MicroRNA profile of bovine monocyte-derived macrophages infected in vitro with Streptococcus agalactiae
ABSTRACT: In a subclinical infection such as bovine streptococcal mastitis, early recognition is a great challenge, and miRNAs profiling could potentially assist in the diagnosis and contribute to the understanding of pathogenicity and defense mechanisms. We have examined the miRNA repertoire during the early phase response of bovine macrophages to in vitro infection with live Streptococcus agalactiae. Next generation sequencing of 20 small RNA libraries from blood monocyte-derived macrophages exposed to two sequence types of S. agalactiae (ST103 and ST12) for 6 hours in vitro was performed. Analyzes of over 356 million of high quality sequence reads, revealed that 17 and 44 miRNAs were differentially expressed (P < 0.05) between the control unchallenged macrophages and the macrophages infected with ST103 and ST12, respectively. We also identified the expression of 31 potentially novel bovine miRNAs.
Project description:We determined dynamic behavior of S. agalactiae transcriptome during growth in THY medium and detected growth phase regulated genes Overall design: Three independent cultures (three biological replicates) of S. agalactiae NEM316 strain in rich laboratory medium were sampled at mid log, late log early and late stationary growth phase.
Project description:Analysis of S. agalactiae response to dltA mutation the role of DltR. This analysis is based on the comparison of the effect of dltA and dltR mutation on S. agalactiae transcription profile.
Project description:Macrophages from cattles with different infectious status of bovine tuberculosis have different responses to in vitro Mycobacterium bovis challenge. This is confirmed in our previous study exploring several immune-related genes using qPCR. Microarrays can help us better understand the differences by screening thousands of genes. Monocytes Derived Macrophages from 3 TB-infected cattles and 3 TB-free cattles were challenged with Mycobacterium bovis at a MOI of 10 at 6 hours, and the control group were the same unchallenged macrophages at 6 hours.
Project description:We determined dynamic behavior of S. agalactiae transcriptome during growth in THY medium and detected growth phase regulated genes Three independent cultures (three biological replicates) of S. agalactiae NEM316 strain in rich laboratory medium were sampled at mid log, late log early and late stationary growth phase.
Project description:In this study, we determined the miRNA expression profile of bovine alveolar macrophages, using next-generation sequencing strategy. On an Illumina HiSeq 2000 machine, we sequenced 8 miRNA libraries, prepared from small RNA fractions of alveolar macrophages isolated from 8 different healthy animals (Bos taurus). From the data, the potential novel miRNAs were predicted, and the expression levels of the known miRNAs were determined. We report that 80 known bovine miRNAs are expressed in bovine alveolar macrophages with >100 reads per million. The most highly expressed miRNA was bta-miR-21, followed by bta-miR-27a. Additionally, one putatively novel bovine miRNA was identified. To our knowledge, this is the first RNA-seq study to profile miRNA expression in bovine alveolar macrophages and provides an important reference dataset for investigating the regulatory roles miRNAs play in this important immune cell type. Examination of bovine alveolar macrophage miRNA profiles, using RNA-seq. Alveolar macrophages were isolated from lung lavages from 8 animals. Small RNA fractions were prepared from the cells using the Qiagen RNeasy Plus mini kit, and miRNA sequencing libraries were prepared using the Epicentre Scriptminer multiplex kit. The sequencing was performed on an Illumina HiSeq 2000 machine.
Project description:Streptococcus agalactiae (Lancefield’s group B Streptococcus, GBS) is a major bacterial species of genus Streptococcus and has medical and veterinary importance by affecting mainly humans (Maione et al., 2005; Johri et al., 2006), cattle (Keefe, 1997) and fish (Mian et al., 2009). The GBS is the most important pathogen for the Nile tilapia, a global commodity of the aquaculture sector, causing outbreaks of septicemia and meningoencephalitis (Hernández et al., 2009; Mian et al., 2009).
Project description:The purpose of this study was to determine what S. agalactiae genes are under the control of the MtaR regulatory protein. Inactivation of the Streptococcus agalactiae mtaR gene resulted in the downregulation of 11 genes and the upregulation of 1 gene in the mtaR mutant. Genes involved in the uptake of methionine and a gene involved in the degradation of peptides were downregulated. Also, genes involved in the transport and biosynthesis of arginine were downregulated. The gene encoding the virulence factor cspA was downregulated, as well as a gene showing similarity to plasminogen activators. Thus, the expression of genes involved in both metabolic functions and virulence are under the influence of mtaR. Overall design: Three cultures of an mtaR insertional mutant (DS101) and three cultures of the isogenic wild-type strain (COH1) were grown in a chemically-defined medium. Cells were disrupted with glass-bead and RNA was isolated by a Trizol method. A custom Affymetrix chip was utilized for microarray analysis. Genes were considered differentially-expressed if they showed a >2.0-fold difference in expression between the strains and a Bayesian (Cyber-T) t-test value of <0.001. The posterior probability of differential expression (PPDE) (< p) value for the genes identified was >0.964.