Characterization and expression analysis of the groESL operon of Bartonella bacilliformis

The groESL operon of Bartonella bacilliformis, a facultative intracellular, Gram-negative bacterium and etiologic agent of Oroya Fever, was characterized. Sequence analysis revealed an operon containing two genes of 294 (groES) and 1632 nucleotides (groEL) separated by a 55-nt intergenic spacer. The operon is preceded by a 72-nt ORF (ORF1) that encodes a hypothetical protein with homology to a portion of the HrcA repressor for groESL. A divergent fumarate hydratase C (fumC) gene lies further upstream. Deduced amino acid sequences for B. bacilliformis GroEL and GroES revealed a high degree of identity with homologues from other Bartonella and alpha-Protebacteria. A single transcriptional start site (TSS) was mapped 79 nucleotides upstream of the groES start codon, regardless of incubation temperature. The TSS was located immediately 5\u27 to a potential controlling inverted repeat of chaperonin expression (CIRCE) element and is preceded by a sigma70-like promoter. The operon is followed by a predicted rho-independent transcriptional terminator. Northern blot analysis indicated that groES and groEL are co-transcribed as a single mRNA of approximately 2.4 kb. A 6-h time course analysis by qRT-PCR showed that groEL expression increases 1.3-fold within 30 min of a temperature upshift from 30 to 37 degrees C, with maximum transcription reached after 60 min (approximately 4.3-fold), followed by a steady decrease to background (30 degrees C) transcription levels by 6 h. Western blot analysis revealed a 1.4- and 1.5-fold increase in GroEL synthesis following a temperature upshift or by inhibiting DNA supercoiling with coumermycin A1, respectively. Functional expression and complementation of temperature-sensitive Escherichia coli groES or groEL mutants with the cloned operon allowed them to grow at otherwise restrictive temperatures

UK B.1.1.7 variant exhibits increased respiratory replication and shedding in nonhuman primates.

The continuing emergence of SARS-CoV-2 variants calls for regular assessment to identify differences in viral replication, shedding and associated disease. In this study, African green monkeys were infected intranasally with either a contemporary D614G or the UK B.1.1.7 variant. Both variants caused mild respiratory disease with no significant differences in clinical presentation. Significantly higher levels of viral RNA and infectious virus were found in upper and lower respiratory tract samples and tissues from B.1.1.7 infected animals. Interestingly, D614G infected animals showed significantly higher levels of viral RNA and infectious virus in rectal swabs and gastrointestinal tract tissues. Our results indicate that B.1.1.7 infection in African green monkeys is associated with increased respiratory replication and shedding but no disease enhancement similar to human B.1.1.7 cases. ONE-SENTENCE SUMMARY: UK B.1.1.7 infection of African green monkeys exhibits increased respiratory replication and shedding but no disease enhancement

Novel neutralizing monoclonal antibodies protect rodents against lethal filovirus challenges

Filoviruses are the causative agents of lethal hemorrhagic fever in human and non-human primates (NHP). The family of Filoviridae is composed of three genera, Ebolavirus, Marburgvirus and Cuevavirus. There are currently no approved vaccines or antiviral therapeutics for the treatment of filovirus infections in humans. Passive transfer of neutralizing antibodies targeting the Ebola virus (EBOV) glycoprotein (GP) has proven effective in protecting mice, guinea pigs and NHP from lethal challenges with EBOV. In this study, we generated two neutralizing monoclonal antibodies (MAbs), termed S9 and M4 that recognize the GP of EBOV or multiple strains of Marburg virus (MARV), respectively. We characterized the putative binding site of S9 as a linear epitope on the glycan cap of the GP1 subunit of the EBOV-GP. The M4 antibody recognizes an unknown conformational epitope on MARV-GP. Additionally, we demonstrated the post-exposure protection potential of these antibodies in both the mouse and guinea pig models of filovirus infection. These data indicate that MAbs S9 and M4 would be good candidates for inclusion in an antibody cocktail for the treatment of filovirus infections

A novel model of lethal Hendra virus infection in African green monkeys and the effectiveness of ribavirin treatment

The henipaviruses, Hendra virus (HeV) and Nipah virus (NiV), are emerging zoonotic paramyxoviruses that can cause severe and often lethal neurologic and/or respiratory disease in a wide variety of mammalian hosts, including humans. There are presently no licensed vaccines or treatment options approved for human or veterinarian use. Guinea pigs, hamsters, cats, and ferrets, have been evaluated as animal models of human HeV infection, but studies in nonhuman primates (NHP) have not been reported, and the development and approval of any vaccine or antiviral for human use will likely require efficacy studies in an NHP model. Here, we examined the pathogenesis of HeV in the African green monkey (AGM) following intratracheal inoculation. Exposure of AGMs to HeV produced a uniformly lethal infection, and the observed clinical signs and pathology were highly consistent with HeV-mediated disease seen in humans. Ribavirin has been used to treat patients infected with either HeV or NiV; however, its utility in improving outcome remains, at best, uncertain. We examined the antiviral effect of ribavirin in a cohort of nine AGMs before or after exposure to HeV. Ribavirin treatment delayed disease onset by 1 to 2 days, with no significant benefit for disease progression and outcome. Together our findings introduce a new disease model of acute HeV infection suitable for testing antiviral strategies and also demonstrate that, while ribavirin may have some antiviral activity against the henipaviruses, its use as an effective standalone therapy for HeV infection is questionable. Copyrigh

Loss of a Biofilm-Inhibiting Glycosyl Hydrolase during the Emergence of Yersinia pestis▿

Yersinia pestis, the bacterial agent of plague, forms a biofilm in the foregut of its flea vector to produce a transmissible infection. The closely related Yersinia pseudotuberculosis, from which Y. pestis recently evolved, can colonize the flea midgut but does not form a biofilm in the foregut. Y. pestis biofilm in the flea and in vitro is dependent on an extracellular matrix synthesized by products of the hms genes; identical genes are present in Y. pseudotuberculosis. The Yersinia Hms proteins contain functional domains present in Escherichia coli and Staphylococcus proteins known to synthesize a poly-β-1,6-N-acetyl-d-glucosamine biofilm matrix. In this study, we show that the extracellular matrices (ECM) of Y. pestis and staphylococcal biofilms are antigenically related, indicating a similar biochemical structure. We also characterized a glycosyl hydrolase (NghA) of Y. pseudotuberculosis that cleaved β-linked N-acetylglucosamine residues and reduced biofilm formation by staphylococci and Y. pestis in vitro. The Y. pestis nghA ortholog is a pseudogene, and overexpression of functional nghA reduced ECM surface accumulation and inhibited the ability of Y. pestis to produce biofilm in the flea foregut. Mutational loss of this glycosidase activity in Y. pestis may have contributed to the recent evolution of flea-borne transmission

Role of the Yersinia pestis Ail Protein in Preventing a Protective Polymorphonuclear Leukocyte Response during Bubonic Plague▿

The ability of Yersinia pestis to forestall the mammalian innate immune response is a fundamental aspect of plague pathogenesis. In this study, we examined the effect of Ail, a 17-kDa outer membrane protein that protects Y. pestis against complement-mediated lysis, on bubonic plague pathogenesis in mice and rats. The Y. pestis ail mutant was attenuated for virulence in both rodent models. The attenuation was greater in rats than in mice, which correlates with the ability of normal rat serum, but not mouse serum, to kill ail-negative Y. pestis in vitro. Intradermal infection with the ail mutant resulted in an atypical, subacute form of bubonic plague associated with extensive recruitment of polymorphonuclear leukocytes (PMN or neutrophils) to the site of infection in the draining lymph node and the formation of large purulent abscesses that contained the bacteria. Systemic spread and mortality were greatly attenuated, however, and a productive adaptive immune response was generated after high-dose challenge, as evidenced by high serum antibody levels against Y. pestis F1 antigen. The Y. pestis Ail protein is an important bubonic plague virulence factor that inhibits the innate immune response, in particular the recruitment of a protective PMN response to the infected lymph node