Crohn’s disease (CD) is a chronic inflammatory bowel disease (IBD) of the digestive tract in humans. There is evidence that Parabacteroides distasonis could contribute to IBD. Here, we present the complete genome sequence of a strain designated CavFT-hAR46, which was isolated from a gut intramural cavernous fistulous tract (CavFT) microlesion in a CD patient.

Chicken embryo fibroblasts (CEFs) are among the most commonly used cells for the study of interactions between chicken hosts and H5N1 avian influenza virus (AIV). In this study, the expression of eleven housekeeping genes typically used for the normalization of quantitative real-time PCR (QPCR) analysis in mammals were compared in CEFs infected with H5N1 AIV to determine the most reliable reference genes in this system. CEFs cultured from 10-day-old SPF chicken embryos were infected with 100 TCID(50) of H5N1 AIV and harvested at 3, 12, 24 and 30 hours post-infection. The expression levels of the eleven reference genes in infected and uninfected CEFs were determined by real-time PCR. Based on expression stability and expression levels, our data suggest that the ribosomal protein L4 (RPL4) and tyrosine 3-monooxygenase tryptophan 5-monooxygenase activation protein zeta polypeptide (YWHAZ) are the best reference genes to use in the study of host cell response to H5N1 AIV infection. However, for the study of replication levels of H5N1 AIV in CEFs, the β-actin gene (ACTB) and the ribosomal protein L4 (RPL4) gene are the best references.

BackgroundThe selection of stably expressed reference genes is a prerequisite when evaluating gene expression, via real-time PCR, in cells in response to viral infections. The objective of our study was to identify suitable reference genes for mRNA expression analysis in chicken embryonic fibroblasts (CEF) after infection with avian leukosis virus subgroup J (ALV-J).FindingsThe expression levels of 11 potential reference genes in CEF infected with ALV-J were determined by real-time PCR. The expression stability of these genes were analyzed and ranked using the geNorm tool. Analysis indicated that the genes RPL30 (ribosomal protein L30) and SDHA (succinate dehydrogenase complex, subunit A) were the most stably expressed genes in the ALV-J infected CEF.ConclusionsThe RPL30 and SDHA were deemed suitable for use as reference genes for real-time PCR analysis of mRNA gene expression during ALV-J infection, whereas commonly used ACTB and GAPDH are unsuitable to be reference genes.

A real-time PCR for detection and quantification of M. ovipneumoniae was developed using 9 recently sequenced M. ovipneumoniae genomes and primers targeting a putative adhesin gene p113. The assay proved to be specific and sensitive (with a detection limit of 22 genomic DNA) and could quantify M. ovipneumoniae DNA over a wide linear range, from 2.2 × 102 to 2.2 × 107 genomes.

Mycoplasma ovipneumoniae is associated with chronic nonprogressive pneumonia in both sheep and goats. Studies concerning its molecular pathogenesis, genetic analysis, and vaccine development have been hindered due to limited genomic information. Here, we announce the first complete genome sequence of this organism.Mycoplasma ovipneumoniae is considered to be a cause of nonprogressive pneumonia of both sheep and goats (1, 4, 7). As well as causing disease in its own right, M. ovipneumoniae predisposes animals to invasion by more serious respiratory pathogens, such as Mannheimia haemolytica, Pasteurella, and parainfluenza 3 virus (3, 5). Although M. ovipneumoniae is an important concern in the sheep and goat industry, little genomic information is available. Therefore, we sequenced the complete genome of M. ovipneumoniae to facilitate related studies in the future.M. ovipneumoniae strain SC01 was isolated from the lung of a goat with pneumonia in Sichuan Province, China. Genomic DNA was extracted and sequenced using the Illumina GA. We constructed a 350-bp library and obtained 216.12 Mb of clean reads (97.69% paired-end reads). The reads were assembled using SOAP de novo (6). Gene prediction was performed using Glimmer3.0. rRNAs were analyzed with rRNAmmer. tRNA sequences were predicted by tRNAscan-SE. The genes were annotated through Blast searches in the databases Swiss-Prot and TrEMBL (release 2010-04), COG (version 2.0), KEGG (release 48.2), and NCBI-NR (release 2010-8-7).The M. ovipneumoniae SC01 genome is 1,020,601 bp in length, with a GC content of 28.85%. The genome contains 864 putative coding sequences (CDSs) with an average gene length of 950 bp, and the mean coding percentage is 80.48%. There is only a single 16S-23S rRNA operon, and the 5S rRNA gene is separate from the 16S-23S rRNA operon. A total of 30 tRNA genes were identified. One prominent feature of the SC01 genome is the frequent usage of UUG as initiation codon. The number of UUG initiation codons is as high as 187, and these codons account for 21.6% of all initiation codons, which is the highest percentage among the mycoplasma genome sequences available so far.M. ovipneumoniae strain SC01 contains several recognizable genes likely to be involved in virulence. Two proteins are highly similar to bacterial toxins, hemolysin A (hlyA) and hemolysin C (hlyC). They are also present in other mycoplasmas, such as M. hyopneumoniae, M. capricolum, and M. conjunctivae (2), although their contribution to pathogenicity in Mycoplasma species is unclear. Niang et al. (7) reported the correlation between ciliostasis induced by M. ovipneumoniae and hydrogen peroxide production. We identified three genes, glpF, glpK, and glpD, involved in glycerol import and production of H 2 O 2 and reactive oxygen species. Mycoplasma infection depends on adherence to host epithelial cells. M. ovipneumoniae SC01 possesses at least 8 CDSs encoding proteins homologous to adhesin-like proteins of M. hyopneumoniae which is phylogenetically closely related to M. ovipneumoniae (8),...

Duck enteritis virus (DEV) is a herpesvirus that causes an acute, contagious, and fatal disease. In the present article, we introduce a quantitative real-time polymerase chain reaction (PCR) assay for DEV DNA using TaqMan technology and a two-step protocol. It was confirmed to be rapid, sensitive, and specific for DEV detection. The primers and probe were designed and directed to the DNA polymerase gene of DEV. The method will provide a valuable tool for rapid laboratory diagnosis of DEV infection. By virtue of its high-throughput format and its ability to accurately quantify the viral DNA, the method may be useful for large epidemiological surveys and clarification of pathogenesis, such as latency and reactivation of the virus.

Newcastle disease virus (NDV), formally recognized as avian paramyxovirus 1 (APMV-1), is the etiological agent of Newcastle disease (ND), an affliction which can cause severe losses in the poultry industry. Better understanding of the molecular basis of viral structural genes involved with production should contribute significantly toward the development of improved prophylactic and therapeutic reagents to control the infection.Here we show that a short hairpin RNA (shRNA) eukaryotic expression vector targeting nucleocapsid (NP) gene of NDV can potently inhibit NDV production in both primary cells and embryonated chicken eggs. Moreover, shRNA specific for NP abolished the accumulation of not only the corresponding mRNA but also P, HN, F, M gene mRNA. The findings reveal that newly synthesized NP mRNA is essential for NDV transcription and replication, and provide a basis for the development of shRNAs as a prophylaxis and therapy for NDV infection in poultry.

Duck hepatitis A virus genotype C (DHAV-C), recognized recently, is one of the pathogens causing fatal duck viral hepatitis in ducklings, especially in Asia. To demonstrate the pathogenesis of the DHAV-C isolate, 3-day-old specific pathogen free ducklings were inoculated subcutaneously with a DHAV-C isolate and the clinical signs were observed. Virus distribution, histological and apoptotic morphological changes of various tissues were examined at different times post inoculation. The serial, characteristic changes included haemorrhage and swelling of the liver. Apoptotic cells and virus antigen staining were found in all of the tissues examined. Where more virus antigen staining was detected, there were more severe histopathological and apoptotic changes. The amount of virus antigen and the histological and apoptotic morphological changes agreed with each other and became increasingly severe with length of time after infection. Apoptotic cells were ubiquitously distributed, especially among lymphocytes, macrophages and monocytes in immune organs such as the bursa of Fabricius, thymus and spleen, and in liver, kidney and cerebral cells. Necrosis was also observed within 72 h post inoculation in all organs examined, except the cerebrum, and was characterized by cell swelling and collapsed plasma membrane. These results suggest that the recent outbreak of disease caused by DHAV-C virus is pantropic, causing apoptosis and necrosis of different organs. The apoptosis and necrosis caused by the DHAV-C field strain in this study is associated with pathogenesis and DHAV-C-induced lesions.