Mycoplasmas are intrinsically resistant to antimicrobials targeting the cell wall (fosfomycin, glycopeptides, or β-lactam antibiotics) and to sulfonamides, first-generation quinolones, trimethoprim, polymixins, and rifampicin. The antibiotics most frequently used to control mycoplasmal infections in animals are macrolides and tetracyclines. Lincosamides, fluoroquinolones, pleuromutilins, phenicols, and aminoglycosides can also be active. Standardization of methods used for determination of susceptibility levels is difficult since no quality control strains are available and because of species-specific growth requirements. Reduced susceptibility levels or resistances to several families of antimicrobials have been reported in field isolates of pathogenic Mycoplasma species of major veterinary interest: M. gallisepticum and M. synoviae in poultry; M. hyopneumoniae , M. hyorhinis , and M. hyosynoviae in swine; M. bovis in cattle; and M. agalactiae in small ruminants. The highest resistances are observed for macrolides, followed by tetracyclines. Most strains remain susceptible to fluoroquinolones. Pleuromutilins are the most effective antibiotics in vitro . Resistance frequencies vary according to the Mycoplasma species but also according to the countries or groups of animals from which the samples were taken. Point mutations in the target genes of different antimicrobials have been identified in resistant field isolates, in vitro -selected mutants, or strains reisolated after an experimental infection followed by one or several treatments: DNA-gyrase and topoisomerase IV for fluoroquinolones; 23S rRNA for macrolides, lincosamides, pleuromutilins, and amphenicols; 16S rRNAs for tetracyclines and aminoglycosides. Further work should be carried out to determine and harmonize specific breakpoints for animal mycoplasmas so that in vitro information can be used to provide advice on selection of in vivo treatments.

Background: Mycoplasma synoviae (MS) is known to cause Eggshell Apex Abnormality (EAA) syndrome characterized by an altered shell surface with increased translucency on the apex. However, no large-scale studies have been conducted to obtain prevalence data of EAA and MS isolates associated to this syndrome. This manuscript reports the results of two field studies performed in the French poultry industry (2015–2017): focusing mainly on investigation of presence and prevalence of EAA in different types of laying hen flocks (phase 1), and isolation of MS strains from EAA-infected flocks (phase 2). Results: The first survey included 77 farms of commercial layers in three French egg-production regions, hosting 40 flocks in alternative systems (ALT) and 56 in furnished cages (FC). Seven flocks (4 FC and 3 ALT) presented EAA clinical signs, giving a prevalence of 7.3% in this studied sample. A second independent field study was conducted to identify MS by in vitro cultivation and PCR in samples from 28 flocks with clinical signs of EAA. Different types of biological specimens were collected in EAA-affected flocks and submitted to the laboratory. M. synoviae was detected in 25/28 flocks, from both production systems (5/5 ALT and 20/23 FC). Detection of MS was significantly higher in tracheal swabs (59%) than in cloacal (10.5%), albumen (3.6%) and egg yolk (1.1%) swabs. It is worth to mention that attempts to clone MS from positive samples were often hampered by the presence of another Mycoplasma species, which showed fast growing behaviour in the selective media used in this study (Frey Medium 4 and Frey Medium 4 supplemented with erythromycin). The use of MALDI-TOF mass spectrometry in combination with next-generation sequencing (NGS) results allowed the identification of this fast growing mycoplasma as Mycoplasma pullorum, which was detected in 14 of the 25 (56%) MS-positive flocks. Conclusions: These results confirmed the presence of the EAA syndrome in MS-positive flocks of layers in France, reared in different regions and in different production systems (ALT and FC). Studies need to be conducted to test whether M. pullorum may influence the expression of clinical signs of EAA in MS-infected layer farms.

Avian mycoplasmas are known pathogens, which cause severe economic losses in poultry flocks. PCR is a rapid, sensitive, and less expensive diagnostic tool than culture for the identification of mycoplasmas in poultry farms. The objective of this study was to determine by PCR the presence of Mycoplasma spp., Mycoplasma gallisepticum (MG), Mycoplasma synoviae (MS), and Mycoplasma pullorum (MP) in laying hens located in the Sfax region, in the South of Tunisia. A total of 781 tracheal swabs were collected from 13 laying-hen farms without clinical signs at the date of sampling. MP was detected by a newly described specific PCR assay. The prevalence calculated from PCR results at the flock level was 100% for Mycoplasma spp., 0% for MG, 84.6% for MS and 61.5% for MP. The overall prevalence at the animal level was 38.7% for Mycoplasma spp., 0% for MG, 25% for MS and 6.4% for MP. The overall prevalence of 100% of avian mycoplasmas in laying-hen farms (38.7% prevalence at the animal level) shows an alarming situation. These results underline the importance of monitoring the emergence and spread of Mycoplasma strains in farms in order to decrease economic losses due to mycoplasmoses.

Mycoplasma infections pose a significant challenge in swine production because of their economic impact and the difficulties associated with diagnosis and treatment. Antimicrobials are often the only available option for combating mycoplasmoses despite reports of resistance to several molecules and the frequent chronic nature of infections, which can reduce treatment efficacy. This study aimed to investigate the use of antimicrobials against swine mycoplasmoses through an international field survey and to review the relevant scientific literature and regulatory recommendations. The survey was led by the MyMIC network, whose main objective is to standardise antimicrobial susceptibility testing (AST) methods for livestock mycoplasmas. The responses of 118 swine veterinarians across 25 countries were analysed. No significant variations in practices were observed between countries. Macrolides, tetracyclines, and pleuromutilins were the most prescribed antimicrobial classes (used in more than 40%, 20% and 5% of clinical cases, respectively), which is consistent with current recommendations. Additional diagnosis with laboratory-based methods was commonly reported, with only 2.5% of veterinarians indicating that they never used it. However, AST was not commonly performed; only 30% of respondents reported requesting it in more than half of the cases. Ineffective or critically important antibiotics, such as beta-lactams or fluoroquinolones, were occasionally prescribed (approximately 1% for each). Occasional deviations from the recommended dosages were also noted. These findings raise concerns regarding resistance selection and potential treatment failures. Our results underscore the need for enhanced diagnosis, especially AST, and for a wider dissemination and adoption of treatment guidelines to support the sustainable management of swine health. By developing and promoting harmonised AST protocols and enabling objective, data-driven prescriptions, MyMIC plays a key role in strengthening communication between swine veterinarians and diagnostic laboratories, thereby supporting international antimicrobial stewardship.

Mycoplasmas are known respiratory pathogens in tortoises, but few studies exist in snakes. To better understand the correlation with clinical signs and co-infections, samples from mycoplasma-positive snakes with and without clinical respiratory disease were analyzed. Oral swabs from 15 snakes (pythons n = 12, boas n = 3) were examined using polymerase chain reaction (PCR) and third-generation sequencing (TGS). Additionally, mycoplasma isolation assays were performed. Pathogens detected by PCR included Mycoplasmas (15/15, 100%), serpentoviruses (9/15, 60%), and Chlamydia sp. (2/15, 13%); those detected by TGS included Mycoplasmas (14/15, 93%), serpentoviruses (10/15, 67%), Chlamydia sp. (1/15, 7%), and 15 different bacterial species. Sequencing of the mycoplasma PCR products revealed a close genetic relationship to Mycoplasmopsis agassizii. TGS identified genetically distinct mycoplasmas and three different serpentoviruses. While mycoplasmas could not be successfully propagated, Brucella intermedia comb. nov. was identified in eight cultures. Respiratory disease in snakes is often multifactorial, involving various pathogens and environmental influences. This study demonstrates that comprehensive diagnostics are essential for understanding disease processes in snakes and improving the detection of diverse pathogens. Further research is needed to improve laboratory diagnostics for infectious diseases in reptiles and to better understand the roles of various pathogens in respiratory diseases in snakes.

Mycoplasmas are major pathogens in cattle, causing respiratory disease, mastitis and arthritis, and leading to significant economic losses. These pathogens’ microbial characteristics complicate diagnosis, treatment, and the performance of certain laboratory assays such as antimicrobial susceptibility testing (AST). Based on a dedicated survey, this study is the first international investigation of diagnostic and treatment strategies used by cattle veterinarians to manage mycoplasma infections. It also assesses how closely these practices align with current scientific evidence and regulatory recommendations. The survey was led by the MyMIC consortium, whose primary objective is to standardize AST methods for livestock mycoplasmas. The survey collected 270 responses from cattle veterinarians across 40 countries worldwide. PCR was the most commonly used diagnostic method, while culture and AST were performed less frequently. Macrolides, florfenicol and tetracyclines were the preferred antimicrobials, in line with current guidelines. However, some prescription practices deviated from recommendations, including the use of ineffective antimicrobial classes (e.g., beta-lactams and trimethoprim/sulfonamides) or of critically important antimicrobial classes as well as inappropriate dosing or duration regimens. These findings emphasize the need for coordinated actions at multiple levels: promoting the use of AST by veterinarians, disseminating updated diagnostic and treatment recommendations, and developing standardized guidelines and procedures for AST in livestock mycoplasmas. The MyMIC consortium has established an inventory of veterinary strategies for antimicrobial use and diagnosis, including resistance surveillance, thereby contributing to improved antimicrobial stewardship and to the health of livestock systems affected by mycoplasmoses. Continued support of this initiative is strongly recommended.