The novel regulator HdrR controls the transcription of the heterodisulfide reductase operon hdrBCA in Methanosarcina barkeri.

Methanogenic archaea play a key role in the global carbon cycle because these microorganisms remineralize organic compounds in various anaerobic environments. The microorganism Methanosarcina barkeri is a metabolically versatile methanogen, which can utilize acetate, methanol, and H2/CO2 to synthesize methane. However, the regulatory mechanisms underlying methanogenesis for different substrates remain unknown. In this study, RNA-seq analysis was used to investigate M. barkeri growth and gene transcription under different substrate regimes. According to the results, M. barkeri showed the best growth under methanol, followed by H2/CO2 and acetate, and these findings corresponded well with the observed variations in genes transcription abundance for different substrates. In addition, we identified a novel regulator, MSBRM_RS03855 (designated as HdrR), which specifically activates the transcription of the heterodisulfide reductase hdrBCA operon in M. barkeri. HdrR was able to bind to the hdrBCA operon promoter to regulate transcription. Furthermore, the structural model analyses revealed a helix-turn-helix domain, which is likely involved in DNA binding. Taken together, HdrR serves as a model to reveal how certain regulatory factors control the expression of key enzymes in the methanogenic pathway.IMPORTANCEThe microorganism Methanosarcina barkeri has a pivotal role in the global carbon cycle and contributes to global temperature homeostasis. The consequences of biological methanogenesis are far-reaching, including impacts on atmospheric methane and CO2 concentrations, agriculture, energy production, waste treatment, and human health. As such, reducing methane emissions is crucial to meeting set climate goals. The methanogenic activity of certain microorganisms can be drastically reduced by inhibiting the transcription of the hdrBCA operon, which encodes heterodisulfide reductases. Here, we provide novel insight into the mechanisms regulating hdrBCA operon transcription in the model methanogen M. barkeri. The results clarified that HdrR serves as a regulator of heterodisulfide reductase hdrBCA operon transcription during methanogenesis, which expands our understanding of the unique regulatory mechanisms that govern methanogenesis. The findings presented in this study can further our understanding of how genetic regulation can effectively reduce the methane emissions caused by methanogens.

The preparation of polyclonal antibody against chlordimeform and establishment of detection by indirect competitive ELISA.

Objective: Chlordimeform is a chemical pesticide that is highly carcinogenic and toxic. The purpose of this study was to establish an enzyme-linked immunosorbent assay (ELISA) method for the detection of chlordimeform in aquaculture and fish farming. METHODS: Chlordimeform was coupled with bovine serum albumin (BSA) and ovalbumin (OVA) as carrier proteins. A chlordimeform-BSA conjugate was used as an immunogen, and chlordimeform-OVA was used as a coating antigen. Chlordimeform-BSA was used to immunize rabbits, and a polyclonal antibody was prepared. An indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) was established to detect chlordimeform. RESULTS: The working range of the established IC-ELISA method for chlordimeform detection was 1-20 ng/mL. The IC50 was 3.126 ng/mL, and the lower limit of detection (LOD) of chlordimeform was 0.637 ng/mL. The recovery of chlordimeform from spiked water samples ranged from 81% to 107%. CONCLUSION: An anti-chlordimeform polyclonal antibody was successfully developed, and a novel IC-ELISA was established to detect chlordimeform in aquaculture.

IL-33 Protects Mice against DSS-Induced Chronic Colitis by Increasing Both Regulatory B Cell and Regulatory T Cell Responses as Well as Decreasing Th17 Cell Response.

BACKGROUND: Previously, we have reported that IL-33 functioned as a protective modulator in dextran sulfate sodium- (DSS-) induced chronic colitis by suppressing Th17 cell response in colon lamina propria and IL-33 induced both regulatory B cells (Bregs) and regulatory T cells (Tregs) in mesenteric lymph nodes (MLNs) of mice with DSS-induced acute colitis. Moreover, we speculated that IL-33 would promote the Treg or Breg responses leading to the attenuation of DSS-induced chronic colitis. So, we investigated the role of IL-33 on Bregs and Tregs in the MLN of DSS-induced chronic colitis mice. METHODS: IL-33 was administered by intraperitoneal injection to mice with DSS-induced chronic colitis. Clinical symptoms, colon length, and histological changes were determined. The production of cytokines was measured by ELISA. The T and B cell subsets were measured by flow cytometry. The expression of mRNA of transcription factors was measured by quantitative real-time PCR. RESULTS: We show that IL-33 treatment increases both Breg and Treg responses in the MLN of mice with DSS-induced chronic colitis. Moreover, IL-33 treatment also decreases Th17 cell response in the MLN of mice with DSS-induced chronic colitis. CONCLUSION: Our data provide clear evidence that IL-33 plays a protective role in DSS-induced chronic colitis, which is closely related to increasing Breg and Treg responses in the MLN of mice as well as suppressing Th17 cell responses.