An updated genome-scale metabolic network reconstruction of Pseudomonas aeruginosa PA14 to characterize mucin-driven shifts in bacterial metabolism

Mucins are present in mucosal membranes throughout the body and play a key role in the microbe clearance and infection prevention. Understanding the metabolic responses of pathogens to mucins will further enable the development of protective approaches against infections. We update the genome-scale metabolic network reconstruction (GENRE) of one such pathogen, Pseudomonas aeruginosa PA14, through metabolic coverage expansion, format update, extensive annotation addition, and literature-based curation to produce iPau21. We then validate iPau21 through MEMOTE, growth rate, carbon source utilization, and gene essentiality testing to demonstrate its improved quality and predictive capabilities. We then integrate the GENRE with transcriptomic data in order to generate context-specific models of P. aeruginosa metabolism. The contextualized models recapitulated known phenotypes of unaltered growth and a differential utilization of fumarate metabolism, while also revealing an increased utilization of propionate metabolism upon MUC5B exposure. This work serves to validate iPau21 and demonstrate its utility for providing biological insights.

Effects of Atrazine on Chernozem Microbial Communities Evaluated by Traditional Detection and Modern Sequencing Technology

Atrazine is a long residual herbicide commonly used in maize fields. Although atrazine can effectively control weeds and improve crop yield, long-term application leads to continuous pollution in the agricultural ecological environment, especially in the soil ecosystem, and its impact on soil microorganisms is still not clear. Four methods were used in the experiment to clarify the effect of atrazine on the bacterial populations of cultivated soil layers of chernozem in a cold region in different periods: high-performance liquid chromatography (HPLC), colorimetry, microplate, and high-throughput sequencing. The level of residual atrazine in cold chernozem decreased from 4.645 to 0.077 mg/kg soil over time, and the residue gradually leached into deep soil and then decreased after accumulating to a maximum value. Atrazine significantly affected the activities of urease and polyphenol oxidase activity in the soil layers at different periods but had no significant effect on sucrase and phosphatase activity. Atrazine significantly reduced the diversity of microbial carbon source utilization and total activity in soil layers of 0–10 and 20–30 cm but only reduced the diversity of microbial carbon source utilization in the 10–20 cm layer. Atrazine had no significant effect on bacterial populations (10–12 phyla, 29–34 genera), but had a slight effect on the relative abundance of various groups. Atrazine significantly reduced the diversity of bacterial populations in cultivated soil layers of chernozem in a cold region, and the diversity of bacterial populations decreased with decreased residue. This lays a foundation for guiding the safe use of herbicides on farmland in Northeast China.

Positioning the Model Bacterial Organelle, the Carboxysome

ABSTRACT Bacterial microcompartments (BMCs) confine a diverse array of metabolic reactions within a selectively permeable protein shell, allowing for specialized biochemistry that would be less efficient or altogether impossible without compartmentalization. BMCs play critical roles in carbon fixation, carbon source utilization, and pathogenesis. Despite their prevalence and importance in bacterial metabolism, little is known about BMC “homeostasis,” a term we use here to encompass BMC assembly, composition, size, copy-number, maintenance, turnover, positioning, and ultimately, function in the cell. The carbon-fixing carboxysome is one of the most well-studied BMCs with regard to mechanisms of self-assembly and subcellular organization. In this minireview, we focus on the only known BMC positioning system to date—the maintenance of carboxysome distribution (Mcd) system, which spatially organizes carboxysomes. We describe the two-component McdAB system and its proposed diffusion-ratchet mechanism for carboxysome positioning. We then discuss the prevalence of McdAB systems among carboxysome-containing bacteria and highlight recent evidence suggesting how liquid-liquid phase separation (LLPS) may play critical roles in carboxysome homeostasis. We end with an outline of future work on the carboxysome distribution system and a perspective on how other BMCs may be spatially regulated. We anticipate that a deeper understanding of BMC organization, including nontraditional homeostasis mechanisms involving LLPS and ATP-driven organization, is on the horizon.

An updated genome-scale metabolic network reconstruction of Pseudomonas aeruginosa PA14 to characterize mucin-driven shifts in bacterial metabolism

Mucins are present in mucosal membranes throughout the body and play a key role in the microbe clearance and infection prevention. Understanding the metabolic responses of pathogens to mucins will further enable the development of protective approaches against infections. We update the genome-scale metabolic network reconstruction (GENRE) of one such pathogen, Pseudomonas aeruginosa PA14, through metabolic coverage expansion, format update, extensive annotation addition, and literature-based curation to produce iPau21. We then validate iPau21 through MEMOTE, growth rate, carbon source utilization, and gene essentiality testing to demonstrate its improved quality and predictive capabilities. We then integrate the GENRE with transcriptomic data in order to generate context-specific models of P. aeruginosa metabolism. The contextualized models recapitulated known phenotypes of unaltered growth and a differential utilization of fumarate metabolism, while also providing a novel insight about an increased utilization of propionate metabolism upon MUC5B exposure. This work serves to validate iPau21 and demonstrate its utility for providing novel biological insights.

Glucose-6-Phosphate Isomerase FgGPI, a β2 Tubulin-Interacting Protein, Is Indispensable for Fungal Development and Deoxynivalenol Biosynthesis in Fusarium graminearum

Glucose-6-phosphate isomerase (GPI) is ubiquitous in most organisms, catalyzing the reversible isomerization of glucose-6-phosphate and fructose-6-phosphate. In this study, we investigated biological and genetic functions of FgGPI in the phytopathogen Fusarium graminearum. We found that hyphal growth, conidial germination, and septa formation were significantly inhibited in FgGPI deletion mutant ∆FgGPI. FgGPI was also positively associated with glucose metabolism, ATP biosynthesis, and carbon source utilization. In addition, pyruvate production, deoxynivalenol (DON) biosynthesis, and virulence were reduced in ∆FgGPI. A coimmunoprecipitation assay demonstrated that FgGPI interacts with Fgβ2. More importantly, the coimmunoprecipitation assay showed that carbendazim-resistant substitutions in β2 tubulin could reduce the interaction intensity between FgGPI and Fgβ2, thereby increasing FgGPI expression and accelerating DON biosynthesis in carbendazim-resistant strains. Taken together, our work revealed the indispensable role of FgGPI in fungal developmental processes, DON biosynthesis, and pathogenicity in F. graminearum.

Studies on Optimization of Growth Parameters for Mass Multiplication of Actinobacteria

Aim: To isolate, characterize and optimize the growth parameters for mass multiplication of Actinobacteria. Place and Duration of Work: The study was carried out in Department of Agricultural Microbiology, GKVK, University of Agricultural Sciences, Bangalore during 2019-20. Methodology: Actinobacterial isolates were characterized morphologically and screened for optimization of growth parameters viz., pH, temperature, salt concentration and utilization of carbon source for their mass multiplication. Results: Forty actinobacterial isolates were enumerated from rhizosphere soil of finger millet, cowpea and also from different organic manures. Color of aerial mycelium in most of the actinobacterial isolates were white, grey or cream with dry, cottony or powdery appearance. All forty isolates were Gram positive, non-acid forming and motile. During optimization of growth parameters, results showed that all the actinobacterial isolates growth was observed good at 30℃, pH 7 and 2 per cent NaCl concentration. Starch was confirmed as the best carbon source for all the actinobacterial isolates during carbon source utilization ability. Conclusion: Based on the results, it is showed that all the actinobacterial isolates enumerated were aerobic, spore-forming, Gram positive bacteria, non-acid forming and motile. Maximum growth of Actinobacterial isolates was obtained at temperature of 30℃, pH 7 and 2 per cent NaCl concentration with the ability of growing on ten different carbon sources during the optimization of nutritional and cultural characterization studies. Among the different carbon sources, starch was confirmed as the best carbon source for all the isolates during the study of carbon source utilization ability.

Soil Enzyme Activity and Microbial Metabolic Function Diversity in Soda Saline–Alkali Rice Paddy Fields of Northeast China

Western Jilin province has the most serious area of soda salinization in Northeast China, which affects and restricts the sustainable development of agriculture. The effects of physico-chemical properties of rhizosphere and non-rhizosphere soil on soil microbial diversity and enzyme activities (polyphenol oxidase, catalase, invertase, amylase) were evaluated in typical soda saline-alkali paddy field. Community-level physiological profile (CLPP) based on Biolog-ECO plates was used to assess the functional diversity of soil microorganisms. Exchangeable sodium percentage (ESP) and pH were negative correlated with the microbial activity (AWCD), soil enzyme activities (amylase, sucrose, and catalase, except for polyphenol oxidase) in rice rhizosphere and non-rhizosphere soil (P < 0.05). The indexes of microbial diversity in rice rhizosphere soil were significantly higher than that of non-rhizosphere soil. The utilization of amino acids by rice rhizosphere microorganisms was relatively high, while non-rhizosphere soil had relatively high utilization of carboxylic acid, phenolic acid, and amine. Among the selected physico-chemical properties, soil organic carbon (SOC) and soil water content (SWC) had the greatest influence on the variation of microbial diversity indexes and enzyme activities in rhizosphere soil. ESP and pH showed a significant positive correlation with carbon source utilization, especially for amine (AM) and phenolic acid (PA) carbon source utilization (P < 0.05) by means of RDA, and the utilization rate of AM and PA carbon sources by rice rhizosphere and non-root soil microorganisms was P1 < P2 < P3.