The Role of Mycoplasma bovirhinis in the Development of Singular and Concomitant Respiratory Infections in Dairy Calves from Southern Brazil.

The role of Mycoplasma bovirhinis in the development of pulmonary disease in cattle is controversial and was never evaluated in cattle from Latin America. This study investigated the respiratory infection dynamics associated with M. bovirhinis in suckling calves from 15 dairy cattle herds in Southern Brazil. Nasal swabs were obtained from asymptomatic (n = 102) and calves with clinical manifestations (n = 103) of bovine respiratory disease (BRD) and used in molecular assays to identify the specific genes of viral and bacterial disease pathogens of BRD. Only M. bovirhinis, bovine coronavirus (BCoV), ovine gammaherpesvirus 2 (OvGHV2), Histophilus somni, Pasteurella multocida, and Mannheimia haemolytica were detected. M. bovirhinis was the most frequently diagnosed pathogen in diseased (57.8%; 59/102) and asymptomatic (55.3%; 57/103) calves at all farms. BCoV-related infections were diagnosed in diseased (52%; 53/102) and asymptomatic (51.4%; 53/103) calves and occurred in 93.3% (14/15) of all farms. Similarly, infectious due to OvGHV2 occurred in diseased (37.2%; 38/102) and asymptomatic (27.2%; /28/103) calves and were diagnosed in 80% (12/15) of all farms investigated. Significant statistical differences were not identified when the two groups of calves were compared at most farms, except for infections due to OvGHV2 that affected five calves at one farm. These results demonstrated that the respiratory infection dynamics of M. bovirhinis identified in Southern Brazil are similar to those observed worldwide, suggesting that there is not enough sufficient collected data to consider M. bovirhinis as a pathogen of respiratory infections in cattle. Additionally, the possible roles of BCoV and OvGHV2 in the development of BRD are discussed.

PCR múltiplex para el diagnóstico de microorganismos atípicos transmitidos sexualmente / Multiplex PCR testing for the atypical microorganisms diagnosis sexually transmitted

Resumen Antecedentes: Las infecciones de transmisión sexual son un problema de salud pública mundial. El análisis rutinario incluye solo pruebas microbiológicas y serológicas para el diagnóstico de patógenos. Los microorganismos atípicos como Chlamydia trachomatis y micoplasmas no son identificados debido a los requerimientos. Además, no es incluida Gardnerella vaginalis, aunque se asocia a la vaginosis bacteriana. Objetivo: Desarrollar una PCR múltiplex para el diagnóstico de C. trachomatis, micoplasmas y G. vaginalis. Método: Se estandarizó la PCR múltiplex utilizando oligonucleótidos para C. trachomatis (gen ompA, orf6 plasmídico), Mycoplasma/Ureaplasma y G. vaginalis (genes rRNA16s). Resultados: Se estandarizaron pruebas de PCR múltiplex para los microorganismos estudiados, optimizándose las concentraciones y condiciones de las reacciones múltiplex. Se obtuvieron PCR dúplex para C. trachomatis (ompA, orf6), Chlamydia/Gardnerella y Chlamydia/micoplasmas y tríplex para Chlamydia/Mycoplasma/Ureaplasma. También un cuádruplex para Chlamydia/Mycoplasma/Ureaplasma/Gardnerella. Los resultados fueron verificados por PCR e hibridación automática (HybriSpot 12) y análisis in silico. Conclusión: Se desarrollaron pruebas de PCR múltiplex con una alta sensibilidad y especificidad para la identificación de C. trachomatis, micoplasmas y G. vaginalis.

Abstract Background: Sexually transmitted infections are a global public health problem. Routine analysis includes microbiological and serological tests for the diagnosis of pathogens. Atypical microorganisms such as Chlamydia trachomatis and mycoplasmas are not determined due to the requirements for their identification. Furthermore, Gardnerella vaginalis is not included despite being associated with bacterial vaginosis. Objective: To develop a multiplex PCR to diagnose Chlamydia, mycoplasmas, and Gardnerella. Method: Standardization of multiplex PCR tests was carried out using oligonucleotides for the identification of Chlamydia (ompA gene, plasmid orf6), Mycoplasma/Ureaplasma and Gardnerella (rRNA16s genes). Results: Multiplex PCR tests were standardized for the microorganisms studied, optimizing the concentrations and conditions of the multiplex reactions. Duplex PCR was obtained for Chlamydia (ompA, orf6), Chlamydia/Gardnerella, and Chlamydia/mycoplasmas, and triplex PCR for Chlamydia/mycoplasmas. Also, a quadruplex for Chlamydia, Mycoplasma/Ureaplasma and Gardnerella. PCR and automatic hybridization verified the results obtained (HybriSpot 12) and in silico analysis. Conclusion: Multiplex PCR tests with high sensitivity and specificity were developed to identify C. trachomatis, mycoplasmas, and G. vaginalis.

Isolation of Mycoplasma salivarium in the empyema of a diabetic patient with deep neck infection and necrotizing mediastinitis: A case report.

Mycoplasma species (spp.) are predominantly found in the human oropharynx, and extracavity infections are rare. Conventional culture limitations hinder Mycoplasma spp. recovery, potentially causing overlooked infections. Molecular techniques reveal their roles in various infections. Mycoplasma pneumoniae causes pneumonia, while Mycoplasma salivarium (M. salivarium) in empyema is scarcely reported. We present a case of a 61-year-old man who suffered from tonsillitis, deep neck infection, necrotizing mediastinitis, and bilateral pleural infections. Mixed pathogens, mainly M. salivarium, were implicated.

Combining Fusion of Cells with CRISPR-Cas9 Editing for the Cloning of Large DNA Fragments or Complete Bacterial Genomes in Yeast.

The genetic engineering of genome fragments larger than 100 kbp is challenging and requires both specific methods and cloning hosts. The yeast Saccharomyces cerevisiae is considered as a host of choice for cloning and engineering whole or partial genomes from viruses, bacteria, and algae. Several methods are now available to perform these manipulations, each with its own limitations. In order to extend the range of yeast cloning strategies, a new approach combining two already described methods, Fusion cloning and CReasPy-Cloning, was developed. The CReasPy-Fusion method allows the simultaneous cloning and engineering of megabase-sized genomes in yeast by the fusion of bacterial cells with yeast spheroplasts carrying the CRISPR-Cas9 system. With this new approach, we demonstrate the feasibility of cloning and editing whole genomes from several Mycoplasma species belonging to different phylogenetic groups. We also show that CReasPy-Fusion allows the capture of large genome fragments with high efficacy, resulting in the successful cloning of selected loci in yeast. We finally identify bacterial nuclease encoding genes as barriers for CReasPy-Fusion by showing that their removal from the donor genome improves the cloning efficacy.

Incidence and Prevalence of Vaginal Infections in Women of Reproductive Age in North Macedonia.

In the available literature on this subject there are many studies which describe the effects of sexually transmitted infections on pregnancy and fertility of women. Because of the frequency of the infections with the atypical bacteria of the Ureaplasma Spp., Mycoplasma Spp., Chlamydia Trachomatis, as well as HPV infections in women of reproductive age, it is easy to underestimate their importance when establishing the basis of the genital health of women of reproductive age. In this prospective analysis, conducted from 2014 to 2018 in the laboratory for HPV and Molecular diagnostics at the University Clinic of Gynaecology and Obstetrics in Skopje, North Macedonia, we analysed the results of 10,387 patients of all ages, of which 973 patients were of reproductive age. A Panel analysis was also conducted (including the above-mentioned pathogens). An HPV analysis was also conducted on 643 patients in this group. Within the group of 643 patients, there was a positive result for HPV in 26.7% of them, while in 40.9% there was a positive result for one or more pathogens on the Panel analysis of bacterial pathogens. The statistical analysis of the results showed that the most frequent of all bacterial pathogens within the Macedonian population of women of reproductive age is Ureaplasma Spp, with an incidence of 33%, followed by Mycoplasma Spp., with 7.8%, while Chlamydia Trachomatis was present in 6.4% of the cases. We should highlight that a co-infection with HPV was present in 18.5% of all the patients where there was analysis of both diagnostic procedures. The analysis of the results in the patients co-infected with HPV and at least one bacterial pathogen on the Panel Analysis, showed a very high statistical correlation (p<001).

Molecular Survey of Hemotropic Mycoplasma spp. and Bartonella spp. in Coatis (Nasua nasua) from Central-Western Brazil.

Even though previous works showed molecular evidence of hemotropic Mycoplasma spp. (hemoplasmas) in ring-tailed coatis (Nasua nasua) from Brazil, Bartonella sp. has not been reported in these mammals so far. The present study aimed to detect the above-mentioned agents in coatis' blood and associated ectoparasites, assessing the association between these infections and red blood parameters. Between March 2018 and January 2019, coati (n = 97) blood samples, Amblyomma sp. ticks (2242 individual ticks, resulting in 265 pools), and Neotrichodectes pallidus louse (n = 59) were collected in forested urban areas from midwestern Brazil. DNA extracted from coatis' blood, and ectoparasite samples were submitted to quantitative PCR (qPCR) (16S rRNA) and conventional PCR (cPCR) (16S rRNA and 23S rRNA) for hemoplasmas and qPCR (nuoG gene) and culturing (only blood) for Bartonella spp. Two different hemoplasma genotypes were detected in blood samples: 71% coatis positive for myc1 and 17% positive for myc2. While 10% of ticks were positive for hemoplasmas (myc1), no louse was positive. The estimated bacterial load of hemoplasmas showed no association with anemia indicators. All coatis were negative for Bartonella sp. in qPCR assay and culturing, albeit two Amblyomma sp. larvae pools, and 2 A. dubitatum nymph pools were positive in the qPCR. The present work showed a high occurrence of hemoplasmas, with two distinct hemoplasma genotypes, in coatis from forested urban areas in midwestern Brazil.

Detection of Mycoplasma spp. in horses with respiratory disorders.

BACKGROUND: Bacteria belonging to the genus Mycoplasma are small-sized, have no cell walls and small genomes. They commonly cause respiratory disorders in their animal hosts. Three species have been found in the respiratory tract of horses worldwide, that is., Mycoplasma (M.) equirhinis, M. pulmonis and M. felis, but their role in clinical cases remains unclear. OBJECTIVES: The aim of this study was to i) develop and validate tools to detect, isolate and identify different Mycoplasma spp. strains in clinical equine respiratory-tract specimens and ii) subsequently define the prevalence of the three species in France depending on sample types and horse characteristics (age, breed, sex). STUDY DESIGN: Validation of a workflow for mycoplasma diagnosis and subsequent prevalence study. METHODS: Mycoplasma-free tracheal wash samples spiked with numerated strains and DNA dilutions were used to validate the culture methods and real-time PCR (rt-PCR) assay. Isolated strains were identified by 16S rRNA gene sequencing. Prevalences were determined on a population of 616 horses with respiratory disorders, sampled in France in 2020. RESULTS: In total, 104 horses (16.9%) were found to be positive for Mycoplasma spp. by at least one method. M. equirhinis was the predominant circulating species, accounting for 85% of the rt-PCR-positive samples and 98% of the 40 cultured strains. MAIN LIMITATION: The proposed pre-enrichment procedure improves the sensitivity of detection but hinders the quantification of the initial mycoplasma load in the clinical specimens. CONCLUSIONS: Prevalence of mycoplasma varied with age, breed, and type of sample.

Comparable outcomes from long and short read random sequencing of total RNA for detection of pathogens in chicken respiratory samples.

Co-infections of avian species with different RNA viruses and pathogenic bacteria are often misdiagnosed or incompletely characterized using targeted diagnostic methods, which could affect the accurate management of clinical disease. A non-targeted sequencing approach with rapid and precise characterization of pathogens should help respiratory disease management by providing a comprehensive view of the causes of disease. Long-read portable sequencers have significant potential advantages over established short-read sequencers due to portability, speed, and lower cost. The applicability of short reads random sequencing for direct detection of pathogens in clinical poultry samples has been previously demonstrated. Here we demonstrate the feasibility of long read random sequencing approaches to identify disease agents in clinical samples. Experimental oropharyngeal swab samples (n = 12) from chickens infected with infectious bronchitis virus (IBV), avian influenza virus (AIV) and Mycoplasma synoviae (MS) and field-collected clinical oropharyngeal swab samples (n = 11) from Kenyan live bird markets previously testing positive for Newcastle disease virus (NDV) were randomly sequenced on the MinION platform and results validated by comparing to real time PCR and short read random sequencing in the Illumina MiSeq platform. In the swabs from experimental infections, each of three agents in every RT-qPCR-positive sample (Ct range 19-34) was detectable within 1 h on the MinION platform, except for AIV one agent in one sample (Ct = 36.21). Nine of 12 IBV-positive samples were assigned genotypes within 1 h, as were five of 11 AIV-positive samples. MinION relative abundances of the test agent (AIV, IBV and MS) were highly correlated with RT-qPCR Ct values (R range-0.82 to-0.98). In field-collected clinical swab samples, NDV (Ct range 12-37) was detected in all eleven samples within 1 h of MinION sequencing, with 10 of 11 samples accurately genotyped within 1 h. All NDV-positive field samples were found to be co-infected with one or more additional respiratory agents. These results demonstrate that MinION sequencing can provide rapid, and sensitive non-targeted detection and genetic characterization of co-existing respiratory pathogens in clinical samples with similar performance to the Illumina MiSeq.

First Report of 'Candidatus Mycoplasma haematomacacae' in Laboratory-Kept Rhesus Monkeys (Macaca mulatta) Maintained in Rio de Janeiro, Brazil.

Health monitoring programs in animals used as experimental models are essential, since only disease-free subjects are considered suitable for research purposes. In laboratory-kept animals, hemoplasmas have been described as an important confounding variable. Different hemoplasma species have been detected infecting non-human primates (NHP) from Brazil. However, the occurrence of hemoplasma species in laboratory-kept NHP in Brazil has not-yet been assessed. Accordingly, this study aimed (i) to screen laboratory-kept rhesus monkeys for hemoplasmas, (ii) to verify if any of the hemoplasma-positive animals demonstrate hematological abnormalities, and (iii) to assess the genotype diversity of hemoplasma species in NHP from Brazil. Five out of eight (62.5%; 95% CI: 3.05-8.63) rhesus monkeys tested positive for hemotropic Mycoplasma spp. by PCR. Sequencing, phylogenetic, distance, and genotype diversity analyses of partial 16S rRNA gene demonstrate that rhesus monkeys were infected by 'Candidatus Mycoplasma haematomacacae' (formerly 'Candidatus Mycoplasma haemomacaque'). Assessments of partial 16S rRNA diversity of hemoplasma species in NHP suggest that at least four genetically diverse groups may occur in Brazil. Although no hematological abnormalities were demonstrated in rhesus monkeys evaluated herein, future studies are needed to elucidate the influence of 'Ca. M. haematomacacae' as a confounding variable on research studies.

Case Report: Hyperammonemic Encephalopathy Linked to Ureaplasma spp. and/or Mycoplasma hominis Systemic Infection in Patients Treated for Leukemia, an Emergency Not to Be Missed.

Background: Hyperammonemic encephalopathy caused by Ureaplasma spp. and Mycoplasma hominis infection has been reported in immunocompromised patients undergoing lung transplant, but data are scarce in patients with hematological malignancies. Case Presentation: We describe the cases of 3 female patients aged 11-16 years old, developing initially mild neurologic symptoms, rapidly evolving to coma and associated with very high ammonia levels, while undergoing intensive treatment for acute leukemia (chemotherapy: 2 and hematopoietic stem cell transplant: 1). Brain imaging displayed cerebral edema and/or microbleeding. Electroencephalograms showed diffuse slowing patterns. One patient had moderate renal failure. Extensive liver and metabolic functions were all normal. Ureaplasma spp. and M. hominis were detected by PCR and specific culture in two patients, resulting in prompt initiation of combined antibiotics therapy by fluoroquinolones and macrolides. For these 2 patients, the improvement of the neurological status and ammonia levels were observed within 96 h, without any long-term sequelae. M. hominis was detected post-mortem in vagina, using 16S rRNA PCR for the third patient who died of cerebral edema. Conclusion: Hyperammonemic encephalopathy linked to Ureaplasma spp. and M. hominis is a rare complication encountered in immunocompromised patients treated for acute leukemia, which can lead to death if unrecognized. Combining our experience with the few published cases (n=4), we observed a strong trend among female patients and very high levels of ammonia, consistently uncontrolled by classical measures (ammonia-scavenging agents and/or continuous kidney replacement therapy). The reversibility of the encephalopathy without sequelae is possible with prompt diagnosis and adequate combined specific antibiotherapy. Any neurological symptoms in an immunocompromised host should lead to the measurement of ammonia levels. If increased, and in the absence of an obvious cause, it should prompt to perform a search for Ureaplasma spp. and M. hominis by PCR as well as an immediate empirical initiation of combined specific antibiotherapy.

Exploiting 16S rRNA-based metagenomics to reveal neglected microorganisms associated with infertility in breeding bulls in Spanish extensive herds.

Bovine infectious infertility represents a problem due to the high impact on animal production and, in many cases, in public health. A lack of information on the characteristics of the bacterial population of the bovine reproductive system can hamper a comprehensive understanding of reproductive pathologies and the role that the microbiome could play. A metagenomic study based on the V3-V4 hypervariable region of the bacterial 16S rRNA gene was performed in 1029 preputial samples from bulls raised in an extensive regimen in Spain (944 from herds with low fertility rates -case group-, and 85 samples from reproductively healthy herds -control group-). The most representative phyla as well as the most 10 abundant bacterial families and their abundance did not show significant differences in both case and control groups. Similarly, the (alpha and beta) diversity of the bacterial populations was similar in both type of herds: the Shannon and Simpson indices show a high diversity of species, while the Bray-Curtis dissimilarity index did not show relevant differences in the bacterial communities. A deeper analysis of the operational taxonomic units showed the presence of one genera, Mycoplasma spp. significantly associated with fertility problems. Our study highlights the promising potential that the application of sequencing techniques (e.g. 16S rRNA-based metagenomics) possesses in examining bovine infertility, as they are able to reveal different pathogens that could go unnoticed using diagnostic approaches for only the main known pathogens.

Detection by multiplex PCR of Mycoplasma species associated with dairy cattle in Argentina / Detección por PCR multiplex de especies de Mycoplasma asociadas a ganado lecheroen Argentina

Abstract There is scarce information about the frequency and epidemiological and clinicalfeatures associated with the presence of Mycoplasma spp. in Argentine dairy herds. The objec-tives of this study were to develop a multiplex PCR for identifying M. bovis and M. canadenseand to describe the frequency of Mycoplasma spp. isolated from clinical samples submitted to adiagnostic laboratory. Of a total of 1548 samples from intramammary infections, bulk tank milkand biological fluids, 38 Mycoplasma isolates were obtained. M. bovis, M. canadense, M. cali-fornicum and M. leachii were detected by using two multiplex PCRs, confirming their presencein clinical conditions in dairy cattle. The techniques used in the present study can be usefulto broaden the knowledge about Mycoplasma infections in cattle, since the search for theseorganisms is not usually included in routine diagnoses.

Resumen Existe poca información sobre la frecuencia, así como las características epidemi-ológicas y clínicas asociadas con la presencia de Mycoplasma en los rodeos lecheros argentinos.Los objetivos de este estudio fueron desarrollar una PCR multiplex para identificar M. bovis yM. canadense y describir la frecuencia de especies de Mycoplasma aisladas de muestras clíni-cas enviadas a un laboratorio de diagnóstico. De un total de 1.548 muestras de infeccionesintramamarias, leche de tanque de frío y fluidos biológicos, se obtuvieron 38 aislamientos de Mycoplasma. Mediante 2 PCR multiplex se detectaron M. bovis, M. canadense, M. californicumy M. leachii, confirmando su presencia en síndromes clínicos en ganado lechero. Las técnicasutilizadas en el presente estudio pueden ser útiles para ampliar el conocimiento sobre las infec-ciones por Mycoplasma en bovinos, ya que la búsqueda de estos organismos no suele incluirseen los diagnósticos de rutina.

Evidence of Mycoplasma spp. transmission by migratory wild geese.

Mycoplasma infections have been found in different species of waterfowl worldwide. However, the question of how the pathogens have been transmitted and dispersed is still poorly understood. Samples collected from clinically healthy greater white-fronted geese (Anser albifrons) (N = 12), graylag geese (Anser anser) (N = 6), taiga bean geese (Anser fabalis) (N = 10), and barnacle geese (Branta leucopsis) (N = 1) were tested for Mycoplasma spp. All Mycoplasma-positive samples were specified by species-specific PCR for Mycoplasma anserisalpingitidis (formerly known as Mycoplasma sp. 1220), M. anseris, M. anatis, and M. cloacale. The presence of Mycoplasma spp. was confirmed in 22 of 29 sampled birds (75.9%). Mycoplasma anserisalpingitidis was the most frequently detected species (15 of 22, 68.2%). However, we did not detect any of the other Mycoplasma spp. typical for geese, among which are M. anatis, M. anseris, and M. cloacale. Phylogenetic analysis revealed that Polish sequences of M. anserisalpingitidis formed a distinct branch, along with 2 Hungarian isolates obtained from domestic geese. Eight of the samples identified as Mycoplasma spp.-positive were negative for the aforementioned Mycoplasma species. A phylogenetic tree constructed based on partial 16S rRNA gene analysis showed that Mycoplasma spp. sequences collected from Polish wild geese represent a distinct phylogenetic group with Mycoplasma sp. strain 2445 isolated from a domestic goose from Austria. The results of our study showed that wild geese could be a reservoir and vector of different species of the Mycoplasma genus that can cause significant economic losses in the domestic goose industry.

Hemotropic mycoplasmas (hemoplasmas) in wild boars, hunting dogs, and hunters from two Brazilian regions.

Haemotropic mycoplasmas (haemoplasmas) are small pleomorphic bacteria infecting erythrocytes of several mammalian species, including human beings. No study to date has focused on the risk of bacteria exposure in hunting activities, particularly in natural environments of highly tick-infested areas. Accordingly, the present study aimed to assess haemoplasma occurrence in the complex encompassing wild boars, hunting dogs and hunters of Brazil. A total of 38/65 (58.5%) wild boars and 94/159 (59.1%) dogs were positive by qPCR for at least one haemoplasma. All 25 hunters were negative. Dogs with high hunting frequency were 2.4 more likely to be infected. Sequencing revealed a probable novel haemoplasma species in wild boars. Although exposure to haemoplasma species was present, the study herein found no evidence of cross-species transmission.

Detection by multiplex PCR of Mycoplasma species associated with dairy cattle in Argentina.

There is scarce information about the frequency and epidemiological and clinical features associated with the presence of Mycoplasma spp. in Argentine dairy herds. The objectives of this study were to develop a multiplex PCR for identifying M.bovis and M.canadense and to describe the frequency of Mycoplasma spp. isolated from clinical samples submitted to a diagnostic laboratory. Of a total of 1548 samples from intramammary infections, bulk tank milk and biological fluids, 38 Mycoplasma isolates were obtained. M. bovis, M. canadense, M.californicum and M.leachii were detected by using two multiplex PCRs, confirming their presence in clinical conditions in dairy cattle. The techniques used in the present study can be useful to broaden the knowledge about Mycoplasma infections in cattle, since the search for these organisms is not usually included in routine diagnoses.

Prevalence of Vector-Borne Pathogens in Reproductive and Non-Reproductive Tissue Samples from Free-Roaming Domestic Cats in the South Atlantic USA.

Reservoir to multiple species of zoonotic pathogens, free-roaming cats (FRCs) interact with domestic and wild animals, vectors, and humans. To assess the potential for feline vector-borne pathogens to be vertically transmitted, this study surveyed ear tip and reproductive tissues of FRCs from two locations in the South Atlantic United States for Anaplasma, Bartonella, Ehrlichia, hemotropic Mycoplasma, and Rickettsia species. We collected ovary (n = 72), uterus (n = 54), testicle (n = 74), and ear tip (n = 73) tissue from 73 cats, and fetal (n = 20) and placental (n = 19) tissue from 11 queens. Pathogen DNA was amplified utilizing qPCR, confirmed by sequencing. Cats were more frequently Bartonella henselae positive on reproductive tissues (19%, 14/73) than ear tip (5%, 4/73; p = 0.02). B. henselae was amplified from fetus (20%, 4/20) and placenta samples (11%, 2/19). Bartonella spp. infection was more common in cats from North Carolina (76%, 26/34) than Virginia (13%, 5/39; p < 0.0001). Fourteen percent (10/73) of both ear tip and reproductive tissues were positive for hemotropic Mycoplasma spp. Anaplasma, Ehrlichia, and Rickettsia spp. DNA was not amplified from any cat/tissue. These findings suggest that B. henselae preferentially infected cats' reproductive tissue and reinforces the importance of investigating the potential for B. henselae vertical transmission or induction of reproductive failure.

Molecular survey and genetic characterization of 'Candidatus Mycoplasma haemolamae' in llamas (Lama glama) and alpacas (Vicugna pacos) from Southern Chile.

This study aimed to perform a molecular survey and identification of hemotropic Mycoplasma spp. in domestic South American Camelids from Southern Chile. Conventional PCR (cPCR) for hemotropic Mycoplasma spp. based on 16S rRNA gene (620bp fragment) was performed in 87 EDTA-blood samples taken from 48 llamas (Lama glama) and 39 and alpacas (Vicugna pacos) from to Temuco, La Araucanía region and Valdivia, Los Rios region, Southern Chile. 16S rRNA hemotropic Mycoplasma PCR-positive were sequenced for species identification, phylogenetic and haplotype analyses, and further tested by cPCR targeting a fragment (160-210 bp) of the RNaseP (rnpB) gene. Based upon 16S rRNA cPCR results, the overall hemotropic Mycoplasma spp. occurrence in Southern camelids was 9.2% (8/87 [95% CI (4.0-17.3%)]), with five positive alpacas (12.8%; 5/39 [95% CI (4.3-27.4%)]) and three llamas (6.3%; 3/48 [95% CI (1.7-17.2%)]). All 16S rRNA PCR-positive samples were negative for the rnpB gene. Obtained 16S sequences presented high identity (99-100%) by BLASTn analysis to 'Candidatus Mycoplasma haemolamae' from an alpaca in the United Kingdom. Phylogenetic and haplotype analyses of the 16s rRNA gene showed high similarity among 'Candidatus M. haemolamae' sequences of this study and the ones from North America, Europe, and Asia evidencing a low diversity of Chilean samples, with only one haplotype detected (#1). Haplotype #1 from South American Camelids in Chile was worldwide distributed and observed in North America, Europe, and Asia. 'Candidatus M. haemolamae' detected for the first time in South American camelids in Southern Chile had low diversity and was worldwide spread.

[Clinical implications of the genus Mycoplasma]. / Implicaciones clínicas de las especies del género Mycoplasma.

Within Mycoplasma genus, M. pneumoniae, M. genitalium, M. hominis or U. urealyticum are the main species that have been traditionally linked to infectious processes. However, there are many other species involved in these conditions and that are, frequently, unfamiliar to healthcare professionals. The aim of this review is to identify all Mycoplasma genus species that have been isolated in human beings and to determine their involvement in infectious pathology.

Occurrence of Mycoplasmas in Galapagos Sea Lions (Zalophus wollebaeki) and their Association with Other Respiratory Pathogens.

During the 2018 breeding season, an outbreak of respiratory disease occurred among Galapagos sea lions (Zalophus wollebaeki) that inhabit rookeries near urban areas with introduced fauna such as dogs and cats. Several sea lions had nasal discharge and respiratory distress and were in poor body condition. Eighteen sea lions were captured for a general health assessment including collection of blood for serology and nasal discharge for culture and PCR. Samples were analyzed for 15 respiratory pathogens known to infect cats, dogs, and marine mammals. There was no evidence for interspecies pathogen transmission between Galapagos sea lions and domestic animals. Several bacterial pathogens associated with respiratory tract infection in the California sea lion (Zalophus californianus) were isolated. Mycoplasma spp. were identified by PCR in nasal discharge samples but were not the species commonly found in cats and dogs.

Mycoplasma species isolated from bovine milk collected from US dairy herds between 2016 and 2019.

Determining the species of mycoplasma isolated from culture-positive milk samples is important for understanding the clinical significance of their detection. Between August 2016 and December 2019, 214,518 milk samples from 2,757 dairy herds were submitted to Quality Milk Production Services (QMPS) at Cornell University for mycoplasma culture. From these samples, 3,728 collected from 204 herds were culture positive. Based on the request of herd managers, owners, or veterinarians, 889 isolates from 98 herds were subjected to molecular identification by PCR and amplicon sequencing. The largest proportion of the identified isolates were from New York State (78.1%), while the others came from the eastern United States (17.8%), Texas (2.0%), and New Mexico (2.1%). As expected, Mycoplasma spp. were the most common (855 isolates, 96.2%) and Acholeplasma spp. accounted for the remainder (34 isolates, 3.8%). Mycoplasma bovis was the most prevalent Mycoplasma species (75.1%), followed by M. bovigenitalium (6.5%), M. canadense (5.9%), M. alkalescens (5%), M. arginini (1.7%), M. californicum (0.1%), and M. primatum (0.1%). A portion of the isolates were confirmed as Mycoplasma spp. other than M. bovis but were not identified at the species level (16 isolates, 1.8%) because further information was not requested by the manager, owner, or veterinarian. Mycoplasma bovis was the only species identified in 59 of the 98 herds. However, more than 1 Mycoplasma sp. was identified in 29 herds, suggesting that herd infection with 2 or more mycoplasmas is not uncommon. Moreover, a Mycoplasma sp. other than M. bovis was the only species identified in 8 herds. From the subset of 889 mycoplasma culture-positive isolates from 98 herds, we determined that over a third of the herds had either more than 1 Mycoplasma sp. or a Mycoplasma sp. other than M. bovis detected in their milk samples. In conclusion, we observed that M. bovis is the most common pathogenic Mycoplasma species found in mastitic milk, but other Mycoplasma species are not uncommon. Our results suggest that it is critical to test milk samples for mycoplasmas using diagnostic tests able to identify both the genus and the species.