Bacteria

Trends Microbiol, 2004 Jul, 12(7), 325 - 36
Strategies of the home-team: symbioses exploited for vector-borne disease control; Rio RV et al.; Symbioses between eukaryotes and unicellular organisms are quite common, with examples copiously disseminated throughout the earth's biota . Arthropods, in particular, owe much of their ecological success to their microbial flora, which often provide supplements either lacking in the limited host diet or which the hosts are unable to synthesize . In addition to harboring beneficial microbes, many arthropods (vectors) also transmit pathogens to the animals and plants upon which they prey . Vector-borne diseases exact a high public health burden and additionally have a devastating impact on livestock and agriculture . Recent scientific discoveries have resulted in the development of powerful technologies for studying the vector's biology, to discover the weak links in disease transmission . One of the more challenging applications of these developments is transgenesis, which allows for insertion of foreign DNA into the insect's genome to modify its phenotype . In this review, we discuss an approach in which the naturally occurring commensal flora of insects are manipulated to express products that render their host environment inhospitable for pathogen transmission . Replacing susceptible insect genotypes with modified counterparts with reduced pathogen transmission ability, might provide a new set of armaments in the battle for vector-borne disease reduction.

Mol Immunol, 2004 Jul, 41(6-7), 577 - 82
The expanding family of MyD88-like adaptors in Toll-like receptor signal transduction; McGettrick AF et al.; Toll-like receptors (TLRs) play an essential role in the detection and elimination of invading microbes . They are type-1 transmembrane receptors, containing extracellular leucine rich repeats and an intracellular Toll/interleukin-1 receptor (TIR) domain . Upon stimulation, these receptors interact with specific TIR domain-containing adaptor proteins . Five such adaptors have been discovered to date, MyD88, Mal (MyD88 adaptor-like)/TIRAP (TIR domain-containing adaptor protein), Trif (TIR-domain-containing adaptor inducing interferon-beta), TRAM (Trif-related adaptor molecule) and SARM (SAM and ARM-containing protein) . Different TLRs use different combinations of these adaptors, leading to the activation of common and unique pathways involved in the elimination of the invading microbe.

Cell Struct Funct, 1999 Oct, 24(5), 405 - 12
Biophysical properties of stable microtubules in neurites revealed by optical techniques; Kurachi M et al.; We tested the stability of microtubules in the neurites of cultured dorsal root ganglion cells by dissolving the cytoplasmic membrane with detergent and exposing them to defined extracellular medium under the microscope . Smooth cytoplasmic filaments visualized after membrane removal were suggested to be microtubules by the preservation of all of the filaments in the presence but not in the absence of taxol . They were further confirmed to be microtubules by immunostaining with anti-tubulin antibody . Significant number of microtubules in the established neurites remained longer than 1 hour after membrane removal . To investigate their stabilization mechanism, we transected the exposed microtubules by laser microbeam irradiation and observed their length changes with video-enhanced microscopy . Microtubule fragments started to shorten on both sides of the transection site, more rapidly from the newly generated plus ends than from the minus ends . The maximal rate as well as the pattern of shortening correlated with the time of transection; microtubules transected later than 30 min after membrane removal shortened at rates less than 20 microm/min and typically with intermittent pauses, while the more labile microtubules included in the earlier transections shortened continuously at higher rates . Microtubules in neurites were thus stabilized by (1) stopping disassembly at local sites including the plus ends, and (2) slowing disassembly along the length . Observations of the course of disassembly also suggested the presence of specialized points along microtubules which is involved in anchoring microtubules to the substratum or transiently stopping disassembly.

Ann Surg, 2004 Jul, 240(1), 132 - 41
Bone marrow norepinephrine mediates development of functionally different macrophages after thermal injury and sepsis; Cohen MJ et al.; OBJECTIVE: We sought to determine the influence of thermal (burn) injury with sepsis and norepinephrine on the clonogenic potential and functional cytokine response to lipopolysaccharide (LPS) stimulation in nonmyeloid committed (CD117) and myeloid committed (ER-MP12) bone marrow progenitor cells . SUMMARY AND BACKGROUND DATA: We have previously demonstrated that norepinephrine stimulated myelopoiesis after burn injury and sepsis, but the site of this stimulation in monocyte development is unknown . In the present study the influence of norepinephrine on the developmental hierarchy of bone marrow cells after thermal injury and sepsis was determined by assessing the clonogenic potential and LPS-stimulated cytokine responses of mature macrophages derived from CD117 and ER-MP12 bone marrow progenitor cells . METHODS: Tissue and bone marrow norepinephrine content was ablated by chemical sympathectomy with 6-hydroxydopamine treatment . CD117 and ER-MP12 bone marrow cells were isolated using antibody-linked magnetic microbeads . Clonogenic potential in response to colony-stimulating factors was determined . Both progenitor cell types were differentiated to mature macrophages in vitro and tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 cytokine responses to LPS provocation were determined . RESULTS: The macrophage- and granulocyte-macrophage colony-stimulating factor responsive clonogenic potential was increased with burn sepsis, suggesting an expansion of both progenitor populations . Such increases were greatly reduced with prior depletion of norepinephrine . TNF-alpha and IL-6 cytokine responses to LPS were markedly influenced by the specific progenitor cells involved as well as the injury conditions and the status of norepinephrine prior to injury . In burn sepsis the depletion of norepinephrine resulted in a dramatic decrease in both IL-6 and TNF-alpha production by both progenitor-derived macrophages . CONCLUSIONS: Depletion of norepinephrine attenuated burn and burn sepsis-induced bone marrow progenitor clonal growth in response to macrophage- and granulocyte-macrophage colony-stimulating factor . Functional phenotypes of bone marrow progenitor-derived macrophages are greatly influenced by norepinephrine and the milieu created by thermal injury and sepsis.

Fish Shellfish Immunol, 2004 Aug, 17(2), 121 - 8
Identification of genes involved in the response of haemocytes of Penaeus japonicus by suppression subtractive hybridization (SSH) following microbial challenge; He N et al.; Penaeus japonicus were injected with a heat-killed microorganism suspension and 291 randomly selected cDNA fragments generated by suppression subtractive hybridization (SSH) were sequenced . A total of 71 cDNA clones corresponding to 25 genes were found to have enhanced expression, of which eight are found for the first time in shrimp . The most abundant gene in the subtractive library was Kunitz-type protease inhibitor, clearly indicating this protease inhibitor in the response . A number of genes encoding signaling molecules, such as Ras-related nuclear protein (Ran), growth factor receptor bound protein (Grb), TGF-beta receptor interacting protein, integrin binding protein and interferon receptor bound protein were found for the first time in the shrimp, and they may be involved in the regulation of the host defense against the injected microbes . Furthermore, cDNAs of chaperonin, proteasome, antioxidant as well as genes associated with actin reorganization, which may be necessary for phagocytosis and encapsulation, were also expressed at a higher level after the challenge . These results may facilitate the understanding of shrimp immune responses.

J Oral Rehabil, 2004 Jul, 31(7), 647 - 52
A 5-year follow-up study on the prosthetic rehabilitation of the elderly in Helsinki, Finland; Nevalainen MJ et al.; In 1990, 364 elderly (76-86 years) inhabitants of Helsinki, Finland, attended a dental and oral examination study that was conducted as part of the Helsinki Aging Study . In spring 1996, these subjects were recalled for a 5-year follow-up . Between the baseline and follow-up examinations, 114 (31%) subjects had deceased (86 women and 28 men), whereas 134 had either moved, were too ill, or refused to participate in the follow-up . Follow-up examination was conducted for 113 subjects (79 women and 34 men), with the participating rate being 46% . Five subjects became edentulous during the follow-up . Of the subjects, 61% had 1-32 teeth at follow-up . In these subjects, the mean number of teeth decreased from 14.9 (+/-8.3) to 13.5 (+/-8.6) (P < 0.0001) . Prosthetic status changed in 40% of the elderly dentate people: 25% received new prostheses whereas 15% lost prostheses that were not replaced . New fixed partial dentures were made in five maxillae and in nine mandibles during the follow-up . Acrylic removable partial dentures (ARPD) were most frequently used: 35% of dentate subjects had an ARPD . Subjects with removable prostheses had higher levels of salivary microbes and higher root caries incidence than those with natural teeth . Furthermore, the presence of removable prostheses at baseline, together with the male gender, was clearly associated with tooth loss during follow-up . This study indicates that fixed rather than removable prostheses should be used in elderly patients . The need for a removable denture ought to be carefully considered.

Notes Rec R Soc Lond, 2004 May, 58(2), 187 - 201
The discovery of microorganisms by Robert Hooke and Antoni Van Leeuwenhoek, fellows of the Royal Society; Gest H; The existence of microscopic organisms was discovered during the period 1665-83 by two Fellows of The Royal Society, Robert Hooke and Antoni van Leeuwenhoek . In Micrographia (1665), Hooke presented the first published depiction of a microganism, the microfungus Mucor . Later, Leeuwenhoek observed and described microscopic protozoa and bacteria . These important revelations were made possible by the ingenuity of Hooke and Leeuwenhoek in fabricating and using simple microscopes that magnified objects from about 25-fold to 250-fold . After a lapse of more than 150 years, microscopy became the backbone of our understanding of the roles of microbes in the causation of infectious diseases and the recycling of chemical elements in the biosphere.

Res Microbiol, 2004 Jun, 155(5), 352 - 9
Adaptive mutation and amplification in Escherichia coli: two pathways of genome adaptation under stress; Hersh MN et al.; The neo-Darwinists suggested that evolution is constant and gradual, and thus that genetic changes that drive evolution should be too . However, more recent understanding of phenomena called adaptive mutation in microbes indicates that mutation rates can be elevated in response to stress, producing beneficial and other mutations . We review evidence that, in Escherichia coli, two separate mechanisms of stress-induced genetic change occur that revert a lac frameshift allele allowing growth on lactose medium . First, compensatory frameshift ("point") mutations occur by a mechanism that includes DNA double-strand breaks and (we have suggested) their error-prone repair . Point mutation requires induction of the RpoS-dependent general stress response, and the SOS DNA damage response leading to upregulation of the error-prone DNA polymerase DinB (Pol IV), and occurs during a transient limitation of post-replicative mismatch repair activity . A second mechanism, adaptive amplification, entails amplification of the leaky lac allele to 20-50 tandem repeats . These provide sufficient beta-galactosidase activity for growth, thereby apparently deflecting cells from the point mutation pathway . Unlike point mutation, amplification neither occurs in hypermutating cells nor requires SOS or DinB, but like point mutation, amplification requires the RpoS-dependent stress response . Similar processes are being found in other bacterial systems and yeast . Stress-induced genetic changes may underlie much of microbial evolution, pathogenesis and antibiotic resistance, and also cancer formation, progression and drug resistance.

Science, 2004 Jul 23, 305(5683), 509 - 13 Epub 2004 Jun 17.
Ecosystem properties and forest decline in contrasting long-term chronosequences; Wardle DA et al.; During succession, ecosystem development occurs; but in the long-term absence of catastrophic disturbance, a decline phase eventually follows . We studied six long-term chronosequences, in Australia, Sweden, Alaska, Hawaii, and New Zealand; for each, the decline phase was associated with a reduction in tree basal area and an increase in the substrate nitrogen-to-phosphorus ratio, indicating increasing phosphorus limitation over time . These changes were often associated with reductions in litter decomposition rates, phosphorus release from litter, and biomass and activity of decomposer microbes . Our findings suggest that the maximal biomass phase reached during succession cannot be maintained in the long-term absence of major disturbance, and that similar patterns of decline occur in forested ecosystems spanning the tropical, temperate, and boreal zones.

Inhal Toxicol, 2004 Jul, 16(8), 509 - 15
The alteration of immune reactions in inbred BALB/c mice following low-level sarin inhalation exposure; Kassa J et al.; To study the influence of low-level sarin inhalation exposure on immune functions, inbred BALB/c mice were exposed to low concentrations of sarin for 60 min in the inhalation chamber . The evaluation of immune functions was carried out using phenotyping of CD3 (T lymphocytes), CD4 (helper T lymphocytes), CD8 (cytotoxic T lymphocytes), and CD19 cells (B lymphocytes) in the lungs, blood, and spleen, lymphoproliferation of spleen cells stimulated in vitro by various mitogens (concanavalin A, lipopolysaccharides), phagocyte activity of peritoneal and alveolar macrophages, production of N-oxides by peritoneal macrophages, and the measurement of the natural killer cell activity at 1 wk following sarin exposure . The results were compared to the values obtained from control mice exposed to pure air instead of sarin . The results indicate that low doses of sarin are able to alter the reaction of immune system at one week following exposure to sarin . While the numbers of CD3 cells in the lungs, blood, and spleen were slightly decreased, an increase in CD19 cells was observed, especially in the lungs and blood . The reduced proportion of T lymphocytes is caused by decay of CD4-positive T cells . Lymphoproliferation was significantly decreased regardless of the mitogen and sarin concentration used . The production of N-oxides by peritoneal macrophages was stimulated after exposure to the highest dose of sarin, whereas their ability to phagocytize the microbes was increased after exposure to the lowest dose of sarin . The natural killer cell activity was significantly higher in the case of inhalation exposure of mice to the highest level of sarin . Thus, not only organophosphorus insecticides but also nerve agents such as sarin are able to alter immune functions even at a dose that does not cause clinically manifested disruption of cholinergic nervous system in the case of inhalation exposure . Nevertheless, the alteration of immune functions following the inhalation exposure to a symptomatic concentration of sarin seems to be more pronounced.

Immunol Rev, 2004 Apr, 198, 106 - 15
A Toll-like receptor in horseshoe crabs; Inamori K et al.; Non-self-recognition of invading microbes relies on the pattern-recognition of pathogen-associated molecular patterns (PAMPs) derived from microbial cell-wall components . Insects and mammals conserve a signaling pathway of the innate immune system through cell-surface receptors called Tolls and Toll-like receptors (TLRs) . Bacterial lipopolysaccharides (LPSs) are an important trigger of the horseshoe crab's innate immunity to infectious microorganisms . Horseshoe crabs' granular hemocytes respond specifically to LPS stimulation, inducing the secretion of various defense molecules from the granular hemocytes . Here, we show a cDNA which we named tToll, coding for a TLR identified from hemocytes of the horseshoe crab Tachypleus tridentatus . tToll is most closely related to Drosophila Toll in both domain architecture and overall length . Human TLRs have been suggested to contain numerous PAMP-binding insertions located in the leucine-rich repeats (LRRs) of their ectodomains . However, the LRRs of tToll contained no obvious PAMP-binding insertions . Furthermore, tToll was non-specifically expressed in horseshoe crab tissues . These observations suggest that tToll does not function as an LPS receptor on granular hemocytes.

Cancer Metastasis Rev, 2004 Aug-Dec, 23(3-4), 323 - 31
Microbeam developments and applications: a low linear energy transfer perspective; Sowa Resat M et al.; In the last few years there has been a significant increase in the number of centers having, or developing, both high- and low-linear energy transfer (LET) microbeams for radiobiological investigations . In this review, we will discuss the various approaches for developing low-LET single cell irradiation devices and consider the instrument designs and how the machines capabilities can be exploited to investigate targeted and non-targeted effects of low-LET irradiation.

Curr Opin Microbiol, 2004 Jun, 7(3), 283 - 9
Virulence gene expression in vivo; Shelburne SA et al.; The ability to identify and isolate bacterial RNA from animals or humans with infections has markedly advanced the capacity to examine microbial gene expression in vivo . This advance has been coupled with the development of quantitative real-time reverse transcription polymerase chain reaction and expression microarrays to allow investigators to accurately measure how organisms are manipulating their genetic expression during actual infections . Though the full ramifications of these technologies have yet to be realized, they promise to open new avenues of therapeutics for a broad range of infectious diseases by allowing researchers to focus on in vivo expressed genes . These developments provide a framework for efficient utilization of the vast amount of information being generated by the accelerating pace of genomic sequencing of microbes.

Curr Opin Microbiol, 2004 Jun, 7(3), 245 - 54
Oligonucleotide microarrays in microbial diagnostics; Bodrossy L et al.; Oligonucleotide microarrays offer a fast, high-throughput alternative for the parallel detection of microbes from virtually any sample . The application potential spreads across most sectors of life sciences, including environmental microbiology and microbial ecology; human, veterinary, food and plant diagnostics; water quality control; industrial microbiology, and so on . The past two years have witnessed a rapid increase of research in this field . Many alternative techniques were developed and validated as seen in 'proof-of-concept' articles . Publications reporting on the application of oligonucleotide microarray technology for microbial diagnostics in microbiology driven projects have just started to appear . Current and future technical and bioinformatics developments will inevitably improve the potential of this technology further.

Curr Opin Microbiol, 2004 Jun, 7(3), 239 - 44
The tritrophic trinity: a source of pollutant-degrading enzymes and its implications for phytoremediation; Singer AC et al.; Barring bioavailability and nutritional limitations, virtually all organic anthropogenic chemicals can be naturally biodegraded . It is to this phenomenon we owe thanks to the long established 'tritrophic trinity' of microbe-plant-insect interactions . Over hundreds of millennia these organisms have coevolved, producing hundreds of thousands of different chemicals that are used to attract, defend, antagonize, monitor and misdirect one another . In comparison, the numbers of truly novel chemicals of anthropogenic origin are negligible . It is only now that we are beginning to appreciate the fortuitous evolution of xenobiotic-degrading enzymes from these interactions . We argue that success in phytoremediation can be hastened through understanding the structure, sources, uses and targets of these secondary metabolites . Owing to recent developments in molecular biology, particularly stable isotope probing, we eagerly anticipate highly significant insights into trophic interactions, particularly in the rhizosphere, providing phytoremediation with a solid mechanistic understanding.

Immunology, 2004 Jul, 112(3), 352 - 63
The increased prevalence of allergy and the hygiene hypothesis: missing immune deviation, reduced immune suppression, or both?
Romagnani S.
Allergic atopic disorders, such as rhinitis, asthma, and atopic dermatitis, are the result of a systemic inflammatory reaction triggered by type 2 T helper (Th2) cell-mediated immune responses against 'innocuous' antigens (allergens) of complex genetic and environmental origin . A number of epidemiological studies have suggested that the increase in the prevalence of allergic disorders that has occurred over the past few decades is attributable to a reduced microbial burden during childhood, as a consequence of Westernized lifestyle (the 'hygiene hypothesis') . However, the mechanisms by which the reduced exposure of children to pathogenic and nonpathogenic microbes results in enhanced responses of Th2 cells are still controversial . The initial interpretation proposed a missing immune deviation of allergen-specific responses from a Th2 to a type 1 Th (Th1) profile, as a result of the reduced production of interleukin-12 and interferons by natural immunity cells which are stimulated by bacterial products via their Toll-like receptors . More recently, the role of reduced activity of T regulatory cells has been emphasized . The epidemiological findings and the experimental evidence available so far suggest that both mechanisms may be involved . A better understanding of this question is important not only from a theoretical point of view, but also because of its therapeutic implications.

Transplant Proc, 2004 May, 36(4), 1199 - 200
Short-term immunosuppression reduces fibrotic cellular infiltration around barium-M-alginate microbeads injected intraportally; Mathe Z et al.; INTRODUCTION: One of the major barriers affecting the viability of encapsulated islets is pericapsular fibrotic infiltration (PFI) . This study aimed to design strategies to reduce PFI around intraportally injected alginate microbeads . METHODS: Empty, highly purified, barium-M-alginate microbeads (400 microm) were injected intraportally into Lewis rats (3000 beads/rat) . Rats (n = 9/group) were treated daily with either rapamycine (RAPA; 1 mg/kg/d p.o.), tacrolimus (TAC; 2 mg/kg/d p.o.), a combination of both, or gadolinium-chloride (GdC13, 20 mg/kg/d i.v., at day -1 and day +4) . Treatment was discontinued at 10 days . Three rats/group were sacrificed at 3, 7, and 42 days after transplantation . Cellular composition of PFI was evaluated by immunohistochemistry . Severity of the reaction to the beads was determined by measuring the thickness of PFI on histology . RESULTS: The main cellular components of PFI in the liver were macrophages and myofibroblasts . There was a significant (P <.05) reduction in the thickness of PFI in all treated groups, even 6 weeks after transplantation . Encapsulated rat islets showed excellent insulin response to glucose in vitro, with a stimulation index of 3.6 +/- 2.0 . CONCLUSION: Combination of highly purified alginate with short-term immunosuppression reduces fibrotic overgrowth around microbeads injected intraportally.

Biochimie, 2004 Apr-May, 86(4-5), 311 - 5
Dihydrodipicolinate synthase (DHDPS) from Escherichia coli displays partial mixed inhibition with respect to its first substrate, pyruvate; Dobson RC et al.; Dihydrodipicolinate synthase (DHDPS, E.C . 4.2.1.52) mediates the first unique reaction of (S)-lysine biosynthesis in plants and microbes-the condensation of (S)-aspartate-beta-semialdehyde ((S)-ASA) and pyruvate . It has been shown that DHDPS is partially feedback inhibited by (S)-lysine; it is suggested that this mechanism regulates flux through the DAP biosynthetic pathway . Others have characterised DHDPS from Escherichia coli with respect to (S)-lysine inhibition . They have concluded that, with respect to pyruvate, the first substrate of the reaction, DHDPS shows uncompetitive inhibition: as such, they further suggest that (S)-lysine inhibits DHDPS via interaction with the binding site for the second substrate, (S)-ASA . Yet, this finding is based on the assumption that (S)-lysine is a fully uncompetitive inhibitor . In light of crystallographic studies, which lead to the proposal that (S)-lysine affects the putative proton-relay of DHDPS, we re-evaluated the inhibition mechanism of DHDPS with respect to (S)-lysine by incorporating the observed hyperbolic inhibition . Our data showed that lysine is not an uncompetitive inhibitor, but a mixed inhibitor when pyruvate and (S)-lysine concentrations were varied . Thus, consistent with the crystallographic data, (S)-lysine must have an effect on the initial steps of the DHDPS reaction, including the binding of pyruvate and Schiff base formation.

Curr Opin Biotechnol, 2004 Jun, 15(3), 191 - 8
Genome analysis of marine photosynthetic microbes and their global role; Hess WR; Four recently completed genome projects on marine Cyanobacteria have started the age of comparative genomics for marine microbes . Cyanobacteria are a group of photoautotrophic bacteria that have traditionally been under-represented in studies of complete genome sequences, as have microbes from the marine environment in general . The new genome information is of crucial importance to understanding their role in oceanic primary production, global carbon cycling and functioning of the biosphere . Marine microbes are a still almost untapped resource for the identification of novel beneficial metabolites and activities . The availability of an increasing number of genome sequences will eventually lead to a sustained development of marine biotechnology.

Curr Opin Biotechnol, 2004 Jun, 15(3), 187 - 90
To BAC or not to BAC: marine ecogenomics; Beja O; Most microbes in the ocean are still resistant to our collective cultivation efforts . Environmental microbial genomics provides science with the means for accessing and assessing the genomes, diversity, evolution and population dynamics of uncultured microorganisms--the ocean's hidden majority.

Med Parazitol (Mosk), 2004 Apr-Jun, (2), 3 - 6
{Seasonal features of the time course of changes in the aggregated state of the causative agent of plague in the organism of Citellophilus tesquorum altaicus (Siphonaptera)}; Bazanova LP et al.; The data of experiments made in a Tuva natural plague focus with the main vector of the flea C . lesquorum altaicus during different seasons are analyzed . Ectoparasites were infected with the plague microbial strain typical for the focus and fed on natural nourishment, such as long-tailed sousliks . An analysis indicated that estimation of the parameter "the proportion of infected insects containing the causative agent at the aggregated state" is of greater informative value than is the recording of only sporadic gizzard "blocks" . In experiments on fleas of a breeding of the current year, the conglomerates of the causative agent of plague were more frequently observed in males . All things being equal, the greatest efficiency of transmission of the microbe to the carriers of infection and the highest block formation in the fleas were observed in summer . The proportion of specimens with microbial conglomerates drastically increased in autumn during the preparation of the body of imagoes for hibernation . At this state, the microbe survived in the organism of a dormant carrier during winter . In spring, in the imagoes infected during the previous season, bacterial conglomerates were observed during the first feeds ten times more frequently than those in the insects of a breeding and infection of the current year.

J Toxicol Environ Health A, 2004 Apr 23-May 28, 67(8-10), 819 - 34
Induced plant signaling and its implications for environmental sensing; De Moraes CM et al.; The sophisticated chemical responses of plants to environmental stimuli have profound implications for the development of remote sensing systems . Recent advances in the investigation of these responses have demonstrated that plants can be developed as reliable reporters of numerous environmental stimuli including soil and water conditions, light levels, temperature, mechanical damage, insect feeding, exposure to pathogens--including those that affect animals and humans--and exposure to airborne chemicals . Researchers are just beginning to understand the full range of plant phenotypic responses to these and other stimuli . These responses often have measurable physiological and molecular components that are readily observed . Other responses (e.g., internal biochemical changes) are less readily assayed, although sensing devices are being developed . Plant volatile emission "signatures" are particularly promising modes of plant reporting that can provide highly specific information regarding a diverse range of environmental variables on short time scales, but new approaches are needed for sensing these responses remotely . Modern molecular techniques promise to allow us to refine plant sensing and reporting, greatly enhancing the potential utility of plants as "sentinels." Continued basic research aimed at characterizing the physiological, biochemical, and molecular responses of plants to environmental stimuli (including airborne chemicals, insects and pathogenic microbes) are essential to achieving that promise.

J Toxicol Environ Health A, 2004 Apr 23-May 28, 67(8-10), 651 - 66
Predictive population dose-response assessment for Cryptosporidium parvum: infection endpoint; Englehardt JD et al.; Data validation of safe doses of pathogens in drinking water consistent with public health goals is not possible due to the number of subjects that would be needed at each dose . Because of this problem, together with the difficulty in extrapolating pathogenic response between species, and the ability of microbes to adapt rapidly, confidence-level-dependent assessments of Cryptosporidium parvum dose-response have been developed . However, these results, even on a relative basis, are dependent on confidence level, and the lack of scientific basis for this choice hampers efforts to set water quality standards . Therefore, a predictive Bayesian dose-response assessment method was proposed previously . In this article, a hierarchical predictive population dose-response Bayesian assessment for C . parvum is presented for the infection endpoint . Available data on the infectivity of three isolates of C . parvum, genotype C, were adjusted for sensitive and antibody-positive subpopulations not proportionately represented in the data, by bootstrap analysis . The diverse mean infectivities of the isolates were used to obtain a predictive distribution for population infectivity, used in turn to obtain the predictive population dose-response function . The predictive result is a distribution of unconditional probability of infection, based on available dose-response information . Information includes theoretical and empirical evidence for the conditional beta-Poisson parametric dose-response function . Results indicate that a dose of 6 x 10(-6) oocysts per exposure corresponds to 10(-4) infections per capita year . An allowable dose corresponding to goals of the SWTR should be increased over this value to reflect the illness endpoint, while potentially being reduced to account for secondary transmission among hosts if important for gastroenteritis in developed countries.

Science, 2004 Jun 11, 304(5677), 1634 - 7
Interactions and self-organization in the soil-microbe complex; Young IM et al.; Soil is the most complicated biomaterial on the planet . As with any material, the physical habitat is of prime importance in determining and regulating biological activity . However, until recently the opaque nature of soil has meant that any interrogation of its interior architecture has been relatively rudimentary, restricted to simple qualitative expressions of the physical heterogeneity that fail to relate to any specific function . However, new techniques and insights into the biophysical and biochemical processes of this inner space are leading to the developments of theoretical frameworks and experimental approaches that will allow us to sustainably manage Earth's most important resource . We introduce the concept that the soil-microbe system is self-organized and suggest new priorities for research based on an integrative approach that combines biochemistry and biophysics.

Tissue Antigens, 2004 Jul, 64(1), 13 - 7
Donor-specific sensitization by cadaveric venous allografts used for arterial reconstruction in peripheral arterial occlusive vascular disease; Balzer KM et al.; The use of allogeneic venous grafts from postmortal organ donors allows for the reconstruction of critically affected arteries in patients with peripheral occlusive vascular disease . We were interested to determine the prevalence and specificity of anti-HLA antibodies in patients after allogeneic vein transplantation . Anti-HLA class I and II alloantibodies were analyzed by flowcytometric analysis using color-coded microbeads coated with HLA antigens including recombinant single antigens . Nine out of 10 patients involving 12 venous allografts were positive for anti-HLA alloantibodies . All antibody-positive patients carried both anti-HLA class I and II alloantibodies . Anti-donor HLA specificity of the anti-HLA alloantibodies was seen in seven out of nine patients for anti-class I antibodies and in eight out of nine patients for anti-HLA class II antibodies . A high rate of donor-specific allosensitization was seen after allogeneic venous transplantation . In conclusion, allosensitization not only includes a humoral response against the constitutively expressed class I antigens but also extends to class II antigens.

Microbiol Mol Biol Rev, 2004 Jun, 68(2), 345 - 61
Microbial responses to microgravity and other low-shear environments; Nickerson CA et al.; Microbial adaptation to environmental stimuli is essential for survival . While several of these stimuli have been studied in detail, recent studies have demonstrated an important role for a novel environmental parameter in which microgravity and the low fluid shear dynamics associated with microgravity globally regulate microbial gene expression, physiology, and pathogenesis . In addition to analyzing fundamental questions about microbial responses to spaceflight, these studies have demonstrated important applications for microbial responses to a ground-based, low-shear stress environment similar to that encountered during spaceflight . Moreover, the low-shear growth environment sensed by microbes during microgravity of spaceflight and during ground-based microgravity analogue culture is relevant to those encountered during their natural life cycles on Earth . While no mechanism has been clearly defined to explain how the mechanical force of fluid shear transmits intracellular signals to microbial cells at the molecular level, the fact that cross talk exists between microbial signal transduction systems holds intriguing possibilities that future studies might reveal common mechanotransduction themes between these systems and those used to sense and respond to low-shear stress and changes in gravitation forces . The study of microbial mechanotransduction may identify common conserved mechanisms used by cells to perceive changes in mechanical and/or physical forces, and it has the potential to provide valuable insight for understanding mechanosensing mechanisms in higher organisms . This review summarizes recent and future research trends aimed at understanding the dynamic effects of changes in the mechanical forces that occur in microgravity and other low-shear environments on a wide variety of important microbial parameters.

Appl Environ Microbiol, 2004 Jun, 70(6), 3650 - 63
Sequence-specific cleavage of small-subunit (SSU) rRNA with oligonucleotides and RNase H: a rapid and simple approach to SSU rRNA-based quantitative detection of microorganisms; Uyeno Y et al.; A rapid and simple approach to the small-subunit (SSU) rRNA-based quantitative detection of a specific group of microorganisms in complex ecosystems has been developed . The method employs sequence-specific cleavage of rRNA molecules with oligonucleotides and RNase H . Defined mixtures of SSU rRNAs were mixed with an oligonucleotide (referred to as a "scissor probe") that was specifically designed to hybridize with a particular site of targeted rRNA and were subsequently digested with RNase H to proceed to sequence-dependent rRNA scission at the hybridization site . Under appropriate reaction conditions, the targeted rRNAs were correctly cut into two fragments, whereas nontargeted rRNAs remained intact under the same conditions . The specificity of the cleavage could be properly adjusted by controlling the hybridization stringency between the rRNA and the oligonucleotides, i.e., by controlling either the temperature of the reaction or the formamide concentration in the hybridization-digestion buffer used for the reaction . This enabled the reliable discrimination of completely matched rRNA sequences from single-base mismatched sequences . For the detection of targeted rRNAs, the resulting RNA fragment patterns were analyzed by gel electrophoresis with nucleotide-staining fluorescent dyes in order to separate cleaved and intact rRNA molecules . The relative abundance of the targeted SSU rRNA fragments in the total SSU rRNA could easily be calculated without the use of an external standard by determining the signal intensity of individual SSU rRNA bands in the electropherogram . This approach provides a fast and easy means of identification, detection, and quantification of a particular group of microbes in clinical and environmental specimens based on rRNA.

Anal Biochem, 2004 Jul 1, 330(1), 70 - 3
Lipase assay in soils by copper soap colorimetry; Saisuburamaniyan N et al.; A simple and sensitive method for the estimation of lipase activity in soils is reported . In this method, 50mg of soil is incubated with emulsified substrate, the fatty acids liberated are treated with cupric acetate-pyridine reagent, and the color developed is measured at 715 nm . Use of olive oil in this protocol leads to an estimation of true lipase activity in soils . The problem of released fatty acids getting adsorbed onto the soil colloids is obviated by the use of isooctane, and separate standards for different soils need not be developed . Among the various surfactants used for emulsification, polyvinyl alcohol is found to be the most effective . Incubation time of 20 min, soil concentration of 50 mg, pH 6.5, and incubation temperature of 37 degrees C were found to be the most suitable conditions for this assay . During the process of enrichment of the soils with oil, interference by the added oil is avoided by the maintenance of a suitable control, wherein 50 mg of soil is added after stopping the reaction . This assay is sensitive and it could be adopted to screen for lipase producers from enriched soils and oil-contaminated soils before resorting to isolation of the microbes by classical screening methods.

Mod Pathol, 2004 Oct, 17(10), 1211 - 6
Correlation between karyotype and quantitative immunophenotype in acute myelogenous leukemia with t(8;21); Khoury H et al.; Acute myelogenous leukemia with t(8;21) is a distinct clinicopathologic entity in which the malignant myeloblasts display a characteristic pattern of surface antigen expression . Quantitative analysis of surface marker expression in patients with this chromosomal abnormality compared to acute myelogenous leukemia patients with a different karyotype has not been reported . From 305 consecutive newly diagnosed acute myelogenous leukemia patients underwent immunophenotyping and cytogenetic analysis at our center; 16 patients (5.2%) had a t(8;21) . Fluorescence intensity values were obtained, using a set of reference microbeads, by conversion of mean channel fluorescence to molecular equivalent of soluble fluorochrome . Patients with t(8;21) displayed higher levels of CD34, HLA-DR and MPO expression (P < 0.001 for each) and lower levels of CD13 (P = 0.03) and CD33 (P = 0.02) expression . In order to study the sensitivity, specificity and predictive value of these markers, molecular equivalent of soluble fluorochrome thresholds were statistically determined . The statistically established threshold for each of the individual markers (CD34 > 60.5 x 10(3), HLA-DR > 176.1 x 10(3), MPO > 735.1 x 10(3), CD13 < 24.3 x 10(3) and CD33 < 17.3 x 10(3)) had a sensitivity of 100%, a specificity of 62-92% and a positive predictive value of 7-45% . In multivariate analysis, two quantitative patterns (CD34 > 60.5 x 10(3) and MPO > 176.1 x 10(3); CD33 < 17.3 x 10(3) and MPO > 176.1 x 10(3)) had a sensitivity, specificity and positive predictive value of 100% . These aberrant phenotypic patterns might help identify patients with t(8;21) at diagnosis and could be useful in minimal residual disease monitoring.

Immunol Res, 2004, 29(1-3), 187 - 96
Modulating TH1/TH2 responses with microbes, dendritic cells, and pathogen recognition receptors; Pulendran B; The adaptive immune system has evolved different types of immune responses, each one effective against a given class of pathogen . For example, Th1 and Th2 responses represent two qualitatively different types of immune responses that are preferentially effective against intracellular microbes and helminths, respectively . Since the original description of Th1 and Th2 T-cell clones (1), we have learned much about the cytokines that influence the type of Th response . Thus, interleukin-4 (IL-4) is known to induce IL-4 production in T cells; conversely IL-12 and interferon-gamma (IFN-gamma) are known to induce IFN-gamma production by T cells . However, the original sources of these cytokines in vivo are less clear . Recent developments from several labs point to a potential role for dendritic cells (DCs) in orchestrating this decision making process . Here, we present our current view of DC development, and then review the evidence for two opposing concepts: (1) that distinct subsets of DCs are predetermined to differentially bias the T-helper response; and (2) that microbes and the local microenvironment are potent modulators of DC function . Thus, nature appears to have evolved different mechanisms to regulate immune responses via DCs.

Biometrics, 2004 Jun, 60(2), 461 - 70
Self modeling with flexible, random time transformations; Brumback LC et al.; Methods for modeling sets of complex curves where the curves must be aligned in time (or in another continuous predictor) fall into the general class of functional data analysis and include self-modeling regression and time-warping procedures . Self-modeling regression (SEMOR), also known as a shape invariant model (SIM), assumes the curves have a common shape, modeled nonparametrically, and curve-specific differences in amplitude and timing, traditionally modeled by linear transformations . When curves contain multiple features that need to be aligned in time, SEMOR may be inadequate since a linear time transformation generally cannot align more than one feature . Time warping procedures focus on timing variability and on finding flexible time warps to align multiple data features . We draw on these methods to develop a SIM that models the time transformations as random, flexible, monotone functions . The model is motivated by speech movement data from the University of Wisconsin X-ray microbeam speech production project and is applied to these data to test the effect of different speaking conditions on the shape and relative timing of movement profiles.

Anal Bioanal Chem, 2004 Jun, 379(4), 622 - 7 Epub 2004 Jun 04.
Ion-beam analysis of CuInSe2 solar cells deposited on polyimide foil; Spemann D et al.; CuInSe(2) (CIS) solar cells deposited on polyimide foil by the Solarion company in a web-coater-based process using sputter and evaporation techniques were investigated in the ion beam laboratory LIPSION of the University of Leipzig by means of Rutherford backscattering spectrometry (RBS) and particle-induced X-ray emission (PIXE) using high-energy broad ion beams and microbeams . From these measurements the composition of the absorber and the lateral homogeneity and film thicknesses of the individual layers could be determined on the basis of some reasonable assumptions . For the first time, quantitative depth profiling of the individual elements was performed by microPIXE measurements on a beveled section prepared by ion-beam etching of a CIS solar cell . Within the CIS absorber layer no significant concentration-depth gradients were found for Cu, In, and Se, in contrast with results from secondary neutral mass spectrometry (SNMS) depth profiling, which was applied to the same samples for comparison . Furthermore, both PIXE and SNMS showed the presence of a remarkable amount of Cd from the CdS buffer layer in the underlying absorber.

Science, 2004 Jun 4, 304(5676), 1463 - 6
Challenges of modeling ocean basin ecosystems; deYoung B et al.; With increasing pressure for a more ecological approach to marine fisheries and environmental management, there is a growing need to understand and predict changes in marine ecosystems . Biogeochemical and physical oceanographic models are well developed, but extending these further up the food web to include zooplankton and fish is a major challenge . The difficulty arises because organisms at higher trophic levels are longer lived, with important variability in abundance and distribution at basin and decadal scales . Those organisms at higher trophic levels also have complex life histories compared to microbes, further complicating their coupling to lower trophic levels and the physical system . We discuss a strategy that builds on recent advances in modeling and observations and suggest a way forward that includes approaches to coupling across trophic levels and the inclusion of uncertainty.

J Microbiol Methods, 2004 Jul, 58(1), 119 - 29
13CO2 pulse labelling of plants in tandem with stable isotope probing: methodological considerations for examining microbial function in the rhizosphere; Griffiths RI et al.; Recently developed 13CO2 pulse labelling and stable isotope probing (SIP) methods offer the potential to track 13C-labelled plant photosynthate into phylogenetic groups of microbial taxa in the rhizosphere, permitting an examination of the link between soil microbial diversity and carbon flow in situ . We tested the feasibility of this approach to detect functional differences in microbial communities utilising recently fixed plant photosynthate in moisture perturbed grassland turfs . Specifically, we addressed two questions: (1) How does moisture perturbation (three treatments; continual wetting, drying, and drying followed by rewetting) affect the assimilation of 13C-labelled exudates carbon into the soil microbial community?; (2) Can 13C deposited in soil from pulse-labelled plants be used to identify microbes utilising plant exudates using SIP methodologies? Net CO2 fluxes showed that prior to 13CO2 pulse labelling, all treatments were photosynthetically active, but differences were observed in night time respiration, indicating moisture treatments had impacted on net CO2 efflux . Measurements of pulse-derived 13C incorporated into soil RNA over 2 months showed that there was only evidence of 13C enrichment in the continuously wetted treatments . However, isotopic values represented only a 0.1-0.2 13C at.% increase over natural abundance levels and were found to be insufficient for the application of RNA-SIP . These findings reveal that in this experimental system, the microbial uptake of labelled carbon from plant exudates is low, and further optimisation of methodologies may be required for application of SIP to natural plant-soil systems where 13C tracer dilution is a consideration .

J Hazard Mater, 2004 Jun 18, 109(1-3), 201 - 11
Performance of BTX degraders under substrate versatility conditions; Maliyekkal SM et al.; A microbial consortium acclimatized with benzene, toluene or xylene (BTX) was employed to study the degradation pattern of these compounds individually under aerobic conditions . Batch and continuous experiments were conducted to evaluate the adaptability of the enriched cultures under substrate versatility conditions . The bio-kinetic parameters obtained under substrate versatility conditions were compared with those of a single substrate condition . Similar degradation patterns were observed for all the substrates with inhibition occurring at higher concentration (approximately 150 mg/L for benzene and xylene, and approximately 200 mg/L for toluene) . Toluene degradation was highest, followed by benzene and xylene in the aqueous phase . Adaptation to a more toxic compound like benzene and xylene improved the utilization of toluene . On the other hand, microbes grown on a less toxic compound (toluene) grew at a lower rate in the presence of more toxic compounds . Suitable kinetic parameters such as micro(max) (maximum specific growth rate per hour), Ks (half saturation constant, mg/L), and KI (threshold substrate inhibition constant, mg/L) were determined using Haldane and Levenspiel substrate inhibition models . The Haldane equation seems to be an adequate expression for the system . The degradation behavior of pollutants in the gas phase was also evaluated using a toluene acclimatized biotrickling filter operated in continuous mode . The biotrickling filter acclimatized with toluene could degrade benzene and xylene with a lower elimination capacity . But, the system could recover its original efficiency quite fast even after a prolonged shock loading . The degradation was better for toluene, followed by benzene and xylene.

Am J Physiol Cell Physiol, 2004 Sep, 287(3), C643 - 54 Epub 2004 Jun 02.
Cytoskeletal mechanics in adherent human airway smooth muscle cells: probe specificity and scaling of protein-protein dynamics; Puig-de-Morales M et al.; We probed elastic and loss moduli in the adherent human airway smooth muscle cell through a variety of receptor systems, each serving as a different molecular window on cytoskeletal dynamics . Coated magnetic microbeads were attached to the cell surface via coating-receptor binding . A panel of bead coatings was investigated: a peptide containing the sequence RGD, vitronectin, urokinase, activating antibody against beta(1)-integrin, nonactivating antibody against beta(1)-integrin, blocking antibody against beta(1)-integrin, antibody against beta(1)-integrin, and acetylated low-density lipoprotein . An oscillatory mechanical torque was applied to the bead, and resulting lateral displacements were measured at baseline, after actin disruption by cytochalasin D, or after contractile activation by histamine . As expected, mechanical moduli depended strongly on bead type and bead coating, differing at the extremes by as much as two orders of magnitude . In every case, however, elastic and loss moduli increased with frequency f as a weak power law, f( x-1) . Moreover, with few exceptions, data could be scaled such that elastic and frictional responses depended solely on the power law exponent x . Taken together, these data suggest that power law behavior represents a generic feature of underlying protein-protein dynamics.

Microb Cell Fact . 2004 Jun 2;3(1):7.
Potential use of sugar binding proteins in reactors for regeneration of CO2 fixation acceptor D-Ribulose-1,5-bisphosphate; Mahato S et al.; Sugar binding proteins and binders of intermediate sugar metabolites derived from microbes are increasingly being used as reagents in new and expanding areas of biotechnology . The fixation of carbon dioxide at emission source has recently emerged as a technology with potentially significant implications for environmental biotechnology . Carbon dioxide is fixed onto a five carbon sugar D-ribulose-1,5-bisphosphate . We present a review of enzymatic and non-enzymatic binding proteins, for 3-phosphoglycerate (3PGA), 3-phosphoglyceraldehyde (3PGAL), dihydroxyacetone phosphate (DHAP), xylulose-5-phosphate (X5P) and ribulose-1,5-bisphosphate (RuBP) which could be potentially used in reactors regenerating RuBP from 3PGA . A series of reactors combined in a linear fashion has been previously shown to convert 3-PGA, (the product of fixed CO2 on RuBP as starting material) into RuBP (Bhattacharya et al., 2004; Bhattacharya, 2001) . This was the basis for designing reactors harboring enzyme complexes/mixtures instead of linear combination of single-enzyme reactors for conversion of 3PGA into RuBP . Specific sugars in such enzyme-complex harboring reactors requires removal at key steps and fed to different reactors necessitating reversible sugar binders . In this review we present an account of existing microbial sugar binding proteins and their potential utility in these operations.

West Afr J Med, 2004 Jan-Mar, 23(1), 54 - 7
Intracerebral abscesses: outcome following management in the CT era; Emejulu JK et al.; BACKGROUND: There are no pathognomonic presenting features of intracerebral abscesses . Their clinical features could be confused with those of a tumour or any other space occupying lesion . STUDY DESIGN: We analyzed the symptoms, predisposing factors, diagnostic modalities, prevalent causative microbes, management and outcome in 11 patients, for whom complete records were available and who were managed for confirmed intracerebral abscess in our Neurosurgical Unit from 1996 to 2000 and compared our findings with those from other series . RESULTS: In these patients focal neurological deficit was the most common clinical feature, trauma was the most predisposing event and Computerised Tomography (CT) Scanning remained the most dependable diagnostic tool . In our practice, burr hole drainage augmented with a minimum of 4 weeks intravenous antibiotics, was the most frequently used treatment modality . CONCLUSION: The outcome was satisfactory (ability to return to pre-morbid activities and duties, with no disabling neurological deficits) in 72.8% cases . There was one death . This modality remains an effective way of treating patients with intracebrebral abscess.

Trends Microbiol, 2004 Jun, 12(6), 259 - 63
The weapon potential of a microbe; Casadevall A et al.; The designation of a microbe as a potential biological weapon poses the vexing question of how such a decision is made given the many pathogenic microbes that cause disease . Analysis of the properties of microbes that are currently considered biological weapons against humans revealed no obvious relationship to virulence, except that all are pathogenic for humans . Notably, the weapon potential of a microbe rather than its pathogenic properties or virulence appeared to be the major consideration when categorizing certain agents as biological weapons . In an effort to standardize the assessment of the risk that is posed by microbes as biological warfare agents using the basic principles of microbial communicability (defined here as a parameter of transmission) and virulence, a simple formula is proposed for estimating the weapon potential of a microbe.

Int J Antimicrob Agents, 2004 Mar, 23(3), 209 - 12
Antibiotic resistance from two perspectives: man and microbe; Hamilton-Miller JM; Despite much effort, antibiotic resistance continues to increase . Looking back, it is clear that this was an inevitable consequence of antibiotic use . From a bacterial viewpoint, the introduction of antibiotics was a tremendous stimulus to evolution . As a survival reaction to stress (selection pressure) bacteria, by means of their extreme biochemical and genetic versatility, have adapted to 21st Century conditions . Resistance can be to some extent contained by less and better use of antibiotics, but ultimately novel approaches to the treatment and prevention of infectious diseases will have to be forthcoming . This will only be achieved if best use is made of alternative resources presently available and most importantly, man's ingenuity must be fully engaged.

Mol Gen Mikrobiol Virusol, 2004, (2), 24 - 8
{Genotyping of the Francisella tularensis strains isolated from natural foci of tularemia in the Rostov region by multilocus VNTR analysis}; Vodop'ianov AS et al.; On the basis of an analysis of the VNTR alleles' distribution in 109 strains of F . tularensis it was established that 19 genotypes of the disease causative agent circulated in the Rostov Region from 1945 to 2002 . The microbe-provoked infection episodes can be divided into polyclonal, monoclonal and cluster ones . A retrospective analysis of the genotypes' distribution is indicative of that strains of similar or of closely-related genotypes circulate simultaneously in the studied territory . All investigated F . tularensis strains could be differentiated into two groups; strains, whose genotypes are encountered almost evenly within the entire Region's territory, belong to group 1; and strains of group 2 displayed a trend towards being geographically bound . Isolations of cultures with similar (close) genotypic features made in prolonged time periods suggest that a part of F . tularensis clones can persist for a long time in environmental foci . A set of strains described by genotype can provide a foundation for a database of the tularemic microbe culture within the geo-information system of the South Federative Okrug of Russia.

Microbes Infect, 2004 May, 6(6), 609 - 16
Innate immune recognition of microbes through Nod1 and Nod2: implications for disease; Carneiro LA et al.; Nod1 and Nod2 are cytosolic proteins involved in intracellular recognition of microbes and their products . Recently, it was shown that these proteins recognize different moieties of bacterial peptidoglycan (PGN) mediating non-specific pathogen resistance and possibly generating signals for the adaptive immune response . Moreover, mutations in the gene encoding Nod2 are associated with increased susceptibility to chronic inflammatory disorders.

Infect Genet Evol, 2004 Jun, 4(2), 159 - 66
The best defence against bioweapons has already been invented by evolution; Rumyantsev SN; Bioweapons are considered from the viewpoint of the mutual evolution of microbe/victim ecological systems . Cases considered include accidental, experimental, and real exploitation of bioweapons as well as other cases in the history of epidemics, and also experimental investigations . It is proposed here that speculations about bioweapon's very high mass annihilating ability are based on over-extrapolation from limited data selected from the ancient history of epidemics and on a false supposition that all people are susceptible to any bioweapon's infectious agent . The history of epidemics, clinical and genetic observations, data from experimental investigations, and results of accidental, experimental, and real exploitation of bioweapons do not confirm the mass annihilating capability of bioweapons . Many people possess constitutional (genetic) immunity presumably formed by natural selection over many human generations . This genetic protection exists in an individual prior to infection . The power of constitutional immunity played an important role during all human evolution . Its protective capability continues to defend humanity from mass annihilation by both epidemics and bioweapons . In this context of constitutional immunity for the majority of individuals, the main goal of modern medicine is to identify and defend the defenseless ones.

J Speech Lang Hear Res, 2004 Apr, 47(2), 342 - 52
Use of electromagnetic midsagittal articulography in the study of swallowing; Steele CM et al.; The tongue functions as the primary articulator during the oropharyngeal stages of swallowing . However, detailed descriptions of the kinematics and spatiotemporal variability of tongue behaviors during swallowing are limited to a handful of analyses of data from the X-ray microbeam database . In this article, a new technique, electromagnetic midsagittal articulography (EMMA), is introduced for the high-resolution description of oral articulatory movements during swallowing . Data from 8 healthy, nondysphagic participants are used to illustrate the methods used for data collection and analysis . Movement data were collected for 3 fleshpoint positions on the tongue (blade, body, dorsum) during sequences of repeated discrete water swallows, and were characterized for variables of spatiotemporal variability and 4 discrete kinematic parameters (movement amplitude, peak velocity, duration, and kinematic stiffness) . These data show that the movement trajectories measured using EMMA are consistent with descriptions from previous X-ray microbeam studies, indicating that EMMA is a feasible method for the detailed study of tongue movements during swallowing.

J Immunol, 2004 Jun 1, 172(11), 6978 - 87
Toll-like receptor 4 signaling by intestinal microbes influences susceptibility to food allergy; Bashir ME et al.; The mechanisms by which signaling by the innate immune system controls susceptibility to allergy are poorly understood . In this report, we show that intragastric administration of a food allergen with a mucosal adjuvant induces allergen-specific IgE, elevated plasma histamine levels, and anaphylactic symptoms in three different strains of mice lacking a functional receptor for bacterial LPS (Toll-like receptor 4 (TLR4)), but not in MHC-matched or congenic controls . Susceptibility to allergy correlates with a Th2-biased cytokine response in both the mucosal (mesenteric lymph node and Peyer's patch) and systemic (spleen) tissues of TLR4-mutant or -deficient mice . TLR4-mutant mice are not inherently impaired in their ability to regulate Th1 cytokine production because they respond to stimulation via TLR9 . Coadministration of CpG oligodeoxynucleotides during sensitization of TLR4-mutant mice with allergen plus CT abrogates anaphylactic symptoms and Ag-specific IgE, and results in a Th1-polarized cytokine response . When the composition of the bacterial flora is reduced and altered by antibiotic administration (beginning at 2 wk of age), TLR4 wild-type mice become as susceptible to the induction of allergy as their TLR4-mutant counterparts . Both allergen-specific IgE and Th2 cytokine responses are reduced in antibiotic-treated mice in which the flora has been allowed to repopulate . Taken together, our results suggest that TLR4-dependent signals provided by the intestinal commensal flora inhibit the development of allergic responses to food Ags.

Am Fam Physician, 2004 May 1, 69(9), 2248 - 9
Sinus and Allergy Health Partnership releases report on adult chronic rhinosinusitis; Ressel G; The report states that chronic rhinosinusitis is an evolving area of study and new information is being assessed on an ongoing basis . Debate continues about potential etiologies and associated conditions, pathophysiology, common inflammatory mediators, and whether or not most cases of chronic rhinosinusitis are associated with infection . Evidence in the most recent literature supports the significant role that microbes (i.e., bacteria, fungi) play in the inflammatory process . The report discusses classification schemes, the role of potential bacterial, allergic, and fungal etiologies, and the controversies about these areas of study.

Sci STKE . 2004 May 11;2004(233):pe21.
During the respiratory burst, do phagocytes need proton channels or potassium channels, or both?
DeCoursey TE.
The NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidase enzyme complex, a crucial component of innate immunity, produces superoxide anion (O2-), which is a precursor to many reactive oxygen species . NADPH oxidase produces O2- by transferring electrons from intracellular NADPH across the membrane to extracellular (or phagosomal) oxygen and is thus electrogenic . It is widely believed that electroneutrality is preserved by proton flux through voltage-gated proton channels . A series of recent papers have challenged several key aspects of this view of the "respiratory burst." The most recent study solidifies the proposal that O2- and other reactive oxygen species produced by phagocytes are not toxic to microbes under physiological conditions . Further, an essential role for high-conductance, Ca2+-activated K+ (maxi-K+) channels in microbe killing is proposed . Finally, the results cast doubt on the widely held view that H+ efflux through voltage-gated proton channels (i) is the main mechanism of charge compensation, and (ii) is essential to continuous O2- production by the NADPH oxidase . My analysis of the new data and of a large body of data in the literature indicates that the proposed role of maxi-K+ channels in the respiratory burst is not yet credibly established . H+ efflux through proton channels thus remains the most viable mechanism for charge compensation and continuous O2- production . The important question of the toxicity of reactive oxygen species in phagocytes and in other cells, which has long been simply taken for granted, is a widespread assumption that deserves critical study.

Vaccine, 2004 Jun 2, 22(17-18), 2106 - 15
Sub-compartmentalization of the gastrointestinal (GI) immune system determined with microbeads that differ in release properties; Cronkhite RI et al.; Immunization of two specific regions of the murine GI tract of two types of mice with ovalbumin (OVA) encapsulated in microbeads with two different pH-sensitive coatings allowed a more precise analysis of this compartment of the mucosal immune system . Acute, chronic and pervasive immunization protocols were utilized in an attempt to stimulate specific types of immunity . Chronic immunization potentiated antibody isotypes influenced by type 2 T helper cells (T(h)2) . Pervasive immunization of both regions of the GI tract mimicked chronic immunization, stimulating high levels of OVA-reactive IgE . Acute immunization was best able to potentiate isotypes influenced by type 1 T helper cells (T(h)1) and a sequential segregated immunization protocol allowed the targeting of T(h)1-like memory responses.

Int J Med Microbiol, 2004 Apr, 293(7-8), 453 - 61
Pathogenomics of mobile genetic elements of toxigenic bacteria; Hacker J et al.; The growing knowledge of genetic diversity and whole genome organization in bacteria shows that pathogenicity islands (PAIs) represent a subtype of a more general genetic element, termed genomic island (GEI), which is widespread among pathogenic and non-pathogenic microbes . These findings mirror the importance of horizontal gene transfer, genome reduction and recombination events as fundamental mechanisms involved in evolution of bacterial variants . GEIs are part of the flexible gene pool and carry selfish genes, but also determinants which may be beneficial under certain conditions thus increasing bacterial fitness and consequently their survival or transmission . In this review, we focus on the role of mobile genetic elements that may also contain toxin-encoding genes for genome variability and evolution of bacteria.

Arch Dermatol Res, 2004 Aug, 296(3), 120 - 4 Epub 2004 May 18.
Differentiation-regulated expression of Toll-like receptors 2 and 4 in HaCaT keratinocytes; Pivarcsi A et al.; Toll-like receptors (TLRs) play an important role in the recognition of pathogens in keratinocytes . In this study, we investigated whether the differentiation state of HaCaT keratinocytes correlates with the expression of TLR2 and TLR4 genes . The expression levels of TLR2 and TLR4 in a HaCaT differentiation model system were determined using quantitative real-time RT-PCR (Q-RT-PCR) and flow cytometry . The progression of keratinocyte differentiation was monitored by determining the level of involucrin gene expression using Q-RT-PCR . The expression levels of TLR2 and TLR4 increased with the stage of differentiation and there were strong correlations between the expression level of the involucrin gene and those of the TLR2 gene ( r=0.809, P<0.0001) and the TLR4 gene ( r=0.568, P<0.02) . Increased cell surface expression of TLR2 and TLR4 was also found in differentiated HaCaT keratinocytes by flow cytometric analysis . Our findings suggest that upregulation of TLR expression during differentiation in keratinocytes could be a part of the differentiation process of keratinocytes and could have biological significance in protecting skin against microbes.

Methods Mol Biol, 2004, 266, 261 - 87
Using the genome to understand pathogenicity; Field D et al.; Genome sequencing, the determination of the complete complement of DNA in an organism, is revolutionizing all aspects of the biological sciences . Genome sequences make available for scientific scrutiny the complete genetic capacity of an organism . With respect to microbes, this means we now have the unprecedented opportunity to investigate the molecular basis of commensal and virulence behavior . We now have genome sequences for a wide range of bacterial pathogens (obligate, facultative, and opportunistic); this has facilitated the discovery of many previously unidentified determinants of pathogenicity and has provided novel insights into what creates a pathogen . In-depth analyses of bacterial genomes are also providing new perspectives on bacterial physiology, molecular adaptation to a preferred niche, and genomic susceptibility to the uptake of foreign DNA, three key factors that can play a significant role in determining whether a species, or a strain, will have pathogenic potential.

Methods Mol Biol, 2004, 266, 71 - 113
Genome plasticity: insertion sequence elements, transposons and integrons, and DNA rearrangement; Bennett PM; Living organisms are defined by the genes they possess . Control of expression of this gene set, both temporally and in response to the environment, determines whether an organism can survive changing conditions and can compete for the resources it needs to reproduce . Bacteria are no exception; changes to the genome will, in general, threaten the ability of the microbe to survive, but acquisition of new genes may enhance its chances of survival by allowing growth in a previously hostile environment . For example, acquisition of an antibiotic resistance gene by a bacterial pathogen can permit it to thrive in the presence of an antibiotic that would otherwise kill it; this may compromise clinical treatments . Many forces, chemical and genetic, can alter the genetic content of DNA by locally changing its nucleotide sequence . Notable for genetic change in bacteria are transposable elements and site-specific recombination systems such as integrons . Many of the former can mobilize genes from one replicon to another, including chromosome-plasmid translocation, thus establishing conditions for interspecies gene transfer . Balancing this, transposition activity can result in loss or rearrangement of DNA sequences . This chapter discusses bacterial DNA transfer systems, transposable elements and integrons, and the contributions each makes towards the evolution of bacterial genomes, particularly in relation to bacterial pathogenesis . It highlights the variety of phylogenetically distinct transposable elements, the variety of transposition mechanisms, and some of the implications of rearranging DNA, and addresses the effects of genetic change on the fitness of the microbe.

Biochem Biophys Res Commun, 2004 Jun 11, 318(4), 1025 - 30
Evidences showing ultraviolet-B radiation-induced damage of DNA in cyanobacteria and its detection by PCR assay; Kumar A et al.; Impact of ultraviolet-B radiation in causing the damages to the DNA of the cyanobacterium, Anabaena strain BT2 has been investigated . Exposure of genomic DNA (in vitro) to UV-B radiation for 1 h did not cause any shift in the absorption peak (lambda(max)) but more than 30% increase in absorbance was noticed in comparison to untreated control DNA (no exposure to UV-B) . This increase in absorbance in a way may be comparable to typical hypochromic effect but there was no decrease in absorbance following transfer of UV-B-treated DNA to fluorescent light or in the dark . That the damaging effect of UV-B radiation on native structure of DNA is indeed real was also evident from the PCR-based assay such as RAPD, rDNA amplification, and ARDRA . Template activity of UV-B-treated genomic DNA was drastically inhibited, there was no amplification in RAPD assay after prior exposure of DNA to UV-B for 60 min . Only one band of approximately 400 bp was observed even after 60 min of exposure which suggests that certain segment of DNA strand is resistant to UV-B effects . Similar to the effects on RAPD profile, amplification of rDNA was significantly inhibited following exposure of genomic DNA to UV-B . Our findings clearly demonstrate that UV-B does affect the DNA of cyanobacteria and the killings of these microbes might be due to the irreversible damages caused to DNA by this high energy radiation . It is felt that PCR assay may be conveniently used for screening the damages caused to DNA by UV-B radiation in cyanobacteria and other microorganisms.

Clin Exp Immunol, 2004 Jun, 136(3), 521 - 6
Expression and subcellular distribution of toll-like receptors TLR4, TLR5 and TLR9 on the gastric epithelium in Helicobacter pylori infection; Schmausser B et al.; Toll-like receptors (TLRs) expressed by mucosal epithelium play an essential role in the defense against microbes by recognizing conserved bacterial molecules . For the first time TLR4, TLR5 and TLR9 have been microanatomically localized in patients with noninflamed gastric mucosa and Helicobacter pylori gastritis by immunohistochemistry . Because polarized expression of TLRs in apical and basolateral epithelial compartments is thought to modulate mucosal immunity, subcellular TLR distribution by gastric epithelium was investigated using confocal microscopy . TLR4, TLR5 and TLR9 were expressed by gastric epithelium in antrum and corpus of all patients with H . pylori gastritis (n = 14) and with noninflamed gastric mucosa (n = 5) . TLR4 was expressed at the apical and the basolateral pole of the gastric epithelium as well in noninflamed gastric mucosa as in H . pylori gastritis . TLR5 and TLR9 expression in the noninflamed gastric mucosa was identical to that of TLR4 with localization at the apical and the basolateral epithelial pole . However, in H . pylori gastritis TLR5 and TLR9 expression on the gastric epithelium changed to an exclusive basolateral localization without detectable expression at the apical pole . In the human stomach, the gastric epithelium expressed TLR4, TLR5 and TLR9, which gives it the possibility to interact with H . pylori . Furthermore, gastric epithelial TLR4 expression is highly polarized in an apical and a basolateral compartment, whereas TLR5 and TLR9 polarization seems to be a process dynamically influenced by H . pylori infection . This polarized and dynamically regulated gastric epithelial expression of TLRs supports a sentinel role for these receptors in the mucosal immunity to H . pylori.

Ying Yong Sheng Tai Xue Bao, 2004 Feb, 15(2), 295 - 8
{Degradation of methyl parathion in soil and Chinese chive by strain DLL-1}; Zhang R et al.; Degradation of methyl parathion in soil and Chinese chive by strain DLL-1 was studied . Usage of methyl parathion at 7.5, 15, and 22.5 kg(a.i.).hm-2 resulted in the average amount of residue of 0.663, 1.270, and 1.901 mg.kg-1 in Chinese chive respectively . The natural degradation rate was 98.94%, 96.44%, and 96.04% corresponding to the 3 levels of usage . The amount of pesticide residue could be significantly decreased through the application of high effective degrading microbial agents . The amount of pesticide residue in Chinese chive and soil was 0.269 and 0.099 mg.kg-1 with the usage of 75 kg.hm-2 of degradation bacterium, which was decreased by 78.82% and 98.68% compared with the control . Increasing the bacterium usage led to the increase of degradation rate . Usage of degradation bacterium more than 75 kg.hm-2 did not increase the degradation rate further . The best time of the application of the degrading microbe was 3 days after the application of the pesticide.

Immunity, 2004 May, 20(5), 551 - 62
Dynamics of blood-borne CD8 memory T cell migration in vivo; Klonowski KD et al.; Memory T cells are distributed throughout the body following infection, but the migratory dynamics of the memory pool in vivo is unknown . The ability of circulating microbe-specific memory T cells to populate lymphoid and nonlymphoid tissues was examined using adoptive transfer and parabiosis systems . While migration of memory CD8 T cells to lymph nodes and peritoneal cavity required G(i)-coupled receptor signaling, migration to the spleen, bone marrow, lung, and liver was independent of this pathway . Following parabiosis, memory T cells rapidly equilibrated into the lymphoid tissues, lung, and liver of each parabiont, implying most memory cells were not obligately tissue resident . Equilibration of memory cell populations was delayed in the brain, peritoneal cavity, and intestinal lamina propria, indicating controlled gating for entry into these tissues . In addition, memory cell migration to the lamina propria required beta7 integrins . Thus, the blood-borne T cell pool serves to maintain the homeostasis of tissue-based memory populations.

Plant Physiol, 2004 May, 135(1), 47 - 58
Proton-transfer-reaction mass spectrometry as a new tool for real time analysis of root-secreted volatile organic compounds in Arabidopsis; Steeghs M et al.; Plant roots release about 5% to 20% of all photosynthetically-fixed carbon, and as a result create a carbon-rich environment for numerous rhizosphere organisms, including plant pathogens and symbiotic microbes . Although some characterization of root exudates has been achieved, especially of secondary metabolites and proteins, much less is known about volatile organic compounds (VOCs) released by roots . In this communication, we describe a novel approach to exploring these rhizosphere VOCs and their induction by biotic stresses . The VOC formation of Arabidopsis roots was analyzed using proton-transfer-reaction mass spectrometry (PTR-MS), a new technology that allows rapid and real time analysis of most biogenic VOCs without preconcentration or chromatography . Our studies revealed that the major VOCs released and identified by both PTR-MS and gas chromatography-mass spectrometry were either simple metabolites, ethanol, acetaldehyde, acetic acid, ethyl acetate, 2-butanone, 2,3,-butanedione, and acetone, or the monoterpene, 1,8-cineole . Some VOCs were found to be produced constitutively regardless of the treatment; other VOCs were induced specifically as a result of different compatible and noncompatible interactions between microbes and insects and Arabidopsis roots . Compatible interactions of Pseudomonas syringae DC3000 and Diuraphis noxia with Arabidopsis roots resulted in the rapid release of 1,8-cineole, a monoterpene that has not been previously reported in Arabidopsis . Mechanical injuries to Arabidopsis roots did not produce 1,8-cineole nor any C6 wound-VOCs; compatible interactions between Arabidopsis roots and Diuraphis noxia did not produce any wound compounds . This suggests that Arabidopsis roots respond to wounding differently from above-ground plant organs . Trials with incompatible interactions did not reveal a set of compounds that was significantly different compared to the noninfected roots . The PTR-MS method may open the way for functional root VOC analysis that will complement genomic investigations in Arabidopsis.

J Biol Chem, 2004 Jul 16, 279(29), 30143 - 9 Epub 2004 May 11.
Selective and efficient immunoprecipitation of the disease-associated form of the prion protein can be mediated by nonspecific interactions between monoclonal antibodies and scrapie-associated fibrils; Morel N et al.; Transmissible spongiform encephalopathies are characterized by the accumulation in brain tissues of an abnormal isoform of the prion protein named PrPsc, which is the only direct marker known for transmissible spongiform encephalopathies . Here we show that PrPsc can be specifically immunoprecipitated by using several monoclonal antibodies (mAbs) of various specificities independently of the properties of their binding site (paratope) . These results strongly suggest that a significant proportion of mAbs can interact with PrPsc aggregates through nonspecific paratope-independent interactions allowing selective immunoprecipitation of PrPsc when these mAbs are immobilized on a polydisperse solid phase like microbeads.

J Biol Chem, 2004 Jul 23, 279(30), 31010 - 7 Epub 2004 May 11.
Drug uptake and modulation of drug resistance in Leishmania by an aquaglyceroporin; Gourbal B et al.; Leishmaniasis is a protozoan parasitic disease that affects 12 million people worldwide . The first line choice for the treatment of this disease is antimonial drugs . In the endemic regions, resistance to this class of drugs is a major impediment to treatment . Microbes often become resistant to drugs by mutation or down-regulation of uptake systems, but the uptake system for the antimonial drugs in Leishmania is unknown . In other organisms, aquaglyceroporins have been shown to facilitate uptake of trivalent metalloids . In this study, we report the identification and characterization of aquaglyceroporins from Leishmania major (LmAQP1) and Leishmania tarentolae (LtAQP1), respectively . These Leishmania proteins have the conserved signature motifs of aquaglyceroporins . Transfection of LmAQP1 into three species of Leishmania, L . tarentolae, Leishmania infantum, and L . major, produced hypersensitivity to both As(III) and Sb(III) in all three strains . Increased production of LmAQP1 was detected by immunoblotting . Drug-resistant parasites with various mutations leading to resistance mechanisms became hypersensitive to both metalloids after expression of LmAQP1 . Increased rates of uptake of As(III) or Sb(III) correlated with metalloid sensitivity of the wild type and drug-resistant transfectants . Transfection of LmAQP1 in a Pentostam-resistant field isolate also sensitized the parasite in the macrophage-associated amastigote form . One allele of LmAQP1 was disrupted in L . major, and the resulting cells became 10-fold more resistant to Sb(III) . This is the first report of the uptake of a metalloid drug by an aquaglyceroporin in Leishmania, suggesting a strategy to reverse resistance in the field.

Biol Sci Space, 2003 Dec, 17(4), 298 - 301
Development of an ion microbeam system for irradiating single plant cell{s}; Yokota Y et al.; An ion microbeam system for irradiating single plant cells was developed to analyze exact biological effects of ion beams . Tobacco BY-2 protoplasts were used as a model of single plant cells . Protoplasts were cultured in thin agarose medium on a specially designed irradiation-vessel, which has a CR-39 nuclear track detector (a 100-micro meter thick sheet) . The colony formation rate of unirradiated protoplasts was 22.7 +- 6.7% (mean +- SE of 3 different experiments) after a month of culture . Protoplasts were irradiated with programmed numbers of 18.3 MeV/u carbon ions that had been collimated by a 20-micro mf micro-aperture . After the irradiation, the positions within the protoplasts that were hit with ions were accurately determined by etching the CR-39 sheet in 13.4M KOH solution at 27 centigrade for 9h . The hit rate of the carbon ion microbeam, i.e., the percent of the ion particles that hit the protoplast that they were aimed at, was 56.9 +- 2.4% (mean +- SE of 7 different replications).

Spectrochim Acta A Mol Biomol Spectrosc, 2004 May, 60(6), 1349 - 57
Search for EPR markers of the history and origin of the insoluble organic matter in extraterrestrial and terrestrial rocks; Gourier D et al.; The insoluble organic matter (IOM) of three carbonaceous meteorites (Orgueil, Murchison and Tagish Lake meteorites) and three samples of cherts (microcrystalline SiO2 rock) containing microfossils with age ranging between 45 million years and 3.5 billion years is studied by electron paramagnetic resonance (EPR) . The age of the meteorites is that of the solar system (4.6 billion years) . The purpose of this work was to determine the EPR parameters, which allow us to discriminate between biogenic and extra terrestrial origin for the organic matter . Such indicators should be relevant for the controversy regarding the biogenicity of the organic matter in the oldest cheroot (3.5 billion years) and in Martian meteorites containing microbe-like microstructures . The organic matter of meteorites contains a high concentration of diradicaloid moieties characterised by a diamagnetic ground state S = 0 and a thermally accessible triplet state S = 1 . The three meteorites exhibit the same singlet-triplet gap (ST gap) DeltaE approximately 0.1 eV . To the best of our knowledge, such diradicaloids are unknown in insoluble organic matter of terrestrial origin . We have also shown that the EPR linewidth of insoluble organic matter in cherts and coals decrease logarithmically with the age of the organic matter . We conclude from this result that the organic matter in the oldest cherts (3.5 billion years) has the same age as their SiO2 matrix, and is not due to a latter contamination by bacteria, as was recently found in meteoritic samples.

J Eukaryot Microbiol, 2004 Mar-Apr, 51(2), 125 - 38
Viral control of phytoplankton populations--a review; Brussaard CP; Phytoplankton population dynamics are the result of imbalances between reproduction and losses . Losses include grazing, sinking, and natural mortality . As the importance of microbes in aquatic ecology has been recognized, so has the potential significance of viruses as mortality agents for phytoplankton . The field of algal virus ecology is steadily changing and advancing as new viruses are isolated and new methods are developed for quantifying the impact of viruses on phytoplankton dynamics and diversity . With this development, evidence is accumulating that viruses can control phytoplankton dynamics through reduction of host populations, or by preventing algal host populations from reaching high levels . The identification of highly specific host ranges of viruses is changing our understanding of population dynamics . Viral-mediated mortality may not only affect algal species succession, but may also affect intraspecies succession . Through cellular lysis, viruses indirectly affect the fluxes of energy, nutrients, and organic matter, especially during algal bloom events when biomass is high . Although the importance of viruses is presently recognized, it is apparent that many aspects of viral-mediated mortality of phytoplankton are still poorly understood . It is imperative that future research addresses the mechanisms that regulate virus infectivity, host resistance, genotype richness, abundance, and the fate of viruses over time and space.

Microbiology, 2004 May, 150(Pt 5), 1507 - 18
Induction of Mycobacterium avium growth restriction and inhibition of phagosome-endosome interactions during macrophage activation and apoptosis induction by picolinic acid plus IFNgamma; Pais TF et al.; Treatment of mouse macrophages with picolinic acid (PA) and gamma-interferon (IFNgamma) led to the restriction of Mycobacterium avium proliferation concomitant with the sequential acquisition of metabolic changes typical of apoptosis, mitochondrial depolarization, annexin V staining and caspase activation, over a period of up to 5 days . However, triggering of cell death by ATP, staurosporine or H(2)O(2) failed to affect mycobacterial viability . In contrast to untreated macrophages where extensive interactions between phagosomes and endosomes were observed, phagosomes from treated macrophages lost the ability to acquire endosomal dextran . N-Acetylcysteine was able to revert both the anti-mycobacterial activity of treated macrophages as well as the block in phagosome-endosome interactions . The treatment, however, induced only a minor increase in the acquisition of lysosomal markers, namely Lamp-1, and did not increase to any great extent the acidification of the phagosomes . These data thus suggest that the anti-mycobacterial activity of PA and IFNgamma depends on the interruption of intracellular vesicular trafficking, namely the blocking of acquisition of endosomal material by the microbe.

Microbiology, 2004 May, 150(Pt 5), 1261 - 70
Spore morphotypes of Thelohania solenopsae (microsporidia) described microscopically and confirmed by PCR of individual spores microdissected from smears by position ablative laser microbeam microscopy; Sokolova YY et al.; Development of Thelohania solenopsae, a parasite of the red imported fire ant (Solenopsis invicta), until recently was thought to include formation of two types of spores: unicellular meiospores, maturing inside sporophorous vesicles in sets of eight (octospores); and Nosema-like binuclear free spores . Megaspores, discovered in 2001, develop primarily in alates and are morphologically distinct from the two previously known types of spores . The role of megaspores in the T . solenopsae life cycle, as well as their existence, has been questioned . The current research includes light and electron microscopic descriptions of the three major spore morphotypes characteristic of T . solenopsae development . In addition, individual octospores and megaspores were isolated into groups of 8-20 from methanol-fixed and Calcofluor-stained smears of the infected ants for subsequent PCR analysis by the laser pressure catapulting function of a position ablative laser microbeam microscope, a technique applied for the first time to research of microsporidia . The PCR-amplified SSU rDNA nucleotide sequences from octospores and megaspores were identical . This, along with the consistency with which megaspores are detected in infected ants, demonstrates that megaspores are integral to the life cycle of T . solenopsae.

Appl Environ Microbiol, 2004 May, 70(5), 3158 - 62
Bacteriophage-based genetic system for selection of nonsplicing inteins; Cann IK et al.; A genetic selection system that detects splicing and nonsplicing activities of inteins was developed based on the ability to rescue a T4 phage strain with a conditionally inactive DNA polymerase . This phage defect can be complemented by expression of plasmid-encoded phage RB69 DNA polymerase . Insertion of an intein gene into the active site of the RB69 DNA polymerase gene renders polymerase activity and phage viability dependent on protein splicing . The effectiveness of the system was tested by screening for thermosensitive splicing mutants . Development of genetic systems with the potential of identifying protein splicing inhibitors is a first step towards controlling proliferation of pathogenic microbes harboring inteins in essential proteins.

Appl Environ Microbiol, 2004 May, 70(5), 3103 - 9
Negative cross-communication among wheat rhizosphere bacteria: effect on antibiotic production by the biological control bacterium Pseudomonas aureofaciens 30-84; Morello JE et al.; Phenazine antibiotic production in the biological control bacterium Pseudomonas aureofaciens 30-84 is regulated in part via the PhzR/PhzI N-acyl homoserine lactone (AHL) system . Previous work showed that a subpopulation of the wheat rhizosphere community positively affected phenazine gene expression in strain 30-84 via AHL signals (E . A . Pierson, D . W . Wood, J . A . Cannon, F . M . Blachere, and L . S . Pierson III, Mol . Plant-Microbe Interact . 11:1078-1084, 1998) . In the present work, a second subpopulation, one that negatively affected phenazine gene expression, was identified from this rhizosphere community . Strain 30-84 grown in conditioned medium (CM) from several strains produced lower levels of phenazines (1.5- to 9.3-fold) than control when grown in CM from the strain 30-84I(1)/I(2) . Growth of the phzB::lacZ reporter strain 30-84Z in this CM resulted in decreased lacZ expression (4.3- to 9.2-fold) compared to growth of the control strain in CM, indicating that inhibition of phzB occurred at the level of gene expression . Preliminary chemical and biological characterizations suggested that these signals, unlike other identified negative signals, were not extractable in ethyl acetate . Introduction of extra copies of phzR and phzI, but not phzI alone, in trans into strain 30-84Z reduced the negative effect on phzB::lacZ expression . The presence of negative-signal-producing strains in a mixture with strain 30-84 reduced strain 30-84's ability to inhibit the take-all disease pathogen in vitro . Together, the results from the previous work on the positive-signal subpopulation and the present work on the negative-signal subpopulation suggest that cross-communication among members of the rhizosphere community and strain 30-84 may control secondary metabolite production and pathogen inhibition.

Appl Environ Microbiol, 2004 May, 70(5), 3082 - 90
Novel forms of structural integration between microbes and a hydrothermal vent gastropod from the Indian Ocean; Goffredi SK et al.; Here we describe novel forms of structural integration between endo- and episymbiotic microbes and an unusual new species of snail from hydrothermal vents in the Indian Ocean . The snail houses a dense population of gamma-proteobacteria within the cells of its greatly enlarged esophageal gland . This tissue setting differs from that of all other vent mollusks, which harbor sulfur-oxidizing endosymbionts in their gills . The significantly reduced digestive tract, the isotopic signatures of the snail tissues, and the presence of internal bacteria suggest a dependence on chemoautotrophy for nutrition . Most notably, this snail is unique in having a dense coat of mineralized scales covering the sides of its foot, a feature seen in no other living metazoan . The scales are coated with iron sulfides (pyrite and greigite) and heavily colonized by epsilon- and delta-proteobacteria, likely participating in mineralization of the sclerites . This novel metazoan-microbial collaboration illustrates the great potential of organismal adaptation in chemically and physically challenging deep-sea environments.

Biochemistry, 2004 May 11, 43(18), 5159 - 69
Structural comparison of MTA phosphorylase and MTA/AdoHcy nucleosidase explains substrate preferences and identifies regions exploitable for inhibitor design; Lee JE et al.; The development of new and effective antiprotozoal drugs has been a difficult challenge because of the close similarity of the metabolic pathways between microbial and mammalian systems . 5'-Methylthioadenosine/S-adenosylhomocysteine (MTA/AdoHcy) nucleosidase is thought to be an ideal target for therapeutic drug design as the enzyme is present in many microbes but not in mammals . MTA/AdoHcy nucleosidase (MTAN) irreversibly depurinates MTA or AdoHcy to form adenine and the corresponding thioribose . The inhibition of MTAN leads to a buildup of toxic byproducts that affect various microbial pathways such as quorum sensing, biological methylation, polyamine biosynthesis, and methionine recycling . The design of nucleosidase-specific inhibitors is complicated by its structural similarity to the human MTA phosphorylase (MTAP) . The crystal structures of human MTAP complexed with formycin A and 5'-methylthiotubercidin have been solved to 2.0 and 2.1 A resolution, respectively . Comparisons of the MTAP and MTAN inhibitor complexes reveal size and electrostatic potential differences in the purine, ribose, and 5'-alkylthio binding sites, which account for the substrate specificity and reactions catalyzed . In addition, the differences between the two enzymes have allowed the identification of exploitable regions that can be targeted for the development of high-affinity nucleosidase-specific inhibitors . Sequence alignments of Escherichia coli MTAN, human MTAP, and plant MTA nucleosidases also reveal potential structural changes to the 5'-alkylthio binding site that account for the substrate preference of plant MTA nucleosidases.

Micron, 2004, 35(6), 431 - 9
Differential-aperture X-ray structural microscopy: a submicron-resolution three-dimensional probe of local microstructure and strain; Yang W et al.; A recently developed differential-aperture X-ray microscopy (DAXM) technique provides local structure and crystallographic orientation with submicron spatial resolution in three-dimensions; it further provides angular precision of approximately 0.01 degrees and local elastic strain with an accuracy of approximately 1.0 x 10(-4) using microbeams from high brilliance third generation synchrotron X-ray sources . DAXM is a powerful tool for inter- and intra-granular studies of lattice distortions and lattice rotations on mesoscopic length scales of tenths of microns to hundreds of microns that are largely above the range of traditional electron microscopy probes . Nondestructive, point-to-point, spatially resolved measurements of local lattice orientations in bulk materials provide direct information on geometrically necessary dislocation density distributions through measurements of the lattice curvature in plastically deformed materials . This paper reviews the DAXM measurement technique and discusses recent demonstrations of DAXM capabilities for measurements of microtexture, local elastic strain, and plastic deformation microstructure.

J Endotoxin Res, 2004, 10(2), 120 - 4
TLR-independent pattern recognition receptors and anti-inflammatory mechanisms; Fraser IP et al.; Pattern recognition receptors recognize molecular patterns associated with the surfaces of microbes and apoptotic cells . These receptors act alone and in concert to bind, phagocytose, and transduce cellular signals derived from these molecular patterns . The outcome of these interactions is dependent on the nature of the ligands, and upon the nature and combination of the ligated receptors . Whereas much attention has been focused on the properties and activities of the Toll-like receptors (TLRs) in this process, many other pattern recognition molecules have been described . Here we review some of these non-TLR receptors and their ligands, and focus attention on the mannose binding lectin, a humoral pattern recognition molecule . In addition, we describe how recognition of apopotic cells via pattern recognition receptors appears to result in responses that differ from those elicited by microbial ligands.

AIDS Res Hum Retroviruses, 2004 Mar, 20(3), 327 - 35
Interaction of mannose-binding lectin with HIV type 1 is sufficient for virus opsonization but not neutralization; Ying H et al.; Mannose-binding lectin (MBL), a microbe-recognition protein in serum, binds to high mannose glycans on HIV-1 gp120 and has been reported to neutralize the cell line-adapted strain HIV(IIIB) . Because HIV primary isolates (PI) are generally more resistant to neutralization by antibodies and considering that PI are produced in primary cells that could alter the number of high mannose glycans on HIV relative to cell lines, we assessed the ability to MBL to neutralize HIV PI . MBL at concentrations up to 50 microg/ml mediated relatively little neutralization (<20%) of HIV PI infection of peripheral blood mononuclear cells (PBMCs) . MBL-neutralizing activity was slightly higher for cell line-adapted HIV infection of the H9 T cell line (up to 64% at 50 microg/ml) . However, this effect was specific for H9 cells since MBL did not neutralize cell line-adapted virus infection of PBMCs, HIV PI infection of the GHOST cell line, or VSV pseudotyped with HIV gp160 from cell line-derived virus or PI . In contrast to its low activity in neutralization assays, MBL efficiently bound infectious HIV PI and opsonized HIV PI for uptake by monocytic cells . These results show that both PI and cell line-adapted HIV, despite binding of MBL, are relatively resistant to neutralization by levels of MBL normally present in serum . However, binding and opsonization of HIV by MBL may alter virus trafficking and viral-antigen presentation during HIV infection.

Clin Chim Acta, 2004 May, 343(1-2), 85 - 92
Differentiation of human umbilical cord blood CD133+ stem cells towards myelo-monocytic lineage; Ruzicka K et al.; BACKGROUND: Characterisation of stem cells by flow cytometry, their expansion and differentiation are presently of major interest for cell engineering as the basis of a therapeutic concept for transplantation . Haematopoietic stem cells (HSC) express CD34, the adhesion structure which binds 2L-selectin, CD117, a receptor for stem cell factor (SCF; c-kit ligand), and CD133, a transmembrane protein belonging to the family of mucoproteins . METHODS: The aim of the present investigation was the systematic investigation of proliferation and differentiation characteristics of umbilical cord blood stem cells (UCBSC) isolated by an immmunomagnetic separation system using CD133 antibody-coated microbeads and to evaluate the effects of different sera and various concentrations, as well as the effects of IL-3 and IL-6 on total cell expansion and differentiation of isolated CD133+ cells . Differentiation patterns were measured by flow cytometry . RESULTS: After the immmunomagnetic separation the yield of CD133+ cells was 0.45+/-0.17 x 10(6) cells/ml; the purity of isolated CD133+ cells was 95.79+/-1.86% . The majority of CD133+ cells coexpressed CD117 . The most pronounced expansion during cultivation of 2 weeks was achieved in media supplemented with 12.5% horse serum plus 12.5% fetal calf serum (FCS) with stem cell factor and interleukine 3; the fold-expansion was 16.67+/-6.20 . During the cultivation period, UCBSC were constantly loosing stem cell markers and differentiated towards myelo-monocyte lineage (granulocytes and/or monocytes) . CONCLUSIONS: These in vitro results demonstrate that thorough investigation of various cultivation conditions is needed for successful expansion and differentiation of stem cells towards different lineages to be used therapeutically for replacement of damaged cells.

Blood, 2004 Aug 15, 104(4), 1066 - 74 Epub 2004 Apr 27.
Tuning the volume of the immune response: strength and persistence of stimulation determine migration and cytokine secretion of dendritic cells; Luft T et al.; Migration to lymph nodes and secretion of cytokines are critical functions of mature dendritic cells (DCs); however, these 2 functions are not necessarily linked . This is the first report showing that quantitative differences in identical signaling pathways determine DC migration and cytokine secretion . Using different polymerized forms of CD40 ligand, we demonstrate that the strength and persistence of CD40 signaling can induce either function . Induction of monocyte-derived DC (MoDC) migration required a weak and transient CD40 signal, whereas strong and persistent CD40 signaling blocked migration and biased toward cytokine secretion . In contrast to MoDCs, CD40 activation of CD1c+ peripheral blood DCs (PBDCs) induced a nonpersistent, intracellular signaling profile resulting in migratory-type DCs unable to secrete interleukin-12p70 (IL-12p70) . Extracellular signal-regulated kinase 1/2 (ERK1/2) and p38K activation synergistically mediated cytokine secretion, whereas migration was enhanced by p38K activation but reduced by persistent ERK1/2 activity . This model of signal strength and persistence also applied when stimulating DCs with intact microbes . Thus, a novel concept emerges in which the type of immune response induced by DCs is tuned by the strength and persistence of DC activating signals.

Anal Biochem, 2004 May 15, 328(2), 113 - 22
Microbead-based electrochemical immunoassay with interdigitated array electrodes; Thomas JH et al.; The objective of this study was to develop a sensitive and miniaturized immunoassay by coupling a microbead-based immunoassay with an interdigitated array (IDA) electrode . An IDA electrode amplifies the signal by recycling an electrochemically redox-reversible molecule . The microfabricated platinum electrodes had 25 pairs of electrodes with 1.6-microm gaps and 2.4-microm widths . An enzyme-labeled sandwich immunoassay on paramagnetic microbeads with mouse IgG as the analyte and beta-galactosidase as the enzyme label was used as the model system . beta-Galactosidase converted p-aminophenyl beta-D-galactopyranoside to p-aminophenol (PAP) . This enzyme reaction was measured continuously by positioning the microbeads near the electrode surface with a magnet . Electrochemical recycling occurred with PAP oxidation to p-quinone imine (PQI) at +290 mV followed by PQI reduction to PAP at -300 mV vs Ag/AgCl . Dual-electrode detection amplified the signal fourfold compared to single-electrode detection, and the recycling efficiency reached 87% . A calibration curve of PAP concentration vs anodic current was linear between 10(-4) and 10(-6)M . A signal from 1000 beads in a 20-microL drop was detectable and the immunoassay was complete within 10 min with a detection limit of 3.5x10(-15)mol mouse IgG.

Diabetes, 2004 May, 53(5), 1285 - 92
From blood monocytes to adipose tissue-resident macrophages: induction of diapedesis by human mature adipocytes; Curat CA et al.; Obesity has been suggested to be a low-grade systemic inflammatory state, therefore we studied the interaction between human adipocytes and monocytes via adipose tissue (AT)-derived capillary endothelium . Cells composing the stroma-vascular fraction (SVF) of human ATs were characterized by fluorescence-activated cell sorter (FACS) analysis and two cell subsets (resident macrophages and endothelial cells {ECs}) were isolated using antibody-coupled microbeads . Media conditioned by mature adipocytes maintained in fibrin gels were applied to AT-derived ECs . Thereafter, the expression of endothelial adhesion molecules was analyzed as well as the adhesion and transmigration of human monocytes . FACS analysis showed that 11% of the SVF is composed of CD14(+)/CD31(+) cells, characterized as resident macrophages . A positive correlation was found between the BMI and the percentage of resident macrophages, suggesting that fat tissue growth is associated with a recruitment of blood monocytes . Incubation of AT-derived ECs with adipocyte-conditioned medium resulted in the upregulation of EC adhesion molecules and the increased chemotaxis of blood monocytes, an effect mimicked by recombinant human leptin . These results indicate that adipokines, such as leptin, activate ECs, leading to an enhanced diapedesis of blood monocytes, and suggesting that fat mass growth might be linked to inflammatory processes.

Vet Clin North Am Small Anim Pract, 2004 May, 34(3), 611 - 22
Matrix metalloproteinase inhibition in corneal ulceration; Brooks DE et al.; The primary objective of current treatment strategies for infectious keratitis is to sterilize the ulcer as rapidly as possible with topically administered antibiotics . Ulcerative processes can proceed in some cases, despite the absence of microbes, as a result of remaining corneal and tear film MMPs . Combining antibiotic therapy with MMP inhibitors can speed corneal healing, because MMPs play an important role in corneal ulceration and stromal liquefaction . MMPs from the rabbit, horse, and human being are inhibited by metal-binding agents EDTA, NAC, and doxycycline as well as by the serum antiprotease alpha2-macroglobulin . It is not yet certain which proteinase inhibitor has the most favorable therapeutic index for clinical use, although we prefer serum because of its effects on multiple types of proteinases . The MMP inhibitors do have significant therapeutic promise in the treatment of corneal ulceration.

Cell Biol Int, 2004, 28(4), 293 - 8
Backward chromosome movement in crane-fly spermatocytes after UV microbeam irradiation of the interzone and a kinetochore; Wong R et al.; Single anaphase chromosomes (in crane-fly spermatocytes) moved backwards after double irradiations with an ultraviolet light (UV) microbeam, first of the interzone and then of a kinetochore: the chromosome irradiated at the kinetochore moved backwards rapidly, across the equator and into the other half-spindle . High irradiation doses at the kinetochore were required to induce backward movement . Single irradiations of kinetochores or interzones were ineffective in inducing backward movements.

Biomaterials, 2004 Sep, 25(21), 5155 - 62
Biodegradability of poly (2-hydroxyethyl methacrylate) in the presence of the J774.2 macrophage cell line; Mabilleau G et al.; The degradation of cross-linked and linear poly(2-hydroxyethyl methacrylate) (pHEMA), was examined in vitro with J774.2 cells . pHEMA microbeads were prepared with both types of polymers . Only cells in contact with the microbeads increased their production of lysosomal enzymes (TRAcP and ANAE) and released large amounts of reactive oxygen species with both types of pHEMA microbeads . Electron microscopy showed that macrophages were able to erode the surface of linear pHEMA but unable to erode the surface of the cross-linked polymer . Cells appeared wrapped by the linear pHEMA surface, but those cultured on the cross-linked polymer were only laying at the surface . After cell culture, the surface roughness of pHEMA slices was observed by atomic force microscopy (AFM) . There was a significant increase in roughness (R(a)) of the surface of linear pHEMA slices cultured with J774.2 cells whereas no difference in R(a) between the surface of cross-linked pHEMA slices could be measured . AFM image of the hydrated materials were done: the surface of linear pHEMA swelled considerably in saline whereas the hydrated cross-linked polymer did not differ from the air-dried appearance . In conclusion, linear pHEMA swells in biological fluids, activates macrophages in close contact with the polymer and can be progressively eroded.

Anal Bioanal Chem, 2004 Jun, 379(3), 419 - 26 Epub 2004 Apr 23.
Biomagnetic separation of Escherichia coli by use of anion-exchange beads: measurement and modeling of the kinetics of cell-bead interactions; Deponte S et al.; Analyses of food-borne pathogens are of great importance in order to minimize the risk of infection for customers . These analyses should be as fast as possible . Any detection method requires enrichment and quantitative analysis of the enriched microbes . Conventional enrichment methods, which take several days, need to be replaced by faster techniques such as biomagnetic separation (BMS) . This technique involves the use of paramagnetic microspheres coated with ligands that have special affinity to the microbes that have to be detected . In the studies reported here, enrichment experiments by BMS were carried out using the non-pathogenic E . coli DSM 498 as a model strain and beads coated with a polyethylenimine (PEI) anion-exchange material . The results show that the number of cells separated, as a proportion of the total, was positively correlated with the bead concentration and the length of the period they were mixed together . In addition, a mathematical model, based on the rate of impact between two different sorts of particles, was developed to describe the proportion of separated cells as a function of incubation time and the concentration, size and density of the beads and cells . This is the first mathematical description of cell-bead interactions to be based on well-understood physicochemical principles . The model was confirmed by separation experiments in which the concentration of beads and the incubation period were varied . The developed model enables optimization of the amount of beads added and the reaction period necessary for complete cell separation and thus minimization of costs in BMS.

J Immunol, 2004 May 1, 172(9), 5304 - 12
CD1d1 displayed on cell size beads identifies and enriches an NK cell population negatively regulated by CD1d1; Huang MM et al.; NK cells destroy microbe-infected cells while sparing healthy cells, and are controlled, in part, by inhibitory receptors specific for class I Ag-presenting molecules . CD1d1, a beta(2)-microglobulin-associated class I-like molecule, binds glycolipids and stimulates NKT cells . We previously demonstrated that target cell lysis by IL-2-activated mouse NK cells is inhibited by target cell expression of CD1d1, suggesting that IL-2-activated NK cells may express a CD1d1-specific inhibitory receptor . We now report that a significant subset of mouse IL-2-activated NK cells specifically binds cell size beads displaying either naturally expressed or recombinant CD1d1 . In contrast, although tetramers of soluble recombinant CD1d1 loaded with alpha-galactosylceramide identify NKT cells, binding of this reagent to resting or IL-2-activated NK cells was undetectable, even with activated NK cells sorted with CD1d1 beads . Cytotoxicity by the CD1d1 bead-separated NK subset was strongly inhibited by CD1d1, compared with the NK cell subset not bound to CD1d1 beads . An Ab that blocks NKT cell recognition of CD1d1 also reverses CD1d1 inhibition of NK lysis, suggesting that TCRs of NKT cells and NK inhibitory receptor(s) may interact with a similar site on CD1d1 . These results provide direct evidence for a physical interaction of NK cells with CD1d1, mediated by a functional, CD1d1-specific low-affinity inhibitory NK receptor . Display of ligands on cell size beads to maximize multivalent interaction may offer an alternative approach to examine NK cell receptor-ligand interactions, particularly those of lower expression and/or lower affinity/avidity that may go undetected using tetrameric reagents.

Plant Biol (Stuttg), 2004 Jan-Feb, 6(1), 91 - 9
Compatible and incompetent Paxillus involutus isolates for ectomycorrhiza formation in vitro with poplar (Populus x canescens) differ in H2O2 production; Gafur A et al.; Isolates of Paxillus involutus (Batsch) Fr . collected from different hosts and environmental conditions were screened for their ability to form ectomycorrhizal symbiosis with hybrid poplar P . x canescens (= Populus tremula L . x P . alba) in vitro . The ability to form ectomycorrhiza varied between the fungal isolates and was not correlated with the growth rate of the fungi on agar-based medium . The isolate MAJ, which was capable of mycorrhiza synthesis under axenic conditions, and the incompetent isolate NAU were characterized morphologically and anatomically . MAJ formed a typical hyphal mantle and a Hartig net, whereas NAU was not able to penetrate the host cell walls and caused thickenings of the outer cell walls of the host . MAJ, but not NAU, displayed strong H2O2 accumulation in the outer hyphal mantle . Increases in H2O2 in the outer epidermal walls and adjacent hyphae of the incompetent isolate were moderate . No increases of H2O2 in response to the mycobionts were found inside roots . Suggested functions of H2O2 production in the outer hyphal mantle of the compatible interaction are: growth regulation of the host's roots, defence against other invading microbes, or increasing plant-innate immunity . The system established here for P . x canescens compatible and incompetent fungal associations will be useful to take advantage of genomic information now available for poplar to study tree-fungal interactions at the molecular and physiological level.

PLoS Biol . 2004 Apr;2(4):E81 . Epub 2004 Apr 13.
Segmentally variable genes: a new perspective on adaptation; Zheng Y et al.; Genomic sequence variation is the hallmark of life and is key to understanding diversity and adaptation among the numerous microorganisms on earth . Analysis of the sequenced microbial genomes suggests that genes are evolving at many different rates . We have attempted to derive a new classification of genes into three broad categories: lineage-specific genes that evolve rapidly and appear unique to individual species or strains; highly conserved genes that frequently perform housekeeping functions; and partially variable genes that contain highly variable regions, at least 70 amino acids long, interspersed among well-conserved regions . The latter we term segmentally variable genes (SVGs), and we suggest that they are especially interesting targets for biochemical studies . Among these genes are ones necessary to deal with the environment, including genes involved in host-pathogen interactions, defense mechanisms, and intracellular responses to internal and environmental changes . For the most part, the detailed function of these variable regions remains unknown . We propose that they are likely to perform important binding functions responsible for protein-protein, protein-nucleic acid, or protein-small molecule interactions . Discerning their function and identifying their binding partners may offer biologists new insights into the basic mechanisms of adaptation, context-dependent evolution, and the interaction between microbes and their environment.

Clin Immunol, 2004 Apr, 111(1), 1 - 15
Vaccines for the prevention of diseases caused by potential bioweapons; Hassani M et al.; The development of vaccines and implementation of vaccination programs are among the most important medical contributions to humanity . To date, vaccination has reduced morbidity and mortality from infectious diseases more than any other specific medical intervention . The intentional use of bioweapons against civilians (bioterrorism), recently highlighted by events around the world, has fueled interest in the development of vaccines for potential microbial agents of bioterror . This review discusses the microbial agents that are considered to pose the greatest risk to the public, the diseases associated with them, and the vaccines that are available for their prevention . The paucity of such vaccines and uncertainty regarding mechanisms of vaccine efficacy and the microbial antigens that elicit protection underscore the need for continued study of host-microbe interaction and the immune response to potential agents of bioterror for the development of new vaccines and immune-based therapies to combat their potential to harm the public.

Environ Pollut, 1997, 97(1-2), 45 - 53
Decomposer communities in contaminated soil: Is altered community regulation a proper tool in ecological risk assessment of toxicants?
Salminen JE, Sulkava PO.
Effects of patchy soil contamination on decomposer organisms, their community regulation and nutrient mineralization were studied in a microcosm experiment . Coniferous forest soil was patchily contaminated with three concentrations of sodiumpentachlorophenate PCP (0, 50 and 500 mg PCP kg(-1) of dry soil) . Abundance of microbes, enchytraeids, nematodes, small oribatids and predatory mites were reduced by the PCP . Direct toxicity of PCP and lowered microbial biomass seemed to affect animal community composition in the most contaminated patches . Some large oribatids which seemed to be tolerant to PCP increased their numbers in the most contaminated patches . Although predatory mites suffered from PCP, no altered predator-prey interactions were observed . At the beginning of the experiment more nutrients were released in the patches with highest PCP concentration and the nutrients accumulated in the soil . Soil decomposer food webs seemed to be mainly bottom-up controlled: PCP strongly affects microbes and hence caused changes in the community structure of soil animals and nutrient cycling . Hence top-down orientated ecological models on community regulation and food web dynamics seem to be unsuitable when assessing effects of pesticides on soil communities.

Environ Pollut, 1997, 97(1-2), 39 - 44
Influence of landfill factors on plants and soil fauna-An ecological perspective; Chan YS et al.; An ecological study was conducted on two landfill sites in Hong Kong . Both sites were contaminated by landfill gas and leachate, as indicated by the high concentrations of CH(4) and CO(2) and the presence of high concentration of NH(4)-N in the cover soil layer . Their plant diversity and performance, as well as the soil and litter animals, were compared with three derelict sites as references . The landfill sites had higher plant coverage and plant diversity, and higher densities of soil and litter animals than the reference sites . Similarly, the microbial activities at the landfill sites were higher than those at reference sites . The results showed that the landfill sites possessed an effective food web, starting from microbes to macroinvertebrates . Woodland establishment seems feasible at these landfill sites, and natural succession appears to take place at a higher rate at these landfill sites when compared with the reference sites.

Environ Pollut, 1994, 86(2), 181 - 8
Effects of mycorrhizae and other soil microbes on revegetation of heavy metal contaminated mine spoil; Shetty KG et al.; The effects of mycorrhizal fungi and other soil microorganisms on growth of two grasses, Andropogon gerardii Vitm . and Festuca arundinacea Schreb., in heavy metal-contaminated soil and mine tailings were investigated . A . gerardii is highly dependent on mycorrhizal fungi in native prairie, while F . arundinacea is a facultative mycotroph and relies on mycorrhizal symbiosis only in extremely infertile soils . Regardless of microbial amendments, neither plant species was able to establish and grow in the mine tailings . Both plant species grew in the moderately contaminated or non-contaminated soils, although A . gerardii grew in these soils only when mycorrhizal . Other soil microbes significantly improved growth of A . gerardii only in uncontaminated soil, but to a lesser extent than mycorrhizae . Although F . arundinacea was more highly colonized by mycorrhizal fungi than A . gerardii, neither microbial amendment affected growth of fescue in any soil . In several treatments mycorrhizal fungi adapted to uncontaminated soil stimulated plant growth more than mycorrhizae adapted to the moderately contaminated soil . However, mycorrhizal fungi adapted to contaminated soil did not increase the productivity of plant growth in contaminated soil more than fungi adapted to uncontaminated soil . A . gerardii plants inoculated with mycorrhizal fungi retained more Zn in roots than in shoots, confirming earlier reports that mycorrhizal fungi alter the translocation pattern of heavy metals in host plants . In contrast, mycorrhizae did not affect translocation patterns in F . arundinaceae, suggesting that the mycorrhizal dependence of a plant species is correlated with the retention of metals in roots . The correlation between mycorrhizal dependence of a plant species and mycorrhizal alteration of translocation pattern may also explain the inconsistent reports of mycorrhizal effects on translocation of heavy metals in plants . Plant response to mycorrhizal symbiosis may therefore provide a useful criterion for the selection of the plant species to be used in revegetation of contaminated sites.

Phys Rev E Stat Nonlin Soft Matter Phys . 2004 Mar;69(3 Pt 1):031113 . Epub 2004 Mar 31.
Fluctuation-response relation in a rocking ratchet; Harada T et al.; The relationship between the positional fluctuation and mechanical response for a thermal ratchet at a nonequilibrium steady state was investigated experimentally and considered theoretically . We constructed a ratchet system using a scanning optical trapping system and microbeads (0.20 microm diameter), which is interpreted as a realization of a rocking ratchet {M . O . Magnasco, Phys . Rev . Lett . 71, 1477 (1993)} . In the experiment, an asymmetric periodic potential (2.5 microm period) was generated by optical trap scanning, which traps a bead . When the potential profile was rocked sinusoidally, diffusion of the bead was rectified in one direction . We confirmed that both the diffusion coefficient and the mobility increased with potential rocking with a positive correlation . To obtain better insight, we performed numerical and theoretical analyses of the corresponding Langevin system . Although there is a positive relationship between the diffusion coefficient and mobility, the diffusion coefficient is greater than the value given in the Einstein relation . This result means that the effective temperature of the thermal ratchet at a nonequilibrium steady state becomes greater than that of the environment . We propose that this elevation of the effective temperature causes a decrease in the energetic efficiency of the thermal ratchet through irreversible dissipation to the heat bath.

J Periodontol, 2004 Mar, 75(3), 353 - 9
Comparison of probes for microbial contamination following use in periodontal pockets of various depths; Holt LA et al.; BACKGROUND: It has been speculated that periodontal probes can transmit periodontal pathogens from site to site . The purpose of this study was to evaluate the potential for bacterial transmission as a function of periodontal probe design . METHODS: Four different periodontal probes were used to measure probing depths ranging from 0 to 3 mm and > or = 4 mm . Following measurement of each pocket, probes were transported to a laboratory and cultured on blood agar, and colony forming units (CFUs) were determined for total aerobic, anaerobic, and dark-pigmented colonies (DPCs) . Eight randomly selected probe tips, representing each probe type and probing depth category, were selected for examination by scanning electron microscopy (SEM) to determine distribution and morphotypes of adhering microbes . RESULTS: Results showed no statistically significant main effect for probes with respect to CFUs . However, there was a statistically significant increase in CFUs for deeper pockets when compared to pockets 0 to 3 mm . SEM observations were consistent among groups, regardless of probe design or probing depth . All probes exhibited a rough surface texture . Microbes were observed as single organisms or in aggregates and were adherent along the entire length of the probe up to the maximum probing depth . CONCLUSIONS: There was no difference in CFUs obtained as a function of periodontal probes . The rough surfaces of the probes used in this study likely promoted bacterial adherence through two different mechanisms: through scraping of the pocket walls by the roughened surfaces of the probe and by the high surface-free energy of the metal probes, facilitating bacterial adherence.

Biotechniques, 2004 Apr, 36(4), 602 - 6, 608-9
Incorporating fluorescent dyes and quantum dots into magnetic microbeads for immunoassays; Mulvaney SP et al.; Microbeads that are both paramagnetic and fluorescently labeled are commercially available in colors spanning the visible spectrum . Although these commercial beads can be bright, polydispersity in both size and fluorescent intensity limit their use in quantitative assays . Very recently, more monodisperse beads have become available, but their large size and surface properties make them less than ideal for some bioassay applications . Here we describe methods to customize commercial nonfluorescent magnetic microparticles with fluorescent dyes and quantum dots (QDs) without affecting their magnetic or surface chemical properties . Fluorescent dyes and 3.3-nm diameter CdSe/ZnS QDs were sequestered within 0.8-micron diameter magnetic beads by swelling the polystyrene matrix of the bead in organic solvent, letting the chromophores partition, and then collapsing the matrix in polar solvents . Chromophore incorporation has been characterized using both UV-visible absorption spectroscopy and fluorescence microscopy, with an average of 3 x 10(8) rhodamine 6G molecules/bead and 6 x 10(4) QDs/bead . The modified beads are uniform in size and intensity, with optical properties comparable to currently available commercial beads . Immunoassay results obtained with our custom fluorescent magnetic microbeads are consistent with those obtained using conventional magnetic microbeads.

J Pediatr Gastroenterol Nutr, 2004 Apr, 38(4), 378 - 88
The hygiene hypothesis of atopic disease--an extended version; Rautava S et al.; The hygiene hypothesis of atopic disease suggests that environmental changes in the industrialized world have lead to reduced microbial contact at an early age and thus resulted in the growing epidemic of atopic eczema, allergic rhinoconjunctivitis, and asthma . The epidemiological findings have been combined with the Th1/Th2 paradigm of immune responsiveness to provide a coherent theory . Recent advances in epidemiology and immunology demonstrate, however, that the hygiene hypothesis may need to be extended in three respects . First, the importance of infections in causing immune deviance may be outweighed by other sources of microbial stimulation, perhaps most importantly by the indigenous intestinal microbiota . Second, immunomodulatory and suppressive immune responses complement the Th1/Th2 paradigm . Third, in addition to protection against atopy, protection against infectious, inflammatory, and autoimmune diseases may also depend upon healthy host-microbe interactions implicated in the hygiene hypothesis.

J Biol Chem, 2004 Jun 18, 279(25), 26605 - 11 Epub 2004 Apr 14.
Innate immunity in a pyralid moth: functional evaluation of domains from a beta-1,3-glucan recognition protein; Fabrick JA et al.; Invertebrates, like vertebrates, utilize pattern recognition proteins for detection of microbes and subsequent activation of innate immune responses . We report structural and functional properties of two domains from a beta-1,3-glucan recognition protein present in the hemolymph of a pyralid moth, Plodia interpunctella . A recombinant protein corresponding to the first 181 amino-terminal residues bound to beta-1,3-glucan, lipopolysaccharide, and lipoteichoic acid, polysaccharides found on cell surfaces of microorganisms, and also activated the prophenoloxidase-activating system, an immune response pathway in insects . The amino-terminal domain consists primarily of an alpha-helical secondary structure with a minor beta-structure . This domain was thermally stable and resisted proteolytic degradation . The 290 residue carboxyl-terminal domain, which is similar in sequence to glucanases, had less affinity for the polysaccharides, did not activate the prophenoloxidase cascade, had a more complicated CD spectrum, and was heat-labile and susceptible to proteinase digestion . The carboxyl-terminal domain bound to laminarin, a beta-1,3-glucan with beta-1,6 branches, but not to curdlan, a beta-1,3-glucan that lacks branching . These results indicate that the two domains of Plodia beta-1,3-glucan recognition protein, separated by a putative linker region, bind microbial polysaccharides with differing specificities and that the amino-terminal domain, which is unique to this class of pattern recognition receptors from invertebrates, is responsible for stimulating prophenoloxidase activation.

J Anim Sci, 2004 Apr, 82(4), 1146 - 56
Site and extent of digestion and amino acid flow to the small intestine in beef cattle consuming limited amounts of forage; Scholljegerdes EJ et al.; Eight Angus x Gelbvieh heifers (445 +/- 74.5 kg) fitted with ruminal and duodenal cannulas were used in a 4 x 4 Latin square double double-crossover designed experiment to assess the effect of restricted forage intake on site and extent of digestion and flow of essential AA amino acids to the small intestine . Heifers were fed chopped (2.54 cm) bromegrass hay (9.2% CP, 64% NDF on an OM basis) at one of four percentages of maintenance (30, 60, 90, and 120%) . Experimental periods were 21 d in length, with 17 d of adaptation followed by 4 d of intensive sample collection, after which maintenance requirements and subsequent level of intake were adjusted for BW change . True ruminal OM, NDF, and N digestion (g/d) decreased linearly (P < 0.001) with decreasing forage intake . When expressed as a percentage of OM intake, true ruminal OM and N digestibility were not affected (P = 0.23 to 0.87), whereas ruminal NDF digestibility tended to increase (P = 0.09) as forage intake decreased . Total and microbial essential amino acid flow to the duodenum decreased linearly (P = 0.001) from 496.1 to 132.1 g/d and 329.1 to 96.0 g/d, as intake decreased from 120 to 30% of maintenance intake, respectively . Although the profile of individual essential amino acids in duodenal digesta (P = 0.001 to 0.07) and isolated ruminal microbes differed (P = 0.001 to 0.09) across treatment, the greatest difference noted for total and microbial essential amino acid profile was only 0.3 percentage units . Because total and microbial flow of essential amino acids to the small intestine decreased as OM intake decreased, but true ruminal degradability of individual essential amino acids (P = 0.17 to 0.99) and digesta essential amino acid profile were comparable across treatments, total essential amino acid supply to the small intestine was predicted using OM intake as the independent variable . The resulting simple linear regression equation was: total essential amino acid flow = (0.055 x OM intake) + 1.546 (r2 = 0.91) . The model developed in this experiment accounted for more of the variation in the data set than the current beef cattle NRC model, which under-predicted total flow of essential amino acids to the duodenum . The prediction equation developed herein can be used to estimate the supply of essential amino acids reaching the small intestine when formulating supplements to compensate for potential amino acid deficiencies resulting from restricted forage intake.

Sci Prog, 2003, 86(Pt 3), 179 - 202
Microbial transformation of metals and metalloids; Raab A et al.; Throughout evolution, microbes have developed the ability to live in nearly every environmental condition on earth . They can grow with or without oxygen or light . Microbes can dissolve or precipitate ores and are able to yield energy from the reduction/oxidation of metal ions . Their metabolism depends on the availability of metal ions in essential amounts and protects itself from toxic amounts of metals by detoxification processes . Metals are metabolised to metallorgano-compounds, bound to proteins or used as catalytic centres of enzymes in biological reactions . Microbes, as every other cell, have developed a whole range of mechanisms for the uptake and excretion of metals and their metabolised compounds . The diversity of microbial metabolism can be illustrated by the fact that certain microbes can be found living on arsenate, which is considered a highly toxic metal for most other forms of live.

Curr HIV Res, 2004 Apr, 2(2), 99 - 111
Can HIV be Cured? Mechanisms of HIV persistence and strategies to combat it; Hamer DH; Stable remission is the ultimate goal of HIV therapy . A review of recent studies on the ability of HIV to persist despite highly active antiretroviral therapy (HAART) and immune stimulation suggests that achieving this goal will require four developments in basic and clinical science . First, more effective antiretroviral therapies, targeted at proteins other than reverse transcriptase and protease, in order to eliminate the cryptic replication that continues despite best available HAART . Second, agents that activate latent HIV gene expression in quiescent CD4 memory T cells, thereby exposing this viral reservoir to therapeutic intervention by a "shock and kill" strategy . Third, molecules such as immunotoxins that specifically recognize HIV-encoded membrane proteins and thereby potentiate the destruction of infected cells . Fourth, and still most distant, novel approaches such as genetically engineered cytotoxic T lymphocytes or anti-HIV microbes to suppress rekindling of infection by residual virus sequestered in anatomical and cellular reservoirs . Although each of these steps will be difficult to achieve, the many benefits of a cure for HIV make this a worthwhile pursuit.

Opt Lett, 2004 Apr 1, 29(7), 736 - 8
Spatially incoherent illumination as a mechanism for cross-talk suppression in wide-field optical coherence tomography; Karamata B et al.; Comparison of two illumination modes for wide-field optical coherence tomography has revealed that spatially coherent illumination generates coherent cross talk, causing significant image degradation, and that spatially incoherent illumination, with an adequate interferometer design, provides an efficient mechanism for suppression of coherent cross talk . This is shown by comparison of a pulsed laser with a thermal light source for a U.S . Air Force resolution target covered with a scattering solution made from microbeads as well as for an ex vivo tooth.

Hum Exp Toxicol, 2004 Feb, 23(2), 81 - 6
A theoretical approach to the role and critical issues associated with bystander effect in risk estimation; Nikjoo H et al.; This paper presents a quantitative biophysical model of the radiation-induced bystander effect . The principle aim of the bystander model is to establish whether bystander signal can be associated with low molecular weight factors that are transmitted by diffusion type processes in the medium surrounding the recipient cells . Cell inactivation and induced oncogenic transformation by microbeam and broadbeam irradiation systems were considered . The biophysical model postulates that the oncogenic bystander response observed in non-hit cells originates from specific signals received from inactivated cells . The bystander signals are assumed to be protein-like molecules spreading in the culture media by Brownian motion . The bystander signals are assumed to switch cells into a state of cell death (apoptotic/mitotic/necrosis) or induced oncogenic transformation modes . The bystander cell survival observed after treatment with the irradiated conditioned medium using broadbeam and the microbeam irradiation modalities were analysed and interpreted in the framework of the Bystander Diffusion Model (BSDM) . The model predictions for cell inactivation and induced oncogenic transformation frequencies agree well with observed data from microbeam and broadbeam experiments . In the case of irradiation with constant fraction of cells, transformation frequency for the bystander effect increases with increasing radiation dose . The BSDM predicts that the bystander effect cannot be interpreted solely as a low-dose effect phenomenon . It is shown that the bystander component of radiation response can increase with dose and can be observed at high doses as well as low doses . The validity of this conclusion is supported by analysis of experimental results from high-LET microbeam experiments.

J Pediatr, 2004 Apr, 144(4), 421 - 9
Innate immunity and toll-like receptors: clinical implications of basic science research; Abreu MT et al.; Humans are constantly exposed to a wide variety of microorganisms that can cause infection . In self-defense, the human host has evolved complex protective mechanisms, and Toll-like receptors (TLRs) have emerged as a central point in defense . These receptors bind molecular structures that are expressed by microbes but are not expressed by the human host, eg, lipopolysaccharides (LPS) or double-stranded RNA (dsRNA) . Activation of these receptors initiates an inflammatory cascade that attempts to clear the offending pathogen and set in motion a specific adaptive immune response . Defects in sensing of pathogens may predispose the host to recurrent infections . The relative rarity of these syndromes of defective innate immunity, however, speaks to the redundancy in sensing of pathogens by the innate immune system . More common, polymorphisms in TLR4 are associated with increased predisposition to severe and recurrent infections but protection against atherosclerotic disease due to diminished inflammation . Toll-like receptor signaling may also contribute to the pathophysiology of disease and injure the host by activating a deleterious immune response such as in sepsis or inflammatory bowel disease (IBD) . The focus of this article is to describe the role of TLRs in the innate immune response in health and disease.

J Eukaryot Microbiol, 2004 Jan-Feb, 51(1), 66 - 72
Identification and sequence analysis of six new members of the NIMA-related kinase family in Chlamydomonas; Bradley BA et al.; The NIMA kinases are an evolutionarily conserved protein family with enigmatic roles in the regulation of mitosis . We report six new members of this family in Chlamydomonas, in addition to the previously identified NIMA-related kinase, Fa2p . Chlamydomonas NIMA-related kinases (CNKs) 1-6 were sequenced from subclones generated by RT-PCR using information from EST libraries and the recently sequenced Chlamydomonas genome . Phylogenetic and bioinformatic approaches were used to determine the relationships of the six new members with known members of the NIMA-related kinase family . Although humans express at least eleven NIMA-related kinases, the eukaryotic microbes that have been studied to date express only one or two members of the family . Thus, the discovery that Chlamydomonas expresses a total of at least seven NIMA-related kinases is intriguing . Our analyses suggest that members of this family may play roles in the assembly and function of cilia.

Int J Mol Med, 2004 May, 13(5), 685 - 90
Detection of germ-cell-tumor-specific gene products in peripheral blood by immunomagnetic tumor cell enrichment followed by RT-PCR; Schrader M et al.; Therapeutic procedures in patients with testicular germ cell tumors (GCT) are determined by the histopathology of the primary tumor and the tumor extension . The aim of our study was to determine whether conventional staging could be supplemented by combining enrichment of disseminated testicular GCT cells from peripheral blood with subsequent detection of germ-cell-specific gene products . Blood samples from 46 patients with GCT of different clinical stages (CS) were examined by RT-PCR before therapy and >/=8 weeks thereafter for alpha-fetoprotein, beta-human chorionic gonadotropin and germ-cell-specific alkaline phosphatase mRNA . In addition, we performed titration experiments to evaluate whether the sensitivity can be improved by previous immunomagnetic tumor cell enrichment with anti-epithelial HEA-125 microbeads . No positive results were found in controls (n=15; specificity 100%) . The overall ratio of positive PCR results in the group of patients with GCTs was 28.26% . The ratio was 35.7% for CS >IIb (n=5/14 patients), 20.0% for CS IIa-b (n=4/20) and 33.3% for CS I (n=4/12) . FACS analysis in titration experiments with GCT cell lines showed that previous immunomagnetic tumor cell enrichment achieved a significant increase ranging up to 185.6 times the initial ratio and thus improved the measuring conditions for detection of tumor-specific transcripts . The sole qualitative RT-PCR of tumor-specific gene products in peripheral blood is not sensitive enough to improve staging in GCT patients . Immunomagnetic enrichment of GCT cells in peripheral blood seems a promising approach for increasing the sensitivity of RT-PCR.

Appl Environ Microbiol, 2004 Apr, 70(4), 2540 - 4
Comparisons of protocols for decontamination of environmental ice samples for biological and molecular examinations; Rogers SO et al.; Drilling and laboratory manipulations of glacial ice cores introduce contemporary microbes and biomolecules onto the cores . We report herein a systematic comparative study of several decontamination protocols . Only treatment with 5% sodium hypochlorite eliminated all external contaminating microbes and nucleic acids while maintaining the integrity of those within the cores.

Appl Environ Microbiol, 2004 Apr, 70(4), 2263 - 70
GC fractionation enhances microbial community diversity assessment and detection of minority populations of bacteria by denaturing gradient gel electrophoresis; Holben WE et al.; Effectively and accurately assessing total microbial community diversity is one of the primary challenges in modern microbial ecology . This is particularly true with regard to the detection and characterization of unculturable populations and those present only in low abundance . We report a novel strategy, GC fractionation combined with denaturing gradient gel electrophoresis (GC-DGGE), which combines mechanistically different community analysis approaches to enhance assessment of microbial community diversity and detection of minority populations of microbes . This approach employs GC fractionation as an initial step to reduce the complexity of the community in each fraction . This reduced complexity facilitates subsequent detection of diversity in individual fractions . DGGE analysis of individual fractions revealed bands that were undetected or only poorly represented when total bacterial community DNA was analyzed . Also, directed cloning and sequencing of individual bands from DGGE lanes corresponding to individual G+C fractions allowed detection of numerous phylotypes that were not recovered using a traditional random cloning and sequencing approach.

J Biotechnol, 2004 Apr 29, 109(3), 217 - 26
Preparation of single rice chromosome for construction of a DNA library using a laser microbeam trap; Liu X et al.; We report the development of a laser micromanipulation system and its application in the isolation of individual rice chromosomes directly from a metaphase cell . Microdissection and flow sorting are two major methods for the isolation of single chromosome . These methods are dependent on the techniques of chromosome spread and chromosome suspension, respectively . In the development of this system, we avoided using chromosome spread and cell suspension was used instead . The cell wall of metaphase rice cell was cut by optical scissors . The released single chromosome was captured by an optical trap and transported to an area without cell debris . The isolated single chromosome was then collected and specific library was constructed by linker adaptor PCR . The average insert size of the library was about 300 bp . Two hundred inserts of chromosome 4 library were sequenced, and 96.5% were aligned to the corresponding sequences of rice chromosome 4 . These results suggest the possible application of this method for the preparation of other subcellular structures and for the cloning of single macromolecule through a laser microbeam trap.

J Mol Biol, 2004 Apr 23, 338(2), 329 - 39
The crystal structure of three site-directed mutants of Escherichia coli dihydrodipicolinate synthase: further evidence for a catalytic triad; Dobson RC et al.; Dihydrodipicolinate synthase (DHDPS, EC 4.2.1.52) catalyses the branchpoint reaction of lysine biosynthesis in plants and microbes: the condensation of (S)-aspartate-beta-semialdehyde and pyruvate . The crystal structure of wild-type DHDPS has been published to 2.5A, revealing a tetrameric molecule comprised of four identical (beta/alpha)(8)-barrels, each containing one active site . Previous workers have hypothesised that the catalytic mechanism of the enzyme involves a catalytic triad of amino acid residues, Tyr133, Thr44 and Tyr107, which provide a proton shuttle to transport protons from the active site to solvent . We have tested this hypothesis using site-directed mutagenesis to produce three mutant enzymes: DHDPS-Y133F, DHDPS-T44V and DHDPS-Y107F . Each of these mutants has substantially reduced activity, consistent with the catalytic triad hypothesis . We have determined each mutant crystal structure to at least 2.35A resolution and compared the structures to the wild-type enzyme . All mutant enzymes crystallised in the same space group as the wild-type form and only minor differences in structure are observed . These results suggest that the catalytic triad is indeed in operation in wild-type DHDPS.

Microbes Infect, 2004 Mar, 6(3), 312 - 8
Survival strategies of Borrelia burgdorferi, the etiologic agent of Lyme disease; Embers ME et al.; To fight, flee or hide are the imperatives of long-term survival by an infectious microbe . Active immune suppression, induction of immune tolerance, phase and antigenic variation, intracellular seclusion, and incursion into immune privileged sites are examples of survival strategies of persistent pathogens . Here we critically review the supporting evidence for possible stratagems utilized by Borrelia burgdorferi, the spirochete that causes Lyme disease, to persist in the mammalian host.

Adv Drug Deliv Rev, 2004 Apr 19, 56(6), 727 - 62
Microbes and their products--physiological effects upon mammalian mucosa; Viswanathan VK et al.; A dynamic array of interactions occurs between pathogens and host mucosal surfaces . The signature molecules unique to microbial pathogens allow the mammalian immune system to recognize them as non-self . This recognition, mediated by the toll-like receptor proteins, results in innate immune responses targeted against the invading organism . Pathogens also elaborate a variety of proteins that actively engage host signaling pathways and subvert them to facilitate their growth and dispersal . These interactions, developed over a long evolutionary period, have been specialized to exquisite detail . These proteins and toxins are either secreted into the medium or directly delivered into host cells by specialized secretion systems . An array of host function alterations is mediated by microbial pathogens including inflammatory responses, secretory responses, alteration of host cytoskeleton, disruption of epithelial tight junctions and apoptosis . The signaling axes involved in these interactions are potential targets for therapeutic strategies against infectious microbes.

Mol Genet Metab, 2004 Apr, 81(4), 313 - 21
NADPH oxidase activity of neutrophil specific granules: requirements for cytosolic components and evidence of assembly during cell activation; Ambruso DR et al.; Neutrophils and other phagocytic cells support host defense by ingesting microbes and destroying them with reactive oxygen species or oxygen independent mechanisms . Production of ROS is initiated by the phagocyte NADPH oxidase (phox), an enzyme system composed of several constituents . During activation of the cell cytosolic phox proteins (p47phox, p67phox, p40phox, and Rac2) translocate to the plasma membrane and specific granules fuse with the plasma membrane increasing the amount of flavocytochrome b(558) . The resultant assembly of phox components results in formation of a complete complex and expression of activity . In this study, we evaluated the oxidase activity of specific granules . In the SDS cell-free system, specific granules expressed oxidase activity in the presence of cytosol in a manner similar to plasma membrane . In contrast to plasma membrane, activity of specific granules was latent, diminishing rapidly over time . In addition, this subcellular fraction contained an inhibitor, possibly related to contamination with azurophilic granules explaining previously published discrepant results . Experiments with recombinant p47phox, p67phox, and dilute cytosol or fractionated cytosol as a source of Rac demonstrated that specific granules have requirements identical to specific granules for oxidase activity . Finally, analysis of neutrophils stimulated with PMA demonstrated translocation of p47phox and to p67phox to specific granules as well as plasma membrane . Both plasma membrane and specific granules from PMA stimulated cells expressed oxidase activity with addition of NADPH demonstrating an assembled oxidase complex . These studies establish a critical role for specific granules as a site for assembly and activation of the oxidase enzyme system and an important constituent for the microbicidal activity of the neutrophil.

Basic Clin Pharmacol Toxicol, 2004 Mar, 94(3), 139 - 43
The influence of single or repeated low-level sarin exposure on immune functions of inbred BALB/c mice; Kassa J et al.; To study the influence of single or repeated low-level sarin inhalation exposure on immune functions, inbred BALB/c mice were exposed to low clinically asymptomatic concentrations of sarin for 60 min . in the inhalation chamber . The evaluation of immune functions was carried out using phenotyping of CD3 (T-lymphocytes), CD4 (helper T-lymphocytes), CD8 (cytotoxic T-lymphocytes) and CD19 (B-lymphocytes) in the lungs, blood and spleen, lymphoproliferation of spleen cells stimulated in vitro by various mitogens (concanavalin A, lipopolysaccharides), phagocyte activity of peritoneal and alveolar macrophages, production of N-oxides by peritoneal macrophages and the measurement of the natural killer cell activity at one week after sarin exposure . The results were compared to the values obtained from control mice exposed to pure air instead of sarin . The results indicate that an asymptomatic dose of sarin is able to alter the reaction of the immune system at one week after exposure to sarin . While the number of CD3 cells in lung was significantly decreased, a slight increase in CD19 cells was observed especially in the lungs after a single sarin inhalation exposure . Lymphoproliferation was significantly decreased regardless of the mitogen and sarin concentration used and the number of low-level sarin exposures . The ability of peritoneal and alveolar macrophages to phagocyte the microbes was also decreased regardless of the number of low-level sarin exposures . The production of N-oxides by peritoneal macrophages was decreased following a single low-level sarin exposure but increased following repeated low-level sarin inhalation exposure . Nevertheless, the changes in the production of N-oxides that reflects a bactericidal activity of peritoneal macrophages was not significant . The natural killer cell activity was significantly higher in the case of inhalation exposure of mice to low concentration of sarin regardless of the number of exposures . Thus, not only organophosphorous insecticides but also nerve agents such as sarin are able to alter immune functions following a single inhalation exposure even at a dose that does not cause clinically manifested intoxication . Generally, the repeated exposure to low concentrations of sarin does not increase the alteration of immune functions compared to the single low-level sarin exposure with the exception of phagocyte activity of alveolar macrophages and natural killer cell activity.

Cell Mol Life Sci, 2004 Mar, 61(6), 682 - 99
The ABC transporter structure and mechanism: perspectives on recent research; Jones PM et al.; ATP-binding cassette (ABC) transporters are multidomain integral membrane proteins that utilise the energy of ATP hydrolysis to translocate solutes across cellular membranes in all phyla . ABC transporters form one of the largest of all protein families and are central to many important biomedical phenomena, including resistance of cancers and pathogenic microbes to drugs . Elucidation of the structure and mechanism of ABC transporters is essential to the rational design of agents to control their function . While a wealth of high-resolution structures of ABC proteins have been produced in recent years, many fundamental questions regarding the protein's mechanism remain unanswered . In this review, we examine the recent structural data concerning ABC transporters and related proteins in the light of other experimental and theoretical data, and discuss these data in relation to current ideas concerning the transporters' molecular mechanism.

Can J Microbiol, 2004 Feb, 50(2), 79 - 89
Genetic and phenotypic traits of streptomycetes used to characterize antibiotic activities of field-collected microbes; Davelos AL et al.; Although antibiotic production may contribute significantly to microbial fitness, there is limited information on the ecology of antibiotic-producing microbial populations in soil . Indeed, quantitative information on the variation in frequency and intensity of specific antibiotic inhibitory and resistance abilities within soil microbial communities is lacking . Among the streptomycetes, antibiotic production is highly variable and resistance to antibiotics is highly specific to individual microbial strains . The objective of this work was to genetically and phenotypically characterize a reference collection of streptomycetes for use in distinguishing inhibition and resistance phenotypes of field-collected microbes . Specifically, we examined inhibition and resistance abilities of all isolates in all possible pairwise combinations, genetic relatedness using BOX-PCR and 16S rDNA sequence analyses, nutrient utilization profiles, and antibiotic induction among all possible three-way combinations of isolates . Each streptomycete isolate possessed a unique set of phenotypic and genetic characteristics . However, there was little correspondence between phenotypic and genetic traits . This collection of reference isolates provides the potential for distinguishing 1024 inhibition and resistance phenotypes in field-collected microbes . Relationships between the genetic and phenotypic characteristics examined may provide preliminary insight into the distinct strategies that microbes use in optimizing their fitness in natural environments.

Environ Microbiol, 2004 May, 6(5), 449 - 61
Community analysis of methanogenic archaea within a riparian flooding gradient; Kemnitz D et al.; Anoxic soils in river floodplains (or riparian soils) are a source of methane emission . However, little is known about the ecology and community structure of archaeal methanogenic microbes, which are a crucial component of methane flux in those habitats . We studied the archaeal community in the vertical profile of four different sites along the River Waal in the Netherlands . These sites differ in their annual flooding regime ranging from never or seldom to permanently flooded . The archaeal community structure has been characterized by terminal restriction fragment length polymorphism (T-RFLP) and comparative sequence analysis of the archaeal SSU rRNA gene and the mcrA gene . The latter gene codes for the alpha-subunit of methyl-coenzyme M reductase . Additionally, the potential methanogenic activity was determined by incubation of soil slurries under anoxic conditions . The community composition differed only slightly with the depth of the soil (0-20 cm) . However, the diversity of archaeal SSU rRNA genes increased with the frequency of flooding . Terminal restriction fragment length polymorphism analysis of mcrA gene amplicons confirmed the results concerning methanogenic archaea . In the never and rarely flooded soils, crenarchaeotal sequences were the dominant group . In the frequently and permanently flooded soils, Methanomicrobiaceae, Methanobacteriaceae, Methanosarcinaceae and the uncultured Rice Clusters IV and VI (Crenarchaeota) were detectable independently from duration of anoxic conditions . Methanosaetaceae, on the other hand, were only found in the permanently and frequently flooded soils under conditions where concentrations of acetate were < 30 microM . The results indicate that methanogens as well as other archaea occupy characteristic niches according to the flooding conditions in the field . Methanosaetaceae, in particular, seem to be adapted (or proliferate at) to low acetate concentrations.

Eur J Immunol, 2004 Apr, 34(4), 1196 - 207
Toll-like receptor stimulation induces airway hyper-responsiveness to bradykinin, an effect mediated by JNK and NF-kappa B signaling pathways; Bachar O et al.; Airway infections induce hyper-responsiveness in asthmatic patients . Toll-like receptors (TLR) mediate inflammatory responses to microbes . Occurrence and effects of TLR2, TLR3 and TLR4 were examined in a mouse organ culture model of asthma focusing on the smooth muscle responses to bradykinin . TLR2, TLR3 and TLR4 mRNA, and TLR2 and TLR4 immunoreactivity were detected in the tracheal muscle layer . Tracheal organ culture for 1 or 4 days with lipopolysaccharide (LPS; TLR2/4 agonist) or polyinosinic polycytidylic acid (poly-I-C; TLR3 agonist) enhanced bradykinin- and {des-Arg(9)}-bradykinin-induced contractions . Simultaneous LPS and poly-I-C treatment resulted in synergistic enhancement of bradykinin-induced contraction . In carbachol-pre-contracted segments TLR stimulation induced less potent relaxations to bradykinin and {des-Arg(9)}-bradykinin . The LPS and poly-I-C enhancement of bradykinin-induced contraction was inhibited by the transcriptional inhibitor actinomycin-D, dexamethasone, the proteasome inhibitor MG-132 and the c-Jun N-terminal kinase (JNK) inhibitor SP600125 . LPS and poly-I-C induced translocation of NF-kappa B p65 to the nucleus and up-regulation of kinin B(1) and B(2) receptor mRNA . In summary, TLR2, TLR3 and TLR4 are expressed in the mouse tracheal smooth muscle . Costimulation of these receptors results in NF-kappa B- and JNK-mediated transcription of B(1) and B(2) receptor, inducing hyper-responsiveness to bradykinin.

Ann Biol Clin (Paris), 2004 Jan-Feb, 62(1), 79 - 83
{Microbeads and nanobeads in microscopy: a tool for spatial, spectral and dynamic settings . Application to molecular co-localization}; Kahn E; This article illustrates a methodology which can be used on cellular and tissular specimens prepared with fluorochromes and analyzed by laser scanning confocal microscopy . Fluorescent beads are used to simulate the fluorochromes and they determine their detectability inside the preparations . They can also be used to verify that the microscopes are set properly so that the resulting images can be analyzed and are reliable to evaluate possible co- localizations . Methods of factor analysis are applied to optical section series obtained on the microscope to characterize the fluorochromes that are used to stain different locations of the specimen . Photophysical properties (emission spectra, decay rates) of the fluorochromes are used to obtain the differentiation . Section series are obtained either by spectral selection via a sequence of filters or by successive scans of the same specimen . Concerning the three- dimensional analysis, differentiation of series that are obtained by z displacement leads to the restoration of focal planes and improves the legibility of specimens . These planes which are not at the same depth when the excitation sources are not aligned can the be superimposed to achieve the co-localization . Copyright John Libbey Eurotext 20003.

Ann Biol Clin (Paris), 2004 Jan-Feb, 62(1), 53 - 7
{Microbeads and flow cytometry: how and why put the "-metry" in immuno-cytometry?}; Poncelet P; In current practice of immunophenotyping, flow cytometers are mainly used as cell counters . These instruments which measure fluorescence intensity on each individual cell with high sensitivity can also count molecules on cells . "Putting the - metry into cytofluorometry" means measuring the expression level of molecules of biological interest on (in) target cells . Candidate molecules are numerous due to the many membrane receptors which biological function depends on the number of accessible molecules . Any modulation of the expression level brings valuable information to the biologist, beit associated to physiological differentiation, pathological state or therapeutic intervention . In order to take the best from such measurements in clinical biology, full reliability is mandatory in terms of time-to-time, platform-to-platform and lab- to-lab reproducibility . One should ban "arbitrary units" as currently provided by the instruments to switch to real units . This paper tends to review the most important features conditioning a reliable quantitation of cellular antigens, with data expressed in terms of number of molecules per cell . In addition to staining tools (antibody-based reagents) and data treatment softwares, calibration tools are mandatory . They are not all equivalent . Fluorescent beads with internal fluorescence are very useful for instrument quality-control but they can not interchange with immunological calibration systems which measure the number of antibody molecules that can bind onto the cells . Numerous application examples illustrate the interest of this "quantitative" dimension of Immunocytometry . Generalization of this approach would help getting a wider access to clinical biology . Copyright John Libbey Eurotext 20003.

Hist Philos Life Sci, 2002, 24(3-4), 467 - 85
Typhus vaccine developments from the First to the Second World War (on Paul Weindling's 'between bacteriology and virology...'); Lindenmann J; After the louse transmission of epidemic typhus had been established (1909), a small microorganism (thought to belong to a new genus, Rickettsia) was shown in enormous numbers in the guts of lice that had fed on human typhus victims . Attempts at cultivating this organism on inert media failed; transfer from louse to louse without loss of virulence for the vertebrate host was successful . Some scientists were not convinced of the etiologic role of Rickettsiae, because the presence of this microbe in blood and organs of victims or of experimentally infected animals was difficult to demonstrate . This uncertainty was dispelled in 1928, when in guinea pigs infected with material from the closely related disease Tabardillo (murine typhus) abundant Rickettsiae were revealed in the tunica vaginalis . Live vaccines, derived from strains of murine typhus and deployed in French North Africa, were considered by outside observers as unsafe . Killed vaccines were derived from the masses of Rickettsiae present in louse guts, in chick embryo yolk sacs or in vertebrate lungs . These developments were not spurned by any 'upswing of virology' but by the threat of typhus in endemic areas and, after 1938, in a war-torn world . Their basis was firmly anchored in bacteriological thought styles and techniques.

Plant Biol (Stuttg), 2004 Mar-Apr, 6(2), 165 - 70
Positive and negative tropic curvature induced by microbeam irradiation of protonemal tip cells of the moss Ceratodon purpureus; Lamparter T et al.; The photoreceptor phytochrome mediates tropic responses in protonemata of the moss Ceratodon purpureus . Under standard conditions the tip cells grow towards unilateral red light, or perpendicular to the electrical vector of polarized light . In this study the response of tip cells to partial irradiation of the apical region was analysed using a microbeam apparatus . The fluence response curve gave an unexpected pattern: whereas a 15-min microbeam with light intensities around 3 micro mol m (-2) s (-1) induced a growth curvature towards the irradiated side, higher light intensities around 100 micro mol m (-2) s (-1) caused a negative response, the cells grew away from the irradiated side . This avoidance response is explained by two effects: the light intensity is high enough to induce photoconversion into the active Pfr form of phytochrome, not only on the irradiated but also on the non-irradiated side by stray light . At the same time, the strong light on the irradiated side acts antagonistically to Pfr . As a result of this inhibition, the growth direction is moved to the light-avoiding side . Such a Pfr-independent mechanism might be important for the phototropic response to distinguish between the light-directed and light-avoiding side under unilateral light.

J Mol Microbiol Biotechnol, 2003, 6(2), 101 - 8
Quantitative analysis of Escherichia coli metabolic phenotypes within the context of phenotypic phase planes; Ibarra RU et al.; In silico models of Escherichia coli metabolism have been developed to predict metabolic behavior and propose experimentally testable hypotheses . However, a thorough assessment of the metabolic phenotype requires well-designed experimentation and reproducible experimental techniques . A method for the quantitative analysis of E . coli metabolism in vivo within the framework of in silico phenotypic phase plane analysis is presented . Using this approach, we have quantitatively studied E . coli metabolism in various environmental conditions and nutritional media . Our experimental methodology, in combination with steady-state metabolic models, can be used to study biological properties and evaluate the metabolic capabilities of microbes .

Med Parazitol (Mosk), 2004 Jan-Mar, (1), 37 - 9
{Differences between two populations of Citellophilus tesquorum altaicus from the Tuva natural focus of plague}; Bazanova LP et al.; Citellophilus tesquorum altaicus fleas from two natural (Karginsky and Barlyksky) populations have been studied . There are differences between the fleas of these populations in susceptibility to plague microbe . The adaptability and abundance of the causative organism in the body are higher in the fleas of the Barlyksky population . The sizes of the head are significantly higher in the females of the Karginsky population . There are no differences in this index between males . According to all 5 studies signs of chaetotaxia, the chaeta abundance is greater in both females and males of the Karginsky population . The fluctuating asymmetry is higher in the Barlyksky fleas that are more susceptible to the studied type of the causative agent of plague.

HNO, 2004 Jul, 52(7), 590 - 8
{Alcohol related diseases of the head and neck}; Riedel F et al.; Chronic consumption of alcohol is an accepted social custom worldwide . In the upper aerodigestive tract, local morphologic, metabolic and functional alterations can be present due such consumption . Gastroesophageal reflux or alterations in sleep structure are typical examples of functional disorders . While alcohol was initially described as a risk enhancer only in smokers, a number of epidemiological studies have now shown that chronic alcohol consumption increases the risk of head and neck cancer independently of exposure to tobacco smoke . In addition, alcohol leads to an accumulation of pathologic microbes within the mucosa, leading to chronic infection . Susceptibility to carcinogens and cell proliferation in the mucosa are increased, resulting in genetic changes with the development of dysplasia, leucoplakia and carcinoma . Chronic alcohol consumption is correlated with an increased risk of cancer and increased mortality in a dose-dependent relationship . A number of biologically plausible mechanisms exist by which alcohol may cause cancer.

Vet Parasitol, 2004 Mar 25, 120(3), 189 - 98
Chemoattraction and chemorepulsion of Strongyloides stercoralis infective larvae on a sodium chloride gradient is mediated by amphidial neuron pairs ASE and ASH, respectively; Forbes WM et al.; Depending on its concentration, sodium chloride acts as either an attractant or a repellant to the infective larvae (L3i) of Strongyloides stercoralis . On a concentration gradient, L3i are attracted to 0.05 M NaCl, but repelled by 2.8M . To test the hypothesis that amphidial neurons ASE and ASH might mediate attraction and repulsion, respectively, these neurons, and control neurons as well, were ablated in hatchling larvae with a laser microbeam . After the larvae attained infectivity (L3i), they were tested on a NaCl gradient . When placed at low salinity, 73.5% of normal controls migrated "up" the gradient, while 26.4% crawled randomly . In contrast, only 20.6% of ASE-ablated L3i migrated "up" the gradient, while 79.4% migrated randomly . Ablation-control ASK-ablated L3i (58.8%) migrated "up" the gradient while 41.1% crawled randomly . When placed at a region of high salinity, 100% of normal control L3i migrated "down" the gradient, whereas 62.5% of ASH-ablated L3i migrated randomly, the remaining 37.5% migrating "down" the gradient . In sharp contrast with ASH-ablated L3i, 94.1% of ablation-control larvae, i.e . ASK-ablated L3i, migrated "down" the gradient . Migration behavior of ASE- and ASH-ablated L3i was significantly different (P < 0.001) from that of ASK-ablated L3i and normal controls . It is noteworthy that 87.5% of ASE-ablated L3i that failed to exhibit chemoattractive behavior were actively chemorepelled from high salinity . Also, 70.0% of ASH-ablated L3i that failed to be chemorepelled from high salinity were capable of chemoattractive behavior, indicating that the worms had retained their behavioral responses except for those associated with the targeted neurons.

Vaccine, 2004 Feb 17, 22(7), 805 - 11
Interactions of commensal gut microbes with subsets of B- and T-cells in the murine host; Jiang HQ et al.; Although mechanisms operative in the induction and maintenance of specific, adaptive immunity, including 'cognate' B/T interactions, have been extensively studied and defined, we still know little about the mechanisms operative in developing and maintaining B- and T-cell dependent 'natural' immunity . Particularly, we are still rather ignorant concerning gut microbial/gut or systemic APC, T cell and B cell interactions that lead to lymphoid cell mediated 'natural' immunity: specific or broadly reactive, activation via TCR and BCR and/or via other receptors such as the TLR series, and whether T/B interactions are operative at this level? Here we will address: (1) the general role of gut microbes in the development and maintenance of the intestinal, humoral immune system; (2) the general role of gut microbes in the development of B1 cell mediated, 'natural' gut IgA and the dependence of these B1 cells on bystander T cell help; (3) the relative contributions of B1 versus B2 cells to gut 'natural' and specific IgA responses; (4) the role for particular 'normal' gut microbes in the initiation of inflammatory bowel diseases (IBD) in mice with a dysregulated immune system; and (5) the possible roles of gut microbes in facilitating oral tolerance, a mechanism likely operative in forestalling or ameliorating IBD . A central theme of this paper is to attempt to define the specificities of activated, functional CD4+ T cells in the gut for Ags of particular, usually benign gut microbes . We will also consider the still-unresolved issue of whether the contributions of B1-derived IgA in the gut to the 'natural' Ab pool are Ag-selected and driven to proliferation/differentiation or whether the main stimuli are not via BCRs but rather other receptors (TLRs, etc.) . The main experimental approach has been to use antigen-free, germ-free, or gnotobiotic (mono- or oligo-associated with precisely known bacterial species) mice.

Nat Prod Rep, 2004 Feb, 21(1), 105 - 21 Epub 2003 Dec 15.
Merging the potential of microbial genetics with biological and chemical diversity: an even brighter future for marine natural product drug discovery; Salomon CE et al.; Marine invertebrates and a growing number of marine bacteria are the sources of novel, bioactive secondary metabolites . Structurally, many of these compounds appear to be biosynthesized by polyketide synthases (PKS) and/or nonribosomal peptide synthetases (NRPS) that have also been found in terrestrial microbes . This review highlights scientific advances from 1999-2003 in the emerging field of molecular genetics of polyketide and nonribosomal peptide natural products isolated from marine organisms . The implications of this research towards the development of marine secondary metabolites as a sustainable source of new drugs are discussed.

Nat Cell Biol, 2004 Mar, 6(3), 227 - 31 Epub 2004 Feb 22.
Kinetochore fibre dynamics outside the context of the spindle during anaphase; Chen W et al.; Chromosomes move polewards as kinetochore fibres shorten during anaphase . Fibre dynamics and force production have been studied extensively, but little is known about these processes in the absence of the spindle matrix . Here we show that laser-microbeam-severed kinetochore fibres in the cytoplasm of grasshopper spermatocytes maintain a constant length while turning over in a polarized manner . Tubulin incorporates at or near the kinetochore and translocates towards severed ends without shortening the fibre . Consequently, the chromosome cannot move polewards unless the severed fibre reattaches to the pole through microtubules . A potential seclusion artefact has been ruled out, as fibres severed inside spindles behave identically despite being surrounded by the spindle matrix . Our data suggest that kinetochore microtubules constantly treadmill during anaphase in insect cells . Treadmilling is an intrinsic property of microtubules in the kinetochore fibre, independent of the context and attachment of the spindle . The machinery that depolymerizes minus ends of kinetochore microtubules is functional in a non-spindle context . Attachment to the pole, however, is required to cause net kinetochore fibre shortening to generate polewards forces during anaphase.

J Leukoc Biol, 2004 Jun, 75(6), 1070 - 8 Epub 2004 Mar 23.
Ascorbate-mediated enhancement of reactive oxygen species generation from polymorphonuclear leukocytes: modulatory effect of nitric oxide; Sharma P et al.; Recent studies from our laboratory have demonstrated that ascorbate potentiated enzymatic synthesis of nitric oxide (NO) from polymorphonuclear leukocytes (PMNs) . NO is known to modulate various function of PMNs such as chemotaxis, adherence, aggregation, and generation of reactive oxygen species (ROS) . The role of ascorbate in the PMN phagocytosis, ROS generation, and apoptosis was thus evaluated in the present study . Ascorbate and its oxidized and cell-permeable analog, dehydroascorbate (DHA), did not affect the phagocytosis but enhanced ROS generation and apoptosis following treatment with Escherichia coli or arachidonic acid . A detailed investigation on the DHA-mediated response indicated that inhibitors of DHA uptake, reduced nicotinamide adenine dinucleotide phosphate oxidase, NO synthase, or ROS scavengers attenuated ROS generation . In DHA-treated cells, enhanced generation of peroxynitrite was also observed; thus, ascorbate-mediated ROS and reactive nitrogen species generation might mediate cytotoxicity toward the ingested microbes and subsequently, augmented PMN apoptosis . Results of the present study have helped in delineating the role of ascorbate in the modulation of NO-mediated ROS generation from PMNs.

Radiat Res, 2004 Apr, 161(4), 397 - 401
The bystander response in C3H 10T1/2 cells: the influence of cell-to-cell contact; Mitchell SA et al.; Although radiation-induced heritable damage in mammalian cells was thought to result from the direct interaction of radiation with DNA, it is now accepted that biological effects may occur in cells that were not themselves traversed by ionizing radiation but are close to those that were . However, little is known about the mechanism underlying such a bystander effect, although cell-to-cell communication is thought to be of importance . Previous work using the Columbia microbeam demonstrated a significant bystander effect for clonogenic survival and oncogenic transformation in C3H 10T(1/2) cells . The present study was undertaken to assess the importance of the degree of cell-to-cell contact at the time of irradiation on the magnitude of this bystander effect by varying the cell density . When 10% of cells were exposed to a range of 2-12 alpha particles, a significantly greater number of cells (P < 0.0001) were inactivated when cells were irradiated at high density (>90% in contact with neighbors) than at low density (<10% in contact) . In addition, the oncogenic transformation frequency was significantly higher in high-density cultures (P < 0.0004) . These results suggest that when a cell is hit by radiation, the transmission of the bystander signal through cell-to-cell contact is an important mediator of the effect, implicating the involvement of intracellular communication through gap junctions.

Microb Ecol, 2004 Apr, 47(3), 252 - 65 Epub 2004 Mar 25.
The calculative nature of microbe-mineral interactions; Caldwell DE et al.; Microorganisms continually redefine themselves at many levels, including the molecule, cell, and community . Although it was initially assumed that this resulted from the genesis of information within DNA alone, it has since been shown that innovation originates at multiple levels . This occurs through calculative units, each unit consisting of two proliferating structures, one nested within the other and each undergoing changes in structural geometry that affect the proliferation rate of the other . For example, the recombination of genetic structures affects the proliferation of community structures, and the recombination of community structures affects the proliferation of genetic structures . The proliferation of a nested series of structures (e.g., genes proliferating within cells, cells proliferating within communities, communities proliferating within ecosystems) results in a logic circuit that calculates the form and function of each structural element in the series . In this situation each element functions as both a habitat and an inhabitant (environment and organism), and it is this dichotomy that determines the balance of nature . Nested geological structures, such as minerals and continents, also proliferate and redefine themselves in much the same way . Microbe-mineral interactions thus link nested biological calculations to an analogous set of nested geological calculations . Examples include the microorganisms involved in the nucleation (proliferation) of ferric hydroxides, carbonates, silicates, and ice crystals.

RNA, 2004 Apr, 10(4), 747 - 53
Selection of AU-rich transiently expressed sequences: reversal of cDNA abundance; Khabar KS et al.; Study of early and transient response gene expression is important for understanding the mechanisms of response to growth stimuli and exogenous agents such as microbes, stress, and radiation . Many of the cytokines, proto-oncogenes, and other transiently expressed gene products are encoded by mRNAs that contain AU-rich elements (AREs) in their 3' untranslated regions (UTRs) . In this article, we describe an approach to selectively synthesize ARE-containing cDNA (ARE-cDNA) using an innovative combination of culture treatment, thermostabilization of reverse transcriptase (RT) by the disaccharide trehalose, and use of optimized ARE-specific oligomers . The monocytic cell line, THP-1, was treated with cycloheximide and endotoxin to enrich for ARE-mediated gene expression followed by the RT procedure . Selection of ARE-cDNA with simultaneous suppression of abundant cDNA was made possible using the procedure as monitored by the preferential expression of IL-8, an ARE-cDNA molecule, over the abundant housekeeping cDNA, beta-actin . The use of trehalose dramatically reversed cDNA abundance, resulting in almost complete suppression of housekeeping cDNA . Finally, construction of specialized ARE-cDNA libraries confirmed the selectivity of ARE-cDNAs and the presence of rare genes . The ability to reverse the abundance of housekeeping and other highly expressed genes toward ARE genes facilitates the discovery and study of rare early response and transiently expressed genes.

Curr Opin Cell Biol, 2004 Feb, 16(1), 86 - 93
Breaking into the epithelial apical-junctional complex--news from pathogen hackers; Vogelmann R et al.; The epithelial apical-junctional complex is a key regulator of cellular functions . In addition, it is an important target for microbial pathogens that manipulate the cell to survive, proliferate and sometimes persist within a host . Out of a myriad of potential molecular targets, some bacterial and viral pathogens have selected a subset of protein targets at the apical-junctional complex of epithelial cells . Studying how microbes use these targets also teaches us about the inherent physiological properties of host molecules in the context of normal junctional structure and function . Thus, we have learned that three recently uncovered components of the apical-junctional complex of the Ig superfamily--junctional adhesion molecule, Nectin and the coxsackievirus and adenovirus receptor--are important regulators of junction structure and function and represent critical targets of microbial virulence gene products.

Curr Opin Microbiol, 2004 Feb, 7(1), 25 - 32
Innate immunity via Toll-like receptors and Nod proteins; Athman R et al.; Host defense against microbes requires the development of an efficient immune response aimed to eradicate the source of infection . Through the expression of a battery of germ-line encoded receptors, including the Toll-like receptors and Nod proteins, the innate immune system, which is a prerequisite to the adaptive immune response, detects microbial motifs and initiates pro-inflammatory signaling . Current research into innate immune function focuses on the nature of the ligands detected by this system, the cell signaling that occurs downstream of receptor activation and finally, how these signals culminate into a tailored adaptive immune response directed to eradicate a specific infection.

Ground Water, 2004 Mar-Apr, 42(2), 167 - 74
Geochemical imaging of flow near an artificial recharge facility, Orange County, California; Clark JF et al.; Critical for the management of artificial recharge operations is detailed knowledge of ground water dynamics near spreading areas . Geochemical tracer techniques including stable isotopes of water, tritium/helium-3 (T/3He) dating, and deliberate gas tracer experiments are ideally suited for these investigations . These tracers were used to evaluate flow near an artificial recharge site in northern Orange County, California, where approximately 2.5 x 10(8) m3 (200,000 acre-feet) of water are recharged annually . T/3He ages show that most of the relatively shallow ground water within 3 km of the recharge facilities have apparent ages < 2 years; further downgradient apparent ages increase, reaching > 20 years at approximately 6 km . Gas tracer experiments using sulfur hexafluoride and xenon isotopes were conducted from the Santa Ana River and two spreading basins . These tracers were followed in the ground water for more than two years, allowing subsurface flow patterns and flow times to be quantified . Results demonstrate that mean horizontal ground water velocities range from < 1 to > 4 km/year . The leading edges of the tracer patch moved at velocities about twice as fast as the center of mass . Leading edge velocities are important when considering the potential transport of microbes and other "time sensitive" contaminants and cannot be determined easily with other methods . T/3He apparent ages and tracer travel times agreed within the analytical uncertainty at 16 of 19 narrow screened monitoring wells . By combining these techniques, ground water flow was imaged with time scales on the order of weeks to decades.

J Immunol, 2004 Apr 1, 172(7), 3989 - 93
Cutting edge: innate immune system discriminates between RNA containing bacterial versus eukaryotic structural features that prime for high-level IL-12 secretion by dendritic cells; Koski GK et al.; RNA derived from bacterial but not eukaryotic sources, when transfected into human monocyte-derived dendritic cell precursors, induces high-level IL-12 secretion in conjunction with dendritic cell maturation stimuli . In vitro-transcribed mRNA that mimics the structure of bacterial mRNA in the lack of a long 3'-poly(A) tail likewise induces IL-12 secretion, but this property is lost upon efficient enzymatic 3'-polyadenylation . Among other tested RNAs, only polyuridylic acid induced IL-12 p70 . This RNA response phenomenon appears biologically distinct from the classically defined response to dsRNA . RNA-transfected APC also polarize T cells in an IL-12-dependent manner toward the IFN-gamma(high)IL-5 (low) Th1 phenotype, suggesting a link between the detection of appropriately structured RNA and the skewing of immune responses toward those best suited for controlling intracellular microbes . RNA structured to emulate bacterial patterns constitutes a novel vaccine strategy to engender polarized Th1-type immune responses.

Ann N Y Acad Sci, 2003 Dec, 1010, 467 - 70
Innate immune collectins bind nucleic acids and enhance DNA clearance in vitro; Palaniyar N et al.; Collectins are a class of innate immune proteins that opsonize microbes and other ligands via their carbohydrate-recognition domains and enhance their clearance by phagocytes . We hypothesized that collectins, such as pulmonary surfactant protein (SP-) A and D and mannose-binding lectin (MBL), could bind nucleic acid, which is a pentameric sugar-based anionic polymer . We have shown that SP-D and MBL bind to both DNA and RNA effectively and also that SP-D enhances the uptake of DNA by human monocytic cells . Therefore, our results suggest that nucleic acids may be a novel class of ligands for collectins . This class of innate immune proteins could enhance the clearance of DNA that is released by necrotic cells or microbial cells . Our findings may have important biological implications, such as the alleviation of DNA-mediated tissue inflammation.

Blood, 2004 Jul 15, 104(2), 453 - 61 Epub 2004 Mar 18.
In vitro-expanded human CD4(+)CD25(+) T-regulatory cells can markedly inhibit allogeneic dendritic cell-stimulated MLR cultures; Godfrey WR et al.; CD4(+)CD25(+) T-regulatory (Treg) cells have been shown to critically regulate self- and allograft tolerance in several model systems . Studies of human Treg cells have been restricted by the small number present in peripheral blood and their naturally hypoproliferative state . To better characterize Treg suppressor cell function, we determined methods for the isolation and expansion of these cells . Stringent magnetic microbead-based purification was required for potent suppressor cell line generation . Culture stimulation with cell-sized Dynabeads coated with anti-CD3 and anti-CD28 monoclonal antibodies, CD4(+) feeder cells, and interleukin 2, provided for marked expansion in cell number (100-fold), with retention and enhancement of suppressor function . The potent Treg cell lines suppressed proliferation in dendritic cell-driven allo-mixed lymphocyte reaction (MLR) cultures by more than 90% . The Treg-derived suppressor cells functioned early in allo-MLR because expression of activation antigens and accumulation of cytokines was nearly completely prevented . Importantly, cultured Treg cells also suppressed activated and matured dendritic cell-driven responses . These results demonstrate that short-term suppressor cell lines can be generated, and they can express a very potent suppressive activity . This approach will enable more detailed biologic studies of Treg cells and facilitate the evaluation of cultured Treg cells as a novel form of immunosuppressive therapy.

Pharmacol Ther, 2004 Mar, 101(3), 193 - 210
Role of atypical bacterial infection of the lung in predisposition/protection of asthma; Hansbro PM et al.; Asthma is a common inflammatory disease of the airways that results in airway narrowing and wheezing . Allergic asthma is characterised by a T-helper cell-type (Th) 2 response, immunoglobulin (Ig) E production, and eosinophilic influx into the airways . Recently, many clinical studies have implicated Mycoplasma pneumoniae and Chlamydia pneumoniae in the development and exacerbation of both chronic and acute asthma . It is widely accepted that M . pneumoniae and C . pneumoniae infections require Th1 immunity for clearance; therefore, according to the hygiene hypothesis, these infections should be protective against asthma . Here, we review the clinical evidence for the association and mechanisms of predisposition to and protection against asthma by these infections . We will examine the following question: Is it the absence of infection or the age of the individual on infection that confers susceptibility or resistance to asthma and does this vary between normal and predisposed individuals? We put forward a hypothesis of the effects of these infections on the development and prevention of asthma and how novel preventative and treatment strategies involving these microbes may be targeted against asthma.

Presse Med, 2004 Feb 28, 33(4), 256 - 61; discussion 269
{Physiopathology of severe sepsis}; Caille V et al.; Regarding the definition . Severe sepsis associates an explosive inflammatory reaction and organ failure . It is secondary to bacterial, fungal or viral infection . It can be at the origin of acute circulatory failure (state of septic shock) . Response of the organism to infection . The presence of certain components of the membrane of pathogenic agents induces the release of various mediators in cascade, notably cytokines . Toll-like receptors (10 cloned in humans) intervene in the detection of microbes and in the inherent and subsequently adaptive immune response . Immune paralysis . The release of pro-inflammatory mediators characterizes the initial phase of sepsis . Persistence of the latter provokes acquired immunodepression, related to an anti-inflammatory profile, and hence to a delayed decrease in hypersensitivity, an incapacity to cope with the infection and the onset of nosocomial infections . The role of the mediators . During sepsis, the cytokines are predominantly pro-inflammatory (TNF-alpha and notably IL-1beta) whereas others, produced concomitantly or subsequently, are predominantly anti-inflammatory (IL-10 in particular) . In fact, the majority of the cytokines have multiple and intrinsic effects, they mediate immune defense but also pathological manifestations . Many other mediators intervene: coagulation or complement systems, contact system, breakdown products of the phospholipid membrane, arachidonic acid metabolites, free radicals and nitrous oxide . Endocrine and metabolic dysregulations . The concept of relative adrenal insufficiency and peripheral syndrome of resistance to glycocorticosteroids have led to hormone replacement therapy during septic shock . Acute insulin resistance has also been described . The role of the endothelium and coagulation . The endothelium plays a key part in the onset of vascular insufficiency during sepsis due to abnormalities in vasomotricity and thrombomodulation . The anticoagulant regulating system is perturbed; there is a decrease in protein C with inactivation of its active form, which has pro-fibrinolytic properties, and a decrease in antithrombin III . Regarding myocardial dysfunction During septic shock there is often severe left ventricular systolic dysfunction, sometimes also involving the right ventricle, largely under-diagnosed despite its severe prognosis, and associated with reduced or even collapsed heart rate.

Mol Gen Mikrobiol Virusol, 2004, (1), 22 - 6
{Analysis of genomic polymorphism of typical and atypical strains of the plague pathogen using polymerase chain reaction with universal primers}; Savostina EP et al.; An analysis of genome polymorphism of the Y . pestis strains by using the method of polymerase chain reaction (PCR) for the tandem repeats of bacteriophage M13 DNA revealed a species similarity of both typical and atypical (according to diagnostic signs) plague-microbe strains . Strain Y . pestis A-1726 with the atypical differential-and-diagnostic properties, without the amplicon specified for Y . pestis and sized 1000 b.p., was identified among 27 analyzed Y . pestis strains . The amplicon profiles of the basic Y . pestis subtype were found to be different from such profiles of other Y . pestis subtypes.

Vestn Ross Akad Med Nauk, 2004, (1), 38 - 40
{The isolation of patients with highly contagious diseases}; Buianov VV et al.; The paper deals with the issue of how to prevent the spread (in the environment) of pathogens of extra dangerous infections, whose source could be a patients (not isolated in Meltzer box) with a highly contagious disease . Autonomous filtering stretchers of a special design are suggested for the transportation of patients; the design is sufficient to protect the habitat and medical personnel against being contaminated with related microbes and does not affect the physiological-and-hygienic indices of an isolated patient.

Vestn Ross Akad Med Nauk, 2004, (1), 30 - 5
{Use of new individual protection substances in Mel'tser boxes}; Buianov VV et al.; Described in the paper are the results of research, experiments and designing of individual protection means for medical personnel attending the patients with extra dangerous infections under the conditions of Meltzer boxes . The authors elaborated pressure suits that can protect the medical personnel from being infected by microbe aerosols . The testing of the mentioned protection suits in infection hospitals of Moscow, Elista, Rostov-on-Don and of other cities was positively assessed.

Anal Chem, 2004 Mar 15, 76(6), 1586 - 95
X-ray fluorescence tomography of individual municipal solid waste and biomass fly ash particles; Camerani MC et al.; Information about Cd distribution inside single municipal solid waste and biomass fly ash particles is fundamental since it affects its leachability . The internal 2D distributions of the main and trace elements in such highly inhomogeneous matrixes were successfully determined by means of the combined synchrotron radiation induced micro X-ray fluorescence (micro-SRXRF) and tomography (micro-SRXRFT) techniques . Scanning micro-SRXRF measurements show Cd elemental distribution within single fly ash particles to be inhomogeneous, but no information can be obtained about its internal distribution . During micro-SRXRFT analysis, single fly ash particles are successively measured by a rotational-translational scan in a VH=2 x 5 microm2 microbeam . The 2D internal elemental distribution images, obtained by the modified simultaneous algebraic reconstruction technique algorithm, provide the size and the location of Cd-containing areas together with the location of other measurable elements . Results showed Cd concentration to be higher in the core of the fly ash particles analyzed rather than on the surface of the particles . Moreover, in both ashes, Ca-containing matrixes are found to be the main Cd-bearing phases . A possible mechanism for Cd adsorption on the fly ash particles is proposed based on the obtained results.

CNS Drugs, 2004, 18(4), 243 - 50
Pharmacokinetic considerations in the treatment of attention-deficit hyperactivity disorder with methylphenidate; Wolraich ML et al.; Methylphenidate is commonly used for the treatment of attention-deficit hyperactivity disorder (ADHD) . Its efficacy in improving the core symptoms of ADHD, as well as some of the aggressive and oppositional behaviours, is well documented, based on a large volume of research . Methylphenidate has a wide margin of safety and relatively mild adverse effects, most commonly appetite suppression and insomnia.Methylphenidate is a rapidly absorbed medication that, in its d-isomer form, readily penetrates the CNS, particularly the striatum . It appears to function by blocking the reuptake of dopamine.Both the plasma concentrations and behavioural effects of methylphenidate demonstrate a time to maximum of between 1 and 3 hours, with the maximum behavioural effects occurring when the plasma concentrations are increasing . Because of the rapid onset of action, the effects of methylphenidate can be dramatic but usually last only about 4 hours with the immediate-release formulation . The behavioural responses of individuals are also highly variable, so that it is necessary to start treatment at a low dosage and increase up to a maximally effective dosage (usually starting at 10-15 mg/day with increases of 10-15mg at weekly intervals to a maximum dosage of 60 mg/day, irrespective of formulation) . Because of the variability in behavioural responses, assessment of plasma concentrations is not clinically useful nor does weight help in deciding an appropriate dosage . The difficulties in administering methylphenidate multiple times a day, particularly during the school day, have been alleviated in the past few years by the development of extended-release preparations with varying behavioural effects lasting 8-12 hours . The 8-hour preparations (Metadate) CD and Ritalin) LA) utilise a microbead technology, while the 12-hour preparation (Concerta) utilises an osmotic pump system . All extended-release formulations effectively control the symptoms of ADHD . While pharmacokinetic differences appear to exist between some of these new formulations, there are currently no clinical data available to demonstrate clinical efficacy differences between them.

J Membr Biol, 2004 Jan 1, 197(1), 1 - 32
What are aquaporins for?
Hill AE, Shachar-Hill B, Shachar-Hill Y.
The prime function of aquaporins (AQPs) is generally believed to be that of increasing water flow rates across membranes by raising their osmotic or hydraulic permeability . In addition, this applies to other small solutes of physiological importance . Notable applications of this 'simple permeability hypothesis' (SPH) have been epithelial fluid transport in animals, water exchanges associated with transpiration, growth and stress in plants, and osmoregulation in microbes . We first analyze the need for such increased permeabilities and conclude that in a range of situations at the cellular, subcellular and tissue levels the SPH cannot satisfactorily account for the presence of AQPs . The analysis includes an examination of the effects of the genetic elimination or reduction of AQPs (knockouts, antisense transgenics and null mutants) . These either have no effect, or a partial effect that is difficult to explain, and we argue that they do not support the hypothesis beyond showing that AQPs are involved in the process under examination . We assume that since AQPs are ubiquitous, they must have an important function and suggest that this is the detection of osmotic and turgor pressure gradients . A mechanistic model is proposed--in terms of monomer structure and changes in the tetrameric configuration of AQPs in the membrane--for how AQPs might function as sensors . Sensors then signal within the cell to control diverse processes, probably as part of feedback loops . Finally, we examine how AQPs as sensors may serve animal, plant and microbial cells and show that this sensor hypothesis can provide an explanation of many basic processes in which AQPs are already implicated . Aquaporins are molecules in search of a function; osmotic and turgor sensors are functions in search of a molecule.

Clin Diagn Lab Immunol, 2004 Mar, 11(2), 255 - 60
Comparison of benchtop microplate beta counters with the traditional gamma counting method for measurement of chromium-51 release in cytotoxic assays; Wallace D et al.; The most traditional method used to measure the lytic activity of cytotoxic T lymphocytes or natural killer (NK) cells is the chromium release assay (CRA) . No study has been reported that systematically compares the traditional gamma counting method with various benchtop microplate scintillation formats to measure chromium release . Here we investigated the utilization of microplate beta counters in comparison with the traditional gamma counting method to quantitate antigen-specific cytolysis, lymphokine-activated killer (LAK) activity, and NK activity in the CRA . Supernatants from standard CRA (n = 7) were directly transferred to a 96-well microplate containing either a solid scintillant (Lumaplate) or a liquid scintillant (flexible beta plate) . Samples were quantified by using two benchtop microplate beta counters, Wallac Microbeta Trilux (Lumalux and Trilux methods, respectively) and Packard TopCount instruments (TopCount method) . These results were then compared with data from an identical assay run in parallel using the traditional gamma counting method (LKB) . The lytic activity for influenza virus-stimulated effectors measured in the benchtop microplate beta counters using Lumalux and Trilux methods exhibited excellent correlations with the one measured in the traditional LKB (r = 0.967 and 0.968, respectively) . The TopCount method demonstrated a similar correlation (r = 0.966) . Similar findings were observed for LAK and NK activity . The 96-well microplate format, specifically the dry-scintillant Lumaplates, offers several advantages over the traditional gamma counting format . Most notable are the reductions in sample volume needed and in the total sample preparation and counting time . Furthermore, this system reduces the amount of dry and mixed radioactive waste generated while using the same instrument for gamma- and beta-emitting isotopes.

Annu Rev Plant Physiol Plant Mol Biol, 1996 Jun, 47, 75 - 100
DNA DAMAGE AND REPAIR IN PLANTS; Britt AB; The biological impact of any DNA damaging agent is a combined function of the chemical nature of the induced lesions and the efficiency and accuracy of their repair . Although much has been learned from microbes and mammals about both the repair of DNA damage and the biological effects of the persistence of these lesions, much remains to be learned about the mechanism and tissue-specificity of repair in plants . This review focuses on recent work on the induction and repair of DNA damage in higher plants, with special emphasis on UV-induced DNA damage products.

Appl Opt, 2004 Feb 10, 43(5), 1063 - 71
Scanning total internal reflection fluorescence microscopy under one-photon and two-photon excitation: image formation; Chon JW et al.; We propose a new type of total internal reflection fluorescence microscopy (TIRFM) called scanning TIRFM (STIRFM) that uses a focused ring-beam illumination and a high-numerical-aperture objective (NA = 1.65) . The evanescent field produced by the STIRFM is focused laterally, producing a small excitation volume that can induce a nonlinear effect such as two-photon absorption . Experimental images of CdSe quantum dot nanocrystals and Rhodamine 6G-doped microbeads show that good lateral and axial resolutions are achieved with the current setup . The theoretical simulation of the focal spot produced in STIRFM geometry shows that the focused evanescent field is split into two peaks because of the depolarization effect of a high numerical-aperture objective lens . However, the point-spread function analysis of both one-photon and two-photon excitation cases shows that the detection of the focus-splitting effect is dependent on the detection pinhole size . The effect of pinhole size on image formation is theoretically investigated and confirmed experimentally with the nanocrystal images.

Shanghai Kou Qiang Yi Xue, 2004 Feb, 13(1), 65 - 8
{The influence of oral health behavior of parent and children to caries risk factors of children}; Gao YX et al.; Caries risk factors such as the number of microbe CFU in plaque and saliva, the content of inorganic ion in plaque and saliva and the capability of remineralization of the teeth, is responsible to forecast the development of caries to some content . On the other hand, oral health behaviours also make a great deal in the development of caries . While in terms of children, the parents have an enormous effect on their behaviours . In order to help forecasting the development of caries more effectively, this paper makes a review about the influence of oral health behaviour of parents and children to caries risk factors of children.

Microbiol Mol Biol Rev, 2004 Mar, 68(1), 132 - 53, table of contents
Diversity of microbial sialic acid metabolism; Vimr ER et al.; Sialic acids are structurally unique nine-carbon keto sugars occupying the interface between the host and commensal or pathogenic microorganisms . An important function of host sialic acid is to regulate innate immunity, and microbes have evolved various strategies for subverting this process by decorating their surfaces with sialylated oligosaccharides that mimic those of the host . These subversive strategies include a de novo synthetic pathway and at least two truncated pathways that depend on scavenging host-derived intermediates . A fourth strategy involves modification of sialidases so that instead of transferring sialic acid to water (hydrolysis), a second active site is created for binding alternative acceptors . Sialic acids also are excellent sources of carbon, nitrogen, energy, and precursors of cell wall biosynthesis . The catabolic strategies for exploiting host sialic acids as nutritional sources are as diverse as the biosynthetic mechanisms, including examples of horizontal gene transfer and multiple transport systems . Finally, as compounds coating the surfaces of virtually every vertebrate cell, sialic acids provide information about the host environment that, at least in Escherichia coli, is interpreted by the global regulator encoded by nanR . In addition to regulating the catabolism of sialic acids through the nan operon, NanR controls at least two other operons of unknown function and appears to participate in the regulation of type 1 fimbrial phase variation . Sialic acid is, therefore, a host molecule to be copied (molecular mimicry), eaten (nutrition), and interpreted (cell signaling) by diverse metabolic machinery in all major groups of mammalian pathogens and commensals.

Appl Environ Microbiol, 2004 Mar, 70(3), 1658 - 68
Diverse organization of genes of the beta-ketoadipate pathway in members of the marine Roseobacter lineage; Buchan A et al.; Members of the Roseobacter lineage, an ecologically important marine clade within the class alpha-Proteobacteria, harbor genes for the protocatechuate branch of the beta-ketoadipate pathway, a major catabolic route for lignin-related aromatic compounds . The genes of this pathway are typically clustered, although gene order varies among organisms . Here we characterize genes linked to pcaH and -G, which encode protocatechuate 3,4-dioxygenase, in eight closely related members of the Roseobacter lineage (pairwise 16S rRNA gene sequence identities, 92 to 99%) . Sequence analysis of genomic fragments revealed five unique pca gene arrangements . Identical gene organization was found for isolates demonstrating species-level identity (i.e., >99% 16S rRNA gene similarity) . In one isolate, six functionally related genes were clustered: pcaQ, pobA, pcaD, pcaC, pcaH, and pcaG . The remaining seven isolates lacked at least one of these genes in their clusters, although the relative order of the remaining genes was preserved . Three genes (pcaC, -H, and -G) were physically linked in all isolates . A highly conserved open reading frame (ORF) was found immediately downstream of pcaG in all eight isolates . Reverse transcription-PCR analysis of RNA from one isolate, Silicibacter pomeroyi DSS-3, provides evidence that this ORF is coexpressed with upstream pca genes . The absence of this ORF in similar bacterial pca gene clusters from diverse microbes suggests a niche-specific role for its protein product in Roseobacter group members . Collectively, these comparisons of bacterial pca gene organization illuminate a complex evolutionary history and underscore the widespread ecological importance of the encoded beta-ketoadipate pathway.

Appl Environ Microbiol, 2004 Mar, 70(3), 1627 - 32
Direct observation of microbial inhibition of calcite dissolution; Luttge A et al.; Vertical scanning interferometry (VSI) provides a method for quantification of surface topography at the angstrom to nanometer level . Time-dependent VSI measurements can be used to study the surface-normal retreat across crystal and other solid surfaces during dissolution or corrosion processes . Therefore, VSI can be used to directly and nondestructively measure mineral dissolution rates with high precision . We have used this method to compare the abiotic dissolution behavior of a representative calcite (CaCO(3)) cleavage face with that observed upon addition of an environmental microbe, Shewanella oneidensis MR-1, to the crystal surface . From our direct observations, we have concluded that the presence of the microbes results in a significant inhibition of the rate of calcite dissolution . This inhibition appears to be a 2nd-order effect that is related to the formation of etch pits . The opening of etch pits was greatly inhibited in the presence of added bacteria, suggesting that the bacterial cells exert their effect by inhibiting the formation of etch pits at high-energy sites at the crystal surface caused by lattice defects, e.g., screw or point dislocations . The experimental methodology thus provides a nondestructive, directly quantifiable, and easily visualized view of the interactions of microbes and minerals during weathering (or corrosion) processes or during mineral precipitation.

Appl Environ Microbiol, 2004 Mar, 70(3), 1328 - 35
Wood-destroying soft rot fungi in the historic expedition huts of Antarctica; Blanchette RA et al.; Three expedition huts in the Ross Sea region of Antarctica, built between 1901 and 1911 by Robert F . Scott and Ernest Shackleton, sheltered and stored the supplies for up to 48 men for 3 years during their explorations and scientific investigation in the South Pole region . The huts, built with wood taken to Antarctica by the early explorers, have deteriorated over the past decades . Although Antarctica has one of the coldest and driest environments on earth, microbes have colonized the wood and limited decay has occurred . Some wood in contact with the ground contained distinct microscopic cavities within secondary cell walls caused by soft rot fungi . Cadophora spp . could be cultured from decayed wood and other woods sampled from the huts and artifacts and were commonly associated with the soft rot attack . By using internal transcribed spacer sequences of ribosomal DNA and morphological characteristics, several species of Cadophora were identified, including C . malorum, C . luteo-olivacea, and C . fastigiata . Several previously undescribed Cadophora spp . also were found . At the Cape Evans and Cape Royds huts, Cadophora spp . commonly were isolated from wood in contact with the ground but were not always associated with soft rot decay . Pure cultures of Cadophora used in laboratory decay studies caused dark staining of all woods tested and extensive soft rot in Betula and Populus wood . The presence of Cadophora species, but only limited decay, suggests there is no immediate threat to the structural integrity of the huts . These fungi, however, are widely found in wood from the historic huts and have the capacity to cause extensive soft rot if conditions that are more conducive to decay become common.

Cell Mol Life Sci, 2004 Mar, 61(5), 537 - 46
Drosophila melanogaster innate immunity: an emerging role for peptidoglycan recognition proteins in bacteria detection; Royet J; Over the past years, parallel studies conducted in mammals and flies have emphasized the existence of common mechanisms regulating the vertebrate and invertebrate innate immune systems . This culminated in the discovery of the central role of the Toll pathway in Drosophila immunity and in the implication of Toll-like receptors (TLRs)/interleukin-1(IL-1) in the mammalian innate immune response . In spite of clear similarities, such as shared intracellular pathway components, important divergences are expected between the two groups, whose last common ancestor lived more than half a billion years ago . The most obvious discrepancies lie in the mode of activation of the signalling receptors by microorganisms . In mammals, TLRs are part of protein complexes which directly recognize microbe-associated patterns, whereas Drosophila Toll functions like a classical cytokine receptor rather than a pattern recognition receptor . Recent studies demonstrate that members of the evolutionarily conserved peptidoglycan recognition protein family play an essential role in microbial sensing during immune response of Drosophila.

J Clin Microbiol, 2004 Mar, 42(3), 1170 - 5
Computer-assisted surveillance for detecting clonal outbreaks of nosocomial infection; Hacek DM et al.; Whole-house surveillance for healthcare-associated infection is no longer the recommended practice because of the large personnel time investment required . We developed a computer-based tracking system using microbiologic data as an aid in detecting potential outbreaks of healthcare-associated infections on a hospital-wide basis . Monthly total isolates of 25 clinically significant hospital pathogens were tallied from 1991 to 1998 to form a database for future comparison . Two different algorithm tools (based on increases of organism numbers over baseline) were applied to determine alert thresholds for suspected outbreaks using this information . The first algorithm (2SD) defined an alert as two standard deviations above the mean monthly number of isolates . The second (MI) defined an alert as either a 100% increase from the baseline organism number over 2 months or a >/=50% increase (compared to baseline) during a three-consecutive-month period . These two methods were compared to standard infection control professional surveillance (ICP) for the detection of clonal outbreaks over 12 months . Overall, a total of seven clonal outbreaks were detected during the 1-year study . Using standard methods, ICP investigated nine suspected outbreaks, four of which were associated with clonal microbes . The 2SD method signaled a suspected outbreak 15 times, of which three were clonal and ICP had detected one . The MI method signaled a suspected outbreak 30 times; four of these were clonal, and ICP had detected one . The sensitivity and specificity values for ICP, 2SD, and MI for detecting clonal outbreaks were 57, 43, and 57% and 17, 83, and 67%, respectively . Statistical methods applied to clinical microbiology laboratory information system data efficiently supplement infection control efforts for outbreak detection.

Biomacromolecules, 2004 Mar-Apr, 5(2), 511 - 5
Microbial scission of sulfide linkages in vulcanized natural rubber by a white rot basidiomycete, Ceriporiopsis subvermispora; Sato S et al.; A white rot basidiomycete, Ceriporiopsis subvermispora, degraded vulcanized natural rubber (NR) sheets on a wood medium . The fungus decreased the total sulfur content of the rubber by 29% in 200 days, accompanied by the cleavage of sulfide bonds between polyisoprene chains . X-ray photoelectron spectroscopy (XPS) demonstrated that C . subvermispora reduced the frequency of S-C bonds by 69% with a concomitant formation of S-O bonds during the culture period . Dipolar decoupling/magic angle spinning (DD/MAS) solid state 13C NMR revealed that the fungus preferentially decomposed monosulfide bonds linked to a cis- and trans-1,4-isoprene backbone but the cleavage of polysulfide bonds was also observed . In contrast, no decrease in weight or devulcanization of rubber was observed in cultures of a white rot fungus, Dichomitus squalens . The oxidative cleavage of sulfide bonds by C . subvermispora demonstrates that ligninolytic basidiomycetes are potential microbes for the biological devulcanization of rubber products.

J Clin Immunol, 2004 Jan, 24(1), 30 - 6
Increased CD8+ T cell apoptosis in scleroderma is associated with low levels of NF-kappa B; Kessel A et al.; Our objectives were (1) to compare lymphocyte subpopulation apoptosis rates in SSc patients versus healthy controls and (2) to compare Bcl-2 and NF-kappa B expression in cultured CD8 lymphocytes of SSc patients versus controls . Peripheral blood samples were obtained from 27 SSc patients meeting the American College of Rheumatology criteria for SSc and 28 healthy individuals . Mononuclear cells were isolated by Ficoll-Hypaque density gradient separation and cultured for 48 hr . For determination of apoptosis within specific cell populations, samples were labeled with PE-conjugated monoclonal antibody to CD8, CD4, and a FITC-conjugated monoclonal antibody to Annexin V . Flow cytometry was carried out with a FACS operating with Cellquest software . CD8+ lymphocytes were positively selected with magnetic microbeads conjugated to antihuman CD8 . CD8 T cells were separated, then incubated with activation for 48 hr, and NF-kappa B and Bcl-2 analysis was carried out using Western immunoblotting . The CD4:CD8 ratio was increased in SSc compared to controls (2.6 +/- 1.13 vs.1.87 +/- 0.76; P = 0.018) . The spontaneous apoptosis rate of SSc CD8 lymphocytes was increased compared to that of controls of (21.9 +/- 13.7 vs . 13.3 +/- 9.9; P = 0.019) . No difference was found in the rate of CD4 apoptosis of SSc patients versus controls (9.8 +/- 5.2 vs . 7.18 +/- 4.89%; P = ns) . The expression of NF-kappa B in SSc CD8 lymphocytes was decreased compared with that of CD8 lymphocytes from healthy controls (144 +/- 13 vs . 188 +/- 11; P = 0.018) . Whereas expression of Bcl-2 was similar in activated CD8+ T cells of SSc patients and healthy controls, CD8+ T cell apoptosis rate was found to be in reverse correlation with expression of NF-kappa B in these cells ( r = - 0.53, P = 0.029) . The increased CD4:CD8 ratio in SSC may result from increased CD8+ T cell apoptosis . Increased SSc CD8 T cell apoptosis is associated with low levels of NF-kappa B.

Environ Pollut, 2004 May, 129(2), 165 - 73
Testing the ecotoxicology of vegetable versus mineral based lubricating oils: 1 . Degradation rates using tropical marine microbes; Mercurio P et al.; Vegetable-derived lubricants (VDL) might be more biodegradable than mineral-derived lubricants (MDL) due to the absence of high molecular weight aromatics, but this remains largely untested in tropical conditions . In this laboratory study, the degradation rates of 2-stroke, 4-stroke and hydraulic VDLs were compared with their MDL counterparts in the presence of mangrove and coral reef microbial communities . While MDLs were comprised largely of unresolved saturated and some aromatic hydrocarbons, their VDL counterparts contained, potentially more degradable, fatty acid methyl esters . Degradation of some VDL was observed by day 7, with the 2-stroke VDL markedly consumed by mangrove microorganisms and the hydraulic VDL degraded by both microorganism communities after this short period . All of the VDL groups were significantly more degraded than the comparable MDLs mineral oil lubricants over 14 days in the presence of either mangrove or coral reef microbial communities . In general the mangrove-sourced microorganisms more efficiently degraded the lubricants than reef-sourced microorganisms.

J Autoimmun, 2004 Mar, 22(2), 95 - 106
T-cell receptor transgenic mice in the study of autoimmune diseases; Lafaille JJ; T cell receptor transgenic mice have been a valuable tool in the study of the immune system, from development to selection to tolerance or pathogenesis . In this manuscript we review the T cell receptor transgenic mouse lines with specificity for self antigens that have been reported before August 2003 . Many such lines have been generated, which have been instrumental in our understanding of, among other aspects, the role regulatory T cells in preventing autoimmunity, the role of microbes in modifying its outcome, the influence of the genetic background, the importance of regional differences in self-antigen concentration, and the importance of differences in antigen deposition between different tissues.

J Nat Prod, 2004 Feb, 67(2), 257 - 68
Natural products from endophytic microorganisms; Strobel G et al.; Endophytic microorganisms are to be found in virtually every plant on earth . These organisms reside in the living tissues of the host plant and do so in a variety of relationships ranging from symbiotic to pathogenic . Endophytes may contribute to their host plant by producing a plethora of substances that provide protection and ultimately survival value to the plant . Ultimately, these compounds, once isolated and characterized, may also have potential for use in modern medicine, agriculture, and industry . Novel antibiotics, antimycotics, immunosuppressants, and anticancer compounds are only a few examples of what has been found after the isolation and culturing of individual endophytes followed by purification and characterization of some of their natural products . The prospects of finding new drugs that may be effective candidates for treating newly developing diseases in humans, plants, and animals are great . Other applications in industry and agriculture may also be discovered among the novel products produced by endophytic microbes.

Ying Yong Sheng Tai Xue Bao, 2003 Oct, 14(10), 1799 - 803
{Main mechanism and affecting factors of phytoremediation of organic contaminated soil}; Lin D et al.; In this paper, the main mechanisms of phytoremediation of organic contaminated soil, such as adsorption and degradation in rhizosphere, plant uptake, and transformation, and phytoremediation model were illuminated in detail, and their affecting factors, including properties of pollutants, plant species, soil properties, coexisted pollutants, and climate conditions were analyzed . The main topics that need further research were prospected, e.g., deepening the mechanism of phytoremediation, consummating phytoremediation models, approaching on the mechanism and appliance of microbe associated phytoremediation systems, building efficient surfactants enhancing phytoremediation systems, and strengthening the research on phytoremediation of combined organic pollution.

Nature, 2004 Feb 26, 427(6977), 853 - 8
The large-conductance Ca2+-activated K+ channel is essential for innate immunity; Ahluwalia J et al.; Neutrophil leukocytes have a pivotal function in innate immunity . Dogma dictates that the lethal blow is delivered to microbes by reactive oxygen species (ROS) and halogens, products of the NADPH oxidase, whose impairment causes immunodeficiency . However, recent evidence indicates that the microbes might be killed by proteases, activated by the oxidase through the generation of a hypertonic, K+-rich and alkaline environment in the phagocytic vacuole . Here we show that K+ crosses the membrane through large-conductance Ca2+-activated K+ (BK(Ca)) channels . Specific inhibitors of these channels, iberiotoxin and paxilline, blocked oxidase-induced 86Rb+ fluxes and alkalinization of the phagocytic vacuole, whereas NS1619, a BK(Ca) channel opener, enhanced both . Characteristic outwardly rectifying K+ currents, reversibly inhibited by iberiotoxin, were demonstrated in neutrophils and eosinophils and the expression of the alpha-subunit of the BK channel was confirmed by western blotting . The channels were opened by the combination of membrane depolarization and elevated Ca2+ concentration, both consequences of oxidase activity . Remarkably, microbial killing and digestion were abolished when the BK(Ca) channel was blocked, revealing an essential and unexpected function for this K+ channel in the microbicidal process.

Cytotherapy, 2004, 6(1), 15 - 22
Ex vivo expansion of natural killer cells for clinical applications; Klingemann HG et al.; BACKGROUND: Immunotherapy with NK cells has been limited by the inability to obtain sufficient numbers of pure NK cells suitable for manipulation and expansion . The goal of this study was to isolate CD56(+) cells (CD3(-)/CD56(+), CD3(+)/CD56(+)) and expand them under culture conditions compliant with current good manufacturing practices . METHODS: Magnetic cell-selection technology, using paramagnetic CD56 microbeads and cell selection columns, was used to isolate a CD56(+) population containing both CD3(-)/56(+) NK (60.6+/-10.8%) and CD3(+)/56(+) NK T cells (30.4+/-8.6%) to initiate the expansion studies . The isolated CD56(+) cells were cultured in X-Vivo10 serum-free media supplemented with 10% human AB serum and 500 U/mL recombinant human IL-2 or 500 U/mL IL-2 plus 10 ng/mL recombinant human IL-15 for 14 days . Cultures were fed fresh media and cytokines every 3-4 days, and were evaluated for cell expansion, phenotype, and cytotoxicity at the end of the culture period . RESULTS: Significant expansion of CD56 cells occurred only during the second week of culture . Although an average of two log expansions was observed, there was substantial cell-expansion variability, depending on the donor, and even when the same donor was tested on different occasions . The cytotoxicity of selected and expanded CD56(+) cells at a low E:T ratio was significantly higher than the starting population, but was comparable to non-separated PBMC expanded for 2 weeks under the same conditions . IL-15 (in combination with IL-2) induced higher killing at the 1:1 E:T ratio than IL-2 alone . Since CD3 cells were not depleted upfront, the expansion of CD3(+)CD56(+) cells was 2-3 times that of CD3(-)CD56(+) cells . NK cells that express the FcgammaRIII (CD16) can mediate Ab-dependent cellular cytotoxicity, and can contribute to enhanced efficacy of MAb treatment . Under the given culture conditions, only moderate expansion of CD56(+)/CD3(-)/CD16(+) cells occurred, with the majority of cells being CD56(+)/CD3(+)/CD16(+) cells . DISCUSSION: Our studies suggest that the positive magnetic cell-separation method provides a good basis for obtaining enriched CD56(+) cells but expansion conditions need to be optimized.

Cytotherapy, 2004, 6(1), 1 - 6
A one-step large-scale method for T- and B-cell depletion of mobilized PBSC for allogeneic transplantation; Barfield RC et al.; BACKGROUND: The presence of T and B cells in allogeneic grafts contributes to GvHD and to EBV-associated lymphoproliferative disease (LPD) . Depletion of T and B cells from the graft decreases the risk of these complications . METHODS: T and B cells were depleted from mobilized peripheral stem cells from volunteer donors (n=5) using anti-CD3 and anti-CD19 Abs conjugated to magnetic microbeads, and the CliniMACS device . The function of the stem cells after depletion was evaluated using colony assays and non-obese diabetic (NOD)/SCID repopulating experiments . RESULTS: The mean mononuclear cell (MNC) count prior to T- and B-cell depletion was 2.19x10(10) (range 1.48-3.53) . After depletion, the mean percentage of contaminating T cells was 0.02% (range 0.01-0.04%) with a mean log(10) depletion of 3.4 (range 3-3.8) . The mean percentage of contaminating B cells was 0.1% (range 0.01-0.4%) with a mean log(10) depletion of 2.2 (range 1.4-3) . The mean recovery of CD3- and CD19-negative MNCs after depletion was 70% (range 54-88%) and the mean recovery of CD34(+) stem cells was 69% (range 52-98%) . The mean number of natural killer (NK) cells after T- and B-cell depletion was 5.2x10(8) (range 2-10x10(8)) . In vitro colony assays and in vivo NOD/SCID repopulation assays showed no negative impact of this method on the function of the hematopoietic stem cells . DISCUSSION: Our results show that the CliniMACS system can be used to efficiently deplete PBSC of T and B cells simultaneously, without adverse effect on the graft.

Cardiovasc Radiat Med, 2003 Jul-Sep, 4(3), 139 - 45
Could X-ray microbeams inhibit angioplasty-induced restenosis in the rat carotid artery?
Dilmanian FA, Kalef-Ezra J, Petersen MJ, Bozios G, Vosswinkel J, Giron F, Ren B, Yakupov R, Antonakopoulos G.
BACKGROUND: Parallel, thin (<100 microm) planes of synchrotron-generated X rays, have been shown to spare normal tissues and preferentially damage tumors in animal models . The aim of the present study was to assess the effect of such microbeams directed unidirectionally on angioplasted rat carotid arteries . METHODS AND MATERIALS: Three groups of Sprague-Dawley rats were studied: (a) rats with normal, untreated arteries, (b) rats treated by balloon angioplasty, but not irradiated, and (c) rats treated with balloon angioplasty and exposed to single fraction, unidirectional, parallel, microbeams an hour after angioplasty . The microbeam array, 15 mm widex7.6 mm high, consisting of 27-microm-wide beam slices, spaced 200 microm center-to-center laterally traversed the damaged artery . The in-depth in-beam dose was 150 Gy, the "valley" dose (dose midway between microbeams resulting mainly from X-ray scattering) was 4.5 Gy on average, and the "integrated" (averaged) dose was 26 Gy . RESULTS: Microbeam irradiation, as given in the present study, was tolerated, but was insufficient to significantly suppress the neointimal hyperplasia . DISCUSSION: The microbeam dose used is considered low . Dose escalation would be necessary to reach conclusive results regarding the X-ray microbeam efficacy to control restenosis.

FEMS Microbiol Rev, 2004 Feb, 28(1), 101 - 12
Exploiting the potential of insects for in vivo pathogenicity testing of microbial pathogens; Kavanagh K et al.; Conventional assays for quantifying the virulence of microbial pathogens and mutants have traditionally relied upon the use of a range of mammalian species . A number of workers have demonstrated that insects can be used for evaluating microbial pathogenicity and provide results comparable to those that can be obtained with mammals since one component of the vertebrate immune system, the innate immune response, remains similar to that found in insects . Larvae of the Greater Wax Moth Galleria mellonella have been used to evaluate the virulence of a range of bacterial and fungal pathogens and a correlation with the virulence of these microbes in mice has been established . This review highlights the similarities of the vertebrate and insect innate immune responses to infection and identifies the potential use of insects for the in vivo evaluation of the microbial pathogenicity.

Nature, 2004 Feb 19, 427(6976), 737 - 40
Multistability in the lactose utilization network of Escherichia coli; Ozbudak EM et al.; Multistability, the capacity to achieve multiple internal states in response to a single set of external inputs, is the defining characteristic of a switch . Biological switches are essential for the determination of cell fate in multicellular organisms, the regulation of cell-cycle oscillations during mitosis and the maintenance of epigenetic traits in microbes . The multistability of several natural and synthetic systems has been attributed to positive feedback loops in their regulatory networks . However, feedback alone does not guarantee multistability . The phase diagram of a multistable system, a concise description of internal states as key parameters are varied, reveals the conditions required to produce a functional switch . Here we present the phase diagram of the bistable lactose utilization network of Escherichia coli . We use this phase diagram, coupled with a mathematical model of the network, to quantitatively investigate processes such as sugar uptake and transcriptional regulation in vivo . We then show how the hysteretic response of the wild-type system can be converted to an ultrasensitive graded response . The phase diagram thus serves as a sensitive probe of molecular interactions and as a powerful tool for rational network design.

Nature, 2004 Feb 19, 427(6976), 731 - 3
Soil biota and exotic plant invasion; Callaway RM et al.; Invasive plants are an economic problem and a threat to the conservation of natural systems . Escape from natural enemies might contribute to successful invasion, with most work emphasizing the role of insect herbivores; however, microbial pathogens are attracting increased attention . Soil biota in some invaded ecosystems may promote 'exotic' invasion, and plant-soil feedback processes are also important . Thus, relatively rare species native to North America consistently demonstrate negative feedbacks with soil microbes that promote biological diversity, whereas abundant exotic and native species demonstrate positive feedbacks that reduce biological diversity . Here we report that soil microbes from the home range of the invasive exotic plant Centaurea maculosa L . have stronger inhibitory effects on its growth than soil microbes from where the weed has invaded in North America . Centaurea and soil microbes participate in different plant-soil feedback processes at home compared with outside Centaurea's home range . In native European soils, Centaurea cultivates soil biota with increasingly negative effects on the weed's growth, possibly leading to its control . But in soils from North America, Centaurea cultivates soil biota with increasingly positive effects on itself, which may contribute to the success of this exotic species in North America.

J Leukoc Biol, 2004 Jul, 76(1), 15 - 24 Epub 2004 Feb 13.
TGF-beta: the perpetrator of immune suppression by regulatory T cells and suicidal T cells; Wahl SM et al.; Innate and adaptive immunity function to eliminate foreign invaders and respond to injury while enabling coexistence with commensal microbes and tolerance against self and innocuous agents . Although most often effective in accomplishing these objectives, immunologic processes are not fail-safe and may underserve or be excessive in protecting the host . Checks and balances to maintain control of the immune system are in place and are becoming increasingly appreciated as targets for manipulating immunopathologic responses . One of the most recognized mediators of immune regulation is the cytokine transforming growth factor-beta (TGF-beta), a product of immune and nonimmune cells . Emerging data have unveiled a pivotal role for TGF-beta as a perpetrator of suppression by CD4(+)CD25(+) regulatory T (Treg) cells and in apoptotic sequelae . Through its immunosuppressive prowess, TGF-beta effectively orchestrates resolution of inflammation and control of autoaggressive immune reactions by managing T cell anergy, defining unique populations of Treg cells, regulating T cell death, and influencing the host response to infections.

Curr Top Med Chem, 2004, 4(5), 499 - 508
Molecules of infectious agents as immunomodulatory drugs; Lavelle EC et al.; Microbes produce a wide range of molecules that can modulate eukaryotic immune responses . These include factors that subvert protective mechanisms in order to facilitate pathogen colonization and persistence . Viral, bacterial and parasite-derived molecules have been identified that can inhibit inflammatory responses . However, in addition to the plethora of microbial factors that suppress immune responses, the most potent immune activators are also of microbial origin . These include the bacterial enterotoxins, parasite-derived excretory-secretory products and viral nucleic acids . In fact, there are examples of immune modulators that can exert either stimulatory or suppressive effects depending on the mode of delivery, dose and experimental model . There is presently great interest in the therapeutic exploitation of these factors, for example as a means to stimulate enhanced immune responses to a new generation of subunit vaccines or to inhibit deleterious immune mediated diseases . This short review, describes representative microbial immunomodulators, their modes of action and the potential for therapeutic application.

Planta, 2004 May, 219(1), 66 - 72 Epub 2004 Feb 12.
Class-1 hemoglobins, nitrate and NO levels in anoxic maize cell-suspension cultures; Dordas C et al.; Nitric oxide (NO) is a reactive gas involved in many biological processes of animals, plants and microbes . Previous work has demonstrated that NO is formed during hypoxia in alfalfa ( Medicago sativa L.) root cultures and that the levels of NO detected are inversely related to the levels of expression of class-1 hemoglobin expressed in the tissue . The objectives of this study were: to examine whether NO is produced in transgenic maize ( Zea mays L.) cell-suspension cultures exposed to anoxic growth conditions; to determine whether a similar relationship existed between a class-1 hemoglobin and the amount of NO detected under these conditions; and, to estimate the route of formation and breakdown of NO in the tissue . Maize cell-suspension cultures, transformed to express the sense or antisense strands of barley hemoglobin were used to overexpress or underexpress class-1 hemoglobin . A maize cell-suspension culture transformed with an empty vector was used as a control . Up to 500 nmol NO (g FW)(-1) was detected in maize cells exposed to low oxygen tensions for 24 h . The steady-state levels of NO in the different cell lines under anoxic conditions had an inverse relationship to the level of hemoglobin in the cells . There was no detectable NO produced under aerobic growth conditions . Spectroscopic data demonstrated that recombinant maize hemoglobin reacted with NO to form methemoglobin and NO(3)(-) . Nitrate was shown to be a precursor of NO in anoxic maize cell-suspension cultures by using (15)NO(3)(-) and electron paramagnetic resonance spectroscopy, suggesting that NO is formed via nitrate reductase during hypoxia . The results demonstrate that NO is produced in plant tissues grown under low oxygen tensions and suggest that class-1 hemoglobins have a significant function in regulating NO levels.

Gut, 2004 Mar, 53 Suppl 2, ii16 - 8
Interface between the intestinal environment and the nervous system; Lundgren O; Possible mechanisms by which the intestinal contents may influence afferent nerve fibres in the lamina propria of the intestinal mucosa are discussed in this brief review . After addressing intestinal epithelial permeability, endocrine and brush cells are discussed, as well as enterocytes, as sensors for luminal microbes.

Morfologiia, 2003, 124(5), 27 - 31
{Pathogenetic mechanisms of bacterial cholangitis in mechanical jaundice (experimental histological study)}; Tret'iakov AA et al.; In 89 rats with experimentally induced mechanical jaundice the infectious process was modeled by intragastric administration of E . coli and B . subtilis . The study included the investigation of the main routes and time intervals of dissemination, organ content of microbes labeled with 3H-thymidine, as well as of structural and functional peculiarities of the interaction of animal tissues with the microbes in cholestasis development dynamics and under the condition of biliary tract decompression . It was established that under the condition of mechanical jaundice viable bacteria from the alimentary tract penetrated the organs of peritoneal cavity through the blood and lymph flow . In the liver, the endogenous agents of surgical infections, and E . coli, in particular, were found to aggravate the inflammatory and necrobiotic processes . B . subtilis did not cause any inflammatory or other pathologic process in the tissue of liver.

Oncogene, 2004 Mar 25, 23(13), 2385 - 400
Genome-wide cDNA microarray analysis of gene expression profiles in pancreatic cancers using populations of tumor cells and normal ductal epithelial cells selected for purity by laser microdissection; Nakamura T et al.; To characterize molecular mechanism involved in pancreatic carcinogenesis, we analysed gene-expression profiles of 18 pancreatic tumors using a cDNA microarray representing 23,040 genes . As pancreatic ductal adenocarcinomas usually contain a low proportion of cancer cells in the tumor mass, we prepared 95% pure populations of pancreatic cancer cells by means of laser microbeam microdissection, and compared their expression profiles to those of similarly purified, normal pancreatic ductal cells . We identified 260 genes that were commonly upregulated and 346 genes that were downregulated in pancreatic cancer cells . Because of the high degree of purity in the cell populations, a large proportion of genes that we detected as upregulated or downregulated in pancreatic cancers were different from those reported in previous studies . Comparison of clinicopathological parameters with the expression profiles indicated that altered expression of 76 genes was associated with lymph-node metastasis and that of 168 genes with liver metastasis . In addition, expression levels of 30 genes were related to the recurrence of disease . These genome-wide expression profiles should provide useful information for finding candidate genes whose products might serve as specific tumor markers and/or as molecular targets for treatment of patients with pancreatic cancer.

Appl Environ Microbiol, 2004 Feb, 70(2), 1190 - 8
Two bacteria phylotypes are predominant in the Suiyo seamount hydrothermal plume; Sunamura M et al.; Microbial diversity and populations in a hydrothermal plume that was present inside the caldera of the Suiyo Seamount, a submarine volcano on the Izu-Bonin Arc, were investigated by performing a phylogenetic analysis of the 16S rRNA gene and by using fluorescence in situ hybridization (FISH) . Corresponding to transmissivity, an indicator of turbidity, the vertical total cell count as determined by 4',6'-diamidino-2-phenylindole (DAPI) staining varied from 5.6 x 10(4) to 1.1 x 10(5) cells ml(-1), and the apparent plume layer was assessed to be at a depth of 1,050 to 1,200 m inside the caldera and to contain 1.0 x 10(5) to 1.1 x 10(5) cells ml(-1) . From microbial samples collected in the plume by an in situ filtration system, the following two major phylogenetic groups, which were closely related to sulfur-oxidizing microbes, were obtained: the SUP05 group belonging to the gamma subclass of the Proteobacteria (13 of 20 clones) and the SUP01 group belonging to the epsilon subclass of the Proteobacteria (5 of 20 clones) . Specific oligonucleotide probes for these groups (SUP05-187 and SUP01-63) were designed and were used with various water samples obtained from the Suiyo Seamount . In the apparent plume layer, up to 66% of the total counts of microbial cells were estimated to be Bacteria cells that hybridized to EUB338, and few cells were identified by the archaeal probe ARCH915 . Almost all Bacteria cells were hard to identify with the known group-specific probes, such as ALF19, GAM42a, and CF319, while 88 to 90% of the Bacteria cells hybridized with SUP05-187 and >98% of them were considered members of the SUP05 and SUP01 populations . In a low-temperature vent fluid emitted from a bivalve-colonized mound, the SUP05 cells accounted for >99% of the Bacteria cells, suggesting that a portion of the plume cells originated on the surface of the seafloor at a depth of about 1,380 m . From further analysis of cell morphology (i.e., cell size and cell elongation index) we inferred that the SUP05 cells were active in the plume layer at a depth of 1,050 to 1,200 m compared to the activity in a near-bottom layer, while many elongated cells were found between these layers . These findings suggest that the morphology and distribution of SUP05 cells have complex relationships with hydrothermal activities and water circulation . Although growth and production rates remain to be defined, we concluded that this Suiyo Seamount caldera has functioned as a natural continuous incubator for these two phylotypes of Bacteria in an aphotic deep-sea environment.

Appl Environ Microbiol, 2004 Feb, 70(2), 1051 - 8
Spatial variation in frequency and intensity of antibiotic interactions among Streptomycetes from prairie soil; Davelos AL et al.; Antibiotic interactions are believed to be significant to microbial fitness in soil, yet little is known of the frequency, intensity, and diversity of antibiotic inhibition and resistance among indigenous microbes . To begin to address these issues, we studied the abilities of streptomycete isolates from prairie soil to inhibit growth and display resistance to antibiotics produced by a test collection of 10 streptomycete isolates . Wide variations in antibiotic inhibition and resistance for prairie isolates among three locations and four soil depths within a 1-m2 plot were revealed . Fewer than 10% of 153 prairie isolates inhibited all 10 test isolates, while more than 40% of the isolates did not inhibit any of the test isolates . No field isolate was resistant to all of the test isolates, nor was any isolate susceptible to all of the test isolates . No correlation between inhibition and resistance phenotypes was found, suggesting that inhibition and resistance are under independent selection . The significant spatial variation in the frequency and intensity of antibiotic inhibition implies that the fitness benefits of antibiotic production are not the same among locations in soil . In contrast, the consistency of resistance over space indicates that its significance to fitness across locations is stable or the costs of maintaining resistance in the absence of selection are small or nonexistent . The spatial clustering of antibiotic inhibitory activity suggests a variable matrix of selection pressures and microbial responses across the soil landscape.

Appl Environ Microbiol, 2004 Feb, 70(2), 831 - 6
Degradation pathway and generation of monohydroxamic acids from the trihydroxamate siderophore deferrioxamine B; Pierwola A et al.; Siderophores are avid ferric ion-chelating molecules that sequester the metal for microbes . Microbes elicit siderophores in numerous and different environments, but the means by which these molecules reenter the carbon and nitrogen cycles is poorly understood . The metabolism of the trihydroxamic acid siderophore deferrioxamine B by a Mesorhizobium loti isolated from soil was investigated . Specifically, the pathway by which the compound is cleaved into its constituent monohydroxamates was examined . High-performance liquid chromatography and mass-spectroscopy analyses demonstrated that M . loti enzyme preparations degraded deferrioxamine B, yielding a mass-to-charge (m/z) 361 dihydroxamic acid intermediate and an m/z 219 monohydroxamate . The dihydroxamic acid was further degraded to yield a second molecule of the m/z 219 monohydroxamate as well as an m/z 161 monohydroxamate . These studies indicate that the dissimilation of deferrioxamine B by M . loti proceeds by a specific, achiral degradation and likely represents the reversal by which hydroxamate siderophores are thought to be synthesized.

Science, 2004 Feb 6, 303(5659), 830 - 2
Role of microbes in the smectite-to-illite reaction; Kim J et al.; Temperature, pressure, and time have been thought to control the smectite-to-illite (S-I) reaction, an important diagenetic process used for petroleum exploration . We demonstrated that microorganisms can promote the S-I reaction by dissolving smectite through reduction of structural Fe(III) at room temperature and 1 atmosphere within 14 days . This reaction typically requires conditions of 300 degrees to 350 degrees C, 100 megapascals, and 4 to 5 months in the absence of microbial activity . These results challenge the conventional concept of the S-I reaction and of reaction kinetic models.

J Immunol, 2004 Feb 15, 172(4), 2201 - 9
IL-10-conditioned dendritic cells, decommissioned for recruitment of adaptive immunity, elicit innate inflammatory gene products in response to danger signals; Nolan KF et al.; Dendritic cells (DCs) are the professional APCs of the immune system, enabling T cells to perceive and respond appropriately to potentially dangerous microbes, while also being able to maintain T cell tolerance toward self . In part, such tolerance can be determined by IL-10 released from certain types of regulatory T cells . IL-10 has previously been shown to render DCs unable to activate T cells and it has been assumed that this process represents a general block in maturation . Using serial analysis of gene expression, we show that IL-10 pretreatment of murine bone marrow-derived DCs alone causes significant changes in gene expression . Furthermore, these cells retain the ability to respond to Toll-like receptor agonists, but in a manner skewed toward the selective induction of mediators known to enhance local inflammation and innate immunity, among which we highlight a novel CXCR2 ligand, DC inflammatory protein-1 . These data suggest that, while the presence of a protolerogenic and purportedly anti-inflammatory agent such as IL-10 precludes DCs from acquiring their potential as initiators of adaptive immunity, their ability to act as initiators of innate immunity in response to Toll-like receptor signaling is enhanced.

J Immunol, 2004 Feb 15, 172(4), 2016 - 20
Cutting edge: dendritic cells copulsed with microbial and helminth antigens undergo modified maturation, segregate the antigens to distinct intracellular compartments, and concurrently induce microbe-specific Th1 and helminth-specific Th2 responses; Cervi L et al.; To examine the ability of dendritic cells (DC) to discriminate between helminth and microbial Ag and induce appropriately polarized Th responses, mouse DC were copulsed with the helminth Ag, schistosome egg Ag (SEA), along with the bacterium Proprionebacterium acnes, Pa, and transferred into wild-type mice . Strikingly, SEA/Pa-copulsed DC induced concurrent Pa-specific Th1 (but not Th2) responses and SEA-specific Th2 (but not Th1) responses . Although DC exposed to both Ag undergo many of the maturation-associated changes that accompany exposure to Pa alone, Pa-induced IL-12 production was inhibited by SEA . Examination of Ag uptake revealed that SEA and Pa are acquired via discrete pathways and enter nonoverlapping intracellular compartments . Data suggest that segregation of SEA and Pa into distinct compartments, coupled with SEA-induced modifications of the DC maturation pathway, are significant components of the ability of DC to interpret signals inherent to SEA and Pa and induce appropriately polarized Th responses.

Mol Microbiol, 2004 Feb, 51(4), 917 - 24
Cellular engineering in a minimal microbe: structure and assembly of the terminal organelle of Mycoplasma pneumoniae; Krause DC et al.; Mycoplasma pneumoniae is a minimal microbe with respect to cell envelope composition, biosynthetic and regulatory capabilities and genome size, yet it possesses a remarkably complex, multifunctional terminal organelle . This membrane-bound extension of the mycoplasma cell is defined by the presence of an electron-dense core that appears as paired, parallel bars oriented longitudinally and enlarging at the distal end to form a terminal button . Most non-cytadhering mutants of M . pneumoniae isolated to date exhibit defects in the architecture of the terminal organelle . Detailed characterization of those mutants has revealed the identities of many component proteins of the terminal organelle as well as the likely order in which some of those components are required . Additional questions regarding the composition of the electron-dense core, the means by which the terminal organelle is duplicated during cell division and the manner in which this process is regulated remain to be answered . Thus, it seems that there is much to be learned about cellular engineering and spatial regulation in these 'simple' cell wall-less bacteria.

Am J Physiol Cell Physiol, 2004 Mar, 286(3), C518 - 28
Extracellular matrix controls myosin light chain phosphorylation and cell contractility through modulation of cell shape and cytoskeletal prestress; Polte TR et al.; The mechanism by which vascular smooth muscle (VSM) cells modulate their contractility in response to structural cues from extracellular matrix remains poorly understood . When pulmonary VSM cells were cultured on increasing densities of immobilized fibronectin (FN), cell spreading, myosin light chain (MLC) phosphorylation, cytoskeletal prestress (isometric tension in the cell before vasoagonist stimulation), and the active contractile response to the vasoconstrictor endothelin-1 all increased in parallel . In contrast, MLC phosphorylation did not increase when suspended cells were allowed to bind FN-coated microbeads (4.5-microm diameter) or cultured on micrometer-sized (30 x 30 microm) FN islands surrounded by nonadhesive regions that support integrin binding but prevent cell spreading . Cell spreading and MLC phosphorylation also both decreased in parallel when the mechanical compliance of flexible FN substrates was raised . MLC phosphorylation was inhibited independently of cell shape when cytoskeletal prestress was dissipated using a myosin ATPase inhibitor in fully spread cells, whereas it increased to maximal levels when microtubules were disrupted using nocodazole in cells adherent to FN but not in suspended cells . These data demonstrate that changes in cell-extracellular matrix (ECM) interactions modulate smooth muscle cell contractility at the level of biochemical signal transduction and suggest that the mechanism underlying this regulation may involve physical interplay between ECM and the cytoskeleton, such that cell spreading and generation of cytoskeletal tension feed back to promote MLC phosphorylation and further increase tension generation.

Expert Rev Vaccines, 2004 Feb, 3(1), 59 - 76
Genome-derived vaccines; De Groot AS et al.; Vaccine research entered a new era when the complete genome of a pathogenic bacterium was published in 1995 . Since then, more than 97 bacterial pathogens have been sequenced and at least 110 additional projects are now in progress . Genome sequencing has also dramatically accelerated: high-throughput facilities can draft the sequence of an entire microbe (two to four megabases) in 1 to 2 days . Vaccine developers are using microarrays, immunoinformatics, proteomics and high-throughput immunology assays to reduce the truly unmanageable volume of information available in genome databases to a manageable size . Vaccines composed by novel antigens discovered from genome mining are already in clinical trials . Within 5 years we can expect to see a novel class of vaccines composed by genome-predicted, assembled and engineered T- and Bcell epitopes . This article addresses the convergence of three forces--microbial genome sequencing, computational immunology and new vaccine technologies--that are shifting genome mining for vaccines onto the forefront of immunology research.

Anal Bioanal Chem, 2004 May, 379(2), 234 - 41 Epub 2004 Jan 31.
Monitoring ground-level air for trace analysis: methods and results; Bieringer J et al.; Trace analysis enables the sensitive detection of radionuclide concentrations in ground-level air in the range of microbecquerel per cubic meter (microBq m(-3)) . Typical sampling intervals of less than one day up to a few days can be used in routine operation . Trace analysis measurements are performed in the framework of the German Integrated Measuring and Information system (IMIS) and the International Monitoring System (IMS) used for verification of the Comprehensive Nuclear-Test-Ban Treaty (CTBT) . Within the environmental monitoring programmes of the German IMIS the Federal Office for Radiation Protection (BfS) performs measurements of aerosol-bound radionuclides and of radioactive noble gases in the atmosphere . Aerosols are collected on filters with high-volume air samplers and analysed by gamma-spectrometry, alpha-spectrometry, and integral measurements of beta-activity, with preceding radiochemical separation . Noble gas samples from 15 sites world-wide are analysed to observe the (85)Kr-release from nuclear fuel reprocessing plants and from seven sites in Germany to monitor the (133)Xe emitted from nuclear power plants . As part of the International Monitoring System (IMS) of the CTBT an automatic aerosol sampling and measuring system and an automatic noble gas sampling and measuring system will be operated by the BfS at Mount Schauinsland near Freiburg . Because of its expertise in noble gas measurements the BfS had been chosen to perform an intercomparison experiment in the BfS laboratory in Freiburg with several automatic noble gas sampling and measurement systems before their installation at IMS sites . To establish quality-assurance programmes for trace analysis performed for the German IMIS close collaboration between the involved German institutions has been established . First steps have been taken to expand cooperation to other European laboratories . Informal data exchange already occurs between trace-analysis laboratories in Europe (Ring of Five) and helps in cases of enhanced activity concentrations to get a rapid overview of the radiological situation and to identify possible sources.

J Allergy Clin Immunol, 2004 Mar, 113(3), 395 - 400 Epub 2004 Feb 01.
Immunologic influences on allergy and the TH1/TH2 balance; Romagnani S; TH2 cell-mediated immune responses against "innocuous" antigens play a triggering role in atopic allergy . Several epidemiologic studies have clearly shown that the reduced microbial exposure of children caused by the westernized lifestyle is responsible for the increased prevalence of allergy that has occurred in the last decades in developed countries ("hygiene hypothesis") . The immunologic changes caused by the reduced exposure to pathogenic and nonpathogenic microbes during childhood are still controversial . The initial interpretation has been a lack of shift of allergen-specific responses from the TH2 to the TH1 phenotype . This is because of reduced production of IL-12 and IFNs by cells of the natural immunity stimulated by bacterial products through their Toll-like receptors (missing immune deviation) . Another interpretation emphasizes the importance of reduced activity of T-regulatory cells (reduced immune suppression) . However, although there are impressive amounts of data in favor of the missing immune deviation, experimental evidence supporting the role of reduced immune suppression in explaining the increased prevalence of allergy is currently weak or even contradictory . The solution to this question is very important not only from a theoretic point of view but also because of its therapeutic implications.

World J Gastroenterol, 2004 Feb 1, 10(3), 389 - 92
Viral replication modulated by synthetic peptide derived from hepatitis B virus X protein; Song CZ et al.; AIM: A strategy for viral vaccine design is the use of conserved peptides to overcome the problem of sequence diversity . At present it is still unclear whether conserved peptide is safe as a candidate vaccine . We reported it here for the first time not only to highlight the biohazard issue and safety importance for viral peptide vaccine, but also to explore the effect of a fully conserved peptide on HBV replication within the carboxyl terminus of HBx . METHODS: We synthesized the fully conserved peptide (CP) with nine residues, FVLGGCRHK . HBV-producing 2.2.15 cells were treated with or without 3.5 microM CP for 36 hours . Quantitative detection of viral DNA was performed by real-time PCR . HBV antigens were determined by enzyme-linked immunoadsorbent assay (ELISA) . Quantitative analyses of p53 and Bax proteins were based on immunofluorescence . Flow cytometry was performed to detect cell cycle and apoptosis . RESULTS: Both extracellular and intracellular copies of HBV DNA per ml were significantly increased after incubation with 3.5 microM of CP . HBsAg and HBeAg in the cultured medium of CP-treatment cells were as abundant as untreated control cells . CP influenced negatively the extracellular viral gene products, and 3.5 microM CP could significantly inhibit intracellular HBsAg expression . In response to CP, intracellular HBeAg displayed an opposite pattern to that of HBsAg, and 3.5 microM CP could efficiently increase the level of intracellular HBeAg . Flow cytometric analyses exhibited no significant changes on cell cycle, apoptosis, p53 and Bax proteins in 2.2.15 cells with or without CP . CONCLUSION: Together with the results generated from the synthetic peptide, we address that the conserved region, a domain of HBx, may be responsible for modulating HBV replication . As conserved peptides from infectious microbes are used as immunogens to elicit immune responses, their latent biological hazard for human beings should be evaluated.

Clin Rev Allergy Immunol, 2004 Feb, 26(1), 15 - 24
Parasites and the hygiene hypothesis: regulating the immune system?
Yazdanbakhsh M, Matricardi PM.
The hygiene hypothesis proposes that the stimulation of the immune system by microbes or microbial products protects from the development of inflammatory diseases; therefore a reduced exposure to infectious agents may explain the rise in allergic and autoimmune diseases in industrialized countries . The contribution of studies on parasites and allergy to our understanding of the hygiene hypothesis has been two-fold . First, several studies have shown an inverse association between exposure to (Toxplasma gondii) or harbouring of parasites (Schistosoma or Intestinal helminths) and allergy . Second, the mechanisms behind such protective effects have provided new insights and theories on the ability of parasite derived molecules to down-regulate immune responses and thereby to control inflammatory diseases such as allergies . In this review, recent findings and new concepts relating to the associations between parasites and allergies at the epidemiological, cellular and molecular level are discussed.

Curr Med Chem, 2004 Jan, 11(2), 241 - 52
Alpha-galactosylceramide: potential immunomodulatory activity and future application; Hayakawa Y et al.; alpha-Galactosylceramide (alpha-GalCer), is a glycolipid which has been identified as a ligand recognized by a special group of immune T cells, known as invariant NKT cells . alpha-GalCer can powerfully activate invariant NKT cells to produce immunoregulatory cytokines, including interferon-gamma and IL-4, and thereby exert a variety of subsequent effects on other cells in the immune system . Recent studies have revealed the mechanism of alpha-GalCer-induced iNKT cell-activation in immune responses to tumors and microbes, and in the suppression of autoimmune diseases . In this review, we discuss the potential immunomodulatory activity of alpha-GalCer and its possible future application for clinical studies in humans.

Int J Phytoremediation, 2003, 5(4), 367 - 79
Phenotypic characterization of microbes in the rhizosphere of Alyssum murale; Abou-Shanab RI et al.; Metal hyperaccumulator plants like Alyssum murale are used for phytoremediation of Ni contaminated soils . Soil microorganisms are known to play an important role in nutrient acquisition for plants, however, little is known about the rhizosphere microorganisms of hyperaccumulators . Fresh and dry weight, and Ni and Fe concentrations in plant shoots were higher when A . murale was grown in non-sterilized compared to sterilized soils . The analysis of microbial populations in the rhizosphere of A . murale and in bulk soils demonstrated that microbial numbers were affected by the presence of the plant . Significantly higher numbers of culturable actinomycetes, bacteria and fungi were found in the rhizosphere compared to bulk soil . A higher percent of Ni-resistant bacteria were also found in the rhizosphere compared to bulk soil . Percentage of acid producing bacteria was higher among the rhizosphere isolates compared to isolates from bulk soil . However, proportions of siderophore producing and phosphate solubilizing bacteria were not affected by the presence of the plant . We hypothesize that microbes in the rhizosphere of A . murale were capable of reducing soil pH leading to an increase in metal uptake by this hyperaccumulator.

Acta Microbiol Immunol Hung, 2003, 50(4), 321 - 30
The molecular basis of infectious diseases: pathogenicity islands and other mobile genetic elements . A review; Hacker J et al.; Bacterial genomes generally consist of stable regions termed core genome, and variable regions that form the so-called flexible gene pool . The flexible part is composed of bacteriophages, plasmids, transposons as well as unstable large regions that have been termed genomic islands . Genomic islands encoding virulence factors of pathogenic bacteria have been designated "pathogenicity islands" . Pathogenicity islands were first discovered in uropathogenic Escherichia coli and presently more than 30 bacterial species carrying pathogenicity islands have been described . This review summarises the current knowledge on bacterial genomic islands and their general features, and discusses their putative role in the evolution of microbes in the light of genomics of pathogenic bacteria.

EMBO Rep, 2004 Feb, 5(2), 207 - 12 Epub 2004 Jan 23.
Parasite-specific immune response in adult Drosophila melanogaster: a genomic study; Roxstrom-Lindquist K et al.; Insects of the order Diptera are vectors for parasitic diseases such as malaria, sleeping sickness and leishmania . In the search for genes encoding proteins involved in the antiparasitic response, we have used the protozoan parasite Octosporea muscaedomesticae for oral infections of adult Drosophila melanogaster . To identify parasite-specific response molecules, other flies were exposed to virus, bacteria or fungi in parallel . Analysis of gene expression patterns after 24 h of microbial challenge, using Affymetrix oligonucleotide microarrays, revealed a high degree of microbe specificity . Many serine proteases, key intermediates in the induction of insect immune responses, were uniquely expressed following infection of the different organisms . Several lysozyme genes were induced in response to Octosporea infection, while in other treatments they were not induced or downregulated . This suggests that lysozymes are important in antiparasitic defence.

J Trauma, 2004 Jan, 56(1), 45 - 51
Patterns of microbiology in intra-abdominal packing for trauma; Granchi TS et al.; INTRODUCTION: This study tracks the microbiology of packs and infections in damage-control trauma patients to determine whether the packs cause infections . METHODS: The peritoneum and abdominal packs were cultured in patients who survived to re-operation . The study recorded all positive cultures, pack count, packing duration, number of operations, and infections . RESULTS: Thirty-five patients were studied . Twenty-eight patients survived; seven died . Packs were cultured in 29 patients . Data for 291 cultures collected . Pack cultures were positive in 20 patients and negative in nine . Positive pack cultures grew skin and gut flora . Twenty-one patients had infections, 14 did not . Organisms from positive pack cultures did not contribute to subsequent infections or mortality . Microbes and sites of infections were consistent with SICU patients . CONCLUSIONS: Intra-abdominal packs are contaminated with skin and gut flora . These contaminants, however, do not contribute to subsequent infections . Pathogens from subsequent infections were typical for ICU infections.

Biotechnol Bioeng, 2004 Feb 5, 85(3), 283 - 92
Exogenous cadherin microdisplay can interfere with endogenous signaling and reprogram gene expression in cultured hepatocytes; Brieva TA et al.; We recently found that the basal micro substrate presentation of E-cadherin, a key cell-cell adhesion molecule in the liver, can modulate hepatocellular proliferative potential and differentiated function (Brieva and Moghe, in press) . In the current study, we established a similar experimental model involving rat hepatocytes cultured on collagen and incorporated 5 microm polystyrene microbeads functionalized with Protein A-anchored E-cadherin/human lgG Fc chimeric fusion constructs . We investigated the cadherin governed dose-response of cell proliferative potential and quantified the underlying changes in intracellular gene signaling processes . Hepatocellular proliferative potential was found to be intensified with an increase in the microdisplay of acellular cadherins and this effect was offset by increased cell seeding density . Notably, we report that following overnight exposure to acellular cadherins, the expression of genes known to mediate the control of cell proliferation, cyclin D1 and c-myc, was upregulated, while the expression of differentiation-related genes, namely albumin and cytochrome p450 II B1, was reduced . The exposure of cell cultures to exogenous cadherins was found to markedly disrupt the localization of endogenous E-cadherin and beta-catenin to junctions at cell-cell contacts and cause a quantitative decrease in the endogenous cadherin protein levels . Based on all of our observations, we propose that the acellular presentation of E-cadherin chimeras competitively disrupts endogenous cadherin containing complexes at cell-cell junctions and increases intracellular cadherin turnover, thereby promoting beta-catenin mediated signaling, which ultimately engenders an increase in cell proliferative potential and a decrease in differentiated function .

J Nutr, 2004 Feb, 134(2), 465 - 72
Molecular ecological analysis of the gastrointestinal microbiota: a review; Zoetendal EG et al.; The gastrointestinal (GI) microbiota of mammals is characterized by its high population density, wide diversity and complexity of interactions . While all major groups of microbes are represented, bacteria predominate . Importantly, bacterial cells outnumber animal (host) cells by a factor of ten and have a profound influence on nutritional, physiological and immunological processes in the host animal . Our knowledge of the molecular and cellular bases of host-microbe interactions is limited, though critically needed to determine if and how the GI microbiota contributes to various enteric disorders in humans and animals . Traditionally, GI bacteria have been studied via cultivation-based techniques, which are labor intensive and require previous knowledge of individual nutritional and growth requirements . Recently, findings from culture-based methods have been supplemented with molecular ecology techniques that are based on the 16S rRNA gene . These techniques enable characterization and quantification of the microbiota, while also providing a classification scheme to predict phylogenetic relationships . The choice of a particular molecular-based approach depends on the questions being addressed . Clone libraries can be sequenced to identify the composition of the microbiota, often to the species level . Microbial community structure can be analyzed via fingerprinting techniques, while dot blot hybridization or fluorescent in situ hybridization can measure abundance of particular taxa . Emerging approaches, such as those based on functional genes and their expression and the combined use of stable isotopes and biomarkers, are being developed and optimized to study metabolic activities of groups or individual organisms in situ . Here, a critical summary is provided of current molecular ecological approaches for studying the GI microbiota.

Rev Mal Respir, 2003 Dec, 20(6 Pt 1), 928 - 39
{Secretory immunity of the airways}; Maury G et al.; INTRODUCTION: Continuous exposure of the respiratory tract to inhaled particles and microbes implies the presence of effective defence mechanisms at a bronchial and alveolar level . STATE OF ART: Among the mechanisms involved secretory mucosal immunity contributes considerably to the defence of the bronchial tree . This immunity depends essentially on the active trans-epithelial transport of IgA involved in both innate non-specific and acquired specific immunity . Recently an IgA receptor has been identified on the surface of phagocytes including alveolar macrophages, establishing a link between alveolar and bronchial defences . PERSPECTIVES: The respiratory mucosa represents a crucial interface between the host and its environment, and should provide in the future a new target for the development of diagnostic and therapeutic tools . CONCLUSIONS: Beyond its function as an anatomical barrier the bronchial epithelium possesses a secretory activity that is essential for the protection of the lung . Despite a better understanding of mucosal immunity this secretory activity and in particular the part played by IgA remains to be elucidated.

Biotechniques, 2004 Jan, 36(1), 139 - 47
Enriching for HIV-infected cells using anti-gp41 antibodies indirectly conjugated to magnetic microbeads; Bahbouhi B et al.; The isolation of a pure population of human immunodeficiency virus (HIV)-infected cells is highly desirable for evaluating the impact of HIV on cellular gene expression . Given that HIV gp41 transmembrane protein is anchored on the surface of HIV-infected cells, we evaluated the use of pooled anti-gp41 monoclonal antibodies (MAbs) and HIV immunoglobulins (HIV-Igs) indirectly conjugated to magnetic microbeads to positively select for infected cells . We demonstrate that pooled anti-gp41 monoclonal antibodies enriched for H9 cells infected with HIV IIIB by approximately 98% . Peripheral blood mononuclear cells (PBMCs) infected with a primary (HIV strain 302151) or laboratory-adapted (IIIB) strain were enriched by 54%-62%, depending on the initial viral inoculum . Using HIV-Ig in this magnetic positive-selection approach was also highly efficient for enriching for H9 cells infected with IIIB but less efficient for infected PBMCs . Both types of antibodies used in the selection process resulted in > 80 viability of selected HIV-infected cells . Analysis of interleukin 2 (IL-2) mRNA expression using real-time reverse transcription PCR (RT-PCR) of the HIV-enriched population demonstrated a higher level of IL-2 mRNA, by approximately four cycles, and an 8-fold increase in IL-2 expression, as evaluated by intracellular staining and flow cytometric analysis, in comparison to gp41-negative cells . Collectively, these data illustrate that antibodies targeting gp41 can be used to enrich for HIV-positive populations . This represents a novel approach for studying the impact of HIV on infected cells and on bystander/uninfected cells.

Clin Exp Rheumatol, 2003 Nov-Dec, 21(6 Suppl 32), S49 - 54
Immune phenomena in localized and generalized Wegener's granulomatosis; Mueller A et al.; Wegener's granulomatosis (WG) is characterized by granulomatous inflammation and systemic vasculitis with a predilection for the lungs and kidneys . In most patients WG begins with a localized organ involvement of the upper respiratory tract that progresses to systemic disease (generalized WG) (1) . Because of the life-threatening nature of systemic vasculitis, much effort has concentrated on elucidating the pathogenesis of the vasculitis . However, based upon a renewed interest in (innate) immune defenses against microbes, a better understanding of the chronic granulomatous inflammation may contribute to a more precise insight into the early genesis of WG . Thus, this review focuses on summarizing and discussing data for a potential pattern of disease, i.e . from localized to generalized WG with a special emphasis on granulomatous lesions of the upper respiratory tract and their alterations during the disease course.

Oecologia, 2004 Mar, 139(1), 98 - 107 Epub 2004 Jan 22.
Decomposition rate of organic substrates in relation to the species diversity of soil saprophytic fungi; Setala H et al.; Despite the great interest concerning the relationship between species diversity and ecosystem functioning, there is virtually no knowledge as to how the diversity of decomposer microbes influences the decomposition rate of soil organic matter . We established a microcosm study in which the number of soil fungi was investigated in relation to the system's ability to (i) degrade raw coniferous forest humus, and (ii) use resources that were either added to the systems or released into the soils after a disturbance (drought) . With the exception of the most diverse treatment, in each of the six replicates of each of the six diversity treatments (1, 3, 6, 12, 24 or 43 taxa), fungal taxa were randomly chosen from a pool of 43 commonly isolated fungal species of raw humus . Two months after initiation of the study CO2 production increased as fungal diversity increased, but in the species-poor end of the diversity gradient only . Addition of various energy resources to the microcosms generally increased the level of soil respiration but did not affect the shape of the diversity-CO2-production curve . Rewetting the soil after severe drought resulted in a rapid flush of CO2, particularly in the most diverse communities . The biomass of the fungi in the non-disturbed soils, and soil NH4-N concentration and soil pH in both disturbed and non-disturbed systems were slightly but significantly higher in the diverse than in the simple systems . Fungal species richness had no influence on the organic matter content of the humus at the end of the experiment . The results suggest that the functional efficiency of fungal communities can increase with the number of fungal taxa . This diversity effect was, however, significant at the species-poor end of the diversity gradient only, which implies considerable functional equivalency (redundancy) among the decomposer fungi.

Arch Environ Health, 2003 May, 58(5), 275 - 83
Microbial exposure and mold-specific serum IgG levels among children with respiratory symptoms in 2 school buildings; Hyvarinen A et al.; In this study, the authors determined the association between serum mold-specific immunoglobulin G (IgG) levels in primary school children (N = 181) with asthma, wheezing, or cough symptoms and exposure to indoor mold in 2 schools, with and without mold damage . Microbial exposure was determined with environmental sampling . Serum IgG antibody concentrations against 24 microbial strains were determined with enzyme-linked immunosorbent assay . Children's microbe-specific IgG levels were often higher in the reference school . There was an association between elevated serum IgG antibody levels to Penicillium notatum and moisture damage in the school . In addition, moisture damage both in school and at home was associated with Penicillium notatum and Eurotium amstelodami IgG levels . These observations comported with microbial findings in the index school . No other positive associations between IgG antibodies and microbial findings for the school buildings were observed . IgG levels in sera of school children did not provide a specific means for assessing the microbial exposure associated with moisture and microbial problems in the school buildings . Thus, IgG analysis cannot be readily suggested as a routine method for the evaluation of these exposures.

Biochem J, 2004 May 1, 379(Pt 3), 765 - 75
Rhesus monkey gastric mucins: oligomeric structure, glycoforms and Helicobacter pylori binding; Linden S et al.; Mucins isolated from the stomach of Rhesus monkey are oligomeric glycoproteins with a similar mass, density, glycoform profile and tissue localization as human MUC5AC and MUC6 . Antibodies raised against the human mucins recognize those from monkey, which thus appear to be orthologous to those from human beings . Rhesus monkey muc5ac and muc6 are produced by the gastric-surface epithelium and glands respectively, and occur as three distinct glycoforms . The mucins are substituted with the histo blood-group antigens B, Le(a) (Lewis a), Le(b), Le(x), Le(y), H-type-2, the Tn-antigen, the T-antigen, the sialyl-Le(x) and sialyl-Le(a) structures, and the expression of these determinants varies between individuals . At neutral pH, Helicobacter pylori strains expressing BabA (blood-group antigen-binding adhesin) bind Rhesus monkey gastric mucins via the Le(b) or H-type-1 structures, apparently on muc5ac, as well as on a smaller putative mucin, and binding is inhibited by Le(b) or H-type-1 conjugates . A SabA (sialic acid-binding adhesin)-positive H . pylori mutant binds to sialyl-Le(x)-positive mucins to a smaller extent compared with the BabA-positive strains . At acidic pH, the microbe binds to mucins substituted by sialylated structures such as sialyl-Le(x) and sialylated type-2 core, and this binding is inhibited by DNA and dextran sulphate . Thus mucin- H . pylori binding occurs via at least three different mechanisms: (1) BabA-dependent binding to Le(b) and related structures, (2) SabA-dependent binding to sialyl-Le(x) and (3) binding through a charge-mediated mechanism to sialylated structures at low pH values.

Radiat Environ Biophys, 2004 Feb, 42(4), 237 - 45 Epub 2004 Jan 20.
Microirradiation of cells with energetic heavy ions; Hauptner A et al.; The ion microprobe SNAKE at the Munich 14 MV tandem accelerator achieves beam focussing by a superconducting quadrupole doublet and can make use of a broad range of ions and ion energies, from 20 MeV protons to 200 MeV gold ions . Because of these properties, SNAKE is particularly attractive for biological microbeam experiments . Here we describe the adaptation of SNAKE for microirradiation of cell samples . This includes enlarging of the focal distance in order to adjust the focal plane to the specimen stage of a microscope, construction of a beam exit window in a flexible nozzle and of a suitable cell containment, as well as development of procedures for on-line focussing of the beam, preparation of single ions and scanning by electrostatic deflection of the beam . When irradiating with single 100 MeV (16)O ions, the adapted set-up permits an irradiation accuracy of 0.91 microm (full width at half maximum) in the x-direction and 1.60 microm in the y-direction, as demonstrated by retrospective track etching of polycarbonate foils . Accumulation of the repair protein Rad51, as detected by immunofluorescence, was used as a biological track detector after irradiation of HeLa cells with geometric patterns of counted ions . Observed patterns of fluorescence foci agreed reasonably well with irradiation patterns, indicating successful adaptation of SNAKE . In spite of single ion irradiation, we frequently observed split fluorescence foci which might be explained by small-scale chromatin movements.

Clin Cancer Res, 2004 Jan 1, 10(1 Pt 1), 76 - 83
Phase I trial of large multivalent immunogen derived from melanoma lysates in patients with disseminated melanoma; Mitchell MS et al.; PURPOSE: The purpose of this research was to determine the toxicity and immunological activity of large multivalent immunogen (LMI), a preparation of tumor cell membranes affixed to amorphous silica microbeads, in patients with melanoma . Experimental Design: Nineteen patients with metastatic (stage IV) melanoma were entered into the study, of whom 15 received the full 3 months of treatment with LMI . LMI was administered without adjuvant, one-half intradermally (i.d.) and the other half s.c . Because we expected little toxicity, we first treated 2 patients at each dose level, 10-, 30-, or 100-million tumor cell equivalents on weeks 0, 4, and 8, and subsequently randomized the remaining 13 patients to receive treatment with one of those dosage schedules, for a total of 19 patients . Two patients who were registered were found to be ineligible because of brain metastases, and 2 others did not complete the course of treatment for reasons other than toxicity . Thus, 15 patients were fully evaluable . Patients with evidence of a clinical response (at least stable disease at the 12-week checkpoint) had the option of continuing treatment at 4-week intervals . Frequencies of cytolytic T cell precursors against HLA-A2 matched melanoma cells, and delayed-type hypersensitivity to a melanoma cell membrane preparation from a component melanoma cell line were performed to measure immunological efficacy, and serum chemistries and complete blood counts were performed every 2 weeks throughout the study to measure possible toxicity . Computed tomography scans were performed pretreatment and at week 12 to measure possible beneficial effects on known lesions . RESULTS: Eight of the 15 evaluable patients had an increase in cytolytic T-cell precursors during the course of therapy, usually by day 42 . No patient had demonstrable delayed-type hypersensitivity to a melanoma membrane preparation before or after treatment . No toxicity of any kind was observed . A degree of clinical effectiveness of LMI was suggested by the elicitation of stable disease in 5 patients at 12 weeks . One patient had >50% regression of a lung nodule but progression of disease to the brain, whereas a second patient had a bona fide partial remission of a 3-cm diameter solitary lung nodule . CONCLUSIONS: LMI was nontoxic, improved immunological reactivity to melanoma cells, and showed evidence of clinical effectiveness (shrinkage of tumor) in 1 patient . Additional studies with LMI with added adjuvant materials, in melanoma and other cancers, appear warranted.

J Eukaryot Microbiol, 2003 Nov-Dec, 50(6), 383 - 402
Extrusomes in ciliates: diversification, distribution, and phylogenetic implications; Rosati G et al.; Exocytosis is, in all likelihood, an important communication method among microbes . Ciliates are highly differentiated and specialized micro-organisms for which versatile and/or sophisticated exocytotic organelles may represent important adaptive tools . Thus, in ciliates, we find a broad range of different extrusomes, i.e ejectable membrane-bound organelles . Structurally simple extrusomes, like mucocysts and cortical granules, are widespread in different taxa within the phylum . They play the roles in each case required for the ecological needs of the organisms . Then, we find a number of more elaborate extrusomes, whose distribution within the phylum is more limited, and in some way related to phylogenetic affinities . Herein we provide a survey of literature and our data on selected extrusomes in ciliates . Their morphology, distribution, and possible function are discussed . The possible phylogenetic implications of their diversity are considered.

Ying Yong Sheng Tai Xue Bao, 2003 Sep, 14(9), 1512 - 6
{Effect of plastic film mulching on soil microbial biomass in spring wheat field in semi-arid loess area}; Song Q et al.; This paper studied the effect of different periods of plastic film mulching (M0-no mulching, M30-mulching for 30 days, M60-mulching for 60 days, and M-mulching for whole growth period) on soil microbial biomass carbon (SMBC) of spring wheat field in semi-arid loess plateau . The mean SMBC in 1999 and 2000 was 335.3 and 259.3 mg.kg-1 dry soil, respectively . It was 29.3% higher in 1999 than in 2000 . The highest SMBC was recorded at the harvest stage in M treatment for the two years . In 1999, a wet year with more rainfall, the SMBC of M60 and M treatments was significantly higher than those of M0 and M30 in the mid-period of growth, reached its top at the end of the growth period . The highest grain yield was also achieved in M60 . It was a dry year in 2000, but rainfall was rich in the latter growth period of spring wheat . SMBC increased at the beginning period of growth, and did not increase during the mid-growth period . It increased again during the latter period of growth, and showed a significant difference among the treatments . At the harvest stage of 2000, SMBC in M0 was the highest among all the treatments . It was similar between M and M60, and lower than that of M30 . In the two years, the ratio of C/N ranged between 7.732-9.042, being lower than the threshold of 11.3, and the ratio of C/P was 300.8-719.6, being higher than the threshold of 300 . The two parameters showed that the increase of SMBC was inhibited because of the competition of soil available nutrients between soil microbes and crops . These indicated that soil organic matter content was so shortage that it became the key factor to restrict crop productivity . Under this condition, increasing crop productivity through the input of chemical fertilizers would conceal the problem of soil degradation, and result in a further decrease of soil quality . A long term plastic film mulching would make the problem more serious.

Ying Yong Sheng Tai Xue Bao, 2003 Sep, 14(9), 1507 - 11
{Parameters of energy flow in wheat-corn intercropping agroecosystem}; Wang J et al.; The parameters of energy flow of primary producers (agricultural crop and weed), consumers (pests and their natural enemies) and decomposer (soil microbe, protozoan and saprozoite) in wheat-corn intercropping agroecosystem in Tibet of China were studied by field investigation and laboratory measurement . The results showed that the wheat-corn intercropping agroecosystem had high primary productivity and energy translation efficiency, and the degression rate of energy flow through food chain was also high . At each nutritive layer of this agroecosystem, relatively less energy was required to product unit energy, and degree of energy utilization of superior nutritive layer from lower nutritive layer was increased gradually.

Trends Cell Biol, 1995 Mar, 5(3), 105 - 9
Phagocytic processing of antigens for presentation by MHC molecules; Harding CV; Phagocytosis plays a major role in the defence of higher organisms against microbial infection not only by allowing ingested microbes to be destroyed by microbicidal mechanisms, but also by providing the basis for processing of their antigens to forms that generate immune responses . This article examines the role of the phagolysosome in antigen processing, and discusses the contributions of both MHC class II and MHC class I molecules to the presentation of antigens derived from phagocytosed material.

Mar Biotechnol (NY), 2003 Sep-Oct, 5(5), 450 - 7
Grouping newly isolated docosahexaenoic acid-producing thraustochytrids based on their polyunsaturated fatty acid profiles and comparative analysis of 18S rRNA genes; Huang J et al.; Seven strains of marine microbes producing a significant amount of docosahexaenoic acid (DHA; C22:6, n-3) were screened from seawater collected in coastal areas of Japan and Fiji . They accumulate their respective intermediate fatty acids in addition to DHA . There are 5 kinds of polyunsaturated fatty acid (PUFA) profiles which can be described as (1) DHA/docosapentaenoic acid (DPA; C22:5, n-6), (2) DHA/DPA/eicosapentaenoic acid (EPA; C20:5, n-3), (3) DHA/EPA, (4) DHA/DPA/EPA/arachidonic acid (AA; C20:4, n-6), and (5) DHA/DPA/EPA/AA/docosatetraenoic acid (C22:4, n-6) . These isolates are proved to be new thraustochytrids by their specific insertion sequences in the 18S rRNA genes . The phylogenetic tree constructed by molecular analysis of 18S rRNA genes from the isolates and typical thraustochytrids shows that strains with the same PUFA profile form each monophyletic cluster . These results suggest that the C20-22 PUFA profile may be applicable as an effective characteristic for grouping thraustochytrids.

Biosci Biotechnol Biochem, 2003 Dec, 67(12), 2655 - 7
Specific incorporation of L-glutamine into volicitin in the regurgitant of Spodoptera litura; Yoshinaga N et al.; Volicitin, {N-(17-hydroxylinolenoyl)-L-glutamine}, was identified as an elicitor of plant volatiles from a Spodoptera exigua regurgitant . It has been proposed that gut microbes synthesize volicitin from glutamine, a predominant amino acid component in the insect gut . However, we found that glutamine was not a major component in the regurgitant of Spodoptera litura, although L-glutamine was exclusively incorporated into volicitin by S . litura fed on diets enriched with various amino acids . This selectivity of glutamine as a substrate was not due to a dominant occurrence in the insect gut.

J Appl Physiol, 2004 May, 96(5), 1701 - 13 Epub 2004 Jan 16.
Role of heat shock protein 27 in cytoskeletal remodeling of the airway smooth muscle cell; An SS et al.; Remodeling of the airway smooth muscle (ASM) cell has been proposed to play an important role in airway hyperresponsiveness . Using a functional assay, we have assessed remodeling of the cultured rat ASM cell and the role of heat shock protein (HSP) 27 in that process . To probe remodeling dynamics, we measured spontaneous motions of an individual Arg-Gly-Asp-coated microbead that was anchored to the cytoskeleton . We reasoned that the bead could not move unless the microstructure to which it is attached rearranged; if so, then its mean square displacement (MSD) would report ongoing internal reorganizations over time . Each bead displayed a random, superdiffusive motion; MSD increased with time as approximately t(1.7), whereas an exponent of unity would be expected for a simple passive diffusion . Increasing concentrations of cytochalasin-D or latrunculin-A caused marked increases in the MSD, whereas colchicine did not . Treatments with PDGF or IL-1beta, but not transforming growth factor-beta, caused decreases in the MSD, the extent of which rank-ordered with the relative potency of these agents in eliciting the phosphorylation of HSP27 . The chemical stressors anisomycin and arsenite each increased the levels of HSP27 phosphorylation and, at the same time, decreased bead motions . In particular, arsenite prevented and even reversed the effects of cytochalasin-D on bead motions . Finally, ASM cells overexpressing phospho-mimicking human HSP27, but not wild-type or phosphorylation-deficient HSP27, exhibited decreases in bead motions that were comparable to the arsenite response . Taken together, these results show that phosphorylated HSP27 favors reduced bead motions that are probably due to stabilization of the actin cytoskeleton.

Trends Plant Sci, 2004 Jan, 9(1), 26 - 32
How plants communicate using the underground information superhighway; Bais HP et al.; The rhizosphere is a densely populated area in which plant roots must compete with invading root systems of neighboring plants for space, water, and mineral nutrients, and with other soil-borne organisms, including bacteria and fungi . Root-root and root-microbe communications are continuous occurrences in this biologically active soil zone . How do roots manage to simultaneously communicate with neighboring plants, and with symbiotic and pathogenic organisms within this crowded rhizosphere? Increasing evidence suggests that root exudates might initiate and manipulate biological and physical interactions between roots and soil organisms, and thus play an active role in root-root and root-microbe communication.

Proc R Soc Lond B Biol Sci, 2003 Dec 22, 270(1533), 2573 - 8
Evolutionary dynamics of escape from biomedical intervention; Iwasa Y et al.; Viruses, bacteria, eukaryotic parasites, cancer cells, agricultural pests and other inconvenient animates have an unfortunate tendency to escape from selection pressures that are meant to control them . Chemotherapy, anti-viral drugs or antibiotics fail because their targets do not hold still, but evolve resistance . A major problem in developing vaccines is that microbes evolve and escape from immune responses . The fundamental question is the following: if a genetically diverse population of replicating organisms is challenged with a selection pressure that has the potential to eradicate it, what is the probability that this population will produce escape mutants? Here, we use multi-type branching processes to describe the accumulation of mutants in independent lineages . We calculate escape dynamics for arbitrary mutation networks and fitness landscapes . Our theory shows how to estimate the probability of success or failure of biomedical intervention, such as drug treatment and vaccination, against rapidly evolving organisms.

Anal Chem, 2004 Jan 15, 76(2), 309 - 14
Microchemical element imaging of yeast and human cells using synchrotron X-ray microprobe with Kirkpatrick-Baez optics; Ortega R et al.; Trace element imaging and speciation analysis in cells and subcellular compartments is a challenging and important objective for modern analytical chemistry in order to better understand the biological chemistry of essential and toxic elements . A focusing system based on Kirkpatrick-Baez design optics mounted on a synchrotron radiation scanning X-ray microscope has been developed at the ESRF and was used for trace element quantitative imaging in single cells . The focused microbeam (1.3 x 3.2 microm(2)) obtained in that way led to a photon flux as bright as 1.5 x 10(11) photons/s at 14 keV . The absolute detection limit of this analytical probe, as measured on standard reference materials, was shown to be 2 x 10(-)(17) g for most elements . Chemical maps of human carcinoma and of Saccharomyces cerevisiae cells were obtained for minor (P, S, Cl, K) and trace elements (Fe, Zn) . Within human cancer cells, chemical elements are homogeneously distributed at the current spatial resolution and correlated with the sample's mass, except Fe, which shows micrometer-sized structures around the cell nucleus, and Zn, which slightly concentrates in the nucleus, while chemical maps of S . cerevisiae show homogeneous pattern distribution at the cellular level.

Genome Res, 2004 Feb, 14(2), 239 - 46 Epub 2004 Jan 12.
Closing the gaps on human chromosome 19 revealed genes with a high density of repetitive tandemly arrayed elements; Leem SH et al.; The reported human genome sequence includes about 400 gaps of unknown sequence that were not found in the bacterial artificial chromosome (BAC) and cosmid libraries used for sequencing of the genome . These missing sequences correspond to approximately 1% of euchromatic regions of the human genome . Gap filling is a laborious process because it relies on analysis of random clones of numerous genomic BAC or cosmid libraries . In this work we demonstrate that closing the gaps can be accelerated by a selective recombinational capture of missing chromosomal segments in yeast . The use of both methodologies allowed us to close the four remaining gaps on the human chromosome 19 . Analysis of the gap sequences revealed that they contain several abnormalities that could result in instability of the sequences in microbe hosts, including large blocks of micro- and minisatellites and a high density of Alu repeats . Sequencing of the gap regions, in both BAC and YAC forms, allowed us to generate a complete sequence of four genes, including the neuronal cell signaling gene SCK1/SLI . The SCK1/SLI gene contains a record number of minisatellites, most of which are polymorphic and transmitted through meiosis following a Mendelian inheritance . In conclusion, the use of the alternative recombinational cloning system in yeast may greatly accelerate work on closing the remaining gaps in the human genome (as well as in other complex genomes) to achieve the goal of annotation of all human genes.

Environ Sci Technol, 2003 Dec 15, 37(24), 5511 - 7
Model of microbial transport and inactivation in the surf zone and application to field measurements of total coliform in Northern Orange County, California; Boehm AB; The classic model of pollutant transport in the surf zone of a long, sandy beach developed by Inman et al . (J . Geophys . Res . 1971, 76, 3493) is altered to account for first-order pollutant inactivation in an effort to understand how rip cell dilution and bacterial inactivation control the length of shoreline adversely impacted by microbial pollution from a point source . A dimensionless number gamma dictates whether physical processes (dilution of microbes in the surf zone by rip cell mixing) or biological processes (microbial inactivation) control the distribution of pollution along the shoreline . Estimates of gamma for beaches in Northern Orange County, California, indicate that dilution is the primary factor controlling total coliform levels surrounding two drains that release nuisance runoff directly onto the beach . It is also shown that, even when alongshore currents are fast, pollutant levels will drop e-fold at distances under 4000 m from the point source due to dilution alone . Because dilution is ultimately controlled by wave climate and shoreline morphology, the results suggest the strategic position of drains and other point sources in high dilution wave environments will reduce potential adverse effects on beach water quality . In addition, the results stress the importance of understanding hydrodynamics when conducting microbial source tracking at wave-dominated marine beaches.

Biotech Histochem, 2003 Jun-Aug, 78(3-4), 191 - 9
Histological investigation of organisms with hard skeletons: a case study of siliceous sponges; Hoffmann F et al.; Siliceous and calcareous sponges commonly are treated with acid to remove the spicules prior to embedding and cutting for histological investigations . Histology of spiculated sponge tissue represents a challenging problem in sponge histotechnology . Furthermore, fluorescence in situ hybridization (FISH), a key method for studying sponge-associated microbes, is not possible after acid treatment . For a broad range of siliceous sponge species, we developed and evaluated methods for embedding in paraffin, methylmethacrylate resins, LR White resin and cryomatrix . Different methods for cutting tissue blocks as well as mounting and staining sections also were tested . Our aim was to enable histological investigations and FISH without prior removal of the spicules . To obtain an overview of tissue and skeleton arrangement, we recommend embedding tissue blocks with LR White resin combined with en bloc staining techniques for large specimens with thick and numerous spicules, but paraffin embedding and subsequent staining for whole small specimens . For FISH on siliceous sponges, we recommend Histocryl embedding if the spicule content is high, but paraffin embedding if it is low . Classical histological techniques are used for detailed tissue examinations.

Bull Exp Biol Med, 2003 Oct, 136(4), 355 - 7
Phagocytic activity and state of bactericidal systems in polymorphonuclear leukocytes from patients with Alzheimer's disease; Davydova TV et al.; Phagocytic activity of peripheral blood neutrophils underwent phase changes in patients with Alzheimer's disease . Neutrophils retained the ability to engulf microbes, but their digestive activity decreased at the early stage of Alzheimer's disease . At the late stage we observed a decrease in the count of phagocytizing neutrophils, reduction of myeloperoxidase activity, and increase in the content of cationic proteins.

Chromosome Res, 2003, 11(8), 771 - 86
'Signalling' between chromosomes in crane-fly spermatocytes studied using ultraviolet microbeam irradiation; Wong R et al.; The present article deals with signals from kinetochores in anaphase crane-fly spermatocytes: when a half-bivalent's kinetochore is irradiated with an ultraviolet microbeam during anaphase, all half-bivalents in the cell stop moving to both poles . Movement blockage is temporary, and different half-bivalent pairs resume movement at different times . Movement stoppage presumably is due to signals arising from the irradiated kinetochores and transmitted to the 'motors' of the other chromosomes . We used a second irradiation (of the interzone) to determine the path of the signal . We reasoned that if irradiation of the interzone blocked transmission of the putative signal, then those chromosomes not receiving the signal should continue to move after irradiation of a kinetochore . Interzone irradiation interfered with the signal in about 20% of the 51 cells irradiated doubly, in that chromosome(s) moving to one pole stopped while chromosome(s) moving to the other pole continued . There was a second indication that interzonal irradiation blocked the signal: in about 30% of the cells in which the kinetochore was irradiated first and interzone second, all half-bivalents resumed movement immediately after the second irradiation.

Appl Environ Microbiol, 2004 Jan, 70(1), 468 - 74
Bacterial diversity in agricultural soils during litter decomposition; Dilly O et al.; Denaturing gradient gel electrophoresis (DGGE) of amplified fragments of genes coding for 16S rRNA was used to study the development of bacterial communities during decomposition of crop residues in agricultural soils . Ten strains were tested, and eight of these strains produced a single band . Furthermore, a mixture of strains yielded distinguishable bands . Thus, DGGE DNA band patterns were used to estimate bacterial diversity . A field experiment performed with litter in nylon bags was used to evaluate the bacterial diversity during the decomposition of readily degradable rye and more refractory wheat material in comparable luvisols and cambisols in northern, central, and southern Germany . The amount of bacterial DNA in the fresh litter was small . The DNA content increased rapidly after the litter was added to the soil, particularly in the rapidly decomposing rye material . Concurrently, diversity indices, such as the Shannon-Weaver index, evenness, and equitability, which were calculated from the number and relative abundance (intensity) of the bacterial DNA bands amplified from genes coding for 16S rRNA, increased during the course of decomposition . This general trend was not significant for evenness and equitability at any time . The indices were higher for the more degradation-resistant wheat straw than for the more easily decomposed rye grass . Thus, the DNA band patterns indicated that there was increasing bacterial diversity as decomposition proceeded and substrate quality decreased . The bacterial diversity differed for the sites in northern, central, and southern Germany, where the same litter material was buried in the soil . This shows that in addition to litter type climate, vegetation, and indigenous microbes in the surrounding soil affected the development of the bacterial communities in the litter.

Appl Environ Microbiol, 2004 Jan, 70(1), 393 - 403
Analysis of dissimilatory sulfite reductase and 16S rRNA gene fragments from deep-sea hydrothermal sites of the Suiyo Seamount, Izu-Bonin Arc, Western Pacific; Nakagawa T et al.; This study describes the occurrence of unique dissimilatory sulfite reductase (DSR) genes at a depth of 1,380 m from the deep-sea hydrothermal vent field at the Suiyo Seamount, Izu-Bonin Arc, Western Pacific, Japan . The DSR genes were obtained from microbes that grew in a catheter-type in situ growth chamber deployed for 3 days on a vent and from the effluent water of drilled holes at 5 degrees C and natural vent fluids at 7 degrees C . DSR clones SUIYOdsr-A and SUIYOdsr-B were not closely related to cultivated species or environmental clones . Moreover, samples of microbial communities were examined by PCR-denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rRNA gene . The sequence analysis of 16S rRNA gene fragments obtained from the vent catheter after a 3-day incubation revealed the occurrence of bacterial DGGE bands affiliated with the Aquificae and gamma- and epsilon-Proteobacteria as well as the occurrence of archaeal phylotypes affiliated with the Thermococcales and of a unique archaeon sequence that clustered with "Nanoarchaeota." The DGGE bands obtained from drilled holes and natural vent fluids from 7 to 300 degrees C were affiliated with the delta-Proteobacteria, genus Thiomicrospira, and Pelodictyon . The dominant DGGE bands retrieved from the effluent water of casing pipes at 3 and 4 degrees C were closely related to phylotypes obtained from the Arctic Ocean . Our results suggest the presence of microorganisms corresponding to a unique DSR lineage not detected previously from other geothermal environments.

Curr Biol, 2004 Jan 6, 14(1), R30 - 2
Immune activation: death, danger and dendritic cells; Pulendran B; Dendritic cells are critical for host immunity, and sense microbes with pathogen recognition receptors . New evidence indicates that these cells also sense uric acid crystals in dead cells, suggesting that the immune system is conscious not only of pathogens, but also of death and danger.

Microb Ecol, 2003 Aug, 46(2), 228 - 37
Comparison of free-living and particle-associated bacterial communities in a coastal lagoon; LaMontagne MG et al.; We analyzed, by terminal restriction fragment length polymorphisms (T-RFLPs) of PCR-amplified 16S rDNA, microbial diversity in water collected during the dry and wet seasons in a human-impacted coastal lagoon . Water samples were fractionated by prefiltration to differentiate particle-associated and free-living microbes . From a sample collected during the dry season, prefiltration removed 23 to 44% of bacteria, as assessed by direct counts and MPN, and 99% of phytoplankton, as assessed by chlorophyll a . Restriction with RsaI yielded fewer peaks than restriction with HhaI . Diversity indices calculated from T-RFLPs were higher in the lagoon than adjoining coastal waters and higher in the particle-associated than the free-living fraction . In the dry season, peaks found only in bulk and particle-associated T-RFLPs were consistent with plastid and cyanobacterial ribotypes . These peaks matched those observed in the sequence of a clone generated from the bulk fraction with plastid and cyanobacterial specific primers . This clone appeared related to plastids found in the diatom genus Skeletonema . Principal component analysis of T-RFLPs suggested that the difference between the free-living and particle-associated fractions in the dry season was less than temporal variability in this lagoon and that these fractions varied significantly only in the wet season . This fractionation of microbial populations into particle-associated and free-living guilds during the wet season, when water residence time in the lagoon is relatively low, suggests an external source of particle-associated bacteria such as erosion of upland soils by runoff.

J Microbiol Methods, 2004 Jan, 56(1), 3 - 15
Real-time reverse transcription PCR analysis of expression of atrazine catabolism genes in two bacterial strains isolated from soil; Devers M et al.; The level of expression of highly conserved, plasmid-borne, and widely dispersed atrazine catabolic genes (atz) was studied by RT-qPCR in two telluric atrazine-degrading microbes . RT-qPCR assays, based on the use of real-time PCR, were developed in order to quantify atzABCDEF mRNAs in Pseudomonas sp . ADP and atzABC mRNAs in Chelatobacter heintzii . atz gene expression was expressed as mRNA copy number per 10(6) 16S rRNA . In Pseudomonas sp . ADP, atz genes were basally expressed . It confirmed atrazine-degrading kinetics indicating that catabolic activity starts immediately after adding the herbicide . atz gene expression increased transitorily in response to atrazine treatment . This increase was only observed while low amount of atrazine remained in the medium . In C . heintzii, only atzA was basally expressed . atzA and atzB expression levels were similarly and significantly increased in response to atrazine treatment . atzC was not expressed even in the presence of high amounts of atrazine . This study showed that atz genes are basally expressed and up-regulated in response to atrazine treatment . atz gene expression patterns are different in Pseudomonas ADP and C . heintzii suggesting that the host may influence the expression of plasmid-borne atrazine-catabolic potential.

J Agric Food Chem, 2003 Jul 30, 51(16), 4719 - 21
Flumioxazin soil persistence and mineralization in laboratory experiments; Ferrell JA et al.; Flumioxazin is an herbicide registered for use in soybean and peanut . However, few published papers concerning the soil persistence of flumioxazin are available . Therefore, laboratory studies were initiated to determine the half-life (t(1/2)) of flumioxazin in Greenville sandy clay loam and Tifton loamy sand soils when incubated at 15 and 25 degrees C . Results indicated that temperature had little effect on flumioxazin persistence . The t(1/2) for the Greenville soil was 17.9 and 16.0 days while the Tifton soil was 13.6 and 12.9 days, at 15 and 25 degrees C, respectively . These data correspond to the greater clay content of the Greenville soil (32%) as compared to the Tifton soil (2%) . Therefore, the Greenville soil had greater soil adsorption and less flumioxazin was generally available to be degraded by soil microorganisms . In soils that were heat treated to reduce microbe populations, 99% of initial flumioxazin was accounted for after 16 days . Mineralization of flumioxazin, measured as 14CO2 evolution, was also greater in the Tifton soil (2.2% after 64 days) than in the Greenville soil (2.0% after 64 days) . From these data, it was concluded that microbes were the most influential factor concerning the degradation of flumioxazin.

Microbiology, 2004 Jan, 150(Pt 1), 171 - 80
Demonstration of antifreeze protein activity in Antarctic lake bacteria; Gilbert JA et al.; Antifreeze proteins (AFPs) are a structurally diverse group of proteins that have the ability to modify ice crystal structure and inhibit recrystallization of ice . AFPs are well characterized in fish and insects, but very few bacterial species have been shown to have AFP activity to date . Thirty eight freshwater to hypersaline lakes in the Vestfold Hills and Larsemann Hills of Eastern Antarctica were sampled for AFPs during 2000 . Eight hundred and sixty six bacterial isolates were cultivated . A novel AFP assay, designed for high-throughput analysis in Antarctica, demonstrated putative activity in 187 of the cultures . Subsequent analysis of the putative positive isolates showed 19 isolates with significant recrystallization inhibition (RI) activity . The 19 RI active isolates were characterized using ARDRA (amplified rDNA restriction analysis) and 16S rDNA sequencing . They belong to genera from the alpha- and gamma-Proteobacteria, with genera from the gamma-subdivision being predominant . The 19 AFP-active isolates were isolated from four physico-chemically diverse lakes . Ace Lake and Oval Lake were both meromictic with correspondingly characteristic chemically stratified water columns . Pendant Lake was a saline holomictic lake with different chemical properties to the two meromictic lakes . Triple Lake was a hypersaline lake rich in dissolved organic carbon and inorganic nutrients . The environments from which the AFP-active isolates were isolated are remarkably diverse . It will be of interest, therefore, to elucidate the evolutionary forces that have led to the acquisition of functional AFP activity in microbes of the Vestfold Hills lakes and to discover the role the antifreezes play in these organisms.

Histol Histopathol, 2004 Jan, 19(1), 251 - 8
Attachment of the soluble complement regulator factor H to cell and tissue surfaces: relevance for pathology; Jozsi M et al.; Complement is a central element of innate immunity and this vital defense system initiates and coordinates immediate immune reactions which attack and eliminate microbes, foreign particles and altered self cells . Newly generated activation products are extremely toxic and consequently, activation is highly restricted in terms of time and space . The initial activation of the alternative complement pathway occurs continuously and the early phase acts indiscriminatoryl and forms on any surface . However, the system discriminates between self and foreign, and therefore allows activation on foreign surfaces e.g . microbes, and restricts activation on host cells . Consequently, self cells and tissues are protected from the harmful activation products . This protection is mediated by specific regulators or inhibitors, which exist in the fluid phase and/or in membrane-bound forms . Here we review a novel mechanism, i.e . the attachment of the soluble complement regulator factor H to the surface of self cells . This attachment, which is demonstrated experimentally by means of immunofluorescense microscopy and by flow cytometry, increases the inhibitory potential at the cell surface and mediates protection by reducing the local formation of toxic inflammatory products . This attachment is highly relevant and has pathophysiological consequences in several human diseases, including Factor H-associated hemolytic uremic syndrome (FH-HUS), membrano-proliferative glomerulonephritis type II, recurrent microbial infections and chronic inflammation, e.g . rheumatoid arthritis and immune evasion of tumor cells . Defects of this safeguard activity have been recently understood in patients with FH-HUS . Point mutations in the Factor H gene occurring in the C-terminus of the protein result in impaired cell binding capacity of Factor H and, consequently, during an inflammatory insult endothelial cells are not properly protected and are damaged.

Biochem Biophys Res Commun, 2004 Jan 16, 313(3), 758 - 64
Mechanical properties of individual focal adhesions probed with a magnetic microneedle; Matthews BD et al.; A permanent magnetic microneedle was developed to apply tensional forces to integrin receptors via ligand-coated magnetic microbeads while optically analyzing the mechanical properties of individual focal adhesions . Force application (130 pN for 3 s) through activated beta1 integrins produced less bead displacement than when unligated integrins were stressed . This strengthening response differed markedly on a bead-by-bead basis, correlated directly with local focal adhesion assembly, and was similar when analyzed at 4 degrees C, indicating that it was due to passive material properties of the cell . Viscoelastic analysis clarified that recruitment of focal adhesion proteins increased the local elastic stiffness of the adhesion complex without changing its viscous behavior . These data indicate that individual focal adhesions exhibit distinct mechanical properties that depend upon local focal adhesion assembly, and that these local variations in micromechanics can be detected and analyzed within living cells using the permanent magnetic microneedle technique.

Bioinformatics, 2004 Jan 1, 20(1), 115 - 9
Essentiality and damage in metabolic networks; Lemke N et al.; Understanding the architecture of physiological functions from annotated genome sequences is a major task for postgenomic biology . From the annotated genome sequence of the microbe Escherichia coli, we propose a general quantitative definition of enzyme importance in a metabolic network . Using a graph analysis of its metabolism, we relate the extent of the topological damage generated in the metabolic network by the deletion of an enzyme to the experimentally determined viability of the organism in the absence of that enzyme . We show that the network is robust and that the extent of the damage relates to enzyme importance . We predict that a large fraction (91%) of enzymes causes little damage when removed, while a small group (9%) can cause serious damage . Experimental results confirm that this group contains the majority of essential enzymes . The results may reveal a universal property of metabolic networks.

Infect Immun, 2004 Jan, 72(1), 498 - 507
Survival strategy of obligately intracellular Ehrlichia chaffeensis: novel modulation of immune response and host cell cycles; Zhang JZ et al.; Ehrlichia chaffeensis is an obligatory intracellular bacterium which resides in an early endosome in monocytes . E . chaffeensis infection in a human monocyte cell line (THP1) significantly altered the transcriptional levels of 4.5% of host genes, including those coding for apoptosis inhibitors, proteins regulating cell differentiation, signal transduction, proinflammatory cytokines, biosynthetic and metabolic proteins, and membrane trafficking proteins . The transcriptional profile of the host cell revealed key themes in the pathogenesis of Ehrlichia . First, E . chaffeensis avoided stimulation of or repressed the transcription of cytokines involved in the early innate immune response and cell-mediated immune response to intracellular microbes, such as the interleukin-12 (IL-12), IL-15, and IL-18 genes, which might make Ehrlichia a stealth organism for the macrophage . Second, E . chaffeensis up-regulated NF-kappaB and apoptosis inhibitors and differentially regulated cell cyclins and CDK expression, which may enhance host cell survival . Third, E . chaffeensis also inhibited the gene transcription of RAB5A, SNAP23, and STX16, which are involved in membrane trafficking . By comparing the transcriptional response of macrophages infected with other bacteria and that of macrophages infected with E . chaffeensis, we have identified few genes that are commonly induced and no commonly repressed genes . These results illustrate the stereotyped macrophage response to other pathogens, in contrast with the novel host response to obligate intracellular Ehrlichia, whose survival depends entirely on a long evolutionary process of outmaneuvering macrophages.

Blood Rev, 2004 Mar, 18(1), 65 - 73
The intriguing contribution of white blood cells to sickle cell disease - a red cell disorder; Okpala I; Sickle cell disease (SCD) is characterized by a point mutation that replaces adenine with thymidine in the sixth codon of the beta-globin gene, a unique morphological abnormality of red blood cells, vaso-occlusion with ischaemic tissue injury, and susceptibility to infections . Vascular lumen obstruction in SCD results from interaction of erythrocytes, leukocytes, platelets, plasma proteins, and the vessel wall . The disease phenotype is a product of various genes and environmental factors acting in concert with the protein lesion underlying the red cell anomaly . The severity of SCD increases with leukocyte count . The biological basis and therapeutic implications of this relationship are discussed . Leukocytes contribute to SCD by adhering to blood vessel walls and obstructing the lumen, aggregating with other blood cells with more effective blockage of the lumen, stimulating the vascular endothelium to increase its expression of ligands for adhesion molecules on blood cells, and causing tissue damage and inflammatory reaction which predispose to vaso-occlusion . Patients with impaired ability of leukocytes to kill microbes are more prone to infections; which precipitate sickle cell crisis . Reduction of leukocyte count ameliorates SCD . Similarly, targeted blockade or reduced synthesis of specific leukocyte adhesion molecules and their ligands might confer clinical benefit in SCD.

Cancer Res, 2003 Dec 1, 63(23), 8437 - 42
Nitric oxide-mediated signaling in the bystander response of individually targeted glioma cells; Shao C et al.; Bystander responses have been reported to be a major determinant of the response of cells to radiation exposure at low doses, including those of relevance to therapy . In this study, human glioblastoma T98G cell nuclei were individually irradiated with an exact number of helium ions using a single-cell microbeam . It was found that when only 1 cell in a population of approximately 1200 cells was targeted, with one or five ions, cellular damage measured as induced micronuclei was increased by 20% . When a fraction from 1% to 20% of cells were individually targeted, the micronuclei yield in the population greatly exceeded that predicted on the basis of the micronuclei yield when all of the cells were targeted assuming no bystander effect was occurring . However when 2-(4-carboxyphenyl)-4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO), a nitric oxide (NO)-specific scavenger was present in the culture medium, the micronuclei yields reduced to the predicted values, which indicates that NO contributes to the bystander effect . By using 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF-FM), NO was detected in situ, and it was found that NO-induced fluorescence intensity in the irradiated population where 1% of cell nuclei were individually targeted with a single helium ion was increased by 1.13 +/- 0.02-fold (P < 0.005) relative to control with approximately 40% of the cells showing increased NO levels . Moreover, the medium harvested from helium ion-targeted cells showed a cytotoxic effect by inducing micronuclei in unirradiated T98G cells, and this bystander response was also inhibited by c-PTIO treatment . The induction of micronuclei in the population could also be decreased by c-PTIO treatment when 100% of cells were individually targeted by one or two helium ions, indicating a complex interaction of direct irradiation and bystander signals.

Astrobiology, 2003 Fall, 3(3), 609 - 18
Modern and ancient extremely acid saline deposits: terrestrial analogs for martian environments?
Benison KC, LaClair DA.
Extremely acid (pH <1) saline lakes and groundwaters existed in the mid-Permian of the mid-continent of North America . Modern counterparts have been found in acid saline lake systems throughout southern Australia . We compare and contrast the Permian Opeche Shale of North Dakota and Nippewalla Group of Kansas to modern Australian salt lakes in southern Western Australia and in northwest Victoria . With the exception of some minor variations in pH, evaporite mineralogy, and water geochemistry, the Permian and modern systems are similar and characterized by: (1) ephemeral saline continental playas hosted by red siliciclastic sediments, (2) evaporite minerals, including abundant sulfates, (3) Al-Fe-Si-rich waters with low pH values, (4) acidophilic microbes, and (5) paucity of carbonates . The composition of these terrestrial systems is strikingly similar to compositional data returned from the martian surface . Specifically, both Earth and martian systems have high amounts of iron oxides and sulfates, and little, if any, carbonates . We propose that the modern and ancient terrestrial acid saline environments may be good analogs for possible environments on Mars.

Astrobiology, 2003 Fall, 3(3), 505 - 29
Near-infrared detection of potential evidence for microscopic organisms on Europa; Dalton JB et al.; The possibility of an ocean within the icy shell of Jupiter's moon Europa has established that world as a primary candidate in the search for extraterrestrial life within our Solar System . This paper evaluates the potential to detect evidence for microbial life by comparing laboratory studies of terrestrial microorganisms with measurements from the Galileo Near Infrared Imaging Spectrometer (NIMS) . If the interior of Europa at one time harbored life, some evidence may remain in the surface materials . Examination of laboratory spectra of terrestrial extremophiles measured at cryogenic temperatures reveals distorted, asymmetric nearinfrared absorption features due to water of hydration . The band centers, widths, and shapes of these features closely match those observed in the Europa spectra . These features are strongest in reddish-brown, disrupted terrains such as linea and chaos regions . Narrow spectral features due to amide bonds in the microbe proteins provide a means of constraining the abundances of such materials using the NIMS data . The NIMS data of disrupted terrains exhibit distorted, asymmetric near-infrared absorption features consistent with the presence of water ice, sulfuric acid octahydrate, hydrated salts, and possibly as much as 0.2 mg cm(-3) of carbonaceous material that could be of biological origin . However, inherent noise in the observations and limitations of spectral sampling must be taken into account when discussing these findings.

Clin Rheumatol, 2003 Dec, 22(6), 381 - 5 Epub 2003 Oct 14.
Joint symptoms and diseases associated with moisture damage in a health center; Luosujarvi RA et al.; Rheumatic diseases do not usually cluster in time and space . It has been proposed that environmental exposures may initiate autoimmune responses . We describe a cluster of rheumatic diseases among a group of health center employees who began to complain of symptoms typically related to moldy houses, including mucocutaneous symptoms, nausea and fatigue, within a year of moving into a new building . Dampness was found in the insulation space of the concrete floor below ground level . Microbes indicating mold damage and actinobacteria were found in the flooring material and in the outer wall insulation . The case histories of the personnel involved were examined . All 34 subjects working at the health center had at least some rheumatic complaints . Two fell ill with a typical rheumatoid factor (RF)-positive rheumatoid arthritis (RA), and 10 had arthritis that did not conform to any definite arthritic syndrome (three met the classification criteria for RA) . Prior to moving into the problem building one subject had suffered reactive arthritis, which had then recurred . Another employee had undiagnosed ankylosing spondylitis and later developed psoriatic arthritis, and another developed undifferentiated vasculitis . A total of 16 subjects developed joint pains, 11 of these after beginning work at the health center . Three subjects developed Raynaud's symptom . Fourteen cases had elevated levels of circulating immune complexes in 1998, 17 in 1999, but there were only three cases in 2001, when the health center had been closed for 18 months . The high incidence of joint problems among these employees suggests a common triggering factor for most of the cases . As some of the symptoms had tended to subside while the health center was closed, the underlying causes are probably related to the building itself and possibly to the abnormal microbial growth in its structures.

Naturwissenschaften, 2003 Dec, 90(12), 558 - 62 Epub 2003 Nov 04.
The infrabuccal pellet piles of fungus-growing ants; Little AE et al.; Fungus-growing ants (Attini) live in an obligate mutualism with the fungi they cultivate for food . Because of the obligate nature of this relationship, the success of the ants is directly dependent on their ability to grow healthy fungus gardens . Attine ants have evolved complex disease management strategies to reduce their garden's exposure to potential parasitic microbes, to prevent the establishment of infection in their gardens, and to remove infected garden sections . The infrabuccal pocket, a filtering device located in the oral cavity of all ants, is an integral part of the mechanisms that leaf-cutter ants use to prevent the invasion and spread of general microbial parasites and the specific fungal-garden parasite Escovopsis . Fungus-growing ants carefully groom their garden, collecting general debris and pathogenic spores of Escovopsis in their infrabuccal pocket, the contents of which are later expelled in dump chambers inside the nest or externally . In this study we examined how a phylogenetically diverse collection of attine ants treat their infrabuccal pellets . Unlike leaf-cutters that deposit their infrabuccal pellets directly in refuse piles, ants of the more basal attine lineages stack their infrabuccal pellets in piles located close to their gardens, and a separate caste of workers is devoted to the construction, management, and eventual disposal of these piles.

Biol Sci Space, 2003 Oct, 17(3), 253 - 4
System of cell irradiation with a precise number of heavy ions (II); Kobayashi Y et al.; A single cell irradiation system has been developed at JAERI-Takasaki to elucidate biological effects of low dose and low dose-rate space radiation . The system has been installed at high-energy collimated heavy-ion microbeam apparatus under a vertical beam line of AVF-cyclotron accelerator . Individual mammalian cultured cells are irradiated in the atmosphere on the cell dish, the bottom of which is made of ion-track-detector TNF-1 (modified CR-39), with a single or precise numbers of 11.5 MeV/u 40Ar13+, 13.0 MeV/u 20Ne7+, and 18.3 MeV/u 12C5+ ions . Positional data of the target cells are obtained at the off-line microscope before irradiation . Targeting and irradiation of the cells are performed automatically at the on-line microscope of the microbeam apparatus according to the obtained data . About 50-100 cells are irradiated in several minutes . The number of ions traversed the cells attached on the ion track detector were counted with a plastic scintillator-photomultiplier tube assembly and a constant fraction discriminator . Immediately after the irradiation, the bottom of the cell dish was etched at 37 degrees C to check the spatial distribution of irradiated ions . The one batch of cell sample is treated in about 1 hr including the phases of cell finding, irradiation, and etching/observing of the ion tracks.

Biol Sci Space, 2003 Oct, 17(3), 251 - 2
Induction of a bystander chromosomal damage of He-ion microbeams in mammalian cells; Suzuki M et al.; We report here a bystander effect in chromosomal damage using He-ion microbeam . Human-hamster hybrid cells were irradiated with a precision He-ion microbeam generated by the Columbia microbeam system . When 20% of the cells were exposed to single He ion, the incidence of cells with chromatid-type breaks detected with the PCC technique was covered wide range from 0 to 6 breaks per cell . In contrast, the distribution showed a mixed two-peak pattern, such as non-exposed and all-cell exposed patterns, under the condition of assuming no bystander effect by treating with an effective inhibitor of cell-cell communication . These findings provide clear evidence that single He-ion irradiated cells can induce bystander chromosomal alterations in neighboring cells not directly hit by He ion.

Br J Haematol, 2004 Jan, 124(1), 72 - 9
Transplantation of a combination of CD133+ and CD34+ selected progenitor cells from alternative donors; Lang P et al.; Positive selected haematopoietic stem cells are increasingly used for allogeneic transplantation with the CD34 antigen employed in most separation techniques . However, the recently described pentaspan molecule CD133 appears to be a marker of more primitive haematopoietic progenitors . Here we report our experience with a new CD133-based selection method in 10 paediatric patients with matched unrelated (n = 2) or mismatched-related donors (n = 8) . These patients received a combination of stem cells (median = 29.3 x 10(6)/kg), selected with either anti-CD34 or anti-CD133 coated microbeads . The proportion of CD133+ selected cells was gradually increased from patient to patient from 10% to 100% . Comparison of CD133+ and CD34+ separation procedures revealed similar purity and recovery of target populations but a lower depletion of T cells by CD133+ selection (3.7 log vs . 4.1 log, P < 0.001) . Both separation procedures produced >90% CD34+/CD133+ double positive target cells . Engraftment occurred in all patients (sustained primary, n = 8; after reconditioning, n = 2) . No primary acute graft versus host disease (GvHD) >/= grade II or chronic GvHD was observed . The patients showed a rapid platelet recovery (median time to independence from substitution = 13.5 d), whereas T cell regeneration was variable . Five patients are alive with a median follow-up of 10 months . Our data demonstrates the feasibility of CD133+ selection for transplantation from alternative donors and encourages further trials with total CD133+ separated grafts.

Kidney Int, 2004 Jan, 65(1), 310 - 22
Single-chain antibody fragment-based adsorbent for the extracorporeal removal of beta2-microglobulin; Grovender EA et al.; BACKGROUND: Dialysis-related amyloidosis (DRA) is a frequent complication of end-stage renal disease (ESRD) that has been associated with the accumulation of beta2-microglobulin (beta2-m) . Removal of beta2-m results in the loss of important proteins due to the nonspecific nature of current therapies . Although whole antibodies can potentially be used to confer specificity to beta2-m removal from blood, single-chain variable region (scFv) antibody fragments could potentially offer several advantages as immunoadsorption ligands due to their size, genetic definition, ability to be expressed by microbes, and amenability for in vitro evolution . METHODS: An antihuman beta2-m scFv was constructed from the BBM.1 hybridoma and expressed by a yeast display vector . The binding affinity of the wild-type scFv fragment was quantified by flow cytometry analysis . Soluble scFv was expressed by a yeast secretion vector, purified, and immobilized onto agarose beads . The binding capacity of the immunoadsorbent was measured by equilibrating samples with saturating quantities of fluorescent beta2-m in serum . RESULTS: The displayed scFv possessed a nanomolar affinity (KD= 0.008 +/- 0.004 mg-beta2-m/L) . The immunoadsorbent exhibited an adsorption site density of 0.41 +/- 0.01 mg beta2-m/mL settled gel . Under saturating conditions, the mass ratio of adsorbed beta2-m to immobilized antibody is 70% greater than any previous literature report for whole antibodies . Preliminary specificity experiments suggest that the scFv-based immunoadsorbent is specific toward human beta2-m . CONCLUSION: Recombinant DNA technology was successfully used to engineer an scFv-based immunoadsorbent . Use of immobilized scFvs during hemodialysis may minimize loss of valuable proteins and facilitate the removal of macromolecules that are significantly larger than the molecular weight cut-off of the membrane.

Anal Chem, 2003 Sep 15, 75(18), 4732 - 9
DNA hybridization and discrimination of single-nucleotide mismatches using chip-based microbead arrays; Ali MF et al.; The development of a chip-based sensor array composed of individually addressable agarose microbeads has been demonstrated for the rapid detection of DNA oligonucleotides . Here, a "plug and play" approach allows for the simple incorporation of various biotinylated DNA capture probes into the bead-microreactors, which are derivatized in each case with avidin docking sites . The DNA capture probe containing microbeads are selectively arranged in micromachined cavities localized on silicon wafers . The microcavities possess trans-wafer openings, which allow for both fluid flow through the microreactors/analysis chambers and optical access to the chemically sensitive microbeads . Collectively, these features allow the identification and quantitation of target DNA analytes to occur in near real time using fluorescence changes that accompany binding of the target sample . The unique three-dimensional microenvironment within the agarose bead and the microfluidics capabilities of the chip structure afford a fully integrated package that fosters rapid analyses of solutions containing complex mixtures of DNA oligomers . These analyses can be completed at room temperature through the use of appropriate hybridization buffers . For applications requiring analysis of < or = 10(2) different DNA sequences, the hybridization times and point mutation selectivity factors exhibited by this bead array method exceed in many respects the operational characteristics of the commonly utilized planar DNA chip technologies . The power and utility of this microbead array DNA detection methodology is demonstrated here for the analysis of fluids containing a variety of similar 18-base oligonucleotides . Hybridization times on the order of minutes with point mutation selectivity factors greater than 10000 and limit of detection values of approximately 10(-13) M are obtained readily with this microbead array system.

J Exp Bot, 2004 Jan, 55(395), 169 - 79 Epub 2003 Dec 12.
Signals for local and systemic responses of plants to pathogen attack; Suzuki H et al.; Activation of plant defences following recognition of pathogen attack involves complex reiterative signal networks with extensive signal amplification and cross-talk . The results of two approaches that have been taken to analyse signalling in plant-microbe interactions are discussed here . Activation tagging with T-DNA harbouring multiple 35S enhancer elements was employed as a gain-of-function approach to dissect signalling related to bacterial pathogen resistance in Arabidopsis thaliana . From a screen of approximately 5000 activation tagged lines, one line was identified as harbouring a T-DNA leading to over-expression of an apoplastic aspartic protease (CDR-1), that resulted in resistance to normally virulent Pseudomonas syringae . The second approach was to screen for loss-of-function mutants in T-DNA tagged populations . From a screen of 11 000 lines, one line, defective in induced resistance-1 (dir-1) lost resistance to normally avirulent P . syringae . Models for action of the products of the CDR-1 and DIR-1 genes suggest involvement of peptide and lipid signals in systemic disease resistance responses in A . thaliana.

Plant Cell, 2004 Jan, 16(1), 229 - 40 Epub 2003 Dec 11.
SETH1 and SETH2, two components of the glycosylphosphatidylinositol anchor biosynthetic pathway, are required for pollen germination and tube growth in Arabidopsis; Lalanne E et al.; Glycosylphosphatidylinositol (GPI) anchoring provides an alternative to transmembrane domains for anchoring proteins to the cell surface in eukaryotes . GPI anchors are synthesized in the endoplasmic reticulum via the sequential addition of monosaccharides, fatty acids, and phosphoethanolamines to phosphatidylinositol . Deficiencies in GPI biosynthesis lead to embryonic lethality in animals and to conditional lethality in eukaryotic microbes by blocking cell growth, cell division, or morphogenesis . We report the genetic and phenotypic analysis of insertional mutations disrupting SETH1 and SETH2, which encode Arabidopsis homologs of two conserved proteins involved in the first step of the GPI biosynthetic pathway . seth1 and seth2 mutations specifically block male transmission and pollen function . This results from reduced pollen germination and tube growth, which are associated with abnormal callose deposition . This finding suggests an essential role for GPI anchor biosynthesis in pollen tube wall deposition or metabolism . Using transcriptomic and proteomic approaches, we identified 47 genes that encode potential GPI-anchored proteins that are expressed in pollen and demonstrated that at least 11 of these proteins are associated with pollen membranes by GPI anchoring . Many of the identified candidate proteins are homologous with proteins involved in cell wall synthesis and remodeling or intercellular signaling and adhesion, and they likely play important roles in the establishment and maintenance of polarized pollen tube growth.

Crit Rev Food Sci Nutr, 2003, 43(6), 645 - 58
Calculation of the non-isothermal inactivation patterns of microbes having sigmoidal isothermal semi-logarithmic survival curves; Peleg M; Sigmoidal isothermal semi-logarithmic survival curves are of two main types; starting with a downward and changing to upward concavity and vice versa . Both can be described by a variety of mathematical models having 3-4 adjustable parameters . The temperature dependence of these models' parameters can be described by empirical models, which account for the progressive change in the sigmoidal shape, including its disappearance at either high or low temperatures . If the temperature history of a heat-treated population of microbial cells or spores ('temperature profile') can be described algebraically, then there is a way to estimate the survival pattern under these non-isothermal conditions without invoking the traditional D and z values, which require forcing straight lines through the curved experimental data . The described method is based on the assumption that the local slope of the non-isothermal survival curve is that of the isothermal curve at the momentary temperature, at a time, which corresponds to the momentary survival ratio . It is similar to the method previously proposed for microbial populations with a 'power law' type isothermal survival curves, except that the time, which corresponds to the momentary survival ratio, is calculated either symbolically or numerically as a procedure incorporated in the governing differential equation . The method's capabilities are demonstrated with simulated survival curves under temperature histories that resemble thermal processing of foods . They include heating to different target temperatures and starting the cooling at different times.

Oecologia, 2004 Oct, 141(2), 317 - 24 Epub 2003 Dec 11.
Precipitation pulse size effects on Sonoran Desert soil microbial crusts; Cable JM et al.; Deserts are characterized by low productivity and substantial unvegetated space, which is often covered by soil microbial crust communities . Microbial crusts are important for nitrogen fixation, soil stabilization and water infiltration, but their role in ecosystem production is not well understood . This study addresses the following questions: what are the CO2 exchange responses of crusts to pulses of water, does the contribution of crusts to ecosystem flux differ from the soil respiratory flux, and is this contribution pulse size dependent? Following water application to crusts and soils, CO2 exchange was measured and respiration was partitioned through mixing model analysis of Keeling plots across treatments . Following small precipitation pulse sizes, crusts contributed 80% of soil-level CO2 fluxes to the atmosphere . However, following a large pulse event, roots and soil microbes contributed nearly 100% of the soil-level flux . Rainfall events in southern Arizona are dominated by small pulse sizes, suggesting that crusts may frequently contribute to ecosystem production . Carbon cycle studies of arid land systems should consider crusts as important contributors because of their dynamic responses to different pulse sizes as compared to the remaining ecosystem components.

Integr Cancer Ther, 2002 Sep, 1(3), 287 - 93; discussion 293
Integration of herbal medicine into modern medical practices: issues and prospects; Fong HH; The integration of herbal medicine into modern medical practices including cancer treatments must take into account the interrelated issues of quality, safety, and efficacy . Quality is the paramount issue because it can affect the efficacy and/or safety of the herbal products being used . Current product quality ranges from very high to very low due to intrinsic, extrinsic, and regulatory factors . Intrinsically, species differences, organ specificity, diurnal and seasonal variations can affect the qualitative and quantitative accumulation of active chemical constituents in the source medicinal plants . Extrinsically, environmental factors; field collection methods such as cultivation, harvest, post-harvest transport and storage; manufacturing practices; inadvertent contamination and substitution; and intentional adulteration are contributing factors to the quality of herbal medicinal products . Source plant materials that are contaminated with microbes, microbial toxins, environmental pollutants, or heavy metals; or finished products that are adulterated with foreign toxic plants or synthetic pharmaceutical agents can lead to adverse events . Substandard source materials or finished products will yield therapeutically less effective agents . Herbal medicine quality can also be attributed to regulatory practices . In a number of countries, herbal medicines are unregulated, which has led to product quality differences . Product quality improvement may be achieved by implementing control measures from the point of medicinal plant procurement under good agricultural practices (GAPs) and the manufacture of the finished botanical products under good manufacturing practices (GMPs), plus post-marketing quality assurance surveillance . The lack of pharmacological and clinical data on the majority of herbal medicinal products is a major impediment to the integration of herbal medicines into conventional medical practices . For valid integration, pharmacological and especially, clinical studies, must be conducted on those plants lacking such data . Adverse events, including drug-herb interaction must also be monitored to promote a safe integration of efficacious herbal medicine into conventional medical practices.

BMC Physiol . 2003 Dec 11;3(1):14.
Delayed leukocytosis after hard strength and endurance exercise: aspects of regulatory mechanisms; Risoy BA et al.; BACKGROUND: During infections, polymorphonuclear neutrophilic granulocytes (PMN) are mobilized from their bone marrow stores, travel with blood to the affected tissue, and kill invading microbes there . The signal(s) from the inflammatory site to the marrow are unknown, even though a number of humoral factors that can mobilize PMN, are well known . We have employed a standardized, non-infectious human model to elucidate relevant PMN mobilizers . Well-trained athletes performed a 60-min strenuous strength workout of leg muscles . Blood samples were drawn before, during and just after exercise, and then repeatedly during the following day . Cortisol, GH, ACTH, complement factors, high-sensitive CRP (muCRP), IL-6, G-CSF, IL-8 (CXCL8) and MIP-1beta (CCL4) were measured in blood samples . PMN chemotaxins in test plasma was assessed with a micropore membrane technique . RESULTS: About 5 hr after the workout, blood granulocytosis peaked to about 150% of baseline . Plasma levels of GH increased significantly 30 min into and 5 min after the exercise, but no increase was recorded for the other hormones . No significant correlation was found between concentrations of stress hormones and the subjects' later occurring PMN increases above their individual baselines . Plasma G-CSF increased significantly - but within the normal range - 65 min after the workout . IL-6 increased very slightly within the normal range, and the chemokines IL-8 and MIP-1beta did not increase consistently . However, we found a significant increase of hitherto non-identified PMN-chemotactic activity in plasma 35, 50, and 60 min after the exercise . No systemic complement activation was detected, and (mu)CRP was within the reference range at rest, 5 h and 23 h after the exercise . After endurance exercise, similar findings were made, except for a cortisol response, especially from non-elite runners . CONCLUSION: Apparently, a multitude of humoral factors can - directly or indirectly - mobilize PMN from marrow to blood; some of the factors are, others are not known to be, chemotactic . Under different conditions, different selections of these mobilizers may be used . In the late granulocytosis after heavy, long-lasting exercise a number of factors thought capable of mimicking the granulocytosis of infectious diseases were apparently irrelevant.

Dent Clin North Am, 2003 Oct, 47(4), 623 - 39
Antibiotic resistance; Pallasch TJ; Through billions of years of evolution, microbes have developed myriad defense mechanisms designed to ensure their survival . This protection is readily transferred to their fellow life forms via transposable elements . Despite very early warnings, humans have chosen to abuse the gift of antibiotics and have created a situation where all microorganisms are resistant to some antibiotics and some microorganisms are resistant to all antibiotics . When antibiotics are used, six events may occur with only one being beneficial: when the antibiotic aids the host defenses to gain control and eliminate the infection . Alternatively, the antibiotic may cause toxicity or allergy, initiate a superinfection with resistant bacteria, promote microbial chromosomal mutations to resistance, encourage resistance gene transfer to susceptible species, or promote the expression of dormant resistance genes.

J Immunol, 2003 Dec 15, 171(12), 6856 - 65
Reduced inflammation and tissue damage in transgenic rabbits overexpressing 15-lipoxygenase and endogenous anti-inflammatory lipid mediators; Serhan CN et al.; PGs and leukotrienes (LTs) mediate cardinal signs of inflammation; hence, their enzymes are targets of current anti-inflammatory therapies . Products of arachidonate 15-lipoxygenases (LO) types I and II display both beneficial roles, such as lipoxins (LXs) that stereoselectively signal counterregulation, as well as potential deleterious actions (i.e., nonspecific phospholipid degradation) . In this study, we examined transgenic (TG) rabbits overexpressing 15-LO type I and their response to inflammatory challenge . Skin challenges with either LTB(4) or IL-8 showed that 15-LO TG rabbits give markedly reduced neutrophil (PMN) recruitment and plasma leakage at dermal sites with LTB(4) . PMN from TG rabbits also exhibited a dramatic reduction in LTB(4)-stimulated granular mobilization that was not evident with peptide chemoattractants . Leukocytes from 15-LO TG rabbits gave enhanced LX production, underscoring differences in lipid mediator profiles compared with non-TG rabbits . Microbe-associated inflammation and leukocyte-mediated bone destruction were assessed by initiating acute periodontitis . 15-LO TG rabbits exhibited markedly reduced bone loss and local inflammation . Because enhanced LX production was associated with an increased anti-inflammatory status of 15-LO TG rabbits, a stable analog of 5S,6R,15S-trihydroxyeicosa-7E,9E,11Z,13E-tetraenoic acid (LXA(4)) was applied to the gingival crevice subject to periodontitis . Topical application with the 15-epi-16-phenoxy-para-fluoro-LXA(4) stable analog (ATLa) dramatically reduced leukocyte infiltration, ensuing bone loss as well as inflammation . These results indicate that overexpression of 15-LO type I and LXA(4) is associated with dampened PMN-mediated tissue degradation and bone loss, suggesting that enhanced anti-inflammation status is an active process . Moreover, they suggest that LXs can be targets for novel approaches to diseases, e.g., periodontitis and arthritis, where inflammation and bone destruction are features.

J Biol Chem, 2004 Feb 20, 279(8), 6613 - 9 Epub 2003 Dec 03.
The soluble and membrane-bound transhydrogenases UdhA and PntAB have divergent functions in NADPH metabolism of Escherichia coli; Sauer U et al.; Pentose phosphate pathway and isocitrate dehydrogenase are generally considered to be the major sources of the anabolic reductant NADPH . As one of very few microbes, Escherichia coli contains two transhydrogenase isoforms with unknown physiological function that could potentially transfer electrons directly from NADH to NADP+ and vice versa . Using defined mutants and metabolic flux analysis, we identified the proton-translocating transhydrogenase PntAB as a major source of NADPH in E . coli . During standard aerobic batch growth on glucose, 35-45% of the NADPH that is required for biosynthesis was produced via PntAB, whereas pentose phosphate pathway and isocitrate dehydrogenase contributed 35-45% and 20-25%, respectively . The energy-independent transhydrogenase UdhA, in contrast, was essential for growth under metabolic conditions with excess NADPH formation, i.e . growth on acetate or in a phosphoglucose isomerase mutant that catabolized glucose through the pentose phosphate pathway . Thus, both isoforms have divergent physiological functions: energy-dependent reduction of NADP+ with NADH by PntAB and reoxidation of NADPH by UdhA . Expression appeared to be modulated by the redox state of cellular metabolism, because genetic and environmental manipulations that increased or decreased NADPH formation down-regulated pntA or udhA transcription, respectively . The two transhydrogenase isoforms provide E . coli primary metabolism with an extraordinary flexibility to cope with varying catabolic and anabolic demands, which raises two general questions: why do only a few bacteria contain both isoforms, and how do other organisms manage NADPH metabolism?

Appl Environ Microbiol, 2003 Dec, 69(12), 7236 - 41
Targeting methanopterin biosynthesis to inhibit methanogenesis; Dumitru R et al.; This paper describes the design, synthesis, and successful employment of inhibitors of 4-(beta-D-ribofuranosyl)aminobenzene-5'-phosphate (RFA-P) synthase, which catalyzes the first committed step in the biosynthesis of methanopterin, to specifically halt the growth of methane-producing microbes . RFA-P synthase catalyzes the first step in the synthesis of tetrahydromethanopterin, a key cofactor required for methane formation and for one-carbon transformations in methanogens . A number of inhibitors, which are N-substituted derivatives of p-aminobenzoic acid (pABA), have been synthesized and their inhibition constants with RFA-P synthase have been determined . Based on comparisons of the inhibition constants among various inhibitors, we propose that the pABA binding site in RFA-P synthase has a relatively large hydrophobic pocket near the amino group . These enzyme-targeted inhibitors arrest the methanogenesis and growth of pure cultures of methanogens . Supplying pABA to the culture relieves the inhibition, indicating a competitive interaction between pABA and the inhibitor at the cellular target, which is most likely RFAP synthase . The inhibitors do not adversely affect the growth of pure cultures of the bacteria (acetogens) that play a beneficial role in the rumen . Inhibitors added to dense ruminal fluid cultures (artificial rumena) halt methanogenesis; however, they do not inhibit volatile fatty acid (VFA) production and, in some cases, VFA levels are slightly elevated in the methanogenesis-inhibited cultures . We suggest that inhibiting methanopterin biosynthesis could be considered in strategies to decrease anthropogenic methane emissions, which could have an environmental benefit since methane is a potent greenhouse gas.

Appl Environ Microbiol, 2003 Dec, 69(12), 7044 - 57
Molecular characterization of community structures and sulfur metabolism within microbial streamers in Japanese hot springs; Nakagawa T et al.; Community structures of submerged microbial slime streamers (SMSS) in sulfide-containing hot springs at 72 to 80 degrees C at Nakabusa and Yumata, Japan, were investigated by molecular analysis based on the 16S rRNA gene . The SMSS were classified into two consortia; consortium I occurred at lower levels of sulfide in the hot springs (less than 0.1 mM), and consortium II dominated when the sulfide levels were higher (more than 0.1 mM) . The dominant cell morphotypes in consortium I were filamentous and small rod-shaped cells . The filamentous cells hybridized with fluorescent oligonucleotide probes for the domain Bacteria, the domain Archaea, and the family Aquificaceae: Our analysis of the denaturing gradient gel electrophoresis (DGGE) bands by using reverse transcription (RT)-PCR amplification with two primer sets (Eub341-F with the GC clamp and Univ907R for the Bacteria and Eub341-F with the GC clamp and Arch915R) indicated that dominant bands were phylogenetically related to microbes in the genus Aquifex: On the other hand, consortium II was dominated by long, small, rod-shaped cells, which hybridized with the oligonucleotide probe S-*-Tdes-0830-a-A-20 developed in this study for the majority of as-yet-uncultivated microbes in the class Thermodesulfobacteria: The dominant DGGE band obtained by PCR and RT-PCR was affiliated with the genus Sulfurihydrogenibium: Moreover, our analysis of dissimilatory sulfite reductase (DSR) gene sequences retrieved from both consortia revealed a high frequency of DSR genes corresponding to the DSR of Thermodesulfobacteria-like microorganisms . Using both sulfide monitoring and (35)SO(4)(2-) tracer experiments, we observed microbial sulfide production and consumption by SMSS, suggesting that there is in situ sulfide production by as-yet-uncultivated Thermodesulfobacteria-like microbes and there is in situ sulfide consumption by Sulfurihydrogenibium-like microbes within the SMSS in the Nakabusa and Yumata hot springs.

Biotechnol Appl Biochem . 2003 Dec 5; {Epub ahead of print}
Human adult craniofacial muscle-derived cells - CD56/NCAM expressing cells appear to contain multipotential stem cells; Sinanan AC et al.; Skeletal muscle has been well characterized as a reservoir of myogenic precursors or satellite cells, with potential to participate in cellular repopulation therapies for muscle dysfunction . Recent evidence however suggests that the post-natal muscle compartment can be considered an alternative to bone marrow, as a source of multipotent cells or muscle-derived stem cells (MDSCs) . MDSCs, when primed with appropriate environmental cues, can differentiate into a variety of non-muscle cells . This study describes the application of a new technique for the isolation of adult human myoblasts and putative MDSCs, based on microbead-immunomagnetic selection of CD56+ cells, derived from craniofacial skeletal muscle, and details changes in morphological/molecular phenotype of the purified cells when maintained in either a myogenic or non-myogenic milieu . Multiple immunofluorescence microscopy and 2- colour flow cytometry analysis of proliferating CD56+ cultures revealed positive staining for myogenic markers (CD56, desmin and M-Cadherin) as well as putative stem cell markers (CD34, CD90, CD106 and FLK-1/VEGFR-2) . Confluent cultures subjected to cycles of adipogenic or osteogenic induction, contained either adipocytes or osteoblasts and myotubes . In conclusion, the CD56+ subpopulation within adult human skeletal muscle is heterogeneous, capable of generating lineage-committed myogenic cells, in addition to multipotent cells, the candidate MDSCs, capable of forming non-muscle tissue such as fat and bone.

J Vet Intern Med, 2003 Nov-Dec, 17(6), 902 - 7
The relationship between endogenous cortisol, blood micronutrients, and neutrophil function in postparturient Holstein cows; Cebra CK et al.; Neutrophil function, blood micronutrients, and cortisol concentrations were measured in 43 clinically healthy postparturient Holstein cows . Estimated 305-day mature equivalent milk production and neutrophil function were related to results of the blood micronutrient concentrations and neutrophil function tests . Cattle had low to normal zinc concentrations; normal to high selenium, vitamin E, and cortisol concentrations; and normal copper concentrations . Blood selenium (P = .03) and zinc (P = .027) concentrations were both significant predictors of neutrophil adhesion, and selenium (P < .001) was a significant predictor of neutrophil cytochrome C reduction (superoxide production) . Fourteen of 20 (70%) cattle with blood selenium concentrations > 300 ng/mL had neutrophil adhesion, and 15 of 20 (75%) had cytochrome C reduction above the mean value for this group . There was also a significant correlation (r = 0.331; P = .037) between cytochrome C reduction and estimated milk production . These findings suggest that neutrophils from postparturient dairy cows with higher blood concentrations of selenium have greater potential to kill microbes, and that cattle with greater superoxide production may have higher milk production.

J Neurosci, 2003 Dec 3, 23(35), 11008 - 14
Ca2+ dependency of N-cadherin function probed by laser tweezer and atomic force microscopy; Baumgartner W et al.; This study was undertaken to provide a biophysical basis for the hypothesis that activity-dependent modulation of cadherin-mediated adhesion by transient changes of extracellular calcium ({Ca2+}e) is causally involved in coordination of synaptic plasticity . Characterization of homophilic N-cadherin binding by atomic force microscopy and laser tweezer trapping of N-cadherin-coated microbeads attached to the cell surface of cultured neuronal cells showed that adhesive activity of N-cadherin is effectively regulated between 0.3 and 0.8 mm {Ca2+}e . Furthermore, we show that an increase of {Ca2+}i, which is known to be essential for induction of synaptic plasticity, causes significant reduction of cadherin-mediated bead adhesion that could be completely suppressed by inhibition of actin depolymerization . The results of this study show that N-cadherin has ideal biophysical properties to serve as a Ca2+-dependent sensor for synaptic activity and, at the same time, is strategically located to control synaptic adhesion . A drop of {Ca2+}e and a concomitant increase of {Ca2+}i may act in concert to modulate N-cadherin-based adhesive contacts at synaptic sites.

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