Project description:Background: Histomonas meleagridis is an anaerobic, intercellular parasite that infects the Galliformes such as turkeys and chickens. In recent years, the reemergence of Histomoniasis has caused serious economic losses as drugs to treat the disease have been banned. At present, studies on H. meleagridis mainly focus on virulence, gene expression analysis, and the innate immunity of the host. However, there are no studies on differential expression of miRNAs (DEMs) in host immune and inflammatory response induced by H. meleagridis infection in chickens. In this study, the expression profile of cecum miRNA at 10 and 15 days post-infection (DPI) with Chinese JSYZ-F strain H. meleagridis was studied by high-throughput sequencing. Results: Compared with the control group, 94 and 127 DEMs were found in the cecum of infected chickens at 10 DPI (CE vs CC) and 15 DPI (CEH vs CCH), respectively, of which 60 DEMs were shared at two-time points. Gene Ontology (GO) enrichment analysis of the target genes of DEMs showed that 881 and 1027 GO terms were significantly enriched at 10 and 15 DPI. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of the target genes of DEMs showed that only 5 and 3 pathways were significantly enriched at 10 and 15 DPI, respectively. The integrated analysis of miRNA–gene network revealed that the DEMs played important roles in the host immune and inflammatory responses to H. meleagridis infection by dynamically regulating the expression of immune and inflammation-related cytokines. Conclusion: Our results not only suggested that host miRNA expression was dynamically altered by H. meleagridis and host, but also revealed that more miRNAs and genes were involved in the later stage of the disease. In addition, host and H. meleagridis regulated the expression of immune and inflammation-related cytokines to respond to H. meleagridis infection. Our results will contribute to future research on miRNA-target interaction during H. meleagridis infection in chickens and provide new ideas for H. meleagridis control.
Project description:Background: Histomonas meleagridis is an anaerobic, intercellular parasite, which infects gallinaceous birds such as turkeys and chickens. In recent years, the reemergence of Histomoniasis has caused serious economic losses as drugs to treat the disease have been banned. At present, research of H. meleagridis focuses on virulence, gene expression analysis, and the innate immunity of the host. However, there are no study on the differentially expressed miRNAs (DEMs) associated with the liver induced by H. meleagridis. In this research, high-throughput sequencing was used to analyze the expression profile of liver miRNA at 10 and 15 days post-infection (DPI) in chickens infected with Chinese JSYZ-F strain H. meleagridis. Results: Compared with the uninfected control, 120 and 118 DEMs were found in the liver of infected chickens at 10 DPI and 15 DPI, respectively, of which 74 DEMs were shared at two-time points. Differentially expressed miRNAs were classified into three types according to the time of infection: L1, 45 miRNAs differentially expressed only at 10 DPI, were predicted to target 1646 genes; L2, 43 miRNAs differentially expressed only at 15 DPI, were predicted to target 2257 genes; L3, 75 miRNAs differentially expressed at both 10 DPI and 15 DPI, were predicted to target 1623 genes. A total of 89, 87 and 41 significantly enriched GO terms (p<0.05) were identified at L1, L2 and L3, respectively. The KEGG pathway analysis of differentially expressed miRNA target genes were shown that 3, 4 and 5 significantly enriched KEGG pathway (p<0.05) were identified at L1, L2 and L3, respectively. Conclusion: This article suggested that liver miRNA expression was dynamically altered by H. meleagridis and host and showed that the expression pattern of the L1 class DEMs were more favourable for controlling the development of the inflammatory response, whereas the L2 class DEMs were more favourable for enhancing the inflammatory response. Inflammation-associated miRNA expression patterns were consistent with the liver inflammatory process after artificial infection. The results of the study laid the foundation for an in-depth analysis of the pathogenic mechanism of H. meleagridis from the perspective of host miRNAs.
Project description:The current study aimed to detect and identify significant differentially expressed proteins between a virulent and an attenuated Histomonas meleagridis strain which was in vitro co-cultivated with Escherichia coli DH5α. Two-dimensional gel electrophoresis (2-DE) was used for proteome visualization , gel image software for computational detection of significantly up-regulated protein spots and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS) for protein identification. The statistical analysis fulfilling two criteria (> or = 3-fold up-regulation and P<0.05) detected 119 differentially expressed protein spots out of which 62 spots were located in gels of the virulent strain and 57 spots in gels of the attenuated strain. The mass spectrometric analysis of 32 spots, up-regulated in gels of the virulent strain, showed that they are of H. meleagridis origin. As opposed to this, the mass spectrometric analysis of 49 protein spots , up-regulated in the gels of the attenuated strain , identified 32 spots as specific to the protozoan. Additionally, the analysis identified a number of E. coli DH5α proteins which were detected as differentially expressed by the computational gel image and statistical analysis.
Project description:De novo transcriptome sequencing of virulent and attenuated Histomonas meleagridis reveals basic insights about biological functions and potential pathogenic mechanisms
Project description:Surface proteome of microbial pathogens is a crucial component of host-pathogen interactions. We applied cell surface biotinylation in association with high-throughput proteomic analysis to identify and quantify the surface proteome of a virulent and an attenuated strain, originating from the same single cell of H. meleagridis. In total 1485 proteins were identified amongst the two strains. Quantification revealed 22 and 45 up-regulated proteins in the virulent and in the attenuated strain, respectively. In the virulent strain proteins such as alpha-amylase, Clan CD family C13 asparaginyl endopeptidase-like cysteine peptidase, two Clan SC family S33, methylesterase-like serine peptidases, LysM peptidoglycan-binding domain-containing protein and surfactant B were up-regulated and could be linked to putative virulence factors involved in the colonization of the host and establishment of an infection. In the attenuated strain structural proteins such as actin-related proteins, fimbrin, coronin, were up-regulated, alongside with various transport and energy production proteins. Our results speak for the protozoan’s adaptation to the in vitro environment, with a clear tendency for up-regulation of mainly metabolic and structural proteins. We present a comprehensive profiling of H. meleagridis surface proteome. These results provide not only a better understanding of the surface molecules that may participate in establishing and maintaining infection but also highlights the pathogen’s in vitro adaptation processes.
Project description:Histomonas meleagridis can cause histomonosis in poultry. Due to the prohibition of effective drugs, the prevention and treatment of the disease requires new strategies. Questions about its pathogenic mechanisms and virulence factors remain puzzling. To address some of these issues, comparative proteomic analysis of TMT was performed on a virulent strain and its attenuated strain of Chinese chicken-origin . A total of 3494 proteins were identified in the experiment, of which 745 proteins were differentially expressed (fold change ≥1.2 or ≤0.8 and p<0.05 ), 192 were up-regulated in virulent strain, and 553 were up-regulated in attenuated strain. Gene ontology (GO) analysis revealed that the differentially expressed proteins were mainly involved in organic material metabolic processes, primary metabolic processes and cellular metabolic processes, exerting protein binding functions, and most were distributed in the cytoplasm and organelle. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these differentially expressed proteins were mainly involved in pathway of Spliceosome, Cytosolic DNA-sensing pathway, Ribosome biogenesis in eukaryotes, RNA transport and Ribosome. In addition, some differentially expressed proteins were quantified by parallel reverse monitoring (PRM) assay to verify the results of TMT analysis. The above results provide some candidate protein-coding genes for further functional verification, which will help to understand the molecular mechanism of pathogenicity and attenuation of H. meleagridis more comprehensively.