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Evidence for an accessory protein function for Toll-like receptor 1 in anti-bacterial responses.

Members of the Toll-like receptor (TLR) family are components of the mammalian anti-microbial response, signaling with a domain closely related to that of IL-1 receptors. In this report the expression and function of TLR1, a TLR of unknown function, are examined. TLR1 is expressed by monocytes, as demonstrated using a novel mAb. Monocytes also express TLR2. TLR1 transfection of HeLa cells, which express neither TLR1 nor TLR2, was not sufficient to confer responsiveness to several microbial extracts. However, cotransfection of TLR1 and TLR2 resulted in enhanced signaling by HeLa cells to soluble factors released from Neisseria meningitidis relative to the response with either TLR alone. This phenomenon was also seen with high concentrations of some preparations of LPS. The N. meningitidis factors recognized by TLR1/TLR2 were not released by N. meningitidis mutant in the LpxA gene. Although LpxA is required for LPS biosynthesis, because cooperation between TLR1 and TLR2 was not seen with all LPS preparations, the microbial component(s) TLR1/2 recognizes is likely to be a complex of LPS and other molecules or a compound metabolically and chemically related to LPS. The functional IL-1R consists of a heterodimer; this report suggests a similar mechanism for TLR1 and TLR2, for certain agonists. These data further suggest that mammalian responsiveness to some bacterial products may be mediated by combinations of TLRs, suggesting a mechanism for diversifying the repertoire of Toll-mediated responses.

Acyltransferases↗

[Inhibitors of oncogene product functions].

Streptomyces and other microorganisms produce antibiotics, and enzyme inhibitors as secondary metabolites. Thus, they could be said as a treasury of organic compounds which have various structures and biological functions. Since oncogene theory has been extensively developed, we have screened oncogene function inhibitors from microorganisms as a new group of microbial secondary metabolites. Erbstatin is an inhibitor of epidermal growth factor (EGF) receptor and p60v-src-associated tyrosine kinase. Its inhibitory pattern vs. peptide is competitive. In cell culture it inhibited both EGF receptor autophosphorylation and internalization. Recently, we have isolated lavendustin A, an extremely potent inhibitor of tyrosine kinase, from Streptomyces. Lavendustin A is a novel compound and about 50 times stronger than erbstatin in inhibiting tyrosine kinase. Oxanosine is an inhibitor of ras oncogene product activity. It induces normal phenotypes in temperature-sensitive Kirsten sarcoma virus-infected rat kidney cells, lowering the intracellular levels of guanine nucleotides. Many oncogenes including src, ras, sis, fms and erbB are known to activate cellular phosphatidylinositol (PI) turnover. Therefore we have screened inhibitors of PI turnover and isolated psi-tectorigenine and pendolmycin from Nocardiopsis and inostamycin from Streptomyces. PI kinase is an enzyme involved in PI turnover pathways. We have isolated 2, 3-dihydroxybenzoic acid from Streptomyces as an inhibitor of PI kinase. These oncogene function inhibitors from microorganisms will be useful for the mechanistic study of oncogene product activities.

1-Phosphatidylinositol 4-Kinase↗

Analysis of BIOLOG GN Substrate Utilization Patterns by Microbial Communities.

BIOLOG GN plates are increasingly used to characterize microbial communities by determining the ability of the communities to oxidize various carbon sources. Studies were done to determine whether the BIOLOG GN plate assay accurately reflects the catabolic potential of the inoculum used. To gain insight into which populations of microbial communities contribute to the BIOLOG patterns, denaturing gradient gel electrophoresis and temperature gradient gel electrophoresis (TGGE) were used to assess the diversity of ribotypes in the inocula and individual wells of BIOLOG plates following incubation. These studies were done with microbial communities from the rhizosphere of potatoes and an activated sludge reactor fed with glucose and peptone. TGGE analyses of BIOLOG wells inoculated with cell suspensions from the potato rhizosphere revealed that, compared with the inoculum, there was a decrease in the number of 16S rRNA gene fragments obtained from various wells, as well as a concomitant loss of populations that had been numerically dominant in the inoculum. The dominant fragments in TGGE gels could be assigned to the gamma subclass of the class Proteobacteria, suggesting that fast-growing bacteria adapted to high substrate concentrations were numerically dominant in the wells and may have been primarily responsible for the patterns of substrate use that were observed. Similarly, the community structure changed in wells inoculated with cells from activated sludge; one or more populations were enriched, but all dominant populations of the inoculum could be detected in at least one well. This study showed that carbon source utilization profiles obtained with BIOLOG GN plates do not necessarily reflect the functional potential of the numerically dominant members of the microbial community used as the inoculum.

Journal Article↗

[Thermophilic microbial communities of deep-sea hydrothermal environments].

The latest publications on the phylogenetic and functional diversity of thermophilic prokaryotes inhabiting thermal deep-sea environments are reviewed. Along with general physicochemical characterization of the biotope studied, certain adaptation mechanisms are discussed that are peculiar to the microorganisms inhabiting it. A separate chapter addresses phylogenetic analysis of deep-sea hydrothermal microbial communities and uncultivated microorganisms recently discovered therein using molecular biological techniques. Physiological groups of thermophilic microorganisms found in deep-sea hydrotherms are considered: methanogens, sulfate-, iron-, and sulfur-reducers, aerobic hydrogen-oxidizing prokaryotes, aerobic and anaerobic organotrophs. In most cases, the isolates represent novel taxa.

Archaea↗

Oligonucleotide microarray for the study of functional gene diversity in the nitrogen cycle in the environment.

The analysis of functional diversity and its dynamics in the environment is essential for understanding the microbial ecology and biogeochemistry of aquatic systems. Here we describe the development and optimization of a DNA microarray method for the detection and quantification of functional genes in the environment and report on their preliminary application to the study of the denitrification gene nirS in the Choptank River-Chesapeake Bay system. Intergenic and intragenic resolution constraints were determined by an oligonucleotide (70-mer) microarray approach. Complete signal separation was achieved when comparing unrelated genes within the nitrogen cycle (amoA, nifH, nirK, and nirS) and detecting different variants of the same gene, nirK, corresponding to organisms with two different physiological modes, ammonia oxidizers and denitrifying halobenzoate degraders. The limits of intragenic resolution were investigated with a microarray containing 64 nirS sequences comprising 14 cultured organisms and 50 clones obtained from the Choptank River in Maryland. The nirS oligonucleotides covered a range of sequence identities from approximately 40 to 100%. The threshold values for specificity were determined to be 87% sequence identity and a target-to-probe perfect match-to-mismatch binding free-energy ratio of 0.56. The lower detection limit was 10 pg of DNA (equivalent to approximately 10(7) copies) per target per microarray. Hybridization patterns on the microarray differed between sediment samples from two stations in the Choptank River, implying important differences in the composition of the denitirifer community along an environmental gradient of salinity, inorganic nitrogen, and dissolved organic carbon. This work establishes a useful set of design constraints (independent of the target gene) for the implementation of functional gene microarrays for environmental applications.

Bacteria↗

Microbial modeling in foods.

Predictive food microbiology is a field of study that combines elements of microbiology, mathematics, and statistics to develop models that describe and predict the growth or decline of microbes under specified environmental conditions. Models can be thought of as having three levels: primary level models describe changes in microbial numbers with time, secondary level models show how the parameters of the primary model vary with environmental conditions, and the tertiary level combines the first two types of models with user-friendly application software or expert systems that calculate microbial behavior under the specified conditions. Primary models include time-to-growth, Gompertz function, exponential growth rate, and inactivation/survival models. Commonly used secondary models are response surface equations and the square root and Arrhenius relationships. Microbial models are valuable tools in planning Hazard Analysis, Critical Control Point (HACCP) programs and making decisions, as they provide the first estimates of expected changes in microbial populations when exposed to a specific set of conditions. This review describes the models currently being developed for food-borne microorganisms, particularly pathogens, and discusses their uses.

Bacteria↗

Amino acid residues affecting drug pump function in Candida albicans--C. albicans drug pump function.

Membrane-located drug transporters are important components in the multidrug resistance of microbial cells and human tissues. In fungi, clinically important resistance to antifungal drugs most often results from the over-expression of efflux pump proteins in the plasma membrane of the resistant cell. This review describes studies of the ATP binding cassette (ABC) family of membrane efflux pumps in the opportunistic human pathogen Candida albicans and, in particular, examines how changes in the polypeptide sequence can affect pump function. The identification of amino acid residues affecting pump function can provide new insights into efflux pump mechanisms and the relationship between structure and function. Such information will be important for the design of pump inhibitors which could supplement existing antifungal drugs.

ATP-Binding Cassette Transporters↗

Bacterial motility in rhizosphere colonization: mechanisms, constraints, and implications for microbial inoculants.

Although the potential of microbial inoculants for sustainable agriculture and environmental restoration has been widely recognized, their field performance remains highly variable and often unpredictable. Current research and development frameworks for microbial inoculants primarily focus on their plant growth-promoting functions and metabolic traits, often overlooking the ecological processes that determine whether introduced strains can successfully disperse, access, and establish within the rhizosphere. Increasing evidence suggests that successful dispersal and establishment cannot be assumed in the highly heterogeneous conditions of soil systems. Here, we summarize the key mechanisms underlying bacterial motility and discuss its role within the broader framework of microbial dispersal, highlighting how motility-mediated processes contribute to rhizosphere colonization. We propose that bacterial motility represents a key mechanistic determinant of biofertilizer efficacy. Its role extends beyond the ability of inoculant strains to physically reach the rhizosphere, encompassing competitive colonization on the root surface, long-term persistence, and the ability to respond to dynamic root-derived chemical gradients associated with newly developing root tissues. We argue that inoculant motility should be elevated from a passive descriptive trait to a core design parameter that can be systematically incorporated and regulated during the development and optimization of microbial inoculants. We outline a multi-tiered strategic framework for next-generation biofertilizer engineering that integrates strain selection, community design, motility regulation, and deployment strategies, thereby unlocking the full potential of synthetic microbial consortia for sustainable agriculture, ecosystem restoration, and climate change mitigation.

Biofertilizer↗

Pollution-induced community tolerance and functional redundancy in a decomposer food web in metal-stressed soil.

Pollution may lead to the development of pollution-induced community tolerance (PICT) in a stressed community. We studied the presence of PICT in soil food webs using soil microcosms. Soil microcosms containing soil invertebrates and microbes were collected from polluted and unpolluted areas and exposed to a range of soil zinc concentrations. A pine seedling was planted in each microcosm to measure the effects of the origin of the community and Zn pollution on above-ground plant production. The effects of the treatments on nutrient content in the soil were also measured. The diversity of soil microarthropods and the soil's mineral nutrient content were low at the Zn-polluted site. We did not observe an increasing Zn tolerance among the soil organisms in the polluted soil. However, low population growth rates of soil invertebrates from the polluted site may indicate the deleterious effects on fitness of long-lasting pollution. In the soil from the nonpolluted site, Zn additions caused changes in the invertebrate food web structure. These changes were explained by the good physiological condition of the animals and their insensitivity to Zn. The fact that the food web structure in soil from the polluted site did not change can be used as a rough indicator of PICT. Structural stability is presumed by the lack of Zn-sensitive species at this site and the inability of populations to acclimate by altering their growth or reproduction patterns in response to changing soil conditions. Although microbial-based soil decomposer systems may have a high functional redundancy, our results indicate that metal stress at the polluted site exceeds the tolerance limits of the system. As a consequence, ecosystem function at this site is endangered. This study also shows that the evolution of metal tolerance by soil decomposer organisms may not be a common reaction to soil pollution, although changes of population and community structure indicated severe metal stress on organisms.

Adaptation, Physiological↗

Immunopharmacological studies of PTT.119, a new synthetic bis-(2-chloroethyl)amino-L-phenylalanine derivative.

The new synthetic tripeptide p-F-Phe-m-bis-(2-chloro-ethyl)amino-Phe-Met-etoxy HCl, PTT.119, was studied for its effects on the host immune system. Doses of PTT.119 ranging from 3 to 18 mg/kg were administered i.p. to recipient mice which, at different times after drug treatment, were tested for allograft response, competence in producing lymphocytes active in lethal graft-versus-host disease, delayed-type hypersensitivity, mitogen responsiveness, humoral antibody production and natural resistance against microbial infections. At therapeutically active dosages, PTT.119 appeared to selectively inhibit functions mediated by B lymphocytes, leaving the majority of those involving T-cell subsets largely unaffected, even at the highest doses employed. Moreover, drug treatment had also a limited impact on the in vivo resistance of mice to microbial infection, which was only affected by a drug injection of 18 mg/kg, a dose well within the confidence limits of the mean lethal dose of the drug.

Animals↗

Effect of high-dose methylprednisolone therapy on phagocyte function in systemic lupus erythematosus.

Circulating phagocytes play a major role in the defence of the host against microbial infection. In an attempt to identify the reason for the unusual susceptibility to infection of patients with systemic lupus erythematosus (SLE) various parameters of phagocytic cell function were assessed kinetically in whole blood, and the accumulation of cells in areas of inflammation was studied in vivo with the skin window technique. The effect on these parameters of conventional therapy with glucocorticoids and pulse therapy with large doses of methylprednisolone were examined. Patients on conventional doses of steroids had no abnormality of phagocyte function that might have predisposed to infection, apart from a reduced accumulation of monocytes in areas of inflammation and decreased lactoferrin secretion. Pulse therapy with methylprednisolone considerably delayed the secretion of lactoferrin and the adherence of neutrophils in most of the patients, as well as impairing bacterial killing and digestion.

Adult↗

Microbial diversity in natural environments: focusing on fundamental questions.

Interactions with Gijs Kuenen and other Dutch scientists have led my lab to fundamental insights into the composition, structure and function of a hot spring cyanobacterial mat community that should influence our thinking about all microbial communities. By focusing on the distribution of molecular sequence variants of predominant mat phototrophs, we have discovered that small-scale sequence variation can be ecologically meaningful. By applying novel cultivation approaches, we have been able to obtain genetically relevant community members and thus to test the hypothesis that closely related sequence variants arose via adaptive evolutionary radiation. By applying the analytical tools of organic geochemistry we have gained insight into the metabolisms of major phototrophic members of the mat community as well as interactions between phototrophic guilds. These observations challenge traditional paradigms about prokaryotic species and cause us to consider evolutionary ecology theory as we develop genome-based methods for high-resolution analysis of the species-like fundamental units comprising microbial communities, and for investigating how such units coordinate the physiological activities within guilds of the community.

Autotrophic Processes↗

Expert systems in the control of animal cell culture processes: potentials, functions, and perspectives.

In recent years, the development of advanced systems for bioprocess monitoring and control has become an area of intensive research. Along with traditional techniques, there are several new approaches which are increasingly being applied to bioprocess operations. Among these, of special note is expert system technology, which provides possibilities for the design of efficient bioprocess control systems with new functional capabilities. This technology has been successfully applied to variety of microbial processes at laboratory and industrial scale. The present paper analyzes the possibility for application of expert systems to animal cell cultures processes whose high complexity is well suited to expert control. The discussion focuses on the organization and the functionality of the intelligent control systems, and covers some practical aspects of their design.

Animals↗

Dental caries and related factors in 88- and 92-year-olds. Cross-sectional and longitudinal comparisons.

Our aim was to compare two groups of 88- and 92-year-olds (n = 92 and n = 40), respectively, with regard to teeth, caries, and salivary and microbial conditions. Oral variables were analyzed in relation to functional capacity and use of cardiovascular agents and psychoactive drugs. Untreated root caries, plaque score, and counts of lactobacilli increased between the ages of 88 and 92 years (P < 0.01). Nine of the 24 longitudinally followed up subjects had lost 1-5 teeth over 4 years, and 17 subjects had developed new caries (DFS). The mean caries increment over 4 years was 1.3 coronal and 3.6 root surfaces, and new DFS per 100 surfaces at risk was 4.3 coronal and 17.5 root surfaces. Plaque score and final pH of buffer capacity increased (P < 0.05 and 0.01, respectively), whereas saliva flow, independent of gender, was unchanged. Use of cardiovascular agents and psychoactive drugs was associated with a deteriorated dental status.

Activities of Daily Living↗

Nadifloxacin downmodulates antigen-presenting functions of epidermal Langerhans cells and keratinocytes.

BACKGROUND: Nadifloxacin is an anti-microbial quinolone derivative widely used for the treatment of acne as a topical agent. This drug has been suggested to have not only anti-bacterial but also anti-inflammatory actions, which may have a beneficial effect on some aspects of inflammatory acne. OBJECTIVE: To further clarify its abilities to modulate skin immunity, we investigated whether nadifloxacin affects the hapten- and superantigen-presenting capacities of epidermal Langerhans cells (LC) and keratinocytes, respectively. METHODS: Immune lymph node CD4+ T cells from trinitrophenyl-sensitized BALB/c mice were cocultured with LC-enriched epidermal cells (LC-EC) that were freshly isolated from syngeneic mice and derivatized with trinitrophenyl hapten in the presence or absence of nadifloxacin. Alternatively, LC-EC were preincubated with nadifloxacin (NDFX), modified with the hapten, and cultured with immune T cells. The effects of nadifloxacin on the surface molecule expression in LC and keratinocytes were also tested by flow cytometry and cellular ELISA. RESULTS: LC-EC cultured with nadifloxacin at 10 microg/ml or more significantly suppressed the antigen-presenting function of LC for T cells. The ability of MHC class II+ keratinocytes to present a superantigen to T cells was suppressed by preincubation of keratinocytes with 30 microg/ml or more of nadifloxacin. These functional reductions in LC and keratinocytes reflected the decreased expression of MHC class II and/or costimulatory molecules. CONCLUSION: Nadifloxacin downmodulates cutaneous immunity by interfering with the antigen-presenting ability of epidermal cells.

Animals↗

Function and recruitment of mucosal regulatory T cells in human chronic Helicobacter pylori infection and gastric adenocarcinoma.

CD4(+)CD25(high) FOXP3-expressing regulatory T cells (Treg) can suppress immune responses to infections and tumors, thereby promoting microbial persistence and tumor progression. However, little is known about the phenotype and function of human mucosal Treg. Therefore, we analyzed the suppressive activity and homing phenotype of Treg in gastric mucosa of Helicobacter pylori-infected gastric adenocarcinoma patients. We found increased numbers of CD4(+)FOXP3(+) Treg in the tumor compared to tumor-free gastric mucosa. Gastric Treg cells were able to suppress H. pylori-induced T cell proliferation and IFN-gamma production. Furthermore, gastric Treg expressed increased levels of l-selectin and CCR4, compared to non-Treg cells, suggesting that these receptors contribute to Treg recruitment. The presence of functional antigen-specific Treg in H. pylori-infected gastric mucosa supports an important role for these cells in suppression of mucosal effector T cell responses, which probably contribute to bacterial persistence and possibly also to gastric tumor progression.

Adenocarcinoma↗

Prostaglandins and reproduction in female farm animals.

Prostaglandins impact on ovarian, uterine, placental, and pituitary function to regulate reproduction in female livestock. They play important roles in ovulation, luteal function, maternal recognition of pregnancy, implantation, maintenance of gestation, microbial-induced abortion, parturition, postpartum uterine and ovarian infections, and resumption of postpartum ovarian cyclicity. Prostaglandins have both positive and negative effects on reproduction; they are used to synchronize oestrus, terminate pseudopregnancy in mares, induce parturition, and treat retained placenta, luteinized cysts, pyometra, and chronic endometritis. Improved therapeutic uses for prostaglandins will be developed when we understand better their involvement in implantation, maintenance of luteal function, and establishment and maintenance of pregnancy.

Animals↗

A rare cause of anemia due to intestinal tuberculosis in a renal transplant recipient.

A renal transplant recipient with stable allograft function presented with massive hemorrhagic diarrhea and severe anemia. No microbial infection could be found in stool cultures. Early colonoscopy showed severe colitis with ulceration. Histological samples confirmed granulomatous inflammation with acid-resistant Ziehl-Neelson-positive microorganisms of mycobacterial type. Polymerase chain reaction (PCR) analysis of native mucosal biopsies specified the infectious organism as Mycobacterium tuberculosis complex. The patient responded well to antimycobacterial therapy and was still asymptomatic after 6 months with a stable graft function. Our case shows that tuberculosis can be a severe clinical problem in transplant recipients. Most of the patients with intestinal tuberculosis, reported to literature, were diagnosed post mortem or after explorative laparotomy and bowel resection. Thus, intestinal tuberculosis should be considered when a transplant recipient shows abdominal symptoms with no clear evidence of another infection. Proper diagnosis and treatment resulted in a beneficial outcome in our patient.

Anemia↗