Evaluation of a Brucella melitensis mutant deficient in O-polysaccharide export system ATP-binding protein as a rough vaccine candidate.

Rough Brucella mutants have been sought as vaccine candidates that do not interfere with the conventional serological diagnosis of brucellosis. In this study, a rough mutant of Brucella melitensis was generated by the disruption of the wzt gene, which encodes the O-polysaccharide (O-PS) export system ATP-binding protein. In vivo, the mutant 16MΔwzt was attenuated and conferred a level of protection against B. melitensis 16M challenge similar to that conferred by the vaccine strain B. melitensis M5 in mice. In pregnant sheep, the mutant 16MΔwzt did not induce abortion. In vitro, 16MΔwzt was more susceptible to polymyxin B and complement-mediated killing than B. melitensis 16M was. Most importantly, although 16MΔwzt had a rough phenotype, it was able to synthesize O-PS and did not induce detectable specific antibodies in sheep. These results suggested that 16MΔwzt deserved to further systematic evaluation as a vaccine for target animal hosts due to its promising features.

Full genome sequence of a bovine enterovirus isolated in china.

We report the full genome sequence of an isolate of bovine enterovirus type B from China. The virus (BEV-BJ001) was isolated from Beijing, China, from fecal swabs of cattle suffering from severe diarrhea. This genome sequence will give useful insight for future molecular epidemiological studies in China.

The Cyclic AMP-binding protein CbpB in Brucella melitensis and its role in cell envelope integrity, resistance to detergent and virulence.

Brucella melitensis possesses an operon with two components: the response regulator OtpR and a putative cAMP-dependent protein kinase regulatory subunit encoded by the BMEI0067 gene. In the previous study, the function of OtpR has been studied, while little is known about the function of the BMEI0067 gene. Using a bioinformatics approach, we showed that the BMEI0067 gene encodes an additional putative cAMP-binding protein, which we refer to as CbpB. Structural modeling predicted that CbpB has a cAMP-binding protein (CAP) domain and is structurally similar to eukaryotic protein kinase A regulatory subunits. Here, we report the characterization of CbpB, a cAMP-binding protein in Brucella melitensis, showed to be involved in mouse persistent infections. ∆cbpB::km possessed cell elongation, bubble-like protrusions on cell surface and its resistance to environmental stresses (temperature, osmotic stress and detergent). Interestingly, comparative real-time qPCR assays, the cbpB mutation resulted in significantly different expression of aqpX and several penicillin-binding proteins and cell division proteins in Brucella. Combined, these results demonstrated characterization of CbpB in B. melitensis and its key role for intracellular multiplication.

Different genotypes of nephropathogenic infectious bronchitis viruses co-circulating in chicken population in China.

Chicken nephropathogenic infectious bronchitis (IB) was prevalent in the most chicken farms during recent years, although the IB vaccination program has been widely performed in China. To characterize the S1 protein of infectious bronchitis virus (IBV) from China, five representative nephropathogenic IB viruses isolated from chickens in different provinces were genetically and phylogenetically analyzed. The results showed that the length of the S1 genes of the isolates were quite different (1,617, 1,620, 1,623, 1,629, and 1,632 nucleotides, respectively). The homology of the nucleotides and amino acids among the five isolates were 76.7%-92.1% and 73.9%-89.5%, respectively, indicating a great variation in S1 genes of the isolates. The variation in S1 genes might affect the antigenicity and pathogenicity of the viruses. Genetically, point mutations, insertions, and deletions in the S1 protein can be observed at many positions, especially at the first 150 amino acids in the N-terminal of the S1 protein. Two motif cleavage sites (R-R-X-R-R/S, H-R-R-R-R/S) were observed in the five sequenced strains. Phylogenetic analysis suggested that they belonged to different lineages. These findings indicated that different genotypes of nephropathogenic IB viruses were co-circulating in the chicken population in China.

Genetic conservation of hemagglutinin gene of H9 influenza virus in chicken population in Mainland China.

The hemagglutinin (HA) genes of 12 H9N2 influenza virus strains isolated from chickens in Mainland China during the period 1995-2002 were genetically analyzed. All the isolates possessed the same amino acid motif -R-S-S-R/G-L- at the cleavage site of HA. Except for the conserved amino acids, as is the case in the other avian influenza viruses, located in the receptor binding site, all of the 12 isolates possessed N at amino acid position 183; A, T, or V at position 190; K at position 137, whereas the representative strains of the other lineage (except Dk/HK/Y280/97-like lineage) virus of H9N2 viruses had H, E, and R at these positions respectively. These could be considered as the partial molecular markers of the H9 viruses isolated from chickens in Mainland China. Phylogenetic analyses showed HA genes of these isolates belonged to that of A/duck/Hong Kong/Y280/97-like virus lineage. No A/quail/Hong Kong/Gl/97-like virus was found in chicken, population since the outbreak of H9N2 influenza in Mainland China in 1992. The available evidence indicates that HA genes of H9 influenza virus circulating in Mainland China during the past years were well conserved.

Phylogenetic analysis of neuraminidase gene of H9N2 influenza viruses prevalent in chickens in China during 1995-2002.

The neuraminidase (NA) genes of 12 H9N2 influenza virus strains isolated from diseased chickens in different farms in mainland China during 1995-2002 were amplified and sequenced. Amino acids at hemadsorbing (HB) site of these isolates are different from those of A/quail/Hong Kong/G1/97-like viruses and A/chicken/Korea/96-like viruses. Neuraminidases of the 12 strains had a deletion of 3 amino acid residues at positions 63-65 as compared to that of A/turkey/Wisconsin/189/66, while those of Korea and Pakistan H9N2 isolates had no deletion. Phylogenetic analyses showed NA gene of these isolates belonged to that of A/duck/Hong Kong/Y280/97-like virus lineage. NA gene of the H9N2 viruses isolated in Korea and Pakistan belonged to lineage different from those of the 12 isolates. The present results indicate that the NA of H9N2 strains isolated in mainland China during the past 8 years were well preserved and the geographical distribution play a significant role in the evolution of the H9N2 influenza viruses.