VOLN27B: A New Head-Tailed Halovirus Isolated from an Underground Salt Crystal and Infecting Halorubrum

A novel halovirus, VOLN27B, was isolated from a drill core sample taken at a depth of approximately 430 m, from a layer formed during the Cretaceous period (Anhui, China). VOLN27B infects the halophilic archaeon Halorubrum sp. LN27 and has a head-tailed morphotype with a contractile tail, typical of myoviruses. The average head diameter is 64 ± 2.0 nm, and uncontracted tails are 15 ± 1.0 × 65 ± 2.0 nm. The latent period is about 10 h. The maturing time of VOLN27B in cells of Halorubrum sp. LN27 was nearly 8 h. The adsorption time of VOLN27B on cells of Halorubrum sp. LN27 was less than 1 min. Virus particles are unstable at pH values less than 5 or when the NaCl concentration is below 12% ( w / v ). VOLN27B and Halorubrum sp. LN27 were recovered from the same hypersaline environment and provide a new virus-host system in haloarchaea.

The Ability of Hop Extracts to Reduce the Methane Production of Methanobrevibacter ruminantium

Background. Methane emissions from agriculture are responsible for over 40% of the world’s greenhouse gas emissions. In the past, antibiotics were used to control methane production by animals, but concerns over the emergence and spread of antibiotic-resistant bacteria to humans have prompted a search for alternative approaches. Hops are the flowers of the hop plant Humulus lupulus. They have been used to feed cattle for many years and are known to contain antibacterial compounds, and their extracts have been shown to kill members of the Mycobacterium spp including Mycobacterium bovis, the causative agent of bovine tuberculosis as well as a number of human pathogens. In this study, hop extracts were studied for their ability to inhibit methane production from Methanobrevibacter ruminantium, a major methane-producing archaeon found in the rumen of cattle. Methods. Methanobrevibacter ruminantium M1T (DSM 1093) was grown at 37°C for 30 days, and the amount of methane produced at different time points during this period was measured using gas chromatography. The archaeon was exposed to commercial hop extracts (tetra-hydro-iso-alpha acid and beta acid) and to aqueous hop extracts of a range of hop variants, and their effect on methane production was determined. Results. All of the extracts reduced the level of methane production of M. ruminantium over the 30-day period compared to the negative control (sterile distilled water). The commercial hop extracts were the most effective at inhibiting methane production over the course of the experiment in contrast to the aqueous extracts, which showed a gradual reduction of inhibition with time. Conclusions. Hops contain compounds which inhibit methane production. Given that hops can be safely fed to cattle, this raises the possibility of rationally designing a feed strategy which could reduce greenhouse gas emissions and protect against bovine tuberculosis. This study recommends that further research be undertaken to further identifying bioactive components from hops and their efficacy against a range of archaea.

Structural and Kinetic Characterization of Hyperthermophilic NADH-Dependent Persulfide Reductase from Archaeoglobus fulgidus

NADH-dependent persulfide reductase (Npsr) has been proposed to facilitate dissimilatory sulfur respiration by reducing persulfide or sulfane sulfur-containing substrates to H2S. The presence of this gene in the sulfate and thiosulfate-reducing Archaeoglobus fulgidus DSM 4304 and other hyperthermophilic Archaeoglobales appears anomalous, as A. fulgidus is unable to respire S0 and grow in the presence of elemental sulfur. To assess the role of Npsr in the sulfur metabolism of A. fulgidus DSM 4304, the Npsr from A. fulgidus was characterized. AfNpsr is specific for persulfide and polysulfide as substrates in the oxidative half-reaction, exhibiting k cat / K m on the order of 104 M-1 s-1, which is similar to the kinetic parameters observed for hyperthermophilic CoA persulfide reductases. In contrast to the bacterial Npsr, AfNpsr exhibits low disulfide reductase activity with DTNB; however, similar to the bacterial enzymes, it does not show detectable activity with CoA-disulfide, oxidized glutathione, or cystine. The 3.1 Å X-ray structure of AfNpsr reveals access to the tightly bound catalytic CoA, and the active site Cys 42 is restricted by a flexible loop (residues 60-66) that is not seen in the bacterial homologs from Shewanella loihica PV-4 and Bacillus anthracis. Unlike the bacterial enzymes, AfNpsr exhibits NADH oxidase activity and also shows no detectable activity with NADPH. Models suggest steric and electrostatic repulsions of the NADPH 2 ′ -phosphate account for the strong preference for NADH. The presence of Npsr in the nonsulfur-reducing A. fulgidus suggests that the enzyme may offer some protection against S0 or serve in another metabolic role that has yet to be identified.

Analysis of a Methanogen and an Actinobacterium Dominating the Thermophilic Microbial Community of an Electromethanogenic Biocathode

Electromethanogenesis refers to the bioelectrochemical synthesis of methane from CO2 by biocathodes. In an electromethanogenic system using thermophilic microorganisms, metagenomic analysis along with quantitative real-time polymerase chain reaction and fluorescence in situ hybridization revealed that the biocathode microbiota was dominated by the methanogen Methanothermobacter sp. strain EMTCatA1 and the actinobacterium Coriobacteriaceae sp. strain EMTCatB1. RNA sequencing was used to compare the transcriptome profiles of each strain at the methane-producing biocathodes with those in an open circuit and with the methanogenesis inhibitor 2-bromoethanesulfonate (BrES). For the methanogen, genes related to hydrogenotrophic methanogenesis were highly expressed in a manner similar to those observed under H2-limited conditions. For the actinobacterium, the expression profiles of genes encoding multiheme c-type cytochromes and membrane-bound oxidoreductases suggested that the actinobacterium directly takes up electrons from the electrode. In both strains, various stress-related genes were commonly induced in the open-circuit biocathodes and biocathodes with BrES. This study provides a molecular inventory of the dominant species of an electromethanogenic biocathode with functional insights and therefore represents the first multiomics characterization of an electromethanogenic biocathode.

Performance Analysis and Microbial Community Evolution of In Situ Biological Biogas Upgrading with Increasing H2/CO2 Ratio

The effect of the amount of hydrogen supplied for the in situ biological biogas upgrading was investigated by monitoring the process and evolution of the microbial community. Two parallel reactors, operated at 37°C for 211 days, were continuously fed with sewage sludge at a constant organic loading rate of 1.5 gCOD∙(L∙d)-1 and hydrogen (H2). The molar ratio of H2/CO2 was progressively increased from 0.5 : 1 to 7 : 1 to convert carbon dioxide (CO2) into biomethane via hydrogenotrophic methanogenesis. Changes in the biogas composition become statistically different above the stoichiometric H2/CO2 ratio (4 : 1). At a H2/CO2 ratio of 7 : 1, the methane content in the biogas reached 90%, without adversely affecting degradation of the organic matter. The possibility of selecting, adapting, and enriching the original biomass with target-oriented microorganisms able to biologically convert CO2 into methane was verified: high throughput sequencing of 16S rRNA gene revealed that hydrogenotrophic methanogens, belonging to Methanolinea and Methanobacterium genera, were dominant. Based on the outcomes of this study, further optimization and engineering of this process is feasible and needed as a means to boost energy recovery from sludge treatment.

Production of Poly(3-Hydroxybutyrate) by Haloarcula, Halorubrum, and Natrinema Haloarchaeal Genera Using Starch as a Carbon Source

Microbial production of bioplastics, derived from poly(3-hydroxybutyrate) (PHB), have provided a promising alternative towards plastic pollution. Compared to other extremophiles, halophilic archaea are considered as cell factories for PHB production by using renewable, inexpensive carbon sources, thus decreasing the fermentation cost. This study is aimed at screening 33 halophilic archaea isolated from three enrichment cultures from Tunisian hypersaline lake, Chott El Jerid, using starch as the sole carbon source by Nile Red/Sudan Black staining and further confirmed by PCR amplification of phaC and phaE polymerase genes. 14 isolates have been recognized as positive candidates for PHA production and detected during both seasons. The identification of these strains through 16S rRNA gene analyses showed their affiliation to Halorubrum, Natrinema, and Haloarcula genera. Among them, three PHB-producing strains, CEJ34-14, CEJ5-14, and CEJ48-10, related to Halorubrum chaoviator, Natrinema pallidum, and Haloarcula tradensis were found to be the best ones reaching values of 9.25, 7.11, and 1.42% of cell dry weight (CDW), respectively. Our findings highlighted that Halorubrum, Natrinema, and Haloarcula genera were promising candidates for PHB production using soluble starch as a carbon source under high salinity (250 g L-1 NaCl).

Flocculation Efficiency and Mechanism of Carbamazepine by Microbial Flocculant Extracted from Klebsiella pneumoniae J1

The microbial flocculant (MFX) extracted from Klebsiella pneumoniae J1 was used to remove carbamazepine in prepared wastewater and domestic sewage. The influence factors and flocculation mechanism were studied. The optimal carbamazepine removal conditions for MFX were pH of 7-8, 7 mL of flocculant, 0.1 mL of coagulant, and 35°C, and the removal rate reached 81.75%. MFX was efficient in the removal of carbamazepine in both domestic sewage (75.03%) and secondary sedimentation tank effluent (69.76%). The pseudo-first-order kinetic equation fitted the adsorption process better than the pseudo-second-order kinetic equation, which suggested that the adsorption was not pure chemical adsorption. The analysis of floc size suggested that the repulsive force between carbamazepine and MFX was weakened under alkalescent conditions, which can help the growth and coherence of flocs and increase the carbamazepine removal efficiency. Enough dosage of MFX can generate larger flocs, but excessive dosage of MFX will decrease the carbamazepine removal rate because of increase in electrostatic repulsion. The analysis of 3D-EEM and FTIR suggested that hydroxyl, amino, and carboxyl in MFX played an important role in the removal of carbamazepine. As an eco-friendly and highly efficient microbial flocculant, MFX has potential for practical applications in carbamazepine removal.

Thermophilic Solid-State Anaerobic Digestion of Corn Straw, Cattle Manure, and Vegetable Waste: Effect of Temperature, Total Solid Content, and C/N Ratio

Thermophilic solid-state anaerobic digestion (SS-AD) of agricultural wastes, i.e., corn straw, cattle manure, and vegetable waste, was carried out in this study. The effects of temperature (40-60°C), initial solid content (ISC, 17.5-32.5%), and C/N ratio (15-32 : 1) on biogas production were evaluated using a Box-Behnken experimental design (BBD) combined with response surface methodology (RSM). The results showed that optimization of process parameters is important to promote the SS-AD performance. All the factors, including interactive terms (except the ISC), were significant in the quadratic model for biogas production with SS-AD. Among the three operation parameters, the C/N ratio had the largest effect on biogas production, followed by temperature, and a maximum biogas yield of 241.4 mL gVS-1 could be achieved at 47.3°C, ISC = 24.81 % , and C / N = 22.35 . After 20 d of SS-AD, the microbial community structure under different conditions was characterized by high-throughput sequencing, showing that Firmicutes, Bacteroidetes, Chloroflexi, Synergistetes, and Proteobacteria dominated the bacterial community, and that Firmicutes had a competitive advantage over Bacteroidetes at elevated temperatures. The biogas production values and relative abundance of OPB54 and Bacteroidia after 20 d of SS-AD can be fitted well using a quadratic model, implying that OPB54 and Bacteroidia play important roles in the methanogenic metabolism for agricultural waste thermophilic SS-AD.

Identification and Characterization of a Newly Isolated Chitinase-Producing Strain Bacillus licheniformis SSCL-10 for Chitin Degradation

Chitinases or chitinolytic enzymes have different applications in the field of medicine, agriculture, and industry. The present study is aimed at developing an effective hyperchitinase-producing mutant strain of novel Bacillus licheniformis. A simple and rapid methodology was used for screening potential chitinolytic microbiota by chemical mutagenesis with ethylmethane sulfonate and irradiation with UV. There were 16 mutant strains exhibiting chitinase activity. Out of the chitinase-producing strains, the strain with maximum chitinase activity was selected, the protein was partially purified by SDS-PAGE, and the strain was identified as Bacillus licheniformis (SSCL-10) with the highest specific activity of 3.4 U/mL. The induced mutation model has been successfully implemented in the mutant EMS-13 (20.2 U/mL) that produces 5-6-fold higher yield of chitinase, whereas the mutant UV-11 (13.3 U/mL) has 3-4-fold greater chitinase activity compared to the wild strain. The partially purified chitinase has a molecular weight of 66 kDa. The wild strain (SSCL-10) was identified as Bacillus licheniformis using 16S rRNA sequence analysis. This study explores the potential applications of hyperchitinase-producing bacteria in recycling and processing chitin wastes from crustaceans and shrimp, thereby adding value to the crustacean industry.