Novel thermo-alkali-stable cellulase-producing Serratia sp. AXJ-M cooperates with Arthrobacter sp. AXJ-M1 to improve degradation of cellulose in papermaking black liquor.

There is an urgent requirement to treat cellulose present in papermaking black liquor since it induces severe economic wastes and causes environmental pollution. We characterized cellulase activity at different temperatures and pH to seek thermo-alkali-stable cellulase-producing bacteria, a natural consortium of Serratia sp. AXJ-M and Arthrobacter sp. AXJ-M1 was used to improve the degradation of cellulose. Notably, the enzyme activities and the degradation rate of cellulose were increased by 30%-70% and 30% after co-culture, respectively. In addition, the addition of cosubstrates increased the degradation rate of cellulose beyond 30%. The thermo-alkali-stable endoglucanase (bcsZ) gene was derived from the strain AXJ-M and was cloned and expressed. The purified bcsZ displayed the maximum activity at 70 °C and pH 9. Mn2+, Ca2+, Mg2+ and Tween-20 had beneficial effects on the enzyme activity. Structurally, bcsZ potentially catalyzed the degradation of cellulose. The co-culture with ligninolytic activities significantly decreased target the parameters (cellulose 45% and COD 95%) while using the immobilized fluidized bed reactors (FBRs). Finally, toxicological tests and antioxidant enzyme activities indicated that the co-culture had a detoxifying effect on black liquor. Our study showed that Serratia sp. AXJ-M acts synergistically with Arthrobacter sp. AXJ-M1 may be potentially useful for bioremediation for black liquor.

Rapid and Broad Immune Efficacy of a Recombinant Five-Antigen Vaccine against Staphylococcus Aureus Infection in Animal Models.

Staphylococcus aureus (S.aureus) is a leading cause of both healthcare-and community-associated infections globally, which result in severe disease and readily developing antibiotic resistance. Developing an efficacious vaccine against S.aureus is urgently required. In the present study, we selected five conserved antigens, including the secreted factors α-hemolysin (Hla), staphylococcal enterotoxin B (SEB) and the three surface proteins staphylococcal protein A (SpA), iron surface determinant B N2 domain (IsdB-N2) and manganese transport protein C (MntC). They were all well-characterized virulence factor of S. aureus and developed a recombinant five-antigen S. aureus vaccine (rFSAV), rFSAV provided consistent protection in S. aureus lethal sepsis and pneumonia mouse models, and it showed broad immune protection when challenged with a panel of epidemiologically relevant S. aureus strains. Meanwhile, rFSAV immunized mice were able to induce comprehensive cellular and humoral immune responses to reduce bacterial loads, inflammatory cytokine expression, inflammatory cell infiltration and decrease pathology after challenge with a sub-lethal dose of S. aureus. Moreover, the importance of specific antibodies in protection was demonstrated by antibody function tests in vitro and in vivo. Altogether, our data demonstrate that rFSAV is a potentially promising vaccine candidate for defensing against S. aureus infection.

Protective efficacy of the chimeric Staphylococcus aureus vaccine candidate IC in sepsis and pneumonia models.

Staphylococcus aureus causes serious sepsis and necrotic pneumonia worldwide. Due to the spread of multidrug-resistant strains, developing an effective vaccine is the most promising method for combating S. aureus infection. In this study, based on the immune-dominant areas of the iron surface determinant B (IsdB) and clumping factor A (ClfA), we designed the novel chimeric vaccine IsdB151-277ClfA33-213 (IC). IC formulated with the AlPO4 adjuvant induced higher protection in an S. aureus sepsis model compared with the single components alone and showed broad immune protection against several clinical S. aureus isolates. Immunisation with IC induced strong antibody responses. The protective effect of antibodies was demonstrated through the opsonophagocytic assay (OPA) and passive immunisation experiment. Moreover, this new chimeric vaccine induced Th1/Th17-skewed cellular immune responses based on cytokine profiles and CD4(+) T cell stimulation tests. Neutralisation of IL-17A alone (but not IFN-γ) resulted in a significant decrease in vaccine immune protection. Finally, we found that IC showed protective efficacy in a pneumonia model. Taken together, these data provide evidence that IC is a potentially promising vaccine candidate for combating S. aureus sepsis and pneumonia.

A novel nitro-dexamethasone inhibits agr system activity and improves therapeutic effects in MRSA sepsis models without antibiotics.

Methicillin-resistant Staphylococcus aureus (MRSA) sepsis is a life-threatening medical condition that involves systemic inflammation throughout the body. Glucocorticoids are widely used in combination with antibiotics in the treatment of MRSA sepsis to fight the overwhelming inflammation. Here, we describe the improved anti-inflammatory properties of a nitric oxide (NO)-releasing derivative of dexamethasone, ND8008. ND8008 affected MRSA biofilm formation, caused biofilm cell death, and reduced the effects of virulence factors, such as α-toxin, by inhibiting the activity of the Staphylococcus aureus accessory gene regulator (agr) system. Dosing of mice with ND8008 (127.4 nmol/kg, i.p.) alone greatly reduced the inflammatory response caused by MRSA blood stream infection and considerably increased the survival rate of septic mice. These findings suggest that this novel NO-releasing derivative of dexamethasone ND8008 could be helpful in the treatment of MRSA sepsis.

Protective Efficacy and Mechanism of Passive Immunization with Polyclonal Antibodies in a Sepsis Model of Staphylococcus aureus Infection.

Staphylococcus aureus (S. aureus) is an opportunistic bacterial pathogen responsible for a diverse spectrum of human diseases, resulting in considerable yearly mortality rates. Due to its rapid acquisition of antibiotic resistance, it becomes increasingly difficult to cure S. aureus infections with conventional antibiotics. Immunotherapy represents a promising alternative strategy to prevent and/or treat the infection. In the present study, passive immunization with polyclonal antibodies targeting three possible S. aureus antigens, Hla, SEB and MntC (termed "SAvac-pcAb") after challenge with lethal dose of S. aureus resulted in reduced bacterial loads, inflammatory cell infiltration and decreased pathology, and was able to provide nearly complete protection in a murine sepsis model. In vitro studies confirmed the direct interaction of SAvac-pcAb with S. aureus bacteria. Additional studies validated that SAvac-pcAb contained both opsonic and neutralizing antibodies that contributed to its protective efficacy. The former mediated opsonophagocytosis in a neutrophil-dependent manner, while the later inhibited the biological functions of Hla and SEB, two major virulence factors secreted by S. aureus. Critically, we demonstrated that SAvac-pcAb was cross-reactive with different clinical strains of S. aureus. These results confirmed the efficacy for treatment of S. aureus infection by passive immunization as an important therapeutic option.

Identification of the immunodominant regions of Staphylococcus aureus fibronectin-binding protein A.

Staphylococcus aureus is an opportunistic bacterial pathogen responsible for a diverse spectrum of human diseases and a leading cause of nosocomial and community-acquired infections. Development of a vaccine against this pathogen is an important goal. The fibronectin binding protein A (FnBPA) of S. aureus is one of multifunctional 'microbial surface components recognizing adhesive matrix molecules' (MSCRAMMs). It is one of the most important adhesin molecules involved in the initial adhesion steps of S. aureus infection. It has been studied as potential vaccine candidates. However, FnBPA is a high-molecular-weight protein of 106 kDa and difficulties in achieving its high-level expression in vitro limit its vaccine application in S. aureus infection diseases control. Therefore, mapping the immunodominant regions of FnBPA is important for developing polyvalent subunit fusion vaccines against S. aureus infections. In the present study, we cloned and expressed the N-terminal and C-terminal of FnBPA. We evaluated the immunogenicity of the two sections of FnBPA and the protective efficacy of the two truncated fragments vaccines in a murine model of systemic S. aureus infection. The results showed recombinant truncated fragment F130-500 had a strong immunogenicity property and survival rates significantly increased in the group of mice immunized with F130-500 than the control group. We futher identified the immunodominant regions of FnBPA. The mouse antisera reactions suggest that the region covering residues 110 to 263 (F1B110-263) is highly immunogenic and is the immunodominant regions of FnBPA. Moreover, vaccination with F1B110-263 can generate partial protection against lethal challenge with two different S. aureus strains and reduced bacterial burdens against non-lethal challenge as well as that immunization with F130-500. This information will be important for further developing anti- S. aureus polyvalent subunit fusion vaccines.

Evaluation of the protective immunity of a novel subunit fusion vaccine in a murine model of systemic MRSA infection.

Staphylococcus aureus is a common commensal organism in humans and a major cause of bacteremia and hospital acquired infection. Because of the spread of strains resistant to antibiotics, these infections are becoming more difficult to treat. Therefore, exploration of anti-staphylococcal vaccines is currently a high priority. Iron surface determinant B (IsdB) is an iron-regulated cell wall-anchored surface protein of S. aureus. Alpha-toxin (Hla) is a secreted cytolytic pore-forming toxin. Previous studies reported that immunization with IsdB or Hla protected animals against S. aureus infection. To develop a broadly protective vaccine, we constructed chimeric vaccines based on IsdB and Hla. Immunization with the chimeric bivalent vaccine induced strong antibody and T cell responses. When the protective efficacy of the chimeric bivalent vaccine was compared to that of individual proteins in a murine model of systemic S. aureus infection, the bivalent vaccine showed a stronger protective immune response than the individual proteins (IsdB or Hla). Based on the results presented here, the chimeric bivalent vaccine affords higher levels of protection against S. aureus and has potential as a more effective candidate vaccine.