Jingliang Su

Complete genome sequence analysis of a predominant infectious bronchitis virus (IBV) strain in China.

Infectious bronchitis (IB) is one of the major diseases in poultry flocks all over the world caused by infectious bronchitis virus (IBV). In the study, the complete genome sequence of strain A2 was sequenced and analyzed, which was a predominant IBV strain in China. The results indicated that there were mutations, insertions, and deletions distributed in the whole genome. The A2 virus had the highest identity to S14 and BJ in terms of full genome, whereas had a further distance to Massachusetts strains. Phylogenetic analysis showed that A2 isolate clustered together with most Chinese strains. The results of this study suggest that strain A2 may play an important role in IBV’s evolution and A2-like IBVs are predominant strains in China.

Isolation and analysis of two naturally-occurring multi-recombination Newcastle disease viruses in China

Two Newcastle disease viruses (NDV), designated QG/Hebei/07 and XD/Shandong/08, were isolated from infected chicken flocks in China in 2007 and 2008, respectively. The results of phylogenetic and recombination analyses on complete NDV genome sequences (excluding terminal segments) show that the QG/Hebei/07 isolate had evidence of recombination in the M and F genes, and recombination in the XD/Shandong/08 isolate in the F, L genes and the non-coding region between the HN and L genes. These two naturally-occurring recombinants we found to be descended from at least three putative parents from vaccine and circulating virus lineages. Moreover, we found that evidence that homologous recombination also occurred between NDV viruses of chicken and swine lineages, while the major putative parent is likely to have been derived from the chicken avirulent vaccine lineage. This study suggests that homologous recombination can occur in all coding and non-coding regions of the NDV genome and a live vaccine strain is capable of recombination with circulating viruses resulting in significant genetic change. The potential role of swine-origin viruses in the evolution of virulent NDV warrants further investigation.

Cephalexin Residue Detection in Milk and Beef by ELISA and Colloidal Gold Based One-Step Strip Assay

An evaluation of a rapid enzyme-linked immunosorbent assay (ELISA) and colloidal gold based one-step strip assay for cephalexin (CEX) residue detection in milk and beef is described. A monoclonal antibody (mAb) against CEX was produced using cephalexin-bovine serum albumin (CEX-BSA) conjugate as the immunogen, which exhibited no cross-reactivity with applied chemicals in the studied concentration range. The detection limit of rapid ELISA was calculated as 0.39 microg/kg in PBS and 19.5 microg/kg in beef and milk, which was quite lower than the European Union Maximum Residue Limit (MRL) of 100 microg/kg in milk and 200 microg/kg in muscle. Spiked samples were detected with a mean recovery of 82.8-124% and coefficient of variation of 4.88-25%, which indicated a good agreement with the spiked concentration. Accuracy and reproducibility were determined using spiked samples with four different final concentrations of 1, 2, 5, and 10 microg/kg of CEX (n = 7). Mean intra-assay variation of 6.67% and inter-assay variation of 10.66% were obtained. In contrast, the strip test for CEX had a visual detection limit of 0.5 microg/kg, which could be evaluated within 3-10 min. However, positive samples should be further quantified by more sensitive and accurate competitive indirect ELISA method. In conclusion, the described strip test is rapid, simple, and cost-effective as well as sensitive and specific enough for reliable and accurate on-site screening.

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.

Molecular types and antibiotic resistance of Staphylococcus aureus isolates from bovine mastitis in a single herd in China

The molecular diversity, antibiotic resistance patterns and presence of resistance genes were determined in Staphylococcus aureus isolates from cases of bovine mastitis in a dairy cattle herd in China. Multiple locus variable number tandem repeat analysis was used for molecular typing. Resistance was determined through minimum inhibitory concentrations and resistance genes were detected by PCR. There was low molecular diversity; one predominant strain (type I) accounted for the majority of cases of S. aureus mastitis in the herd and this strain had a high frequency of resistance to penicillin and tetracycline. The most prevalent resistance genes were blaZ, ermC and tetM.

Isolation and characterization of a reovirus causing spleen necrosis in Pekin ducklings.

High rates of mortality for Pekin ducklings have been recorded in several duck farms in China since 2006. Dead ducklings were characterized by spleen necrosis, suggesting microbial infection as a cause of disease. Laboratory investigations led to the isolation of a virus strain from the spleen tissues of dead ducklings, designated DRV-HC. Subsequent experimental infections with DRV-HC resulted in marked spleen necrosis in the ducklings similar to those observed in the natural outbreaks. Electron microscopy of the cultured DRV-HC revealed viral particles that were non-enveloped and icosahedral with a mean diameter of approximately 72 nm. Agar gel precipitating tests showed that the isolate shared a common group-specific antigen with chicken reovirus S1133. DNA sequencing revealed that this isolate was closely related to Muscovy duck reoviruses. Experimental infection with DRV-HC resulted in death of young chicks with necrotic foci in the liver and spleen. To the best of our knowledge, this is the first report of the isolation of a duck reovirus with high virulence in Pekin ducklings and SPF chickens.

Duck Tembusu virus exhibits neurovirulence in BALB/c mice

BackgroundDuck Tembusu virus is a member of the Ntaya group in the genus Flavivirus. The virus has been responsible for severe duck egg-drop syndrome in China since 2010. Its emergence and rapid spread have caused great economic loss for the poultry industry. The epidemiology of the virus infection and the potential threat to public health is of great concern because of the infective and zoonotic nature of flaviviruses.ResultsIn this study, the pathogenicity of duck Tembusu virus in BALB/c mice was investigated. Infected mice developed clinical signs, including loss of appetite, ruffled hair, weight loss, disorientation, blindness and paralysis of hind limbs from six days post- infection following intracerebral inoculation. Morbidity was 100%, with mortality ranging from 20 to 80% in three- to eight-week-old mice. High virus titers were recovered from the brain, and the virus was distributed in several organs. Histologically, there was widespread non-suppurative encephalitis in the brain. Lymphocyte depletion in the spleen was observed, along with fatty degeneration in the liver and kidney.ConclusionsOur results demonstrate, for the first time, that duck Tembusu virus is highly neurovirulent in BALB/c mice. The mouse model used in this work was able to produce Tembusu virus infection and could be useful for elucidating some of the aspects of the pathophysiology of other flavivirus infections.

Antibiotic resistance of Streptococcus agalactiae from cows with mastitis

The aim of this study was to characterise the phenotypic and genotypic antibiotic resistance patterns of Streptococcus agalactiae isolated from cows with mastitis in China. Antibiotic resistance was based on minimum inhibitory concentrations and detection of resistance genes by PCR. S. agalactiae isolates most frequently exhibited phenotypic resistance to tetracycline, while the resistance genes most frequently detected were ermB, tetL and tetM. Resistance genes were detected in some susceptible isolates, whereas no resistance genes could be detected in some resistant isolates, indicating that the resistance genotype does not accurately predict phenotypic resistance.

Full Genome Sequences of Two Reticuloendotheliosis Viruses Contaminating Commercial Vaccines

Abstract Reticuloendotheliosis virus (REV) fragments are a common contaminant in some commercial vaccines such as fowl poxvirus (FPV) and Mareks disease virus. However, only those strains integrating or containing a near-intact REV provirus are more likely to cause problems in the field. We confirm here, by PCR assays and animal experiments, that vaccines against FPV and herpes virus of turkeys were contaminated with full genome sequences of REV. Further, we determined the complete proviral sequence of two REV isolates from contaminated vaccines. Two REV isolates (REV-99 and REV-06) present in the vaccines were both replication competent, and their proviral genome was 8286 nucleotides in length with two identical long terminal repeats (LTR). The complete genome in these two REV isolates shared 99.8% identity to APC-566 and fowl poxvirus REV proviral inserts (FPV-REV). REV-99 and REV-06 LTR showed over 99% identity to chicken syncytial virus (CSV), but an identity of only 75.8 % and 78.0%, respectively, to SNV. Alignments with other available REV gag, pol, and env sequences revealed high similarity at the nucleotide level. The results further indicated that the prototype CSV may be the most-important REV contaminant in the commercial vaccines, and distinct genotypes of REVs may cocirculate in chicken flocks of China at the present time.

An infectious full-length cDNA clone of duck Tembusu virus, a newly emerging flavivirus causing duck egg drop syndrome in China.

Duck Tembusu virus (TMUV) is a recently identified pathogenic flavivirus that causes severe egg drop and encephalitis in Chinese ducks and geese. It has been found to be most closely related to the mosquito-origin Tembusu virus and chicken Sitiawan virus reported in Malaysia. However, the ecological characteristics and the pathogenesis of duck TMUV are largely unknown. We report the construction of full-length cDNA clone of duck TMUV strain JXSP. The virus genome was reverse transcribed, amplified as seven overlapping fragments and successively ligated into the low copy number vector pWSK29 under the control of a T7 promoter. Transfection of BHK-21 cells with the transcribed RNA from the full-length cDNA clone resulted in production of highly infectious progeny virus. In vitro growth characteristics in BHK-21 cells and virulence in ducklings and BALB/c mice were similar for the rescued and parental viruses. This stable infectious cDNA clone will be a valuable tool for studying the genetic determinants of duck TMUV.