to respond to ESAT-6 and/or CFP-10 . ELISA and ELISPOT yielded comparable results, as did whole antigen and peptides (P<.0001) . These results may be relevant for the development of novel assays for diagnosis of tuberculosis.

J Infect Dis, 2002 Dec 15, 186(12), 1733 - 9 Epub 2002 Nov 22.
Molecular epidemiology of Bartonella henselae infection in human immunodeficiency virus-infected patients and their cat contacts, using pulsed-field gel electrophoresis and genotyping; Chang CC et al.; Bartonella henselae causes severe disease in immunocompromised individuals . B . henselae was isolated from 12 human immunodeficiency virus (HIV)-infected individuals with bacillary angiomatosis and/or peliosis hepatis and from their 15 cat contacts . Specific associations between the 2 B . henselae genotypes, individual pulsed-field gel electrophoresis (PFGE) patterns, and different clinical syndromes and pathogenicity were investigated . The role of cat contacts as the source of human infection was also examined . Three of the 4 patients with B . henselae genotype I infection, but none of the 8 patients with genotype II infection, had hepatosplenic vascular proliferative lesions (P=.018) . Four of 5 human-cat pairs had closely-related PFGE patterns and concordant results by 16S rDNA typing, which strongly suggests that human infection was caused by the cat contact . These results corroborate the major role of cats in the transmission of B . henselae to humans and suggest that B . henselae genotypes may induce different pathological features in HIV-infected patients.

J Invertebr Pathol, 2002 Oct, 81(2), 122 - 6
Assessment of the efficacy of Japanese Bacillus thuringiensis isolates against the cigarette beetle, Lasioderma serricorne (Coleoptera: Anobiidae); Tsuchiya S et al.; A total of 2,652 Japanese isolates of Bacillus thuringiensis, belonging to at least 54 H serogroups, were examined for assessment of the toxicity against the cigarette beetle, Lasioderma serricorne (Coleoptera: Anobiidae) . When tested with spore/parasporal inclusion mixtures, strong larvicidal activities were associated with 28 isolates (1.1%) . Serologically, these toxic isolates fell into 4 known H serovars: thuringiensis (9 isolates), kurstaki (2), kenyae (2), and darmstadiensis (15) . Purified parasporal inclusions of the 10 selected isolates exhibited no larvicidal activity, while the supernatants of liquid cultures showed larvicidal and/or growth inhibitory effects . The activities were fully retained after heat treatment at 100 degrees C for 10 min . Overall results suggest that beta-exotoxin (or thuringiensin)-related substances are responsible for the toxicity of the present B . thuringiensis isolates against the cigarette beetle.

FEMS Microbiol Lett, 2002 Nov 19, 217(1), 115 - 9
The enterotoxin T (BcET) from Bacillus cereus can probably not contribute to food poisoning; Choma C et al.; A polymerase chain reaction (PCR) fragment from strain NVH 38 (containing bceT) was cloned, sequenced and expressed in Escherichia coli . This sequence showed 50-60% identity to the original . When this bceT clone was expressed in E . coli no biological activity was found in either supernatants or cell extracts . Cell extracts from the Bacillus cereus strains (NVH 38 and B-4ac) were also negative on Vero cells . Neutralisation of supernatant from B . cereus B-4ac using a monoclonal antibody (reacting with NheB and HblD) abolished the activity . A test for cytotoxic enterotoxins was negative for both cell extracts and supernatants . Our data suggest that BcET either has an unknown type enterotoxic action or non at all.

J Nat Prod, 2002 Nov, 65(11), 1664 - 6
Limnazine, the first bacterial azine derivative from Bacillus sp . GW90a; Asolkar RN et al.; A novel dimeric azine derivative designated as limnazine (1) has been isolated from the culture broth of an aquatic Bacillus sp . strain GW90a . The structure has been elucidated, on the basis of spectral data, as N,N'-bis(2,2,6-trimethylchroman-4-ylidene)azine and was additionally confirmed by synthesis . Limnazine (1) is inactive against algae, fungi, and bacteria.

Parasitol Res, 2002 Dec, 88(12), 1090 - 2 Epub 2002 Jun 28.
Antitrichomonal strains of Bacillus thuringiensis; Kondo S et al.; Parasporal inclusion proteins from a total of 816 Bacillus thuringiensis strains isolated in Japan were examined for antitrichomonal activity against Trichomonas vaginalis . Ten strains of B . thuringiensis inhibited the growth of T . vaginalis in 48 h cultures at 37 degrees C . Moreover, the B622 and B626 strains clearly showed trichomonacidal effects against T . vaginalis . The H antigen serotypes of both strains were identified as H13/29 (pakistani/amagiensis) . The parasporal inclusion protein from both strains consisted of three major polypeptides of 77, 45 and 25 kDa.

Urol Int, 2002, 69(4), 263 - 5
The management of T1G3 bladder cancer; Bogdanovic J et al.; INTRODUCTION: The treatment of T1G3 bladder cancer is still a controversial issue . Nowadays, intravesical bacillus Calmette-Guerin (BCG) instillation is considered to be the treatment of choice for patients with high-grade superficial bladder tumour after transurethral resection of all visible tumour . The aim of this retrospective study was to determine the effects and results of this approach, recurrence and progression rates in patients with T1G3 superficial bladder tumours . MATERIALS AND METHODS: 43 patients (28 male, 15 female; mean age 65.5 years, range 21-82) with T1G3 TCC (transitional cell carcinoma) bladder tumour underwent transurethral resection and subsequent intravesical BCG according to Morales protocol, in the period 1993-1998 at our institution . The mean follow-up period was 52.5 (range 30-96) months . RESULTS: After one or more initial courses of therapy, 33 patients were disease-free . Twelve patients (27.90%) had recurrent tumour after a median of 7 (range 3-46) months . After a second course of BCG treatment, 6 patients had no evidence of disease, 3 patients had progression and 3 had recurrence . Progression occurred in 7 (16.27%) patients after a median of 19 (range 3-43) months . Five patients underwent radical cystectomy and the remaining 2 underwent bladder-preserving therapies . Two patients died of TCC and 3 due to disease-unrelated conditions . CONCLUSION: Intravesical BCG instillation can be recommended as treatment modality for responders with T1G3 TCC bladder tumour . The benefit of the second course of intravesical BCG therapy has to be confirmed in further investigations .

Vaccine, 2002 Nov 22, 21(1-2), 7 - 14
A novel TB vaccine; towards a strategy based on our understanding of BCG failure; Agger EM et al.; The protection afforded by the currently available tuberculosis vaccine, bacillus Calmette-Guerin (BCG) is insufficient and new vaccine strategies are urgently needed . Progress in our understanding of the immunological deficits of BCG combined with novel knowledge on genetics of mycobacteria has paved the way for promising new vaccine strategies . These include recombinant modified BCG vaccines, attenuated strains of Mycobacterium tuberculosis, and various non-live candidates such as DNA and subunit vaccines . Decisive for transforming technical progress into a novel tuberculosis (TB) vaccine strategy is the recent advance in our understanding of the failure of BCG in the third world and the interaction between this vaccine and environmental mycobacteria.

AIDS, 2002 Nov 22, 16(17), 2285 - 93
Rapid detection of active and latent tuberculosis infection in HIV-positive individuals by enumeration of Mycobacterium tuberculosis-specific T cells; Chapman AL et al.; OBJECTIVES: An accurate test for Mycobacterium tuberculosis infection is urgently needed . The tuberculin skin test (TST) lacks sensitivity, particularly in HIV-infected individuals, and has poor specificity because of antigenic cross-reactivity with Bacillus Calmette-Guerin (BCG) vaccination . ESAT-6 and CFP-10 are antigens expressed in Mycobacterium tuberculosis, but not in Mycobacterium bovis BCG and most environmental mycobacteria . We investigated whether T cells specific for these antigens could serve as accurate markers of M . tuberculosis infection in an area of high tuberculosis and HIV prevalence . METHODS: Using the rapid ex-vivo enzyme-linked immunospot (ELISPOT) assay for IFN-gamma, we enumerated T cells specific for ESAT-6, CFP-10 and purified protein derivative (PPD) in blood samples from 50 Zambian tuberculosis patients, 75 healthy Zambian adults, and 40 healthy UK residents . TSTs were performed in 49 healthy Zambian adults . RESULTS: All (100%; n = 11) and 90% (n = 39) of HIV-negative and HIV-positive tuberculosis patients, respectively, had detectable ESAT-6- or CFP-10-specific T cells . The ESAT-6/CFP-10-based ELISPOT assay was positive in 37 out of 54 HIV-negative healthy Zambians, suggesting a 69% prevalence of latent M . tuberculosis infection . Fewer HIV-positive Zambians possessed ESAT-6/CFP-10-specific T cells, but the impact of HIV infection was less on this assay than on the PPD-based ELISPOT or TST . CONCLUSION: The ESAT-6/CFP-10-based ELISPOT assay detects active tuberculosis in HIV-positive individuals with high sensitivity . It is more specific, and possibly more sensitive, than PPD-based methods of detecting latent M . tuberculosis infection, and may potentially improve the targeting of isoniazid preventative therapy to HIV-positive individuals with latent tuberculosis infection .

Proc Natl Acad Sci U S A, 2002 Dec 10, 99(25), 16006 - 11 Epub 2002 Nov 19.
The 2B domain of the Escherichia coli Rep protein is not required for DNA helicase activity; Cheng W et al.; The Escherichia coli Rep protein is a 3' to 5' SF1 DNA helicase required for replication of bacteriophage phiX174 in E . coli, and is structurally homologous to the E . coli UvrD helicase and the Bacillus stearothermophilus PcrA helicase . Previous crystallographic studies of Rep protein bound to single-stranded DNA revealed that it can undergo a large conformational change consisting of an approximately 130 degrees rotation of its 2B subdomain about a hinge region connected to the 2A subdomain . Based on crystallographic studies of PcrA, its 2B subdomain has been proposed to form part of its duplex DNA binding site and to play a role in duplex destabilization . To test the role of the 2B subdomain in Rep-catalyzed duplex DNA unwinding, we have deleted its 2B subdomain, replacing it with three glycines, to form the RepDelta2B protein . This RepDelta2B protein can support phiX174 replication in a rep(-) E . coli strain, although the growth rate of E . coli containing the repDelta2B gene is approximately 1.5-fold slower than with the wild-type rep gene . Pre-steady-state, single-turnover DNA unwinding kinetics experiments show that purified RepDelta2B protein has DNA helicase activity in vitro and unwinds an 18-bp DNA duplex with rates at least as fast as wild-type Rep, and with higher extents of unwinding and higher affinity for the DNA substrate . These studies show that the 2B domain of Rep is not required for DNA helicase activity in vivo or in vitro, and that it does not facilitate DNA unwinding in vitro.

Curr Gastroenterol Rep, 2002 Dec, 4(6), 448 - 54
Helicobacter pylori-Associated Diseases; Sanders MK et al.; Since the initial report 20 years ago by Marshall and Warren of an unidentified curved bacillus located on the gastric epithelium of patients with chronic active gastritis, the discovery of Helicobacter pylori and its association with a number of gastrointestinal diseases has revolutionized the field of gastroenterology . Although the association of H . pylori infection with peptic ulcer disease, chronic gastritis, mucosa-associated lymphoid tissue lymphoma, and gastric adenocarcinoma has been well documented over the past two decades, other areas remain less clear, including the role of H . pylori in gastropathy associated with nonsteroidal anti-inflammatory drugs, gastroesophageal reflux disease, and both uninvestigated and nonulcer dyspepsia . Although these areas still remain somewhat controversial, recent reports further clarify the role of H . pylori in these conditions . A review of the recent literature regarding H . pylori-associated diseases is presented along with recommendations for diagnosis and treatment of H . pylori infection.

Biochem J, 2003 Mar 15, 370(Pt 3), 971 - 8
Recombinantly expressed isoenzymic aminopeptidases from Helicoverpa armigera (American cotton bollworm) midgut display differential interaction with closely related Bacillus thuringiensis insecticidal proteins; Rajagopal R et al.; Several investigators have independently identified membrane-associated aminopeptidases in the midgut of insect larvae as the initial interacting ligand to the insecticidal crystal proteins of Bacillus thuringiensis . Though several isoenzymes of aminopeptidases have been identified from the midgut of an insect and their corresponding cDNA cloned, only one of the isoform has been expressed heterologously and studied for its binding to Cry toxins . Here we report the cloning and expression of two aminopeptidases N from Helicoverpa armigera (American cotton bollworm) (HaAPNs) . The full-length cDNA of H . armigera APN1 (haapn1) is 3205 bp in size and encodes a 1000-amino-acid protein, while H . armigera APN2 (haapn2) is 3116 bp in size and corresponds to a 1012-amino-acid protein . Structurally these proteins show sequence similarity to other insect aminopeptidases and possess characteristic aminopeptidase motifs . Both the genes have been expressed in Trichoplusia ni (cabbage looper) cells using a baculovirus expression vector . The expressed aminopeptidases are membrane-associated, catalytically active and glycosylated . Ligand-blot analysis of both these aminopeptidases with bioactive Cry1Aa, Cry1Ab and Cry1Ac proteins displayed differential interaction . All the three toxins bound to HaAPN1, whereas only Cry1Ac interacted with HaAPN2 . This is the first report demonstrating differential Cry-toxin-binding abilities of two different aminopeptidases from a susceptible insect.

Infect Immun, 2002 Dec, 70(12), 6996 - 7003
Identification of novel Mycobacterium tuberculosis antigens with potential as diagnostic reagents or subunit vaccine candidates by comparative genomics; Cockle PJ et al.; An independent review for the British government has concluded that the development of a cattle vaccine against Mycobacterium bovis holds the best long-term prospects for tuberculosis control in British herds . The development of complementary diagnostic tests to differentiate between vaccinated and infected animals is necessary to allow the continuation of test-and-slaughter-based control policies alongside vaccination . Vaccination with M . bovis bacillus Calmette-Guerin (BCG), the only available vaccine, results in tuberculin purified protein derivative sensitivity and has shown varying vaccine efficacies in cattle . Thus, identification of more-specific reagents to distinguish between vaccination and infection, as well as the identification of subunit vaccine candidates for improved tuberculosis vaccines, is a research priority . In the present study, we applied comparative genomics to identify M . bovis-Mycobacterium tuberculosis antigens whose genes had been deleted in BCG Pasteur . In total, 13 open reading frames (ORFs) from the RD1, RD2, and RD14 regions of the M . tuberculosis genome were selected . Pools of overlapping peptides spanning these ORFs were tested in M . bovis-infected (n = 22), BCG-vaccinated (n = 6), and unvaccinated (n = 10) control cattle . All were recognized in infected cattle, with responder frequencies varying between 16 and 86% . In particular, eight highly immunogenic antigens were identified whose potentials as diagnostic reagents or as subunit vaccines warrant further study (Rv1983, Rv1986, Rv3872, Rv3873, Rv3878, Rv3879c, Rv1979c, and Rv1769).

J Chromatogr A, 2002 Oct 11, 973(1-2), 197 - 202
Determination of poly-beta-hydroxybutyric acid in Bacillus thuringiensis by capillary zone electrophoresis with indirect ultraviolet absorbance detection; He J et al.; A new capillary electrophoresis method for determining poly-beta-hydroxybutyric acid (PHB) in Bacillus thuringiensis was established . Poly-beta-hydroxybutyric acid in samples was hydrolyzed by sulphuric acid and neutralized by Ba(OH)2 . The content of produced beta-hydroxybutyrate was then determined by capillary zone electrophoresis (CZE) with indirect UV detection at 254 nm . With 5 mM p-hydroxybenzoate and 0.5 mM tetradecyltrimethylammonium bromide (TTAB) at pH 8.0 as carrier electrolyte, beta-hydroxybutyrate can be determined within 6 min . Standard regression equation was made by beta-hydroxybutyrate, and the linear range was 2-1000 microg/ml . The relative standard deviations (RSDs) for migration time and peak area are both less than 1.0% . The detection limit for beta-hydroxybutyrate was 0.2 microg/ml, which is two to three orders of magnitude lower than that of the gas chromatography (GC) method . The capillary electrophoresis method was successfully applied to determine poly-beta-hydroxybutyric acid in fermentation broth and single colony . The added standard recovery was 96%.

Appl Microbiol Biotechnol, 2002 Nov, 60(3), 313 - 9 Epub 2002 Oct 05.
An immobilised catalase peroxidase from the alkalothermophilic Bacillus SF for the treatment of textile-bleaching effluents; Fruhwirth GO et al.; A catalase peroxidase (CP) from the newly isolated Bacillus SF was used to treat textile-bleaching effluents . The enzyme was stable at high pH values and temperatures, but was more sensitive to deactivation by hydrogen peroxide than monofunctional catalases . Based on the Michaelis-Menten kinetics of the CP, a model was developed to describe its deactivation characteristics . The enzyme was immobilised on various alumina-based carrier materials with different shapes and the specific activity increased with the porosity of the carrier . The shape of the carrier had an important influence on the release of oxygen formed during the catalase reaction from the packed-bed reactor and Novalox saddles were found to be the most suitable shape . Bleaching effluent was treated in a horizontal packed-bed reactor containing 10 kg of the immobilised CP at a textile-finishing company . The treated liquid (500 l) was reused within the company for dyeing fabrics with various dyes, resulting in acceptable colour differences of below Delta E*=1.0 for all dyes.

Drug Ther Bull, 2002 Oct, 40(10), 78 - 80
BCG, TB and the UK; Bacillus macerans cyclomaltodextrin glucanotransferase transglycosylation reactions with different molar ratios of D-glucose and cyclomaltohexaose; Laboratory of Carbohydrate Chemistry and Enzymology, 4252 Molecular Biology Bldg, Iowa State University, Ames, IA 50011, USAIt was found that Bacillus macerans cyclomaltodextrin glucanotransferase (CGTase) reacts with cyclomaltohexaose (alpha-cyclodextrin, alpha-CD) to give a series of cyclomaltooligosaccharides (cyclomaltodextrins, CDs), having seven to more than 20 D-glucose residues and maltooligosaccharides (maltodextrins, MDs) from G5 to G12+ . When D-glucose (Glc) was added to the alpha-CD at very low molar ratios (1:100) of Glc to alpha-CD, the predominant products (95%) were CDs, some of which were macrocyclic MDs with 20-60 D-glucose residues, along with MDs that also had high molecular weights, containing 10-75 D-glucose residues and gave a blue iodine-iodide color . As the molar ratio of Glc to alpha-CD was increased, the amount of CDs progressively decreased and MDs proportionately increased in the range of G2-G12 . At 25 mM alpha-CD and Glc to alpha-CD molar ratio of 1:1, a 75% yield of MDs, G1-G12, each in approximately equal amounts, was obtained; and at 20 mM and a 5:1 ratio, a 97% yield of MDs, G2-G9, was obtained but in unequal amounts . At higher ratios (10:1), the CDs completely disappeared, and at very high ratios (50:1 to 100:1) only low-molecular-weight MDs, G2-G4, were formed .

Anal Chem, 2002 Nov 1, 74(21), 5681 - 7
A microarray immunoassay for simultaneous detection of proteins and bacteria; Delehanty JB et al.; We report the development and characterization of an antibody microarray biosensor for the rapid detection of both protein and bacterial analytes under flow conditions . Using a noncontact microarray printer, biotinylated capture antibodies were immobilized at discrete locations on the surface of an avidin-coated glass microscope slide . Preservation of capture antibody function during the deposition process was accomplished with the use of a low-salt buffer containing sucrose and bovine serum albumin . The slide was fitted with a six-channel flow module that conducted analyte-containing solutions over the array of capture antibody microspots . Detection of bound analyte was subsequently achieved using fluorescent tracer antibodies . The pattern of fluorescent complexes was interrogated using a scanning confocal microscope equipped with a 635-nm laser . This microarray system was employed to detect protein and bacterial analytes both individually and in samples containing mixtures of analytes . Assays were completed in 15 min, and detection of cholera toxin, staphylococcal enterotoxin B, ricin, and Bacillus globigii was demonstrated at levels as low as 8 ng/mL, 4 ng/mL, 10 ng/mL, and 6.2 x 10(4) cfu/mL, respectively . The assays presented here are very fast, as compared to previously published methods for measuring antibody-antigen interactions using microarrays (minutes versus hours).

Curr Microbiol, 2003 Jan, 46(1), 43 - 6
Mannose-specific lectin activity of parasporal proteins from a lepidoptera-specific Bacillus thuringiensis strain; Wasano N et al.; Lectin activity, agglutinating sheep erythrocytes, was associated with parasporal inclusion proteins from a Lepidoptera-specific isolate of Bacillus thuringiensis serovar galleriae (H5ab) . The activity was generated when parasporal inclusions were solubilized in an alkaline condition . Proteolytic processing was not required for generation of the lectin activity; the activity level was not affected by the presence/absence of the three proteases (trypsin, chymotrypsin, and proteinase K) . SDS-PAGE analysis revealed that (1) alkali-solubilized parasporal inclusion proteins consisted of two major components of 130 kDa and 65 kDa, and (2) proteinase K treatment of alkali-solubilized proteins yielded a single major protein of 60 kDa . Lectin activity of our isolate was strongly inhibited by preincubation with D-mannose, but not with the six other monosaccharides: D-galactose, D-glucose, L-fucose, N-acetyl- D-glucosamine, N-acetyl- D-galactosamine, and N-acetylneuraminic acid . In contrast, D-mannose did not inhibit the in vivo larvicidal activity of the proteins against the silkworm, Bombyx mori.

Curr Microbiol, 2003 Jan, 46(1), 33 - 8
Cloning and characterization of two novel crystal protein genes from a Bacillus thuringiensis serovar dakota strain; Kim HS et al.; Two genes encoding the 32- and 40-kDa polypeptides of Bacillus thuringiensis strain 90-F-45-14 crystals were cloned, expressed in an acrystalliferous B . thuringiensis strain, and sequenced . The polypeptides had deduced molecular weights of 30,319 and 33,885, respectively . The amino acid sequence of the 32-kDa protein was 37.7% identical to the known sequence of a non-insecticidal parasporal protein in B . thuringiensis serovar thompsoni crystals . The sequence of the cloned 40-kDa protein was 37.0% and 22.3% identical to that of the existing Cry protein classes, Cry15Aa1 and Cry23Aa1, respectively . Thus, this protein constitutes a novel protein class, Cry33Aa1 . The open reading frames of the two genes were located on the predominant plasmid of 17,629 bp (=11,752 MDa) in the same orientation, and they were separated by the sequence of 32 nucleotides . The two proteins are likely produced simultaneously from a single transcript to form spherical crystals.

Urology, 2002 Nov, 60(5), 816 - 21
Association of cyclooxygenase-2 expression with prognosis of stage T1 grade 3 bladder cancer; Kim SI et al.; OBJECTIVES: To determine whether the expression of cyclooxygenase-2 (COX-2) has prognostic significance in Stage T1G3 transitional cell carcinoma of the bladder, the most unfavorable subgroup in terms of recurrence and disease progression . METHODS: Thirty-seven consecutive patients with initial T1G3 transitional cell carcinoma, who had undergone complete transurethral resection, followed by 6 weeks of intravesical instillation of bacille Calmette-Guerin (BCG), and with at least 1 year of follow-up, were enrolled in the study . Paraffin-embedded cancer tissue samples were immunohistochemically stained for COX-2, and possible correlations with clinicopathologic features, such as age, shape and multiplicity of tumor, recurrence, and progression were examined . RESULTS: The median follow-up was 27 months (range 12 to 67) . Sixteen patients (43.2%) experienced recurrence and 6 (16.2%) had progression defined as muscle invasion . Of 37 specimens, 16 (43.2%) stained positive for COX-2, defined as 5% or greater of positively stained cancer cells . COX-2 expression was statistically significant in predicting both recurrence (P = 0.0493) and disease progression (P = 0.0272) . Patient age and the shape and multiplicity of tumors were not significantly predictive of recurrence or progression . CONCLUSIONS: In a pathologically homogeneous group of T1G3 transitional cell carcinoma of the bladder, the expression of COX-2 correlated with recurrence and progression . Thus, patients with COX-2 positive superficial bladder cancer may need to be followed up more vigorously . Additional studies on the mechanistic implications of COX-2 with respect to recurrence and progression and the possible application of a COX-2 inhibitor to prevent recurrence and progression of superficial bladder cancer are warranted.

J Agric Food Chem, 2002 Nov 20, 50(24), 7076 - 8
Rapid degradation of the Cry1F insecticidal crystal protein in soil; Herman RA et al.; The gene for the core Cry1F insecticidal crystal protein (ICP) from Bacillus thuringiensis Berliner (Bt) has been incorporated into the genome of maize plants, Zea mays L . Plants expressing this ICP are protected from attack by various Lepidopteran pests including the European corn borer, Ostrinia nubilalis (Hubner) . The stability of the Cry1F ICP in soil was assessed in a laboratory study designed to determine the persistence of the active protein residue in soil over time, using insect bioassay as the analytical quantification method . The GI(50) (concentration estimated to inhibit growth by 50%) rose at each consecutive incubation interval, indicating a consistent decline in Cry1F activity over time . The residue data were poorly described by a first-order model when fit to either the full data or a truncated data set where the last interval (28 days) was excluded . Data were well described by a shift-log model, and this model predicted DT(50) (time until 50% decay) and DT(90) (time until 90% decay) values of 0.6 and 6.9 days, respectively . This rapid degradation rate was consistent with other Bt proteins evaluated in our laboratory.

Am J Clin Pathol, 2002 Nov, 118(5), 742 - 8
Diagnosis of Wihipple disease by immunohistochemical analysis: a sensitive and specific method for the detection of Tropheryma whipplei (the Whipple bacillus) in paraffin-embedded tissue; Baisden BL et al.; Whipple disease is a rare infection characterized clinically by diarrhea, fever, weight loss, arthralgia, malabsorption, and other systemic manifestations . The etiologic agent, Tropheryma whipplei, has been cultured only rarely . By using a polyclonal rabbit antibody produced against a cultured strain of T whipplei, tissue sections from 18 patients with Whipple disease were studied . Specimens from patients with histologic mimics and other infections served as control specimens . Immunostaining was identified in all 18 patients . Granular immunostaining was observed similar to that in periodic acid-Schiff (PAS) stains . In 2 patients, immunostaining was identified in specimens negative by H&E and PAS stains . In 4 patients studied before and after antibiotic therapy, immunostaining was retained but diminished in intensity and quantity . Immunostaining was not identified in any control specimen . Immunohistochemical analysis is a sensitive and specific method for the diagnosis of Whipple disease in paraffin-embedded tissue and may provide new opportunities to investigate the pathogenesis of the infection.

Methods Find Exp Clin Pharmacol, 2002 Sep, 24(7), 431 - 55
Gateways to Clinical Trials; Bayes M et al.; Gateways to Clinical Trials is a guide to the most recent clinical trials in current literature and congresses . The data in the following tables has been retrieved from the Clinical Studies knowledge area of Prous Science Integrity, the drug discovery and development portal, This issue focuses on the following selection of drugs: Adalimumab, aeroDose insulin inhaler, agomelatine, alendronic acid sodium salt, aliskiren fumarate, alteplase, amlodipine, aspirin, atazanavir; Bacillus Calmette-Guerin, basiliximab, BQ-788, bupropion hydrochloride; Cabergoline, caffeine citrate, carbamazepine, carvedilol, celecoxib, cyclosporine, clopidogrel hydrogensulfate, colestyramine; Dexamethasone, diclofenac sodium, digoxin, dipyridamole, docetaxel, dutasteride; Eletriptan, enfuvirtidie, eplerenone, ergotamine tartrate, esomeprazole magnesium, estramustine phosphate sodium; Finasteride, fluticasone propionate, fosinopril sodium; Ganciclovir, GBE-761-ONC, glatiramer acetate, gliclazide, granulocyte-CSF; Heparin sodium, human isophane insulin (pyr), Hydrochlorothiazide; Ibuprofen, inhaled insulin, interferon alfa, interferon beta-1a; Laminvudine, lansoprazole, lisinopril, lonafarnib, losartan potassium, lumiracoxib; MAb G250, meloxicam methotrexate, methylprednisolone aceponate, mitomycin, mycophenolate mofetil; Naproxen sodium, natalizumab, nelfinavir mesilate, nemifitide ditriflutate, nimesulide; Omalizumab, omapatrilat, omeprazole, oxybutynin chloride; Pantoprazole sodium, paracetamol, paroxetine, pentoxifylline, pergolide mesylate, permixon, phVEGF-A165, pramipexole hydrochloride, prasterone, prednisone, probucol, propiverine hydrochloride; Rabeprazole sodium, resiniferatoxin, risedronate sodium, risperidone, rofecoxib rosiglitazone maleate, ruboxistaurin mesilate hydrate; Selegiline transdermal system, sertraline, sildenafil citrate, streptokinase; Tadalafil, tamsulosin hydrochloride, technosphere/Insulin, tegaserod maleate, tenofovir disoproxil fumarate, testosterone heptanoate, testosterone undecanoate, tipifarnib, tolterodine tartrate, topiramate, troglitazone; Ursodeoxycholic acid; Valdecoxib, valsartan, vardenafil, venlafaxine hydrochloride, VX-745.

Biochemistry, 2002 Nov 19, 41(46), 13717 - 24
Proteolytic cleavage of the developmentally important cadherin BT-R1 in the midgut epithelium of Manduca sexta; Candas M et al.; BT-R1 (M(r) = 210 kDa) represents a new type of insect cadherin that is expressed specifically in the midgut epithelium during growth and development of Manduca sexta larvae . It also is a target receptor for the Cry1A toxins of the entomopathogenic bacterium Bacillus thuringiensis . Expression of BT-R1, which varies during larval development, correlates with the abundance of the protein and with the differential cleavage of the molecule at each developmental stage . The cleavage of BT-R1 is calcium dependent, and consequently, Ca2+ directly influences the structural integrity of BT-R1 . Indeed, removal of calcium ions by chelating agents promotes cleavage of the BT-R1 ectodomain, resulting in formation of fragments that are similar to those observed during larval development . Partial purification of proteins from brush border membrane vesicles (BBMVs) by gel filtration chromatography hinders the cleavage of BT-R1 in the presence of EDTA and EGTA, indicating that there is specific proteolytic activity associated with the BBMV . This specific proteolytic cleavage of BT-R1 not only alters the integrity of BT-R1 but it most likely is implicated in cell adhesion events during differentiation and development of M . sexta midgut epithelium . We propose a model for calcium-dependent protection of BT-R1 as well as a cleavage pattern that may modulate the molecular interactions and adhesive properties of its ectodomain . Molecular characterization of such a protection mechanism should lead to a better understanding of how the function of specific cadherins is modulated during tissue differentiation and insect development.

J Bacteriol, 2002 Dec, 184(23), 6602 - 14
Arginine operator binding by heterologous and chimeric ArgR repressors from Escherichia coli and Bacillus stearothermophilus; Ghochikyan A et al.; Bacillus stearothermophilus ArgR binds efficiently to the Escherichia coli carAB operator, whereas the E . coli repressor binds very poorly to the argCo operator of B . stearothermophilus . In order to elucidate this contradictory behavior between ArgRs, we constructed chimeric proteins by swapping N-terminal DNA-binding and C-terminal oligomerization domains or by exchanging the linker peptide . Chimeras carrying the E . coli DNA-binding domain and the B . stearothermophilus oligomerization domain showed sequence-nonspecific rather than sequence-specific interactions with arg operators . Chimeras carrying the B . stearothermophilus DNA-binding domain and E . coli oligomerization domain exhibited a high DNA-binding affinity for the B . stearothermophilus argCo and E . coli carAB operators and repressed the reporter-gene transcription from the B . stearothermophilus PargCo control region in vitro; arginine had no effect on, and indeed even decreased, their DNA-binding affinity . With the protein array method, we showed that the wild-type B . stearothermophilus ArgR and derivatives of it containing only the exchanged linker from E . coli ArgR or carrying the B . stearothermophilus DNA-binding domain along with the linker and the alpha4 regions were able to bind argCo containing the single Arg box . This binding was weaker than binding to the two-box operator but was no longer arginine dependent . Several lines of observations indicate that the alpha4 helix in the oligomerization domain and the linker peptide can contribute to the recognition of single or double Arg boxes and therefore to the operator DNA-binding specificity in similar but not identical ArgR repressors from two distant bacteria.

Chest, 2002 Nov, 122(5), 1609 - 14
Factors associated with the treatment of latent tuberculosis infection among health-care workers at a midwestern teaching hospital; Shukla SJ et al.; STUDY OBJECTIVE: To assess factors associated with initiating therapy and compliance with treatment for latent tuberculosis infection among health-care workers with positive tuberculin skin test results . DESIGN: Prospective cohort study . SETTING: An urban midwestern teaching hospital in St . Louis, MO . Study population: Health-care workers with positive tuberculin skin test results . MEASUREMENTS: (1) Rates of initiating therapy for latent tuberculosis infection among all health-care workers with positive tuberculin skin test results, and (2) compliance rates with therapy for latent tuberculosis infection among health-care workers with recent tuberculin skin test conversion . RESULTS: A total of 440 tuberculin skin test-positive health-care workers were evaluated from January 1, 1994, to May 1, 2000 . Of those evaluated, 1 health-care worker had presumed active tuberculosis, 1 had no record of being evaluated, 1 had missing records, and 33 were not recommended isoniazid therapy, leaving 404 workers for analysis . Overall, 396 of 404 health-care workers (98%) with positive tuberculin skin test results initiated isoniazid therapy . In univariate analysis, bacille Calmette-Guerin (BCG) vaccination (p = 0.02) and foreign birth (p = 0.03) were significantly associated with not initiating isoniazid therapy . Compliance data were available for 388 of 404 health-care workers (96%) . Of these, 318 of 388 health-care workers (82%) were compliant with 6 months of therapy . BCG vaccination (odds ratio {OR}, 3.5; 95% confidence interval {CI}, 1.8 to 7.1) and symptoms while receiving therapy (OR, 4.5; 95% CI, 2.0 to 10.1) were significantly associated with noncompliance in multivariate analysis . Among new converters, Asian race (p = 0.006), foreign birth (p = 0.01), BCG vaccination (p = 0.006), and symptoms while receiving therapy (p < 0.001) were significantly associated with noncompliance in univariate analysis . CONCLUSION: This hospital had a high rate of initiating isoniazid therapy for tuberculosis infection among their health-care workers, and a high rate of compliance with therapy . These rates of initiation and completion of isoniazid therapy were much higher than those previously reported in the literature . This may be largely due to a focused program, which includes active follow-up of health-care workers with positive tuberculin skin test results, consisting of physician counseling and monthly phone consultations by nurses, along with free services and medications provided on-site.

Di Yi Jun Yi Da Xue Xue Bao, 2001, 21(12), 906 - 907
Effects of gentiopicrooside injections on experimental hepatic injury; Tong L et al.; OBJECTIVE: To investigate the protective and jaundice-relieving effects of gentiopicrooside (GPS) injections in mouse and rat models of chemical-induced and immune-mediated hepatic injury . METHODS: Mouse models of chemical-induced liver injury were established by CCl4 injections into the abdominal cavity, mouse models of immune-mediated liver damage by bacillus Calmette-Guerin (BCG) and lipopolysaccharide (LPS) and rat models of jaundice by oral alpha-naphthyliso-thiocyanate (ANIT) . Treatment with GPS injections was administered and both of enzyme activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the Serum were measured in the models . The serum level of total bilirubin was determined in the jaundice models . RESULTS: Compared with those of the untreated models, the enzyme activities of ALT and AST were significantly reduced in groups with a 10 day GPS treatment (P<0.001, P<0.05) . Higher dosage of GPS showed more conspicuous effects in relieving the jaundice . CONCLUSION: GPS can be administered as the antagonist against CC14-induced liver injury and offers protection against immune-mediated liver damage.

Am J Gastroenterol, 2002 Nov, 97(11), 2756 - 62
Helicobacter pylori antigen stool test and 13C-urea breath test in patients after eradication treatments; Perri F et al.; OBJECTIVES: Current guidelines recommend either the urea breath test (UBT) or the Helicobacter pylori antigen stool test (HpSA) for monitoring H . pylori infection . The aim of this study was to evaluate the agreement between the two tests in patients after treatment . METHODS: After eradication treatments, patients were tested with both UBT and HpSA . Cut-off values (delta value over baseline at 30') for UBT were positive (> or = 5 per thousand), indeterminate (3.01-4.99 per thousand), and negative (< or = 3 per thousand) . Cut-off values (absorbance at 450 nm) for HpSA test were positive (> or = 0.160), indeterminate (0.159-0.140), and negative (< 0.140) . Patients with either discordant or indeterminate tests underwent repeat endoscopy with multiple gastric biopsies for rapid urease test (RUT), culture, histology, and immunohistochemistry to detect H . pylori and to assess the ratio between coccoid and bacillary forms . RESULTS: A total of 458 patients were studied . Of these, 422 (92.2%) had concordant tests, three (0.6%) indeterminate tests (one on UBT and two on HpSA), and 33 (7.2%) discordant tests . A total of 28 patients (25 with discordant and three with indeterminate tests) underwent endoscopy . The HpSA was inaccurate in 24 cases (18 false negative, four false positive, and two indeterminate results), whereas the UBT was inaccurate in four cases (two false positive, one false negative, and one indeterminate results) . Biopsy-based tests showed no bacillary or coccoid forms in all five endoscoped patients who were negative on UBT and positive on HpSA, but in one in whom the ratio between coccoid and bacillary forms was 3:1 in the antrum and corpus . CONCLUSIONS: UBT and HpSA test give discordant or indeterminate results in nearly 8% of patients after treatment . The HpSA test is less accurate than the UBT . Coccoid forms do not cause false positive HpSA results.

Meded Rijksuniv Gent Fak Landbouwkd Toegep Biol Wet, 2001, 66(2a), 425 - 35
First steps to improve cotton crop management in Thailand; Renou A et al.; Thai cotton growers yearly face serious pest problems consisting of heavy infestation associated with resistance to pesticides leading to expensive and harmful pest control practices . Thanks to a multi discipline research work carried out from 1991 to 1996, a decision-oriented model for cotton crop management was developed and compared to farmers' practices during the 1988 cotton growing season 1998 in Klang Hat district, Thailand . This model, based on plant monitoring, involved a new cotton cultivar (DORA 11 as a replacement for Sri Samrong 60), seed treatment with imidacloprid (at 3.5 g of a.i./kg of seeds), a threshold-based spray of mepiquat chloride, a cotton growth regulator (Pix at 1.5 l/ha) and a set of rules to selectively control with chemical and biological insecticides the major pests the American bollworm (Helicoverpa armigera Hubner) and the cotton leafhopper (Amrasca biguttula biguttula Ishida) . This package was more friendly towards environment than farmers' practices since seed treatment does not harm beneficials at the beginning of cotton growing season while 1 and 5 sprays were saved against respectively the American bollworm and sucking pests and more than 50% of the sprays directed against American bollworms involved biological insecticides (Bacillus thuringiensis toxins potentialized by a low dosage of endosulfan) . Threshold-based sprays against the main pests, the use of a cotton growth regulator, earliness of cotton cultivar and seed treatment were accountable for savings in pesticide sprays . Involving a biological insecticide and endosulfan at the beginning of cotton growing season and using the same quantity of active ingredient at each spray during the cotton growing season also improved the management of pyrethroid resistance in American bollworm populations was also . There was no significant difference between yields inside farmers' fields and innovation plots . However, thanks to cheaper pest control practices economic results were better . Lastly few improvements of this package are proposed.

Meded Rijksuniv Gent Fak Landbouwkd Toegep Biol Wet, 2001, 66(2a), 259 - 67
Pest insect control in organically-produced crops of field vegetables; Collier RH et al.; In the UK, the demand for organic vegetable and salad crops is increasing, mainly as a result of the requirements of the multiple retailers . However, approximately 85% of the organic fruit and vegetable produce sold in the UK is imported . A major constraint to growing field vegetable crops, and particularly organically-produced crops, is the reduction in crop yield and quality caused by pest insects . This paper will consider the control techniques currently available to organic growers and other techniques that may become available in the future . Growing plant varieties with complete or even partial resistance to pest insects can be an effective way of reducing crop damage . There are already varieties of carrot, with resistance to carrot fly, and lettuce, with resistance to certain pest aphid species, which are available commercially . Cultural techniques to exclude, deter or avoid pest insects are also being used by some organic growers . Although isolating new crops from sources of infestation can be a highly effective control strategy, many organic growers cannot use it, as the land converted for organic production is still limited . Various crop covers can be used to prevent pest insects from damaging field crops, but to be effective such covers have to be in place before the pests enter the crop . Several researchers have tried to develop techniques to prevent pest insects from finding their host-plants . No technique involving semiochemicals has been sufficiently successful to be used in field vegetable production in the UK . Other studies have shown that the numbers of pest insects found on crop plants are reduced considerably when the crop is allowed to become weedy, is intercropped with another plant species, or is undersown with a living mulch . Hence, work is now needed to select background plant species that will both reduce pest insect numbers and cause the least reduction in yield to the harvested crop plants . There is also a need to obtain a better understanding of "companion planting", a practice used frequently by organic growers . To date, microbial control is the only biological technique that has been used successfully in field vegetable crops in the UK . However, only the toxicant produced by one microbial agent, the bacterium Bacillus thuringiensis, has so far been registered for use . The use of bacteria, fungi and viruses to control pests of field vegetable crops certainly has possibilities . However, in many cases there are still problems to be overcome to select pathogens that are compatible with, or can still be effective in, the wide fluctuations in temperature, humidity and soil moisture that occur under field conditions . Attempts are now being made to use entomopathogenic nematodes and predatory arthropods to control one major pest insect, the cabbage root fly . Techniques developed to improve the timing of application of various crop protection procedures in systems of conventional vegetable production apply equally well to organic production, despite the choice of control options being more limited . In particular, models to forecast the timing of pest insect attacks could be used to great effect, to indicate the best times to plant, protect and harvest a specific crop to minimise pest insect damage.

Bioinformatics, 2002 Nov, 18(11), 1534 - 7
A motif of a microbial starch-binding domain found in human genethonin; Janecek S; The sequence of the starch-binding domain present in 10% of amylolytic enzymes of microbial origin and classified as the carbohydrate-binding module family 20, was identified in the equivalent part of sequence of human genethonin, a skeletal muscle protein of unknown function . The sequence identity between the starch-binding domain from Bacillus sp . strain 1011 cyclodextrin glucanotransferase and the corresponding segment of human genethonin was higher than 28% . The amino acid residues known to be involved in the raw starch binding were found to be conserved in the genethonin sequence . The three-dimensional structure of the genethonin 'starch-binding domain' was modelled and its eventual function briefly discussed.

Phytochemistry, 2002 Nov, 61(6), 693 - 8
Mulberry anthracnose antagonists (iturins) produced by Bacillus amyloliquefaciens RC-2; Hiradate S et al.; Bacillus amyloliquefaciens strain RC-2 produced seven antifungal compounds (1-7) secreted into the culture filtrate . These compounds inhibited the development of mulberry anthracnose caused by the fungus, Colletotrichum dematium . Chemical structural analyses by NMR and FAB-MS revealed that all these compounds were iturins (cyclic peptides with the following sequence: L-Asn --> D-Tyr --> D-Asn --> L-Gln --> L-Pro --> D-Asn --> L-Ser --> D-beta-amino acid -->) and compounds 1-6 are identical to iturins A-2-A-7, respectively . Compound 7 (iturin A-8) is a new iturin, which has a -(CH(2))(10)CH(CH(3))CH(2)CH(3) group as a side chain in the beta-amino acid in the molecule.

J Nutr, 2002 Nov, 132(11), 3441S - 2S
Research needs to improve agricultural productivity and food quality, with emphasis on biotechnology; Thomson JA; Research into agricultural productivity, especially for crops in the developing world, should include resistance to plant viruses, fungi and the parasitic weed Striga . It must also include research into the development of resistance to Bacillus thuringiensis (Bt) toxin-expressing crops . Drought- and heat-tolerant crops, and those that can combat the problems of soil deficiencies, are required, and vaccine production in plants should be a high priority . Research into food quality should include the equivalent of "golden rice" in maize, the enhancement of the production of phytosterols and improved qualities of vegetable oils.

Insect Mol Biol, 2002 Dec, 11(6), 619 - 25
Transgenic Drosophila reveals a functional in vivo receptor for the Bacillus thuringiensis toxin Cry1Ac1; Gill M et al.; The bacterium Bacillus thuringiensis synthesizes toxins (delta-endotoxins) that are highly specific for insects . Once ingested, the activated form of the toxin binds to a specific receptor(s) located on the midgut epithelial cells, inserts into the membrane causing the formation of leakage pores and eventual death of the susceptible insect larvae . Manduca sexta larvae are highly susceptible to Cry1Ac1, a toxin that is believed to bind M . sexta Aminopeptidase N, a glycoprotein located on the apical membrane . However, the binding data obtained to date only support the interaction of Cry1Ac1 with APN in vitro . To explore the in vivo role of APN, we have utilized the GAL4 enhancer trap technique to drive the expression of M . sexta APN in both midgut and mesodermal tissues of Cry1Ac1 insensitive Drosophila larvae . Transgenic Drosophila fed the toxin were now killed, demonstrating that APN can function as a receptor for Cry1Ac1 in vivo.

J Chemother, 2002 Aug, 14(4), 336 - 41
Daptomycin morphostructural damage in Bacillus cereus visualized by atomic force microscopy; Braga PC et al.; Daptomycin is a novel, rapidly bactericidal in vitro antibiotic that is under investigation for the treatment of serious Gram-positive infections . Although daptomycin appears to disrupt membrane function, the precise mechanism of action has not been fully elucidated . Atomic force microscopy (AFM) is an innovative technique that allows high-resolution visualization and digital image manipulation of cell surface structures in 3 dimensions without the use of photons and electrons . The aim of this study was to use AFM to investigate the morphostructural changes in Bacillus cereus that occur upon daptomycin administration . The effects of daptomycin at 4x and 8x the minimal inhibitory concentration were visualized during an 8-hour incubation period . Atomic force microscopy images showed aberrant bacterial surface formations, including flattening and shrinking of cells and leakage of cytoplasm through the membrane . In addition to structural changes, the destabilization of flagella was also observed . These results support previous data suggesting that daptomycin disrupts membrane function.

Arch Microbiol, 2002 Dec, 178(6), 450 - 6 Epub 2002 Aug 29.
Localization of Mn(II)-oxidizing activity and the putative multicopper oxidase, MnxG, to the exosporium of the marine Bacillus sp . strain SG-1; Francis CA et al.; Dormant spores of the marine Bacillus sp . strain SG-1 catalyze the oxidation of manganese(II), thereby becoming encrusted with insoluble Mn(III,IV) oxides . In this study, it was found that the Mn(II)-oxidizing activity could be removed from SG-1 spores using a French press and recovered in the supernatant following centrifugation of the spores . Transmission electron microscopy of thin sections of SG-1 spores revealed that the ridged outermost layer was removed by passage through the French press, leaving the remainder of the spore intact . Comparative chemical analysis of this layer with the underlying spore coats suggested that this outer layer is chemically distinct from the spore coat . Taken together, these results indicate that this outer layer is an exosporium . Previous genetic analysis of strain SG-1 identified a cluster of genes involved in Mn(II) oxidation, the mnx genes . The product of the most downstream gene in this cluster, MnxG, appears to be a multicopper oxidase and is essential for Mn(II) oxidation . In this study, MnxG was overexpressed in Escherichia coli and used to generate polyclonal antibodies . Western blot analysis demonstrated that MnxG is localized to the exosporium of wild-type spores but is absent in the non-oxidizing spores of transposon mutants within the mnx gene cluster . To our knowledge, Mn(II) oxidation is the first oxidase activity, and MnxG one of the first gene products, ever shown to be associated with an exosporium.

Toxicon, 2002 Nov, 40(11), 1515 - 39
Plant toxic proteins with insecticidal properties . A review on their potentialities as bioinsecticides; Carlini CR et al.; To meet the demands for food of the expanding world population, there is need of new ways for protecting plant crops against predators and pathogens while avoiding the use of environmentally aggressive chemicals . A milestone in this field was the introduction into crop plants of genes expressing Bacillus thuringiensis entomotoxic proteins . In spite of the success of this new technology, however, there are difficulties for acceptance of these 'anti-natural' products by the consumers and some concerns about its biosafety in mammals . An alternative could be exploring the plant's own defense mechanisms, by manipulating the expression of their endogenous defense proteins, or introducing an insect control gene derived from another plant . This review deals with the biochemical features and mechanisms of actions of plant proteins supposedly involved in defense mechanisms against insects, including lectins, ribosome-inactivating proteins, enzymes inhibitors, arcelins, chitinases, ureases, and modified storage proteins . The potentialities of genetic engineering of plants with increased resistance to insect predation relying on the repertoire of genes found in plants are also discussed . Several different genes encoding plant entomotoxic proteins have been introduced into crop genomes and many of these insect resistant plants are now being tested in field conditions or awaiting commercialization .

Int Dent J, 2002 Oct, 52(5), 330 - 6
Re-emergence of tuberculosis and its variants: implications for dentistry; Samaranayake P; Tuberculosis is one of the deadliest scourges of mankind and, overall, one third of the global population is infected with this mycobacterium or its variants . The advent of the human immunodeficiency virus (HIV) pandemic has accelerated its spread inexorably whilst the multi-drug resistant strains of the bacillus have hampered disease management . Given the alarming spread of the disease, there appears to be a significant potential for occupationally acquired tuberculous infection amongst health care workers, including dental care workers . This review addresses the basic microbiology and the pathogenesis of tuberculosis, its oral manifestations, mycobacteria other than tuberculosis (MOTT), multi-drug resistant tuberculosis (MDR-TB), management including aspects of the global programme on tuberculosis (DOTS programme), the potential for occupationally acquiring the disease and finally, the infection control measures that are available for dental and other health care workers.

Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 2002 Nov, 34(6), 690 - 6
Purification and characterization of a novel chitinase from Bacillus brevis; Li S et al.; An extracellular chitinase secreted by Bacillus brevis was purified to homogeneity by a combination of ammonium sulfate precipitation, Phenyl-Sepharose hydrophobic-interaction chromatography and DEAE anion-exchange chromatography . On SDS-polyacrylamide gel electrophoresis analysis, the purified enzyme showed a mass of 85 kD even in the presence of beta mercaptoethanol, but shifted to 48 kD when heated in boiling water or treated with 8 mol/L urea at 50 degrees for 10 min . The depolymerization of subunits was accompanied with the loss of chitinase activity, and removing denaturing factors by dialysis could restore the dimer structure and enzymatic activity . The enzyme had an isoelectric point of 5.5 and an optimal temperature of 60 degrees, and was most active at pH 8.0 . The enzymatic activity was stable at pH 6-10, and inhibited by Ag(+) . Ten N-terminal amino acids were determined to be AVSNSKIIGY, demonstrating that the purified enzyme was a novel one . The hydrolysis pattern of the purified enzyme indicated that the chitinase was an endochitinase . The extraordinary thermo-stability and high resistance to proteolysis provide the enzyme with a good prospect to be used as a new tool for biocontrol.

Atherosclerosis, 2002 Dec, 165(2), 231 - 40
The magnitude of the immune response to heat shock protein-65 following BCG immunisation is associated with the extent of experimental atherosclerosis; Lamb DJ et al.; Several studies have reported associations between coronary heart disease (CHD) and infection . Recent studies have implicated immune responses to heat shock protein(s) (HSP) as a contributary factor . Using an immunisation model, we have assessed the relationship between the immune responses to HSP and subsequent atherosclerosis . Rabbits were immunised with bacillus Calmette-Guerin (BCG) vaccine (n=10) or saline (n=10) and subsequently fed a 0.25-1.0% cholesterol diet for 10 weeks . Plasma levels of IgG specific for mycobacterial antigen A60 and human HSP-60, but not for human HSP-70, rose following BCG immunisation, reaching a peak after 8 weeks . The percentage aortic area covered by atherosclerotic plaque was greater in animals immunised with BCG (30.5+/-3.8) compared to saline treated animals (16.4+/-2.6) (P<0.05) . Furthermore, the individual titres of anti-HSP-60 in the BCG-immunised animal antibodies at week 8 (prior to starting the cholesterol diet) correlated with the percentage aortic area covered by plaque after 18 weeks (R2=0.72; P<0.05) . No correlation was found between anti-A60 antibody titres and plaque area . Antiserum from BCG-immunised, but not control, animals stained heat-shocked endothelial cells . These data suggest that immune responses to HSP may be implicated in the relationship between specific infections and CHD.

Biochemistry, 2002 Nov 12, 41(45), 13370 - 7
Kinetic evidence for a substrate-induced fit in phosphonoacetaldehyde hydrolase catalysis; Zhang G et al.; Phosphonoacetaldehyde hydrolase (phosphonatase) from Bacillus cereus catalyzes hydrolytic P-C bond cleavage of phosphonoacetaldehyde (Pald) via a Schiff base intermediate formed with Lys53 . A single turnover requires binding of Pald to the active site of the core domain, closure of the cap domain containing the Lys53 over the core domain, and dissociation of the products following catalysis . The ligand binding and dissociation steps occur from the "open conformer" (domains are separated and the active site is solvent-exposed), while catalysis occurs from the "closed conformer" (domains are bound together and the active site is sequestered from solvent) . To test the hypothesis that bound substrate stabilizes the closed conformer, thus facilitating catalysis, the rates of chemical modification of Lys53 in the presence and absence of inert substrate and/or product analogues were compared . Acetylation of Lys53 with 2,4-dinitrophenylacetate (DNPA) resulted in the loss of enzyme activity . The pseudo-first-order rate constant for inactivation varied with pH . The pH profile of inactivation is consistent with a pK(a) of 9.3 for Lys53 . The inhibitors tungstate and vinyl sulfonate, which are known to bind to active site residues comprising the core domain, protected Lys53 from acetylation . These results are consistent with a dynamic equilibrium between the open and closed conformations of phosphonatase and the hypothesis that ligand binding stabilizes the closed conformation required for catalytic turnover.

Yakushigaku Zasshi, 2002, 37(1), 84 - 94
{A 50-year year history of new drugs in Japan: the developments of antituberculosis drugs and their influences on the epidemiological aspects}; Ozawa H et al.; Drugs used in the treatment of tuberculosis (Tb) in Japan are investigated . Especially the chemotherapy for Tb and its influences on epidemiological aspects are discussed . 1 . Various drugs were used for Tb patients before the World War II, but none was effective in curing this infectious disease . Creosote and guajacol groups were used frequently to relieve symptoms of pulmonary tuberculosis, but the disease could not cure itself . Because of the sacrifice of young patients, the mortality rate of Tb from 1935 to 1950 was ranked as the worst in Japan . So until the advent of chemotherapeutic drugs, Tb was known as the most formidable fatal plague . 2 . Streptomycin (SM), the first effective chemotherapeutic drug, was imported into Japan and widely used from 1947 . PAS in 1950 and isoniazid (INH) in 1952 were introduced to Tb therapy . The triple combination therapy of these drugs was considered the most favorable regimen for Tb from the 1950s to the early 1970s . Excellent results were obtained in this period . The mortality rate of Tb had dropped rapidly from its peak to half in 1952 and to a fourth in 1956 (Figs . 2, 3) . 3 . Several anti-Tb drugs, such as pyrazinamide ethionacide, ethambutol, and some antibiotics (kanamycin, cyloserine, and capreomycin) had been discovered and used in practice . These were not used singly, because of their weak clinical efficacies and severe side effects . They were mostly used to prevent the development of bacillus resistance to SM or INH . In the guidelines of the Japanese Society for Tuberculosis, in 1974 . drugs used for Tb could be divided into two major categories: first-line and second-line groups . The second-line drugs included those that prevent a high resistance to the main (first-line) drugs . 4 . Rifampicin (RFP), the most valuable drug for Tb, was introduced in therapy in Japan in 1971 . RFP has a low incidence of severe side-effects, but because of the rapidity with which resistance may develop, it cannot be used alone . RFP in combination with INH is the most effective therapy for all forms of the disease . The guidelines, newly proposed in 1986 by the Japanese Society for Tuberculosis recommended the short 6-month course of treatment that used combination of RFP and INH . The advent of RFP had contributed to the cure of the individual patient, but it did not effect the mortality or the morbidity rate of Tb . Chemotherapy is the most effective means of suppressing tuberculosis, which was formerly nearly always fatal, but it could not completely eradicate the disease . Preventing the development of resistance to chemotherapeutic drugs might be a special problem.

Yakushigaku Zasshi, 2002, 37(1), 76 - 83
{A 50-year history of new drugs in Japan: the developments of antileprosy drugs and their epidemiological aspects}; Ozawa H et al.; The developments of antileprosy drugs and their influences on the epidemiological aspects of Hansen's disease (leprosy) in Japan are investigated . 1 . Hydnocarpus oil (Daifushi-Yu) products were the only useful drugs for the treatment of Hansen's disease (leprosy) in Japan from the early 1900's to just after the World War II . In those days leprosy was considered to be incurable malady . 2 . The chemotherapy of leprosy, progressing from 1943 in the United States, was introduced to Japan in 1948 . Promin(R) (sulfoxone sodium) for injection in 1948 and Diasone(R), and Promizole(R) for oral use in 1949 were available for the treatment of leprosy patients in the National Hansen's Disease Sanatorium in Japan . Because DDS (dapsone, diaphenylsulfone) was proved to be the main ingredient of sulfone drugs, since 1958 it has been the drug of choice for all forms of leprosy . Monotherapy with DDS has continued for more than 30 years, and sulfone-resistant bacillus has appeared occasionally . 3 . Clofazimine (a new type of chemotherapeutic drug) and rifampicin (an antibiotic for tuberculosis) was added to therapy treatment for leprosy in 1996 . In 1983, WHO recommended multidrug therapy (MDT) to prevent resistance to sulfones . The Japanese Leprosy Association published "Guidlines" for the Treatment of Hansen's Disease in Japan" in 2000, which proposes a multidrug therapy with rifampicin, DDS, and clofazimine for a 6-month or 2-year treatment . 4 . The number of leprosy patients has slowly decreased since the application of chemotherapy with sulfone drugs, and newly infected patients in Japan have decreased to less than 10 per million persons (Figs . 1 & 2) . Therefore the "Leprosy Prevention Law" (1953), which compels the controlled isolation of patients, was abolished in 1996 . Effective chemotherapy with sulfone and other drugs has changed incurable leprosy to a curable infective disease.

Nucleic Acids Res, 2002 Nov 1, 30(21), 4692 - 9
Site-specific incorporation of an unnatural amino acid into proteins in mammalian cells; Sakamoto K et al.; A suppressor tRNA(Tyr) and mutant tyrosyl-tRNA synthetase (TyrRS) pair was developed to incorporate 3-iodo-L-tyrosine into proteins in mammalian cells . First, the Escherichia coli suppressor tRNA(Tyr) gene was mutated, at three positions in the D arm, to generate the internal promoter for expression . However, this tRNA, together with the cognate TyrRS, failed to exhibit suppressor activity in mammalian cells . Then, we found that amber suppression can occur with the heterologous pair of E.coli TyrRS and Bacillus stearothermophilus suppressor tRNA(Tyr), which naturally contains the promoter sequence . Furthermore, the efficiency of this suppression was significantly improved when the suppressor tRNA was expressed from a gene cluster, in which the tRNA gene was tandemly repeated nine times in the same direction . For incorporation of 3-iodo-L-tyrosine, its specific E.coli TyrRS variant, TyrRS(V37C195), which we recently created, was expressed in mammalian cells, together with the B.stearothermophilus suppressor tRNA(Tyr), while 3-iodo-L-tyrosine was supplied in the growth medium . 3-Iodo-L-tyrosine was thus incorporated into the proteins at amber positions, with an occupancy of >95% . Finally, we demonstrated conditional 3-iodo-L-tyrosine incorporation, regulated by inducible expression of the TyrRS(V37C195) gene from a tetracycline-regulated promoter.

J Clin Microbiol, 2002 Nov, 40(11), 4143 - 7
Rapid and specific detection of Mycobacterium tuberculosis from acid-fast bacillus smear-positive respiratory specimens and BacT/ALERT MP culture bottles by using fluorogenic probes and real-time PCR; Miller N et al.; A real-time PCR assay using the LightCycler (LC) instrument for the specific identification of Mycobacterium tuberculosis complex (MTB) was employed to detect organisms in 135 acid-fast bacillus (AFB) smear-positive respiratory specimens and in 232 BacT/ALERT MP (MP) culture bottles of respiratory specimens . The LC PCR assay was directed at the amplification of the internal transcribed spacer region of the Mycobacterium genome with real-time detection using fluorescence resonance energy transfer probes specific for MTB . The results from the respiratory specimens were compared to those from the Amplicor M . tuberculosis PCR test . Specimens from MP culture bottles were analyzed by Accuprobe and conventional identification methods . MTB was cultured from 105 (77.7%) respiratory AFB smear-positive specimens; 103 of these samples were positive by LC PCR and Amplicor PCR . Two samples negative in the LC assay contained rare numbers of organisms; both were positive in the Amplicor assay . Two separate samples negative by Amplicor PCR contained low and moderate numbers of AFB, respectively, and both of these were positive in the LC assay . There were 30 AFB smear-positive respiratory specimens that grew mycobacteria other than tuberculosis (MOTT), and all tested negative in both assays . Of the 231 MP culture bottles, 114 cultures were positive for MTB and all were positive by the LC assay . The remaining 117 culture bottles were negative in the LC assay and grew various MOTT . This real-time MTB assay is sensitive and specific; a result was available within 1 h of having a DNA sample available for testing.

BioDrugs, 2002, 16(5), 349 - 72
Potential of interferon-alpha in solid tumours: part 2; Santhanam S et al.; The second part of this review examines the use of recombinant interferon-alpha (rIFNalpha) in the following solid tumours: superficial bladder cancer, Kaposi's sarcoma, head and neck cancer, gastrointestinal cancers, lung cancer, mesothelioma and ovarian, breast and cervical malignancies . In superficial bladder cancer, intravesical rIFNalpha has a promising role as second-line therapy in patients resistant or intolerant to intravesical bacille Calmette-Guerin (BCG) . In HIV-associated Kaposi's sarcoma, rIFNalpha is active as monotherapy and in combination with antiretroviral agents, especially in patients with CD4 counts >200/mm(3), no prior opportunistic infections and nonvisceral disease . rIFNalpha has shown encouraging results when used in combination with retinoids in the chemoprevention of head and neck squamous cell cancers . It is effective in the chemoprevention of hepatocellular cancer in hepatitis C-seropositive patients . In neuroendocrine tumours, including carcinoid tumour, low-dosage (</=3 MU) or intermediate-dosage (5 to 10 MU) rIFNalpha is indicated as second-line treatment, either with octreotide or alone in patients resistant to somatostatin analogues . Intracavitary IFNalpha may be useful in malignant pleural effusions from mesothelioma . Similarly, intraperitoneal IFNalpha may have a role in the treatment of minimal residual disease in ovarian cancer . In breast cancer, the only possible role for IFNalpha appears to be intralesional administration for resistant disease . IFNalpha may have a role as a radiosensitising agent for the treatment of cervical cancer; however, this requires confirmation in randomised trials . On the basis of current evidence, the routine use of rIFNalpha is not recommended in the therapy of head and neck squamous cell cancers, upper gastrointestinal tract, colorectal and lung cancers, or mesothelioma . Pegylated IFNalpha (peginterferon-alpha) is an exciting development that offers theoretical advantages of increased efficacy, reduced toxicity and improved compliance . Further data from randomised studies in solid tumours are needed where rIFNalpha has activity, such as neuroendocrine tumours, minimal residual disease in ovarian cancer, and cervical cancer . A better understanding of the biological mechanisms that determine response to rIFNalpha is needed . Studies of IFNalpha-stimulated gene expression, which are now feasible, should help to identify molecular predictors of response and allow us to target therapy more selectively to patients with solid tumours responsive to IFNalpha.

Mar Environ Res, 2002 Sep-Dec, 54(3-5), 755 - 9
Esterases as biomarkers in Nereis (Hediste) diversicolor exposed to temephos and Bacillus thuringiensis var . israelensis used for mosquito control in coastal wetlands of Morbihan (Brittany, France); Fourcy D et al.; Since 1998, a biomonitoring programme has been implemented to assess the potential impact of chemical mosquito control on macroinvertebrates of the coastal wetlands of Morbihan (Brittany, France) . Acetylcholinesterase and carboxylesterases were used as biomarkers to assess the effects of Abate 500e (a.i . temephos) and Vectobac 12 AS (a.i . endotoxins of Bacillus thuringiensis var . israelensis, Bti) in Nereis (Hediste) diversicolor . Esterase inhibition revealed a marked impact of temephos, suggesting preferential contamination of the worms through the food . In Bti-exposed N . diversicolor, random variations of esterase activities were observed, that could not be attributed to the larvicide . However, esterases only reflected indirect physiological effects of Bti, and further investigations are needed to identify biomarkers more specific of Bti endotoxins.

Appl Environ Microbiol, 2002 Nov, 68(11), 5711 - 7
Altered Glycosylation of 63- and 68-kilodalton microvillar proteins in Heliothis virescens correlates with reduced Cry1 toxin binding, decreased pore formation, and increased resistance to Bacillus thuringiensis Cry1 toxins; Jurat-Fuentes JL et al.; The binding and pore formation abilities of Cry1A and Cry1Fa Bacillus thuringiensis toxins were analyzed by using brush border membrane vesicles (BBMV) prepared from sensitive (YDK) and resistant (YHD2) strains of Heliothis virescens . 125I-labeled Cry1Aa, Cry1Ab, and Cry1Ac toxins did not bind to BBMV from the resistant YHD2 strain, while specific binding to sensitive YDK vesicles was observed . Binding assays revealed a reduction in Cry1Fa binding to BBMV from resistant larvae compared to Cry1Fa binding to BBMV from sensitive larvae . In agreement with this reduction in binding, neither Cry1A nor Cry1Fa toxin altered the permeability of membrane vesicles from resistant larvae, as measured by a light-scattering assay . Ligand blotting experiments performed with BBMV and 125I-Cry1Ac did not differentiate sensitive larvae from resistant larvae . Iodination of BBMV surface proteins suggested that putative toxin-binding proteins were exposed on the surface of the BBMV from resistant insects . BBMV protein blots probed with the N-acetylgalactosamine-specific lectin soybean agglutinin (SBA) revealed altered glycosylation of 63- and 68-kDa glycoproteins but not altered glycosylation of known Cry1 toxin-binding proteins in YHD2 BBMV . The F1 progeny of crosses between sensitive and resistant insects were similar to the sensitive strain when they were tested by toxin-binding assays, light-scattering assays, and lectin blotting with SBA . These results are evidence that a dramatic reduction in toxin binding is responsible for the increased resistance and cross-resistance to Cry1 toxins observed in the YHD2 strain of H . virescens and that this trait correlates with altered glycosylation of specific brush border membrane glycoproteins.

J Agric Food Chem, 2002 Nov 6, 50(23), 6775 - 9
Efficiency of mannose-binding plant lectins in controlling a homopteran insect, the red cotton bug; Roy A et al.; Yield losses of different crops due to the attack of various classes of insects are a worldwide problem . Sucking type homopteran pests causing damage to many crop species are not controlled by commonly known insecticidal proteins, namely, Bacillus thuringiensis delta-endotoxin (Bt) . This study describes the purification of mannose-binding lectins from three different monocotyledonous plants (Allium sativum, Colocasia esculenta, and Diffenbachia sequina) and their effects on a homopteran insect, the red cotton bug . All of them had a detrimental effect on the growth and development of the insect, where A . sativum bulb lectin showed the highest mortality of all, in particular . The same bulb lectin not only affected the growth and fecundity of the insect but also imparted drastic changes in the color, weight, and size, even on the second generation of the insects which have been reared on artificial diet supplemented with a sublethal dose of the lectin . Thus, this finding opens up a possibility of using this lectin as an important component in crop management.

Crit Rev Biotechnol, 2002, 22(3), 225 - 44
Positive selection vectors; Choi YJ et al.; This review describes information concerning positive selection vectors on their mechanism, classification, property, and limitation . A total of 72 positive selection vectors collected were discussed . Positive selection vectors can reduce background and directly screen transformants containing cloned DNA fragments . The mechanisms to perform positive selection include insertional inactivation and the replacement of functional genes of the vectors . In general, the former is much more convenient than the latter . The functional genes are controlled either by their promoters or by heterologous promoters introduced . On the basis of the structures, positive selection vectors could be classified into five groups . The positive selection vectors are commonly based on the mechanisms of lethal genes and the sensitivity of compounds . The vectors, with molecular weights ranging from 2.6 to 17.0 kb, have diverse genetic markers and wide host ranges, including Escherichia coli, Bacillus, Streptomyces, lactic acid bacteria, yeasts, and mammalian cells . Although some limitations exist for using some positive selection vectors, they are useful in recombinant DNA experiments.

J Infect Dis, 2002 Nov 15, 186(10), 1448 - 57 Epub 2002 Oct 23.
Investigation of the relationships between immune-mediated inhibition of mycobacterial growth and other potential surrogate markers of protective Mycobacterium tuberculosis immunity; Hoft DF et al.; Tuberculosis (TB) vaccine development is hindered by the lack of clear surrogate markers of protective human immunity to Mycobacterium tuberculosis . This study evaluated the hypothesis that immune-mediated inhibition of mycobacterial growth would more directly correlate with protective TB immunity than other immunologic responses . Bacille Calmette-Guerin (BCG) vaccination, known to induce partial protection against TB, was used as a model system to investigate mechanistic relationships among different parameters of antigen-specific immunity . Effects of primary and booster intradermal BCG vaccinations were assessed in 3 distinct assays of mycobacterial inhibition . Correlations between vaccine-induced growth inhibition and other immune responses were analyzed . BCG significantly enhanced all antigen-specific responses . Peak responses occurred at 2 months after boosting . Statistical analyses suggested that each assay measured unique aspects of mycobacterial immunity . Despite previous evidence that type 1 immune responses are essential for TB immunity, interferon-gamma production did not correlate with mycobacterial inhibition . These results have important implications for TB vaccine development.

Thorax, 2002 Nov, 57(11), 964 - 6
Tobacco smoking and pulmonary tuberculosis; Kolappan C et al.; BACKGROUND: The prevalence of tuberculosis in adult men in India is 2-4 times higher than in women . Tobacco smoking is prevalent almost exclusively among men, so it is possible that tobacco smoking may be a risk factor for developing pulmonary tuberculosis . A nested case control study was carried out to study the association between tobacco smoking and pulmonary tuberculosis . METHODS: A tuberculosis disease survey was carried out in two Panchayat unions in the Tiruvallur district of Tamil Nadu in India . Eighty five men aged 20-50 years with bacteriological tuberculosis (smear and/or culture positive) were selected as cases and 459 age matched men without tuberculosis were selected randomly as controls . Information on smoking status, type of tobacco smoked, quantity of tobacco smoked, and duration of tobacco smoking was collected from cases and controls using a questionnaire . RESULTS: The estimated crude odds ratio (OR) of the association between tobacco smoking and bacillary tuberculosis was 2.48 (95% confidence interval (CI) 1.42 to 4.37), p<0.001.The age adjusted OR (Mantel-Hanszel estimate) was 2.24.(95% CI 1.27 to 3.94), p<0.05 . The ORs for mild (1-10 cigarettes/day), moderate (11-20/day), and heavy (>20/day) smokers were 1.75, 3.17, and 3.68, respectively (p<0.0001 test for linear trend) . The ORs for smokers with <10 years, 11-20 years, and >20 years of smoking were 1.72, 2.45, and 3.23, respectively (p<0.0001 test for linear trend) . CONCLUSION: There is a positive association between tobacco smoking and pulmonary (bacillary) tuberculosis (OR 2.5) . The association also shows a strong dose-response relationship.

Biochem J, 2003 Feb 1, 369(Pt 3), 697 - 703
Cytotoxic activity of Bacillus thuringiensis Cry proteins on mammalian cells transfected with cadherin-like Cry receptor gene of Bombyx mori (silkworm); Tsuda Y et al.; Cry1Aa, an insecticidal protein produced by Bacillus thuringiensis, has been shown to bind to cadherin-like protein, BtR175, in Bombyx mori (silkworm) midgut . We previously reported three variant alleles of BtR175 (BtR175a, b and c) . When transiently expressed in COS7 cells, all the three BtR175 variants bound to Cry1Aa . We stably expressed BtR175b in HEK293 cells . These BtR175b-expressing cells swelled and died in the presence of activated Cry1Aa in a dose- and time-dependent manner, showing that BtR175b itself can impart Cry1Aa-susceptibility to mammalian cells . These cells were more susceptible to Cry1Aa than to Cry1Ab and Cry1Ac . Since dispersed B . mori midgut cells were reported to be highly susceptible to Cry1Ac, this result suggested that other Cry1Ac-specific receptor(s) were simultaneously working with BtR175 in the midgut cells . Advantages are also discussed of applying these transfected mammalian cells to toxicity assays of mutant Cry proteins.

J Econ Entomol, 2002 Oct, 95(5), 1018 - 26
Inheritance of resistance to Bt toxin crylac in a field-derived strain of pink bollworm (Lepidoptera: Gelechiidae); Tabashnik BE et al.; Laboratory selection with Cry1Ac, the Bacillus thuringiensis (Bt) toxin in transgenic cotton, initially produced 300-fold resistance in a field-derived strain of pink bollworm, Pectinophora gossypiella (Saunders), a major cotton pest . After additional selection increased resistance to 3,100-fold, we tested the offspring of various crosses to determine the mode of inheritance of resistance to Cry1Ac . The progeny of reciprocal F1 crosses (resistant male x susceptible female and vice versa) responded alike in bioassays, indicating autosomal inheritance . Consistent with earlier findings, resistance was recessive at a high concentration of Cry1Ac . However, the dominance of resistance increased as the concentration of Cry1Ac decreased . Analysis of survival and growth of progeny from backcrosses (F1 x resistant strain) suggest that resistance was controlled primarily by one or a few major loci . The progression of resistance from 300- to 3,100-fold rules out the simplest model with one locus and two alleles . Overall the patterns observed can be explained by either a single resistance gene with three or more alleles or by more than one resistance gene . The pink bollworm resistance to Cry1Ac described here fits "mode 1" resistance, the most common type of resistance to Cry1A toxins in Lepidoptera.

J Econ Entomol, 2002 Oct, 95(5), 878 - 92
Biotechnology and the European corn borer: measuring historical farmer perceptions and adoption of transgenic Bt corn as a pest management strategy; Pilcher CD et al.; A 3-yr, multi-state survey of farmers who had planted transgenic Bacillus thuringiensis (Bt) corn was conducted to evaluate perceptions of Bt corn performance and its utility as a management option for European corn borer, Ostrinia nubilalis (Hubner) . A questionnaire was sent to farmers in Illinois, Iowa, Kansas, Minnesota, Nebraska, and Pennsylvania who had grown Bt corn during the growing seasons of 1996, 1997, or 1998 . There were 7,427 usable questionnaires returned with the following response percentages: 1996 (42.1%), 1997 (35.0%), and 1998 (22.6%) . Adoption rates, based on percentage of acreage planted to Bt corn, increased dramatically from 1996 (10.5%) to 1998 (40.7%) . The states growing the highest percentage of Bt corn were Minnesota, Iowa, and then Nebraska However, Illinois, was adopting Bt corn at the fastest rate . Historical use of insecticides did not influence the adoption of Bt corn . In addition, of those farmers who used insecticides to control European corn borer, the percentage that decreased their use of insecticides nearly doubled from 13.2% (1996) to 26.0% (1998) over this 3-yr period . The primary reason farmers planted Bt corn was to eliminate the yield loss caused by European corn borer . Scouting for European corn borers decreased from 91% (scouting 2.2 times a year) in 1996 to 75% (scouting 1.8 times a year) in 1998 . The percentage of farmers not scouting for European corn borers increased from 9.6% (1996) to 25% (1998) . Most farmers believed yields of Bt hybrids were either similar to or greater than the yields of non-Bt hybrids . Minnesota farmers perceived the greatest yield advantages . Farmers are becoming more aware of insect resistance management guidelines; however, they also clearly show preferences for having the flexibility to use different spatial plantings of Bt and non-Bt corn . Finally, after having planted Bt corn and obtained excellent control of European corn borer, most farmers believed that this insect had been causing more yield loss than they previously had suspected in their non-Bt corn . The data represented here provide an historical foundation for how transgenic Bt corn was used by farmers during the first 3 yr of commercial availability, their initial perceptions on the performance of this technology, and their attitudes regarding management of the European corn borer.

Biotechnol Bioeng, 2002 Dec 30, 80(7), 755 - 61
Laboratory-scale continuous reactor for soluble selenium removal using selenate-reducing bacterium, Bacillus sp . SF-1; Fujita M et al.; A model continuous flow bioreactor (volume 0.5 L) was constructed for removing toxic soluble selenium (selenate/selenite) of high concentrations using a selenate-reducing bacterium, Bacillus sp . SF-1, which transforms selenate into elemental selenium via selenite for anaerobic respiration . Model wastewater contained 41.8 mg-Se/L selenate and excess lactate as the carbon and energy source; the bioreactor was operated as an anoxic, completely mixed chemostat with cell retention time between 2.2-95.2 h . At short cell retention times selenate was removed by the bioreactor, but accumulation of selenite was observed . At long cell retention times soluble selenium, both selenate and selenite, was successfully reduced into nontoxic elemental selenium . A simple mathematical model is proposed to evaluate Se reduction ability of strain SF-1 . First-order kinetic constants for selenate and selenite reduction were estimated to be 2.9 x 10(-11) L/cells/h and 5.5 x 10(-13) L/cells/h, respectively . The yield of the bacterial cells by selenate reduction was estimated to be 2.2 x 10(9) cells/mg-Se .

Curr Microbiol, 2002 Dec, 45(6), 405 - 9
Intracellular proteases of Bacillus thuringiensis subsp . kurstaki and a protease-deficient mutant Btk-q; Reddy YC et al.; The commencement of intracellular protease synthesis was studied by gelatin zymography in Bacillus thuringiensis ( Btk) HD1, Btk HD73, and a protease-deficient mutant Btk-q derived from the former strain . By gelatin zymography, a 92-kDa protease was detected first at 3 h of sporulation, which continued until 48 h, whereas two other proteases of mol wt 78 and 69 kDa were detectable from 6 h onwards and continued until 48 h of growth in Btk HD1 . Similar studies revealed the presence of two major intracellular proteases in Btk HD73 by gelatin zymography, which first appeared at 6 h of sporulation and continued until 48 h of growth . The quantitative azocasein assay confirmed that the total protease activity increases from 3 to 21 h, thereafter reaching a plateau up to 48 h of growth examined, in HD1 and HD73 strains . Btk-q, a protease-deficient mutant, showed traces of protease activity by azocasein analysis that could not be detected by gelatin zymography . The free amino acid pool content was also increased parallel to the way that the protease activity increased in all three strains . However, this increase was found to be low (16-fold) in Btk-q when compared with Btk HD1 and HD73 strains . The following amino acids were detected by paper chromatography in Btk HD1: DL-alanine, L-glutamic acid, L-aspartic acid, tyrosine, tryptophan/methionine/valine, arginine, leucine/norleucine/isoleucine, and glycine, whereas only DL-alanine, L-glutamic acid, and L-aspartic acid were in Btk-q at 24 and 48 h, when the protease activity was maximum.

Int J Clin Oncol, 2002 Oct, 7(5), 289 - 93
Complications of bacillus Calmette-Guerin therapy in superficial urothelial cancer: clinical analysis and implications; Suzuki S et al.; BACKGROUND: Intravesical bacillus Calmette-Guerin (BCG) therapy has been proven to be effective in the prophylaxis and treatment of superficial bladder cancer . However, several complications of BCG therapy have been reported . The aim of this study was to clarify the impact of BCG treatment-related side effects on the clinical outcome of patients with superficial urothelial cancer . METHODS: We reviewed the medical records of 33 patients who underwent BCG instillation therapy in our department . After complete endoscopic tumor resection, intravesical or intrapelvic instillation of BCG (80 mg of the Tokyo strain) was performed every week for 8 weeks . BCG treatment-related side effects were classified as minor (persistence of symptoms or low-grade fever for less than 48 h) or major (persistence of symptoms or low-grade fever for more than 48 h, or high fever) . Risk factors for major side effects and relationships between the occurrence of major side effects and subsequent tumor progression were evaluated . RESULTS: In total, there were 43 courses of intravesical and intrapelvic instillations of BCG in 33 patients, 20 (46%) of which were associated with major side effects . Risk factors associated with the occurrence of major side effects could not be detected . Subsequent tumor progression was observed in 3 of the 16 patients (19%) with major side effects and in 10 of the 17 patients (59%) without them . Nine patients who discontinued BCG therapy because of major side effects experienced no tumor progression . Progression-free survival was significantly higher in patients with major side effects than in those without them . CONCLUSIONS: These results suggest that BCG therapy should be discontinued whenever major side effects occur, because this does not necessarily lead to an unfavorable outcome regarding tumor progression.

Infect Control Hosp Epidemiol, 2002 Oct, 23(10), 595 - 9
Failure to implement respiratory isolation: why does it happen?
Iwata K, Smith BA, Santos E, Polsky B, Sordillo EM.
BACKGROUND: Respiratory isolation for 90% of individuals with acid-fast bacillus (AFB)-smear-positive tuberculosis (TB) is a recommended performance indicator in recent Infectious Diseases Society of America and Centers for Disease Control and Prevention guidelines . However, compliance with respiratory isolation reported from multiple centers in the United States and Europe falls short of that goal . OBJECTIVE: To identify missed clues in TB patients who are not appropriately isolated . DESIGN: Retrospective survey . SETTING: A 900-bed voluntary hospital . PATIENTS: All patients with AFB-smear-positive TB admitted between January 1995 and December 1999 who were not appropriately isolated . RESULTS: There were 173 TB cases admitted, including 106 with pulmonary TB . AFB smears were positive in 82 cases; 24 (29%) of these were not appropriately isolated . During the study period, the number of TB cases declined, but the proportion of appropriately isolated patients did not change . Most isolation failure cases were men (median age, 45.5 years); 21 of these patients were black, 2 were Hispanic white, and 1 was Asian, but none was non-Hispanic white . All isolation failure cases had at least one characteristic predictive of TB that could have been elicited at admission (eg, abnormal chest radiograph findings consistent with TB, fever, weight loss, a history of TB, a positive result on tuberculin skin test, hemoptysis, and human immunodeficiency virus infection) . CONCLUSION: Consistent with experiences at other hospitals, we found that the rate of isolation failure remained unchanged despite an overall decline in TB cases . In our experience, almost all isolation failures could be avoided by careful review of the history, physical examination, and chest radiograph for characteristics classically considered predictive of TB.

Infect Control Hosp Epidemiol, 2002 Oct, 23(10), 591 - 4
Tuberculin skin test conversion among medical students at a teaching hospital in Rio de Janeiro, Brazil; Silva VM et al.; OBJECTIVE: To describe the cumulative incidence of and risk factors for tuberculosis (TB) infection among medical students . DESIGN: In 1999, a cohort study of medical students with negative results (induration < 10 mm) on tuberculin skin test (TST) was performed . Students who had undergone two-step testing in 1998 were retested . SETTINGS: University and teaching hospital and referral center for TB and acquired immunodeficiency syndrome, and the Health Sciences Building of the Medical School of the Federal University of Rio de Janeiro, Brazil . PARTICIPANTS: A sample of 618 consecutive medical students with negative TST results who had been tested 12 months before were approached . Information about sociodemographic characteristics, bacille Calmette-Guerin vaccination history, and potential exposures to TB was obtained using a standardized questionnaire . Four hundred fourteen (67%) students completed the study . Students were at two different levels of their training programs (juniors = no contact with patients; seniors = intensive contact with patients) . RESULTS: Of 414 participants, 16 (3.9%; 95% confidence interval, 1.06% to 12.1%) had converted to a positive reaction after 1 year . In a multivariate logistic regression analysis, higher level of clinical training was confirmed to be an independent factor associated with TST conversion (odds ratio, 4.77; 95% confidence interval, 1.01 to 22.46; P= .048) . CONCLUSION: Senior medical students are at increased risk of Mycobacterium tuberculosis infection in this setting . Therefore, a program of routine tuberculin skin testing and specific TB infection control guidelines are needed for this population.

Infect Control Hosp Epidemiol, 2002 Oct, 23(10), 584 - 90
Tuberculin skin testing among healthcare workers in the University of Malaya Medical Centre, Kuala Lumpur, Malaysia; Tan LH et al.; OBJECTIVES: To determine the occupational risk of Mycobacterium tuberculosis infection among healthcare workers (HCWs) and to examine the utility of tuberculin skin testing in a developing country with a high prevalence of bacille Calmette-Guerin vaccination . DESIGN: Tuberculin skin test (TST) survey . SETTING: A tertiary-care referral center and a teaching hospital in Kuala Lumpur, Malaysia . PARTICIPANTS: HCWs from medical, surgical, and orthopedic wards . INTERVENTION: Tuberculin purified protein derivative RT-23 (State Serum Institute, Copenhagen, Denmark) was used for the TST (Mantoux method) . RESULTS: One hundred thirty-seven (52.1%) and 69 (26.2%) of the HCWs tested had indurations of 10 mm or greater and 15 mm or greater, respectively . Medical ward HCWs were at significantly higher risk of a positive TST reaction than were surgical or orthopedic ward HCWs (odds ratio, 2.18; 95% confidence interval, 1.33 to 3.57; P = .002 for TST positivity at 10 mm or greater) (odds ratio, 2.61; 95% confidence interval, 1.44 to 4.70; P = .002 for TST positivity at 15 mm or greater) . A previous TST was a significant risk factor for a positive TST reaction at either 10 mm or greater or 15 mm or greater, but a duration of employment of more than 1 year and being a nurse were only significantly associated with a positive TST reaction at a cut-off point of 15 mm or greater . CONCLUSIONS: HCWs at the University of Malaya Medical Centre had an increased risk for M . tuberculosis infection that was significantly associated with the level of occupational tuberculosis exposure . A TST cut-off point of 15 mm or greater may correlate better with M . tuberculosis infection than a cut-off point of 10 mm or greater in settings with a high prevalence of bacille Calmette-Guerin vaccination.

Biosci Biotechnol Biochem, 2002 Sep, 66(9), 1873 - 9
Cloning, sequencing, and expression of the gene from bacillus circulans that codes for a heparinase that degrades both heparin and heparan sulfate; Yoshida E et al.; The gene, designated hep, coding for a heparinase that degrades both heparin and heparan sulfate, was cloned from Bacillus circulans HpT298 . Nucleotide sequence analysis showed that the open reading frame of the hep gene consists of 3,150 bp, encoding a precursor protein of 1,050 amino acids with a molecular mass of 116.5 kDa . A homology search found that the deduced amino acid sequence has partial similarity with enzymes belonging to the family of acidic polysaccharide lyases that degrade chondroitin sulfate and hyaluronic acid . Recombinant mature heparinase (111.2 kDa) was produced by the addition of IPTG from Escherichia coli harboring pETHEP with an open reading frame of the mature hep gene and was purified to homogeneity by SDS-polyacrylamide gel electrophoresis . Analyses of substrate specificity and degraded disaccharides indicated that the recombinant enzyme acts on both heparin and HS, as does heparinase purified from the wild-type strain.

Biosci Biotechnol Biochem, 2002 Sep, 66(9), 1806 - 18
Purification and characterization of glucosyltransferase and glucanotransferase involved in the production of cyclic tetrasaccharide in Bacillus globisporus C11; Nishimoto T et al.; Glucosyltransferase and glucanotransferase involved in the production of cyclic tetrasaccharide (CTS; cyclo {-->6}-alpha-D-glucopyranosyl-(1-->3)-alpha-D-glucopyranosyl-(1-->6)-alpha-D-glucopyranosyl-(1-->3)-alpha-D-glucopyranosyl-(1-->)) from alpha-1,4-glucan were purified from Bacillus globisporus C11 . The former was a 1,6-alpha-glucosyltransferase (6GT) catalyzing the a-1,6-transglucosylation of one glucosyl residue to the nonreducing end of maltooligosaccharides (MOS) to produce alpha-isomaltosyl-MOS from MOS . The latter was an isomaltosyl transferase (IMT) catalyzing alpha-1,3-, alpha-1,4-, and alpha,beta-1,1-intermolecular transglycosylation of isomaltosyl residues . When IMT catalyzed alpha-1,3-transglycosylation, alpha-isomaltosyl-(1-->3)-alpha-isomaltosyl-MOS was produced from alpha-isomaltosyl-MOS . In addition, IMT catalyzed cyclization, and produced CTS from alpha-isomaltosyl-(1-->3)-alpha-isomaltosyl-MOS by intramolecular transglycosylation . Therefore, the mechanism of CTS synthesis from MOS by the two enzymes seemed to follow three steps: 1) MOS-->alpha-isomaltosyl-->MOS (by 6GT), 2) alpha-isomaltosyl-MOS-->alpha-isomaltosyl-(1-->3)-alpha-isomaltosyl-MOS (by IMT), and 3) alpha-isomaltosyl-(1-->3)-alpha-isomaltosyl-MOS-->CTS + MOS (by IMT) . The molecular mass of 6GT was estimated to be 137 kDa by SDS-PAGE . The optimum pH and temperature for 6GT were pH 6.0 and 45 degrees C, respectively . This enzyme was stable at from pH 5.5 to 10 and on being heated to 40 degrees C for 60 min . 6GT was strongly activated and stabilized by various divalent cations . The molecular mass of IMT was estimated to be 102 kDa by SDS-PAGE . The optimum pH and temperature for IMT were pH 6.0 and 50 degrees C, respectively . This enzyme was stable at from pH 4.5 to 9.0 and on being heated to 40 degrees C for 60 min . Divalent cations had no effect on the stability or activity of this enzyme.

Pest Manag Sci, 2002 Oct, 58(10), 1022 - 8
Susceptibility of field-collected populations of the southwestern corn borer, Diatraea grandiosella, to Bacillus thuringiensis; Trisyono YA et al.; The susceptibilitity of newly hatched larvae of laboratory-adapted and field-collected populations of the Southwestern corn borer, Diatraea grandiosella (Dyar) (Lepidoptera: Crambidae), to a Bacillus thuringiensis protein (Cry1Ab) was examined using a larval feeding bioassay . D grandiosella populations were collected from five states: Missouri, Kansas, Texas, Tennessee and Kentucky . Using larval mortality as the end-point of the bioassay, the magnitude of differences in the susceptibility of the laboratory-adapted and the field-collected populations to Cry1Ab protein varied from one to 46 times depending on the time of observation and the standard of comparison (LC50 or LC95) . However, significant differences in susceptibility to Cry1Ab protein among these populations were not detected when the comparisons were based on growth inhibition (EC50 or EC95); the magnitude of differences was less than fourfold . Either using larval mortality or larval growth inhibition, the results indicated that the field-collected populations of D grandiosella were susceptible to Cry1Ac, and differences in susceptibility may reflect natural variation among populations . The bioassay using larval growth inhibition offers advantages over that using larval mortality, including giving more accurate representation of the toxicological effects of the toxin.

Rev Chir Orthop Reparatrice Appar Mot, 2002 Sep, 88(5), 522 - 5
{Tuberculosis of the talus: a rare localization of Koch's bacillus}; Boussouga M et al.; A 66-year-old woman complained of pain in the left rear foot and malleoli . Standard x-ray and magnetic resonance imaging as well as laboratory tests were non-contributive . Pathology study of a bone biopsy led to the diagnosis of talar tuberculosis . A 12-month regimen combining rifampicin, isoniazid and pyrazinamide was given . Clinical improvement was achieved although the patient developed significant reflex dystrophy . The risk of degeneration compromises the functional future of the ankle joint.

Dermatology, 2002, 205(3), 289 - 92
Diagnostic odyssey of a cutaneous mycobacteriosis rare in central Europe; Fischer TW et al.; Cutaneous infection with Mycobacterium chelonae is an uncommon disease, although this atypical mycobacterium is an acid-fast bacillus ubiquitous in the environment . It is often misdiagnosed and treated as a fungal or common bacterial infection . We report a case of disseminated atypical mycobacterial skin infection of a 72-year-old woman who was treated with different topical and systemic antimycotic and antibiotic drugs over a period of 5 months without remarkable improvement . Eventually, repeated tissue cultures on special medium and performance of PCR led to the diagnosis of M . chelonae infection . The patient was treated successfully with oral clarithromycin within 8 weeks . In case of abscessing cutaneous infection, M . chelonae should be considered in the differential diagnosis of prolonged disease when common antibiotics are not effective after 2-4 weeks of treatment .

J Bacteriol, 2002 Nov, 184(22), 6301 - 15
Genes coding for a new pathway of aerobic benzoate metabolism in Azoarcus evansii; Gescher J et al.; A new pathway for aerobic benzoate oxidation has been postulated for Azoarcus evansii and for a Bacillus stearothermophilus-like strain . Benzoate is first transformed into benzoyl coenzyme A (benzoyl-CoA), which subsequently is oxidized to 3-hydroxyadipyl-CoA and then to 3-ketoadipyl-CoA; all intermediates are CoA thioesters . The genes coding for this benzoate-induced pathway were investigated in the beta-proteobacterium A . evansii . They were identified on the basis of N-terminal amino acid sequences of purified benzoate metabolic enzymes and of benzoate-induced proteins identified on two-dimensional gels . Fifteen genes probably coding for the benzoate pathway were found to be clustered on the chromosome . These genes code for the following functions: a putative ATP-dependent benzoate transport system, benzoate-CoA ligase, a putative benzoyl-CoA oxygenase, a putative isomerizing enzyme, a putative ring-opening enzyme, enzymes for beta-oxidation of CoA-activated intermediates, thioesterase, and lactone hydrolase, as well as completely unknown enzymes belonging to new protein families . An unusual putative regulator protein consists of a regulator protein and a shikimate kinase I-type domain . A deletion mutant with a deletion in one gene (boxA) was unable to grow with benzoate as the sole organic substrate, but it was able to grow with 3-hydroxybenzoate and adipate . The data support the proposed pathway, which postulates operation of a new type of ring-hydroxylating dioxygenase acting on benzoyl-CoA and nonoxygenolytic ring cleavage . A beta-oxidation-like metabolism of the ring cleavage product is thought to lead to 3-ketoadipyl-CoA, which finally is cleaved into succinyl-CoA and acetyl-CoA.

FEMS Microbiol Lett, 2002 Oct 8, 215(2), 261 - 5
Use of fluorescence ratio imaging microscopy and flow cytometry for estimation of cell vitality for Bacillus licheniformis; Hornbaek T et al.; For Bacillus licheniformis SJ4628, an organism widely used in the enzyme industry, methods for determination of cell vitality at a single cell level using 5(6)-carboxyfluorescein diacetate succinimidyl ester in combination with fluorescence ratio imaging microscopy and flow cytometry were developed . Immediately after inoculation and during growth, changes in intracellular pH values determined by fluorescence ratio imaging microscopy and in green fluorescence intensities determined by flow cytometry were observed . Correlations between the capacity to multiply and intracellular pH or green fluorescence intensity were demonstrated . Populations of cells not having a pH gradient or exhibiting low fluorescence intensities had significantly longer lag phases than populations of cells with a pH gradient and high fluorescence intensities.

J Nat Prod, 2002 Oct, 65(10), 1447 - 51
Tupuseleiamides and basiliskamides, new acyldipeptides and antifungal polyketides produced in culture by a Bacillus laterosporus isolate obtained from a tropical marine habitat; Barsby T et al.; Laboratory cultures of PNG 276, a Bacillus laterosporus isolate obtained from coastal waters off Papua New Guinea, have been shown to produce the novel metabolites basiliskamide A (1), basiliskamide B (2), tupuseleiamide A (3), and tupusleiamide B (4) . The structures of 1 to 4 were elucidated by analysis of spectroscopic data and chemical degradation . Basiliskamides A (1) and B (2) show potent in vitro anti-Candida activity.

Appl Biochem Biotechnol, 2002 Jul-Dec, 102-103(1-6), 303 - 13
Immobilization of alpha-amylase from Bacillus circulans GRS 313 on coconut fiber; Dey G et al.; A simple and inexpensive method for immobilizing alpha-amylase from Bacillus circulans GRS 313 on coconut fiber was developed . The immobilization conditions for highest efficiency were optimized with respect to immobilization pH of 5.5, 30 degrees C, contact time of 4 h, and enzyme to support a ratio of 1:1 containing 0.12 mg/mL of protein . The catalytic properties of the immobilized enzyme were compared with that of the free enzyme . The activity of amylase adsorbed on coconut fiber was 38.7 U/g of fiber at its optimum pH of 5.7 and 48 degrees C, compared with the maximum activity of 40.2 U/mL of free enzyme at the optimum pH of 4.9 and 48 degrees C . The reutilization capacity of the immobilized enzyme was up to three cycles.

Appl Biochem Biotechnol, 2002 Jul-Dec, 102-103(1-6), 213 - 26
Phenotypic and genetic diversity of Bacillus thuringiensis strains isolated in India active against Spodoptera litura; Prabagaran SR et al.; Bacillus thuringiensis strains isolated from different agroclimatic regions of India were found to harbor cry1 family genes . Of 831 strains 18 that were found to produce 130- and 68-kDa mol wt proteins in sodium dodecyl sulfate polyacry1amide gel electrophoresis were subjected to bioassay against second instar larvae of Spodoptera litura . According to the time response curve, while the highest toxic activity against S . litura was observed in PBT-782 with an LT50 of 25.46 h, strains PBT-372, PBT-574, PBT-801, and PBT-716 in descending order of merit had LT50 values of 36.81, 48.18, 50.35, and 73.53 h . The results of the field experiment testing the efficacy of different B . thuringiensis strains in controlling S . litura larvae infecting peanut plants showed that the chemical insecticide chlorpyriphos was the most effective in controlling S . litura throughout the study period . However, among B . thuringiensis strains, PBT-372 was superior . All the B . thuringiensis strains except PBT-689 were found to contain cry1Ac1-type gene . However, only nine strains contained cry1Aa1 gene . While cry1Ab1 was present only in PBT-372 and PBT-689, cry1Ca1 was present in PBT-574, PBT-688, PBT-689, and PBT-695 . cry1Da1 was detected only in PBT-688 and PBT-692 . None of the strains contained cry1Ba1 and cry1Ea1 genes . When polymerase chain reaction analysis using cry1Ca1 primer was performed, PBT-695 produced an unexpected 739-bp product, which showed 33% homology with cry1Ca1 gene between nucleotides 1819 and 2107 . Our results indicated that among the field-collected B . thuringiensis strains, PBT-372 harbors multiple cry-type genes and could be employed for biological control of insects.

Indian J Environ Health, 2001 Oct, 43(4), 174 - 5
Physico-chemical characteristics and algae of a percolation tank of Pimpalgaon; Shastri Y et al.; Physico-Chemical characteristics and algae of a percolation tank of Pimpalgaon (Dabhadi) have been studied from January to December 1997 . The studies reveal that in all 92 algal taxa belonging to four classes i.e . Cyanophyceae, Chlorophyceae, Bacillariophyceae and Euglenophyceae were recorded . Cyanophyceae members show luxurient growth in summer . Euglenoids show better growth during winter and when pH 9.09.

Mol Genet Genomics, 2002 Oct, 268(2), 240 - 8 Epub 2002 Aug 15.
Identification and characterization of the non-PTS fru locus of Bacillus megaterium ATCC 14581; Chiou CY et al.; A genetic locus that is adjacent to the gene encoding the small acid-soluble protein SASP C-4 of Bacillus megaterium has been identified . This locus, designated fru, contains a beta-fructosidase gene (fruA), a gene encoding a hydrophobic protein that is closely related to non-PTS sugar permeases of the proton symport type (fruP), and a gene coding for a transcriptional regulator of the LacI/GalR family (fruR) . The FruA protein can hydrolyze sucrose and raffinose, but not maltose, isomaltose, trehalose, melibiose or lactose . The transcription initiation site of fruP has been mapped and the fruP promoter identified . Gel mobility shift assays showed that the FruR protein can bind specifically to a DNA fragment containing the fruP promoter region . DNase I footprinting analysis has defined the FruR binding site . Disruption of fruR led to high-level constitutive expression of fruPA, but had no effect on expression from the fruR promoter itself, indicating that FruR acts as a repressor of fruPA expression, but does not autoregulate its own synthesis . Interestingly, expression of fruPA in B . megaterium was not induced by sucrose, raffinose, fructose or inulin, whereas the constitutive expression of fruPA in a fruR mutant was repressed by both glucose and sucrose . Possible physiological implications of these findings are discussed.

J Urol, 2002 Nov, 168(5), 1964 - 70
Intravesical bacillus Calmette-Guerin reduces the risk of progression in patients with superficial bladder cancer: a meta-analysis of the published results of randomized clinical trials; Sylvester RJ et al.; PURPOSE: We determine if intravesical bacillus Calmette-Guerin (BCG) reduces the risk of progression after transurethral resection to stage T2 disease or higher in patients with superficial (stage Ta, T1 or carcinoma in situ) bladder cancer . MATERIALS AND METHODS: A meta-analysis was performed of the published results of randomized clinical trials comparing transurethral resection plus intravesical BCG to either resection alone or resection plus another treatment other than BCG . RESULTS: We identified 24 trials with progression information on 4,863 patients . Based on a median followup of 2.5 years and a maximum of 15 years, 260 of 2,658 patients on BCG (9.8%) had progression compared to 304 of 2,205 patients in the control groups (13.8%), a reduction of 27% in the odds of progression on BCG (OR 0.73, p = 0.001) . The percent of patients with progression was low (6.4% of 2,880 patients with papillary tumors and 13.9% of 403 patients with carcinoma in situ, reflecting the short followup and relatively low risk patients entered in many of the trials . The size of the treatment effect was similar in patients with papillary tumors and in those with carcinoma in situ . However, only patients receiving maintenance BCG benefited . There was no statistically significant difference in treatment effect for either overall survival or death due to bladder cancer . CONCLUSIONS: Intravesical BCG significantly reduces the risk of progression after transurethral resection in patients with superficial bladder cancer who receive maintenance treatment . Thus, it is the agent of choice for patients with intermediate and high risk papillary tumors and those with carcinoma in situ.

FEMS Microbiol Lett, 2002 Sep 24, 215(1), 163 - 8
Microbial survival of space vacuum and extreme ultraviolet irradiation: strain isolation and analysis during a rocket flight; Saffary R et al.; We have recovered new isolates from hot springs, in Yellowstone National Park and the Kamchatka Peninsula, after gamma-irradiation and exposure to high vacuum (10(-6) Pa) of the water and sediment samples . The resistance to desiccation and ionizing radiation of one of the isolates, Bacillus sp . strain PS3D, was compared to that of the mesophilic bacterium, Deinococcus radiodurans, a species well known for its extraordinary resistance to desiccation and high doses of ionizing radiation . Survival of these two microorganisms was determined in real and simulated space conditions, including exposure to extreme UV radiation (10-100 nm) during a rocket flight . We found that up to 15 days of desiccation alone had little effect on the viability of either bacterium . In contrast, exposure to space vacuum ( approximately 10(-6) Pa) decreased cell survival by two and four orders of magnitude for Bacillus sp . strain PS3D and D . radiodurans, respectively . Simultaneous exposure to space vacuum and extreme UV radiation further decreased the survival of both organisms, compared to unirradiated controls . This is the first report on the isolated effect of extreme UV at 30 nm on cell survival . Extreme UV can only be transmitted through high vacuum, therefore its penetration into the cells may only be superficial, suggesting that in contrast to near UV, membrane proteins rather than DNA were damaged by the radiation.

FEMS Microbiol Lett, 2002 Sep 24, 215(1), 109 - 114
Interaction between Cry9Ca and two Cry1A delta-endotoxins from Bacillus thuringiensis in larval toxicity and binding to brush border membrane vesicles of the spruce budworm, Choristoneura fumiferana Clemens; Pang AS et al.; A genetically altered variant of Cry9Ca from Bacillus thuringiensis shows high potency against the spruce budworm, Choristoneura fumiferana Clemens . Its activity, as measured by feeding inhibition in frass-failure assays, is estimated to be four to seven times greater than B . thuringiensis subsp . kurstaki HD-1, the strain currently used in commercial products to control this insect . Bioassays against budworm of mixtures of the modified Cry9Ca and two of the Cry1A endotoxin proteins produced by HD-1 show neither synergism nor antagonism . Experiments with brush border membrane vesicles from budworm midgut revealed that Cry9Ca and the Cry1A toxins share a common binding site and that bound Cry9Ca can be displaced from the membrane to some extent by the Cry1A toxins . However, it is uncertain whether the binding site is actually the receptor molecule or a membrane protein associated with pore formation.

FEMS Microbiol Lett, 2002 Sep 24, 215(1), 47 - 51
Pathogenic potential of fifty Bacillus weihenstephanensis strains; Stenfors LP et al.; The aim of this study was to evaluate the food poisoning potential of strains of the new species in the Bacillus cereus group, B . weihenstephanensis . Fifty strains were tested for cytotoxicity in a Vero cell assay, and 23 of the strains were also tested for production of enterotoxin components with commercial antibody kits, and for presence of enterotoxin gene components by polymerase chain reaction (PCR) . The majority of the strains (72%) were not cytotoxic, although all of the strains that were tested with PCR and commercial kits had part of at least one of the B . cereus enterotoxins Hbl, Nhe or CytK.

Control Clin Trials, 2002 Oct, 23(5), 540 - 53
Design of the Brazilian BCG-REVAC trial against tuberculosis: a large, simple randomized community trial to evaluate the impact on tuberculosis of BCG revaccination at school age; Barreto ML et al.; This paper describes the design and baseline results of a large and simple randomized controlled trial of the protection against tuberculosis of a dose of Bacillus Calmette Guerin (BCG) vaccination given to school children in a population with a high coverage of neonatal BCG (The Brazilian BCG-REVAC trial) . The study started in 1996 and is a pair-matched and stratified-cluster randomized controlled trial with no placebo . The study population consists of children aged 7-14 years enrolled in 763 state schools from the cities of Salvador and Manaus, Brazil . Schools were the unit of randomization . Identifying information was collected for 354,708 school children . The final study population, after exclusions on the basis of age, BCG scar readings and absence from school on the day of the study visit, consists of 242,401 children, of whom 125,403 are in intervention schools . Follow-up relies on ascertainment of cases diagnosed at the health services and notified to the tuberculosis control program surveillance system . Blindness is guaranteed during linkage and validation of cases . Analysis is planned for the next 12 months, where efficacy will be estimated by calculating incidence of tuberculosis in the vaccine and control groups, taking into consideration the cluster design . The intervention studied, a second BCG vaccination, is widely used, although the World Health Organization does not recommend it on the basis of absence of evidence of protection or lack of protection . The results of the trial will make it possible for BCG revaccination practice to be informed by evidence . This is an example of a large simple and relatively inexpensive effectiveness trial, resulting from good collaboration between academia and health and education services enabling developing countries to define policies that are relevant for their reality .

Microb Cell Fact . 2002 Sep 12;1(1):3.
Optimisation of batch culture conditions for cyclodextrin glucanotransferase production from Bacillus circulans DF 9R; Rosso AM et al.; BACKGROUND: The extracellular enzyme cyclodextrin glucanotransferase (CGTase) synthesizes cyclic malto-oligosaccharides called cyclodextrins (CDs) from starch and related alpha-1,4-glucans . CGTases are produced by a variety of bacteria, mainly Bacillus species, by submerged culture in complex medium . CGTases differ in the amount and types of CDs produced . In addition, CGTase production is highly dependent on the strain, medium composition and culture conditions . Therefore we undertook this study with a newly isolated strain of Bacillus circulans . RESULTS: CGTase activity produced from Bacillus circulans DF 9R was optimised in shake flasks using a combination of conventional sequential techniques and statistical experimental design . Effects of nutrients, including several carbon, nitrogen and mineral sources, were assayed . The selected minimal medium consisted of 1.5 % cassava starch, 0.4 % ammonium sulphate, 0.1 M phosphate buffer, 0.002 % MgSO4 and 0.002 % FeSO4 . The optimal concentrations of carbon and nitrogen sources were determined using a central composite design . Maximum CGTase activity obtained in supernatants was 5.8 U/mL in 48 h of incubation . Optimal conditions for enzyme production also included an initial pH of 8.3 and 37 degrees C as the incubation temperature.Cell growth and CGTase production profile were not linked to each other, suggesting that enzyme production/secretion is not growth-associated but mainly a late-log phase event . CONCLUSION: We have screened conditions for optimal CGTase production . The selected minimal medium contained starch, ammonium, Mg2+ and Fe2+ as essential nutrients . As an additional advantage, this medium does not require complex nitrogen sources with varying and unknown composition.

J Appl Microbiol, 2002, 93(5), 772 - 80
Isolation and characterization of a novel Bacillus strain from coffee phyllosphere showing antifungal activity; Nair JR et al.; AIMS: The isolation and characterization of a novel coffee-associated Bacillus mojavensis strain, designated as strain AB1, and its survival on the coffee phyllosphere . METHODS AND RESULTS: A pair of 16S rDNA primers was designed to amplify a highly variable region within the 16S rDNA gene of Bacillus spp., with the purpose of identifying the AB1 isolate through PCR and sequence analysis . By this method, AB1 was identified as a strain of B . mojavensis . Bioassays were carried out to characterize the broad spectrum antifungal activity of AB1 . Plant colonization studies revealed that AB1 could colonize the coffee phyllosphere better than Bacillus thuringiensis . CONCLUSIONS: These studies suggest that AB1 could be a new strain of B . mojavensis . AB1 is also shown to have antifungal activity against a wide spectrum of pathogenic fungi . The antifungal metabolite of AB1 has been partially characterized as a thermostable, protease- and alkali-resistant substance that is secreted into the surrounding medium . SIGNIFICANCE AND IMPACT OF THE STUDY: As far as is known, this is the first strain of B . mojavensis which has been identified as inhabiting the coffee phyllosphere . The study highlights the potential use of AB1 as an antifungal agent in the coffee crop and as a delivery agent of the insecticidal toxin of B . thuringiensis to the coffee phyllosphere . The 16S rRNA identification strategy discussed could also be used in the identification of other new Bacillus strains.

C R Biol, 2002 Aug, 325(8), 851 - 3; discussion 879-83
{The plague}; Carniel E; The plague has been one of the most devastating diseases of human history . Despite major advances in diagnosis, prevention, and treatment, it has not been possible to eradicate this infection . Plague is still active in Africa, in Asia and in the Americas, and is classified as a currently re-emerging disease . The plague is mainly a disease of rodents, which is transmitted by fleabites . Humans develop two main clinical forms: bubonic plague (following bites of infected fleas, lethal in 50-70% of the cases in less than a week if an appropriate treatment is not started rapidly), and pneumonic plague (after inhalation of infected droplets, lethal in less than three days in 100% of cases without immediate treatment) . Y . pestis, the causative agent of plague, is usually sensitive to most antibiotics, but the first multi-resistant strain was recently described . No efficient and safe vaccines are currently available . The plague bacillus is one of the few organisms that could be used for biological warfare.

Prikl Biokhim Mikrobiol, 2002 Sep-Oct, 38(5), 552 - 5
{Induced phytophthora resistance in transgenic potato tubers}; Ozeretskovskaia OL et al.; Resistance of transgenic cultivars based on the expression of one or more resistance genes is sooner or later broken by pathogens whose race-producing rates are high . Thus, combining transgenesis with elicitor-induced resistance is a promising approach . The elicitor-induced resistance is based on the expression of multiple resistance genes, which can prevent the adaptation of pathogens to transgenic races, maintain the stability of cultivars, and increase their lifespan . In this work, we used transgenic potato cultivars Temp and Superior transformed with Bacillus thuringiensis delta-endotoxin gene and Lukyanovskii transformed with leukocyte alpha-interferon gene . Arachidonic acid (10(-8) M) and soluble chitosan (5 kDa, 100 micrograms/ml) were used as elicitors for tuber treatment . Our data showed that pretreatment with elicitors causes a 15-25% increase in both the systemic prolonged resistance of potato tubers to Phytophthora infestans and their ability to repair mechanical damage.

Prikl Biokhim Mikrobiol, 2002 Sep-Oct, 38(5), 540 - 7
{Features of the kinetics of accumulating entomocidal exotoxins of Bacillus thuringiensis subsp . thuringiensis during fermentation stage of bitoxibacillin production}; Efimtsev EI et al.; Accumulation of Bacillus thuringiensis subsp . thuringiensis beta-exotoxin (BET) in the course of industrial fermentation (a stage in the production of the entomocidal biopreparation bitoxibacillin) has been studied . It has been demonstrated in model experiments that the decrease in the content of BET in the culture fluid is accounted for by the toxin interaction with an attendant product, the exogenous metabolite (EM) . EM has been isolated from the culture fluid and characterized . EM causes alkalization of the medium, exerts entomocidal effects (in Musca domestica) and fails to form salts on treatment with BaCl2 . The absorption spectrum of EM is similar to that of BET, showing a maximum at lambda = 259 nm . The light-absorbing chromophore is a pyrimidine or purine base . A method for quantitative determination of both exotoxins (BET and EM) in bacterial preparations has been developed.

Prikl Biokhim Mikrobiol, 2002 Sep-Oct, 38(5), 502 - 8
{Preparation of an active strain of Bacillus licheniformis--producer of thermostable alpha-amylase}; Tsurikova NV et al.; A highly potent strain of Bacillus licheniformis 103 that synthesized thermostable alpha-amylase with temperature and pH optima of 90-95 degrees C and 6.0-8.5, respectively, was obtained by mutagenesis and selection . The composition of fermentation media and conditions for submerged cultivation of the producer were optimized . alpha-Amylase whose activity reached 260 U/ml was obtained in laboratory fermenters.

Cell Microbiol, 2002 Sep, 4(9), 557 - 69
Activation of Rac, Cdc42 and other downstream signalling molecules by Bartonella bacilliformis during entry into human endothelial cells; Verma A et al.; Bartonella bacilliformis is an intracellular bacterial pathogen of human endothelial cells . In vitro incubation of B . bacilliformis with human endothelial cells leads to the formation of filamentous actin extensions (filopodia) within 30 min, followed by formation of membrane rufflings or lamellipodia within 1 h of incubation . By immunofluorescence, F-actin phalloidin staining and anti-Rac antibodies were shown to co-localize in the membrane rufflings, indicating the recruitment of activated Rac at lamellipodia . Preincubation of endothelial cells with the Clostridial toxin, TcdB-10463, which inactivates the Rho-family GTPases, Rho, Rac and Cdc42, inhibited the entry of B . bacilliformis by 50-90% . Preincubation of endothelial cells with the Clostridial toxin, TcsL-1522, which specifically inactivates Rac and, to a lesser extent, Cdc42, but not Rho, inhibited entry by 30-40% . A 3.4-5.0-fold increase in activated (GTP-bound) -intracellular Rac and Cdc42 was observed in affinity precipitation assays . Increased kinase activity of p21-activated kinase (PAK), a specific downstream effector of activated Rac/Cdc42 was also observed during the time course of infection . Activation of SAPK/JNK-1 and 2, and p38 MAPKs in signalling pathways, was also detected during infection with Bartonella, as was increased binding activity of AP-1 transcription factor.

Biochemistry, 2002 Oct 29, 41(43), 12967 - 74
Reversible ligand-induced dissociation of a tryptophan-shift mutant of phosphofructokinase from Bacillus stearothermophilus; Riley-Lovingshimer MR et al.; The biophysical properties of a tryptophan-shifted mutant of phosphofructokinase from Bacillus stearothermophilus (BsPFK) have been examined . The mutant, designated W179Y/Y164W, has kinetic and thermodynamic properties similar to the wild-type enzyme . A 2-fold decrease in kcat is observed, and the mutant displays a 3-fold smaller K(0.5) for the substrate, fructose-6-phosphate (Fru-6-P), as compared to the wild-type enzyme . The dissociation constant for the inhibitor, phospho(enol)pyruvate (PEP), increases 2-fold, and the coupling parameter, Q(ay), decreases 2-fold . This suggests that while the mutant displays a slightly decreased affinity for PEP, PEP is still an effective inhibitor once bound . The new position of the tryptophan in W179Y/Y164W is approximately 6 A from the Fru-6-P portion of the active site . A 25% decrease in fluorescence intensity is observed upon Fru-6-P binding, and an 80% decrease in fluorescence intensity is observed with PEP binding . In addition, the intrinsic fluorescence polarization increases from 0.327 +/- 0.001 to 0.353 +/- 0.001 upon Fru-6-P binding, but decreases to 0.290 +/- 0.001 when PEP binds . Most notably, the presence of PEP induces dissociation of the tetramer . Dissociation of the tetramer into dimers occurs along the active site interface and can be monitored by the loss in activity or the loss in tryptophan fluorescence that is observed when the enzyme is titrated with PEP . Activity can be protected or recovered by incubating the enzyme with Fru-6-P . Recovery of activity is enzyme concentration dependent, and the rate constant for association is 6.2 +/- 0.3 M(-1) x s(-1) . Ultracentrifugation experiments revealed that in the absence of PEP the mutant enzyme exists in an equilibrium between the dimer and tetramer forms with a dissociation constant of 11.8 +/- 0.5 microM, while in the presence of PEP the enzyme exists in equilibrium between the dimer and monomer forms with a dissociation constant of 7.5 +/- 0.02 microM . A 3.1 A crystal structure of the mutant enzyme suggests that the amino acid substitutions have not dramatically altered the tertiary structure of the enzyme . While it is clear that wild-type BsPFK exists as a tetramer under these same conditions, these results suggest that quaternary structural changes probably play an important role in allosteric communication.

Arch Tierernahr, 2002 Feb, 56(1), 23 - 31
Investigations on genetically modified maize (Bt-maize) in pig nutrition: chemical composition and nutritional evaluation; Reuter T et al.; The objective of the present study was to determine the composition and the nutritional value of parental and transgenic maize seeds fed to pigs . The parental maize line was genetically modified to incorporate a gene from Bacillus thuringiensis (Bt) expressing a toxin against the European corn borer (Ostrinia nubilalis) . Both (parental and transgenic) maize lines were analyzed for crude nutrients, starch, sugar, non-starch polysaccharides (NSP), amino acids, fatty acids, as well as for selected minerals . Furthermore, four complete diets were mixed and were analyzed for the same nutrients and some selected ingredients . The diets contained 70% maize to attain a high effect level . To evaluate the feeding value of one variety of genetically modified maize (transgenic) compared to the feeding value of the unmodified maize (parental) line, a balance study with twelve pigs was designed . Three collecting periods were used for each maize line each with six animals . The collected faeces were analyzed for crude nutrients . All measured parameters were virtually the same (e.g . crude protein 11.59% vs . 11.06% in DM), especially the digestibility of crude protein (85.8 +/- 2.3% vs . 86.1 +/- 1.8%), the amount of nitrogen-free-extract (92.8 +/- 0.6% vs . 93.2 +/- 0.6%) and the metabolizable energy (15.7 +/- 0.2% vs . 15.8 +/- 0.2% MJ/kg DM) for both maize lines . Compared to the parental line, the chemical composition and digestibility of crude nutrients and energy content were not significantly affected by the genetic modification of maize . Therefore, from the view of a nutritional assessment, the genetically modified maize can be regarded as substantially equivalent to the parental maize line.

J Biochem Mol Biol Biophys, 2002 Oct, 6(5), 365 - 9
Cloning of high activity xylanase gene from Bacillus pumilus PJ19; Hamzah A et al.; The xylanase gene from Bacillus pumilus PJ19 amplified by polymerase chain reaction (PCR) was cloned into pCRII vector and transformed into Escherichia coli strain INValphaF' . Starting from an ATG as an initiator codon, an open reading frame coding for 202 amino acids was obtained . The recombinant xylanase sequence showed a 96% homology with the xylanase sequence from B . pumilus IPO strain and had an estimated molecular weight of 22,474 . Xylanase activity expressed by E . coli INValphaF' harboring the cloned gene was located primarily in the cytoplasmic fraction.

J Biochem Mol Biol Biophys, 2002 Oct, 6(5), 325 - 34
Characterization of alkaline thermoactive cellulase-free xylanases from alkalophilic Bacillus (NCL 87-6-10); Balakrishnan H et al.; Two alkaline xylanases designated as "A" and "C", respectively, were isolated from the culture filtrates of the alkalophilic Bacillus grown on a wheat bran-yeast extract medium . The two xylanases occurred in the culture filtrate in a ratio of 10:90 . These xylanases were purified to homogeneity on a CM-Sephadex matrix followed by further separation of Xylanase "A" on a phenyl sepharose column and preparative electrophoresis . The two xylanases differed considerably in their physico-chemical properties, kinetics and in their mode of action . Xylanase "C" had a molecular weight of 25,000 as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis and was a cationic protein with a pI of 8.9 . In contrast xylanase "A" had a molecular weight of 45,000 with a pI of 5.3 . The two xylanases showed distinct differences in their hydrolysis pattern . Xylanase "A" produced comparatively larger amounts of small molecular weight oligosaccharides and xylose namely xylotriose (X(3)), xylobiose (X(2)) and xylose even in the initial stages of hydrolysis (2 and 5 h) while xylanase "C" produced negligible amounts of X(2) and no xylose for the same period of incubation . At 24 h only traces of xylose was produced by xylanase "C" while substantial amounts of the monomer was produced by xylanase A in 24 h . Xylanase "A" had a broad pH optimum ranging from pH 6.0-10.0 at 40-60 degrees C while xylanase "C" had an optimum pH of 8.0 at 40-60 degrees C . Xylanases "A" and "C" differed in their K(m) and V(max) values . Xylanase "A" had a K(m) of 1.67 mg/ml and a V(max) of 3.85 x 10(2) micromol/ml/min, whereas xylanase "C" had a K(m) of 10 mg/ml and a V(max) of 1.43 x 10(4) micromol/ml/min.

Transfusion, 2002 Aug, 42(8), 1027 - 31
Detecting bacteria in platelet concentrates by use of reagent strips; Werch JB et al.; BACKGROUND: Iatrogenic infection of immunosuppressed or immunocompromised hosts secondary to receipt of blood components containing bacteria may result in serious adverse outcomes . Measurement of pH and glucose by use of inexpensive reagent strips has been proposed as a practical means of screening for bacteria in platelet concentrate (PC) units . STUDY DESIGN AND METHODS: Glucose and pH were measured in 3093 PC units by use of reagent strips (Multistix, Bayer Corp.) to screen for bacterial content . Any PC classified by the reagent strip method as containing bacteria was subsequently cultured to confirm the presence and quantity of bacteria present . RESULTS: Thirty of 3093 PC units were classified as containing bacteria by the reagent strip method . Two of the 30 PC units positive by the reagent strip method were also positive by standard bacterial culture technique . Bacillus cereus was isolated from both units . CONCLUSION: Screening PC units by the reagent strip method resulted in 9.7 units per 1000 being wasted, but prevented two patients from receiving a PC unit containing B . cereus.

Am J Pharmacogenomics, 2002, 2(3), 189 - 96
Comparative genomics in the fight against tuberculosis: diagnostics, epidemiology, and BCG vaccination; Mostowy S et al.; Although the causative agent of tuberculosis, Mycobacterium tuberculosis, has been known for some 120 years, the disease continues to plague humanity . In 1998, the sequencing of M . tuberculosis H37Rv enabled tuberculosis researchers to draw comparisons between it and other species of the closely-related M . tuberculosis complex, including bacillus Calmette-Guerin (BCG), the vaccine administered to prevent human tuberculosis . These efforts have uncovered genomic variability that potentially encodes the discrepant phenotypes displayed by species . Due to the infrequency of single nucleotide polymorphisms (SNPs) and other modes of genomic change, large sequence polymorphisms (LSPs) have presented themselves as the most obvious form of genomic variability among species . This review discusses genomic polymorphism among species of the M . tuberculosis complex as revealed through comparative genomics . Attention is drawn towards the impact of comparative genomics in generating several exciting hypotheses towards diagnosis, epidemiology, and prevention of tuberculosis disease.

Int J Food Microbiol, 2002 Nov 15, 79(1-2), 65 - 73
Acid tolerance response is low-pH and late-stationary growth phase inducible in Bacillus cereus TZ415; Jobin MP et al.; The acid tolerance of foodborne pathogen Bacillus cereus TZ415 was examined . B . cereus was more tolerant to an acid challenge at pH 4.0 when cells were grown at low pH in regulated batch cultures of rich J Broth (JB) medium . The pH-inducible acid tolerance response (ATR) was maximal at pH 5.0, a sublethal growth condition inducing a remarkable cell elongation . During growth at regulated pH 7.0 and 6.0, B . cereus TZ415 became more acid sensitive from lag to stationary growth phase and the acid tolerance of cells reached its maximum level in late-stationary growth phase . The ATR induced at pH 5.5 and 5.0 was not affected by growth phase . Cellular protein profiles were analysed as a function of growth phase and medium pH . The Hemolysin BL (HBL) enterotoxin was only detected when cells were grown at pH 7.0.

Int J Food Microbiol, 2002 Nov 15, 79(1-2), 17 - 26
Analysis of the heat-adaptive response of psychrotrophic Bacillus weihenstephanensis; Periago PM et al.; The heat-adaptive response of the psychrotrophic spoilage bacterium Bacillus weihenstephanensis DSM11827 is described . It is demonstrated that vegetative cells of B . weihenstephanensis adapts to heat exposure at 47 degrees C by prior exposure to heat at the nonlethal temperature of 38 degrees C . For this adaptive response, protein synthesis is required and maximum adaptation was noted after 15 min to 2 h prior exposure at 38 degrees C . By using two-dimensional gel electrophoresis (2D-E), an overview of the heat-shock proteins (HSPs) of B . weihenstephanensis was obtained and it was shown that the production of 15 proteins increased upon exposure to 38 degrees C . In more detail, the use of specific antibodies revealed induction of the HSPs DnaK, DnaJ, GroEL, ClpC, ClpP and ClpX of B . weihenstephanensis . In addition, also pre-exposure to other stresses than heat, such as exposure to a high salt concentration, low pH, a high ethanol concentration or low temperature, resulted in development of increased heat tolerance of B . weihenstephanensis, and during these conditions, an increased production of some HSPs was noted . This phenomenon of cross-protection might be of substantial importance in relation to the design of safe minimal processing regimes.

Ann N Y Acad Sci, 2002 Oct, 969, 97 - 101
Global analysis of Brucella melitensis proteomes; Mujer CV et al.; Brucella melitensis is a facultative, intracellular, gram-negative cocco-bacillus that causes Malta fever in humans and brucellosis in animals . There are at least six species in the genus, and the disease is classified as zoonotic because several species infect humans . Using 2-D gel electrophoresis and mass spectrometry, we have initiated (i) a comprehensive mapping and identification of all the expressed proteins of B . melitensis virulent strain 16M, and (ii) a comparative study of its proteome with the attentuated vaccinal strain Rev 1 . Comprehensive proteome maps of all six Brucella species will be generated in order to obtain vital information for vaccine development, identification of pathogenicity islands, and establishment of host specificity and evolutionary relatedness.

J Microbiol Immunol Infect, 2002 Sep, 35(3), 152 - 8
Prevention of Der p2-induced allergic airway inflammation by Mycobacterium-bacillus Calmette Guerin; Tsai JJ et al.; Epidemiologic studies suggest an inverse correlation between infection and development of allergy . The purpose of this study was to test the hypothesis whether a preexisting T helper 1 (Th1)-type immune response elicited by Mycobacterium bovis-bacillus Calmette-Guerin (BCG) immunization could suppress allergic airway inflammation induced by the mite allergen Dermatophagoides pteronyssinus group 2 (Der p2) in an animal model . C57BL/6 mice were immunized with subcutaneous injection of BCG, then intraperitoneal Der p2 emulsified in alum . Der p2-specific immunoglobulin G1 and cytokine production from splenocytes were measured after Der p2 sensitization, and pulmonary function and airway inflammation were determined after inhalation challenge with Der p2 . The intraperitoneal Der p2 with alum injection was able to induce Der p2-specific immunoglobulin G1 production, which could be downregulated by the pretreatment with BCG + Der p2 . The inoculation of BCG + Der p2 caused splenocytes to produce more interferon-gamma, and this level was higher than that elicited by Der p2 or buffer alone . The positive interferon-gamma-staining CD4 cells were also increased after activation by phorbol myristate acetate and ionomycin . Lung pathology examination found decreased airway inflammation (associated with the best pulmonary function and least airway desquamation) in the mice inoculated with BCG + Der p2 . In this Der p2-induced allergy model, BCG inoculation with Der p2 can cause a Th1-type immune response that hinders Der p2-induced allergic sensitization and the development of airway inflammation.

Chest, 2002 Oct, 122(4), 1467 - 70
Feasibility and value of video-assisted thoracoscopic surgery wedge excision of small pulmonary nodules in patients with malignancy; Burdine J et al.; PURPOSE: Advances in CT scanning have presented physicians with the challenge of diagnosing small (< 10 mm) or deep (> 5 mm) pulmonary nodules (SmPNs) in patients with known malignancies during workup or follow-up . Wedge excision of SmPNs is difficult with video-assisted thoracoscopic surgery (VATS) and often requires the performance of a thoracotomy . The value of the early detection of metastatic disease must be weighed against the morbidity (ie, thoracotomy) that is necessarily involved in obtaining the information . Little is known about the incidence of metastases in this subset of patients . We describe a VATS technique that allows the reliable excisional biopsy of SmPNs and present our findings in this patient population . METHODS: Using CT scan localization, 150 micro Ci technetium sulfur colloid is injected into the area of the pulmonary nodule . Additional blue dye is injected at the lung surface . During VATS, a sterile gamma probe is used to identify the area of radioactivity and plan placement of staple lines performed by an endostapling instrument . Palpation and the presence of radioactivity in the specimen supported the resection of the correct nodule, and CT scan findings confirmed the procedure . Between March 2000 and January 2001, 17 patients with known malignancies and SmPNs underwent VATS excisional biopsies . Six patients received a new diagnosis of malignancy, and 11 patients were in follow-up of a previously treated malignancy . The malignancies included the following: breast (four patients), head and neck (four patients), pancreas (two patients), lymphoma (two patients), lung (one patient), prostate (one patient), rectal (one patient), seminoma (one patient), and urethral (one patient) . RESULTS: All lesions were successfully resected on the first try . Nodules were removed from 10 segments and all lobes . The mean (+/-/SD) nodule size was 9.2 +/- 3.6 mm, and the mean depth was 9.4 +/- 5.2 mm . Fourteen of 17 nodules (82.4%) could be neither seen nor felt using standard VATS techniques . Diagnoses included metastatic (four patients), new primary lung cancer (one patient), acid-fast bacillus (one patient), granuloma (seven patients), carcinoid (two patients), and inflammatory pseudotumor (two patients) . Among these lesions, 29.4% were malignant, and 35.3% of patients received a diagnosis that altered their therapy . Five of 12 SmPNs (41.7%) < 10 mm in size were malignant . The median length of hospital stay was 2 days . Patients returned to full activity within 1 week . CONCLUSION: VATS excision of SmPNs after CT scan localization with radiolabeled technetium is reliable, reproducible, and associated with minimal morbidity . The technique prevented thoracotomies in 82.4% of patients . Despite the small size of these lesions, malignancy was found 29.4% of the time . This technique allows the early diagnosis of SmPNs, with low morbidity, in patients with known malignancies . Clinical implications: The reliability of this technique, the high incidence of malignancy, and the reduction in morbidity from undergoing excisional biopsy procedures will encourage the clinician to strive for earlier and more aggressive diagnoses of SmPNs.

Chest, 2002 Oct, 122(4), 1299 - 301
Tuberculin test measurement: variability due to the time of reading; Singh D et al.; STUDY OBJECTIVES: It is recommended that the PPD Mantoux tuberculin test be read at 48 or 72 h . We have compared the measurements at these time points . DESIGN: A 10-tuberculin unit (TU) PPD Mantoux test was administered to 116 healthy subjects (76.7% with bacillus Calmette-Guerin scars) . PARTICIPANTS: One hundred sixteen healthy adult volunteers were recruited (health service employees, 29 volunteers; general public, 87 volunteers) . RESULTS: The measurements made at 72 h were significantly higher than those made at 48 h (median, 9.4 vs 4.95 mm, respectively; p = 0.017) . In those subjects with induration at either or both time points (n = 69), the readings taken at 72 h were on average 1.7 mm (95% confidence interval, 0.4 to 3.0 mm) larger than those at 48 h . Using an induration of > 15 mm diameter to define a positive result, there were more positive test results at 72 h (36) compared to 48 h (28), with the results from 10 subjects (8.5%) changing because of the time of measurement . CONCLUSIONS: This study demonstrates that, in adults, the size of the 10-TU Mantoux reaction is significantly larger at 72 h compared to the reaction at 48 h . In clinical practice, tuberculin tests should be read at 72 h as negative tests at 48 h may be false results.

J Biol Chem, 2002 Dec 6, 277(49), 46849 - 51 Epub 2002 Oct 10.
Silencing of midgut aminopeptidase N of Spodoptera litura by double-stranded RNA establishes its role as Bacillus thuringiensis toxin receptor; Rajagopal R et al.; Insecticidal crystal proteins of Bacillus thuringiensis bind to receptors in the midgut of susceptible insects leading to pore formation and death of the insect . The identity of the receptor is not clearly established . Recently a direct interaction between a cloned and heterologously expressed aminopeptidase (slapn) from Spodoptera litura and the Cry1C protein was demonstrated by immunofluorescence and in vitro ligand blot interaction . Here we show that administration of slapn double-stranded RNA to S . litura larvae reduces its expression . As a consequence of the reduced expression, a corresponding decrease in the sensitivity of these larvae to Cry1C toxin was observed . The gene silencing was retained during the insect's moulting and development and transmitted to the subsequent generation albeit with a reduced effect . These results directly implicate larval midgut aminopeptidase N as receptor for Bacillus thuringiensis insecticidal proteins.

Prep Biochem Biotechnol, 2002 Aug, 32(3), 277 - 85
Screening and simple purification of alanine dehydrogenase in Bacillus strains; Baysal SH et al.; Alanine dehydrogenase (EC 1.4.1.1), in the presence of NAD+, catalyzes the reversible deamination of L-alanine . Screening of alanine dehydrogenase in bacillus strains was carried out to develop its utilization as an industrial and analytical catalyst . Eight bacillus strains were used, including Bacillus megaterium LA 199 which abundantly produces enzymes . Alanine dehydrogenase was purified simply from Bacillus megaterium LA 199 by heat treatment at pH 5.4, followed by DEAE-Sepharose CL-6B and Sepharose CL-2B chromotography . The enzyme consisted of six subunits with an identical molecular mass of 42.5 kDa . The Km were 1.17 x 10(-2) mM for NADH and 5.12 x 10(-2) mM for pyruvate.

Parasitol Res, 2002 Nov, 88(11), 979 - 83 Epub 2002 Jul 12.
The helminth community of Talpa romana (Thomas, 1902) (Insectivora, Talpidae) in southern Italy; Milazzo C et al.; The helminth parasite community of Talpa romana in Calabria (southern Italy ) was studied . The helminth fauna comprised six species: Ityogonimus ocreatus (Goeze 1782), Staphylocistis bacillaris (Goeze 1782), Capillaria talpae (Siebold 1850), Parastrongyloides winchesi (Morgan 1928), Spirura talpae (Gmelin 1790), and Tricholinstowia linstowi (Travassos 1918) . All species except S . bacillaris were dominant in this community . The helminths are all stenoxenous species of Paleartic Talpaspp . This paper is the first quantitative approach to the helminth community of T . romana and reveals typical characteristics of an isolationist community . This can be explained by genetic and paleogeographic events.

Microb Ecol, 2002 Nov, 44(4), 372 - 80 Epub 2002 Oct 14.
Comparison of eukaryotic phytobenthic community composition in a polluted river by partial 18S rRNA gene cloning and sequencing; Dorigo U et al.; We compared the species composition in phytobenthic communities at different sampling sites in a small French river presenting polluted and unpolluted areas . For each sampling point, the total DNA was extracted and used to construct an 18S rRNA gene clone library after PCR amplification of a ca 400 bp fragment . Phytobenthic community composition was estimated by random sequencing of several clones per library . Most of the sequences corresponded to the Bacillariophyceae and Chlorophyceae groups . By combining phylogenetic and correspondence analyses, we showed that our molecular approach is able to estimate and compare the species composition at different sampling sites in order to assess the environmental impact of xenobiotics on phytobenthic communities . Changes in species composition of these communities were found, but no evident decrease in the diversity . We discuss the significance of these changes with regard to the existing level of pollution and their impact on the functionality of the ecosystem . Our findings suggest that it is now possible to use faster molecular methods (DGGE, ARISA.) to test large numbers of samples in the context of ecotoxicological studies, and thus to assess the impact of pollution in an aquatic ecosystem.

Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 2002 Oct, 94(4), 494 - 8
Dialister pneumosintes can be a suspected endodontic pathogen; Siqueira JF Jr et al.; OBJECTIVE: Dialister pneumosintes is a nonmotile, nonfermentative, non-spore-forming, obligately anaerobic, gram-negative bacillus that has been associated with some infections in the human body . A species-specific nested polymerase chain reaction assay was used to investigate the occurrence of D pneumosintes in primary root canal infections . STUDY DESIGN: Samples were collected from 32 teeth with carious lesions, necrotic pulps, and radiographic evidence of periradicular bone destruction . Twenty-two teeth were asymptomatic, and 10 cases were diagnosed as acute apical periodontitis . DNA extracted from the samples was initially amplified with universal 16S ribosomal DNA primers . A second round of amplification used the first polymerase chain reaction products to specifically detect D pneumosintes . RESULTS: This bacterial species was detected in 17 of 22 asymptomatic cases (77.3%) and in 4 of 10 root canals associated with acute apical periodontitis (40%) . No relationship was found between the presence of this bacterial species and the occurrence of symptoms . In general, D pneumosintes was detected in 21 of 32 root canal samples (65.6%) . CONCLUSION: This study is the first to report a high prevalence of D pneumosintes in root canal infections of humans . Because of this high prevalence and the apparent pathogenicity of the microorganism, we suggest the inclusion of D pneumosintes in the selected group of putative endodontic pathogens.

J Bacteriol, 2002 Nov, 184(21), 5848 - 54
An ABC transporter from Bacillus thuringiensis is essential for beta-exotoxin I production; Espinasse S et al.; beta-Exotoxin I is a nonspecific insecticidal metabolite secreted by some Bacillus thuringiensis strains . Several studies of B . thuringiensis strains that have lost the capacity to produce beta-exotoxin I have suggested that there is a strong correlation between high levels of beta-exotoxin I production and the ability to synthesize crystal proteins . In this study, we showed that a mutant strain, B . thuringiensis 407-1(Cry(-))(Pig(+)), with no crystal gene, produced considerable amounts of beta-exotoxin I together with a soluble brown melanin pigment . Therefore, beta-exotoxin I production can take place after a strain has lost the plasmids bearing the cry genes, which suggests that these curable plasmids probably contain determinants involved in the regulation of beta-exotoxin I production . Using a mini-Tn10 transposon, we constructed a library of strain 407-1(Cry(-))(Pig(+)) mutants . We screened for nonpigmented mutants with impaired beta-exotoxin I production and identified a genetic locus harboring two genes (berA and berB) essential for beta-exotoxin I production . The deduced amino acid sequence of the berA gene displayed significant similarity to the ATP-binding domains of the DRI (drug resistance and immunity) family of ATP-binding cassette (ABC) proteins involved in drug resistance and immunity to bacteriocins and lantibiotics . The berB gene encodes a protein with six putative transmembrane helices, which probably constitutes the integral membrane component of the transporter . The demonstration that berAB is required for beta-exotoxin I production and/or resistance in B . thuringiensis adds an adenine nucleotide analog to the wide range of substrates of the superfamily of ABC proteins . We suggest that berAB confers beta-exotoxin I immunity in B . thuringiensis, through active efflux of the molecule.

J Am Acad Orthop Surg, 2002 Sep-Oct, 10(5), 312 - 20
Musculoskeletal manifestations of human immunodeficiency virus infection; Biviji AA et al.; Musculoskeletal manifestations of the human immunodeficiency virus (HIV) are common and are sometimes the initial presentation of the disease . Knowledge of the conditions affecting muscle, bone, and joints in HIV-infected patients is essential for successful management . Myopathies may be caused by pyogenic infection (eg, pyomyositis), idiopathic inflammation (eg, polymyositis), or drug effect (eg, AZT myopathy) . Characteristic skeletal infections, such as tuberculosis and bacillary angiomatosis, require a high index of suspicion for accurate diagnosis . Neoplastic processes, such as non-Hodgkin's lymphoma and Kaposi's sarcoma, occur more frequently as the immune system deteriorates . Inflammatory and reactive arthropathies are more prevalent in HIV-positive than HIV-negative individuals and include Reiter's syndrome, psoriatic arthritis, HIV-associated arthritis, painful articular syndrome, acute symmetric polyarthritis, and hypertrophic osteoarthropathy . Patients with atypical musculoskeletal complaints and a suspected history of exposure should be tested for HIV.

J Antibiot (Tokyo), 2002 Aug, 55(8), 707 - 14
Identification of L-glutamine: 2-deoxy-scyllo-inosose aminotransferase required for the biosynthesis of butirosin in Bacillus circulans; Tamegai H et al.; Using inverse PCR, two new genes (btrN and btrS) were identified upstream of the putative glycosyltransferase gene btrM in the butirosin-biosynthetic btr gene cluster of Bacillus circulans . The upstream gene btrS showed significant homology with stsC of Streptomyces griseus, which encodes L-glutamine:scyllo-inosose aminotransferase in the biosynthesis of streptomycin . The function of BtrS was further confirmed by heterologous expression in Escherichia coli and chemical identification of the conversion of 2-deoxy-scyllo-inosose into 2-deoxy-scyllo-inosamine . The identification of BtrS as L-glutamine:2-deoxy-scyllo-inosose aminotransferase is the first report of the aminotransferase gene responsible for 2-deoxystreptamine biosynthesis.

Clin Chest Med, 2002 Sep, 23(3), 553 - 67
Epidemiology of human pulmonary infection with nontuberculous mycobacteria; Marras TK et al.; A great deal of study has gone into the assessment of the epidemiology of NTM infection and disease in many different parts of the world . Review of the available studies provides insight into the frequency of this clinical problem as well as important limitations in current data . Study methods have varied greatly, undoubtedly leading to differing biases . In general, reported rates of infection and disease are likely underestimates, with the former probably less accurate than the latter, given that people without significant symptoms are not likely to have intensive investigations to detect infection . Pulmonary NTM is a problem with differing rates in various parts of the world . North American rates of infection and disease have been reported to range from approximately 1-15 per 100,000 and 0.1-2 per 100,000, respectively (see Table 1) . Rates have been observed to increase with coincident decreases in TB . MAC has been reported most commonly, followed by rapid growers and M kansasii . Generally similar rates have been reported in European studies, with the exception of extremely high rates in an area of the Czech Republic where mining is the dominant industry (see Table 2) . These studies have also shown marked geographic variability in prevalence . The only available population-based studies have been in South Africa and report extremely high rates of infection, three orders of magnitude greater than studies from other parts of the world (see Table 3) . This undoubtedly reflects the select population with an extremely high rate of TB and resultant bronchiectasis leading to NTM infection . Rates in Japan and Australia were similar to those reported in Europe and North America and also show significant increases over time (see Table 3) . Specific risk factors have been identified in several studies . CF and HIV, mentioned above, are two important high-risk groups . Other important factors include underlying chronic lung disease, work in the mining industry, warm climate, advancing age, and male sex . Aside from HIV and CF, mining with associated high rates of pneumoconiosis and previous TB may be the most important historically, reported in studies worldwide {63} . A recurring observation is the increase in rates of infection and disease . The reason for this is unclear but may be caused by any of several contributing factors . The possibility exists that the apparent increase is either spurious or less significant than studies would suggest . Changes in clinician awareness leading to increased investigations, or laboratory methods leading to isolation and identification of previously unnoticed organisms, could play a role in this trend, and studies have been published that support {67} and refute {31} this argument . We believe such factors may contribute to but do not explain the significant increases that have been observed . A true increase could be related to the host, the pathogen, or some interaction between the two . Host changes leading to increased susceptibility could play an important role, with increased numbers of patients with inadequate defenses from diseases such as HIV infection, malignancy, or simply advanced age {31} . An increase in susceptibility could also relate to the decrease in infection with two other mycobacteria . It has been speculated that infection with TB {29,38} and Bacillus Calmette-Guerin (BCG) {19,68} may provide cross-immunity protecting against NTM infection . Many investigations have observed decreasing rates of TB concomitant with the increases in NTM . In addition, studies from Sweden {68} and the Czech Republic {19} have found that children who were not vaccinated with BCG had a far higher rate of extrapulmonary NTM infection . Potential changes in the pathogens include increases in NTM virulence, and it has been argued that this should be considered as a possible contributing factor {69} . Finally, an interaction between the host and pathogen could involve a major increase in pathogen exposure or potential inoculum size . This may be occurring secondary to the increase in popularity of showering as a form of bathing {66}, a habit that greatly increases respiratory exposure to water contaminants . Several limitations of our review should be noted . We reviewed English-language reports and abstracts, probably leading to fewer data from non-English speaking regions, which may explain the paucity of studies from Africa, Eastern Europe, and most Asian nations . The heterogeneity of study methods in identifying cases and the lack of a uniformly applied definition of disease makes it difficult to compare rates between studies . Finally, the lack of systematic reporting of NTM infection in most nations limits the ability to derive accurate estimates of infection and disease . Regardless, there are more than adequate data to conclude that NTM disease rates vary widely depending on population and geographic location . NTM disease is clearly a major problem in certain groups, including patients with underlying lung disease and also in individuals with impaired immunity . The rates of NTM infection and disease are increasing, so the problem will likely continue to grow and become a far more important issue than current rates suggest.

Nat Biotechnol, 2002 Nov, 20(11), 1135 - 9 Epub 2002 Oct 07.
Laboratory evolution of a soluble, self-sufficient, highly active alkane hydroxylase; Glieder A et al.; We have converted cytochrome P450 BM-3 from Bacillus megaterium (P450 BM-3), a medium-chain (C12-C18) fatty acid monooxygenase, into a highly efficient catalyst for the conversion of alkanes to alcohols . The evolved P450 BM-3 exhibits higher turnover rates than any reported biocatalyst for the selective oxidation of hydrocarbons of small to medium chain length (C3-C8) . Unlike naturally occurring alkane hydroxylases, the best known of which are the large complexes of methane monooxygenase (MMO) and membrane-associated non-heme iron alkane monooxygenase (AlkB), the evolved enzyme is monomeric, soluble, and requires no additional proteins for catalysis . The evolved alkane hydroxylase was found to be even more active on fatty acids than wild-type BM-3, which was already one of the most efficient fatty acid monooxgenases known . A broad range of substrates including the gaseous alkane propane induces the low to high spin shift that activates the enzyme . This catalyst for alkane hydroxylation at room temperature opens new opportunities for clean, selective hydrocarbon activation for chemical synthesis and bioremediation.

J Pediatr Hematol Oncol, 2002 Oct, 24(7), 569 - 71
Brain abscesses resulting from Bacillus cereus and an Aspergillus-like mold; Psiachou-Leonard E et al.; An 11-year-old boy with alveolar rhabdomyosarcoma of the thigh experienced three instances of catheter-related bacteremia resulting from After two episodes of seizures, two low-density lesions in the right parietal lobe and the left corpus callosum with enhanced pericavitary opacity were detected . The catheter was removed . A brain biopsy sample grew and revealed dichotomously branched septate hyphae compatible with The patient was treated with ceftriaxone and liposomal amphotericin B for 12 and 52 weeks, respectively, until biopsy-confirmed resolution of the infections.

Microbiology, 2002 Oct, 148(Pt 10), 3223 - 33
Dissimilation of {(13)C}methanol by continuous cultures of Bacillus methanolicus MGA3 at 50 degrees C studied by (13)C NMR and isotope-ratio mass spectrometry; Pluschkell SB et al.; Using a continuous culture of Bacillus methanolicus MGA3 limited by 100 mM methanol in the feed and growing at a dilution rate D=0.25 h(-1), transients in dissolved methanol were studied to determine the effects of methanol toxicity and the pathway of methanol dissimilation to CO(2) . Steady-state cultures were disturbed by pulses of methanol resulting in a rapid change in concentration of 6.4-12.8 mM . B . methanolicus MGA3 responded to a sudden increase in available methanol by a transient decline in the biomass concentration in the reactor . In most cases the culture returned to steady state between 4 and 12 h after pulse addition . However, at a methanol pulse of 12.8 mM, complete biomass washout occurred and the culture did not return to steady state . Integrating the response curves of the dry biomass concentration over a 12 h time period showed that a methanol pulse can cause an average transient decline in the biomass yield of up to 22% . (13)C NMR experiments using labelled methanol indicated that the transient partial or complete biomass washout was probably caused by toxic accumulation of formaldehyde in the culture . These experiments also showed accumulation of formate, indicating that B . methanolicus possesses formaldehyde dehydrogenase and formate dehydrogenase activity resulting in a methanol dissimilation pathway via formate to CO(2) . Studies using isotope-ratio mass spectrometry provided further evidence of a methanol dissimilation pathway via formate . B . methanolicus MGA3, growing continuously under methanol limitation, consumed added formate at a rate of approximately 0.85 mmol l(-1) h(-1) . Furthermore, significant accumulation of (13)CO(2) in the reactor exhaust gas was measured in response to a pulse addition of {(13)C}formic acid to the bioreactor . This indicates that B . methanolicus dissimilates methanol carbon to CO(2) in order to detoxify formaldehyde by both a linear pathway to formate and a cyclic mechanism as part of the RuMP pathway.

Microbiology, 2002 Oct, 148(Pt 10), 3059 - 67
Ligation of arabinogalactan to peptidoglycan in the cell wall of Mycobacterium smegmatis requires concomitant synthesis of the two wall polymers; Hancock IC et al.; To study the late events of cell wall assembly in Mycobacterium smegmatis, specific in vivo radiolabelling of exponentially growing liquid cultures over periods of less than one cell generation were carried out . N-Acetyl-{(14)C}glucosamine was used to label peptidoglycan and {(14)C}glucose to label arabinogalactan and arabinomannan . Over periods of several generations, radioactive cell wall material was turned over as soluble autolysis products into the culture fluid . However, turnover of newly synthesized and labelled cell wall was delayed for about one cell generation, implying inside-to-outside growth of the wall as observed in BACILLUS: Little radioactive wall material was released into the culture fluid during the first generation of labelling in growing cultures, but the addition of amoxicillin plus the beta-lactamase inhibitor clavulanic acid, at the minimum inhibitory concentration of amoxicillin, led to the release of radioactive peptidoglycan that could be isolated by gel filtration chromatography and contained nearly 3 mol alanine per glutamic acid residue, indicating that it was linear, un-crosslinked peptidoglycan that had never been substantially cross-linked to the cell wall due to inhibition of transpeptidation by amoxicillin . This peptidoglycan had no covalently attached arabinogalactan . Radioactive arabinogalactan was synthesized and released from the amoxicillin-treated bacteria without attachment to peptidoglycan . The results indicate that during growth, incorporation of arabinogalactan into the cell wall requires its ligation to newly synthesized peptidoglycan and that the peptidoglycan must be undergoing concomitant cross-linking to the inner surface of the cell wall . Inhibition of peptidoglycan transpeptidation prevents ligation of arabinogalactan to peptidoglycan and its consequent incorporation into the wall.

J Mol Biol, 2002 Oct 11, 323(1), 115 - 23
Directed evolution of barnase stability using proteolytic selection; Pedersen JS et al.; We report the construction of a phage-displayed repertoire of mutants of the ribonuclease barnase from Bacillus amyloliquefaciens . The construction was guided by the natural variability between two closely related ribonucleases, barnase and binase from Bacillus intermedius . This repertoire was selected using a proteolytic selection method, allowing sorting of the library according to the resistance of the mutants toward proteolysis . Susceptibility toward proteolysis has been correlated with flexibility and unfolding, and is thus expected to yield mutants with increased thermal stability.Enrichment of barnase mutants with specific combinations of amino acid residues at four of the randomised positions was observed . Three of these enriched amino acid residues are present in neither barnase nor binase . For some of the mutations, the improvement in proteolytic stability does not lead to a pronounced improvement in thermodynamic stability, indicating that the factors governing the proteolytic stability in some cases may be different from those governing the thermodynamic stability, e.g . propensity to local unfolding.The results obtained add important knowledge to a novel use of phage display technology for selection of thermodynamically stable proteins . Only by carefully establishing the parameters that can be adjusted, and recognising the influence this will have on the outcome of selection, will it be possible to realise the powerful technique of proteolytic selection.

BMC Microbiol . 2002 Oct 07;2(1):30.
Mycosin-1, a subtilisin-like serine protease of Mycobacterium tuberculosis, is cell wall-associated and expressed during infection of macrophages; Dave JA et al.; BACKGROUND: Exported proteases are commonly associated with virulence in bacterial pathogens, yet there is a paucity of information regarding their role in Mycobacterium tuberculosis . There are five genes (mycP1-5) present within the genome of Mycobacterium tuberculosis H37Rv that encode a family of secreted, subtilisin-like serine proteases (the mycosins) . The gene mycP1 (encoding mycosin-1) was found to be situated 3700 bp (four ORF's) from the RD1 deletion region in the genome of the attenuated vaccine strain M . bovis BCG (bacille de Calmette et Guerin) and was selected for further analyses due to the absence of expression in this organism . RESULTS: Full-length, 50 kDa mycosin-1 was observed in M . tuberculosis cellular lysates, whereas lower-molecular-weight species were detected in culture filtrates . A similar lower-molecular-weight species was also observed during growth in macrophages . Mycosin-1 was localized to the membrane and cell wall fractions in M . tuberculosis by Western blotting, and to the cell envelope by electron microscopy . Furthermore, M . tuberculosis culture filtrates were shown to contain a proteolytic activity inhibited by mixed serine/cysteine protease inhibitors and activated by Ca2+, features typical of the subtilisins . CONCLUSIONS: Mycosin-1 is an extracellular protein that is membrane- and cell wall-associated, and is shed into the culture supernatant . The protein is expressed after infection of macrophages and is subjected to proteolytic processing . Although proteolytically active mycosin-1 could not be generated recombinantly, serine protease activity containing features typical of the subtilisins was detected in M . tuberculosis culture filtrates.

Rheumatology (Oxford), 2002 Oct, 41(10), 1119 - 25
Reactive arthritis after BCG immunotherapy: T cell analysis in peripheral blood and synovial fluid; Bartolome Pacheco MJ et al.; OBJECTIVE: To investigate the pathogenic mechanism of reactive arthritis after instillation of Calmette-Guerin bacillus (BCG) . Although the clinical features of reactive arthritis after BCG therapy are well described, only a few reports have studied the possible pathogenic mechanisms . METHODS: We analysed by flow cytometry the phenotype and T-cell receptor (TCR) expression of peripheral blood (PB) and synovial fluid (SF) T cells in a patient who developed reactive arthritis (ReA) following intravesical BCG immunotherapy for bladder cancer . The proliferative response of short-term T-cell lines (TCL) from PB of this patient to mycobacterial antigens was tested by bromodeoxyuridine incorporation . RESULTS: CD4(+) and CD8(+) SF T cells with activated and memory phenotype were observed at the onset of arthritis . We were able to detect BV-restricted expansion of CD8(+) T cells in PB (BV17) and in SF (BV5S1 and BV12) . The percentage of PB and SF CD8(+) T cells that expanded diminished when the symptoms remitted . The strongest response of CD4(+) TCL from the patient in vitro was obtained for human hsp-60 in an inversely dose-dependent manner . Very important was the finding that CD8(+) TCL from the patient demonstrated no proliferative response to any antigenic challenge that was reversed after the addition of exogenous interleukin 2 . CONCLUSION: Although the identity of the stimulating antigen that led to the expansions observed in this patient is not clarified by the present data, both CD4(+) and CD8(+) T cells might play a role in the development of ReA following intravesical administration of BCG.

Biotechnol Prog, 2002 Sep-Oct, 18(5), 1087 - 94
Validation and use of an enzymic time-temperature integrator to monitor thermal impacts inside a solid/liquid model food; Guiavarc'h YP et al.; Heat denaturation kinetics of Bacillus licheniformis alpha-amylase, equilibrated at 81% equilibrium relative humidity at 4 degrees C (BLA81), was studied with help of isothermal and nonisothermal conditions by monitoring the decrease in enthalpy associated with the heat denaturation of the enzyme . Due to its low water content, BLA81 denaturation could be studied in the range of 118-124 degrees C . Two batches of BLA81 were successfully validated under nonisothermal conditions allowing the determinations of process values (reference temperature of 121.1 degrees C) in the range of 1-15 min . In a second step, BLA81 was used as a time-temperature integrator (TTI) to investigate potential differences of process values received by freely moving spherical particles as compared to a centrally fixed particle (single-position impact) inside cans containing water as brine . Results showed that the process value received by freely moving particles can be from 5.6% (4 rpm) to 19.7% (8 rpm) smaller than the process value received by the centrally fixed sphere . This means that evaluating the process value by means of a particle fixed at the critical point in a package can lead to potentially overestimations of the actual process value with possible hazardous quality/safety implications . These results highlight the potentials of the TTI technology to monitor the safety of heat-processed agitated solid/liquid foodstuffs.

Water Sci Technol, 2002, 46(4-5), 361 - 6
Ozonation of domestic secondary effluent for recycling and reuse--a pilot plant study; Ni CH et al.; In this study, a pilot plant combined ozonation with sand filtration is established . Wastewater from the secondary effluent was taken as the sample for looking into the feasibility of domestic wastewater recovery and recycling . At the beginning, a series of hydraulic analysis and ozone transfer tests was conducted in the bubble column reactor . From these tests, it was found that when the gas flow is controlled to within 0.2 to approximately 1.0 L/min and liquid flow within 0.5 to approximately 2.5 L/min, using series connected mixing tank model for our calculation, the result J (CSTR) is 1 to approximately 2 . From the ozone transfer test it is known that the smaller the gas flow, the better the transfer rate, and the same pattern occurs on ozone gas concentration . After sand filtration and ozone treatment, the G/L ratio within the ozone column reactor can be maintained within 0.2 to approximately 0.4 and the ozone dosage within 8 to approximately 12 mg/L . The removal rate for coliform bacillus, BOD, turbidity and color is 99.96%, 62.2%, 89.6% and 67% respectively . After ozonation treatment, coliform bacillus content can be controlled under 10 CFU/mL, BOD under 10 mg/L, turbidity within 2.0 to approximately 2.5 (NTU), and color within 10.3 to approximately 13.7 degree . The recycled water is almost colorless and odorless, and is capable of reaching the reference standard for recycled water.

Allergy, 2002 Nov, 57(11), 1059 - 62
Tuberculin responses in children with allergic diseases; Ozmen S et al.; BACKGROUND: The prevalence of allergic disorders has been increasing over the last 30 years, especially in developed countries . One factor associated with this rise may be the decline of many childhood infections . We investigated tuberculin responses in allergic children in order to see the development of delayed-type hypersensitivity reactions to tubercule bacillus infection . METHODS: The study sample was composed of 106 allergic and 100 nonallergic children vaccinated with bacille Calmette-Guerin (BCG) . The standard Mantoux test was applied to all children . The reactions were read after 72 h by measuring the diameter of the wheal . RESULTS: The wheal size was 6.29 +/- 5.09 mm (mean +/- SD) in allergic children, and 2.79 +/- 2.96 mm in nonallergic children . The difference between the two groups was significant (P < 0.001) . In children with a single BCG scar, the mean purified protein derivative (PPD) wheal size for allergic children was 4.77 +/- 4.79 mm, and for nonallergic children it was 2.48 +/- 3.19 mm . The mean PPD wheal sizes in allergic and nonallergic children who had been vaccinated twice were 8.35 +/- 4.80 mm and 3.33 +/- 2.44 mm, respectively . This difference was statistically significant (P < 0.05) . In 27.35% of the allergic children and 6% of the nonallergic children, the positive tuberculin responses (PPD > or = 10 mm) were recorded . The difference was significant (P < 0.05) . CONCLUSIONS: Our results showed that response to tuberculin in BCG-immunized allergic children is higher than in BCG-immunized nonallergic children.

Lett Appl Microbiol, 2002, 35(4), 311 - 5
Inhibition of adhesion of enterotoxigenic Escherichia coli K88 by soya bean tempe; Kiers JL et al.; AIMS: Tempe is a traditional fungal fermented food made from soaked and cooked soya beans . It has been associated with antidiarrhoeal characteristics . This study investigated potential inhibitory effects of tempe on enterotoxigenic Escherichia coli (ETEC) K88 . METHODS AND RESULTS: Soya beans were soaked, cooked and subsequently fermented using several Rhizopus spp . Water-soluble filter-sterile extracts were tested for their ability to inhibit growth of E . coli and several indicator microorganisms and to inhibit adhesion of ETEC K88 . Antimicrobial activity was found against Bacillus stearothermophilus only . ETEC K88-induced haemagglutination of hamster red blood cells was strongly inhibited by a number of tempe extracts and hardly by the cooked soya bean extract . Furthermore, several tempe extracts were able to inhibit adhesion of ETEC K88 to piglet small intestinal brush-border membranes . CONCLUSIONS: Tempe appeared to interfere with ETEC K88 adhesion rather than showing growth inhibitory properties . SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicate that tempe could exert an antagonistic effect against ETEC through inhibition of adhesion and might therefore have a protective effect against ETEC K88 infection in pigs . Hence, tempe could have potential to use as a feed supplement in the diet of weaned piglets.

J Agric Food Chem, 2002 Oct 9, 50(21), 6245 - 52
Angiotensin I-converting enzyme inhibitory peptides derived from wakame (Undaria pinnatifida) and their antihypertensive effect in spontaneously hypertensive rats; Sato M et al.; Seven kinds of angiotensin I-converting enzyme (ACE) inhibitory peptides were isolated from the hydrolysates of wakame (Undaria pinnatifida) by Protease S "Amano" (from Bacillus stearothermophilus) by using three-step high-performance liquid chromatography (HPLC) on a reverse-phase column . These peptides were identified by amino acid composition analysis, sequence analysis, and liquid chromatography-mass spectrometry (LC-MS), as Val-Tyr (IC(50) = 35.2 microM), Ile-Tyr (6.1 microM), Ala-Trp (18.8 microM), Phe-Tyr (42.3 microM), Val-Trp (3.3 microM), Ile-Trp (1.5 microM), and Leu-Trp (23.6 microM) . These peptides have resistance against gastrointestinal proteases in vitro . Each peptide was determined to have an antihypertensive effect after a single oral administration in spontaneously hypertensive rats (SHR) . Among them, the blood pressure significantly decreased by Val-Tyr, Ile-Tyr, Phe-Tyr, and Ile-Trp in a dose of 1 mg/kg of body weight (BW) . The present study showed that antihypertensive effect in the hydrolysates of wakame by Protease S "Amano" was attributed to these peptides.

J Genet, 2002 Apr, 81(1), 5 - 11
Cloning and characterization of an insecticidal crystal protein gene from Bacillus thuringiensis subspecies kenyae; Misra HS et al.; A sporulating culture of Bacillus thuringiensis subsp . kenyae strain HD549 is toxic to larvae of lepidopteran insect species such as Spodoptera litura, Helicoverpa armigera and Phthorimaea operculella, and a dipteran insect, Culex fatigans . A 1.9-kb DNA fragment, PCR-amplified from HD549 using cryII-gene-specific primers, was cloned and expressed in E . coli . The recombinant protein produced 92% mortality in first-instar larvae of Spodoptera litura and 86% inhibition of adult emergence in Phthorimaea operculella, but showed very low toxicity against Helicoverpa armigera, and lower mortality against third-instar larvae of dipteran insects Culex fatigans, Anopheles stephensi and Aedes aegypti . The sequence of the cloned crystal protein gene showed almost complete homology with a mosquitocidal toxin gene from Bacillus thuringiensis var . kurstaki, with only five mutations scattered in different regions . Amino acid alignment with different insecticidal crystal proteins using the MUTALIN program suggested presence of the conserved block 3 region in the sequence of this protein . A mutation in codon 409 of this gene that changes a highly conserved phenylalanine residue to serine lies in this block.

Protein Expr Purif, 2002 Oct, 26(1), 82 - 8
Cloning and expression of the vegetative insecticidal protein (vip3V) gene of Bacillus thuringiensis in Escherichia coli; Doss VA et al.; A genomic library of Bacillus thuringiensis var . kurstaki (B.t.k.) was constructed and a positive clone harboring the full-length gene encoding a novel vegetative insecticidal protein (Vip3V) was characterized . The vip3V gene was subcloned into pET-22b(+) vector and overexpressed in Escherichia coli to an extent of about 30% of the total protein . While transcription was influenced by the T7 promoter of the vector, synthesis of Vip3V in E . coli host occurred from the B.t.k . ribosomal binding site (rbs) found 917bp downstream of the insert and not from the E . coli rbs of the vector . The expressed Vip3V protein was found in the soluble and periplasmic fractions as well as in the inclusion bodies . A simplified anion-exchange chromatographic method for the purification of Vip3V using step gradient or one-step elution was developed . The purified protein showed broad-spectrum activity against some of the lepidopteran larvae tested and did not show any activity against the larvae of silkworm (Bombyx mori) and mosquito (Culex quinquefasciatus).

J Clin Microbiol, 2002 Oct, 40(10), 3641 - 7
Genetic classification and differentiation of Bartonella species based on comparison of partial ftsZ gene sequences; Zeaiter Z et al.; Currently, 19 species are recognized in the genus Bartonella, 7 of which are involved in an increasing variety of human diseases . Development of molecular tools for detection, identification, and subtyping of strains and isolates has promoted research on Bartonella spp . We amplified and sequenced the portion of the ftsZ gene encoding the N-terminal region of the cell division protein for 13 Bartonella species: Bartonella alsatica, B . birtlesii, B . doshiae, B . elizabethae, B . grahami, B . koehlerae, B . schoenbuchensis, B . taylorii, B . tribocorum, Bartonella vinsonii subsp . arupensis, Bartonella vinsonii subsp . berkhoffii, Bartonella vinsonii subsp . vinsonii, and B . bovis Bermond et al.("B . weissii") . Phylogenetically derived trees revealed four statistically supported groups, indicating that sequencing of the ftsZ gene is a useful tool for identifying evolutionary relationships among Bartonella species . Furthermore, we amplified and sequenced the portion of the ftsZ gene encoding the C-terminal region of the protein for 4 B . bacilliformis isolates, 14 B . clarridgeiae isolates, 14 B . quintana isolates, and 30 B . henselae isolates that were obtained from different geographic regions, hosts, and clinical specimens . B . clarridgeiae and B . quintana sequences were highly conserved, while those of the four B . bacilliformis isolates differed from the type strain at 5 positions . Among B . henselae strains isolated from cats and patients, only two genotypes were detected: Houston and Marseille . Among 80 clinical samples we detected Bartonella spp . in 35 (43.75%) and found the assay to be comparable to that of a combined intergenic-spacer-region- and pap31-based PCR assay . Our results show the usefulness of the portion of the ftsZ gene encoding the C-terminal region for diagnosis of Bartonella infections . More samples should be tested to study its usefulness for epidemiological investigations.

J Clin Microbiol, 2002 Oct, 40(10), 3606 - 12
Identification of Bartonella bacilliformis genotypes and their relevance to epidemiological investigations of human bartonellosis; Birtles RJ et al.; Genotypic diversity among 26 isolates of Bartonella bacilliformis obtained from different areas of Peru, and at different times, was assessed by comparison of DNA sequences derived from 16S-23S ribosomal DNA intergenic spacer regions (ISR) and a citrate synthase gene (gltA) fragment and by amplified fragment length polymorphism (AFLP) analysis . gltA comparison divided the isolates into two groups, whereas ISR comparison revealed six sequences . AFLP analysis using a selective primer delineated five profiles that correlated well with those obtained by sequence comparison . Combination of all three data sets divided the isolates into six genotypes . One of these genotypes was common to isolates collected from a large area in western Peru that corresponded to the region of endemicity for bartonellosis; however, isolates belonging to two other genotypes were also found within this region . Two of these genotypes were found in isolates isolated more than 35 years apart . The remaining three genotypes were each specifically associated with three outbreaks of bartonellosis that have recently occurred in areas where the disease had not previously been recognized . Demonstration of the unique nature of these isolates indicates that the outbreaks with which they were associated did not result from the introduction of disease by individuals who acquired their infection in the recognized region of endemicity . The sources of these outbreaks remain unknown . A consensus approach to bacterial typing using comparative sequence analysis of multiple genetic loci and the pan-genomic sampling of AFLP appears to offer a well-supported assessment of B . bacilliformis diversity, and the genotypic differences identified appear to have epidemiological significance.

Life Sci, 2002 Oct 18, 71(22), 2667 - 80
Slight influence of the estrous cycle stage on the mucosal and systemic specific antibody response induced after vaginal and intraperitoneal immunization with protoxin Cry1Ac from Bacillus thuringiensis in mice; Moreno-Fierros L et al.; Since the immune response appears to be variable according to the hormonal stage of the mammalian female, the aim of this study was to determine whether estrous cycle stage modifies the mucosal and systemic immune responses induced by intraperitoneal and vaginal immunization of mice with protoxin Cry1Ac . We tested the influence of three immunizations on the specific antibody response elicited at estrus and diestrus, that were the same estrous cycle stages at which the antigen was applied . Both intraperitoneal and vaginal immunization of mice with Cry1Ac either at estrus or diestrus induces specific antibody responses at serum, vagina and large intestine . The stage of the estrous cycle have little or non influence in the magnitude of the response induced, since only at serum the IgM was slightly higher at estrus than at diestrus by both routes . At the large intestine only the IgA response elicited via the intraperitoneal route changed, being higher at diestrus, whereas at the vagina IgA response induced did not change significantly due to the cycle stage . Present results suggest that Cry1Ac may be used as an antigen carrier as it can elicit antibody responses at systemic level and at several mucosal sites including the vagina that are not modified significantly throughout the reproductive cycle.

Dig Dis Sci, 2002 Sep, 47(9), 2045 - 8
Colonic tuberculosis; Villanueva Saenz E et al.; Tubercle bacillus was discovered in 1882 by Robert Koch . With the introduction of chemotherapy for tuberculosis in the 1940s the incidence of this entity decreased . The incidence of the tuberculosis of the colon began to increase at the 1980s with the rise in numbers of patients considered as high risk for this entity, such as HIV-infected individuals, patients with chronic renal disease, and immunosuppressed patients with prolonged steroid therapy . We report on two patients with history of chronic abdominal pain and weight loss with a palpable mass in the right lower quadrant . In one patient chest radiography revealed a miliary reticulonodular pattern . In both, abdominal CT scan showed retroperitoneal lymphadenopathy and colonic wall thickness . Colonoscopic examination showed ulcerative lesions and ileocecal valve disruption . Microscopic examination of intestinal content showed evidenced M . tuberculosis . Tuberculosis of the colon should be suspected in patients suffering from chronic abdominal pain and weight loss.

Can J Public Health, 2002 Sep-Oct, 93(5), 353 - 5
An outbreak of Bacillus cereus implicating a part-time banquet caterer; Gaulin C et al.; CONTEXT AND OBJECTIVES: In the aftermath of a party, 70% (25 of 36) of attendees had gastroenteritis . The objectives of this study were to identify a risk factor associated with the food during the banquet and to identify measures of control for avoiding this kind of outbreak in the future . METHOD: A retrospective cohort study was used . We tried to reach by telephone all guests who had attended this banquet . A standardized questionnaire was used to provide information about identification of a risk factor, especially in relation to food . RESULTS: The cohort study has shown that potato salad served at the party was significantly associated with the disease . The mayonnaise used to prepare the salad was analyzed and Bacillus cereus was isolated (10(3) bacteria per gram) . DISCUSSION: Bacillus microorganisms are usually found in decaying organic matter, dust, soil, vegetables and water . The bacteria has a remarkable ability to survive strong environmental stresses . There are strains of B . cereus that can cause food poisoning episodes with infective doses as low as 10(3) to 10(4) bacteria per gram . B . cereus is an infrequently reported cause of foodborne illnesses in Quebec and in North America but this may be due to underreporting of episodes . In this outbreak, bacterial multiplication was facilitated at several points in the interval between the preparation of the meal and the consumption of the banquet by the guests . Because the spores are ubiquitous and resistant to inactivation with most food grade disinfectants, temperature control should be the main focus of B . cereus outbreak prevention . CONCLUSION: The meal was prepared by a restaurateur who was inexperienced in catering services and temperature control in particular when food is served outside the restaurant . This outbreak underscores the importance of maintaining meticulous hygienic procedures in food processing . Restaurateurs who offer catering services should be familiar with the constraints that are specific to this sector of the food industry.

J Biol Chem, 2002 Dec 6, 277(49), 46966 - 73 Epub 2002 Sep 25.
Identification of a magnesium-dependent NAD(P)(H)-binding domain in the nicotinoprotein methanol dehydrogenase from Bacillus methanolicus; Hektor HJ et al.; The Bacillus methanolicus methanol dehydrogenase (MDH) is a decameric nicotinoprotein alcohol dehydrogenase (family III) with one Zn(2+) ion, one or two Mg(2+) ions, and a tightly bound cofactor NAD(H) per subunit . The Mg(2+) ions are essential for binding of cofactor NAD(H) in MDH . A B . methanolicus activator protein strongly stimulates the relatively low coenzyme NAD(+)-dependent MDH activity, involving hydrolytic removal of the NMN(H) moiety of cofactor NAD(H) (Kloosterman, H., Vrijbloed, J . W., and Dijkhuizen, L . (2002) J . Biol . Chem . 277, 34785-34792) . Members of family III of NAD(P)-dependent alcohol dehydrogenases contain three unique, conserved sequence motifs (domains A, B, and C) . Domain C is thought to be involved in metal binding, whereas the functions of domains A and B are still unknown . This paper provides evidence that domain A constitutes (part of) a new magnesium-dependent NAD(P)(H)-binding domain . Site-directed mutants D100N and K103R lacked (most of the) bound cofactor NAD(H) and had lost all coenzyme NAD(+)-dependent MDH activity . Also mutants G95A and S97G were both impaired in cofactor NAD(H) binding but retained coenzyme NAD(+)-dependent MDH activity . Mutant G95A displayed a rather low MDH activity, whereas mutant S97G was insensitive to activator protein but displayed "fully activated" MDH reaction rates . The various roles of these amino acid residues in coenzyme and/or cofactor NAD(H) binding in MDH are discussed.

FEMS Microbiol Lett, 2002 Sep 10, 214(2), 189 - 93
The ribulose monophosphate pathway operon encoding formaldehyde fixation in a thermotolerant methylotroph, Bacillus brevis S1; Yurimoto H et al.; The hps and phi genes encoding 3-hexulose-6-phosphate synthase and 6-phospho-3-hexuloisomerase, the key enzymes of the ribulose monophosphate (RuMP) pathway for formaldehyde fixation, were cloned from the chromosomal DNA of a thermotolerant methylotroph, Bacillus brevis S1 . Enzyme induction and Northern blot analyses revealed that both the hps and phi genes are induced by methanol or ethanol, and that their expression is controlled polycistronically at the transcription stage . Sequence analysis also suggested that the hps and phi genes constitute an RuMP operon . The gene organization of the RuMP operon and its surrounding region are unique among bacteria possessing the RuMP pathway genes.

J Med Entomol, 2002 Sep, 39(5), 729 - 35
High resistance to Bacillus sphaericus binary toxin in culex pipiens (Diptera: Culicidae): the complex situation of west Mediterranean countries; Nielsen-Leroux C et al.; This study was aimed at clarifying the nature of the resistance to Bacillus sphaericus Neide (Bs) that Culex pipiens L . has developed in west Mediterranean countries, France, and Tunisia . Two recessive and sex-linked mutants, sp-1R and sp-2R, were previously detected in southern France . Here, the Tunisian resistance was also shown to involve a single recessive and sex-linked gene that was temporarily named sp-T(R) . In addition, sp-1R, sp-2R, and sp-T(R) were shown to separately confer a similar high resistance level (> 5,000-fold) in the homozygous state . Knowing that sp-1R resistance does not alter the binding of Bs binary toxin to its specific receptor, we investigated this character in sp-2RR and sp-T(RR) homozygotes . This was performed by in vitro experiments in which larval brush border membrane fractions (BBMF) were exposed to the 125I-Bin2 toxin of B . sphaericus strain 1593 . The toxin-receptor binding was found disrupted by sp-2R but not by sp-T(R) . Comparing the binding kinetics among nine Culex pipiens strains of diverse origins revealed that the Bs receptors of sp-1RR and Sp-T(RR) homozygous larvae were displaying the highest affinity toward Bs binary toxins . These results are discussed with regard to alternative assumptions on the dynamics of high Bs-resistance and on the emerging possibilities to test them in a near future.

JK Pract, 2000 Apr-Jun, 7(2), 153 - 5
Current status of passive tuberculosis detection in a rural district of J & K; Gaash B et al.; PIP: Tuberculosis affects one-fifth of mankind and kills more people than any other infectious disease; yet misdiagnosis and under-reporting are rampant . The situation in rural areas is especially disappointing . Despite decades of a national-level program for control of tuberculosis, no improvement has occurred in detection and reporting . Analysis and comparison of the records of the District Tuberculosis Centre in a rural district have shown almost no improvement in the dismal performance over the last full decade . The failure of passive detection of cases, especially the bacillary positive cases, has suggested that active detection coupled with epidemiological investigation is needed to boost routine case detection and to find out the bottlenecks in the implementation of the National Tuberculosis Program in rural areas .

Vaccine Wkly . 1996 Jan 15;:15-6.
TB vaccines . Study finds BCG efficacious in Central African Republic; Complete sequence and organization of pBtoxis et al.; Cardiff School of Biosciences, Cardiff University, Cardiff, United Kingdom . Berry@cf.ac.uk

The entire 127,923-bp sequence of the toxin-encoding plasmid pBtoxis from Bacillus thuringiensis subsp . israelensis is presented and analyzed . In addition to the four known Cry and two known Cyt toxins, a third Cyt-type sequence was found with an additional C-terminal domain previously unseen in such proteins . Many plasmid-encoded genes could be involved in several functions other than toxin production . The most striking of these are several genes potentially affecting host sporulation and germination and a set of genes for the production and export of a peptide antibiotic.

Appl Environ Microbiol, 2002 Oct, 68(10), 4894 - 9
Inactivation of the elongation factor Tu by mosquitocidal toxin-catalyzed mono-ADP-ribosylation; Schirmer J et al.; The mosquitocidal toxin (MTX) produced by Bacillus sphaericus strain SSII-1 is an approximately 97-kDa single-chain toxin which contains a 27-kDa enzyme domain harboring ADP-ribosyltransferase activity and a 70-kDa putative binding domain . Due to cytotoxicity toward bacterial cells, the 27-kDa enzyme fragment cannot be produced in Escherichia coli expression systems . However, a nontoxic 32-kDa N-terminal truncation of MTX can be expressed in E . coli and subsequently cleaved to an active 27-kDa enzyme fragment . In vitro the 27-kDa enzyme fragment of MTX ADP-ribosylated numerous proteins in E . coli lysates, with dominant labeling of an approximately 45-kDa protein . Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry combined with peptide mapping identified this protein as the E . coli elongation factor Tu (EF-Tu) . ADP ribosylation of purified EF-Tu prevented the formation of the stable ternary EF-Tuaminoacyl-tRNAGTP complex, whereas the binding of GTP to EF-Tu was not altered . The inactivation of EF-Tu by MTX-mediated ADP-ribosylation and the resulting inhibition of bacterial protein synthesis are likely to play important roles in the cytotoxicity of the 27-kDa enzyme fragment of MTX toward E . coli.

Harv AIDS Rev . 1995 Fall;:2-9.
AIDS in India; Shreedhar J; PIP: A major HIV epidemic is underway in India, home to 900 million people and the world's second largest population . The director-general of the Indian Council of Medical Research expects India by the year 2000 to be the country with the largest number of HIV infections, with some experts predicting 5 million people to be infected with HIV in India by the year 2000 . Others predict 30-55 million to be infected . Although HIV is increasingly spreading to typically low-risk group populations, it is the female sex workers and their clients, long distance truck drivers, men who have sex with men, blood transfusion donors and recipients, and IV drug users throughout the country who are both the reservoirs of HIV and vectors of transmission to the general population . For example, 52% of sex workers in Bombay in 1994 were found to be infected with HIV . Studies indicate that India's long-distance truck drivers average 200 sexual encounters per year; at any given time, 70% of them have STDs . Preliminary surveys estimate that almost 33% are infected with HIV . HIV seroprevalence among truckers in Madras requesting HIV testing because they have STDs increased from almost 60% in 1993 to 91% in 1995 . Moreover, the illegal status of homosexuality in India has created an underground culture in which HIV and STDs are rampant; one 1995 study in the Sangli district of Maharashtra found 50% of men who have sex with men to be infected with HIV . Half of India's blood for transfusion is drawn from commercial donors . A Bombay study, however, found 86% of such donors screened in 1992 to be HIV-seropositive and not all blood banks comply with mandatory screening laws . As widespread HIV infection evolves into a multitude of AIDS cases, India's health care system and economy will be heavily taxed, and the number of tuberculosis (TB) cases greatly increased . More than half the population carries the TB bacillus . The government by 1992 had drafted a national prevention and control plan and formed the National AIDS Control Organization (NACO), but much remains to be accomplished in the prevention and control of HIV/AIDS . Nongovernmental organizations are working to strengthen government efforts .

Salubritas . 1986 Jul;9(2):3.
How to give an immunization.
{Bacterial present in the xylem of coffee (Rubiaceae: coffea arabica) with "Crespera" disease}
Vargas Cartagena L, Sanchez E, Vargas M, Solorzano A, Hernandez F, Iwasawa H, Freer E.

Unidad de Microscopia Electronica, Universidad de Costa Rica, Apdo 2060 San Pedro Montes de Oca, San Jose, Costa Rica . vargmora@racsa.co.cr

"Crespera" is an infectious disease of coffee plants that affects both the coffee production and the economy of the coffee producer countries . This disease affects morphologically the plant: long and narrow leaves with wavy borders and marginal necrosis; strong chlorosis results in drying of the leave, and leads to bad conditions of the plant . The internodes are short, producing the appearance of multiple sprouts in the axial sprout, the flowers can turn greenish, and the plant can present branches with severe symptoms, and branches without apparent symptoms at the same time . As a result, the coffee bean production decreases strikingly . The aim of this work was to determine the occurrence of the possible causative agent in the coffee plants using transmission electron microscopy . Normal and infected plants were compared looking at the leaves, central vein, lateral veins and petiole . It was determined that xylematic vessels show the presence of gram negative bacilliform bacteria (of thick-wavy walls), with dimensions of 0.3-0.5 micron diameter and 1-4 microns, length . The control plants did not show bacteria in the xylem.

Chemistry, 2002 Sep 16, 8(18), 4129 - 37
Chemically modified "polar patch" mutants of subtilisin in peptide synthesis with remarkably broad substrate acceptance: designing combinatorial biocatalysts; Matsumoto K et al.; A significant enhancement of the applicability of the serine protease subtilisin Bacillus lentus (SBL) in peptide synthesis was achieved by using the strategy of combined site-directed mutagenesis and chemical modification to create chemically modified mutant (CMM) enzymes . The introduction of polar and/or homochiral auxiliary substituents, such as X=oxazolidinones, alkylammonium groups, and carbohydrates at position 166 at the base of the primary specificity S(1) pocket created SBL CMMs S166C-S-X with strikingly broad structural substrate specificities . These CMMs are capable of catalyzing the coupling reactions of not only L-amino acid esters but also D-amino acid esters as acyl donors with glycinamide to give the corresponding dipeptides in good yields . These powerful enzymes are also applicable to the coupling of L-amino acid acyl donors with alpha-branched acyl acceptor, L-alaninamide . Typical increases in isolated yields of dipeptides of 60-80 % over SBL-WT (e.g . 0 % yield of Z-D-Glu-GlyNH(2) using SBL-WT-->74 % using S166C-S-(CH(2))(2) NMe(3) (+)) demonstrate the remarkable synthetic utility of this "polar patch" strategy . Such wide-ranging systems displaying broadened and therefore similarly high, balanced yields of products (e.g . 91 % Z-L-Ala-GlyNH(2) and 86 % yield of Z-D-Ala-GlyNH(2) using S166C-S-(3R,4S)-indenooxazolidinone) may now allow the use of biocatalysts in parallel library synthesis.

Mutat Res, 2002 Sep 26, 520(1-2), 141 - 50
Chromosome aberration assays for the study of cyclophosphamide and Bacillus thuringiensis in Oxya chinensis (Orthoptera: Acrididae); Ren Z et al.; Chromosome aberrations induced by an anti-neoplastic drug, cyclophosphamide (CP) and a bioinsecticide, Bacillus thuringiensis (B.t.) were examined using grasshoppers as an animal model, with injection as the route of exposure . Oxya chinensis (Thunberg), having a small number (2n male symbol =23) of large-sized chromosomes in males, was used for this purpose . The fifth instar nymphs were treated with various concentrations of CP (2, 5 and 10 mg/ml) and B.t . (0.55, 1.83 and 5.50 IU/ml) by injection into the abdomen, using physiological saline and distilled water as negative controls, respectively . The chromosomal preparations were made from the spermatogonia of the specimen testis at different intervals after dosing (24 and 48 h) . The effect of the high dose of CP (10 mg/ml) in O . chinensis was also analyzed at the 42-h time point . The chromosome aberrations observed were mainly chromatid and chromosome breaks . CP induced a dose- and time-dependents increase in the number of chromosome aberrations (CAs) per cell and in the percentage of aberrant cells . The strongest effect was seen when grasshoppers were injected with the highest dose and cells were analyzed at the 48-h time point . The results show that CP induced a significant increase in the frequency of CAs in testicular cells of O . chinensis with the three doses employed, compared to the negative control . Our results suggest that there exists in the grasshopper an enzyme system analogous to liver-S9 fraction, and that CP may be used as a positive control in genotoxicity test in this species . In addition, the evaluation of the chromosome aberrations induced by B.t . in the grasshoppers' testicular cells showed that B.t . may induce chromosome aberrations, mainly chromatid and chromosome breaks, in spermatogonia . By statistical analysis, B.t . showed significant dose-effect relationships and it may be mutagenic in this species . Recent research has focused on the development of biological insecticides to protect cereal crops against damage by insect species, such as beetles and grasshoppers . The present studies may contribute to our knowledge of entomological genotoxicity in grasshoppers and provide reference for the research on the mechanism of B.t . toxicity.

J Biochem Mol Biol, 2002 Mar 31, 35(2), 236 - 8
Relationship between acrylamide concentration and enzymatic activity in an improved single fibrin zymogram gel system; Choi NS et al.; Based on the zymography analysis, Bacillus sp . DJ-4 (screened from Doen-Jang, a Korean traditional fermented food) secretes seven extracellular fibrinolytic enzymes (EFEs; 68, 64, 55, 45, 33, 27, and 13 kDa) in culture broth . These seven EFEs were analyzed by newly applied SDSfibrin zymography combined with gradient polyacrylamide (SDS-FZGP) . This improved gel system was used with a 5-20% acrylamide gradient in a fibrin zymogram gel for the separation of proteins with molecular masses from below 10 kDa to over 100 kDa on one gel plate . Using this system, high molecular weight bands (HMWBs) were clearly and sharply resolved . We also examined the relationship between an acrylamide concentration and the enzymatic activity of EFE using densitometric analysis.

J Biochem Mol Biol, 2002 Mar 31, 35(2), 228 - 35
The purification and characterization of a Bacillus stearothermophilus methionine aminopeptidase (MetAP); Chung JM et al.; Methionine aminopeptidase (MetAP) catalyzes the removal of an amino-terminal methionine from a newly synthesized polypeptide . The enzyme was purified to homogeneity from Bacillus stearothermophilus (KCTC 1752) by a procedure that involves heat precipitation and four sequential chromatographs (including DEAESepharose ion exchange, hydroxylapatite, Ultrogel AcA 54 gel filtration, and Reactive red 120 dye affinity chromatography) . The apparent molecular masses of the enzyme were 81,300 Da and 41,000 Da, as determined by gel filtration chromatography and sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE), respectively . This indicates that the enzyme is comprised of two identical subunits . The MetAP specifically hydrolyzed the N-terminal residue of Met-Ala-Ser that was used as a substrate, and exhibited a strong preference for Met-Ala- Ser over Leu-Gly-Gly, Leu-Ser-Phe, and Leu-Leu-Tyr . The enzyme has an optimal pH at 8.0, an optimal temperature at 80 degrees C, and pI at 4.1 . The enzyme was heat-stable, as its activity remained unaltered when incubated at 80 degrees C for 45 min . The Km and Vmax values of the enzyme were 3.0 mM and 1.7 mmol/min/mg, respectively . The B . stearothermophilus MetAP was completely inactivated by EDTA and required Co(2+) ion(s) for activation, suggesting the metal dependence of this enzyme

J Biochem Mol Biol, 2002 May 31, 35(3), 306 - 12
Effects of salts on the conformation and catalytic properties of d-amino acid aminotransferase; Ro HS; The effects of salts on the biochemical properties of D-amino acid aminotransferase from Bacillus sp . YM-1 have been studied to elucidate both the inhibitory effects of salts on the activity and the protective effects of salts on the substrate-induced inactivation . The results from UV-visible spectroscopy studies on the reaction of the enzyme with D-serine revealed that salt significantly reduced the rate of the formation of the quinonoid intermediate and its accumulation . The kinetic and spectroscopy studies of the reaction with alpha-{(2)H}-DL-serine in different concentrations of NaCl provided evidence that the rate-limiting step was changed from the deprotonation of the external aldimine to another step(s), presumably to the hydrolysis of the ketimine . Gel filtration chromatography data in the presence of NaCl showed that the enzyme volume was reduced sharply with the increasing NaCl concentration, up to 100 mM . An additional increase of the NaCl concentration did not affect the elution volume, which suggests that the enzyme has a limited number of salt-binding groups . These results provide detailed mechanistic evidence for the way salts inhibit the catalytic activity of Damino acid aminotransferase

JK Pract, 2000 Jan-Mar, 7(1), 12 - 5
Childhood tuberculosis (Part-I) . Epidemiology, pathogenesis, clinical profile; Mubarik M et al.; PIP: This paper focuses on the pathogenesis and clinical profile of tuberculosis (TB), a bacterial infection caused by Mycobacterium tuberculosis characterized by granuloma formation in infected tissues and by cell-mediated hypersensitivity . TB remains a major cause of morbidity and mortality worldwide and is the most common cause of death from a single infectious disease, particularly in children . Nearly 40 million children are likely to be exposed to the risk of TB and nearly 3-4 million children below age 5 years are estimated to be infected and may progress to disease . TB accounts for 10-15% of all pediatric deaths in a number of Indian hospitals . Factors such as low socioeconomic status, infections, drugs such as steroids, and age predisposes a person to TB . Childhood TB is classified as follows: 1) asymptomatic mantoux positive, 2) symptomatic mantoux positive, 3) primary pulmonary complex, 4) progressive pulmonary disease, 5) disseminated TB, 6) cervical and abdominal TB, 7) tubercular meningitis, 8) progressive bacillus Calmette-Guerin (BCG) disease, and 9) congenital tuberculosis . Among the various symptom complexes of childhood TB are failure to gain weight or weight loss, sudden onset of fever with erythema nodosum and phlyctenular conjunctivitis, failure to gain weight with respiratory wheezing . The direct and indirect impact of HIV infection on the development of TB in children is also discussed .

AIDS Anal Afr, 1997 Oct, 7(5), 5 - 7
Tuberculosis in the HIV / AIDS era: interactions, impacts and solutions; Floyd K et al.; PIP: Tuberculosis (TB) is a health problem of global significance, with 2 billion people infected with the tubercle bacillus and 8 million new cases each year, of which approximately 2.5 million die . TB accounts for an estimated 7% of all avoidable adult deaths worldwide . In 1995, there were an estimated 1.5 million new cases of TB in sub-Saharan Africa; the annual caseload is projected to grow to 2.1 million by 2000 . The growth in population and poverty, as well as the HIV/AIDS epidemic, are behind the rise in TB cases . Countries with high rates of HIV infection have reported far faster increases in cases than those with low rates of infection, rates of HIV infection among TB patients are much higher than the rate of infection in the general population, and TB has become the most important opportunistic disease and one of the most important causes of death among HIV-positive patients in Africa . The link between the HIV/AIDS and TB epidemics, how TB makes HIV both a public and individual health problem, challenges for health services, TB treatment, the delivery of care, prevention, developing an effective response, and a future research agenda are discussed .

AIDS Wkly Plus . 1995 Nov 20-27;:24-5.
Tuberculosis epidemic poses international threat . World Health Organization; Tuberculosis: a snap shot picture; PIP: In India almost 40% of the population is infected with tuberculosis (TB); 0.4% are sputum-positive infectious cases, 2-2.5 million new cases occur annually, and mortality amounts to 50/100,000 population . The National Tuberculosis Program (NTP) and its District TB Program (DTP) aim to detect all TB cases and treat them effectively as part of the general health services, to vaccinate most children with bacillus Calmette-Guerin, to manage planning and implementation, and to carry out proper recording and reporting of cases . Health education is also carried out in order to enlighten the community, patients, children, students, and medical personnel on various aspects of TB using booklets, pamphlets, TV, and newspaper advertisements . Among resources rendering anti-TB services are 390 district TB centers, 17,850 rural health centers, 330 other clinics, and 17 TB demonstration and training centers; there are approximately 47,000 beds available . International assistance has been obtained from the Swedish International Development Agency, who has supplied X-ray units, anti-TB drugs, and vehicles, since 1979 . The World Health Organization (WHO) has assisted by providing consultants, fellowships, and equipment for the National TB Institute in Bangalore and the TB Research Center in Madras . These are also helped by WHO to conduct short courses and training courses for health administrators and college teachers . Some of the problems the NTP faces include: completion of implementation of the DTP in 80 districts and in 25% of peripheral health institutions; nonavailability of trained personnel and vehicles in DTP clinics; overburdened laboratory technicians in 25-30% of primary health care; lack of adequate quantities of drugs, material, and equipment for TB treatment; and lack or shortage of beds . The trends of TB demonstrate that cases tend to concentrate in higher age groups; prevalence in younger people and in newborn children is low and on the decline; and there is a shift from the acute type to the chronic fibrotic type .

IDRC Rep . 1995 Jan;22(4):21.
Malaria control in a nutshell: Palmira Ventosilla; Zighelboim A; PIP: Palmira Ventosilla, a 35-year old Peruvian microbiologist, and her team of researchers at the Alexander von Humboldt Tropical Medicine Institute in Lima, with funding from IDRC, have developed a method of controlling malaria through biological control of mosquito larvae . Bacillus thuringiensis var . israelensis H-14 (Bti) is a naturally occurring bacterium that kills the larvae of Anopheles; it is harmless to humans, but expensive to buy from commercial dealers . The team discovered an inexpensive way to produce the bacterium by growing it in coconuts and releasing it into ponds where the mosquito larvae flourished . However, the community was not easily persuaded to change their lifestyles by foreigners, especially women . Children were easier to persuade than adults . An educational program using posters, comics, and games was developed by Jorge Velez . Lucy Harman and Mark Snyder worked on information sessions designed for adults . While the adults were unwilling, the children were eager; they taught the technique to their families . Several short videos were produced for use at community meetings . The 3 major schools of Salitral, the town where the program is based, are involved and the whole community has been reached; future plans include expansion to more towns, schools, and ponds .

AIDS Wkly . 1994 Oct 3;:14-5.
Prevention of disease the answer; The biotechnology revolution and new vaccines; The author reviews the principles underlying immunization and explains the basis of currently available vaccines . He then describes 2 new biotechnologies, namely peptide synthesis and genetic engineering, which have the capacity to produce both molecular vaccines and live, attenuated vaccines, of an extraordinary range . He mentions the limitations in the strength of some of the experimental vaccines and research being performed to overcome these . Examples are given of some of the vaccines in the research pipeline -- such as those against malaria, hepatitis B and a variety of diarrheal diseases including typhoid, cholera, and bacillary dysentery . He makes reference to new ways of using old vaccines, such as an inhaled aerosolized version of the measles vaccine.

Health Technol Dir, 1983 Jan, 3(1), 1 - 6
Immunization: a key to primary health care; Program for Appropriate Technology in Health PATH; PIP: Focus of this discussion is on some of the problems enountered by national immunization programs and on the technology that is available now or that will be in the near future to help solve these problems . 4 basic aspects of immunization services are examined: the safety, effectiveness, and stability of vaccines; the cold chain, i.e., the transportation, storage, and handling of heat-sensitives vaccines from manufacturer to health worker in the field; vaccination equipment and sterilization for correct administration of immunization; and program management--schedules, records, training, resource allocation . The section devoted to vaccines focuses on immunization against 6 of (diphtheria, whooping cough, tetanus, measles, polio, and tuberculosis) against 6 of the major killers of children in developing countries: BCG, DPT (diphtheria-pertussis-tetanus), measles and poli vaccines, and tetanus toxiod . The bacillus of Calmette and Guerline (BCG) is considered a very safe vaccine . Questions about the effectiveness of BCG in preventing tuberculosis have been raised throughout its 60-year history . Different studies have produced conflicting results, some showing BCG to be highly effective and others showing no positive effect . Diphtheria toxioid, a very safe and relatively stable vaccine, is very effective in protecting against the development of diphtheria . Live attenuated measles virus vaccine is a safe, highly effective vaccine, but it requires careful handling and storage to prevent damage due to excessive heat or light exposure . The vaccine used for pertussis (whooping cough) is a saline suspension of killed Bordetella pertussis bacteria . The vaccine usually is administered as part of the triple DPT vaccine . Concerns about its safety have led to greatly reduced levels of use in some European countries in recent years . Its effectiveness also has been questioned . 2 types of polio vaccine are available: a live, attenuated vaccine given orally (Sabin) and a killed or inactivated vaccine injected intramuscularly (Salk) . Both vaccines are trivalent, i.e., they contain all 3 major strains of polio virus . Both types of vaccine are quite safe . Effectiveness of the vaccines seems to vary in different places . Tetanus toxoid is safe and relatively stable . The 3 doses of DPT given to infants provide long-lasting protection against tetanus . To combine the components of vaccine and equipment into an effective immunizaton service, a program manager must consider several issues: program goals and the development of an appropriate and feasible immunization schedule; systems for keeping records and for training health and cold chain personnel; analysis of the costs of each option; and the development of a method of evaluation .

Diarrhoea Dialogue . 1984 Feb;(16):7.
Typhoid vaccines: outlook for the future; Levine MM et al.; PIP: A new oral typhoid vaccine is undergoing trials among children in Egypt and Chile . Unlike the common injectable killed typhoid vaccine, which causes fever, pain and swelling at the injection site, this live oral vaccine has no side effects . It was developed from a new strain of typhoid, the Ty21a bacillus . The Chilean trials will attempt to determine the efficacy of Ty21a given in enteric coated capsules, to evaluate the efficacy of fewer doses, and to assess the efficacy of this vaccine in an area where typhoid is particularly widespread and lethal . 150,000 school children in Santiago, Chile will follow 5 protocols: 1) 3 doses within 1 week; 2) 3 doses with NaHC03 (sodium bicarbonate, an antacid) within 1 week, both in gelatin capsules; 3) 3 doses as in group 1, separated by 3 weeks; 4) 3 doses as in group 2, separated by 3 weeks; 5) placebo . Each dose contains 1-3 thousand million live, freeze-dried bacteria . Trials in Egypt where children took antacid before each of 3 doses of reconstituted vaccine showed 96% vaccine efficacy after 3 years of surveillance .

Diarrhoea Dialogue . 1984 Feb;(16):3.
Rotaviruses: expectations for a vaccine; Flewett TH; PIP: An attempt is made to answer 2 questions: is it possible to develop a vaccine capable of preventing acute infectious diarrhea; and would this rotavirus vaccine make a significant impression on the number of fatal cases of acute infectious diarrhea . The isolation of rotaviruses in tissue culture to develop less powerful strains for use as a live oral vaccine has been accomplished . At least 4 serotypes exist which can be distinguished from each other by specific antibody . No one knows yet as to whether infection by 1 serotype can provide immunity against infection by a different serotype . The techniques of genetic engineering suggest new possibilities for vaccine development . Rotavirus antigen could be implanted in common bacteria which normally colonize the small bowel . They would reproduce the antigen and thus immunize the patient . Alternatively, the strain of typhoid bacillus, Ty21a, used in the new antityphoid vaccine might carry the rotavirus antigen . The effectiveness of such vaccines is unknown, but the technology to conduct the experiment exists . Recently, new kinds of rotavirus have been identified . 2 serotypes of "standard" piglet rotavirus are well established and share a common group antigen with the rotaviruses commonly found in children, and, of late, 2 new piglet rotaviruses have been discovered . Additionally, recent reports describe new human rotavirus strains in Australia, China, Brazil, France, and Britain, and these are serologically distinct but cannot be detected by current ELISA tests . Prior to marketing a diarrhea vaccine, fully adequate field trails need to guarantee its effectiveness . It will be necessary to conduct these field trials over several years because of the variation in viruses and the periodic nature of viral diarrhea . As reliable vaccination against viral diarrhea seems technically unlikely to become available in the near future, the availability of the inexpensive, simple, and life-saving technique of oral rehydration must be universally publicized and applied .

EPI Newsl . 1979 May;1(1):7.
Vaccination schedules in PAHO member countries.
{Hydrolytic enzymes and spore formation in Bacillus intermedius}
Sharipova MR, Balaban NP, Gabdrakhmanova LA, Shilova MA, Kadyrova IuM, Rudenskaia GN, Leshchinskaia IB.

Kazan State University, ul . Kremlevskaya 18, Kazan, 420008 Russia . margarita.sharipova@ksu.ru

The investigation of the activity of extracellular hydrolytic enzymes and sporulation in the bacterium Bacillus intermedius 3-19 showed that the activity of ribonuclease is maximal in the glucose-containing growth medium, in which sporulation is suppressed . At the sporulation stages II-IV, the synthesis of phosphatase was not regulated by the factors that influence this synthesis in the phase of growth retardation . Caseinolytic activity exhibited two peaks . The first peak was observed when thiol-dependent proteinase began accumulating in the medium . The second peak corresponded to the late stages of sporulation, i.e., the stages of spore maturation and the autolysis of sporangium . The regulatory relationship between proteinase synthesis and sporulation and the possible role of extracellular phosphatases and proteinases in the sporulation are discussed.

Haematologia (Budap), 2002, 32(1), 79 - 80
Shewanella alga bacteremia and associated cellulitis in a patient with multiple myeloma; Krsnik I et al.; Shewanella alga is a gram-negative bacillus found in all types of water as well as in a variety of tainted food . It has rarely been associated with human disease, either in healthy or in immunocompromised patients . We report a 66-year-old man with a multiple myeloma who developed a cellulitis in both forearms in the course of a Shewanella bacteremia . He had a renal insufficiency and was moderately neutropenic after chemotherapy (vincristine, adriamycin, dexamethasone) . Outcome was good after treatment with ceftazidime-amikacin despite all the risk factors . Shewanella isolation may be clinically significant . Haematological patients constitute a group of risk as increasingly aggressive chemotherapy regimens are used . Clinical outcome is not necessarily bad.

J Antibiot (Tokyo), 2002 Jul, 55(7), 650 - 4
An unexpected inhibitory effect of rapamycin against germination of spores of Bacillus brevis strain Nagano; Kim WS et al.; Rapamycin is used in medicine as an immunosuppressive agent (Sirolimus; Rapamune) although discovered as an antifungal agent . It is thought not to have antibacterial activity . Surprisingly, we found that rapamycin inhibits the germination of Bacillus brevis Nagano spores, but is inactive against Bacillus brevis Nagano vegetative cells . Surprisingly rapamycin did not show antimicrobial activity against other Bacillus strains, including other gramicidin S-producing Bacillus brevis strains such as ATCC 9999 and BI-7, whether tested as spores or vegetative cells.

Chem Commun (Camb), 2001 Nov 7, (21), 2194 - 5
Conversion of pyrrole to pyrrole-2-carboxylate by cells of Bacillus megaterium in supercritical CO2; Matsuda T et al.; Pyrrole was converted to pyrrole-2-carboxylate in supercritical CO2 using cells of Bacillus megaterium PYR 2910, and the yield of the carboxylation reaction in supercritical CO2 was 12 times higher than that under atmospheric pressure.

Protein Sci, 2002 Oct, 11(10), 2370 - 81
PrfA protein of Bacillus species: prediction and demonstration of endonuclease activity on DNA; Rigden DJ et al.; The prfA gene product of Gram-positive bacteria is unusual in being implicated in several cellular processes; cell wall synthesis, chromosome segregation, and DNA recombination and repair . However, no homology of PrfA with other proteins has been evident . Here we report a structural relationship between PrfA and the restriction enzyme PvuII, and thereby produce models that predict that PrfA binds DNA . Indeed, wild-type Bacillus stearothermophilus PrfA, but not a catalytic site mutant, nicked one strand of supercoiled plasmid templates leaving 5'-phosphate and 3'-hydroxyl termini . This activity, much lower on linear or relaxed circular double-stranded DNA or on single-stranded DNA, is consistent with a role for this protein in chromosome segregation, DNA recombination, or DNA repair.

Nucleic Acids Res, 2002 Sep 15, 30(18), 4051 - 60
Site-directed mutagenesis reveals roles for conserved amino acid residues in the hexameric DNA helicase DnaB from Bacillus stearothermophilus; Soultanas P et al.; Site-directed mutagenesis studies on conserved amino acid residues within motifs H1, H1a, H2 and H3 of the hexameric replicative helicase DnaB from Bacillus stearothermophilus revealed specific functions associated with these residues . In particular, residues that coordinate a bound Mg2+ in the active site (T217 and D320) are important for the function of the enzyme but are not required for the formation of stable hexamers . A conserved glutamic acid (E241) in motif H1a is likely to be involved in the activation of a water molecule for in line attack on the gamma-phosphate of the bound nucleotide during catalysis . A conserved glutamine (Q362) in motif H3 acts as a gamma-phosphate sensor and mediates the conformational coupling of nucleotide- and DNA-binding sites . The nature of the residue at this position is also important for the primase-mediated activation of DnaB, suggesting that primase uses the same conformational coupling pathway to induce its stimulatory effect on the activity of DnaB . Together, these mutations reveal a conservation of many aspects of biochemical activity in the active sites of monomeric and hexameric helicases.

Antimicrob Agents Chemother, 2002 Oct, 46(10), 3193 - 6
Bactericidal activity of rifampin-amikacin against Mycobacterium ulcerans in mice; Dega H et al.; To identify the most active curative treatment of Buruli ulcer, two regimens incorporating the use of rifampin (RIF) were compared with the use of RIF alone in a mouse footpad model of Mycobacterium ulcerans infection . Treatments began after footpad swelling from infection and continued for 12 weeks with five doses weekly of one of the following regimens: (i) 10 mg of RIF/kg alone; (ii) 10 mg of RIF/kg and 100 mg of amikacin (AMK)/kg; and (iii) 10 mg of RIF/kg, 100 mg of clarithromycin (CLR)/kg, and 50 mg of sparfloxacin (SPX)/kg . The activity of each regimen was assessed in terms of the reduction of the average lesion index and acid-fast bacillus (AFB) and CFU counts . All three regimens displayed various degrees of bactericidal activity against M . ulcerans . The ranking of bactericidal activity was found to be as follows: RIF-AMK > RIF-CLR-SPX > RIF . RIF-AMK was able to cure M . ulcerans-infected mice and prevent relapse 26 weeks after completion of treatment . To determine the impact of different rhythms of administration of RIF-AMK on the suppression of M.ulcerans growth, mice were given the RIF-AMK combination for 4 weeks but doses were administered either 5 days a week or twice or once weekly . After completion of treatment, the mice were kept under supervision for 30 additional weeks . M . ulcerans was considered to have grown in the footpad if swelling was visually observed and harvests contained more than 5 x 10(5) AFB per footpad . The proportion of mice in which growth of M . ulcerans occurred, irrespective of drug dosage, was compared with the control mice to determine the proportion of M . ulcerans killed . Each dosage of RIF-AMK was bactericidal for M . ulcerans (P < 0.001), but the effect was significantly stronger in mice treated 5 days per week . The promising results of RIF-AMK treatment in M . ulcerans-infected mice support the clinical trial that is currently in progress under World Health Organization auspices in Ghana.

J Invertebr Pathol, 2002 May, 80(1), 55 - 63
Toxicity of Bacillus thuringiensis insecticidal proteins for Helicoverpa armigera and Helicoverpa punctigera (Lepidoptera: Noctuidae), major pests of cotton; Liao C et al.; The susceptibilities of the major pests of cotton in Australia, Helicoverpa armigera and Helicoverpa punctigera, to some insecticidal proteins from Bacillus thuringiensis were tested by bioassay . A commercial formulation, DiPel, and individual purified insecticidal proteins were tested . H . armigera was consistently more tolerant to B . thuringiensis insecticidal proteins than was H . punctigera, although both were susceptible to only a limited range of these proteins . Only Cry1Ab, Cry1Ac, Cry2Aa, Cry2Ab, and Vip3A killed H . armigera at dosages that could be considered acceptable . There was no significant difference in the toxicities of Cry1Fa and Cry1Ac for H . punctigera but Cry1Fa had little toxicity for H . armigera . The five instars of H . armigera did not differ significantly in their susceptibility to DiPel on the basis of LC(50) . However, there were significant differences in the susceptibility to Cry1Ac and Cry2Aa of three strains of H . armigera . Bioassays conducted with Cry1Ac and Cry2Aa showed that there was a small but significant negative interaction between these delta-endotoxins.

Res Microbiol, 2002 Jul-Aug, 153(6), 379 - 83
Destabilized green fluorescent protein for monitoring transient changes in mycobacterial gene expression; Triccas JA et al.; The green fluorescent protein (GFP) is a useful reporter for the study of gene expression and protein localisation within living cells . The stability of GFP permits its intracellular accumulation and detection, but renders it less useful for assessing transient changes in gene expression . We have developed a destabilized form of GFP for monitoring gene expression in mycobacteria . By fusing to the C-terminal end of GFP an 11 amino acid peptide encoded by the E . coli ssrA gene, we have developed a form of GFP that exhibits gradual, time-dependent degradation within the fast-growing species Mycobacterium smegmatis . This unstable variant of GFP detected transient changes in the activity of the stress-induced Mycobacterium tuberculosis sigE promoter; by contrast, unmodified GFP only detected a delayed 'switch-on' of this promoter upon exposure to acid stress . Both forms of the protein displayed equivalent stability in the slow-growing species Mycobacterium bovis bacille Calmette-Guerin (BCG), suggesting differing recognition of the ssrA-encoded peptides in slow- and fast-growing mycobacteria . This system will facilitate studies exploring dynamic changes in mycobacterial gene expression.

J Infect Dis, 2002 Oct 1, 186(7), 983 - 90 Epub 2002 Sep 03.
Epidemiology of endemic Bartonella bacilliformis: a prospective cohort study in a Peruvian mountain valley community; Chamberlin J et al.; Bartonella bacilliformis has caused debilitating illness since pre-Incan times, but relatively little is known about its epidemiology . A population-based, prospective cohort investigation was conducted in a Peruvian community with endemic bartonellosis . By use of house-to-house and hospital surveillance methods, cohort participants were monitored for evidence of bartonellosis . Of 690 participants, 0.5% had asymptomatic bacteremia at study initiation . After 2 years of follow-up, the incidence of infection was 12.7/100 person-years . The highest rates were in children <5 years old, and there was a linear decrease in incidence with increasing age . Seventy percent of cases were clustered in 18% of households . Age and bartonellosis in a family member were the best predictors of B . bacilliformis infection . There were multiple clinical presentations and significant subclinical infection . A cost-effective control strategy should include vector control and surveillance efforts focused on children and clusters of households with highest endemicity.

Curr Microbiol, 2002 Nov, 45(5), 346 - 9
Bioremediation of an industrial effluent containing monocrotophos; Bhadbhade BJ et al.; Almost 30% of the precious agricultural output of India is lost owing to pest infestation . In India, pesticide consumption for protecting crops is about 3% of the total world consumption . Monocrotophos (MCP), an organophosphorus insecticide, is widely used to control insects on crops . Being readily water soluble and highly toxic, its removal from wastewater generated during manufacture becomes inevitable . Bioremediation of wastewater containing MCP by Arthrobacter atrocyaneus, Bacillus megaterium, and Pseudomonas mendocina was highest at pH 8.0, but maximum reduction in Chemical Oxygen Demand (COD) was at pH 7.0 . Removal of MCP and reduction in COD by B . megaterium and Ps . mendocina were highest at 35 degrees C, while with A . atrocyaneus, it was maximum at 30 degrees C, under aerated culture condition and inoculum density of 10(8) cells/ml . Use of pure cultures for bioremediation of effluent containing MCP appears to be the first such attempt.

Curr Microbiol, 2002 Nov, 45(5), 309 - 14
Cloning and expression of cyt2Ba7 gene from a soil-isolated Bacillus thuringiensis; Yu J et al.; Bacillus thuringiensis ( Bt) cyt genes coding hemolytic and cytolytic toxins constitute a gene family, which are divided into two groups: cyt1 and cyt2 . A novel cyt2 gene was detected from a soil-isolated Bt strain T301, which was highly homologous to cyt2Ba1 and finally designated cyt2Ba7 . Until now, Cyt2Ba has not been expressed alone in Bt or other hosts . In this study, the cyt2Ba7 gene was cloned into the vector pQE30 and expressed as a fusion protein with 6xHistidine residues in Escherichia coli . Unlike cyt1A, cyt2Ba7 was freely expressed and formed cytoplasmic inclusions without the need for a "helper" protein . The 6xHis-tagged Cyt2Ba7 was purified in one step by Ni-NTA affinity chromatography, examined cytolytic activity on Sf9 cells, and developed as an antigen to obtain the antiserum against Cyt2Ba by subcutaneous injection into rabbits . This gene was also cloned into the Bt-E . coli shuttle vector pHT3101 and expressed in Bt strain 4Q7 . Immunoblotting analysis revealed that the antiserum was remarkably selective and specific to Cyt2Ba.

Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 1996, 28(6), 640 - 646
Purification and Characterization of DNA Helicase BstH1 from Bacillus stearothermophilus; Zhang XH et al.; We have partially purified a DNA helicase BstH1 from Bacillus stearothermophilus through Polymin P precipitation, ammonia sulfate precipitation and column chromatographic steps with Pheny1-Sepharose, DEAE-cellulose, phosphocellulose, FPLC Mono Q and Superose 12 . Bsth1 possesses a DNA-Dependent ATPase activity in the presence of Mg(2+) . The ATPase activity of BstH1 is differentially stimulated by the presence of different types of nucleic acids . BstH1 has an optimal ATPase activity at 55 degrees . The DNA helicase activity of BstH1 requires a 3'-terminal single-stranded DNA binding site to initiate the unwinding reaction in the 3' right curved arrow 5'direction . BstH1 can unwind blunt-ended duplex DNA in a concentration-dependent manner.

BJU Int, 2002 Oct, 90(6), 554 - 7
Recurrence and progression in stage T1G3 bladder tumour with intravesical bacille Calmette-Guérin (Danish 1331 strain); Kulkarni JN et al.; OBJECTIVE: To report recurrence and progression rates in patients with T1G3 superficial bladder carcinoma treated with intravesical bacille Calmette-Guerin (BCG, Danish 1331 strain) after complete transurethral resection . PATIENTS AND METHODS: Data from the records of 111 patients with T1G3 bladder carcinoma treated between January 1991 and December 1999 were analysed for recurrence, progression, salvage therapy and survival . RESULTS: Of the 111 patients with T1G3 bladder tumours, 69 had intravesical BCG therapy, 20 radical cystectomy and 22 only transurethral resection (TUR) . Of the 69 patients receiving BCG therapy 37 (54%) had no recurrence, and 24 (35%) had a recurrence that was not muscle-invasive (Ta/T1) and were treated with TUR only . The remaining eight (12%) progressed to muscle invasion and had salvage cystectomy . During the follow-up six patients died, four from disease and three from other causes, while the remaining 63 are alive and well . Of the other 42 patients, 15 are alive after radical cystectomy and 18 after TUR . CONCLUSION: This series further confirms the benefits of intravesical BCG (Danish 1331) in an adjuvant setting; furthermore, this treatment facilitates bladder preservation by reducing recurrences and delaying the progression in many patients.

Eur J Biochem, 2002 Sep, 269(18), 4577 - 85
The molecular surface of proteolytic enzymes has an important role in stability of the enzymatic activity in extraordinary environments; Yamagata Y et al.; It is scientifically and industrially important to clarify the stabilizing mechanism of proteases in extraordinary environments . We used subtilisins ALP I and Sendai as models to study the mechanism . Subtilisin ALP I is extremely sensitive to highly alkaline conditions, even though the enzyme is produced by alkalophilic Bacillus, whereas subtilisin Sendai from alkalophilic Bacillus is stable under conditions of high alkalinity . We constructed mutant subtilisin ALP I enzymes by mutating the amino acid residues specific for subtilisin ALP I to the residues at the corresponding positions of amino acid sequence alignment of alkaline subtilisin Sendai . We observed that the two mutations in the C-terminal region were most effective for improving stability against surfactants and heat as well as high alkalinity . We predicted that the mutated residues are located on the surface of the enzyme structures and, on thebasis of three-dimensional modelling, that they are involved in stabilizing the conformation of the C-terminal region . As proteolytic enzymes frequently become inactive due to autocatalysis, stability of these enzymes in an extraordinary environment would depend on the conformational stability of the molecular surface concealing scissile peptide bonds . It appeared that the stabilization of the molecular surface structure was effective to improve the stability of the proteolytic enzymes.

J Invest Dermatol, 2002 Sep, 119(3), 570 - 5
Inverse association between melanoma and previous vaccinations against tuberculosis and smallpox: results of the FEBIM study; Pfahlberg A et al.; Various forms of immunotherapy utilizing bacille Calmette-Guerin vaccine or vaccinia vaccine have been evaluated in clinical trials on melanoma patients . The effect of the "natural" application of these vaccinations, administered to provide protection against tuberculosis and smallpox, has, however, never been studied in epidemiologic investigations on risk factors for melanoma . In a case-control study comprising 11 institutions in seven countries we recruited 603 incident melanoma cases and 627 population controls frequency matched to the cases with respect to sex, age, and ethnic origin within each center to assess this relationship to obtain insights into the prevention of melanoma . Exposure information, incorporating also detailed ascertainment of potential confounding variables, was obtained in standardized personal interviews at the study subject's home . We found an inverse association between melanoma risk and previous bacille Calmette-Guerin vaccine/vaccinia vaccination depicted by an adjusted odds ratio of 0.44 (95% confidence interval: 0.26-0.72) for those vaccinated against tuberculosis and smallpox compared with subjects without a positive history of either vaccination . A variety of subgroup analyses showing a consistent pattern of results make it unlikely that the observed inverse association is a spurious finding . We conclude that bacille Calmette-Guerin vaccination and vaccinia vaccination may lower melanoma risk . Current immunologic theory of melanoma development provides a sound basis for understanding the biologic plausibility of the findings that have to be confirmed in future studies.

Water Res, 2001 Nov, 35(16), 3807 - 16
Wastewater treatment sludge as a raw material for the production of Bacillus thuringiensis based biopesticides; Montiel MD et al.; Seven wastewater sludges of different origins and types were used as an alternate culture medium for producing Bacillus thuringiensis variety kurstaki HD-1 . The sludge samples were used under three different preparations: without pre-treatment, with acid treatment (hydrolysed sludge) and the supernatant obtained after centrifugation of the hydrolysed sludge . The sludge composition varied widely with origin and the type of sludge . Growth and sporulation were evaluated by the total viable cell count and spore count of the preparations . Growth, sporulation and endotoxin production were affected by the sludge origin . Hydrolysed sludge gave the highest viable cell and spore counts while the liquid phase (supernatant) gave the lowest . Non-hydrolysed primary sludge from Valcartier was unable to sustain bacterial growth because of its low pH . Bioassays were conducted against larvae of spruce budworm to evaluate entomotoxic potential of the preparations obtained . In general, sludge hydrolysis increased the entomotoxicity yields . Similar entomotoxicity was observed in Black Lake secondary sludge (4100 IU/microL) as that obtained in the reference soya medium (3800 IU/microL) . The use of the sludge supernatant (liquid phase) was not recommended due to the low entomotoxic potential obtained.

Biol Pharm Bull, 2002 Sep, 25(9), 1169 - 74
Suppression of NO production in activated macrophages in vitro and ex vivo by neoandrographolide isolated from Andrographis paniculata; Batkhuu J et al.; In this study, we investigated the in vitro and ex vivo suppressive effects of Andrographis paniculata on nitric oxide (NO) production in mouse peritoneal macrophages elicited by bacillus Calmette-Guein (BCG) and stimulated by lipopolysaccharide (LPS) . Incubation of BCG-induced macrophages with the methanol extract of A . paniculata reduced LPS stimulated NO production . The diterpene lactones andrographolide and neoandrographolide were isolated as active components from the extract . These compounds suppressed NO production in a concentration-dependent manner in the concentration range from 0.1 to 100 microM and their IC50 values were 7.9 and 35.5 microM . Neoandrographolide also suppressed NO production by 35 and 40% when the macrophages were collected after oral administration of neoandrographolide at doses of 5 and 25 mg/kg/d and LPS stimulated NO production was examined . However, andrographolide did not reduce NO production on oral administration at the same doses . These results indicate that neoandrographolide, which inhibited NO production both in vitro and ex vivo may play an important role in the use of A . paniculata as an anti-inflammatory crude drug.

Clin Infect Dis, 2002 Oct 1, 35(7), 796 - 801 Epub 2002 Sep 11.
Risk of tuberculin skin test conversion among health care workers: occupational versus community exposure and infection; Larsen NM et al.; A prospective observational cohort study to assess rates of and risk factors for tuberculin skin test (TST) conversion among health care workers (HCWs) was conducted at an urban hospital located in a high tuberculosis-incidence area in 1994-1998 . All hospital employees undergoing required testing every 6 months were included . A total of 69 (1.2%) of 5773 susceptible employees had a documented TST conversion (overall rate, 0.38 per 100 person-years worked) . No significant difference existed in conversion rates among employees with frequent, limited, or no patient contact . HCWs with a TST conversion lived in zip codes with higher tuberculosis case rates (P< or =.05) . In multivariate analysis, TST conversion was associated with history of bacille Calmette-Guerin vaccination (relative risk {RR}, 11.63), annual salary <$20,000 (RR, 3.67), and increasing age . In the setting of an effective tuberculosis infection-control program, TST conversion rates were low, and risk of conversion among HCWs was associated most strongly with nonoccupational factors.

Infect Immun, 2002 Oct, 70(10), 5381 - 9
Contribution of membrane-damaging toxins to Bacillus endophthalmitis pathogenesis; Callegan MC et al.; Membrane-damaging toxins are thought to be responsible for the explosive clinical course of Bacillus endophthalmitis . This study analyzed the contribution of phosphatidylinositol-specific phospholipase C (PI-PLC) and phosphatidylcholine-specific phospholipase C (PC-PLC) to the pathogenesis of experimental Bacillus endophthalmitis . Isogenic mutants were constructed by insertion of lacZ into Bacillus thuringiensis genes encoding PI-PLC (plcA) and PC-PLC (plcB) . Rabbit eyes were injected intravitreally with 2 log(10) CFU of strain BT407 (wild type), the PI-PLC mutant (BTplcA::lacZ), or the PC-PLC mutant (BTplcB::lacZ) . The rates of decrease in retinal responses of eyes infected with the isogenic mutants were similar to that of wild type, with all infections resulting in elimination of retinal function by 18 h . Strain BT407 caused a significant increase in the latency of retinal responses at 6 h, but strains BTplcA::lacZ and BTplcB::lacZ did not . All strains elicited significant inflammatory cell influx into the anterior chamber by 12 h . Histologically, eyes infected with each strain were indistinguishable throughout the infection course . In this model, neither PI-PLC nor PC-PLC had an effect on the course or severity of experimental Bacillus endophthalmitis . Alterations in retinal responses early in infection may mark the beginnings of specific photoreceptor or glial cell dysfunction.

Plant Physiol, 1996 Nov, 112(3), 919 - 929
Induction of Defense-Related Ultrastructural Modifications in Pea Root Tissues Inoculated with Endophytic Bacteria; Benhamou N et al.; The stimulation exerted by the endophytic bacterium Bacillus pumilus strain SE34 in plant defense reactions was investigated at the ultrastructural level using an in vitro system in which root-inducing T-DNA pea (Pisum sativum L.) roots were infected with the pea root-rotting fungus Fusarium oxysporum f . sp . pisi . In nonbacterized roots, the pathogen multiplied abundantly through much of the tissue including the vascular stele, whereas in prebacterized roots, pathogen growth was restricted to the epidermis and the outer cortex In these prebacterized roots, typical host reactions included strengthening the epidermal and cortical cell walls and deposition of newly formed barriers beyond the infection sites . Wall appositions were found to contain large amounts of callose in addition to being infiltrated with phenolic compounds . The labeling pattern obtained with the gold-complexed laccase showed that phenolics were widely distributed in Fusarium-challenged, bacterized roots . Such compounds accumulated in the host cell walls and the intercellular spaces as well as at the surface or even inside of the invading hyphae of the pathogen . The wall-bound chitin component in Fusarium hyphae colonizing bacterized roots was preserved even when hyphae had undergone substantial degradation . These observations confirm that endophytic bacteria may function as potential inducers of plant disease resistance.

Plant Physiol, 1996 Feb, 110(2), 355 - 363
Fructan Accumulation and Sucrose Metabolism in Transgenic Maize Endosperm Expressing a Bacillus amyloliquefaciens SacB Gene; Caimi PG et al.; Over 40,000 species of plants accumulate fructan, {beta}-2-1- and {beta}-2-6-linked polymers of fructose as a storage reserve . Due to their high fructose content, several commercial applications for fructans have been proposed . However, plants that accumulate these polymers are not agronomically suited for large-scale cultivation or processing . This study describes the transformation of a Bacillus amyloliquefaciens SacB gene into maize (Zea mays L.) callus by particle bombardment . Tissue-specific expression and targeting of the SacB protein to endosperm vacuoles resulted in stable accumulation of high-molecular-weight fructan in mature seeds . Accumulation of fructan in the vacuole had no detectable effect on kernel development or germination . Fructan levels were found to be approximately 9-fold higher in sh2 mutants compared to wild-type maize kernels . In contrast to vacuole-targeted expression, starch synthesis and endosperm development in mature seeds containing a cytosolically expressed SacB gene were severely affected . The data demonstrate that hexose resulting from cytosolic SacB activity was not utilized for starch synthesis . Transgenic seeds containing a chimeric SacB gene provide further evidence that the dominant pathway for starch synthesis in maize endosperm is through uridine diphosphoglucose catalyzed by the enzyme sucrose synthase.

Biosci Biotechnol Biochem, 2002 Jul, 66(7), 1538 - 45
Significance of the 20-kDa subunit of heterodimeric 2-deoxy-scyllo-inosose synthase for the biosynthesis of butirosin antibiotics in Bacillus circulans; Tamegai H et al.; A gene (btrC2) encoding the 20-kDa subunit of 2-deoxy-scyllo-inosose (DOI) synthase, a key enzyme in the biosynthesis of 2-deoxystreptamine, was identified from the butirosin-producer Bacillus circulans by reverse genetics . The deduced amino acid sequence of BtrC2 closely resembled that of YaaE of B . subtilis, but the function of the latter has not been known to date . Instead, BtrC2 appeared to show sequence similarity to a certain extent with HisH of B . subtilis, an amidotransferase subunit of imidazole glycerol phosphate synthase . Disruption of btrC2 reduced the growth rate compared with the wild type, and simultaneously antibiotic producing activity was lost . Addition of NH4Cl to the medium complemented only the growth rate of the disruptant, and both the growth rate and antibiotic production were restored by addition of yeast extract . In addition, a heterologous co-expression system of btrC2 with btrC was constructed in Escherichia coli . The simultaneously over-expressed BtrC2 and BtrC constituted a heterodimer, the biochemical features of which resembled those of DOI synthase from B . circulans more than those of the recombinant homodimeric BtrC . Despite the similarity of BtrC2 to HisH the heterodimer showed neither aminotransfer nor amidotransfer activity for 2-deoxy-scyllo-inosose as a substrate . All the observations suggest that BtrC2 is involved not only in the secondary metabolism, but also in the primary metabolism in B . circulans . The function of BtrC2 in the butirosin biosynthesis appears to be indirect, and may be involved in stabilization of DOI synthase and in regulation of its enzyme activity.






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