Metagenomics indicates abundance of biofilm related genes and horizontal transfer of multidrug resistant genes among bacterial communities in nano zinc oxide polluted soil.

The unsafe and reckless disposal of metal oxide nanoparticles like ZnO (nZnO) into the soil could seriously impact bacterial behavioural responses and functions. Under such stress, biofilm formation is considered to be a robust mechanism for bacterial survival in soil. We examined the response of bacterial metagenomes in soils exposed to varying levels of Zn (50, 200, 500 and 1000 mg kg-1) as nano Zn oxide (nZnO) in terms of biofilm genesis and regulation and their co-occurrences with multidrug resistance genes (MDRGs) and mobile genetic elements (MGEs). The size-specific effects of nZnO were verified using its bulk counterpart (bZnO). Both nZnO and bZnO facilitated profusion of biofilm related genes (BGs) especially at higher Zn levels (500 and 1000 mg kg-1 Zn), though maximum abundance was registered at a comparatively lower level under nZnO. In general, nZnO favoured an enhancement of genes involved in exopolysaccharide biosynthesis and attachment, while bZnO favoured genes related to capsule formation, chemotaxis and biofilm dispersion. Co-occurrence network analysis revealed significant positive correlations between abundances of BGs, MDRGs and MGEs, indicating an enhanced probability for horizontal gene transfer of MDRGs in nZnO polluted soils.

Soil polluted with nano ZnO reveals unstable bacterial communities and decoupling of taxonomic and functional diversities.

Due to relentless production and disposal of nano zinc oxide (nZnO), it has become critical to comprehend the serious risks large-scale accumulation of nZnO pose to bacterial communities in soil. The primary objective was to evaluate the changes in bacterial community structure and associated functional pathways through predictive metagenomic profiling and subsequent validation through Quantitative Realtime PCR in soil spiked with nZnO (0, 50, 200, 500 and 1000 mg Zn kg-1) and similar levels of bulk ZnO (bZnO). The results revealed that soil microbial biomass-C, -N, -P, soil respiration and enzyme activities decreased markedly at higher ZnO levels. The alpha diversity decreased with increasing ZnO level, with more impact under nZnO, while beta diversity analyses indicated a distinct dose- dependent separation of bacterial communities. The dominant taxa including Proteobacteria, Bacterioidetes, Acidobacteria and Planctomycetes significantly increased in abundance, while Firmicutes, Actinobacteria and Chloroflexi decreased in abundance with elevated nZnO and bZnO levels. Redundancy analysis indicated that changes in bacterial community structure instilled a greater dose- rather than size- specific response on key microbial parameters. Predicted key functions did not show a dose- specific response, and at 1000 mg Zn kg-1, methane metabolism as well as starch and sucrose metabolism were attenuated, while functions involving two component systems and bacterial secretion systems were enhanced under bZnO indicating better stress avoidance mechanism than under nZnO. Realtime PCR and microbial endpoint assays confirmed the metagenome derived taxonomic and functional data, respectively. Taxa and functions that varied substantially under stress were established as bioindicators to predict nZnO toxicity in soils. Taxon-function decoupling indicated that the soil bacterial communities deployed adaptive mechanisms under high ZnO, with lesser buffering capacity and resilience of communities under nZnO.

Augmented cellulase production by Bacillus subtilis strain MU S1 using different statistical experimental designs.

The production of cellulase by Bacillus subtilis MU S1, a strain isolated from Eravikulam National Park, was optimized using one-factor-at-a-time (OFAT) and statistical methods. Physical parameters like incubation temperature and agitation speed were optimized using OFAT and found to be 40 °C and 150 rpm, respectively, whereas, medium was optimized by statistical tools. Plackett-Burman design (PBD) was employed to screen the significant variables that highly influence cellulase production. The design showed carboxymethyl cellulose (CMC), yeast extract, NaCl, pH, MgSO4 and NaNO3 as the most significant components that affect cellulase production. Among these CMC, yeast extract, NaCl and pH showed positive effect whereas MgSO4 and NaNO3 were found to be significant at their lower levels. The optimum levels of the components that positively affect enzyme production were determined using response surface methodology (RSM) based on central composite design (CCD). Three factors namely CMC, yeast extract and NaCl were studied at five levels whilst pH of the medium was kept constant at 7. The optimal levels of the components were CMC (13.46 g/l), yeast extract (8.38 g/l) and NaCl (6.31 g/l) at pH 7. The maximum cellulase activity in optimized medium was 566.66 U/ml which was close to the predicted activity of 541.05 U/ml. Optimization of physical parameters and medium components showed an overall 3.2-fold increase in activity compared to unoptimized condition (179.06 U/ml).

Isolation and Characterization of Cellulase Producing Bacteria from the Gut of Termites (Odontotermes and Heterotermes Species).

Aim: To identify, characterize and compare the cellulolytic potentials of strains isolated from the gut of Odontotermes and Heterotermes species. Methodology: Termites were collected, identified, surface sterilized and used as source of cellulase producers. Enrichment of cellulose utilizers were done using liquid media containing carboxy methyl cellulose (CMC) as the sole source of carbon and their cellulolytic potentials were confirmed using congo red plate screening method. The isolates showing considerable zone of clearance were biochemically characterized. Three effective isolates were further identified by 16S rRNA gene sequence analysis. Growth curves of the strains were constructed under six different conditions (static and shaking at 25°C, 37°C and 45°C). Their cellulase activities- endoglucanase, FPase and β-glucosidase, assayed at 18 hours of incubation were compared under the above mentioned conditions. Results: Five isolates showing significant zone of clearance were selected, out of which three belonged to Bacillus and one each to Staphylococcus and Enterobacter sp. The three Bacillus sp. which were dominant cellulase producers were found to belong to Bacillus cereus (strain HT from Heterotermes sp. and ODO1, ODO2 from Odontotermes sp.). All the isolates showed high growth at 37°C under shaker condition. Bacillus cereus ODO2 displayed a higher cellulolytic potential compared to strain HT and ODO1. The endoglucanase, FPase and β-glucosidase activities of ODO2 were 5.06 U/mg, 2.52 U/mg and 6.01 U/mg respectively. ODO2 showed optimum specific activity at 37°C in shaker condition, whereas ODO1 and HT preferred static at same temperature. Conclusion: The strains obtained in the present study are potent cellulase producers and thus can find application in food, animal feed, textile, fuel and chemical industries. Optimization of media and genetic modification of the strains can further improve their efficiency. All the three isolates are promising in view of use in future.