Serological responses to smallpox A33 antigen and monkeypox A35 antigen in healthy people and people living with HIV-1 from Guangzhou, China.

People are expected to have been previously vaccinated with a Vaccinia-based vaccine, as until 1980 smallpox vaccination was a standard protocol in China. It is unclear whether people with smallpox vaccine still have antibody against vaccinia virus (VACV) and cross-antibody against monkeypox virus (MPXV). Herein, we assessed the binding antibodies with antigen of VACV-A33 and MPXV-A35 in the general population and HIV-1 infected patients. Firstly, we detected VACV antibody with A33 protein to evaluate the efficiency of smallpox vaccination. The result show that 29% (23 of 79) of hospital staff (age ≥ 42 years) and 63% (60 of 95) of HIV-positive patients (age ≥ 42 years) from Guangzhou Eighth People's Hospital were able to bind A33. However, among the subjects below 42 years of age, 1.5% (3/198) of the hospital volunteer samples and 1% (1/104) of the samples from HIV patients were positive for antibodies against A33 antigen. Then, we assessed the specific cross-reactive antibodies against MPXV A35 protein. 24% (19 of 79) hospital staff (age〉 = 42 years) and 44% (42 of 95) of HIV-positive patients (age〉 = 42 years) were positive. 98% (194/198) of the hospital staff and 99% (103/104) of the HIV patients had no A35-binding antibodies. Further, we found significant sex differences for the reactivity to A35 antigen were observed in HIV population, but no significant sex differences in hospital staff. Further, we analyzed the positivity rate of anti-A35 antibody of men who have sex with men (MSM) and non-MSM in HIV patients (age〉 = 42years). We found that 47% of no-MSM population and 40% of MSM population were positive for A35 antigen, with no significant difference. Lastly, we found only 59 samples were positive for anti-A33 IgG and anti-A35 IgG in all participants. Together, we demonstrated A33 and A35 antigens binding antibodies were detected in HIV patients and general population who were older than 42 years, and cohort studies only provided data of serological detection to support early response to monkeypox outbreak.

Transcription factor GlbHLH regulates hyphal growth, stress resistance, and polysaccharide biosynthesis in Ganoderma lucidum.

Basic helix-loop-helix (bHLH) transcription factors (TFs) participate in many physiological and cellular processes in eukaryotes. However, their functions remain unclear in the macro basidiomycete Ganoderma lucidum (G. lucidum). In this study, a gene encoding bHLH TF, GlbHLH, was identified in G. lucidum. The knockdown of GlbHLH by RNA interference reduced hyphal growth, hyphal branching, and resistant to osmotic, oxidative, and cell wall stress. The content of cell wall components ß-1,3 glucan and chitin and the expression of their synthesis genes were decreased in the GlbHLH knockdown strains. The knockdown of GlbHLH led to an increase of intracellular reactive oxygen species by decreasing the enzyme activity and gene expression of antioxidant enzymes. Furthermore, the production of intracellular polysaccharides and extracellular polysaccharides was greatly decreased in the GlbHLH mutants. These results suggested that GlbHLH is involved in hyphal growth, stress response, and polysaccharide biosynthesis in G. lucidum.

Enhanced exopolysaccharide production in submerged fermentation of Ganoderma lucidum by Tween 80 supplementation.

Bioactive polysaccharides extracted from Ganoderma lucidum (G. lucidum) have been widely applied in food and medicine for their multiple functions. In this study, G. lucidum exopolysaccharide (EPS) production in submerged fermentation was stimulated by Tween 80. The addition of 0.25% Tween 80 on day 3 gave a maximum production of mycelial biomass and EPS, with an increase of 19.76 and 137.50%, respectively. Analysis of fermentation kinetics showed that glucose was consumed faster after adding Tween 80, while the expression of EPS biosynthesis-related genes and ATP generation were greatly improved. Moreover, Tween 80 resulted in the significant accumulation of reactive oxygen species and increased cell membrane and cell wall permeability. The EPS from Tween 80-containing medium had higher contents of carbohydrate and uronic acid, lower molecular weight, and higher antioxidant activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals than those of EPS produced in the absence of Tween 80. This study provides further evidence to clarify the stimulatory effects of Tween 80 in fermentation and provides a guide for the production of bioactive G. lucidum EPS.

Effects of Christensenella minuta lipopolysaccharide on RAW 264.7 macrophages activation.

Christensenella minuta (C. minuta) is a gram-negative gastrointestinal bacterium associated with weight loss. However, recent studies have shown that C. minuta might be a potential pathogen and thus limited its application in the control of obesity. Research into the genetic characteristics and pathogenicity of C. minuta remain elusive. As a major virulence factor of gram-negative bacteria, lipopolysaccharide (LPS) can induce various diseases. In this study, we report the complete genome sequence of C. minuta and have also identified some genes related to LPS biosynthesis. The structure of C. minuta LPS, detected by SDS-PAGE, was different from that of Escherichia coli (E. coli) LPS. The incubation of RAW 264.7 macrophages with C. minuta LPS resulted in lower levels of cellular proliferation, phagocytosis and nuclear factor-kappa B (NF-κB) activation as compared to incubation with E. coli LPS. Furthermore, the expression of pro-inflammatory cytokines, as well as nitric oxide and reactive oxygen species production, was induced in C. minuta LPS-treated cells but to a much lower extent than that by E. coli LPS. These findings show that C. minuta LPS acts as a weak agonist for RAW 264.7 macrophages and can only trigger a weak inflammatory response through the NF-κB signalling pathway. In conclusion, these results suggest that the toxicity of C. minuta LPS is significantly attenuated due to its atypical structure and weak agonist activity for RAW 264.7 macrophages.

Novel Cysteine Desulfidase CdsB Involved in Releasing Cysteine Repression of Toxin Synthesis in Clostridium difficile.

Clostridium difficile, a major cause of nosocomial diarrhea and pseudomembranous colitis, still poses serious health-care challenges. The expression of its two main virulence factors, TcdA and TcdB, is reportedly repressed by cysteine, but molecular mechanism remains unclear. The cysteine desulfidase CdsB affects the virulence and infection progresses of some bacteria. The C. difficile strain 630 genome encodes a homolog of CdsB, and in the present study, we analyzed its role in C. difficile 630Δerm by constructing an isogenic ClosTron-based cdsB mutant. When C. difficile was cultured in TY broth supplemented with cysteine, the cdsB gene was rapidly induced during the exponential growth phase. The inactivation of cdsB not only affected the resistance of C. difficile to cysteine, but also altered the expression levels of intracellular cysteine-degrading enzymes and the production of hydrogen sulfide. This suggests that C. difficile CdsB is a major inducible cysteine-degrading enzyme. The inactivation of the cdsB gene in C. difficile also removed the cysteine-dependent repression of toxin production, but failed to remove the Na2S-dependent repression, which supports that the cysteine-dependent repression of toxin production is probably attributable to the accumulation of cysteine by-products. We also mapped a δ54 (SigL)-dependent promoter upstream from the cdsB gene, and cdsB expression was not induced in response to cysteine in the cdsR::ermB or sigL::ermB strain. Using a reporter gene fusion analysis, we identified the necessary promoter sequence for cysteine-dependent cdsB expression. Taken together, these results indicate that CdsB is a key inducible cysteine desulfidase in C. difficile which is regulated by δ54 and CdsR in response to cysteine and that cysteine-dependent regulation of toxin production is closely associated with cysteine degradation.