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Mycoplasma synoviae (MS) infection is a serious threat to poultry industry in China. Tilmicosin is a semisynthetic macrolide antibiotic used only in animals and has shown potential efficacy against MS, but there were no reported articles concerning the pharmacokinetics/pharmacodynamics (PK/PD) interactions of tilmicosin against MS in vitro and vivo. This study aimed to assess the antibacterial activity of tilmicosin against MS in vitro and in vivo using PK/PD model to provide maximal efficacy. The minimum inhibitory concentration (MIC) and killing rates of different drug concentrations were measured using the microdilution method in vitro. Then, tilmicosin was administered orally to the MS‐infected chickens at doses of 7.5 and 60 mg/kg, and the PK parameters of tilmicosin in joint dialysates were determined using high‐pressure liquid chromatography/tandem mass spectrometry (HPLC‐MS/MS) combined with the microdialysis technique. The antibacterial effect (△E) was calculated when the infected chickens were administered a single oral dose of tilmicosin at 4, 7.5, 15, 30, and 60 mg/kg b.w. The PK and PD data were fitted using the Sigmoid Emax model to evaluate the PK/PD interactions of tilmicosin against MS. The bactericidal activity of tilmicosin against MS was concentration dependent. Furthermore, the PK/PD index of AUC0–72h/MIC exhibited the most optimal fitting results (R2 = .98). The MS load decreased by 1, 2, and 3 Log10 CFU/mL, then AUC/MIC was determined as 13.99, 20.53, and 28.23 h, respectively, and the bactericidal effect can be achieved when the dose of MS‐infected chickens is at 31.64 mg/kg b.w. The findings of this study hold significant implications for optimizing the treatment regimen for MS infection. [ABSTRACT FROM AUTHOR]

Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) are the primary agents responsible for mycoplasma disease in poultry. MG has been identified as a significant cause of chronic respiratory disease in chickens, while MS has been linked to the development of tenosynovitis, joint swelling and other symptoms in chickens, leading to considerable economic losses for the poultry industry. Unfortunately, there is no specific drug for treatment and vaccination is the most important way to control the disease. There are some different types of vaccines, including live vaccines, inactivated vaccines, sub-unit vaccines and vector vaccines. This paper provides a comprehensive review of the development of vaccines for MG and MS. [ABSTRACT FROM AUTHOR]

[Objective] Mycoplasma gallisepticum（MG）and Mycoplasma synoviae（MS）are the two most harmful mycoplasmas to poultry. Early diagnosis of MG and MS was a key to prevent and control avian mycoplasma. Through combining recombinase polymerase amplification（RPA）technology with lateral flow dipstick（LFD）, a rapid detection method for MG and MS with convenient operation, rapid response and visual results was established. [Method] Specific primers and probes were designed based on MG mgc2 gene and MS vlhA gene, and a dual LFD-RPA rapid detection method was established by optimizing reaction time, reaction temperature and primer ratio, and its sensitivity, specificity and stability were evaluated. Meanwhile, 120 clinical samples were tested. [Result] The amplification was done by dual LFD-RPA detection method at 37℃ for 5-10 min, and the optimal primer ratio of RPA-MG-F2/R2 and RPA-MS-F1/R1 was 0.6 ∶ 1.4. The minimum detection limits of MG and MS were 3.75 copies/μL and 3.46 copies/μL, respectively. This method and the PCR method recommended by OIE were used to detect 120 samples, and the coincidence rate was 93.3%. [Conclusion] The amplification can be done by the dual LFDRPA detection method of MG and MS under constant temperature conditions. It has simple operation, rapid response, high sensitivity, strong specificity and good stability, and the results can be observed without using any instrument. It is suitable for the field rapid detection of MG and MS infection alone or mixed infection at the primary level. [ABSTRACT FROM AUTHOR]

Abstract Background The impact of chickens on maintaining the economy and livelihood of rural communities cannot be overemphasized. In recent years, mycoplasmosis has become one of the diseases that affect the success of South African chicken production. Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) are the most prevalent strains of Mycoplasma in South Africa. MG and MS are significant respiratory pathogens affecting the productivity of chickens. The present study aimed to molecularly detect using qPCR and characterize the presence of MG and MS using phylogenetic analysis. The phylogenetic analysis was utilized to clarify general evolutionary relationships between related taxa of different MG and MS observed in tracheal swabs from South African chicken breeds. Methods Forty-five tracheal swabs of the Lohmann Brown (n = 9), Rhode Island Red (n = 9), Ovambo (n = 9), Venda (n = 9), and Potchefstroom Koekoek (n = 9) breeds were collected from symptomatic chickens present in the commercial farm. To detect MG and MS, DNA was extracted from tracheal swabs and faecal samples, and qPCR was performed with a 16 s rRNA (310 bp) and vlhA (400 bp) gene fragment. Following the sequencing of all the amplicons, MG, and MS dendrograms showing the evolutionary relationships among the five South African chicken breeds and the GeneBank reference population were constructed. Results The qPCR revealed the presence of MG and MS in 22% (2/9) of the tracheal swab samples tested for MS only in Rhode Island Red breeds; 66.6% (6/9) and 33% (3/9) of the tested samples in Ovambo breeds; and 11.1% (1/9) and 44.4% (4/9) of the tested samples in Venda breeds. No MG or MS were detected in the Lohmann Brown or Potchefstroom Koekoek breed. Furthermore, qPCR revealed the presence of MG in pooled faecal samples from Lohmann Brown and Ovambo breeds. Eight different bacterial isolates were recognized from both samples. Four isolates were of the 16 s ribosomal ribonucleic acid (rRNA) gene (named PT/MG51/ck/00, PT/MG48/ck/00, PT/MG41/ck/00 and PT/MG71/ck/00) gene of Mycoplasma gallisepticum, and the other was Mycoplasma Synoviae variable lipoprotein hemagglutinin A (vlhA) gene (named PT/MSA22/ck/01, PT/MS41/ck/01, PT/MS74/ck/01 and PT/MS46/ck/01) which were available in GenBank. These isolates were successfully sequenced with 95–100% similarity to the isolates from the gene bank. Conclusion The study revealed the presence of both MG and MS in the chicken breeds sampled. Furthermore, the different breeds of chicken were found to be susceptible to infection under the intensive or commercial management system. Therefore, continuous surveillance is encouraged to prevent the spread and outbreak of MG and MS in the poultry industry in South Africa.

Background: The impact of chickens on maintaining the economy and livelihood of rural communities cannot be overemphasized. In recent years, mycoplasmosis has become one of the diseases that affect the success of South African chicken production. Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) are the most prevalent strains of Mycoplasma in South Africa. MG and MS are significant respiratory pathogens affecting the productivity of chickens. The present study aimed to molecularly detect using qPCR and characterize the presence of MG and MS using phylogenetic analysis. The phylogenetic analysis was utilized to clarify general evolutionary relationships between related taxa of different MG and MS observed in tracheal swabs from South African chicken breeds. Methods: Forty-five tracheal swabs of the Lohmann Brown (n = 9), Rhode Island Red (n = 9), Ovambo (n = 9), Venda (n = 9), and Potchefstroom Koekoek (n = 9) breeds were collected from symptomatic chickens present in the commercial farm. To detect MG and MS, DNA was extracted from tracheal swabs and faecal samples, and qPCR was performed with a 16 s rRNA (310 bp) and vlhA (400 bp) gene fragment. Following the sequencing of all the amplicons, MG, and MS dendrograms showing the evolutionary relationships among the five South African chicken breeds and the GeneBank reference population were constructed. Results: The qPCR revealed the presence of MG and MS in 22% (2/9) of the tracheal swab samples tested for MS only in Rhode Island Red breeds; 66.6% (6/9) and 33% (3/9) of the tested samples in Ovambo breeds; and 11.1% (1/9) and 44.4% (4/9) of the tested samples in Venda breeds. No MG or MS were detected in the Lohmann Brown or Potchefstroom Koekoek breed. Furthermore, qPCR revealed the presence of MG in pooled faecal samples from Lohmann Brown and Ovambo breeds. Eight different bacterial isolates were recognized from both samples. Four isolates were of the 16 s ribosomal ribonucleic acid (rRNA) gene (named PT/MG51/ck/00, PT/MG48/ck/00, PT/MG41/ck/00 and PT/MG71/ck/00) gene of Mycoplasma gallisepticum, and the other was Mycoplasma Synoviae variable lipoprotein hemagglutinin A (vlhA) gene (named PT/MSA22/ck/01, PT/MS41/ck/01, PT/MS74/ck/01 and PT/MS46/ck/01) which were available in GenBank. These isolates were successfully sequenced with 95–100% similarity to the isolates from the gene bank. Conclusion: The study revealed the presence of both MG and MS in the chicken breeds sampled. Furthermore, the different breeds of chicken were found to be susceptible to infection under the intensive or commercial management system. Therefore, continuous surveillance is encouraged to prevent the spread and outbreak of MG and MS in the poultry industry in South Africa. [ABSTRACT FROM AUTHOR]

The significance of Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) in the poultry industry underscores the critical importance of diagnosing avian mycoplasmosis within the field of veterinary medicine. The present study aimed to compare various diagnostic methods for detecting MG and MS in Egyptian Chicken Flocks. A total of 360 samples were collected from breeder, layer, and broiler chickens from four governorates in Egypt. Conventional isolation methods and polymerase chain reaction (PCR) were used for the direct detection of MG and MS, while serum plate agglutination test (SPA) and Enzyme-linked immunosorbent assay (ELISA) were used for detecting antibodies against MG and MS. The highest detection rate of MG was found in commercial layers, followed by breeders, and broilers. Regarding MS, the highest detection rate was found in breeders, followed by commercial layers, and broilers. By comparing the used diagnostic methods, MG and MS were determined by the SPA test (40% and 31.1% respectively), ELISA test (31.7% and 23.6%), PCR (16.7% and 11.7%), and by the conventional culture method (10.8% and 3.9%). It could be concluded that the serological methods and PCR gave better sensitivity than culture methods and can be used in the diagnosis of avian mycoplasmosis. [ABSTRACT FROM AUTHOR]

Beside biosecurity, vaccination is important for Mycoplasma synoviae (MS) control as it has been shown to contribute to the reduction of economic impact and, experimentally, also lessens horizontal transmission. In this study, the effect of MS live vaccination on horizontal transmission was quantified under field conditions by analysing 4-year MS monitoring data from non-MS-vaccinated broiler and layer breeders and MS-vaccinated broiler breeders with good biosecurity in single-age housing systems. Flocks were monitored at 20 and 30 weeks of age and every 12 weeks thereafter. At every sampling, 60 blood samples or 24 tracheal swabs were tested using rapid plate agglutination test and ELISA serially or MS DIVA PCR, respectively. The MS incidence rate was calculated and the association with vaccination was analysed by logistic regression. The average MS incidence rate per 1000 weeks was 11.6 cases for non-MS-vaccinated broiler breeders and decreased from 29.6 to 5.6 cases with successive vaccinated production cycles. In non-MS-vaccinated layer breeders it was 3.6. A significant negative association with MS incidence was found after vaccinating four to six successive production cycles compared to non-MS-vaccinated or only one production cycle vaccinated breeders (odds ratio (OR) = 0.23, P = 0.05 & OR = 0.12, P = 0.01, respectively). A significant negative association with MS in non-MS-vaccinated layer breeders (OR = 0.29, P = 0.00) was observed compared to non-MS-vaccinated broiler breeders, possibly due to more controlled contact structures within the layer breeder industry. The results suggest that vaccination and control of contacts contribute to the reduction of between-farm MS transmission. [ABSTRACT FROM AUTHOR]

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Toll-like receptors (TLRs) are one of the important immunes signaling pathways that participate in the activation of host immune response after detecting microbial pattern molecules. Despite important expression profile TLRs genes in chicken erythrocytes, studies have been lacking. This study investigated the expression profile of TLRs pathway immune gene in chicken erythrocytes in response to Mycoplasma synoviae. The purpose of the current in-vitro study was to determine the chicken erythrocytes interaction with M. synoviae using a transmission electron microscope (TEM). The mRNA gene expression of TLR1, 2, 3, 4, 5, 7, 15, MHC I, II, and MyD88 in M. synoviae infected chicken erythrocytes was determined using quantitative real-time PCR (qRT-PCR) at four different time intervals (0, 2, 6 and 10 h) post-infection and compared to uninfected controls. The mRNA expression of TLRs such as TLR1, 2, 3, 15, and MHC II were significantly upregulated at 6 and 10 h post-infection in infected chicken erythrocytes. However, significantly upregulated expression of TLR5 and MHC I were noted at 2, 6, and 10 h while TLR4 and MyD88 mRNA expression was also significantly upregulated but at different time intervals. This study provides the first evidence of upregulated expression of TLR signaling pathway genes in M. synoviae infected chicken erythrocytes. These results provide new insights on M. synoviae infection resistance mechanisms and the role of TLR signaling immune genes in the control of the host immune response. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae is one of the most important pathogens in the poultry industry and often causes diseases of a chronic and persistent nature. However, there is limited data available on Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway genes expression in chicken erythrocytes infected with Mycoplasma synoviae (M. synoviae). Therefore, the aim of the current in-vitro study was to determine the interaction between chicken erythrocytes and M. synoviae using Transmission electron microscope (TEM) and further to investigate the mRNA gene expression of MyD88 (Myeloid differentiation primary response 88), CCL5 (C-C Motif Chemokine Ligand 5), MDA5 (melanoma differentiation-associated protein 5) IKBKE (inhibitor of nuclear factor kappa-B kinase subunit epsilon), NFKBIA (NF-kappa-B inhibitor alpha), NFKBIE (NF-kappa-B inhibitor epsilon), Interferon Alpha (IFN-α), cMGF (chicken myelomonocytic growth factor), and TRAF6 (Tumor necrosis factor receptorassociated factor 6) in chicken erythrocytes infected with M. synoviae using quantitative real-time PCR (qRT-PCR) at four different time intervals (0, 2, 6 and 10 h) post-infection and compared to uninfected controls. The results indicated that M.synoviae interacted efficiently in chicken erythrocytes, which strongly induced the up-regulation of NF-κB pathway and other immune system genes in response to early bacterial infection such as NFKBIA, NFKBIE, IKBKE, CCL5, MDA5, MyD88, and TRAF6 at different h's interval. Whereas cMGF and IFN-α expression were significantly downregulated during early time intervals such as 0 h, 2 h and 6 h while later on the expression level significantly increased. These results will lead to increased insights on M. synoviae infection resistance mechanisms and the role of NF-κB signaling pathway and other immune system genes in the control of the host immune response. [ABSTRACT FROM AUTHOR]

ABSTRACT: Mycoplasma synoviae (MS) is an essential pathogenic mycoplasma in poultry worldwide, posing a serious threat to the poultry industry's health. Timely detection is imperative for early diagnosis, prevention, and control of MS infection. Current laboratory methods for MS detection are generally complicated, time-consuming, and require sophisticated equipment. Therefore, a simple and rapid method is urgently needed. This study developed a novel real-time fluorescence-based recombinase-aided amplification (RF-RAA) technique for detecting MS nucleic acids, enabling target gene amplification within 20 min at 39°C. The RF-RAA outcomes are interpretable in 2 modalities: real-time fluorescence monitoring employing a temperature-controlled fluorescence detector or direct visual inspection facilitated by a portable blue light transilluminator. This method exhibits robust specificity, demonstrating no cross-reactivity with various common poultry pathogens, and achieves high sensitivity, detecting as low as 10 copies/μL for the standard plasmid. Seventy-one clinical samples of chicken throat swabs were detected by RF-RAA and real-time fluorescence quantitative polymerase chain reaction (qPCR) methods. The diagnostic coincidence rates of qPCR with RF-RAA (fluorescence monitoring) and RF-RAA (visual observation) were determined to be 100% and 97.2% (69/71), respectively. In conclusion, the RF-RAA method developed in this study provides a rapid and visually observable approach for MS detection, offering a novel technique to diagnosing MS infection, especially in resource-limited settings.

ABSTRACT: Mycoplasma synoviae (MS) is a contagious pathogen that poses a significant threat to the poultry industry. Detection plays an important role in the prevention and control of MS, particularly in differentiating between wild-type MS and live attenuated vaccine strains for vaccination selection and culling of animals with wild-type only. The live attenuated ts+ vaccine strain MS-H is recognized as the most effective and widely used vaccine. In this study, we have developed a method called double enzyme-activated differentiation probes PCR (DEA-probes PCR) for the differentiation of MS-H vaccine strain from wild-type strain by targeting the single nucleotide polymorphism (SNP) of the 367th nucleotide in the Obg gene sequence. We developed 2 modified probes with the ribonucleotide insert. When the probe perfectly complements with the target, the ribonuclease H2 (RNase H2) will cleave the ribonucleotide, resulting in the generation of fluorescent signal. With a detection limit of 5.8 copies/µL, the DEA-probes PCR method demonstrates 100% specificity in distinguishing wild-type MS from MS-H strains in 1 h. The method demonstrated great performance in real application of 100 superior palate cleft swab samples from chickens in poultry farms. Twenty-eight samples were detected as MS positive, consistent with the results of the Chinese industry standard method. Additionally, our method was able to distinguish 19 wild-type MS strains from 9 MS-H vaccine strains. The DEA-probes PCR method is rapid, specific and sensitive for SNP detection, overcoming the misidentification in MS detection and differentiation. It can be also applied to the differentiation of infected from vaccinated animals (DIVA) for other pathogens.

Mycoplasma synoviae (MS) is an economically important pathogen in the poultry industry. Vaccination is an effective method to prevent and control MS infections. Currently two live attenuated MS vaccines are commercially available, the temperature-sensitive MS-H vaccine strain and the NAD-independent MS1 vaccine strain. Differentiation of vaccine strains from wild-type (WT) strains is crucial for monitoring MS infection, especially after vaccination. In this study, we developed a Taqman duplex real-time polymerase chain reaction (PCR) method to identify MS1 vaccine strains from WT strains. The method was specific and did not cross-react with other avian pathogens. The sensitivity assay indicated that no inhibition occurred between probes or between mixed and pure templates in duplex real-time PCR. Compared with the melt-based mismatch amplification mutation assay (MAMA), our method was more sensitive and rapid. In conclusion, the Taqman duplex real-time PCR method is a useful method for the diagnosis and differentiation of WT-MS and MS1 vaccine strains in a single reaction. [ABSTRACT FROM AUTHOR]

The MS-H vaccine, containing a live strain of Mycoplasma synoviae, is a feasible option for controlling M. synoviae infection in poultry flocks. A comprehensive understanding of vaccinated chickens, including strain differentiation and immune response mechanisms, is required to optimize vaccination strategy. This study aimed to verify the PCR-RFLP molecular assay as a convenient technique for detecting the MS-H vaccine strain and to characterize the immune response mechanisms in experimental layer-type chickens receiving one of three different vaccination programmes; a single dose at either 9 or 12 weeks of age or two doses at both 9 and 12 weeks of age. The PCR-RFLP assay, using restriction enzyme TasI to digest vlhA gene-targeted PCR amplicons, was performed to evaluate vaccine administration by detecting the MS-H vaccine strain in vaccinated chickens and differentiating it from non-vaccine strains such as WVU1853 reference strain and Thai M. synoviae field strains. Results demonstrated that vaccination in layer-type chickens, whether as one or two doses, stimulated immune response mechanisms with no significant advantages of two administrations over a single administration. Serological responses in vaccinated chickens, examined by RPA test and ELISA, were initially detected at 2 weeks post-vaccination, continuously increased, and then remained at the baseline levels from 6 to 9 weeks post-vaccination. Cellular immune responses against both homologous and heterologous antigens, examined by the MTS tetrazolium assay, were similar in the early period post-vaccination, whereas cellular immune response against the homologous MS-H antigen was improved in the late period post-vaccination. [ABSTRACT FROM AUTHOR]

1. Mycoplasma synoviae (MS) is the primary causative agent of synovitis in avian species. In order to investigate the pathogenicity and immunological responses associated with MS in specific pathogen-free chicken embryos, a series of generations (F1, F95, F120, F160 and F200) of MS were introduced into 7-day-old SPF chicken embryos and subsequent mortality rates were recorded and analysed 2. Reverse transcription-quantitative polymerase chain reaction was performed to detect expression of heat shock proteins HSP27, HSP40, HSP60, HSP70 and HSP90 and inflammatory factors interleukin (IL)-1β, caspase-1 and IL-18 in the tracheal tissue. 3. The results showed that the mortality rate of SPF chicken embryos decreased with an increase in the number of passages, with the highest being 80% (8/10) for F1 generation and the lowest being 10% (1/10) for F200. The expression of HSP27, IL-1β, HSP40, caspase-1, HSP70 and HSP90 showed a significant downregulation trend with an increase in the generation (except IL-18; P < 0.05). The HSP60 expression was significantly upregulated with increasing generations (P < 0.05). 4. A relationship between pathogenicity and the number of passages was observed and the decrease in pathogenicity appeared to be associated with HSP and genes related to inflammatory factors. The present work offers a scientific foundation for screening potential MS strains that might be employed to develop attenuated vaccines. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae (M. synoviae) is one of the major poultry pathogens causing infectious synovitis, airsacculitis, a high incidence of shell breakage, and egg production loss. However, the pathogenesis of M. synoviae remains unclear. Adhesion of mycoplasmas to host cells is a crucial step in infection and colonization. The purpose of this study was to determine the adhesive function of a putative P80 family lipoprotein (LP78) and evaluate its application in the detection of antibodies against M. synoviae. Recombinant LP78 (rLP78) was expressed in the supernatant component of Escherichia coli and mouse antirLP78 serum was prepared. Bioinformatic analysis and western blotting results revealed that LP78 was conservative among M. synoviae strains. It was distributed not only in the cytoplasm but also on the membrane of M. synoviae through western blotting and indirect immunofluorescence (IFA). The adherence of M. synoviae to DF-1 cells was significantly inhibited by mouse anti-rLP78 serum (p < 0.01). IFA revealed that rLP78 adhered to DF-1 cells, and this adherence was prevented by mouse anti-rLP78 serum. Furthermore, rLP78 was found to bind to the DF-1 cells membrane proteins in a dose-dependent manner by enzymelinked immunosorbent assay (ELISA). Screening of DF-1 cells membrane proteins by western blotting showed that proteins with molecular weight of 35-40 kDa and 55-70 kDa bound to rLP78. Moreover, rLP78 was identified to be a fibronectin/plasminogen binding protein. The sensitivity and specificity of rLP78-based iELISA were 85.7 and 94.1%, respectively. The maximum dilution of positive serum (HI titer, 1:128) detected via rLP78-based iELISA was 1:6,400, whereas that detected using a commercial ELISA kit was 1:12,800-1:25,600. Both rLP78-based iELISA and the commercial ELISA kit detected seroconversion after 7 days of challenge and immunization. No cross-reactivity with positive sera against other avian pathogens was observed in rLP78-based iELISA. Collectively, these results indicate that LP78 is a fibronectin/plasminogen-binding adhesion protein of M. synoviae and a potential diagnostic antigen. The present study will facilitate a better understanding of the pathogenesis of M. synoviae and the development of new diagnostic. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae (MS) is a primary avian pathogen prevalent worldwide that causes airsacculitis and synovitis in birds. Vaccination is recommended as the most cost-effective strategy in the control of MS infection. Novel alternative vaccines are needed for eradicating and controlling MS infection in flocks. DnaK, enolase, elongation factor Tu (EF-Tu), MSPB, NADH oxidase and LP78 are the major immunogenic antigens of MS and are promising targets for subunit vaccine candidates. In the present study, genes encoding DnaK, enolase, EF-Tu, MSPB, LP78, and NADH oxidase were cloned and expressed in Escherichia coli. Enzyme-linked immunosorbent assay showed that the six recombinant proteins were recognized by convalescent sera, indicating that they were expressed during infection. Two injections of the six subunit vaccines induced a robust antibody response and increased the concentrations of IFN-γ and IL-4, especially rEnolase and rEF-Tu. The proliferation of peripheral blood lymphocytes was enhanced in all of the immunized groups. Chickens immunized with rEnolase, rEF-Tu, rLP78, and rMSPB conferred significant protection against MS infection, as indicated by significantly lower DNA copies in the trachea, lower scores of air sac lesions, and lesser tracheal mucosal thickness than that in the challenge control. Especially, rEnolase provided the best protective efficacy, followed by rEFTu, rMSPB, and rLP78. Our finds demonstrate that the subunit vaccines and bacterin can only reduce the lesions caused by MS infection, but not prevent colonization of the organism. Our findings may contribute to the development of novel vaccine agents against MS infection. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae, which causes the disease known as chicken synovitis, causes serious immunosuppression. We developed a rapid insulated isothermal polymerase chain reaction (iiPCR) assay for on-site detection of M. synoviae using a primer and probe set targeting the variable lipoprotein and haemagglutinin (vlhA) gene. In addition, the specificity, sensitivity, repeatability, and clinical detection of this method were evaluated. Our iiPCR assay detected M. synoviae clinical isolates and samples successfully and produced negative results on Mycoplasma galliscepticum, avian viral arthritis, Escherichia coli, Salmonella, Staphylococcus aureus and Corynebacterium, indicating that the PCR reactions were specific. Additionally, our iiPCR assay detected the prepared positive standard plasmid diluted 10 times (1.00 × 10-1 - 1.00 × 10-10) as a template. The undiluted positive plasmid was positive and double distilled water was negative indicating that the PCR reactions were sensitive, respectively. Finally, the vlhA positive standard plasmid with dilution multiple of 1.00 × 10-4 - 1.00 × 10-6 was repeatedly detected three times to evaluate the repeatability of the iiPCR method established in this experiment showing that the iiPCR of M. synoviae is repeatable. The established iiPCR was also used to detect 50 chicken joint enlargement samples. The thermostatic detection PCR established in this experiment was comparable to a reference real-time PCR (qPCR). [ABSTRACT FROM AUTHOR]

The pathogenic mycoplasmas are among the bacteria causing significant losses in the poultry industry worldwide. Mycoplasma gallisepticum (MG) and M. synoviae (MS) are economically important pathogens causing chronic respiratory disease, decreased growth, egg production and hatchability rates, and significant downgrading of carcasses. Effective diagnosis of infection with these species in poultry is highly requisite considering their two routes of spreading—horizontal and vertical. Their prevalence and molecular epidemiology were investigated in 184 turkey flocks in Poland. Tracheal samples were selected from 144 broiler flocks and 40 turkey breeder flocks collected in 2015–2023. The prevalence of MG was determined by real-time PCR targeting the 16S rRNA gene and PCR targeting the mgc2 gene, and MS was determined by a 16–23S rRNA real-time PCR and a vlhA gene PCR. Further identification and molecular characterization were carried out using PCR and sequencing. M. gallisepticum and M. synoviae were found in 8.33% and 9.72% of turkey broiler flocks respectively. The phylogenetic analysis of MG isolates in most cases showed high similarity to the ts-11-like strains. MS isolates showed high similarity to strains isolated from flocks of laying hens causing EAA. Additional tests detected Ornithobacterium rhinotracheale, Gallibacterium anatis, Enterococcus faecalis and Enterococcus faecium, Staphylococcus aureus and Riemerella anatipestifer. These secondary pathogens could have significantly heightened the pathogenicity of the mycoplasma infections studied. [ABSTRACT FROM AUTHOR]

Background: Mycoplasma synoviae is an important poultry pathogen causing heavy economic losses Worldwide. Subclinical persistence of this pathogen is the major issue to control its prevalence. Aim: This study aimed to determine the molecular and cross immunogenicity of Mycoplasma synoviae among broiler in five Districts of Khyber Pakhtunkhwa. Methods: This study was conducted by collecting 434 specimen samples from 40 broiler farms and desi poultry in five districts of Khyber Pakhtunkhwa. Specimen samples from the broiler birds (n=150), broiler farm environment (n= 264) and desi poultry birds (n=20) were aseptically collected and serially passaged in Modified Frey's broth. The homologous and heterologous antibody reactions were studied in rabbits. Prior to inoculation into rabbits the MS isolates were inactivated by formalin and adjuvanted with Montanide. Results: The overall turbidity prevalence in Frey's broth was observed as 109/434 (25.11%) samples and these turbidity positive samples were shifted on Frey's agar. After the appearance of classic fried egg colonies, the Biochemical confirmation was supported by production of Catalase and Phosphatase, reduction of Tetrazolium, Film and spot Assay and fermentation of Glucose for specie differentiation in avian mycoplasma. The MS prevalence percentage was recorded as 2% (9/434) through biochemical tests. The PCR results showed 0.5% MS prevalence with two field isolates (named as MS-1 and MS-2). Both MS-1 and MS-2 field isolates showed similar value (42.2) of homologous GMT titers. While the heterologous GMT for MS-1 serum against MS-2 isolate was lower (27.9) as compared to MS-2 serum against MS1 isolate (38.9). No titer was detected in control group (Group-III). Conclusion: In conclusion the results indicated the existence of MS in broiler birds and high homologous titers recorded between fields isolates which is a perpetual menace to poultry. [Open Vet J 2023; 13(10.000): 1299-1307]

Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) are the primary agents responsible for mycoplasma disease in poultry. MG has been identified as a significant cause of chronic respiratory disease in chickens, while MS has been linked to the development of tenosynovitis, joint swelling and other symptoms in chickens, leading to considerable economic losses for the poultry industry. Unfortunately, there is no specific drug for treatment and vaccination is the most important way to control the disease. There are some different types of vaccines, including live vaccines, inactivated vaccines, sub-unit vaccines and vector vaccines. This paper provides a comprehensive review of the development of vaccines for MG and MS.

Background: Mycoplasma synoviae (MS) is an important poultry pathogen causing heavy economic losses Worldwide. Subclinical persistence of this pathogen is the major issue to control its prevalence. Aim: This study aimed to determine the molecular and cross-immunogenicity of MS among broilers in five Districts of Khyber Pakhtunkhwa (KP). Methods: This study was conducted by collecting 434 specimen samples from 40 broiler farms and desi poultry in five districts of KP. Specimen samples from the broiler birds (n = 150), broiler farm environment (n = 264), and desi poultry birds (n = 20) were aseptically collected and serially passaged in Modified Frey's broth. The homologous and heterologous antibody reactions were studied in rabbits. Before inoculation into rabbits, the MS isolates were inactivated by formalin and adjuvanted with Montanide. Results: The overall turbidity prevalence in Frey's broth was observed as 109/434 (25.11%) samples, and these turbiditypositive samples were shifted on Frey's agar. After the appearance of classic fried egg colonies, the Biochemical confirmation was supported by the production of catalase and phosphatase, reduction of tetrazolium, film and spot assay, and fermentation of glucose for species differentiation in avian mycoplasma. The MS prevalence percentage was recorded as 2% (9/434) through biochemical tests. The PCR results showed 0.5% MS prevalence with two field isolates (named MS-1 and MS-2). Both MS-1 and MS-2 field isolates showed similar values (42.2) of homologous geometric mean titer (GMT). While the heterologous GMT for MS-1 serum against MS-2 isolate was lower (27.9) as compared to MS-2 serum against MS1 isolate (38.9). No titer was detected in the control group (Group-III). Conclusion: In conclusion, the results indicated the existence of MS in broiler birds and high homologous titers recorded between field isolates, which is a perpetual menace to poultry. [ABSTRACT FROM AUTHOR]

Featured Application: The proposed AI-supported classification method, together with the described portable multispectral fibre-optics reflectometer, is recommended for use as a fast-warning detection tool against eggshell changes caused by Mycoplasma synoviae in flocks of birds. Other application areas are eggs wholesalers and distributors, veterinarians, sanitary stations, border services, etc. Rapid detection of Mycoplasma synoviae (MS) in a flock is crucial from the perspective of animals' health and economic income. MS are highly contagious bacteria that can cause significant losses in commercial poultry populations when its prevalence is not limited. MS infections can cause losses associated with a range of clinical symptoms related to the respiratory, mobility and reproductive systems. Lesions related to the reproductive system and changes in the eggshell result in losses associated with infection or embryo death or complete destruction of the eggs. The authors propose using spectral measurements backed up by an AI data processing algorithm to classify eggs' origin: from healthy hens or MS-infected ones. The newest obtained classification factors are F-scores for white eggshells of 99% and scores for brown eggshells of 99%—all data used for classification were obtained using a portable multispectral fibre-optics reflectometer. The proposed method may be used directly on the farm by staff members with limited qualifications, as well as by veterinary doctors, assistants, or customs officers. Moreover, this method is scalable to mass production of eggs. Standard methods such as serological tests require either specialized staff or laboratory equipment. Implementation of a non-destructive optical measurement method, which is easily adapted for use on a production line, is economically reasonable. [ABSTRACT FROM AUTHOR]

Mycoplasma gallisepticum (MG) and M. synoviae (MS) can infect many bird species. Susceptible species of birds are responsible for spillover infections from poultry to wild birds. In Malaysia, previous studies reported the detection of MG in free-flying birds, but there is a lack of information on the characterization and antimicrobial susceptibility profile of these isolates. Therefore, this study aims to molecular characterize and assess the minimum inhibitory concentration (MIC) of MG and MS isolated from different bird species, including aviary, free-flying, and pet birds. Altogether 54 choanal slit swab samples were collected. All the swab samples were subjected to isolation and polymerase chain reaction (PCR) detection. Positive samples were subjected to molecular characterization and MIC assay. Molecular characterization was done using targeted gene sequencing, and microdilution MIC assay was employed to determine the antimicrobial susceptibility profile of the isolates. Using both culture and PCR techniques, 5.6% (3/54) of the samples were positive for MG, while MS was detected in 29.6% (16/54) and 40.7% (22/54) of samples using culture and PCR, respectively. The partial sequence of the mgc2 gene showed a distinct pattern of this gene in one of the MG isolates. Phylogenetic analysis of this isolate indicated that this isolate is a progeny of a Thailand poultry strain. Microdilution MIC assay indicated 100% resistance to erythromycin and chlortetracycline. The findings of this study indicate the possibility of avian interspecies transmission of MG and MS in Malaysia and the need for antimicrobial treatment dosage optimization. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

ABSTRACT: Mycoplasma synoviae (MS) is an important pathogen in laying hens and causes serious economic losses in poultry production. Rapid, accurate and specific detection is important for the prevention and control of MS. Argonaute from Pyrococcus furiosus (PfAgo) is emerging as a nucleic acid detector that works via “dual-step” sequence-specific cleavage. In this study, an MS detection method combining recombinase polymerase amplification (RPA) and PfAgo was established. Through elaborate design and screening of RPA primers and PfAgo gDNA and condition optimization, amplification and detection procedures can be completed within 40 min, whereas the results were superficially interpreted under UV and blue light. The sensitivity for MS detection was 2 copies/µL, and the specificity results showed no cross reaction with other pathogens. For the detection of 31 clinical samples, the results of this method and qPCR were completely consistent. This method provides a reliable and convenient method for the on-site detection of MS that is easy to operate without complex instruments and equipment.

Abstract Background Multi‐causal respiratory infections are more commonly observed than uncomplicated cases with single agents in the commercial poultry industry. Recently, increased mortality rates associated with respiratory clinical signs have been reported in Iranian broiler farms. Objectives The present study aimed to determine the spectra of avian mycoplasmas (Mycoplasma gallisepticum, MG and Mycoplasma synoviae, MS) and Ornithobacterium rhinotracheale (ORT) in the broiler farms with the multi‐causal respiratory disease (MCRD) from 2017 to 2020. Methods Trachea and lung tissue samples were collected from 70 broiler flocks presenting increased mortality and acute respiratory disease. MG, MS, and ORT were detected by performing polymerase chain reaction with primers complementary to the 16S rRNA, vlhA, and 16S rRNA genes, respectively. Results Genetic materials of MG, MS, and ORT were detected in five, three, and five of the 70 flocks. Based on the phylogenetic analysis of the complete mgc2 coding sequences, all MG strains formed a distinct cluster along with other Iranian MG isolates. According to the phylogenetic analysis of the partial vlhA gene of MS strains, two isolates were located along with Australian and European strains. In addition, one of them displayed an out‐group association with MS isolates from Jordan. Phylogenetic analysis of Iranian ORT strains using a partial sequence of the 16S rRNA gene showed a distinct group among the other ORT strains. Conclusions The results indicate that MG, MS, and ORT are not predominantly responsible for the MCRD. However, continuous monitoring of poultry flocks could be significant for obtaining valuable information related to different MG, MS, and ORT strains and designing effective control strategies.

The unreasonable use of antibiotics is one of the important causes of antimicrobial resistance (AMR) that poses a huge public health threat. Magnolol is a traditional Chinese medicine exhibiting antibacterial-, antifungal-, anti-inflammatory-, and antioxidant activities. However, it is unclear whether magnolol has an inhibitory effect on mycoplasma. This study found that magnolol showed excellent inhibitory activity against various mycoplasmas. Magnolol showed dose-dependent inhibition of Mycoplasma synoviae growth and biofilm formation in vitro. Magnolol caused severely sunken and wrinkled M. synoviae cell membranes at the minimum inhibitory concentration, and an enlarged cell diameter. The chicken embryo infection model showed that magnolol significantly reduced M. synoviae pathogenicity in vivo. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the citrate cycle, glycolysis/gluconeogenesis, and pyruvate metabolism were significantly disturbed at the minimum inhibitory concentration of magnolol. Interestingly, 41% of differential metabolites were in the categories of lipids and lipid-like molecules. Protegenin A was up-regulated 58752-fold after magnolol treatment. It belongs to fatty acyls, and destroys cell membrane integrity and cell activity. Ghosphatidylethanolamine, phosphatidylglycerol, phosphatidic acid, and phosphatidylserine related to membrane maintenance and stress response were widely down-regulated. Collectively, our results illustrate the feasibility of magnolol as a phytochemical compound to treat mycoplasma infection.

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.

Background: Multi‐causal respiratory infections are more commonly observed than uncomplicated cases with single agents in the commercial poultry industry. Recently, increased mortality rates associated with respiratory clinical signs have been reported in Iranian broiler farms. Objectives: The present study aimed to determine the spectra of avian mycoplasmas (Mycoplasma gallisepticum, MG and Mycoplasma synoviae, MS) and Ornithobacterium rhinotracheale (ORT) in the broiler farms with the multi‐causal respiratory disease (MCRD) from 2017 to 2020. Methods: Trachea and lung tissue samples were collected from 70 broiler flocks presenting increased mortality and acute respiratory disease. MG, MS, and ORT were detected by performing polymerase chain reaction with primers complementary to the 16S rRNA, vlhA, and 16S rRNA genes, respectively. Results: Genetic materials of MG, MS, and ORT were detected in five, three, and five of the 70 flocks. Based on the phylogenetic analysis of the complete mgc2 coding sequences, all MG strains formed a distinct cluster along with other Iranian MG isolates. According to the phylogenetic analysis of the partial vlhA gene of MS strains, two isolates were located along with Australian and European strains. In addition, one of them displayed an out‐group association with MS isolates from Jordan. Phylogenetic analysis of Iranian ORT strains using a partial sequence of the 16S rRNA gene showed a distinct group among the other ORT strains. Conclusions: The results indicate that MG, MS, and ORT are not predominantly responsible for the MCRD. However, continuous monitoring of poultry flocks could be significant for obtaining valuable information related to different MG, MS, and ORT strains and designing effective control strategies. [ABSTRACT FROM AUTHOR]

The pathogenic mycoplasmas are among the bacteria causing significant losses in the poultry industry worldwide. Mycoplasma gallisepticum (MG) and M. synoviae (MS) are economically important pathogens causing chronic respiratory disease, decreased growth, egg production and hatchability rates, and significant downgrading of carcasses. Effective diagnosis of infection with these species in poultry is highly requisite considering their two routes of spreading—horizontal and vertical. Their prevalence and molecular epidemiology were investigated in 184 turkey flocks in Poland. Tracheal samples were selected from 144 broiler flocks and 40 turkey breeder flocks collected in 2015–2023. The prevalence of MG was determined by real-time PCR targeting the 16S rRNA gene and PCR targeting the mgc2 gene, and MS was determined by a 16–23S rRNA real-time PCR and a vlhA gene PCR. Further identification and molecular characterization were carried out using PCR and sequencing. M. gallisepticum and M. synoviae were found in 8.33% and 9.72% of turkey broiler flocks respectively. The phylogenetic analysis of MG isolates in most cases showed high similarity to the ts-11-like strains. MS isolates showed high similarity to strains isolated from flocks of laying hens causing EAA. Additional tests detected Ornithobacterium rhinotracheale, Gallibacterium anatis, Enterococcus faecalis and Enterococcus faecium, Staphylococcus aureus and Riemerella anatipestifer. These secondary pathogens could have significantly heightened the pathogenicity of the mycoplasma infections studied.

Abstract Background Mycoplasma synoviae (MS) is an important pathogen causing respiratory diseases and arthritis in chickens and turkeys, thus, resulting in serious economic losses to the poultry industry. Membrane-associated proteins are thought to play important roles in cytoadherence and pathogenesis. NADH oxidase (NOX) is an oxidoreductase involved in glycolysis, which is thought to be a multifunctional protein and potential virulence factor in some pathogens. However, little is known regarding the NOX of MS (MSNOX). We previously demonstrated that MSNOX was a metabolic enzyme distributed in not only the cytoplasm but also the MS membrane. This study was aimed at exploring NOX’s potential as a diagnostic antigen and its role in MS cytoadherence. Results Western blots and ELISAs indicated that recombinant MSNOX (rMSNOX) protein reacted with sera positive for various MS isolates, but not MG isolates or other avian pathogens, thus, suggesting that rMSNOX is a potential diagnostic antigen. In addition, rabbit anti-rMSNOX serum showed substantial complement-dependent mycoplasmacidal activity toward various MS isolates and MG Rlow. MSNOX protein was found not only in the cytoplasm but also on the membrane of MS through suspension immunofluorescence and immunogold electron microscopy assays. Indirect immunofluorescence assays indicated that rMSNOX adhered to DF-1 cells, and this adherence was inhibited by rabbit anti-rMSNOX, but not anti-MG serum. Furthermore, indirect immunofluorescence and colony counting assays confirmed that the rabbit anti-rMSNOX serum inhibited the adherence of various MS isolates but not MG Rlow to DF-1 cells. Moreover, plasminogen (Plg)- and fibronectin (Fn)-binding assays demonstrated that rMSNOX bound Plg and Fn in a dose-dependent manner, thereby further confirming that MSNOX may be a putative adhesin. Conclusion MSNOX was identified to be a surface immunogenic protein that has good immunoreactivity and specificity in Western blot and ELISA, and therefore, may be used as a potential diagnostic antigen in the future. In addition, rMSNOX adhered to DF-1 cells, an effect inhibited by rabbit anti-rMSNOX, but not anti-MG serum, and anti-rMSNOX serum inhibited the adherence of various MS isolates, but not MG Rlow, to DF-1 cells, thus indicating that the inhibition of adherence by anti-MSNOX serum was MS specific. Moreover, rMSNOX adhered to extracellular matrix proteins including Plg and Fn, thus suggesting that NOX may play important roles in MS cytoadherence and pathogenesis. Besides, rabbit anti-rMSNOX serum presented complement-dependent mycoplasmacidal activity toward both MS and MG, indicating the MSNOX may be further studied as a potential protective vaccine candidate.

Abstract Mycoplasma synoviae (M. synoviae) infections have become an increasingly serious concern in China because they cause huge economic losses to the poultry industry. Antibiotic treatment is one of control strategies that can be used to contain clinical outbreaks in M. synoviae-free flocks, especially because the bacteria can be transmitted through eggs. To understand M. synoviae infection status in farms of central China and the antibiotic susceptibility of the circulating strains in vivo and in vitro, 485 samples were collected from five provinces from 2019 to 2021. Fifty-two strains were isolated and identified. Determination of the minimum inhibitory concentration (MIC) of eight antibiotics (tylvalosin, tiamulin, tilmicosin, lincomycin, enrofloxacin, chlortetracycline, doxycycline and tylosin) for isolates showed that tylvalosin, doxycycline and tiamulin were effective against 52 clinical isolates (MIC values ≤ 0.0625–0.25 μg/mL, ≤0.0625–1 μg/mL, and 0.25–2 μg/mL, respectively). Tilmicosin, enrofloxacin and lincomycin had high MIC90 values (>32 μg/mL). An artificial M. synoviae infection model was established in chickens for evaluation of the short-term therapeutic effect of these antibiotics. After 5 days of medication, doxycycline (200 mg/L) showed a superior ability to inhibit M. synoviae compared with other groups, as did tylvalosin (200 mg/L). Furthermore, the therapeutic efficacy of tylvalosin (0.4 μg/mL) on intra-embryo-injected M. synoviae was higher than that of tiamulin at the same dose. A combination of MIC values determined in vitro and therapeutic effects observed in vivo revealed that tylvalosin and doxycycline had the best therapeutic effects. Tylvalosin also showed better inhibitory effects on the vertical transmission of M. synoviae than tiamulin.

Background and Aim: The Indian and global poultry industries suffer significant economic losses due to Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) infections, which adversely affect egg production, hatchability, weight gain, and feed efficiency in farms, thus decreasing the overall production efficiency. This study aimed to determine the percent positivity and phylogenetic analysis of MG, MS, and co-infection of both mycoplasmas in commercial poultry farms across different states of India from 2017 to 2021. Materials and Methods: A total of 3620 tracheal or choacal swabs were collected from breeder and layer farms showing clinical signs of avian mycoplasma infections from commercial poultry farms across India, and the percent positivites for MG, MS, and co-infection of both mycoplasmas were determined by Polymerase chain reaction using the 16S rRNA and vlhA genes amplification, respectively. Phylogenetic analysis was carried out by sequencing the mgc2 and vlhA genes of 2 samples of MG and 24 samples of M. synoviae to gain insight into the genetic variability of Indian strains. The data were then compared with other Indian strains, vaccines strains, and strains from other countries. Results: Our data shows the percent positivity of MG, MS, and co-infection of both MG and MS was 6.43%, 23.61%, and 15.49%, respectively. The phylogenetic relationship between MG and MS was determined using the vlhA and mgc2 genes, revealing two samples of MG and 24 samples of MS clustered with other Indian strains. M. synoviae MSM22 and previously studied M. synoviae MGS 482 clustered with vaccine strain M. synoviae MS-H. Conclusion: Mycoplasma synoviae infections in breeder, layer, and in both is predominant compared to MG across the states investigated in India. Sequenced samples of MG and MS showed evolutionary relationships with the previously studied Indian strains of MG and MS. These findings support our view that monitoring chickens for avian mycoplasma infections are of paramount significance. It further lends credence to the contention that such information will pave the way for the development of a home-grown vaccination control program and thus safeguard the poultry sector against mycoplasma infections.

ABSTRACT: Mycoplasma synoviae (M. synoviae) is an important pathogen in poultry industry and has led to major economic losses. Understanding the epidemiology is crucial to improve control and eradication program of M. synoviae. In this study, 487 samples suspected with M. synoviae infection were collected from August 2020 to June 2021 in China. Among 487 samples, 324 samples were MS positive, the positive rate was 66.53%, and 104 strains were isolated from 324 positive samples. The multilocus sequence typing (MLST) method based on seven housekeeping genes was used to conduct genotyping 104 M. synoviae strains isolated, and the 104 isolates belonged to 8 sequence types (STs) after MLST genotyping, and ST-34 had the highest proportion. After BURST analysis, all 104 isolates were divided into group 12 with other 56 strains isolated from China. Phylogenetic tree constructed by neighbor-joining method showed that nearly all of Chinese isolates (160 isolates) clustered together and separated from other reference isolates (217 isolates) in the PubMLST database. In conclusion, this study suggested that the M. synoviae strains in China were highly similar and independent of abroad strains.

ABSTRACT: Mycoplasma synoviae (MS) infection is a serious threat to poultry industry in China, thus it is essential to study the pharmacokinetics (PK) in the target site of MS-infected chickens, but there are no relevant reports at present. The aim of this study was to compare the PK of tilmicosin in plasma and joint dialysate in MS-infected chickens. The MS infection model was established by evaluating the influence factors of the susceptibility of chicken species, day age of chicken, infection routes, infection cycle, infection dose, and stress response. The clinical symptoms, pathogen isolation, PCR identification, and ELISA antibody were detected to determine whether the MS infection model has been successfully established. Eight-week-old Mahuang chickens were challenged with MS by joint combined with footpad, 2 mL each time, twice a day for 5 d, then the MS infection model was successfully established. The infection group was orally administrated a single dose of 15 mg/kgbody weight (b. w.) tilmicosin. The joint dialysate was collected by the microdialysis technique, then the concentration of tilmicosin in plasma samples and joint dialysate were determined by triple quadrupole high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). There was no significant difference in elimination half-life (t1/2) and the mean residence time (MRT) of dialysate and plasma. In contrast, the time of the area under the concentration-time curve (AUC) and the (maximum concentration of tilmicosin in plasma) Cmax of tilmicosin in plasma was 2.1 and 1.4 times higher than in dialysate. The distribution coefficient of tilmicosin in joint and plasma (AUCdialysate/AUCplasma) was 0.51. In conclusion, tilmicosin concentration in joints of MS-infected chicken was much lower than that of plasma, which may result in the poor clinical effect and drug resistance. The study could provide a reference for the clinical use of tilmicosin against MS.

Rapid detection of Mycoplasma synoviae (MS) in a flock is crucial from the perspective of animals’ health and economic income. MS are highly contagious bacteria that can cause significant losses in commercial poultry populations when its prevalence is not limited. MS infections can cause losses associated with a range of clinical symptoms related to the respiratory, mobility and reproductive systems. Lesions related to the reproductive system and changes in the eggshell result in losses associated with infection or embryo death or complete destruction of the eggs. The authors propose using spectral measurements backed up by an AI data processing algorithm to classify eggs’ origin: from healthy hens or MS-infected ones. The newest obtained classification factors are F-scores for white eggshells of 99% and scores for brown eggshells of 99%—all data used for classification were obtained using a portable multispectral fibre-optics reflectometer. The proposed method may be used directly on the farm by staff members with limited qualifications, as well as by veterinary doctors, assistants, or customs officers. Moreover, this method is scalable to mass production of eggs. Standard methods such as serological tests require either specialized staff or laboratory equipment. Implementation of a non-destructive optical measurement method, which is easily adapted for use on a production line, is economically reasonable.

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.

The growth properties of the nutrient medium for the cultivation of pathogenic mycoplasmas depend on the type of blood serum it is supplemented with. Comparative tests of two cell-free nutrient media supplemented with bovine and porcine blood sera for the cultivation of strains “S6” Mycoplasma gallisepticum and “WVU 1853” Mycoplasma synoviae were performed. Growth properties of the tested nutrient media were assessed by determining the activity of the resulting biomass in the hemagglutination and agglutination assays, as well as by determining the concentration of viable cells after the 9th passage. It has been shown that a cell-free nutrient medium supplemented with the porcine blood serum is optimal for the cultivation of pathogenic mycoplasma species causing infectious diseases in birds. The hemagglutinating activity of the Mycoplasma gallisepticum culture reached 5 HAU log2 after 72 hours of cultivation, the agglutinating activity of Mycoplasma synoviae reached 5 AU log2 during the 88-hour incubation period, the concentration of viable cells of both strains was 106 CFU/cm3. The low growth properties of the medium prepared with the addition of bovine blood serum are most likely associated with its biochemical composition, which contains 5–20 times more provitamin A than the porcine blood serum, and high density lipoprotein cholesterol. On the contrary, in the porcine blood serum, most of the lipoproteins have a low density, containing a large amount of fatty acids and cholesterol, which are the main structural elements of mycoplasma cells. The obtained test results are of practical value and can be used in the technology of cultivation of pathogenic species of avian mycoplasmas in the production of diagnostic and preventive tools.

Background: Mycoplasma synoviae (MS) is an important pathogen causing respiratory diseases and arthritis in chickens and turkeys, thus, resulting in serious economic losses to the poultry industry. Membrane-associated proteins are thought to play important roles in cytoadherence and pathogenesis. NADH oxidase (NOX) is an oxidoreductase involved in glycolysis, which is thought to be a multifunctional protein and potential virulence factor in some pathogens. However, little is known regarding the NOX of MS (MSNOX). We previously demonstrated that MSNOX was a metabolic enzyme distributed in not only the cytoplasm but also the MS membrane. This study was aimed at exploring NOX's potential as a diagnostic antigen and its role in MS cytoadherence. Results: Western blots and ELISAs indicated that recombinant MSNOX (rMSNOX) protein reacted with sera positive for various MS isolates, but not MG isolates or other avian pathogens, thus, suggesting that rMSNOX is a potential diagnostic antigen. In addition, rabbit anti-rMSNOX serum showed substantial complement-dependent mycoplasmacidal activity toward various MS isolates and MG Rlow. MSNOX protein was found not only in the cytoplasm but also on the membrane of MS through suspension immunofluorescence and immunogold electron microscopy assays. Indirect immunofluorescence assays indicated that rMSNOX adhered to DF-1 cells, and this adherence was inhibited by rabbit anti-rMSNOX, but not anti-MG serum. Furthermore, indirect immunofluorescence and colony counting assays confirmed that the rabbit anti-rMSNOX serum inhibited the adherence of various MS isolates but not MG Rlow to DF-1 cells. Moreover, plasminogen (Plg)- and fibronectin (Fn)-binding assays demonstrated that rMSNOX bound Plg and Fn in a dose-dependent manner, thereby further confirming that MSNOX may be a putative adhesin. Conclusion: MSNOX was identified to be a surface immunogenic protein that has good immunoreactivity and specificity in Western blot and ELISA, and therefore, may be used as a potential diagnostic antigen in the future. In addition, rMSNOX adhered to DF-1 cells, an effect inhibited by rabbit anti-rMSNOX, but not anti-MG serum, and anti-rMSNOX serum inhibited the adherence of various MS isolates, but not MG Rlow, to DF-1 cells, thus indicating that the inhibition of adherence by anti-MSNOX serum was MS specific. Moreover, rMSNOX adhered to extracellular matrix proteins including Plg and Fn, thus suggesting that NOX may play important roles in MS cytoadherence and pathogenesis. Besides, rabbit anti-rMSNOX serum presented complement-dependent mycoplasmacidal activity toward both MS and MG, indicating the MSNOX may be further studied as a potential protective vaccine candidate. [ABSTRACT FROM AUTHOR]

Avian mycoplasmosis mainly caused by Mycoplasma gallisepticum and M. synoviae is an economically important disease of poultry industry. It causes huge economic losses in terms of decrease in weight gain, feed conversion efficiency, egg production, hatchability; increase in embryo mortality, carcass condemnation, prophylaxis and treatment cost in broiler, layer and breeder flocks. The disease is caused by four major pathogenic mycoplasmas viz., M. gallisepticum (MG), M. synoviae (MS), M. meleagradis (MM) and M. iowae (MI). The MG and MS are World Organization for Animal Health listed respiratory pathogens. MG causes chronic respiratory disease in chicken and infectious sinusitis in turkey; however, MS causes synovitis and airsacculitis in birds. The infection is transmitted both horizontally and vertically. Prevention and control measures of avian mycoplasmosis mainly comprises of biosecurity, treatment and vaccination. For vaccination of birds, inactivated bacterins, live attenuated and/or recombinant live poxvirus vaccines are commercially available against MG and MS infection. The present systematic review summarizes the different epidemiological studies carried out on MG and MS infection in poultry in different geographical locations of India and abroad over the last decade (2010–2020), economic impact, diagnosis and prevention and control. [ABSTRACT FROM AUTHOR]

The proper classification of the origins of food products is a crucial issue all over the world nowadays. In this paper, the authors present a device—a multispectral portable fibre-optic reflectometer and signal processing patch—together with a machine-learning algorithm for the classification of the origins of chicken eggshells in the case of Mycoplasma synoviae infection. The sensor device was developed based on previous studies with a continuous spectrum in transmittance and selected spectral lines in reflectance. In the described case, the sensor is based on the integration of reflected spectral data from short spectral bands from the VIS and NIR region, which are produced by single-colour LEDs and introduced to the sample via a fibre bundle. The measurement is carried out in a sequence, and the reflected signal is pre-processed to be put in the machine learning algorithm. The support vector machine algorithm is used together with three different types of data normalization. The obtained results of the F-score factor for classification of the origins of samples show that the percentages of eggs coming from Mycoplasma synoviae infected hens are up to 87% for white and 96% for brown eggshells. [ABSTRACT FROM AUTHOR]

Objective: Mycoplasma synoviae (M. synoviae) is the second most clinically important mycoplasma species, causing significant economic losses to the poultry industry worldwide. The aim of this study is to investigate the presence of antibodies against M. synoviae by indirect ELISA in commercial broiler and commercial layer flocks. Methods: A total of 957 blood samples were collected from vena subcutanae ulnaris under aseptic conditions by random sampling from unvaccinated commercial broiler (n=380) and commercial layer (n=577) flocks in the Central Anatolia Region and their serums were extracted. Anti-M.synoviae antibodies were detected by using commercial indirect ELISA kit. Results: The seropositivity rates were determined 40.76% (n=155) and 54.59% (n=315) in 380 sera from 21 commercial broiler flocks and 577 sera from 33 commercial layer flocks, respectively. Conclusion: In comparison to broilers, M. synoviae infection was shown to be more widespread in poultry breeding, particularly in commercial layer flocks, in the Central Anatolia Region. To avoid economic loss due to M. synoviae in both breeding types, we conclude that it is critical to improve the biosecurity of poultry breeder flocks, conduct regular follow-ups with monitoring studies, especially in breeder flocks, and treat infected herds as soon as possible. [ABSTRACT FROM AUTHOR]

Mycoplasmosis is one of the most important diseases in the poultry industry. mycoplasma synoviae is one of the important disease-causing factors in chickens, which causes respiratory tract infection and synovitis. and leads to significant economic losses to the poultry industry all over the world. In this study, mycoplasma infection of broiler flocks in Ahvaz slaughterhouse was investigated using molecular method. For this purpose, 200 samples of lung and trachea tissues and joint fluid of hock joint were collected from 20 broiler flocks with macroscopic symptoms and 20 samples from the mentioned parts from apparently healthy broiler chickens of each flock in the winter season. Tissue DNA extraction was performed. The polymerase chain reaction (PCR) was performed to detect Vlha gene of mycoplasma synoviae. Of the 200 samples, 55 lungs and 45 tracheas from flocks with macroscopic symptoms and 4 lung and trachea samples from apparently healthy birds were positive. In the molecular examination of joint samples, no positive cases were observed for the presence of mycoplasma synoviae. The PCR products of positive samples were sent to Gene FanAvaran Company for sequencing. The sequence of a part of the Vlha gene of this isolate was compared with other isolates in the gene bank and it was about 99% similar to the previous Iranian isolates (IRG11/C/09, IRG6/C/08, IRG1/C/08). According to the results of the present study, mycoplasma synoviae is common among broiler flocks under investigation in Ahvaz slaughterhouse. Therefore, the policy makers of the health system of the poultry industry should prevent the economic losses caused by this disease with preventive measures. [ABSTRACT FROM AUTHOR]

Abstract Background In the past decade, Mycoplasma synoviae (M. synoviae) infection has become widely prevalent in China, has caused serious economic losses and has become one of the most important diseases in the chicken industry. Medication is a general approach for the control of M. synoviae infection, but antibiotics are sometimes ineffective in clinical practice. To investigate the sensitivity of M. synoviae to antimicrobials commonly used in the treatment of M. synoviae infection, the antibiotic susceptibility of 32 M. synoviae strains isolated from China from 2016 to 2019 were determined using the minimum inhibitory concentration (MIC) method. Results All isolates had low MIC values for the combination of lincomycin and spectinomycin, pleuromutilin, and macrolides. However, the M. synoviae isolates displayed variance in MICs for doxycycline hydrochloride with a range of 0.25 to 8 μg/mL, and oxytetracycline hydrochloride with a range of 0.5 to 8 μg/mL. Three and one M. synoviae isolates showed intermediate MIC values to doxycycline hydrochloride and oxytetracycline hydrochloride, respectively. High MIC values for enrofloxacin were detected in all isolates with MICs ranging from 4 to 32 μg/mL. Furthermore, comparison of the parC QRDR identified a mutation at nucleotide position 254 (C254T) resulting in a Thr 85 Ile amino acid change in all M. synoviae isolates and the reference strain ATCC 25204 being resistant to enrofloxacin. Moreover, mutations at Glu 804 Gly and Thr 686 Ala of gyrA QRDR were identified in all M. synoviae isolates and ATCC 25204. The mutation in the QRDR of the parE gene resulted in amino acid changes at positions 197 (Pro to Ser) in 27/32 M. synoviae isolates. Conclusion Three nonsynonymous mutations in gyrA and parE were first identified to be related to enrofloxacin resistance. Our results showed that M. synoviae resistance to enrofloxacin is widespread.

Mycoplasma synoviae (MS) is an essential pathogenic mycoplasma in poultry worldwide, posing a serious threat to the poultry industry's health. Timely detection is imperative for early diagnosis, prevention, and control of MS infection. Current laboratory methods for MS detection are generally complicated, time-consuming, and require sophisticated equipment. Therefore, a simple and rapid method is urgently needed. This study developed a novel real-time fluorescence-based recombinase-aided amplification (RF-RAA) technique for detecting MS nucleic acids, enabling target gene amplification within 20 min at 39°C. The RF-RAA outcomes are interpretable in 2 modalities: real-time fluorescence monitoring employing a temperature-controlled fluorescence detector or direct visual inspection facilitated by a portable blue light transilluminator. This method exhibits robust specificity, demonstrating no cross-reactivity with various common poultry pathogens, and achieves high sensitivity, detecting as low as 10 copies/μL for the standard plasmid. Seventy-one clinical samples of chicken throat swabs were detected by RF-RAA and real-time fluorescence quantitative polymerase chain reaction (qPCR) methods. The diagnostic coincidence rates of qPCR with RF-RAA (fluorescence monitoring) and RF-RAA (visual observation) were determined to be 100% and 97.2% (69/71), respectively. In conclusion, the RF-RAA method developed in this study provides a rapid and visually observable approach for MS detection, offering a novel technique to diagnosing MS infection, especially in resource-limited settings. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae (MS) is a contagious pathogen that poses a significant threat to the poultry industry. Detection plays an important role in the prevention and control of MS, particularly in differentiating between wild-type MS and live attenuated vaccine strains for vaccination selection and culling of animals with wild-type only. The live attenuated ts+ vaccine strain MS-H is recognized as the most effective and widely used vaccine. In this study, we have developed a method called double enzyme-activated differentiation probes PCR (DEA-probes PCR) for the differentiation of MS-H vaccine strain from wild-type strain by targeting the single nucleotide polymorphism (SNP) of the 367th nucleotide in the Obg gene sequence. We developed 2 modified probes with the ribonucleotide insert. When the probe perfectly complements with the target, the ribonuclease H2 (RNase H2) will cleave the ribonucleotide, resulting in the generation of fluorescent signal. With a detection limit of 5.8 copies/µL, the DEA-probes PCR method demonstrates 100% specificity in distinguishing wild-type MS from MS-H strains in 1 h. The method demonstrated great performance in real application of 100 superior palate cleft swab samples from chickens in poultry farms. Twenty-eight samples were detected as MS positive, consistent with the results of the Chinese industry standard method. Additionally, our method was able to distinguish 19 wild-type MS strains from 9 MS-H vaccine strains. The DEA-probes PCR method is rapid, specific and sensitive for SNP detection, overcoming the misidentification in MS detection and differentiation. It can be also applied to the differentiation of infected from vaccinated animals (DIVA) for other pathogens. [ABSTRACT FROM AUTHOR]

Mycoplasma synoviae causes infectious synovitis and respiratory tract infections in chickens and is responsible for significant economic losses in the poultry industry. Effective attachment and colonisation of the trachea is critical for the persistence of the organism and progression of the disease it causes. The respiratory tract infection is usually sub-clinical, but concurrent infection with infectious bronchitis virus (IBV) is known to enhance the pathogenicity of M. synoviae. This study aimed to explore differentially expressed genes in the tracheal mucosa, and their functional categories, during chronic infection with M. synoviae , using a M. synoviae -IBV infection model. The transcriptional profiles of the trachea were assessed 2 weeks after infection using RNA sequencing. In chickens infected with M. synoviae or IBV, only 1 or 8 genes were differentially expressed compared to uninfected chickens, respectively. In contrast, the M. synoviae -IBV infected chickens had 621 upregulated and 206 downregulated genes compared to uninfected chickens. Upregulated genes and their functional categories were suggestive of uncontrolled lymphoid cell proliferation and an ongoing pro-inflammatory response. Genes associated with anti-inflammatory effects, pathogen removal, apoptosis, regulation of the immune response, airway homoeostasis, cell adhesion and tissue regeneration were downregulated. Overall, transcriptional changes in the trachea, 2 weeks after infection with M. synoviae and IBV, indicate immune dysregulation, robust inflammation and a lack of cytotoxic damage during chronic infection. This model provides insights into the pathogenesis of chronic infection with M. synoviae. • Mycoplasma synoviae causes respiratory tract infections in chickens. • The tracheal response to infection with M. synoviae was assessed at 2 weeks post infection. • Transcriptional changes were indicative of uncontrolled lymphoid cell proliferation and robust inflammation. • This analysis provides insights into the pathogenesis of chronic infection with M. synoviae. [ABSTRACT FROM AUTHOR]