Exploring Mycoplasma ovipneumoniae NXNK2203 infection in sheep: insights from histopathology and whole genome sequencing.

BACKGROUND: Mycoplasma ovipneumoniae (M. ovipneumoniae) is a significant pathogen causing respiratory infections in goats and sheep. This study focuses on investigating vulnerability of Hu sheep to M. ovipneumoniae infection in the context of late spring's cold weather conditions through detailed autopsy of a severely affected Hu sheep and whole genome sequencing of M. ovipneumoniae. RESULTS: The autopsy findings of the deceased sheep revealed severe pulmonary damage with concentrated tracheal and lung lesions. Histopathological analysis showed tissue degeneration, mucus accumulation, alveolar septum thickening, and cellular necrosis. Immunohistochemistry analysis indicated that M. ovipneumoniae was more in the bronchi compared to the trachea. Genome analysis of M. ovipneumoniae identified a 1,014,835 bp with 686 coding sequences, 3 rRNAs, 30 tRNAs, 6 CRISPRs, 11 genomic islands, 4 prophages, 73 virulence factors, and 20 secreted proteins. CONCLUSION: This study investigates the vulnerability of Hu sheep to M. ovipneumoniae infection during late spring's cold weather conditions. Autopsy findings showed severe pulmonary injury in affected sheep, and whole genome sequencing identified genetic elements associated with pathogenicity and virulence factors of M. ovipneumoniae.

Mesomycoplasma ovipneumoniae from goats with respiratory infection: pathogenic characteristics, population structure, and genomic features.

BACKGROUND: Mycoplasma ovipneumoniae is a critical pathogen that causes respiratory diseases that threaten Caprini health and cause economic damage. A genome-wide study of M. ovipneumoniae will help understand the pathogenic characteristics of this microorganism. RESULTS: Toxicological pathology and whole-genome sequencing of nine M. ovipneumoniae strains isolated from goats were performed using an epidemiological survey. These strains exhibited anterior ventral lung consolidation, typical of bronchopneumonia in goats. Average nucleotide identity and phylogenetic analysis based on whole-genome sequences showed that all M. ovipneumoniae strains clustered into two clades, largely in accordance with their geographical origins. The pan-genome of the 23 M. ovipneumoniae strains contained 5,596 genes, including 385 core, 210 soft core, and 5,001 accessory genes. Among these genes, two protein-coding genes were annotated as cilium adhesion and eight as paralog surface adhesins when annotated to VFDB, and no antibiotic resistance-related genes were predicted. Additionally, 23 strains carried glucosidase-related genes (ycjT and group_1595) and glucosidase-related genes (atpD_2), indicating that M. ovipneumoniae possesses a wide range of glycoside hydrolase activities. CONCLUSIONS: The population structure and genomic features identified in this study will facilitate further investigations into the pathogenesis of M. ovipneumoniae and lay the foundation for the development of preventive and therapeutic methods.

Antibiotic treatment of Mycoplasma ovipneumoniae in domestic sheep (Ovis aries): Working at the livestock-wildlife interface in Yukon, Canada.

Domestic sheep (Ovis aries) can carry the bacterium Mycoplasma ovipneumoniae (M. ovipneumoniae) in their upper respiratory tract, often with little effect on health and productivity. However, for bighorn sheep (Ovis canadensis) populations, there is a link between M. ovipneumoniae infection and pneumonia, poor lamb recruitment, and high fatality rate. Because of these outcomes, preventing transmission of M. ovipneumoniae to free-ranging wild sheep has garnered interest from both the livestock and wildlife sectors. We hypothesized that treatment with intranasal and systemic enrofloxacin would reduce the prevalence of M. ovipneumoniae-positive animals in a flock of domestic sheep. Initially, the prevalence decreased in the treated group; but by 34 d post-treatment, the number of M. ovipneumoniae-positive sheep returned to near pretreatment prevalence. Key clinical message: Test-and-slaughter is a method used to reduce the risk of transmission of pneumonia-causing M. ovipneumoniae from domestic sheep and goats to free-ranging wild sheep. In an effort to find an alternative, we used enrofloxacin to treat a flock of M. ovipneumoniae-positive domestic sheep; however, long-term reduction of M. ovipneumoniae prevalence in the flock was not achieved.

Traitement antibiotique de Mycoplasma ovipneumoniae chez le mouton domestique (Ovis aries): travail à l'interface bétail-faune au Yukon, Canada. Les moutons domestiques (Ovis aries) peuvent être porteurs de la bactérie Mycoplasma ovipneumoniae (M. ovipneumoniae) dans leurs voies respiratoires supérieures, avec souvent peu d'effets sur la santé et la productivité. Cependant, pour les populations de mouflons d'Amérique (Ovis canadensis), il existe un lien entre l'infection à M. ovipneumoniae et la pneumonie, un faible recrutement d'agneaux et un taux de mortalité élevé. En raison de ces résultats, la prévention de la transmission de M. ovipneumoniae aux moutons sauvages en liberté a suscité l'intérêt des secteurs de l'élevage et de la faune sauvage. Nous avons émis l'hypothèse qu'un traitement par enrofloxacine intranasale et systémique réduirait la prévalence d'animaux positifs à M. ovipneumoniae dans un troupeau de moutons domestiques. Initialement, la prévalence a diminué dans le groupe traité; mais 34 jours après le traitement, le nombre de moutons positifs à M. ovipneumoniae est revenu à une prévalence proche de celle précédant le traitement.Message clinique clé :L'essai et l'abattage sont une méthode utilisée pour réduire le risque de transmission de M. ovipneumoniae, responsable de la pneumonie, des moutons et chèvres domestiques aux moutons sauvages en liberté. Dans le but de trouver une alternative, nous avons utilisé l'enrofloxacine pour traiter un troupeau de moutons domestiques positifs à M. ovipneumoniae; cependant, aucune réduction à long terme de la prévalence de M. ovipneumoniae dans le troupeau n'a été obtenue.(Traduit par Dr Serge Messier).

DNA hybridization-induced fluorescence variation in ThT: a new strategy of developing aqueous sensors for MO genes.

Mycoplasma ovipneumoniae (MO), a disease-causing pathogen with some of the highest levels of morbidity and mortality, can spread silently at the herd level. A novel alternative nanoprobe for MO was prepared using porous metal-organic frameworks (MOFs) as the scaffold and hairpin DNA with specific sequences of MO and G4 segments as the probe. This preparation was based on the strong fluorescence emission by ThT (thioflavin T) in the limitation of G-quadruplexes with a cavity structure. The use of MOFs effectively limited the folding behavior of G4 as a part of the probe to improve the defect of the strong background signal caused by the free-state G4Probe in a buffer. The results from the selectivity experiment showed that only a trace amount of the target with lower ΔG could be the "key" to the highly efficient triggering of the release behavior of the G4Probe from MOFs and the subsequent change in the fluorescence behavior of ThT. The DNA targets could be determined by observing the change in the signal. More importantly, the probe showed a low detection limit and a good linear correlation between the concentration of target DNA ranging from 10-10 M to 10-6 M not only in buffer but also in natural complex media. Moreover, the operation involved in the whole strategy was simple and the total cost was low. These findings demonstrated the value of the probe in further clinical diagnosis. This study reports the successful construction of a ΔG-sensitive sensor for MO for the first time.

Comparative analysis on lung transcriptome of Mycoplasma ovipneumoniae (Mo) - infected Bashbay sheep and argali hybrid sheep.

BACKGROUND: Bashbay sheep (Bbs) has a certain degree of resistance to Mycoplasma ovipneumoniae (Mo), however, Argali hybrid sheep (Ahs) is susceptible to Mo. To understand the molecular mechanisms underlying the difference of the susceptibility for Mo infection, RNA-sequencing technology was used to compare the transcriptomic response of the lung tissue of Mo-infected Bbs and Ahs. RESULTS: Six Bbs and six Ahs were divided into experimental group and control group respectively, all of them were experimentally infected with Mo by intratracheal injection. For collecting lung tissue samples, three Bbs and three Ahs were sacrificed on day 4 post-infection, and the others were sacrificed on day 14 post-infection. Total RNA extracted from lung tissue were used for transcriptome analyses based on high-throughput sequencing technique and bioinformatics. The results showed that 212 (146 up-regulated, 66 down-regulated) DEGs were found when comparing transcriptomic data of Bbs and Ahs at 4th dpi, besides, 311 (158 up-regulated, 153 down-regulated) DEGs were found at 14th dpi. After GO analysis, three main GO items protein glycosylation, immune response and positive regulation of gene expression were found related to Mo infection. In addition, there were 20 DEGs enriched in these above items, such as SPLUC1 (BPIFA1), P2X7R, DQA, HO-1 and SP-A (SFTPA-1). CONCLUSIONS: These selected 20 DEGs associated with Mo infection laid the foundation for further study on the underlying molecular mechanism involved in high level of resistance to Mo expressed by Bbs, meanwhile, provided deeper understandings about the development of pathogenicity and host-pathogen interactions.

NOD2/c-Jun NH2-Terminal Kinase Triggers Mycoplasma ovipneumoniae-Induced Macrophage Autophagy.

Mycoplasma ovipneumoniae belongs to Mycoplasma, a genus containing the smallest self-replicating microorganisms, and causes infectious pleuropneumonia in goats and sheep. Nucleotide-binding oligomerization domain-containing protein (NOD2), an intracellular pattern recognition receptor, interacts with muramyl dipeptide (MDP) to recognize bacterial peptidoglycans and is involved in autophagy induction. However, there have been no reports about NOD recognition of mycoplasmas or M. ovipneumoniae-induced autophagy. In this study, we sought to determine the role of NOD2 in M. ovipneumoniae-induced autophagy using Western blotting, immunofluorescence, real-time PCR (RT-PCR), and color-changing unit (CCU) analysis. M. ovipneumoniae infection markedly increased NOD2 but did not increase NOD1 expression in RAW 264.7 cells. Treating RAW 264.7 cells with MDP significantly increased colocalization of M. ovipneumoniae and LC3, whereas treatment with NOD inhibitor, NOD-IN-1, decreased colocalization of M. ovipneumoniae and LC3. Furthermore, suppressing NOD2 expression with small interfering RNA (siRNA)-NOD2 failed to trigger M. ovipneumoniae-induced autophagy by detecting autophagy markers Atg5, beclin1, and LC3-II. In addition, M. ovipneumoniae infection significantly increased the phosphorylated c-Jun NH2-terminal kinase (p-JNK)/JNK, p-Bcl-2/Bcl-2, beclin1, Atg5, and LC3-II ratios in RAW 264.7 cells. Treatment with JNK inhibitor, SP600126, or siRNA-NOD2 did not increase this reaction. These findings suggested that M. ovipneumoniae infection activated NOD2, and both NOD2 and JNK pathway activation promoted M. ovipneumoniae-induced autophagy. This study provides new insight into the NOD2 reorganization mechanism and the pathogenesis of M. ovipneumoniae infection.IMPORTANCEM. ovipneumoniae, which lacks a cell wall, causes infectious pleuropneumonia in goats and sheep. In the present study, we focused on the interaction between NOD and M. ovipneumoniae, as well as its association with autophagy. We showed for the first time that NOD2 was activated by M. ovipneumoniae even when peptidoglycans were not present. We also observed that both NOD2 and JNK pathway activation promoted M. ovipneumoniae-induced autophagy.

Development and validation of the isothermal recombinase polymerase amplification assays for rapid detection of Mycoplasma ovipneumoniae in sheep.

BACKGROUND: Mycoplasmal pneumonia is an important infectious disease that threatens sheep and goat production worldwide, and Mycoplasma ovipneumoniae is one of major etiological agent causing mycoplasmal pneumonia. Recombinase polymerase amplification (RPA) is an isothermal nucleic acid amplification technique, and RPA-based diagnostic assays have been described for the detection of different types of pathogens. RESULTS: The RPA assays using real-time fluorescence detection (real-time RPA) and lateral flow strip detection (LFS RPA) were developed to detect M. ovipneumoniae targeting a conserved region of the 16S rRNA gene. Real-time RPA was performed in a portable florescence scanner at 39 °C for 20 min. LFS RPA was performed in a portable metal bath incubator at 39 °C for 15 min, and the amplicons were visualized with the naked eyes within 5 min on the lateral flow strip. Both assays were highly specific for M. ovipneumoniae, as there were no cross-reactions with other microorganisms tested, especially the pathogens involved in respiratory complex and other mycoplasmas frequently identified in ruminants. The limit of detection of LFS RPA assay was 1.0 × 101 copies per reaction using a recombinant plasmid containing target gene as template, which is 10 times lower than the limit of detection of the real-time RPA and real-time PCR assays. The RPA assays were further validated on 111 clinical sheep nasal swab and fresh lung samples, and M. ovipneumoniae DNA was detected in 29 samples in the real-time RPA, 31 samples in the LFS RPA and 32 samples in the real-time PCR assay. Compared to real-time PCR, the real-time RPA and LFS RPA showed diagnostic specificity of 100 and 98.73%, diagnostic sensitivity of 90.63 and 93.75%, and a kappa coefficient of 0.932 and 0.934, respectively. CONCLUSIONS: The developed real-time RPA and LFS RPA assays provide the attractive and promising tools for rapid, convenient and reliable detection of M. ovipneumoniae in sheep, especially in resource-limited settings. However, the effectiveness of the developed RPA assays in the detection of M. ovipneumoniae in goats needs to be further validated.

Loop-mediated isothermal amplification-lateral-flow dipstick (LAMP-LFD) to detect Mycoplasma ovipneumoniae.

In order to establish a rapid detection method for Mycoplasma ovipneumoniae, this study used the loop-mediated isothermal amplification (LAMP) technique to carry out nucleic acid amplification and chromatographic visualization via a lateral flow dipstick (LFD) assay. The M. ovipneumoniae elongation factor TU gene (EF-TU) was detected using a set of specific primers designed for the EF-TU gene, and the EF-TU FIP was detected by biotin labeling, which was used in the LAMP amplification reaction. The digoxin-labeled probe specifically hybridized with LAMP products, which were visually detected by LFD. Here, we established the M. ovipneumoniae LAMP-LFD rapid detection method and tested the specificity, sensitivity, and clinical application of this method. Results showed that the optimized LAMP performed at 60 °C for 60 min, and LFD can specifically and visually detect M. ovipneumoniae with a minimum detectable concentration at 1.0 × 102 CFU/mL. The sensitivity of LAMP-LFD was 1000 times that of the conventional PCR detection methods, and the clinical lung tissue detection rate was 86% of 50 suspected sheep infected with M. ovipneumoniae. In conclusion, LAMP-LFD was established in this study to detect M. ovipneumoniae, a method that was highly specific, sensitive, and easy to operate, and provides a new method for the prevention and diagnosis of M. ovipneumoniae infection.

Mycoplasma ovipneumoniae in Wildlife Species beyond Subfamily Caprinae.

Elucidating the emergence of Mycoplasma ovipneumoniae-associated respiratory disease in ruminants requires identification of the pathogen host range. This bacterium was thought to be host restricted to subfamily Caprinae, but we describe its identification in healthy moose, caribou, and mule deer and diseased mule and white-tailed deer, all species in subfamily Capreolinae.

Age-specific infectious period shapes dynamics of pneumonia in bighorn sheep.

Superspreading, the phenomenon where a small proportion of individuals contribute disproportionately to new infections, has profound effects on disease dynamics. Superspreading can arise through variation in contacts, infectiousness or infectious periods. The latter has received little attention, yet it drives the dynamics of many diseases of critical public health, livestock health and conservation concern. Here, we present rare evidence of variation in infectious periods underlying a superspreading phenomenon in a free-ranging wildlife system. We detected persistent infections of Mycoplasma ovipneumoniae, the primary causative agent of pneumonia in bighorn sheep (Ovis canadensis), in a small number of older individuals that were homozygous at an immunologically relevant genetic locus. Interactions among age-structure, genetic composition and infectious periods may drive feedbacks in disease dynamics that determine the magnitude of population response to infection. Accordingly, variation in initial conditions may explain divergent population responses to infection that range from recovery to catastrophic decline and extirpation.

Contact and contagion: Probability of transmission given contact varies with demographic state in bighorn sheep.

Understanding both contact and probability of transmission given contact are key to managing wildlife disease. However, wildlife disease research tends to focus on contact heterogeneity, in part because the probability of transmission given contact is notoriously difficult to measure. Here, we present a first step towards empirically investigating the probability of transmission given contact in free-ranging wildlife. We used measured contact networks to test whether bighorn sheep demographic states vary systematically in infectiousness or susceptibility to Mycoplasma ovipneumoniae, an agent responsible for bighorn sheep pneumonia. We built covariates using contact network metrics, demographic information and infection status, and used logistic regression to relate those covariates to lamb survival. The covariate set contained degree, a classic network metric describing node centrality, but also included covariates breaking the network metrics into subsets that differentiated between contacts with yearlings, ewes with lambs, and ewes without lambs, and animals with and without active infections. Yearlings, ewes with lambs, and ewes without lambs showed similar group membership patterns, but direct interactions involving touch occurred at a rate two orders of magnitude higher between lambs and reproductive ewes than between any classes of adults or yearlings, and one order of magnitude higher than direct interactions between multiple lambs. Although yearlings and non-reproductive bighorn ewes regularly carried M. ovipneumoniae, our models suggest that a contact with an infected reproductive ewe had approximately five times the odds of producing a lamb mortality event of an identical contact with an infected dry ewe or yearling. Consequently, management actions targeting infected animals might lead to unnecessary removal of young animals that carry pathogens but rarely transmit. This analysis demonstrates a simple logistic regression approach for testing a priori hypotheses about variation in the odds of transmission given contact for free-ranging hosts, and may be broadly applicable for investigations in wildlife disease ecology.

Capsular Polysaccharide is a Main Component of Mycoplasma ovipneumoniae in the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells.

Mycoplasma ovipneumoniae (M. ovipneumoniae) is characterized as an etiological agent of primary atypical pneumonia that specifically infects sheep and goat. In an attempt to better understand the pathogen-host interaction between the invading M. ovipneumoniae and airway epithelial cells, we investigated the host inflammatory responses against capsular polysaccharide (designated as CPS) of M. ovipneumoniae using sheep bronchial epithelial cells cultured in an air-liquid interface (ALI) model. Results showed that CPS derived from M. ovipneumoniae could activate toll-like receptor- (TLR-) mediated inflammatory responses, along with an elevated expression of nuclear factor kappa B (NF-κB), activator protein-1 (AP-1), and interferon regulatory factor 3 (IRF3) as well as various inflammatory-associated mediators, representatively including proinflammatory cytokines, such as IL1ß, TNFα, and IL8, and anti-inflammatory cytokines such as IL10 and TGFß of TLR signaling cascade. Mechanistically, the CPS-induced inflammation was TLR initiated and was mediated by activations of both MyD88-dependent and MyD88-independent signaling pathways. Of importance, a blockage of CPS with specific antibody led a significant reduction of M. ovipneumoniae-induced inflammatory responses in sheep bronchial epithelial cells. These results suggested that CPS is a key virulent component of M. ovipneumoniae, which may play a crucial role in the inflammatory response induced by M. ovipneumoniae infections.

Serological and molecular survey of sheep infected with Mycoplasma ovipneumoniae in Xinjiang, China.

Mycoplasma pneumonia is one of the most important infectious diseases that threaten sheep production. In order to investigate the epidemic status of Mycoplasma ovipneumoniae infection in sheep, indirect hemagglutination assay was used to analyze 1679 serum samples collected from four different breeds of sheep (Kazak sheep, Hu sheep, Merino sheep, and Duolang sheep) in six regions in Xinjiang between 2012 and 2014. One thousand one hundred sixty-nine sheep nasal swabs and 180 lungs were PCR analyzed. The results showed that the average positive rates of the serum samples were 17.75 %. The positive rates were between 9.76 and 30.61 % in the four breeds. Among them, the Hu sheep had a significantly higher rate than other breeds (P < 0.05). The average positive rates of nasal swabs and lungs were 10.18 and 28.89 %, respectively. Based on the phylogenetic trees of 16S RNA gene, the isolates were closest to those strains isolated from inland areas of China, indicating that these epidemic isolates came from the trans-province introductions. Our survey suggests that quarantine is necessary for sheep imported from inland, and effective immunization should be implemented in sheep susceptible to M. ovipneumoniae in Xinjiang, China.

Seroprevalence and molecular detection of Mycoplasma ovipneumoniae in goats in tropical China.

In the present study, the seroprevalence and genetic identification of Mycoplasma ovipneumoniae infection in goats were investigated in Hainan Province, tropical China between October 2012 and October 2013. A total of 1,210 serum samples collected from 16 herds in various administrative regions in tropical China were evaluated using indirect hemagglutination assay (IHA). Antibodies to M. ovipneumoniae were tested in (31.7 %, 95 % confidence interval (CI) 29-34.3) 383 of 1,210 serum samples (IHA titer ≥1:16). The M. ovipneumoniae seroprevalence ranged from 26.8 % (95 % CI 20.8-32.9) to 39 % (95 % CI 30.8-47.2) among different regions in tropical China, and the difference was statistically significant (P < 0.01). The seroprevalence of M. ovipneumoniae infection in goats was higher in winter (46.1 %, 95 % CI 39.6-52.5) and spring (33.8 %, 95 % CI 28.3-39.3) than in autumn (27.5 %, 95 % CI 22.6-32.3) and summer (24.7 %, 95 % CI 20.3-29.1), and the difference was statistically significant (P < 0.01). In addition, DNA was extracted from nasal swab; lung samples and the 16S rRNA gene sequences were amplified by polymerase chain reaction (PCR) and then sequenced. Twenty-four of 329 (7.3 %) nasal swab samples and 73 of 280 (26.1 %) pneumonic lung tissues were found to contain M. ovipneumoniae, respectively. The results of the present survey indicate that M. ovipneumoniae infection is highly prevalent in goats in tropical China. This is the first report of the comprehensive survey of M. ovipneumoniae prevalence in goats in China.

Development of loop-mediated isothermal amplification for detection of Mycoplasma ovipneumoniae in goats, Capra hircus.

Mycoplasma ovipneumoniae is an important pathogen that causes respiratory disease in goats and sheep, leading to significant economic losses in the livestock industry. A quick and robust diagnostic test will aid in early diagnosis and treatment of the disease. Loop-mediated isothermal amplification (LAMP) offers several advantages over traditional PCR, including faster amplification, simpler operation, and lower equipment requirements, making it a promising tool for use in basic livestock units where resources and infrastructure may be limited. The present study reports on developing a LAMP assay to rapidly detect M. ovipneumoniae in goats (Capra hircus) targeting the 16S rRNA gene. LAMP was optimized to perform at 60 °C for 75 min. The result was visualized by a change in colour from deep pink to orange and further confirmed by gel electrophoresis, which gave a typical ladder-like pattern. The detection limit of the assay was as low as 0.04 ng/µl, indicating the high sensitivity of the assay. The test failed to amplify DNA isolated from healthy goat blood, Mycoplasma arginini DNA, and Staphylococcus aureus DNA. The sensitivity, specificity, and accuracy of the assay were 97.73% and 94.83%, and 96.08%, respectively. The study concludes that the developed loop-mediated isothermal amplification assay is a practical and reliable tool for field-level diagnosis of M. ovipneumoniae infections in goats, with high sensitivity and specificity under resource-limited conditions.

Evaluating the transmission dynamics and host competency of aoudad (Ammotragus lervia) experimentally infected with Mycoplasma ovipneumoniae and leukotoxigenic Pasteurellaceae.

Feral populations of aoudad (Ammotragus lervia) occur in Texas bighorn sheep (Ovis canadensis) habitat and pose several conceptual ecological threats to bighorn sheep re-establishment efforts. The potential threat of disease transmission from aoudad to bighorn sheep may exacerbate these issues, but the host competency of aoudad and subsequent pathophysiology and transmissibility of pneumonic pathogens involved in the bighorn sheep respiratory disease complex is largely unknown. Because the largest population-limiting diseases of bighorn sheep involve pathogens causing bronchopneumonia, we evaluated the host competency of aoudad for Mycoplasma ovipneumoniae and leukotoxigenic Pasteurellaceae. Specifically, we described the shedding dynamics, pathogen carriage, seroconversion, clinical patterns, and pathological effects of experimental infection among wild aoudad held in captivity. We found that aoudad are competent hosts capable of maintaining and intraspecifically transmitting Mycoplasma ovipneumoniae and Pasteurellaceae and can shed the bacteria for 53 days after exposure. Aoudad developed limited clinical signs and pathological findings ranged from mild chronic lymphohistiocytic bronchointerstitial pneumonia to severe and acute suppurative pneumonia, similarly, observed in bighorn sheep infected with Mycoplasma spp. and Pasteurellaceae bacteria, respectively. Furthermore, as expected, clinical signs and lesions were often more severe in aoudad inoculated with a combination of Mycoplasma ovipneumoniae and Pasteurellaceae as compared to aoudad inoculated with only Mycoplasma ovipneumoniae. There may be evidence of interindividual susceptibility, pathogenicity, and/or transmissibility, indicated by individual aoudad maintaining varying severities of chronic infection who may be carriers continuously shedding pathogens. This is the first study to date to demonstrate that aoudad are a conceptual disease transmission threat to sympatric bighorn sheep populations due to their host competency and intraspecific transmission capabilities.

Pathology of Chronic Mycoplasma ovipneumoniae Carriers in a Declining Bighorn Sheep (Ovis canadensis) Population.

Bighorn sheep (Ovis canadensis) across North America commonly experience population-limiting epizootics of respiratory disease. Although many cases of bighorn sheep pneumonia are polymicrobial, Mycoplasma ovipneumoniae is most frequently associated with all-age mortality events followed by years of low recruitment. Chronic carriage of M. ovipneumoniae by adult females serves as a source of exposure of naïve juveniles; relatively few ewes may be responsible for maintenance of infection within a herd. Test-and-remove strategies focused on removal of adult females with evidence of persistent or intermittent shedding (hereafter chronic carriers) may reduce prevalence and mitigate mortality. Postmortem confirmation of pneumonia in chronic carriers has been inadequately reported and the pathology has not been thoroughly characterized, limiting our understanding of important processes shaping the epidemiology of pneumonia in bighorn sheep. Here we document postmortem findings and characterize the lesions of seven ewes removed from a declining bighorn sheep population in Wyoming, USA, following at least two antemortem detections of M. ovipneumoniae within a 14-mo period. We confirmed that 6/7 (85.7%) had variable degrees of chronic pneumonia. Mycoplasma ovipneumoniae was detected in the lung of 4/7 (57.1%) animals postmortem. Four (57.1%) had paranasal sinus masses, all of which were classified as inflammatory, hyperplastic lesions. Pasteurella multocida was detected in all seven (100%) animals, while Trueperella pyogenes was detected in 5/7 (71.4%). Our findings indicate that not all chronic carriers have pneumonia, nor do all have detectable M. ovipneumoniae in the lung. Further, paranasal sinus masses are a common but inconsistent finding, and whether sinus lesions predispose to persistence or result from chronic carriage remains unclear. Our findings indicate that disease is variable in chronic M. ovipneumoniae carriers, underscoring the need for further efforts to characterize pathologic processes and underlying mechanisms in this system to inform management.

Characterization and epidemiologic analysis of mycoplasmal pneumonia of sheep in Qinghai Province.

Mycoplasmal pneumonia in sheep and goats usually result covert but huge economic losses in the sheep and goat industry. The disease is prevalent in various countries in Africa and Asia. Clinical manifestations in affected animals include anorexia, fever, and respiratory symptoms such as dyspnea, polypnea, cough, and nasal discharge. Due to similarities with other respiratory infections, accurate diagnosis can be challenging, and isolating the causative organism is often problematic. However, the utilization of molecular techniques, such as PCR, allows for rapid and specific identification of pathogens. Thus, a goat infection model with Mycoplasma was established and the pathogen was tested using PCR. The results indicated that this approach could be effectively utilized for the rapid detection of mycoplasma in clinical settings. Additionally, the prevalence of contagious pleuropneumonia of sheep in Qinghai Province was further investigated through PCR analysis. A total of 340 nasal swabs were collected from 17 sheep farms in Qinghai province. Among these samples, 84 tested positive for Mycoplasma mycoides subsp. capri (Mmc) and 148 tested positive for Mycoplasma ovipneumoniae (Movi), resulting in positive rates of 24.71% and 43.53% respectively. Furthermore, our investigation revealed positive PCR results for nasal swabs, trachea, and lung samples obtained from sheep exhibiting symptoms suggestive of mycoplasma infection. Moreover, three distinct strains were isolated from these positive samples. Additionally, the inflammatory cytokines of peripheral blood mononuclear cells (PBMCs) were assessed using RT-PCR. The findings demonstrated a high susceptibility of sheep to Movi in Qinghai province, with infected sheep displaying an inflammatory response. Consequently, the outcomes of this study will furnish valuable epidemiological insights for the effective prevention and control of this disease within Qinghai Province.

Comparative genomic analysis of Brazilian Mesomycoplasma ovipneumoniae strains revealed genomic differences associated with the geographic origin and health status and mutations in the gyrA.

Sheep respiratory disease (SRD) is a multifactorial illness commonly affecting sheep. Mesomycoplasma (Mycoplasma) ovipneumoniae is one of the most important etiological agents of SRD and should be better understood, especially in countries where it was recently detected, such as Brazil. Also, the intensive use of quinolones in mycoplasmal infections increases the selective pressure for resistance to this drug class, and no data about antimicrobial resistance in Brazil is available. Therefore, this study aimed to perform a comparative genomic analysis of newly isolated Brazilian M. ovipneumoniae strains, identify point mutations in target genes that may be associated with antibiotic resistance, and perform a phylogenomic analysis of these strains with available genome representatives of M. ovipneumoniae. Glucose-fermenting fried egg-like colonies identified as M. ovipneumoniae were obtained after a culture of tracheobronchial lavage from infected sheep. The genomes were sequenced, de novo assembled and comparatively evaluated. Important putative virulence factors were detected in all isolates: the analysis of the average nucleotide homology of all these genes with the M. ovipneumoniae ATCC 29419 revealed associations between clpB, lgt, tuf, and dnaJ genes and geographic location. In addition, nucleotide substitutions in a few positions of the Quinolone-Resistant Determinant Region of the gyrA gene, including the Ser83Ala, were detected. The phylogenomic analysis showed that the Brazilian isolates belonged to two different clades corresponding to geographic location, and the isolates from São Paulo showed high similarity, which differs from isolates from Rio de Janeiro. This first genomic analysis of the Brazilian M. ovipneumoniae genomes demonstrates strain segregation according to location and health status, reinforcing the importance of continuous surveillance and diagnostics of this bacteria causing sheep respiratory disease in the Brazilian flocks.

Identification by culture, PCR, and immunohistochemistry of mycoplasmas and their molecular typing in sheep and lamb lungs with pneumonia in Eastern Turkey.

This study used cultures, polymerase chain reaction (PCR), and immunoperoxidase to examine samples from 216 lungs from sheep and lambs with macroscopic pneumonia lesions for the presence of Mycoplasma species. DNA was extracted from lung tissue samples and broth cultures with the help of a DNA extraction kit and replicated using genus-specific and species-specific primers for mycoplasma. The lung samples were examined by the immunoperoxidase method using hyperimmune Mycoplasma ovipneumoniae serum. The randomly amplified polymorphic DNA (RAPD) test was used for the molecular typing of M. ovipneumoniae isolates. Mycoplasma was isolated in the cultures of 80 (37.03 %) of a total of 216 lung samples. Genus-specific mycoplasma DNA was identified by PCR in 96 (44.44 %) samples in broth cultures and 36 (16.66 %) directly in the lung tissue. Of these 96 cases in which genus-specific identification was made, 57 (59.37 %) were positive for reaction with species-specific primers for M. ovipneumoniae and 31 (32.29 %) for Mycoplasma arginini. The DNA of neither of the latter two species could be identified in the remaining eight samples (8.33 %) where mycoplasma had been identified. As for the immunoperoxidase method, it identified M. ovipneumoniae in 61 of 216 lung samples (28 %). Positive staining was concentrated in the bronchial epithelium cell cytoplasm and cell surface. RAPD analysis resulted in 15 different profiles. Our results suggest that PCR methods could be successfully used in the diagnosis of mycoplasma infections as an alternative to culture method and identifying this agent at the species level.