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Substrate-inhibitor cooperative interactions with microbial dihydrofolate reductases.

Cooperativity in the binding of two substrates to an enzyme is a now well-established phenomenon. The x-ray crystallographic structure of the E. coli DHFR binary TMP complex compared with the ternary enzyme-NADPH-TMP complex suggests without too imaginative extrapolation, that the conformational changes resulting from the binding of one ligand aid in favorably positioning potential binding sites for the second ligand. Of greater importance is the fact that the extent to which inhibitor binding is enhanced by the binding of NADPH varies from species to species. To a significant extent, for example, the selectivity of TMP is enhanced by the increase in its binding to the E. coli enzyme when NADPH is present as compared with several mammalian enzymes. The reverse, negative cooperativity (a decrease in binding of a substance when moving from the binary to a ternary complex), is perhaps less common and certainly less well studied. The present paper deals with one such enzyme, the DHFR from C. albicans, and by reference to another, that from S. cerevisiae, where it is shown that the binding of substrates exhibit strong negative cooperativity. It was of interest also to determine the relationship between inhibitor/NADPH cooperativity and the relative insensitivity of N. gonorrhoeae to TMP. Equilibrium studies show that the binding of TMP in binary complex with this enzyme is exceedingly poor and that a 2,200-fold cooperative effect brings the gonococcal enzyme Ki within one order of magnitude of the E. coli enzyme Ki. Even so, it takes synergism of another sort (with sulfamethoxazole) and high doses to make co-trimoxazole therapy feasible for treating gonorrhoeae. The comparative results on the gonococcal enzyme for a family of near relatives of TMP are of interest also for the reason that the structure-activity relationships with this enzyme are quite different from those of the E. coli and other microbial enzymes. Finally, it should be pointed out that although the negative cooperativity found for the candida and saccharomyces enzymes is relatively large, it is the values of the substrate Michaelis constants that are physiologically relevant. The Km values of the yeast enzymes are within the range for other DHFR and therefore the intracellular activity of the enzymes should not be compromised.

Candida albicans

Cross-domain cooperation drives nutrient acquisition and metabolism in the bark beetle holobiont.

Microbial symbiosis underpins host adaptation, yet mechanisms of metabolic integration in holobionts remain unclear. Using metatranscriptomics, genomics, and metabolic assays, we investigated gut microbiome interactions in the European spruce bark beetle (Ips typographus). We observed metabolic complementarity among symbionts and host, forming cross-domain networks that support nutrient acquisition. Nitrogen recycling revealed strong interdependence: no single partner possessed a complete uric acid degradation pathway, but combined evidence supports a distributed pathway spanning beetle, Bacteria, and fungi. Additionally, bacterial nitrate reduction to ammonia indicates a potential nitrogen influx, making otherwise inaccessible inorganic nitrogen available to the host. Shaped by microbial interactions, symbionts also likely supply specific amino acids, while vitamin metabolism showed cross-domain co-metabolism, with Bacteria as main producers of B vitamins, while host and fungi modulated interconversion. Carbohydrate degradation was highly partitioned; bacteria target xylan and pectin, while fungi contribute to glucan breakdown. Crucially, our data provide indirect evidence that the beetle may contribute to complete cellulose degradation, highlighting an underappreciated host role in lignocellulose processing. In terms of enzymatic functional diversity, the bacteriome emerged as the most important microbiome component-an observation that contrasts with the traditional focus on fungi and underscores the need to consider bacterial contributions in insect symbioses. Despite life-stage variation, core metabolic functions remained stable. Overall, metabolic interdependence, rather than microbial composition alone, structures holobiont function. These results highlight functional redundancy and ecological resilience, emphasizing the importance of microbial cooperation and host-microbe metabolic evolution.

Bark beetle

Cross-Kingdom Siderophores: Biosynthesis, Ecology, and Biotechnological Applications.

Microbial siderophores are high-affinity iron-binding compounds which are produced by bacteria, fungi, and actinomycetes to obtain iron and survive and interact with different species in an iron-deficient environment. While the conventional research on siderophore systems deals mainly with the study within the same taxa, modern researchers have increased their inclination toward cross-kingdom integration of siderophore behavior and their impact on host-associated environments. This can be largely attributed to differences in biosynthetic gene clusters, receptor systems, and regulatory networks, which produce distinct genotype-to-phenotype results determining microbial cooperation and competition. Current advancements in genomic research, together with omics studies like transcriptomics, proteomics, and metabolomics, have created newer insights into how siderophores function. However, the present literature evidences multiple major gaps in multi-omics data because the link between genomes and metabolomes remains weak due to inconsistent regulatory data sets and failure in identifying producer-consumer relationships in polymicrobial systems. Additionally, major constraints like molecular instability, delivery system limitations, host toxicity, limitations in upscaling, and regulatory issues delimit the use of siderophores in medical treatment, agricultural practices, and environmental biotechnology. This review aims to bridge the existing knowledge about siderophore biochemistry, biosynthesis, ecological functions, and genetic regulation across kingdoms while integrating multi-omics outlook with translational considerations. Thus, by connecting molecular mechanisms with evolutionary cross-talk, this study aims to provide a system-level framework in the world of siderophore-mediated iron uptake and therefore shapes future directions in emerging fields of microbial engineering, precision therapies, and sustainable biotechnology.

Fur regulation

Standardized susceptibility testing of fluconazole: an international collaborative study.

An international collaborative study of broth dilution (MIC) and disk diffusion susceptibility testing of fluconazole was conducted by using a chemically defined medium (High-Resolution Antifungal Assay Medium; Oxoid Ltd., Basingstoke, United Kingdom) and standard test methods performed in eight reference laboratories. Ten yeast isolates were tested by each test method in duplicate on each of 3 separate days. The intralaboratory reproducibility of the MIC test was excellent; 95.7% of the replicate tests (n = 220) were within 2 doubling dilutions of the other values in the set for the eight laboratories. The intralaboratory reproducibility of the disk test was also good, with 91% of the replicate tests (n = 234) agreeing with each other within an arbitrarily chosen value of 4 mm. Interlaboratory agreement of MIC test results was acceptable, with 84% of the MICs agreeing within 2 doubling dilutions. In contrast, the interlaboratory agreement of the disk test was not good, with only 59% of test results agreeing within 4 mm. Comparison of the rank order of MICs obtained in each laboratory with the reference rank order gave an agreement of 70 to 80% (median, 80%) with the MIC test and 70 to 90% (median, 80%) with the disk test. These preliminary results are encouraging for the development of standardized testing methods for testing fluconazole.

Culture Media

Worldwide study of cefoperazone susceptibility.

In vitro susceptibility to cefoperazone of more than one million clinical aerobic bacterial isolates was evaluated in 369 hospitals in Japan, the United States, Canada, France, Austria, and Hungary. Standard versions of the disk diffusion method were used according to interpretive criteria approved in the respective countries, and comparisons were made among the countries and among various body sites. Susceptibility of organisms varied little within most species among the countries. Minor differences in methodology and in interpretive criteria may explain some of the variations observed among countries and hospitals. The susceptibility of key pathogens to cefoperazone among countries ranged from 91% to 98% for Escherichia coli, 73% to 91% for Pseudomonas aeruginosa, 90% to 95% for Klebsiella pneumoniae, 73% to 92% for Enterobacter cloacae, 74% to 92% for Staphylococcus aureus, and 91% to 97% for Proteus mirabilis. Susceptibility of isolates from the urinary tract did not vary markedly from that in other body sites.

Cefoperazone

Carbon metabolic homogenization is linked to microbial competition and antimicrobial resistance in soils under forest-to-cropland conversion.

Global agricultural expansion by converting natural forests into croplands often leads to soil functional homogenization and antimicrobial resistance enhancement, threatening ecosystem services. However, the associations between microbial carbon metabolic homogenization and antimicrobial resistance remain largely unknown. Here, we collected 240 paired forest and cropland soil samples from the most intensively farmed Yangtze River Basin in China, and constructed a novel framework based on microbial functional traits to decipher the role of carbon metabolic homogenization on antimicrobial resistance via microbial competition for metabolites. Using genome-scale metabolic models, we found that carbon metabolic homogenization was associated with a shift in microbial interactions from cooperation toward competition, with a 45.6% increase in competitive interactions that coincided with a 35.6% higher antimicrobial resistance gene (ARG) diversity. This shift was accompanied by smaller genome sizes and higher 16S rRNA copy numbers, indicating fast-growing, resource-acquisitive microbial strategies. Metabolic transfer analyses further revealed less cooperation relationships among microbial communities in cropland soils than in forest soils, indicating an intensified battle for communal metabolites and an attenuated exchange for complementary metabolites. Together, these findings provide a new framework to understand the association between carbon metabolic homogenization and soil antimicrobial resistance risks from the perspective of microbial traits and interactions under land use change.

Soil Microbiology

Ultrastructural features of microbial colony organization.

The ultrastructure of microbial colonies was studied. Inside the colonies three types of intercellular contacts were demonstrated. In the colonies of Gram-negative bacteria, the cells were found to be connected by tight adhesions of outer membranes of the cell walls and membrane bridges. In the colonies of Gram-positive bacteria, the intercellular contacts were formed by fusion of peptidoglucan layers of the cell walls. Bacterial cells were differentiated by the presence of a capsule-like envelope. The obtained data indicate the existence of elements of cellular cooperation and specialization in microbial colonies.

Bacterial Adhesion

Enterotoxigenic Escherichia coli diarrhea of travelers: a prospective study of American Peace Corps volunteers.

Travelers' diarrhea was studied prospectively in a group of 39 American Peace Corps Volunteers (PCVs) during their first five weeks in Kenya. Twenty-seven developed diarrheal disease and 12 remained well. Multiple episodes were documented in 11 of the symptomatic volunteers. Enterotoxigenic Escherichia coli of many serotypes producing heat-labile and/or heat-stable enterotoxin were isolated from 17 of the 27 volunteers with diarrhea and from 1 of the 12 well volunteers. The enterotoxigenic E. coli were more likely to be antibiotic sensitive than the non-enterotoxigenic E. coli. A serum antibody rise to the heat-labile toxin (LT) was detected in six symptomatic volunteers, five of whom had a positive culture for LT-producing E. coli, and from one asymptomatic, culture negative volunteer. Salmonella cubana was isolated from two volunteers, and three volunteers had serologic evidence of infection with human reovirus-like (rotavirus) agent. This study confirms the role of enterotoxigenic E. coli as a major cause of travelers' diarrhea and suggests that the disease is similar in widely separated geographic areas.

Adult

The beneficial effects of localized tumor necrosis factor production in BCG infection.

Inflammatory responses to infectious agents involve different cell populations, including monocytes/macrophages, granulocytes, eosinophils, mastocytes and lymphocytes, which realize a cooperative network aimed at microbial clearance. The recruitment and activation of these inflammatory cells is mediated by a series of cytokines synthesized by distinct elements of the immune system. In particular, the production of tumor necrosis factor (TNF) is known to be induced during the course of various infectious processes. TNF is now recognized to be the major effector of gram-negative endotoxic shock. Experimental and clinical studies have documented the systemic release of TNF during bacterial and parasitic infections, and TNF is considered to account for some of the severe metabolic and tissular damages associated with such disease states. In contrast, we have shown that during BCG infections TNF is produced transiently and focally by granulomas in response to mycobacterial challenge and that it contributes to their containment and elimination. These observations suggest that the localized release of TNF may play a pivotal role in defense mechanisms against microorganisms, while its overproduction, leading to systemic release during severe infections, might be held responsible for a wide range of tissular injuries.

Animals

[Choleragen and neuraminidase production by Vibrio in vitro].

In vitro studies demonstrated a high degree of the rank correlation between the synthesis of cholera exotoxin and neuraminidase by cholera vibrios (V. cholerae 569B). The appearance of these biochemically-active materials proved to depend on the growth phase of the microbial population. A possibility of cooperation between them in the pathogenesis of cholera is suggested.

Enterotoxins