Mycoplasma synoviae is a significant cause of respiratory disease and synovitis among chickens, and has an adverse economic impact on broiler breeding efforts. The present study was designed to develop a systematic understanding of the role that M. synoviae lipid-associated membrane proteins (LAMPs) may play in the virulence of this pathogen. Bioinformatics tools were used to identify 146 predicted membrane proteins and lipoproteins in the M. synoviae proteome. Then, Triton X-114 was used to extract LAMPs that were subsequently identified via LC–MS/MS. This approach enabled the detection of potential LAMPs, and the top 200 most abundant proteins detected using this strategy were subject to further analysis. M. synoviae cells (100 MOI) were exposed to chicken fibroblasts (DF-1) and macrophages (HD-11) in a 1:1 mixed culture. Analysis of LAMP transcripts identified 72 up-regulated LAMP genes which were analyzed in depth by bioinformatics. GO analysis revealed these genes to be enriched in the nucleotide binding, sulfur amino acid transmembrane transporter activity, tRNA binding, rRNA modification, and transition metal ion transport pathways. Moreover, KEGG enrichment analysis suggested that these genes were enriched in the biosynthesis of secondary metabolites, carbon metabolism, glycolysis/gluconeogenesis, and nitrogen metabolism pathways.

Mycoplasma synoviae infection rates in chickens are increasing worldwide. Genomic studies have considerably improved our understanding of M. synoviae biology and virulence. However, approximately 20% of the predicted proteins have unknown functions. In particular, the M. synoviae ATCC 25204 genome has 663 encoding DNA sequences, among which 155 are considered encoding hypothetical proteins (HPs). Several of these genes may encode unknown virulence factors. This study aims to reannotate all 155 proteins in M. synoviae ATCC 25204 to predict new potential virulence factors using currently available databases and bioinformatics tools. Finally, 125 proteins were reannotated, including enzymes (39%), lipoproteins (10%), DNA-binding proteins (6%), phase-variable hemagglutinin (19%), and other protein types (26%). Among 155 proteins, 28 proteins associated with virulence were detected, five of which were reannotated. Furthermore, HP expression was compared before and after the M. synoviae infection of cells to identify potential virulence-related proteins. The expression of 14 HP genes was upregulated, including that of five virulence-related genes. Our study improved the functional annotation of M. synoviae ATCC 25204 from 76% to 95% and enabled the discovery of potential virulence factors in the genome. Moreover, 14 proteins that may be involved in M. synoviae infection were identified, providing candidate proteins and facilitating the exploration of the infection mechanism of M. synoviae.

Mycoplasma bovis is one of the most important pathogens in animal husbandry, and the current infection and morbidity rates are increasing year by year, causing great losses to the farming industry and seriously affecting animal welfare. In this study, we took tracheal tissues from calves infected with M. bovis to make pathological tissue sections for observation, and selected tracheal tissues for transcriptome sequencing to screen differentially expressed genes based on the threshold |log2FoldChange| > 1 and Padjust < 0.05 and functional enrichment, to explore in depth the potential mechanisms of bovine tracheal damage caused by bovine tracheitis. Experiments were conducted to observe the changes in tracheal tissues after M. bovis infection through pathological sections of the trachea of M. bovis-infected calves. From the transcriptome sequencing results, we mined the main differential genes and important metabolic pathways of M. bovis causing damage to the trachea of calves. It was found that the cricoid cartilage tissue of the trachea was congested and hemorrhagic after M. bovis infection in calves, and the pathological sections showed localized necrosis of epithelial cells, disorganization, high inflammatory cell infiltration in the interepithelial and lamina propria, and some epithelial cell detachment. Transcriptome sequencing identified 4199 DEGs, including 1378 up-regulated genes and 2821 down-regulated genes. KEGG enrichment analysis indicated that the differential genes were enriched to 59 significantly differing signaling pathways, and a number of important metabolic pathways related to tracheitis induced by M. bovis-infected calves were unearthed. The major ones included IL-17, the Toll-like receptor, JAK/STAT, the PI3K-Akt signaling pathway, etc. In this study, we found that M. bovis infection of calves caused inflammatory damage to the trachea, and transcriptome sequencing results also showed significant differences in the expression of key genes such as IL-6 inflammatory factor, CASP8, and APOA1.

Background: Mycoplasma synoviae infection in chickens is associated with development of synovitis and arthritis, leading to a decline in poultry production and significant economic repercussions. However, the mechanism of this infection remains imperfectly known. Results: Prokaryotic transcriptome sequencing was used to identify differentially expressed genes (DEGs) in M. synoviae when exposed to mixed cultures of chicken macrophages and chicken embryo fibroblast cells, as well as M. synoviae cultured in vitro . Of 416 DEGs, 262 are upregulated and 122 are downregulated. Ten DEGs were randomly selected and subjected to verification through quantitative reverse transcriptase PCR. The upregulated DEGs exhibited significant enrichment in various Gene Ontology (GO) terms, notably transmembrane transport, transporter activity, and membrane protein complex. Additionally, these DEGs were enriched in Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, including ABC transporters, quorum sensing, aminoacyl-tRNA biosynthesis, DNA replication, and the bacterial secretion system. The 5’ untranslated region, transcription starting point and noncoding small RNA (sRNA) were also revealed, thirty-one upregulated genes were predicted to interact with sRNA NZ_CP011096_predRNA9. Conclusion: These findings revealed the expression of M. synoviae transcripts when exposed, which beneficial for the treatment of M. synoviae’s target proteins study, will also help to enhance understanding of mechanisms underlying M. synoviae infection.

Mycoplasma synoviae (M. synoviae) is an avian pathogen responsible for respiratory disease and synovitis. The VlhA (variable lipoprotein hemagglutinin) family of surface adhesins plays a critical role in host cell attachment, yet the specific residues and structural determinants governing this interaction remain incompletely understood. In this study, we selected the C-terminal lipid-associated membrane hemagglutinin (LAM HA) domain within the VlhA family as the bait protein, based on its conserved C-terminal region. Yeast two-hybrid screening identified 18 LAM HA-interacting host proteins, and molecular docking pinpointed five residues (S83, R85, Y88, N124, K192) as the most frequent interaction hotspots. To assess their collective functional relevance, we constructed a combined deletion mutant lacking these five candidate residues. At the optimal pH of 6.0–6.5, wild-type LAM HA showed the highest titer (1:2), significantly exceeding that at pH 7.0–7.5 (no activity) and pH 5.0–5.5 (1:1) (p < 0.05), whereas the deletion mutant displayed a reduced titer (from 2 to 1; p = 0.05). Secondary structure analysis at the same pH revealed decreased α-helix content (from 7.90% to 7.57%), along with increased β-sheet (from 38.92% to 38.83%) and random coil (from 10.83% to 10.44%). Molecular dynamics simulations revealed that the deletion mutant exhibited elevated RMSF (from 2.83 ± 1.88 Å to 4.18 ± 2.65 Å) and Rg (from 40.48 ± 1.31 Å to 54.16 ± 4.47 Å) at pH 6.0–6.5, while RMSD showed a slight decrease (11.17 ± 1.33 Å vs. 12.14 ± 0.21 Å), collectively indicating compromised structural stability of the mutant. Collectively, these findings suggest that the LAM HA domain contributes to hemagglutination activity through pH-sensitive conformational stability and specific residue clusters—a property that may be relevant to M. synoviae colonization in the acidic microenvironments of the respiratory tract and synovial fluid during infection.