Nature, 2001 Mar 1, 410(6824), 77 - 81 Isotopic evidence for microbial sulphate reduction in the early Archaean era; Shen Y et al.; Sulphate-reducing microbes affect the modern sulphur cycle, and may be quite ancient, though when they evolved is uncertain . These organisms produce sulphide while oxidizing organic matter or hydrogen with sulphate . At sulphate concentrations greater than 1 mM, the sulphides are isotopically fractionated (depleted in 34S) by 10-40/1000 compared to the sulphate, with fractionations decreasing to near 0/1000 at lower concentrations . The isotope record of sedimentary sulphides shows large fractionations relative to seawater sulphate by 2.7 Gyr ago, indicating microbial sulphate reduction . In older rocks, however, much smaller fractionations are of equivocal origin, possibly biogenic but also possibly volcanogenic . Here we report microscopic sulphides in approximately 3.47-Gyr-old barites from North Pole, Australia, with maximum fractionations of 21.1/1000, about a mean of 11.6/1000, clearly indicating microbial sulphate reduction . Our results extend the geological record of microbial sulphate reduction back more than 750 million years, and represent direct evidence of an early specific metabolic pathway--allowing time calibration of a deep node on the tree of life. Mutat Res, 2001 Mar 1, 474(1-2), 159 - 68 Polar alteration of short tandem repeats (STRs) in mammalian cells; Suzuki A et al.; Instability of short tandem repeats (STRs) in DNA during replication is observed in all organisms examined, and is causatively involved in various human diseases . We explore the mechanisms involved in instability by examining length changes occurring during the replication of {(CA)(20)TA}(n) and {(CAG)(20)TAG}(n), in human cells . We show that the majority of alterations consist of an insertion or deletion of one repeat unit, and base substitutions or length changes involving many repeat units are rare . We also show that length changes of two-tract STRs are biased toward the 3'-end of the repeat tract, in reference to lagging strand synthesis . There are some differences between our observations and previous observations in microbes, e.g . the orientation effect was not observed in this study . The results of this study are discussed in terms of the molecular mechanisms leading to alterations in repeat tracts. Microbiol Mol Biol Rev, 2001 Mar, 65(1), 106 - 18 Biosynthesis of polyketides in heterologous hosts; Pfeifer BA et al.; Polyketide natural products show great promise as medicinal agents . Typically the products of microbial secondary biosynthesis, polyketides are synthesized by an evolutionarily related but architecturally diverse family of multifunctional enzymes called polyketide synthases . A principal limitation for fundamental biochemical studies of these modular megasynthases, as well as for their applications in biotechnology, is the challenge associated with manipulating the natural microorganism that produces a polyketide of interest . To ameliorate this limitation, over the past decade several genetically amenable microbes have been developed as heterologous hosts for polyketide biosynthesis . Here we review the state of the art as well as the difficulties associated with heterologous polyketide production . In particular, we focus on two model hosts, Streptomyces coelicolor and Escherichia coli . Future directions for this relatively new but growing technological opportunity are also discussed. J Immunol, 2001 Mar 15, 166(6), 4237 - 43 Environmental modulation of autoimmune arthritis involves the spontaneous microbial induction of T cell responses to regulatory determinants within heat shock protein 65; Moudgil KD et al.; Both genetic and environmental factors are believed to be involved in the induction of autoimmune diseases . Adjuvant arthritis (AA) is inducible in susceptible rat strains by injection of Mycobacterium tuberculosis, and arthritic rats raise T cell responses to the 65-kDa mycobacterial heat-shock protein (Bhsp65) . We observed that Fischer 344 (F344) rats raised in a barrier facility (BF-F344) are susceptible to AA, whereas F344 rats maintained in a conventional facility (CV-F344) show significantly reduced incidence and severity of AA, despite responding well to the arthritogenic determinant within Bhsp65 . The acquisition of protection from AA can be circumvented if rats are maintained on neomycin/acidified water . Strikingly, naive unimmunized CV-F344 rats but not BF-F344 rats raised T cell responses to Bhsp65 C-terminal determinants (BCTD) (we have previously shown that BCTD are involved in regulation of acute AA in the Lewis rat); however, T cells of naive CV-F344 and BF-F344 gave a comparable level of proliferative response to a mitogen, but no response at all to an irrelevant Ag . Furthermore, adoptive transfer into naive BF-F344 rats of splenic cells of naive CV-F344 rats (restimulated with BCTD in vitro) before induction of AA resulted in a considerably reduced severity of AA . These results suggest that spontaneous (inadvertent) priming of BCTD-reactive T cells, owing to determinant mimicry between Bhsp65 and its homologues in microbial agents in the conventional environment, is involved in modulating the severity of AA in CV-F344 rats . These results have important implications in broadening understanding of the host-microbe interaction in human autoimmune diseases. Br J Cancer, 2001 Mar 2, 84(5), 674 - 9 Direct evidence for a bystander effect of ionizing radiation in primary human fibroblasts; Belyakov OV et al.; Bystander responses underlie some of the current efforts to develop gene therapy approaches for cancer treatment . Similarly, they may have a role in strategies to treat tumours with targeted radioisotopes . In this study we show direct evidence for the production of a radiation-induced bystander response in primary human fibroblasts . We utilize a novel approach of using a charged-particle microbeam, which allows individual cells within a population to be selected and targeted with counted charged particles . Individual primary human fibroblasts within a population of 600-800 cells were targeted with between 1 and 15 helium ions (effectively, alpha-particles) . The charged particles were delivered through the centre of the nucleus with an accuracy of +/- 2 micrometer and a detection and counting efficiency of greater than 99% . When scored 3 days later, even though only a single cell had been targeted, typically an additional 80-100 damaged cells were observed in the surviving population of about 5000 cells . The yield of damaged cells was independent of the number of charged particles delivered to the targeted cell . Similar results of a 2-3-fold increase in the background level of damage present in the population were observed whether 1 or 4 cells were targeted within the dish . Also, when 200 cells within one quadrant of the dish were exposed to radiation, there was a 2-3-fold increase in the damage level in an unexposed quadrant of the dish . This effect was independent of the presence of serum in the culture medium and was only observed when a cell was targeted, but not when only the medium was exposed, confirming that a cell-mediated response is involved . Anal Biochem, 2001 Mar, 290(2), 214 - 20 Miniaturization of a hepatitis C virus RNA polymerase assay using a -102 degrees C cooled CCD camera-based imaging system; Zheng W et al.; Innovations in detection technologies have allowed us to develop a novel assay in 1536-well plate format and assess the advantages of screen miniaturization compared with conventional high-throughput compound screening in 96- or 384-well plates . An HCV RNA polymerase assay has been miniaturized in 1536-well plates by using a new detection technology known as LEADseeker homogeneous imaging system . It uses a -102 degrees C cooled charge-coupled device (CCD) camera and newly designed scintillation proximity microparticles . The miniaturized assay used europium-doped streptavidin-coated yttrium oxide (YO(x)) or polystyrene (PS) microspheres to capture biotin-labeled {(3)H}RNA product transcripts . Beads in proximity to the radioisotope convert the emitted beta(-) particles into photons having wavelengths in the red region of the visible spectrum, optimal for detection by the CCD camera . Because the camera collects light from all wells of the plate simultaneously, 1536-well plates are imaged as rapidly as 384-well plates, on the order of 10 min per plate . The assay has a signal to background of approximately 20-fold, satisfactory for high-throughput robotics screening . The enzyme kinetics and potency of a known inhibitor were similar to those obtained from the conventional assay using scintillation proximity assay (SPA) beads and a scintillation plate counter . Furthermore, the newly developed microbeads (emitting at 610 to 620 nm) are less prone to quenching effects caused by yellow-colored compounds, than conventional SPA beads or scintillation fluid (emitting at 400 to 480 nm region) . Thus, the LEADseeker imaging system is a useful new tool for miniaturization of assays for high-throughput screening . Zh Mikrobiol Epidemiol Immunobiol, 2001 Jan-Feb, (1), 104 - 8 {Effect of microalgae on viability of microorganisms in the natural and artificial environment}; Gorobets OB et al.; The data of literature on complicated relationships between microorganisms and microalgae in algobacterial associations with microalgae often playing the leading role are analyzed . Under these conditions inhibiting and stimulating substances synthesized by microalgae, as well as their nutritional value, may have an essential impact on the state of the bacterial population . Apparently that microalgae may become the main substrate in the development of new culture media (ecologically pure culture media, media for the reversion of the noncultured forms of bacteria into the vegetative state, media for the prolonged preservation of microbes in the noncultured form). Appl Microbiol Biotechnol, 2001 Jan, 55(1), 1 - 9 Microbial formation, biotechnological production and applications of 1,2-propanediol; Bennett GN et al.; This short review covers metabolic pathways, genetics and metabolic engineering of 1,2-propanediol formation in microbes . 1,2-Propanediol production by bacteria and yeasts has been known for many years and two general pathways are recognized . One involves the metabolism of deoxyhexoses, where lactaldehyde is formed during the glycolytic reactions and is then reduced to 1,2-propanediol . The second pathway derives from the formation of methylglyoxal from dihydroxyacetonephosphate and its subsequent reduction to 1,2-propanediol . The enzymes involved in the reduction of methylglyoxal can generate isomers of lactaldehyde or acetol, which can be further reduced by specific reductases, giving chiral 1,2-propanediol as the product . The stereospecificity of the enzymes catalyzing the two reduction steps is important in deriving a complete pathway . Through genetic engineering, appropriate combinations of enzymes have been brought together in Escherichia coli and yeast to generate 1,2-propanediol from glucose . The optimization of these strains may yield microbial processes for the production of this widely used chemical. Prikl Biokhim Mikrobiol, 2001 Jan-Feb, 37(1), 123 - 8 {Rapid assay for the assessment of a potential of chemical biocides to microbial destructors of industrial materials}; Novikov IA et al.; A colorimetric rapid assay for estimating the biocide potential of various chemicals towards metal biocorrosive and petroleum product degrading microbes was developed based on the reducing potential of live microbial cell . A water-soluble organic redox indicator, blue in the oxidized form and pink in the reduced form, was used as an indicator of the reducing potential of microbial cells . Once added to a suspension of vital microbial cells, it was reduced and changed in color . A good correlation between the results of this assay and viability control was obtained by employing surfactants and heavy metal ions. Nature, 2001 Feb 22, 409(6823), 1092 - 101 Life in extreme environments; Rothschild LJ et al.; Each recent report of liquid water existing elsewhere in the Solar System has reverberated through the international press and excited the imagination of humankind . Why? Because in the past few decades we have come to realize that where there is liquid water on Earth, virtually no matter what the physical conditions, there is life . What we previously thought of as insurmountable physical and chemical barriers to life, we now see as yet another niche harbouring 'extremophiles' . This realization, coupled with new data on the survival of microbes in the space environment and modelling of the potential for transfer of life between celestial bodies, suggests that life could be more common than previously thought . Here we examine critically what it means to be an extremophile, and the implications of this for evolution, biotechnology and especially the search for life in the Universe. Folia Biol (Praha), 2001, 47(1), 36 - 9 Analysis of paternal alleles in nucleated red blood cells enriched from maternal blood; Hromadnikova I et al.; The purpose of our study was to identify paternal alleles in NRBC enriched from maternal peripheral blood for detection of the presence of foetal cells in the maternal circulation and to establish a reliable non-invasive method which should allow following genetic testing . For enrichment of foetal cells from peripheral maternal blood we combined Ficoll-Paque density gradient centrifugation and MACS . Maternal leukocytes were firstly depleted using anti-CD14 and anti-CD45 microbeads . NRBC were sorted from the CD14-/CD45- fraction by positive selection using CD71 microbeads . Paternal alleles in the CD14-/CD45-/CD71+ fraction were indicated by the PCR method using HLA (DRB1, DQB1, DQA1) and Polymarker System (LDLR, GYPA, HBGG, D7S8, GC) as genetic markers . Different paternal alleles of studied 8 loci were detected in 13 out of 19 samples of cells enriched from maternal peripheral blood between the 13th and 36th week of gestation . Our results demonstrate that foetal cells enriched from maternal peripheral blood may be used as a source of foetal DNA for prenatal diagnosis, paternity testing and other application. Plant Cell Physiol, 2001 Feb, 42(2), 129 - 37 An arabinogalactan protein(s) is a key component of a fraction that mediates local intercellular communication involved in tracheary element differentiation of zinnia mesophyll cells; Motose H et al.; Local intercellular communication is involved in tracheary element (TE) differentiation of zinnia (Zinnia elegans L.) mesophyll cells and mediated by a proteinous macromolecule, which was designated xylogen . To characterize and isolate xylogen, a bioassay system to monitor the activity of xylogen was developed, in which mesophyll cells were embedded in microbeads of agarose gel at a low (2.0-4.3x10(4) cells ml(-1)) or high density (8.0-9.0x10(4) cells ml(-1)) and microbeads of different cell densities were cultured together in a liquid medium to give a total density of 2.1-2.5x10(4) cells ml(-1) . Without any additives, the frequency of TE differentiation was much smaller in the low-density microbeads than in the high-density microbeads . This low level of TE differentiation in the low-density microbeads was attributable to the shortage of xylogen . When cultures were supplemented with conditioned medium (CM) prepared from zinnia cell suspensions undergoing TE differentiation, the frequency of TE differentiation in the low-density microbeads increased remarkably, indicating the activity of xylogen in the CM . The xylogen activity in CM was sensitive to proteinase treatments . Xylogen was bound to galactose-specific lectins such as Ricinus communis agglutinin and peanut agglutinin, and precipitated by beta-glucosyl Yariv reagent . These results indicate that xylogen is a kind of arabinogalactan protein. Proc Natl Acad Sci U S A, 2001 Feb 27, 98(5), 2164 - 9 Truncated hexa-octahedral magnetite crystals in ALH84001: presumptive biosignatures; Thomas-Keprta KL et al.; McKay et al . {(1996) Science 273, 924-930} suggested that carbonate globules in the meteorite ALH84001 contained the fossil remains of Martian microbes . We have characterized a subpopulation of magnetite (Fe(3)O(4)) crystals present in abundance within the Fe-rich rims of these carbonate globules . We find these Martian magnetites to be both chemically and physically identical to terrestrial, biogenically precipitated, intracellular magnetites produced by magnetotactic bacteria strain MV-1 . Specifically, both magnetite populations are single-domain and chemically pure, and exhibit a unique crystal habit we describe as truncated hexa-octahedral . There are no known reports of inorganic processes to explain the observation of truncated hexa-octahedral magnetites in a terrestrial sample . In bacteria strain MV-1 their presence is therefore likely a product of Natural Selection . Unless there is an unknown and unexplained inorganic process on Mars that is conspicuously absent on the Earth and forms truncated hexa-octahedral magnetites, we suggest that these magnetite crystals in the Martian meteorite ALH84001 were likely produced by a biogenic process . As such, these crystals are interpreted as Martian magnetofossils and constitute evidence of the oldest life yet found. Curr Opin Gastroenterol, 2001 Mar, 17(2), 171 - 176 Nutrition and the mucosal immune system; Cunningham-Rundles S; Gut-associated lymphoid tissue is the dominant site for the initiation of mucosal immune response . Mucosal immunity depends on regulatory signals; nutritional elements, including fats, amino acids, and micronutrients, are critical cofactors for these signals . Nutrients specifically affect lymphocyte influx and migration, mononuclear cell activation, and the differentiated expression of immune response . The molecular basis of nutrient action has been shown to involve effects on receptor regulation, adhesion molecule expression, and the pattern of cytokine production . The gastrointestinal mucosal immune system is the major site for host interaction with microbes and provides a barrier against systemic access for food antigens and microbes . Nutrient metabolism has unique and direct impact on the host defense system of gut-associated lymphoid tissue and therefore has potential for widely disseminated impact on systemic immune response. Br J Ophthalmol, 2001 Mar, 85(3), 336 - 40 A comparison of cyst age and assay method of the efficacy of contact lens disinfectants against Acanthamoeba; Kilvington S et al.; AIMS: (i) To determine effect of Acanthamoeba cyst age, method of production, and (ii) to assay technique on the efficacy of multipurpose solutions (MPS) and hydrogen peroxide based contact lens disinfectants . (iii) To establish if MPS can remove mature cysts from contact lenses according to the ISO/DIS 14729 regimen test for microbe removal . METHODS: Immature and mature cysts of A polyphaga were tested against the MPS Opti-Free express and the hydrogen peroxide based solutions Oxysept 1Step and Oxysept 1 using two assay methods . Simulated patient regimen testing was performed with the Opti-Free express and Complete using mature cysts inoculated on to group I or group IV lenses . RESULTS: Immature cysts were sensitive to disinfection by all solutions . No killing was observed with mature cysts with Opti-Free express, while immature cysts yielded a 1-2 log reduction in viability . Oxysept 1Step gave a 1.1 (SD 0.3) log reduction in mature cysts after 6 hours . Oxysept 1 gave a 2.4 (0.3) log reduction in mature cysts after 4 hours and a 3.8 (0.5) log reduction after 6 hours . Patient regimen testing using Opti-Free express and Complete resulted in no recovery of viable mature cysts from the contact lenses or from the soaking solutions . CONCLUSION: Cyst age but not method of production used in this study influences the efficacy of contact lens disinfectants against Acanthamoeba . MPS are effective in removing cysts from contact lens surfaces and may have a role in the prevention of acanthamoeba keratitis. Ann Biomed Eng, 2001 Jan, 29(1), 1 - 8 Controlled cell deformation produces defined areas of contact between cells and ligand-coated surfaces; Patrick SM et al.; A method which allows precise control of the time of initiation and the area of contact of T cells with immobilized ligands has been developed . Cells are trapped in an asymmetric film that can be quantitatively thinned by reducing the film's capillary pressure . Ligands adsorbed to the base of the apparatus are forced into close contact with the cells as the air-liquid interface is drawn down . Using interference microscopy and microbeads to indicate the film height, the amount of thinning can be controlled to within 1 microm . In this study, this system was used to produce contact areas of 182 and 356 microm2 between T cells and anti-CD3 coated surfaces . These contact areas were measured using fluorescent dye exclusion microscopy . This apparatus can be used for quantitative studies of T cell activation, as is reported in Patrick et al., J . Immunol . Method . 24:97-108, 2000. J Chromatogr A, 2001 Feb 9, 909(1), 53 - 60 Detection of the cyclic nitramine explosives hexahydro-1,3,5-trinitro- 1,3,5-triazine (RDX) and octahydro- 1,3,5,7-tetranitro- 1,3,5,7-tetrazine (HMX) and their degradation products in soil environments; Groom CA et al.; The cyclic nitramine explosives hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazine (HMX) were examined in field and microcosm soil samples to determine their patterns of degradation and environmental fates . A number of analytical techniques, including solid-phase microextraction with on-fiber derivatization, gas chromatography-mass spectrometry, gas chromatography with electron-capture detection, liquid chromatography-mass spectrometry, and micellar electrokinetic chromatography were required for the analyses . Two different classes of intermediates were detected, both of which lead ultimately to the formation of nitrous oxide (N2O) and carbon dioxide (CO2) . The first class was identified as the nitroso derivatives formed by the sequential reduction of -NO2 functional groups . The second class of intermediates, which was favored at higher humidities and in the presence of anaerobic sludge amendments, consisted of ring cleavage products including bis-(hydroxymethyl)-nitramine and methylenedinitramine . Rye-grass (Lolium perenne) present in field samples was found to extract and accumulate HMX from soil without further degradation . In all cases (excepting the plant samples), the indigenous microbes or amended domestic anaerobic sludge consortia degraded the cyclic nitramine explosives eventually to produce N2O and CO2. Proc R Soc Lond B Biol Sci, 2001 Feb 7, 268(1464), 303 - 9 Microbial diversity, producer-decomposer interactions and ecosystem processes: a theoretical model; Loreau M; Interactions between the diversity of primary producers and that of decomposers--the two key functional groups that form the basis of all ecosystems--might have major consequences on the functioning of depauperate ecosystems . I present a simple ecosystem model in which primary producers (plants) and decomposers (microbes) are linked through material cycling . The model considers a diversity of plant organic compounds and a diversity of microbial species . Nutrient recycling efficiency from organic compounds to decomposers is then the key parameter that controls ecosystem processes (primary productivity, secondary productivity, producer biomass and decomposer biomass) . The model predicts that microbial diversity has a positive effect on nutrient recycling efficiency and ecosystem processes through either greater intensity of microbial exploitation of organic compounds or functional niche complementarity, much like in plants . Microbial niche breadth and overlap should not affect ecosystem processes unless they increase the number of organic compounds that are decomposed . In contrast, the model predicts that plant organic compound diversity can only have a negative effect or, at best, no effect on ecosystem processes, at least in a constant environment . This creates a tension between the effects of plant diversity and microbial diversity on ecosystem functioning, which may explain some recent experimental results. Nature, 2001 Feb 1, 409(6820), 630 - 3 Polarity controls forces governing asymmetric spindle positioning in the Caenorhabditis elegans embryo; Grill SW et al.; Cell divisions that create daughter cells of different sizes are crucial for the generation of cell diversity during animal development . In such asymmetric divisions, the mitotic spindle must be asymmetrically positioned at the end of anaphase . The mechanisms by which cell polarity translates to asymmetric spindle positioning remain unclear . Here we examine the nature of the forces governing asymmetric spindle positioning in the single-cell-stage Caenorhabditis elegans embryo . To reveal the forces that act on each spindle pole, we removed the central spindle in living embryos either physically with an ultraviolet laser microbeam, or genetically by RNA-mediated interference of a kinesin . We show that pulling forces external to the spindle act on the two spindle poles . A stronger net force acts on the posterior pole, thereby explaining the overall posterior displacement seen in wild-type embryos . We also show that the net force acting on each spindle pole is under control of the par genes that are required for cell polarity along the anterior-posterior embryonic axis . Finally, we discuss simple mathematical models that describe the main features of spindle pole behaviour . Our work suggests a mechanism for generating asymmetry in spindle positioning by varying the net pulling force that acts on each spindle pole, thus allowing for the generation of daughter cells with different sizes. Cell Mol Life Sci, 1999 Nov 30, 56(9-10), 742 - 54 Antibiotic resistance in microbes; Mazel D et al.; The treatment of infectious disease is compromised by the development of antibiotic-resistant strains of microbial pathogens . A variety of biochemical processes are involved that may keep antibiotics out of the cell, alter the target of the drug, or disable the antibiotic . Studies have shown that resistance determinants arise by either of two genetic mechanisms: mutation and acquisition . Antibiotic resistance genes can be disseminated among bacterial populations by several processes, but principally by conjugation . Thus the overall problem of antibiotic resistance is one of genetic ecology and a better understanding of the contributing parameters is necessary to devise rational approaches to reduce the development and spread of antibiotic resistance and so avoid a critical situation in therapy--a return to a pre-antibiotic era. Australas Phys Eng Sci Med, 2000 Sep, 23(3), 88 - 93 Monte Carlo calculation for microplanar beam radiography; Company FZ et al.; In radiography the scattered radiation from the off-target region decreases the contrast of the target image . We propose that a bundle of collimated, closely spaced, microplanar beams can reduce the scattered radiation and eliminate the effect of secondary electron dose, thus increasing the image dose contrast in the detector . The lateral and depth dose distributions of 20-200 keV microplanar beams are investigated using the EGS4 Monte Carlo code to calculate the depth doses and dose profiles in a 6 cm x 6 cm x 6 cm tissue phantom . The maximum dose on the primary beam axis (peak) and the minimum inter-beam scattered dose (valley) are compared at different photon energies and the optimum energy range for microbeam radiography is found . Results show that a bundle of closely spaced microplanar beams can give superior contrast imaging to a single macrobeam of the same overall area. Ann Med, 2000 Dec, 32(9), 585 - 7 The future of fluoroquinolones; Drlica K; The mutant prevention concentration (MPC) is a new measure of antibiotic potency above which a microbe must attain two concurrent resistance mutations for growth . For some C-8-methoxy fluoroquinolone-pathogen combinations, the value of MPC is below human serum drug concentration achieved with standard doses . Although untested clinically, such a low value of MPC, coupled with high serum concentration, should allow these fluoroquinolones to restrict severely the selection of resistant mutants when used as monotherapy . Compounds that cannot meet the MPC-pharmacokinetic criterion will enrich resistant mutants unless they are a part of combination therapy . Separation of fluoroquinolones into groups suitable for monotherapy or for combination therapy, followed by appropriate adminstration, may help extend the lifespan of the fluoroquinolones. J Vet Med B Infect Dis Vet Public Health, 2000 Dec, 47(10), 727 - 37 Relationship between the amounts of antibodies to Actinobacillus pleuropneumoniae serotype 2 detected in blood serum and in fluids collected from muscles of pigs; Wallgren P et al.; An indirect ELISA method, previously used to detect antibodies to Actinobacillus pleuropneumoniae serotype 2 in serum of pigs, was further developed aiming to measure antibodies to the microbe in muscle fluids . Serum and muscle fluid were collected from Specific Pathogen Free (SPF) pigs as well as from SPF pigs challenged with A . pleuropneumoniae which were either treated with effective antibiotics or left as infected controls . The antibody responses measured in serum correlated well to the clinical signs of respiratory disease observed and to pathological lesions found at necropsy performed 17 days post-infection . The amounts of antibodies monitored in serum and in muscle fluid collected from the diaphragm and the thigh, respectively, were compared . Higher concentrations of antibodies were assessed in serum than in diaphragm fluid, which in turn contained more antibodies per ml than fluid collected from the thigh . The amount of antibodies to A . pleuropneumoniae measured in fluid from the diaphragm diluted 1/50 correlated well with the quantity measured in serum diluted 1/1000 (r2 = 0.87; P < 0.001) . When validated by using serum antibody responses as a standard, the specificity of the ELISA employed in fluid from the diaphragm was found to be 100% . The sensitivity was determined to be 88% when calculated on seropositive pigs (A450 = 0.3 in serum diluted 1/1000) . That figure increased to 97% if calculated on pigs expressing pronounced amounts of serum antibodies (A450 > or = 0.5). J Neuropathol Exp Neurol, 2001 Jan, 60(1), 33 - 48 Effects of sodium and chloride on neuronal survival after neurite transection; Rosenberg LJ et al.; An in vitro investigation was undertaken to study the roles of Na+ and Cl- in mammalian spinal cord (SC) neuron deterioration and death after injury involving physical disruption of the plasma membrane . Individual SC neurons in monolayer cultures were subjected to UV laser microbeam transection of a primary dendrite . Neurons lesioned in modified ionic environments (MIEs) where 50%-75% of the NaCl was replaced with sucrose had higher survival (65%-75%) than neurons lesioned in medium with normal (125 mM) NaCl (28%; p < 0.001) . Subsequent experiments found a comparable increase in lesioned neuron survival in MIEs in which only Na+ was replaced with specific ionic substitutes; however, replacement of Cl- was not protective . Electron microscope examinations of neurons fixed <16 min after lesioning showed a dramatic decrease in vesiculation of the smooth endoplasmic reticulum and Golgi apparatus in the low NaCl or low Na+ MIEs . It is hypothesized that Na+ entry after membrane disruption may stimulate elevation of {Ca+2}i leading to ultrastructural disruption and death of injured neurons . The results of these studies suggest that a low NaCl MIE may be useful as an irrigant to limit damage spread and cell death within CNS tissues during surgery or after trauma. Br Poult Sci, 2000 Dec, 41(5), 575 - 83 Investigation of hygiene aspects during air chilling of poultry carcases using a model rig; Allen VM et al.; 1 . An experimental rig, designed and built to simulate conditions found in commercial poultry chilling systems, was used to investigate the effects of varying air temperature and chilling duration, and the effect of chlorinated water sprays, on the microbial load present on the skin and in the body cavity of freshly eviscerated poultry carcases; deep muscle and skin temperatures were monitored during chilling at three different temperatures . 2 . During dry chilling for 2 h, total viable microbe counts (TVC) and counts of coliforms and pseudomonads from the body cavity fell by between half and one log unit; smaller reductions were observed in samples from the breast skin . 3 . The situation changed when chlorinated water sprays (50, 100 or 250 ppm available chlorine) were applied for the first hour of chilling; spraying carcases enhanced the reduction in numbers on the skin; the effect was most pronounced with 250 ppm chlorine; conversely in the body cavity, the general effects of sprays was to increase contamination by up to one log unit . 4 . There was no evidence that sprays increased the rate of chilling . 5 . When carcases were held overnight in the rig at 11 degrees C after chilling, microbe counts on dry-chilled carcases remained stable, but increased on carcases that had been sprayed with chlorinated water. J Behav Med, 2000 Dec, 23(6), 519 - 29 Serum cholesterol concentrations and mood states in violent psychiatric patients: an experience sampling study; Hillbrand M et al.; The well-documented negative association between serum cholesterol and aggressive behavior has led Kaplan to propose a cholesterol-serotonin hypothesis of aggression . According to this hypothesis, low dietary cholesterol intake leads to depressed central serotonergic activity, which itself has been reported in numerous studies of violent individuals . In the present study, 25 violent psychiatric patients participated in a microbehavioral experience sampling procedure to examine differences in self-reports of affective and cognitive experiences as a function of serum cholesterol concentrations . For 7 days, they wore signaling devices that emitted an average of seven signals a day . Following each signal, patients filled out a mood questionnaire . Total serum cholesterol (TSC) concentration was positively associated with measures of affect, cognitive efficiency, activation, and sociability, suggesting a link between low TSC and dysphoria . These findings are consistent with the cholesterol-serotonin hypothesis and with the substantive literature linking both aggression and depression to depressed central serotonergic activity. J Food Prot, 2001 Jan, 64(1), 94 - 8 Changes in K value and microorganisms of tilapia fillet during storage at high-pressure, normal temperature; Ko WC et al.; This study determined the effect of high-pressure, normal temperature (25 degrees C) storage on tilapia fillets . After pressurization, the fillets were stored at normal condition (1 atm {1 atm = 101.29 kPa} and 25 degrees C) for 12 h to evaluate the changes of microbes and K value . The fillets stored at 2,000 atm for 12 h still kept the K value, a freshness index that represents putrefaction with the value beyond 60%, still below 40%, and the K value of the meat without pressurization was up to 92% . Total plate counts of the fillets stored at below 1,000 atm for 12 h were maintained at the value of 4.7 log CFU/g of meat, which was similar to the initial level . However, the counts were obviously decreased to about 2.0 log CFU/g of meat for the fillets stored at above 2,000 atm . The same effect was obtained for psychrophilic bacteria . Enzymes and microbes reactivated apparently after 12 h of normal condition storage of mild pressurized fillets . The study demonstrates that high-pressure storage can inhibit the putrefaction of tilapia meat but no longer after that. Electrophoresis, 2001 Jan, 22(1), 23 - 8 Hundredfold productivity of genome analysis by introduction of microtemperature-gradient gel electrophoresis; Biyani M et al.; Genome profiling, which employs temperature-gradient gel electrophoresis (TGGE) for DNA analysis, has recently been developed in identifying species by genotype . However, the performance of this technology like the general applications of TGGE was, though highly informative, limited in its ability due to methodological reasons . This study demonstrates that minimization of the gel for TGGE, to around one-tenth of its conventional size (approximately 2 cm), can be successfully introduced, resulting in a hundredfold higher performance (total evaluation of time, cost, and degree of parallel operations) than that of the conventional . Reproducibility was evaluated from the measures of the pattern similarity scores (PaSS) between band patterns (genome profiles) obtained with the conventional TGGE, and that with micro-TGGE (microTGGE) developed here, after extracting a set of featuring points from genome profiles . Size minimization, which leads to the reduction of the amount of samples required (cost-saving), is another great advantage, enhancing the employment of multicolor fluorescence technology . Since the further development of microbe-related fields such as epidemiology and microbial ecology inevitably require knowledge based on the identification of a great number of species and strains, microbe-related fields will receive the most optimal benefits from the technological improvements attained here. Nature, 2001 Jan 11, 409(6817), 188 - 91 Nitrogen limitation of microbial decomposition in a grassland under elevated CO2; Hu S et al.; Carbon accumulation in the terrestrial biosphere could partially offset the effects of anthropogenic CO2 emissions on atmospheric CO2 . The net impact of increased CO2 on the carbon balance of terrestrial ecosystems is unclear, however, because elevated CO2 effects on carbon input to soils and plant use of water and nutrients often have contrasting effects on microbial processes . Here we show suppression of microbial decomposition in an annual grassland after continuous exposure to increased CO2 for five growing seasons . The increased CO2 enhanced plant nitrogen uptake, microbial biomass carbon, and available carbon for microbes . But it reduced available soil nitrogen, exacerbated nitrogen constraints on microbes, and reduced microbial respiration per unit biomass . These results indicate that increased CO2 can alter the interaction between plants and microbes in favour of plant utilization of nitrogen, thereby slowing microbial decomposition and increasing ecosystem carbon accumulation. Anal Chem, 2001 Jan 1, 73(1), 1 - 7 Surface plasmon resonance imaging measurements of DNA and RNA hybridization adsorption onto DNA microarrays; Nelson BP et al.; Surface plasmon resonance (SPR) imaging is a surface-sensitive spectroscopic technique for measuring interactions between unlabeled biological molecules with arrays of surface-bound species . In this paper, SPR imaging is used to quantitatively detect the hybridization adsorption of short (18-base) unlabeled DNA oligonucleotides at low concentration, as well as, for the first time, the hybridization adsorption of unlabeled RNA oligonucleotides and larger 16S ribosomal RNA (rRNA) isolated from the microbe Escherichia coli onto a DNA array . For the hybridization adsorption of both DNA and RNA oligonucleotides, a detection limit of 10 nM is reported; for large (1,500-base) 16S rRNA molecules, concentrations as low as 2 nM are detected . The covalent attachment of thiol-DNA probes to the gold surface leads to high surface probe density (10(12) molecules/cm2) and excellent probe stability that enables more than 25 cycles of hybridization and denaturing without loss in signal or specificity . Fresnel calculations are used to show that changes in percent reflectivity as measured by SPR imaging are linear with respect to surface coverage of adsorbed DNA oligonucleotides . Data from SPR imaging is used to construct a quantitative adsorption isotherm of the hybridization adsorption on a surface . DNA and RNA 18-mer oligonucleotide hybridization adsorption is found to follow a Langmuir isotherm with an adsorption coefficient of 1.8 x 10(7) M(-1). Theriogenology, 2000 Nov 1, 54(8), 1281 - 4 Mummification of fetal membranes in the bovine vagina: a case report; Jalakas M; Papyraceous, mummified fetal membranes were found in the vagina of a cow at the end of the fourth month after a normal delivery . The uterus and cervix were without any pathological changes . The cow conceived on Day 36 after removal of the mummified fetal membranes upon the second insemination . Apart from the uncommon nature of the case, it proves that after calving the bovine vagina can maintain a sterile environment and that the constrictive mechanism of the vagina can be strong enough to hinder for a long time the discharge of fetal membranes and access of microbes to the vagina. Cell Stress Chaperones, 2000 Nov, 5(5), 458 - 61 Regulation of immune activity by mild (fever-range) whole body hyperthermia: effects on epidermal Langerhans cells; Ostberg JR et al.; Inflammation of the skin and systemic fever, both of which occur with injury or infection, include a hyperthermic component that many believe constitutes a physiological stress . Such increases in local or systemic body temperature may also have a regulatory effect on immune function . Langerhans cells (LCs), the dendritic cells of the skin, continuously monitor the extracellular matrix of the skin by taking up particles and microbes that they then carry to draining lymph nodes for presentation to T lymphocytes . We hypothesize that the thermal element of inflammation and/or fever may help regulate the activation and migration of LCs out of the epidermis . To test this hypothesis, Balb/ c mice were exposed to a mild (39.8 degrees C +/- 0.2 degrees C), long-duration (6 hours) whole body hyperthermia (WBH) treatment, which mimics the thermal component of fever . The number of LCs and their morphology were analyzed at various time points up to 7 days after the initiation of WBH . The LCs of the ear epidermis were visualized using a fluorescein isothiocyanate-conjugated antibody specific for the major histocompatibility complex (MHC) class II molecule and confocal microscopy . Although MHC class II staining was diffuse on the surface of the LC body and dendritic extensions of both WBH and control samples, the WBH-treated LCs exhibited a more punctate morphology with fewer dendritic processes compared with control LCs . A significant decrease in the number of LCs was also observed 1 to 5 days after WBH treatment . Furthermore, in vitro heating of Balb/c ear skin cultures at 40 degrees C for 6 to 8 hours enhanced the numbers of viable LCs that migrated into the culture wells . These results suggest that WBH treatment stimulates epidermal LCs in the absence of foreign antigen. Nippon Jibiinkoka Gakkai Kaiho, 2000 Nov, 103(11), 1218 - 26 {Isolation of FDC-lymphocyte clusters from human tonsillar tissues}; Takemoto N; Follicular B cells and follicular dendritic cells (FDCs) form FDC-lymphocyte clusters and play a central role in events related to humoral immunity in the lymphoid follicle (LF) . The secondary LF is divided into five zones, each of which exhibits functional differences . However, the distribution of the clusters across the five follicular zones remains unclear . We here report a procedure for isolating FDC-lymphocyte clusters from fixed tonsillar tissues and compare of the structure of clusters isolated from three follicular zones: the mantle, light and dark zones . First, the germinal centers (GCs) of the secondary LFs were removed under a stereoscope, and the GCs were enzymatically digested for 20, 30, 40 and 50 minutes at 37 degrees C . The FDC-lymphocyte clusters were then isolated using a discontinuous density gradient and a Magnetic Particle Concentrator, followed by microbeads . The number of isolated medium sized clusters composed of 6-25 cells was greatest when the samples were incubated for 40 minutes . To detect the mantle, light and dark zones, and GCs, isolated FDC-lymphocyte clusters from each zone were immunostained . Their cell structures were then compared . The clusters were composed mainly of B cells (comprising about 80% of the cells in each cluster, on average), T cells, natural killer/T cells and macrophages were also observed, but less frequently . The proportions of CD 45 RO-positive cells and CD4-positive cells were clearly different for each zone, with CD4-positive cells in the majority . No clear differences in isolated clusters from fixed and unfixed tonsillar tissues were observed . Our data indicate that this procedure is suitable for isolating FDC-lymphocyte clusters from fixed lymphoid tissues and that the proportions of cells composing the clusters differ in the three follicular zones. Traffic, 2000 Apr, 1(4), 295 - 300 Pathways for lipid antigen presentation by CD1 molecules: nowhere for intracellular pathogens to hide; Sugita M et al.; A crucial feature of peptide antigen presentation by major histocompatibility complex (MHC) class I and II molecules is their differential ability to sample cytosolic and extracellular antigens . Intracellular viral infections and bacteria that are taken up in phagosomes, but then escape from the endocytic compartment efficiently, enter the class I pathway via the cytosol . In contrast, phagosome-resident bacteria yield protein antigens that are sampled deep in the endocytic compartment and presented in a vacuolar acidification-dependent pathway mediated by MHC class II molecules . Despite this potential for antigen sampling, microbes have evolved a variety of evasive mechanisms that affect peptide transport in the MHC class I pathway or blockade of endosomal acidification and inhibition of phagosome-lysosome fusion that may compromise the MHC class II pathway of antigen presentation . Thus, besides MHC class I and II, a third lineage of antigen-presenting molecules that bind lipid and glycolipid antigens rather than peptides exists and is mediated by the family of CD1 proteins . CD1 isoforms (CD1a, b, c, and d) differentially sample both recycling endosomes of the early endocytic system and late endosomes and lysosomes to which lipid antigens are differentially delivered . These CD1 pathways include vacuolar acidification-independent pathways for lipid antigen presentation . These features of presenting lipid antigens, independently monitoring various antigen-containing intracellular compartments and avoiding certain evasive techniques employed by microbes, enable CD1 molecules to provide distinct opportunities to function in host defense against the microbial world. Plant J, 2001 Feb, 25(3), 281 - 93 The spatial expression patterns of a phosphate transporter (MtPT1) from Medicago truncatula indicate a role in phosphate transport at the root/soil interface; Chiou TJ et al.; The movement of phosphate from the soil into plant root cells is the first of many crucial transport events required to supply phosphorous (P) to cells throughout the plant . In addition to the ability to acquire phosphate from the soil, the majority of the vascular plants are able to form arbuscular mycorrhizal associations in which phosphate may be delivered to the cortex via a fungus . Previously, we cloned two phosphate transporter genes, MtPT1 and MtPT2 from Medicago truncatula roots . Complementation of a yeast phosphate transport mutant revealed that MtPT1 is a functional phosphate transporter and Northern analyses revealed that MtPT1 is expressed exclusively in roots (Liu et al., 1998, Mol . Plant-Microbe Interact . 11, 14--22) . Utilising an antibody specific for MtPT1, we have analysed the accumulation and spatial expression patterns of the MtPT1 transporter . MtPT1 transcript and protein levels show close correlation and increase dramatically in the roots in response to phosphate starvation . MtPT1 protein levels decrease in roots during development of a symbiosis with arbuscular mycorrhizal (AM) fungi, indicating that this transporter is not involved in symbiotic phosphate transport . Membrane fractionation and analysis of a MtPT1/GFP fusion protein revealed that MtPT1 is located in the plasma membrane, while in situ hybridisation and immunolocalisation demonstrate the presence of MtPT1 transcripts and protein in the epidermal cells and root hairs of M . truncatula roots . MtPT1 shows expression patterns consistent with a role specifically in the acquisition of phosphate from the soil and is distinct from the other phosphate transporter of this class described to date. Cell Microbiol, 1999 Nov, 1(3), 205 - 14 Phagocytic processing of antigens for presentation by class II major histocompatibility complex molecules; Ramachandra L et al.; Microbes and other particulate antigens (Ags) are internalized by phagocytosis and then reside in plasma membrane-derived phagosomes . The contribution of phagosomes to the degradation of Ags has long been appreciated . It has been unclear, however, whether peptides derived from these degraded antigens bind class II major histocompatibility complex (MHC-II) molecules within phagosomes or within endocytic compartments that receive Ag fragments from phagosomes . Recent experiments have demonstrated that phagosomes containing Ag-conjugated latex beads express a full complement of Ag-processing molecules, e.g . MHC-II molecules, invariant chain, H2-DM and proteases sufficient to degrade bead- associated Ag . These phagosomes mediate the formation of peptide-MHC-II complexes, which are transported to the cell surface and presented to T cells . Phagosomes acquire both newly synthesized and plasma membrane-derived MHC-II molecules, but the formation of peptide-MHC-II complexes in phagosomes primarily involves newly synthesized MHC-II molecules . The content and traffic of phagosomal proteins vary considerably with the type of Ag ingested . Pathogenic microbes can alter phagosome composition and function to reduce Ag processing . For example, Mycobacterium tuberculosis blocks the maturation of phagosomes and reduces the ability of infected cells to present exogenous soluble protein Ags. Radiat Res, 2001 Mar, 155(3), 402 - 8 The bystander effect in radiation oncogenesis: II . A quantitative model; Brenner DJ et al.; There is strong evidence that biological response to ionizing radiation has a contribution from unirradiated "bystander" cells that respond to signals emitted by irradiated cells . We discuss here an approach incorporating a radiobiological bystander response, superimposed on a direct response due to direct energy deposition in cell nuclei . A quantitative model based on this approach is described for alpha-particle-induced in vitro oncogenic transformation . The model postulates that the oncogenic bystander response is a binary "all or nothing" phenomenon in a small sensitive subpopulation of cells, and that cells from this sensitive subpopulation are also very sensitive to direct hits from alpha particles, generally resulting in a directly hit sensitive cell being inactivated . The model is applied to recent data on in vitro oncogenic transformation produced by broad-beam or microbeam alpha-particle irradiation . Two parameters are used in analyzing the data for transformation frequency . The analysis suggests that, at least for alpha-particle-induced oncogenic transformation, bystander effects are important only at small doses-here below about 0.2 Gy . At still lower doses, bystander effects may dominate the overall response, possibly leading to an underestimation of low-dose risks extrapolated from intermediate doses, where direct effects dominate. Radiat Res, 2001 Mar, 155(3), 397 - 401 The bystander effect in radiation oncogenesis: I . Transformation in C3H 10T1/2 cells in vitro can be initiated in the unirradiated neighbors of irradiated cells; Sawant SG et al.; It has long been accepted that radiation-induced genetic effects require that DNA be hit and damaged directly by the radiation . Recently, evidence has accumulated that in cell populations exposed to low doses of alpha particles, biological effects occur in a larger proportion of cells than are estimated to have been traversed by alpha particles . The end points observed include chromosome aberrations, mutations and gene expression . The development of a fast single-cell microbeam now makes it possible to expose a precisely known proportion of cells in a population to exactly defined numbers of alpha particles, and to assay for oncogenic transformation . The single-cell microbeam delivered no, one, two, four or eight alpha particles through the nuclei of all or just 10% of C3H 10T1/2 cells . We show that (a) more cells can be inactivated than were actually traversed by alpha particles and (b) when 10% of the cells on a dish are exposed to alpha particles, the resulting frequency of induced transformation is not less than that observed when every cell on the dish is exposed to the same number of alpha particles . These observations constitute evidence suggesting a bystander effect, i.e., that unirradiated cells are responding to damage induced in irradiated cells . This bystander effect in a biological system of relevance to carcinogenesis could have significant implications for risk estimation for low-dose radiation. Toxicol Appl Pharmacol, 2001 Feb 15, 171(1), 61 - 9 Inflammatory responses in mice after intratracheal instillation of spores of Streptomyces californicus isolated from indoor air of a moldy building; Jussila J et al.; Microbial growth in buildings is associated with respiratory symptoms in the occupants . However, the specific effects of the microbes and the way they provoke clinical manifestations are poorly understood . In the current study, mice were exposed via intratracheal instillation to single doses of the spores of Streptomyces californicus, isolated from indoor air of a moisture-damaged building (2.2 x 10(7), 1.1 x 10(8), and 3.3 x 10(8) spores), or lipopolysaccharide (50 microg) . Inflammation and toxicity in lungs were evaluated 24 h later . The time course of the effects was explored with the dose of 1.1 x 10(8) spores for up to 7 days . The microbial spores elevated proinflammatory cytokine (i.e., TNFalpha and IL-6) levels in bronchoalveolar lavage fluid (BALF) and in serum in a dose- and time-dependent manner and evoked expression of inducible nitric oxide synthase in BAL cells . Both TNFalpha and IL-6 responses peaked at 6 h after instillation, but TNFalpha leveled off more quickly than IL-6 . The cytokine surge was followed by inflammatory cell recruitment into airways . Moreover, the spores increased dose- and time-dependently total protein, albumin, hemoglobin, and lactate dehydrogenase concentrations in BALF during the first 24 h . Histopathological examination of lungs confirmed the inflammatory changes . With the exception of macrophage and lymphocyte numbers, all parameters returned to control level at 7 days . In summary, these observations indicate that the spores of S . californicus are capable of provoking an acute inflammation in mouse lungs and can cause cytotoxicity . Thus, S . californicus can be considered as a species with potential to cause adverse health effects in occupants of moisture-damaged buildings. Proc Natl Acad Sci U S A, 2001 Feb 13, 98(4), 1376 - 80 Interaction of the herbicide glyphosate with its target enzyme 5-enolpyruvylshikimate 3-phosphate synthase in atomic detail; Schonbrunn E et al.; Biosynthesis of aromatic amino acids in plants, many bacteria, and microbes relies on the enzyme 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase, a prime target for drugs and herbicides . We have identified the interaction of EPSP synthase with one of its two substrates (shikimate 3-phosphate) and with the widely used herbicide glyphosate by x-ray crystallography . The two-domain enzyme closes on ligand binding, thereby forming the active site in the interdomain cleft . Glyphosate appears to occupy the binding site of the second substrate of EPSP synthase (phosphoenol pyruvate), mimicking an intermediate state of the ternary enzyme.substrates complex . The elucidation of the active site of EPSP synthase and especially of the binding pattern of glyphosate provides a valuable roadmap for engineering new herbicides and herbicide-resistant crops, as well as new antibiotic and antiparasitic drugs. J Hosp Infect, 2000 Dec, 46(4), 241 - 56 The index of microbial air contamination; Pasquarella C et al.; The standard index of microbial air contamination (IMA) for the measurement of microbial air contamination in environments at risk is described . The method quantifies the microbial flow directly related to the contamination of surfaces coming from microbes that reach critical points by falling on to them . The index of microbial air contamination is based on the count of the microbial fallout on to Petri dishes left open to the air according to the 1/1/1 scheme (for 1h, 1m from the floor, at least 1m away from walls or any obstacle) . Classes of contamination and maximum acceptable levels have been established . The index of microbial air contamination has been tested in many different places: in hospitals, in food industries, in art galleries, aboard the MIR space station and also in the open air . It has proved to be a reliable and useful tool for monitoring the microbial surface contamination settling from the air in any environment . Cytometry, 2001 Feb 1, 43(2), 154 - 60 The "vanishing counting bead" phenomenon: effect on absolute CD34+ cell counting in phosphate-buffered saline-diluted leukapheresis samples; Brando B et al.; BACKGROUND: Using a single-platform protocol to count absolute CD34+ hematopoietic precursor cell (HPC) levels with different reference microbeads, we recorded occasionally artifactually high CD34+ HPC counts in some leukapheresis bags, whereas dual-platform calculations were always consistent . Abnormal countings were observed only when phosphate-buffered saline (PBS)-diluted leukapheresis samples were vortexed before analysis . A large series of blood samples analyzed similarly for CD34+ and CD4+ absolute counts did not show any sample or vortexing effect . With the volumetric absolute counting cytometer Partec-PAS, lower counts were also observed when different reference beads were vortexed before the instrument checking procedures . The counting abnormality was caused by a drop in microbead concentration (the "vanishing bead phenomenon") . This phenomenon reduced the total and relative bead event number in experimental and routine samples and in calibration procedures . This altered the bead denominator used to calculate absolute CD34+ HPC levels and it also reduced the concentration of standard calibration beads . METHODS: Using the Partec-PAS to measure volumetrically the actual bead concentration, we studied the vanishing bead phenomenon . Different types of counting and reference microbeads were resuspended in media with or without proteins or cells . Replicates were submitted either to gentle manual mixing or to vortexing before counting . RESULTS: Vortex agitation almost invariably induced the vanishing bead phenomenon when beads were resuspended in saline media or when an insufficient protein concentration was present, such as in diluted leukapheresis samples . Different bead types showed various degrees of sensitivity to vortexing . The bead disappearance was not caused by bubble formation or disruption . The addition of small amounts of protein completely prevented the vanishing bead phenomenon . The causative effect of the electrostatic charging of tube induced by vortexing is hypothesized . CONCLUSIONS: Sample suspensions containing counting beads for single-platform analysis must be resuspended in media with protein supplements to prevent the vanishing bead phenomenon and to ensure accurate counting . Immunology, 2001 Jan, 102(1), 2 - 7 Co-option of endocytic functions of cellular caveolae by pathogens; Shin JS et al.; It is increasingly becoming clear that various immune cells are infected by the very pathogens that they are supposed to attack . Although many mechanisms for microbial entry exist, it appears that a common route of entry shared by certain bacteria, viruses and parasites involves cellular lipid-rich microdomains sometimes called caveolae . These cellular entities, which are characterized by their preferential accumulation of glycosylphosphatidylinositol (GPI)-anchored molecules, cholesterol and various glycolipids, and a distinct protein (caveolin), are present in many effector cells of the immune system including neutrophils, macrophages, mast cells and dendritic cells . These structures have an innate capacity to endocytoze various ligands and traffic them to different intracellular sites and sometimes, back to the extracellular cell surface . Because caveolae do not typically fuse with lysosomes, the ligands borne by caveolar vesicles are essentially intact, which is in marked contrast to ligands endocytozed via the classical endosome-lysosome pathway . A number of microbes or their exotoxins co-opt the unique features of caveolae to enter and traffic, without any apparent loss of viability and function, to different sites within immune and other host cells . In spite of their wide disparity in size and other structural attributes, we predict that a common feature among caveolae-utilizing pathogens and toxins is that their cognate receptor(s) are localized within plasmalemmal caveolae of the host cell. Immunol Cell Biol, 2001 Feb, 79(1), 62 - 6 Genetic 'budget' of viruses and the cost to the infected host: a theory on the relationship between the genetic capacity of viruses, immune evasion, persistence and disease; Chaston TB et al.; The nature of the pathogen-host relationship is recognized as being a dynamic coevolutionary process where the immune system has required ongoing adaptation and improvement to combat infection . Under survival pressure from sophisticated immune responses, adaptive processes for microbes, including viruses, have manifested as immune evasion strategies . This paper proposes a theory that virus immune evasion can be broadly classified into 'acquisition' or 'erroneous replication' strategies . Acquisition strategies are characteristic of large genome dsDNA viruses, which (i) replicate in the cell nucleus; (ii) have acquired host genes that can be used to directly manipulate responses to infection; (iii) are often latent for the lifetime of the host; and (iv) have little or no serious impact on health . Alternatively, erroneous replication strategies are characteristic of small genome RNA viruses, which are recognized as being the cause of many serious diseases in humans . It is proposed that this propensity for disease is due to the cytoplasmic site of replication and truncated temporal relationship with the host, which has limited or removed the evolutionary opportunity for RNA viruses to have acquired host genes . This has resulted in RNA viruses relying on error-prone replication strategies which, while allowing survival and persistence, are more likely to lead to disease due to the lack of direct viral control over potentially host-deleterious inflammatory and immune responses to infection. Plant Sci, 2000 Dec 7, 160(1), 1 - 13 Organic acid metabolism in plants: from adaptive physiology to transgenic varieties for cultivation in extreme soils; Lopez-Bucio J et al.; During the last 20 years increasing experimental evidence has associated organic acid metabolism with plant tolerance to environmental stress . Current knowledge shows that organic acids not only act as intermediates in carbon metabolism but also as key components in mechanisms that some plants use to cope with nutrient deficiencies, metal tolerance and plant-microbe interactions operating at the root-soil interphase . In this review we summarize recent knowledge on the physiology and occurrence of organic acids in plants and their special relevance concerning nitrate reduction, phosphorus and iron acquisition, aluminum tolerance and soil ecology . We also discuss novel findings in relation to the biotechnological manipulation of organic acids in transgenic models ranging from cell cultures to whole plants . This novel perspective of organic acid metabolism and its potential manipulation may represent a way to understand fundamental aspects of plant physiology and lead to new strategies to obtain crop varieties better adapted to environmental and mineral stress. Cancer Lett, 2001 Jan, 162 Suppl, S11 - S16 Predictive laboratory diagnostics in oncology utilizing blood-borne cancer cells--current best practice and unmet needs; Brandt BH et al.; The aim of laboratory diagnostics in oncology is to improve the clinical outcome of cancer by allowing earlier detection . Molecular knowledge of cancer should increase the number of risk and prognostic factors and will allow development of methods for detection and elimination of even very small tumors . Thus, the race for the specific tumor antigen in peripheral blood and the race for the blood-borne cancer cell happened simultaneously . The direct detection of the cells which have the highest probability to harbor all the properties mandatory to be life-threatening, conceivably metastatic, would be the most promising way to find the target structure of malignancy . Methods applying enrichment techniques based on density, morphology, tissue specific protein and tumor-associated protein detection enabled multi-parametric analysis of those blood-borne cancer cells . In exemplary studies it was demonstrated that the count of cell clusters positive for the tissue-specific proteins cytokeratin and prostate-specific antigen (PSA) from the peripheral blood of prostate cancer patients and a combination of a tissue-specific protein, a oncogenic receptor protein cytokeratin and p185(c-erbB-2) from the peripheral blood of breast cancer patients is related to the stage of the diseases . Breast cancer patients who presented with cytokeratin/p185(c-erbB-2) positive cell clusters showed a decrease of those cells under adriamycin adjuvant therapy . Nevertheless, additional molecular markers are required to characterize the functional properties of blood-borne cancer cells . Therefore, the genome of the cells can be investigated using a procedure for indirectly detecting aberrations of defined gene locations, i.e . multiplex microsatellite polymerase chain reaction . Up to now, the methods applied to the separation of blood-borne cancer cells are time-consuming and rather expensive . They consist of an initial enrichment step of density gradient centrifugation or buffy coat preparation followed by a specific isolation step using superparamagnetic microbeads coupled to antibodies, filter techniques or multi-parametric flow cytometry . Novel technologies have to be applied using miniaturization, integration and parallel-processing techniques based on those used in the computer industry to overcome the drawbacks. J Clin Virol, 2001 Jan, 20(1-2), 1 - 6 Automation of laboratory testing for infectious diseases using the polymerase chain reaction-- our past, our present, our future; Jungkind D; While it is an extremely powerful and versatile assay method, polymerase chain reaction (PCR) can be a labor-intensive process . Since the advent of commercial test kits from Roche and the semi-automated microwell Amplicor system, PCR has become an increasingly useful and widespread clinical tool . However, more widespread acceptance of molecular testing will depend upon automation that allows molecular assays to enter the routine clinical laboratory . The forces driving the need for automated PCR are the requirements for diagnosis and treatment of chronic viral diseases, economic pressures to develop more automated and less expensive test procedures similar to those in the clinical chemistry laboratories, and a shortage in many areas of qualified laboratory personnel trained in the types of manual procedures used in past decades . The automated Roche COBAS AMPLICOR system has automated the amplification and detection process . Specimen preparation remains the most labor-intensive part of the PCR testing process, accounting for the majority of the hands-on-time in most of the assays . A new automated specimen preparation system, the COBAS AmpliPrep, was evaluated . The system automatically releases the target nucleic acid, captures the target with specific oligonucleotide probes, which become attached to magnetic beads via a biotin-streptavidin binding reaction . Once attached to the beads, the target is purified and concentrated automatically . Results of 298 qualitative and 57 quantitative samples representing a wide range of virus concentrations analyzed after the COBAS AmpliPrep and manual specimen preparation methods, showed that there was no significant difference in qualitative or quantitative hepatitis C virus (HCV) assay performance, respectively . The AmpliPrep instrument decreased the time required to prepare serum or plasma samples for HCV PCR to under 1 min per sample . This was a decrease of 76% compared to the manual specimen preparation method . Systems that can analyze more samples with higher throughput and that can answer more questions about the nature of the microbes that we can presently only detect and quantitate will be needed in the future. Aquat Toxicol, 2001 Mar, 52(1), 61 - 71 The use of microangiography in detecting aberrant vasculature in zebrafish embryos exposed to cadmium; Cheng SH et al.; Embryonic vascular patterns in zebrafish (Danio rerio) could be visualised by confocal microscopy coupled with microinjected fluorescent microbeads . This microangiographic technique was adopted here, for the first time, to study the effects of cadmium on cardiovascular development in zebrafish embryos . Zebrafish embryos were incubated in culture medium containing 100 microM cadmium from 5 h post fertilisation (hpf) to 48 hpf . At 48 hpf, embryos were examined for viability and occurrence of malformations . The 100 microM cadmium caused 32.21 +/- 3.65% mortality and 20.33 +/- 4.04% visible malformations in surviving embryos . In the remaining embryos with no visible signs of malformations, further assessments for less obvious abnormalities were performed . Assessments on craniofacial development were made by digital measurements on areas of brains and eyes . Cardiac development was assessed by immunostaining the heart with the antibody MF20 specific for myosin heavy chain . Body lengths of the embryos were also measured . Embryonic development of brains, eyes, hearts and body lengths of visibly healthy embryos in the cadmium treatment group showed no significant difference from the controls . Embryonic vasculature of these visibly healthy embryos was then studied by microinjecting fluorescent microbeads of diameter 0.02 microm into the circulation . All the cadmium treated embryos showed localised vascular defects in the dorsal aortae, segmental and cranial vessels while none of the control embryos showed any aberrant patterns in the networking of the vasculature . Improved image analyses on the anterior regions revealed that cadmium treated embryos had markedly less complex networks of cranial vessels with fewer vessels perfusing the craniofacial regions . The number of branch points in the vascular network was counted . In untreated embryos, there were 135.6 +/- 51 branches in the vasculature in entire body . In the cadmium treated embryos, there were 64.5+/-31 branches . The difference was significant when assessed with Student's t-test . It appeared that although cadmium did not cause any signs of external malformations in these visibly healthy embryos, nonetheless induced impaired branching and anastomsis of the cranial vessels . This study revealed, for the first time, that vital vascular structures in fish embryos could be affected by exposure to cadmium . This technique allowed visualisation of vascular anomalies in embryos showing no external signs of malformations . The impairment of anatomical features during embryonic development might serve as meaningful health endpoints in ecotoxicological studies and in risk assessment. Environ Res, 2001 Feb, 85(2), 135 - 44 Exposure assessment of indoor allergens, endotoxin, and airborne fungi for homes in southern Taiwan; Su HJ et al.; This study was undertaken to examine the seasonal variations of domestic Der p 1, Der p 2, and endotoxin on mattress and airborne fungal concentrations in homes of asthmatic and nonasthmatic children in southern Taiwan, where temperature and relative humidity are usually high throughout the year . A group of asthmatic children (10-12 years old) were selected randomly based on a citywide questionnaire survey . The nonasthmatic children were chosen to be in the comparison group by matching in age, gender, and proximity of residence . Environmental sampling of domestic microbes was conducted once a month for a year . Twelve calendar months were grouped into spring, summer, fall, and winter according to weather data (mainly average temperature and humidity) from the Central Weather Bureau . Dust samples from a child's mattress and airborne samples from a child's bedroom were collected and analyzed for allergens of Der p 1 and Der p 2, endotoxin, and fungi respectively . Results show that about 65% of children's mattresses in our region have Der p 1 levels greater than 2 microg/g . It is also apparent that most airborne fungal concentrations found in homes of either asthmatic or nonasthmatic children are higher than the recommended levels of concern . The predominant genera are Cladosporium, Aspergillus, Penicillium, Alternaria, and yeast . In addition, seasonal effects seem to be a critical factor for the concentrations and distributions of domestic endotoxin in these study homes . The implication of long-term exposure to these high levels of environmental microbes and how their effects vary with seasons remain to be further characterized . Plant Physiol, 2001 Feb, 125(2), 728 - 37 LEPS2, a phosphorus starvation-induced novel acid phosphatase from tomato; Baldwin JC et al.; Phosphate (Pi) is one of the least available plant nutrients found in the soil . A significant amount of phosphate is bound in organic forms in the rhizosphere . Phosphatases produced by plants and microbes are presumed to convert organic phosphorus into available Pi, which is absorbed by plants . In this study we describe the isolation and characterization of a novel tomato (Lycopersicon esculentum) phosphate starvation-induced gene (LePS2) representing an acid phosphatase . LePS2 is a member of a small gene family in tomato . The cDNA is 942 bp long and contains an open reading frame encoding a 269-amino acid polypeptide . The amino acid sequence of LePS2 has a significant similarity with a phosphatase from chicken . Distinct regions of the peptide also share significant identity with the members of HAD and DDDD super families of phosphohydrolases . Many plant homologs of LePS2 are found in the databases . The LePS2 transcripts are induced rapidly in tomato plant and cell culture in the absence of Pi . However, the induction is repressible in the presence of Pi . Divided root studies indicate that internal Pi levels regulate the expression of LePS2 . The enhanced expression of LePS2 is a specific response to Pi starvation, and it is not affected by starvation of other nutrients or abiotic stresses . The bacterially (Escherichia coli) expressed protein exhibits phosphatase activity against the synthetic substrate p-nitrophenyl phosphate . The pH optimum of the enzyme activity suggests that LePS2 is an acid phosphatase. Am J Clin Nutr, 2001 Feb, 73(2 Suppl), 393S - 398S Quality assurance criteria for probiotic bacteria; Tuomola E et al.; Acid and bile stability and intestinal mucosal adhesion properties are among the criteria used to select probiotic microbes . The quality control of probiotic cultures in foods traditionally has relied solely on tests to ensure that an adequate number of viable bacteria are present in the products throughout their shelf lives . Viability is an important factor, but not the only criterion for quality assurance . To be effective, probiotic strains must retain the functional health characteristics for which they were originally selected . Such characteristics include the ability to survive transit through the stomach and small intestine and to colonize the human gastrointestinal tract . In vitro test protocols can be readily adopted to examine the maintenance of a strain's ability to tolerate acidic conditions, survive and grow in the presence of bile, and metabolize selective substrates . Molecular techniques are also available to examine strain stability . Adhesion characterization may be an important quality-control method for assessing gut barrier effects . Adhesion has been related to shortening the duration of diarrhea, immunogenic effects, competitive exclusion, and other health effects . Adhesion properties should be carefully monitored, including adhesion to intestinal cells (eg, Caco-2) and human intestinal mucus . This article outlines the types of in vitro testing that can be used to ensure quality control of functional probiotic strains. J Immunol, 2000 Apr 15, 164(8), 3946 - 9 Cutting edge: infection by the agent of human granulocytic ehrlichiosis prevents the respiratory burst by down-regulating gp91phox; Banerjee R et al.; The agent of human granulocytic ehrlichiosis (HGE) is an emerging tick-borne pathogen that resides in neutrophils and can be cultured in a promyelocytic (HL-60) cell line . In response to microbes, polymorphonuclear leukocytes normally activate the NADPH oxidase enzyme complex and generate superoxide anion (O2-) . However, HL-60 cells infected with HGE bacteria did not produce O2- upon activation with PMA . RT-PCR demonstrated that HGE organisms inhibited mRNA expression of a single component of NADPH oxidase, gp91phox, and FACS analysis showed that plasma membrane-associated gp91phox protein was reduced on the infected cells . Infection with HGE organisms also decreased gp91phox mRNA levels in splenic neutrophils in a murine model of HGE, demonstrating this phenomenon in vivo . Therefore, HGE bacteria repress the respiratory burst by down-regulating gp91phox, the first direct inhibition of NADPH oxidase by a pathogen. Int J Radiat Biol, 1999 Aug, 75(8), 1015 - 9 Lethality of single-track events: comparison between calculations and experimental data; Sato Y et al.; Recent results on cell killing by microbeams were analysed using three parameters (k, L, L1), where k is the mean number of lethal particles per cell nucleus, L {keV/microm} is the track-average LET (linear energy transfer) in cells, and L1 {keV/microm} is a critical value for inducing lethal damage by a single track . Analysis showed that calculations are consistent with two data sets . The existence of a quadratic dependence on LET of cellular effects is confirmed in the high-LET region between 30 and 500 keV/microm . L1 approximately 150 keV/microm was found to give the best fit and the sensitive area of the cell nucleus was determined as approximately 50 microm2 for Chinese hamster V79 cells . In microbeam experiments with several MeV alpha-particles, the relationship between k and L for V79 cells can be expressed as L = approximately 150(k)-1/2 . For a given survival level, the difference in the required dose (or L) between the microbeam and broad-beam experiments is also analytically described. Transplantation, 2000 Dec 27, 70(12), 1752 - 60 Involvement of multiple subpopulations of human bone marrow cells in the regulation of allogeneic cellular immune responses; Mathew JM et al.; BACKGROUND: The identity of the cells in the human bone marrow that function as effective regulators of in vitro and possibly in vivo cellular immune responses is not well established . METHODS: Cell subpopulations were isolated from cadaver donor vertebral-body bone marrow cells (DBMC) by using immuno-magnetic microbeads and were tested as inhibitors (modulators) in cell-mediated lympholysis (CML) and mixed lymphocyte reaction (MLR) responses of normal peripheral blood lymphocytes stimulated with irradiated cadaver donor spleen cells . RESULTS: Compared with spleen cells as controls, un-irradiated T-cell depleted DBMC inhibited both the MLR and CML responses of allogeneic responder cells in a dose dependent manner (as in our previous reports) . The inhibition was also mediated by a number of purified subpopulations including pluripotent CD34+ stem cells, and their CD34 negative early progeny of both lymphoid and myeloid lineages . These included DBMC enriched for non-T-cell lymphoid precursors (NT-LP/DBMC; i.e., DBMC depleted of CD3, CD15, and glycophorin-A positive cells) and DBMC positively selected for CD38+, CD2+, CD5+, and CD1+ lymphoid cells (all were depleted of CD3+ cells) as well as CD33+ (but CD15 negative) myeloid precursors . However, positively selected CD19+ B-cells and CD15+ myeloid cells did not inhibit the MLR and CML responses . The NT-LP/DBMC that had been repeatedly stimulated with irradiated allogeneic peripheral blood lymphocytes caused the strongest inhibition of the MLR and CML responses of the same allogeneic cells with 200 times fewer modulator cells needed than uncultured DBMC (P<0.001) . Flow cytometric analysis revealed that majority of cells in these cell lines had become CD3+ TcR-alphabeta+ CD4+ and CD28+ cells . CONCLUSION: A variety of less differentiated cells of various lineages residing in the human bone marrow are immunoregulatory in vitro . Among them, there is at least one subset that can undergo differentiation in vitro into regulatory T cells that can be maintained in long-term cultures. FASEB J, 2001 Jan, 15(1), 59 - 69 Surfactant protein A (SP-A): the alveolus and beyond; Khubchandani KR et al.; Surfactant protein A (SP-A) is the major protein component of pulmonary surfactant, a material secreted by the alveolar type II cell that reduces surface tension at the alveolar air-liquid interface . The function of SP-A in the alveolus is to facilitate the surface tension-lowering properties of surfactant phospholipids, regulate surfactant phospholipid synthesis, secretion, and recycling, and counteract the inhibitory effects of plasma proteins released during lung injury on surfactant function . It has also been shown that SP-A modulates host response to microbes and particulates at the level of the alveolus . More recently, several investigators have reported that pulmonary surfactant phospholipids and SP-A are present in nonalveolar pulmonary sites as well as in other organs of the body . We describe the structure and possible functions of alveolar SP-A as well as the sites of extra-alveolar SP-A expression and the possible functions of SP-A in these sites. Ann Biomed Eng, 2000, 28(10), 1184 - 93 A chamber to permit invasive manipulation of adherent cells in laminar flow with minimal disturbance of the flow field; Levitan I et al.; An obstacle to real-time in vitro measurements of endothelial cell responses to hemodynamic forces is the inaccessibility of the cells to instruments of measurement and manipulation . We have designed a parallel plate laminar flow chamber that permits access to adherent cells during exposure to flow . The "minimally invasive flow device" (MIF device) has longitudinal slits (1 mm wide) cut in the top plate of the chamber to allow insertion of a recording, measurement, or stimulating instrument (e.g., micropipette) into the flow field . Surface tension forces at the slit openings are sufficient to counteract the hydrostatic pressure generated in the chamber and thus prevent overflow . The invasive probe is brought near to the cell surface, makes direct contact with the cell membrane, or enters the cell . The slits provide access to a large number (and choice) of cells . The MIF device can maintain physiological levels of shear stress (<1-15 dyn/cm2) without overflow in the absence and presence of fine instruments such as micropipettes used in electrophysiology, membrane aspiration, and microinjection . Microbead trajectory profiles demonstrated negligible deviations in laminar flow near the surface of target cells in the presence of microscale instruments . Patch-clamp electrophysiological recordings of flow-induced changes in membrane potential were demonstrated . The MIF device offers numerous possibilities to investigate real-time endothelial responses to well-defined flow conditions in vitro including electrophysiology, cell surface mechanical probing, local controlled chemical release, biosensing, microinjection, and amperometric techniques. J Indian Med Assoc, 2000 Jul, 98(7), 377 - 80 Evolution of drug resistance and interpretation of drug sensitivity test; Pal D; Though with the advent of more and more numbers of antibiotics, infection control has become easier but it has been established more resistant strains of microbes are appearing . These resistant strains are becoming major public health problem . Drug misuse is one of the factors for resistant strains . While discussing mechanisms of drug resistance, gene transfer plays an important part . Drug resistance can be prevented by taking several strategies . There are several methods for performing antibiotic sensitivity test e.g., diffusion methods and dilution methods which are discussed elaborately. Mem Inst Oswaldo Cruz, 2000, 95 Suppl 1, 153 - 8 Fungal infections in the immunocompromised host; Wanke B et al.; In recent years many remarkable changes occurred in our way of life, producing opportunities for microbes . All these changes are related to the recent emergence of previously unrecognized diseases, or the resurgence of diseases that, at least in developed countries, were thought to be under control . This concept is reviewed regarding fungal infections and their agents in the immunocompromised host . The changing pattern of these infections, the portals of entry of fungi into the human host, fungal pathogenicity and the main predisposing factors are analyzed . Opportunistic fungal infections in cancer, organ transplant and acquired immunodeficiency syndrome patients are reviewed, specially candidiasis and aspergillosis. Vaccine, 2000 Dec 8, 19(9-10), 1038 - 46 Respiratory syncytial virus infection of gene gun vaccinated mice induces Th2-driven pulmonary eosinophilia even in the absence of sensitisation to the fusion (F) or attachment (G) protein; Bembridge GP et al.; Complete protection against respiratory syncytial virus (RSV) infection was induced in mice vaccinated on two occasions with 2.5 microg of DNA, encoding the fusion (F) protein of RSV, precipitated onto gold microbeads . In contrast, immunisation with DNA encoding the attachment (G) protein of RSV resulted in a significant reduction in viral load following infection, but did not afford complete protection . Gene gun delivery of DNA-F elicited a T helper-2 (Th2) biased immune response that could not be modulated by the co-delivery of plasmids encoding IL-2, IL-12 or IFNgamma . Similarly gene gun delivery of DNA-G primed a Th2 response . Thus, all gene gun vaccinated mice produced a predominant Th2 biased pulmonary immune response characterised by the production of IL-4 and IL-5 with little IFNgamma following RSV challenge . Analysis of bronchoalveolar lavage (BAL) cells, 5 days post challenge, indicated that there was only a two-fold increase in the number of inflammatory cells in vaccinated compared with control animals . Despite the strong Th2 cytokine bias of lung lymphocytes and the predominant recruitment of CD4(+) T cells, following challenge, there was not a marked pulmonary eosinophilic response (range from 2 to 7% of BAL) . In contrast, the BAL from mice vaccinated with control plasmid contained significantly more eosinophils than any other group. Indian J Pediatr, 1999, 66(1 Suppl), S124 - 34 Diarrhoea and malnutrition interaction; Patwari AK; Epidemiological studies have demonstrated a marked negative relationship between diarrhoea and physical growth and development of a child . Each day of illness due to diarrhoea produces a weight deficit of 20-40 gms . Poor nutrition is associated with more serious prolonged diarrhoea . 'Catch-up growth' often does not occur in malnourished children . Malnutrition, particularly wasting, is a strong predictor of diarrhoeal duration and the prolonged illness could exacerbate nutritional faltering, thereby increasing the subsequent risk of death . Poor appetite, vomiting, deliberate withholding of food resulting in poor intake; malabsorption of macro and micronutrients; hastening of intestinal transit time; disturbance of metabolic and endocrine functions; and direct loss of protein and other nutrients in gastrointestinal tract are some of the known mechanisms which have an impact on the nutrition during an episode of diarrhea . In addition diarrhoea of infectious origin causes cytokine induced malnutrition which results from the actions of proinflammatory cytokines like tumour necrosis factor and interleukin 1, 6 and 8 . Preexisting malnutrition is associated with decreased turnover of epithelial cells resulting in delayed recovery which may prolong an episode of infectious diarrhoea by itself as well as by promoting tissue invasion by other enteropathogens . Malnutrition may also alter protective host factors and thereby favour intestinal colonization by the pathogenic microbes . Mucosal damage varying from moderately severe changes to flat lesions indistinguishable from those of celiac disease may occur in kwashiorkar . Diarrhoea malnutrition interaction represents a dangerous web which can be distangled by prevention of disease transmission by promoting exclusive breast feeding, hygienic weaning practices, safe drinking water and handwashing, improved host defences by breast feeding, improved nutrition, measles vaccine and other vaccines against enteropathogens in the offing; and promotion of standard case management with special emphasis on nutritional support and rehabilitation. J Biol Chem, 2001 Apr 6, 276(14), 11100 - 12 Epub 2000 Dec 15. Cuticular pro-phenoloxidase of the silkworm, Bombyx mori . Purification and demonstration of its transport from hemolymph; Asano T et al.; Pro-phenoloxidase (proPO) in insects is implicated in the defense against microbes and wounding . The presence of proPO in the cuticle was suggested more than 30 years ago, but it has not been purified . The extract of cuticles of the silkworm, Bombyx mori, was shown to contain two proPO isoforms (F-type and S-type proPOs, which have slightly different mobilities in polyacrylamide gel electrophoresis under nondenaturing conditions) . The two isoforms were purified to homogeneity . From hemolymph of the same insect, two types of proPO with the same electrophoretic mobilities as those of cuticular isoforms were separated and were shown to be different at five amino acid residues in one of their subunits . The isoforms in the hemolymph and cuticle were activated by a specific activating enzyme . The resulting active phenoloxidases exhibited almost the same substrate specificities and specific activities toward o-diphenols . The substrate specificities and the susceptibilities to inhibitors, including carbon monoxide, indicated that the purified proPO isoforms were not zymogens of laccase-type phenoloxidase . The proPO in hemolymph was shown to be transported to the cuticle . This demonstration was corroborated by the failure to detect proPO transcripts by Northern analysis of total RNA from epidermal cells . In reversed-phase column chromatography, cuticular and hemolymph proPOs gave distinct elution profiles, indicating that some yet to be identified modification occurs in hemolymph proPO and results in the formation of cuticular proPO . There was little transportation of cuticular proPO to the cuticle when it was injected into the hemocoel . The nature of the modification is described in the accompanying paper (Asano, T., and Ashida, M . (2001) J . Biol . Chem . 276, 11113-11125). AANA J, 2000 Jun, 68(3), 233 - 6 Laryngoscope handles: a potential for infection; Simmons SA; Laryngoscope handles do not usually come in direct contact with the patient's mucous membranes . Consequently, routine disinfection of laryngoscope handles is not currently standard practice unless gross contamination is clearly evident . Recent reports indicate that apparently clean handles may be contaminated with blood or body fluids . No report examined microbes on handles . The present article describes the incidence and types of microbes on laryngoscope handles after their use in the operating rooms of a 502-bed medical center in northwestern Pennsylvania . Twenty laryngoscope handles were cultured on Mueller Hinton 5% sheep blood agar plates . The plates were incubated at 37 degrees C for 48 hours and examined for growth . The identification, incidence, and susceptibility patterns of organisms were determined . Microorganisms were present on all 20 laryngoscope handles . Nine different types were isolated; some strains were resistant to multiple antibiotics . Organisms were categorized as contaminants or opportunistic pathogens . The presence of opportunistic pathogens places anesthesia providers and patients at risk of nosocomial infections . Based on the recommendations of the 1997 American Association of Nurse Anesthetists' Infection Control Guide and the results of the present study, institutional guidelines should be established for the use of disposable laryngoscope covers, high-level (destroying all microorganisms with the exception of high numbers of bacterial spores) disinfection, or sterilization of laryngoscope equipment between each patient use. Cancer J, 2000 Nov-Dec, 6(6), 343 - 50 Radiation, the two-edged sword: cancer risks at high and low doses; Hall EJ; Diagnostic Radiology and Radiation Oncology represent beneficial uses of radiation . The possible price tag is that radiation can cause, as well as diagnose or cure, cancer . Cancer risks at high doses are well known from epidemiologic studies of the Japanese survivors . Risks at low doses must be extrapolated from the high-dose data . The standard-setting bodies recommend a linear no-threshold extrapolation, but this is controversial . A knowledge of mechanisms, while not replacing epidemiology as a source of radiation-induced cancer risks, may provide insights into the shape of the dose-response relationships, and therefore on the validity of the linear extrapolation . The spectrum of second malignancies in radiotherapy patients sheds some light on mechanisms . More useful are new experiments involving the single-particle microbeam; these have demonstrated a bystander effect, that is, biologic effects in cells that are not directly irradiated, as well as mutations in cells where the cytoplasm, but not the nucleus, has been traversed by an alpha-particle. Pediatr Clin North Am, 2000 Dec, 47(6), 1197 - 209 Immune function; Fleisher TA et al.; The innate and the adaptive immune systems have evolved to provide a rapid and specific means for protecting hosts against the many microbes experienced over a lifetime . These two immune responses interact cooperatively to enhance the host defense . Defects in either of these two pathways can have devastating consequences, as evidenced {figure: see text} by primary immune deficiencies, many of which are discussed in this issue of the Pediatric Clinics of North America . The immune system has a central role in the pathogenesis of many disorders that involve an inflammatory response, including allergic and autoimmune diseases . New and more effective therapies for these many disorders will develop as the understanding of the immune system and its many secreted mediators continues to increase. Semin Thromb Hemost, 2000, 26(5), 589 - 94 The hepatic microvascular responses to sepsis; Ring A et al.; The liver is believed to play a major role in the initiation of multiorgan failure, the most lethal complication in the clinical course of sepsis . Microbes and their virulence factors enter the hepatic circulation where they first activate sinusoidal endothelial cells and Kupffer cells to produce proinflammatory mediators, including TNF-alpha, IL-1, IL-6, reactive oxygen metabolites, and eicosanoids . These mediators cause not only microbial killing, but also structural and functional liver damage concerning mainly the parenchymal cells . Leukocytes are targeted to the liver sinusoids by chemoattractants and, like platelets, tether to the sinusoidal endothelial cells, which are in a procoagulant state of inflammatory activation . Clogging of the sinusoids by these cells leads to a decrease of blood flow through the sinusoids, which is further aggravated by endothelin-1 effectuating the constriction of hepatic stellate cells in the sinusoids . In contrast, both nitric oxide (NO) and carbon monoxide (CO) act as antagonists of endothelin-1 by mediating relaxation of sinusoidal vessels . By maintaining an adequate sinusoidal perfusion, both NO and CO are hepatoprotective during the early, hyperdynamic phase of sepsis characterized by an increased cardiac output and moderate peripheral vasodilation . However, during the late, hypodynamic phase of sepsis, massive overproduction of NO by the inducible NO synthase leads to circulatory collapse, which inevitably includes breakdown of the liver circulation. Haematologia (Budap), 2000, 30(3), 149 - 57 A solid phase and microtiter plate hemagglutination method for pretransfusion compatibility testing; Sandler SG et al.; Most hospital blood transfusion services perform routine pretransfusion compatibility tests (ABO/D typings, antibody detection tests and crossmatches) using standard test tube methods . Recently, alternative technologies, including microtiter plate methods, solid phase red cell adherence (SPRCA) assays, gel tests, microbead columns and affinity column assays have become available . While the increased sensitivity of these new serological technologies is an important advantage, cost savings and automated testing are also important benefits . Our hospital's Transfusion Service converted from manual test tube methods for compatibility testing to manual microtiter plate and SPRCA methods and, subsequently, to automated microtiter plate and SPRCA methods . The conversion was facilitated by using commercially-marketed reagent kits and a fully-automated blood typing analyzer . The automated blood typing system was linked electronically to a hand-held combination bar code reader/portable data terminal that enabled positive identification of patients' bar code wrist bands, personal identification badges, and bar code labels on patients' blood samples and blood components . This bar code identification system has been implemented in the hospital's outpatient Infusion Service . Thus, the conversion to microtiter plate and SPRCA assays enhanced transfusion safety not only by increasing the sensitivity of serological testing, but also by standardizing compatibility testing, supporting electronic record keeping, and linking the laboratory analyzer to a bar code identification system. Am J Physiol Gastrointest Liver Physiol, 2001 Jan, 280(1), G1 - 6 Microbes and microbial toxins: paradigms for microbial-mucosal interactions I . Pathophysiological aspects of enteric infections with the lumen-dwelling protozoan pathogen Giardia lamblia; Eckmann L et al.; Giardia lamblia is one of the most important causes of waterborne diarrheal disease worldwide, and giardiasis is the most common protozoan infection of the human small intestine . Symptomatic infection is characterized by diarrhea, abdominal pain, and malabsorption, leading to malnutrition and weight loss, particularly in children . The pathogen resides strictly in the lumen of the small intestine, and infection is typically not accompanied by significant mucosal inflammation . Clinical and experimental studies indicate that B cell-dependent host defenses, particularly IgA, are important for controlling and clearing Giardia infection, although B cell-independent mechanisms also contribute to this outcome . In contrast to antigiardial host defenses, much less is known about the pathophysiological mechanisms underlying the clinical symptoms of giardiasis, partly because of the current lack of suitable model systems . In addition to being an important human enteric pathogen, Giardia is an interesting model organism for gaining basic insights into genetic innovations that led to evolution of eukaryotic cells, since it belongs to the earliest diverging eukaryotic lineage known . The completion of the giardial genome project will increase understanding of the basic biology of the protozoan and will help us to better understand host pathogen-interactions as a basis for developing new vaccination and therapeutic strategies. Insect Mol Biol, 2000 Dec, 9(6), 661 - 73 Wolbachia infections in native and introduced populations of fire ants (Solenopsis spp.); Shoemaker DD et al.; Wolbachia are cytoplasmically inherited bacteria that induce a variety of effects with fitness consequences on host arthropods, including cytoplasmic incompatibility, parthenogenesis, male-killing and feminization . We report here the presence of Wolbachia in native South American populations of the fire ant Solenopsis invicta, but the apparent absence of the bacteria in introduced populations of this pest species in the USA . The Wolbachia strains in native S . invicta are of two divergent types (A and B), and the frequency of infection varies dramatically between geographical regions and social forms of this host . Survey data reveal that Wolbachia also are found in other native fire ant species within the Solenopsis saevissima species complex from South America, including S . richteri . This latter species also has been introduced in the USA, where it lacks Wolbachia . Sequence data reveal complete phylogenetic concordance between mtDNA haplotype in S . invicta and Wolbachia infection type (A or B) . In addition, the mtDNA and associated group A Wolbachia strain in S . invicta are more closely related to the mtDNA and Wolbachia strain found in S . richteri than they are to the mtDNA and associated group B Wolbachia in S . invicta . These data are consistent with historical introgression of S . richteri cytoplasmic elements into S . invicta populations, resulting in enhanced infection and mtDNA polymorphisms in S . invicta . Wolbachia may have significant fitness effects on these hosts (either directly or by cytoplasmic incompatibility) and therefore these microbes potentially could be used in biological control programmes to suppress introduced fire ant populations. Radiat Res, 2001 Jan, 155(1 Pt 1), 122 - 6 Long-term genomic instability in human lymphocytes induced by single-particle irradiation; Kadhim MA et al.; Recent evidence suggests that genomic instability, which is an important step in carcinogenesis, may be important in the effectiveness of radiation as a carcinogen, particularly for high-LET radiations . Understanding the biological effects underpinning the risks associated with low doses of densely ionizing radiations is complicated in experimental systems by the Poisson distribution of particles that can be delivered . In this study, we report an approach to determine the effect of the lowest possible cellular radiation dose of densely ionizing alpha particles, that of a single particle traversal . Using microbeam technology and an approach for immobilizing human T-lymphocytes, we have measured the effects of single alpha-particle traversals on the surviving progeny of cells . A significant increase in the proportion of aberrant cells is observed 12-13 population doublings after exposure, with a high level of chromatid-type aberrations, indicative of an instability phenotype . These data suggest that instability may be important in situations where even a single particle traverses human cells. Radiat Res, 2001 Jan, 155(1 Pt 1), 89 - 94 Microdosimetry of a 25 keV electron microbeam; Wilson WE et al.; Electron microbeam experiments are planned or under way to explore in part the question regarding whether the bystander effect is a general phenomenon or is restricted to high-LET radiation . Since low-LET radiations scatter more readily compared to high-LET radiations, identifying bystander cells and assessing the potential dose that they may receive will be crucial to the interpretation of radiobiological results . This paper reports on initial calculations of the basic information needed for a stochastic model of the penetration of energetic electrons in tissue-like matter; the model will be used to predict doses delivered to adjacent regions in which bystander cells may reside . Results are presented of calculations of the stochastics of energy deposition by 25 keV electrons slowing down in a homogeneous water medium . Energy deposition distributions were scored for 1-micrometer spheres located at various penetration and radial distances up to 10 micrometer from the point of origin . The energy of 25 keV was selected because experiments are planned for that energy . At 25 keV there is a high probability that the entire electron track will be contained within a typical mammalian cell . Individual tracks are scored because of their primacy; data for higher doses can be obtained by convoluting single-track distributions . The event frequency decreases approximately exponentially after the first micrometer to 1% at about 8 micrometer of penetration . Radially, the 1% contour extends to 3.5 micrometer at a penetration of 5.5 micrometer . The frequency-mean energy deposited decreases from 1.5 to 1 keV/micrometer at a penetration of 3.5 micrometer, then increases back to about 1.5 at a penetration of 6.5 micrometer . The mean energy increases to about 3 keV/micrometer at a radial distance of 8.5 micrometer. Infect Immun, 2001 Jan, 69(1), 494 - 500 Microtubule- and dynein-mediated movement of Orientia tsutsugamushi to the microtubule organizing center; Kim SW et al.; The host cell microfilaments and microtubules (MTs) are known to play a critical role in the life cycles of several pathogenic intracellular microbes by providing for successful invasion and promoting movement of the pathogen once inside the host cell cytoplasm . Orientia tsutsugamushi, an obligate intracellular bacterium, enters host cells by induced phagocytosis, escapes to the cytosol, and then replicates in the cytosol . ECV304 cells infected with O . tsutsugamushi revealed the colocalization of the MT organizing center (MTOC) and cytosolic orientiae by indirect immunofluorescence assay . Using immunofluorescence microscopy in the presence and absence of MT-depolymerizing agents (colchicine and nocodazole), it was shown that the cytosolic oriential movement was mediated by MTs . By transfection study (overexpression of dynamitin {also called p50}, which is known to associate with dynein-dependent movement), the movement of O . tsutsugamushi to the MTOC was also mediated by dynein, the minus-end-directed MT-related motor . Although the significance of this movement in the life cycle of O . tsutsugamushi was not proven, we propose that the cytosolic O . tsutsugamushi bacteria use MTs and dyneins to propel themselves from the cell periphery to the MTOC. Arch Environ Contam Toxicol, 2001 Jan, 40(1), 10 - 7 The effect of mercury and PCBs on organisms from lower trophic levels of a Georgia salt marsh; Wall VD et al.; We examined several indicators of salt marsh function, focusing on primary producers, microbes, and grass shrimp, at a Superfund site (LCP) contaminated with mercury and polychlorinated biphenyls (PCBs) and a reference site (Cross-River) in Georgia . Primary production of Spartina alterniflora was assessed by measuring peroxidase activity (POD), glutathione concentration (tGSH), photosynthesis (A(net)), and transpiration (E) . Microbial populations were assessed by measuring living-fungal standing crop (as ergosterol) and Microtox(R) . Grass shrimp (Palaemonetes pugio) reproductive potential was determined by measuring individual egg mass, average egg area, brood size, and brood mass of gravid females . Comparison of the sites suggested that P . pugio reproduction was affected at the LCP site, but we were unable to document clear negative effects on other organisms we investigated . Due to natural environmental gradients, the Cross-River site may not have been a perfect control for the LCP site . Therefore, data from just the LCP site were reanalyzed using multiple regression . Fungal biomass was related to methylmercury concentrations, but the direction of the relationship differed between wholly dead shoots (positive) and partially dead shoots (negative) . S . alterniflora POD was positively related to methylmercury concentrations . S . alterniflora A(net) and E were negatively related to elevation and salinity, respectively . Despite high levels of contamination at the LCP site, our results provided only suggestive evidence for impacts on organisms at lower trophic levels. J Bacteriol, 2001 Jan, 183(1), 131 - 8 Recombinational repair is critical for survival of Escherichia coli exposed to nitric oxide; Spek EJ et al.; Nitric oxide (NO(.)) is critical to numerous biological processes, including signal transduction and macrophage-mediated immunity . In this study, we have explored the biological effects of NO(.)-induced DNA damage on Escherichia coli . The relative importance of base excision repair, nucleotide excision repair (NER), and recombinational repair in preventing NO(.)-induced toxicity was determined . E . coli strains lacking either NER or DNA glycosylases (including those that repair alkylation damage {alkA tag strain}, oxidative damage {fpg nei nth strain}, and deaminated cytosine {ung strain}) showed essentially wild-type levels of NO(.) resistance . However, apyrimidinic/apurinic (AP) endonuclease-deficient cells (xth nfo strain) were very sensitive to killing by NO(.), which indicates that normal processing of abasic sites is critical for defense against NO(.) . In addition, recA mutant cells were exquisitely sensitive to NO(.)-induced killing . Both SOS-deficient (lexA3) and Holliday junction resolvase-deficient (ruvC) cells were very sensitive to NO(.), indicating that both SOS and recombinational repair play important roles in defense against NO(.) . Furthermore, strains specifically lacking double-strand end repair (recBCD strains) were very sensitive to NO(.), which suggests that NO(.) exposure leads to the formation of double-strand ends . One consequence of these double-strand ends is that NO(.) induces homologous recombination at a genetically engineered substrate . Taken together, it is now clear that, in addition to the known point mutagenic effects of NO(.), it is also important to consider recombination events among the spectrum of genetic changes that NO( . ) can induce . Furthermore, the importance of recombinational repair for cellular survival of NO(.) exposure reveals a potential susceptibility factor for invading microbes. Microbes Infect, 2000 Nov, 2(13), 1609 - 18 Leukocyte-facilitated entry of intracellular pathogens into the central nervous system; Drevets DA et al.; Microbes use numerous strategies to invade the central nervous system . Leukocyte-facilitated entry is one such mechanism whereby intracellular pathogens establish infection by taking advantage of leukocyte trafficking to the central nervous system . Key components of this process include peripheral infection and activation of leukocytes, activation of cerebral endothelial cells with or without concomitant infection, and trafficking of infected leukocytes to and through the blood-brain or blood-cerebrospinal fluid barrier. Kekkaku, 2000 Oct, 75(10), 599 - 602 {Prospect of chemotherapy in the 21st century}; Omura S; "Golden Era in Chemotherapy" has begun with the discovery of penicillin in the early 1940's and lasted for two decades during which many antibiotics were discovered . However, the once-believed bright prospect that every infectious disease could be eliminated on the earth by the discovery of antibiotics had to be canceled owing to the emerging of drug-resistant microbes . It was indeed a rat race . We are now at the point when we have to seek another way to combat infectious diseases: One possible way might be not to eradicate the microbes but to coexist with them so long as they do no harm to the human hosts . The first step of infection with pathogens to the host is the adherence of the microbes to the surface of host cells . Therefore, the method how to inhibit this adhesion of microbes to the host cells may provide a new tool to prevent the development of infectious diseases without elimination of microbes from the host . This is just an example of strategy by which humans and pathogens coexist at peace and should be taken into consideration for the development of new-type antibiotics or "anti-infective drugs" in the 21st century . The analysis of genome sequences has been accelerated recently for various pathogenic bacteria one by one . New targets in the pathogenic microbes for the development of new antibiotics can, therefore, be determined from the genetic point of view . The discovery of antibiotics has indeed been the history of collection of innumerable species and/or strains of bacteria from the soils to search for the biologically active anti-pathogenic agents . The current progress in the technology of molecular genetics, however, will certainly make it possible to search for active molecules by DNA technology; bacterial DNA but not whole microorganisms from the soil is to be transformed into the conventional bacteria and searched for active molecules with combat against pathogens. Kekkaku, 2000 Oct, 75(10), 595 - 8 {Immunology in the 20th century--progress made in research on infectious and immunological diseases}; Kishimoto T; The new era of the modern medicine was opened 100 years ago by Robert Koch and Louis Pasteur who demonstrated that various infectious diseases were caused by their respective microbes . Koch discovered Mycobacterium tuberculosis, the causative agent of tuberculosis . The first breakthrough in the modern medicine to combat against infectious diseases was the discovery of anti-diphtheria toxin antibody by E.A . von Behring and S . Kitasato . The concept of immunity--immune from disease--has thus been established . The immune response between antigen and antibody sometimes provides the host with a harmful effect . The concept of allergy was introduced by Richet and