Detection Of Mycoplasma Research Articles

Molecular Detection and Characterization of Mycoplasma spp. in Marine Mammals, Brazil.

Mycoplasma spp. are wall-less bacteria able to infect mammals and are classified as hemotropic (hemoplasma) and nonhemotropic. In aquatic mammals, hemoplasma have been reported in California sea lions (Zalophus californianus) and river dolphins (Inia spp.). We investigated Mycoplasma spp. in blood samples of West Indian manatees (Trichechus manatus), pinnipeds (5 species), and marine cetaceans (18 species) that stranded or were undergoing rehabilitation in Brazil during 2002-2022. We detected Mycoplasma in blood of 18/130 (14.8%) cetaceans and 3/18 (16.6%) pinnipeds. All tested manatees were PCR-negative for Mycoplasma. Our findings indicate that >2 different hemoplasma species are circulating in cetaceans. The sequences from pinnipeds were similar to previously described sequences. We also detected a nonhemotropic Mycoplasma in 2 Franciscana dolphins (Pontoporia blainvillei) that might be associated with microscopic lesions. Because certain hemoplasmas can cause disease and death in immunosuppressed mammals, the bacteria could have conservation implications for already endangered aquatic mammals.

Detection of Mycoplasma pneumoniae in hospitalized children with pneumonia in Laos

Mycoplasma pneumoniae has been described worldwide as an important cause of community-acquired pneumonia. From December 2013 to December 2014, 461 children admitted to Mahosot Hospital, Vientiane, Laos, with acute respiratory infection were investigated for upper respiratory microorganisms using probe-based real-time polymerase chain reaction (PCR) (FTD33). M. pneumoniae was detected by FTD33 in the upper respiratory tract of three patients, two girls and one boy, 5.7 and 3.9 years old and 13.6 years old, respectively. They presented with clinical features compatible with M. pneumoniae infection. The two girls were also positive for other potential pathogens. The boy had abnormal pulmonary auscultation, and one of the girls had significant anaemia. These results suggest that enhancement of diagnostic systems for M. pneumoniae detection and analysis of its antibiotic resistance profile is needed to raise awareness and improve understanding of the epidemiology of M. pneumoniae infection in Laos, enable targeted therapy, and inform treatment guidelines.

Detection of Microbiological Contamination in Cell Cultures and their Raw Materials: a 6 years report of this activity.

Contaminations in cell cultures are a source of concern for biotechnology laboratories, bio products manufacturers and cell banks as they involve economical loss, time loss and non-valid results. The main issues are fungi, bacteria (including mycoplasmas) and viruses. In this study, we present a retrospective of cell cultures quality control and their raw materials carried out at INMETRO throughout 6 years. From 2014 to 2019, different cell lines and raw materials derived from LABIO, from partner institutions of research projects, or from cooperation agreements were assessed. The detection of mycoplasmas were made using the biochemical method MycoAlert Plus. Contamination by bacteria and fungi were detected by a microbiological culturing according to the Brazilian Pharmacopoeia. Among the 944 assays carried out to detect bacteria and fungi; 5.6% presented contamination. Among 1143 assays of mycoplasmas detection; 10.8% presented contamination. Regarding raw materials assessment, 0.7% and 7.7% were contaminated with mycoplasmas and bacteria/fungi, respectively. In total, 8% of all samples presented contamination: 5.6% by mycoplasmas, 2.4% by bacteria/fungi and 0.1% by both. The dissemination of Good Practices on Cells Culture, the availability of these assays and the support of the implementation given to laboratories which are interested have been our contribution to improve control quality in the activity of cell cultures.

A Study on Mycoplasmal and Viral Infections in Bovine Keratoconjunctivitis

Background: Infectious bovine keratoconjunctivitis (IBK or “pink eye”) is the most common infectious ocular disease in cattle worldwide. In addition to Moraxella bovis as the principal causative agent, infectious bovine rhinotracheitis virus (BHV-1) and Mycoplasma species probably act as risk factors for IBK. Objectives: This study aimed to evaluate the association between the detection of Mycoplasma sp., bovine herpesvirus 1 (BHV-1), and bovine viral diarrhea virus (BVDV) in the conjunctival sac of the eye and IBK. Methods: Polymerase chain reaction (PCR) was employed to detect Mycoplasma sp., BHV-1, and BVDV in samples collected from IBK-affected and healthy eyes. Results: Based on the PCR results, Mycoplasma sp. was detected in 63.6% and 47.2% of IBK-affected and healthy eyes, respectively. BHV-1 was detected in 59.1% and 36.1% of affected and healthy eyes, respectively. BVDV was detected in 65.9% and 58.3% of affected and healthy eyes, respectively. BHV-1 was the only agent significantly (P<0.05) associated with IBK lesions (isolated from 59.1% of affected vs 36.1% of healthy eyes). Conclusion: Based on the study results, BHV-1 may be a risk factor in the pathogenesis of infectious bovine keratoconjunctivitis, and mechanisms other than immune depression might be involved in its pathogenicity.

Establishment and Performance Evaluation of Multiplex PCR-Dipstick DNA Chromatography for Mycoplasma pneumoniae and Chlamydia pneumoniae Rapid Detection.

Nasopharyngeal swab samples of 300 children with an acute respiratory tract infection were detected by a multiplex PCR-dipstick chromatography assay, and the results were compared with the DNA sequencing and serum IgM antibody assay. A multiplex PCR-dipstick DNA assay can specifically detect Mycoplasma pneumoniae and Chlamydia pneumoniae and shows a good specificity, with a minimum detection limit of 10 CFU/mL, respectively. Using DNA sequencing results as the gold standard, the sensitivity, specificity, positive predictive value, and negative predictive value of the multiplex PCR-dipstick DNA chromatography assay for the diagnosis of Mycoplasma pneumoniae were 96.61%, 100%, 100%, and 99.18% respectively, and those of Chlamydia pneumoniae were 95.24%, 100%, 100%, and 99.64% respectively. There was no statistical significance MP and CP diagnosis by the multiplex PCR-dipstick DNA assay and DNA sequencing (MP: P = 0.5; CP: P = 1.0), and the two assays had very high statistical consistency (MP: kappa = 0.979; CP: kappa = 0.974). The positive rate of the multiplex PCR-dipstick chromatography assay was significantly higher than that of the serum IgM antibody assay, with MP (17.7% vs. 13.3%), CP (5.7% vs. 3.3%), and mixed infection of MP and CP (1.3% vs. 0.67%). A multiplex PCR-dipstick chromatography assay was successfully established for the joint detection of Mycoplasma pneumoniae and Chlamydia pneumoniae within 2 hours. It is simple, fast, sensitive, accurate, cost-effective with good diagnostic performance, which can be used for small laboratories and point-of-care diagnosis.

Detection of Mycoplasma bovigenitalium and Mycoplasma tauri in Holstein Friesian dairy cows with subclinical endometritis

Abstract Mycoplasma spp. is the cause of serious cattle health disorders that lead to poor reproductive efficiency. Chronic seminal vesiculitis, infertility, vulvovaginitis and dystocia were observed in Mycoplasma spp. infection. The objectives of the present study were to investigate the frequency o f Mycoplasmas in the uterus of postpartum dairy cows and its potential role in the occurrence of subclinical endometritis. Our study included 102 Holstein Friesian dairy cows 22-32 days postpartum. Uterine samples for cytological, bacteriological, and molecular analysis were provided using Cytoprint A I®. Metricheck® was used for the assessment of cervicovaginal mucus. After detection of Mycoplasma spp. genome in uterine samples, the swabs of the vulva, vagina, nasal swab, and samples of milk were sampled in order to exclude possible contamination with other microorganisms as potential trigger of subclinical endometritis. The genome of Mycoplasma spp. in the uterus was confirmed in 4 cows (2.40%). Sequencing of the 16S RNA revealed that detected mycoplasmas belonged to Mycoplasma bovigenitalium and Mycoplasma tauri species, with a prevalence of 1.80% and 0.6%, respectively. The cytological evaluation showed a 69.90%, 54.87% and 48.33% of polymorphonuclear cells for M. bovigenitalium positive cows, and 61.64% for M. tauri positive cow. Bacteriological examination revealed Trueperella pyogenes in uterine samples in all 4 observed cows. This is the first study where M. tauri was detected in the uterus of a live cow with subclinical enodmetritis. In this paper we hypothesized that M. bovigenitalium and M. tauri may have a certain role in the etiology of subclinical enodmetritis.

Molecular detection of hemotropic Mycoplasma and Bartonella species in lice from sheep and goats of Mexico.

The knowledge of lice associated with small ruminants, especially sheep and goats, is scarce. In Mexico, there are historical reports of six species of chewing and sucking lice associated with Capra hircus and Ovis canadensis. However, the reports did not analyze the ecology of the infestations or the presence of potentially pathogenic bacteria. For this reason, the objectives of this study were i) to identify the species of lice associated with sheep and goats in three states of the Mexican Republic, ii) to characterize the infestations, and iii) to identify the presence of bacterial pathogens. From October 2019 to August 2021, six ranches with sheep and goats were sampled in the states of Hidalgo and Veracruz. Hosts were visually inspected, and lice were retrieved with forceps. The specimens were sexed and identified using morphological taxonomic keys. DNA extraction was performed individually, and a fragment of the cytochrome oxidase subunit 1 gene (COI) was amplified for the molecular identification of the specimens. Subsequently, Anaplasma, Bartonella, Ehrlichia, Mycoplasma, and Rickettsia were molecularly detected. Additionally, the infestations were characterized by calculating the prevalence and mean abundances. We collected 563 specimens of three species, Bovicola caprae, Bovicola ovis, and Linognathus africanus. The highest infestation levels were recorded for B. ovis (66.7%; 4.4) from Veracruz. Additionally, two Bartonella species were detected: Bartonella mellophagi in B. ovis and Bartonella capreoli in L. africanus. In contrast, Mycoplasma ovis was detected exclusively in one pool of B. ovis. This study provides new bacterial-ectoparasite associations and highlights the possible role of these neglected ectoparasites as vectors in the populations of sheep and goats from Mexico.

Development and Evaluation of a New qPCR Assay for the Detection of Mycoplasma in Cell Cultures.

In recent years, cell culture has become an important tool not only in research laboratories, but also in diagnostic and biotechnological development laboratories. Mycoplasma contamination is present in up to 35% of cell cultures used in research and in cell therapies. This fact represents a significant problem since such contamination can cause disastrous effects on eukaryotic cells by altering their cellular parameters, which, in turn, can lead to unreliable experimental results. For this reason, it is mandatory to carry out continuous testing for the presence of Mycoplasma in cell culture and the development of appropriate methodologies for this purpose. An ideal detection methodology should be fast, sensitive, and reliable. In this study, we propose an alternative detection method based on real-time PCR in conjunction with a novel combination of primers and probes that have been improved to increase their efficiency. The new PCR method demonstrates 100% sensitivity and specificity results in the detection of common Mycoplasma species that contaminate cell cultures. Whilst 11 of 45 tested supernatants were positive for Mycoplasma (24.4%) using the new PCR method (corresponding to 5 of the 14 lines tested (35.71%)), only 10 of 45 supernatants showed positive results with the commercial Venor®GeM qEP and Plasmotest® kit. In addition, the new PCR method exhibits a high capacity to detect less-frequent Mycoplasma species, such as those related to the M. mycoides cluster. The use of an alternative Mycoplasma-detection method in cell culture labs can guarantee the detection of Mycoplasma contamination, especially in cases when dubious results are recorded.

Establishment of a Mycoplasma hyorhinis challenge model in 5-week-old piglets.

Mycoplasma hyorhinis is an emerging swine pathogen with high prevalence worldwide. The main lesions caused are arthritis and polyserositis, and the clinical manifestation of the disease may result in significant economic losses due to decreased weight gain and enhanced medical costs. We aimed to compare two challenge routes to induce M. hyorhinis infection using the same clinical isolate. Five-week-old, Choice hybrid pigs were inoculated on 2 consecutive days by intravenous route (Group IV-IV) or by intravenous and intraperitoneal routes (Group IV-IP). Mock-infected animals were used as control (control group). After the challenge, the clinical signs were recorded for 28 days, after which the animals were euthanized. Gross pathological and histopathological examinations, PCR detection, isolation, and genotyping of the re-isolated Mycoplasma sp. and culture of bacteria other than Mycoplasma sp. were carried out. The ELISA test was used to detect anti-M. hyorhinis immunoglobulins in the sera of all animals. Pericarditis and polyarthritis were observed in both challenge groups; however, the serositis was more severe in Group IV-IV. Statistically significant differences were detected between the challenged groups and the control group regarding the average daily weight gain, pathological scores, and ELISA titers. Additionally, histopathological scores in Group IV-IV differed significantly from the scores in the control group. All re-isolated strains were the same or a close genetic variant of the original challenge strain. Our results indicate that both challenge routes are suitable for modeling the disease. However, due to the evoked more severe pathological lesions and the application being similar to the hypothesized natural route of infection in Group IV-IV, the two-dose intravenous challenge is recommended by the authors to induce serositis and arthritis associated with M. hyorhinis infection.

High throughput Luminex beads based multiplex assay for identification of six major bacterial pathogens of mastitis in dairy animals.

Bovine mastitis is caused by over 150 different microorganisms. Specific identification and quantification of multiple bacteria in a single milk sample becomes essential for rapid intervention. In the present study a Luminex beads based multiplex assay emphasizing on the precise identification of six major bacterial pathogens of mastitis was developed. Assay was developed in two triplex sets, triplex 1 comprised of Streptococcus agalactiae, Streptococcus dysgalactiae and Streptococcus uberis while triplex 2 consisted of Staphylococcus aureus, E. coli and Klebsiella pneumoniae. The analytical sensitivity was 10 6 copies per reaction mixture for all the six bacteria. A 100% analytical specificity was observed for simultaneous detection of these bacteria. Clinical milk samples from 100 bovine quarters were tested for validation. The analytical sensitivity was similar to the findings reported earlier in real time PCR multiplex assay targeting the DNA of the 11 most common bacterial species or groups in mastitis. The analytical specificity of the optimized assay was 100% similar to reported earlier for simultaneous detection of Mycoplasma spp. and for seven entric viruses of humans.The developed assay indicates a concept proof of a rapid, cost effective high throughput diagnostic tool for identification of major bacteria causing mastitis.

Rapid and sensitive detection of waterfowl mycoplasmas using TaqMan assays.

Waterfowl-specific mycoplasmas cause significant economic losses worldwide. However, only limited resources are available for the specific detection of three such bacteria, Mycoplasma anatis, M. anseris and M. cloacale. We developed species-specific TaqMan assays and tested their reliability across 20 strains of the respective target species as well as 84 non-target avian bacterial strains. Furthermore, we analysed 32 clinical DNA samples and compared the results with those of previously published conventional PCRs. The TaqMan assays showed 100% specificity and very high sensitivity, enabling the detection of target DNA as low as either 10 or 100 copies/μl concentration, depending on the assay. Importantly, we found that while the here developed TaqMan assays are reliable for species-specific detection of M. anatis, the previously published conventional PCR assay may give false positive results. In conclusion, the new assays are reliable, sensitive and suitable for clinical diagnostics of the target species.

Regeneration of T cells from human-induced pluripotent stem cells for CAR-T cell medicated immunotherapy.

Background: Chimeric antigen receptor (CAR) T cell treatment involves in vitro production of T cells from patient blood with synthetic receptors specific to a cancer antigen. They circumvent the major histocompatibility complex to recognize the tumor antigen, reducing hematologic malignancy remission rates by 80%. Considering the efficacy of CAR-T treatment, the present work aimed at generating functional clusters of differentiation (CD)8 + T cells from human induced pluripotent stem cells (hiPSC) and to generate hiPS-CAR-T cells with high antigen-specific cytotoxicity. Methods: The Alkaline phosphatase assay and MycoEasy rapid mycoplasma detection kit was implemented for detection of hiPSCs and mycoplasma, respectively. The CD34+ HSPCs were harvested in AggreWellTM 400 using a 37-micron reversible strainer. Likewise, the lymphoid progenitor and CD4+CD8+ DP T cells were also harvested. The Cell Counting Kit-8 (CCK-8) assay was used to mark cytotoxicity and ELISA was used to detect IFN-γ secretion. Further, flow cytometry and transwell chambers were used to assess cell cycle, and migration and invasion. Finally, the in vivo antitumor effects of the CAR-T cells were evaluated using experimental animals (mice). Results: Results revealed that a serum-free, feeder layer-free differentiation system significantly yielded hiPSC-based T cell immunotherapy with interleukin-2, interleukin-15, and activators at the differentiation stage to promote the maturation of these cells into human induced pluripotent stem (hiPS)-T cells. The infection of hiPSCs with the CD19 CAR lentivirus resulted in the production of the hiPSC-CAR-T cells. We validated the function of hiPS-CAR-T cells in vivo and in vitro experimentation which revealed no significant differences in cell morphology and function between hiPSC-derived hiPS-CAR-T cells and peripheral blood-derived CAR-T cells. Conclusion: This study developed a culture method that is efficient and clinically useful to make functional CD8+ T cells from hiPSC and to get hiPS-CAR-T cells with high antigen-specific cytotoxicity that are not very different from CAR T cells found in peripheral blood. As a result, our findings may open the way for the clinical use of hiPSC to create functional CD8+ T and hiPS-CAR-T cells cells for use in cell-based cancer therapy.

Significance of the determination of DNA load of drug-resistant mycoplasma pneumoniae and 23sRNA gene mutation locus in children

Purpose: To determine the significance of the detection of Mycoplasma pneumoniae (MP)-DNA load and 23sRNA gene mutation locus in children with drug-resistant MP pneumonia. 
 Methods: A total of 158 children with MP pneumonia received drug sensitivity tests. The patients were divided into resistance group and non-resistance group. The MP-DNA load index (MPLI) and mutation rate of 23sRNA gene at 2063 locus were assessed and compared between the two groups: the MPLI-negative group and the MPLI-positive group, based on whether MPLI was greater than 6.12. The association of MPLI of all the patients and the 23sRNA gene mutation at 2063 locus in the resistance group, as well as clinical indicators were analyzed. 
 Results: The MPLI of the resistance group was lower than that of the non-resistance group. In the MPLI-positive group, the duration of disease, defervescence time, disappearance time of cough and expectoration, disappearance time of chest opacity, and length of stay were all longer than those of the MPLI-negative group, while the proportion of cases with extrapulmonary complications and the white blood cell (WBC) count were higher than those of the MPLI-negative group. The mutation rate of 23sRNA gene at 2063 locus in the resistance group was higher than that in non-resistance group (p < 0.05). The defervescence time, disappearance time of cough and expectoration, disappearance time of chest opacity and length of stay were longer in the mutation-positive group than those in the mutation-negative group (p < 0.05). 
 Conclusion: The mutation rate of 23sRNA gene at 2063 locus is higher in children with drug-resistant MP pneumonia. Furthermore, low MPLI and 23sRNA gene mutations at 2063 locus are associated with the duration of disease, disappearance time of clinical symptoms and other clinical indicators.

CRISPR/Cas9-based point-of-care lateral flow biosensor with improved performance for rapid and robust detection of Mycoplasma pneumonia

Screening of acute respiratory infections causes serious challenges in urgent point-of-care scenarios where conventional methods are impractical and alternative techniques suffer from low accuracy, poor robustness, and reliance on sophisticated instruments. As an improvement to this paradigm, we report a point-of-care lateral flow biosensor (LFB) based on the recognition property of clustered regularly interspaced short palindromic repeats (CRISPR)/associated protein 9 (Cas9) and apply it to the detection of Mycoplasma pneumoniae (M. pneumoniae). The designed biosensor employs CRISPR/Cas9 for secondary recognition after preamplification of target gene using specific primer set, avoiding false positives caused by nontarget factors. The high amplification efficiency and low applicable temperatures of recombinase polymerase amplification brings the detection limit of the biosensor to 3 copies even at a preamplification temperature of 25 °C. Its practical application is further demonstrated with 100% accuracy by testing with 43 M. pneumoniae-infected specimens and 80 uninfected specimens. Additionally, the entire detection, including pretreatment, preamplification, CRISPR/Cas9 recognition, and visual analysis, can be completed in 30 min. Featured with the combination of CRISPR/Cas9 and LFB, the biosensor we developed herein ensures excellent convenience, accuracy, and robustness, which endows promising point-of-care screening potential for infectious pathogens.

Molecular detection of Mycoplasma spp. from camel’s milk

The dromedary camels are essential because of their multiple uses for transport, wool, meat, and milk production. Camel’s milk is considered exceptional nutrition, so it uses for therapeutic and high nutrition. Mastitis may affect camel’s milk production, cause significant economic loss, and is associated with zoonotic disease. The study aimed to detect the prevalence of Mycoplasma spp. especially M. bovis in camel’s milk using the polymerase chain reaction (PCR) method as a primary molecular diagnostic technique. Fifty milk samples were collected from she camels suffering from subclinical mastitis in Iraq. The result of the current study declared that 26.66% of the camel’s milk samples were positive for Mycoplasma. In contrast, the prevalence rate of Mycoplasma (M.) bovis in all the samples was 61.53%. The study concludes that Mycoplasma and especially M. bovis are considered one of the bacteria that cause subclinical mastitis in camels, and using the PCR method is regarded as a more rapid, simple, and current for detecting the Mycoplasma bacteria.

The association of host and vector characteristics with Ctenocephalides felis pathogen and endosymbiont infection.

Surveillance of the fleas and flea-borne pathogens infecting cats is important for both human and animal health. Multiple zoonotic Bartonella and Rickettsia species are known to infect the most common flea infesting cats and dogs worldwide: Ctenocephalides felis, the cat flea. The ability of other flea species to transmit pathogens is relatively unexplored. We aimed to determine cat host and flea factors independently associated with flea Bartonella and Rickettsia infection. We also assessed flea and cat infection by flea-host pair and location. To accomplish these aims, we performed qPCR for the detection of Bartonella, hemotropic Mycoplasma, Rickettsia, and Wolbachia DNA using paired cat and flea samples obtained from free-roaming cats presenting for spay or neuter across four locations in the United States. A logistic regression model was employed to identify the effect of cat (sex, body weight, geographic location, and Bartonella, hemotropic Mycoplasma, and Rickettsia spp., infection) and flea (clade and Rickettsia and Wolbachia infection) factors on C. felis Bartonella clarridgeiae infection. From 189 free roaming cats, we collected 84 fleas: Ctenocephalides felis (78/84), Cediopsylla simplex (4/84), Orchopeas howardi (1/84), and Nosopsyllus fasciatus (1/84). Ctenocephalides felis were phylogenetically assigned to Clades 1, 4, and 6 by cox1 gene amplification. Rickettsia asembonensis (52/84) and B. clarridgeiae (16/84) were the most common pathogenic bacteria detected in fleas. Our model identified host cat sex and weight as independently associated with B. clarridgeiae infection in fleas. Rickettsia asembonensis (52/84), Rickettsia felis (7/84) and Bartonella henselae (7/84) were detected in specific clades: R. felis was detected only in Clades 1 and 6 while B. henselae and R. asembonensis were detected only in Clade 4. Wolbachia spp., also displayed clade specificity with strains other than Wolbachia wCfeT only infecting fleas from Clade 6. There was poor flea and host agreement for Bartonella spp., infection; however, there was agreement in the Bartonella species detected in cats and fleas by geographic location. These findings reinforce the importance of considering reservoir host attributes and vector phylogenetic diversity in epidemiological studies of flea-borne pathogens. Widespread sampling is necessary to identify the factors driving flea-borne pathogen presence and transmission.

Progress in detection of Mycoplasma pneumoniae infection

肺炎支原体是引起儿童社区获得性肺炎的主要病原菌。近年来，大环内酯类耐药肺炎支原体感染（MRMPI）比率的上升已成为全球关注的问题，MRMPI可能会延长症状持续时间和治疗时间，甚至增加并发症和后遗症的发生风险。因此，进行准确、快速、经济的肺炎支原体感染和MRMPI检测对于临床诊治至关重要。本文就肺炎支原体感染尤其是MRMPI感染检测的研究进展进行了综述。.

Detection of Macrolide Resistant Mycoplasma pneumoniae in Children with Lower Respiratory Tract Infection by Sanger Sequencing Targeting Domain V Region of 23S rRNA Gene

Mycoplasma pneumoniae is recognized as the leading cause of community-acquired lower respiratory tract infection in children, accounting for a significant proportion of pediatric mortality. Macrolides are the first-line treatment for M. pneumoniae infections. However, the extensive use of macrolides in clinical practice resulted in the emergence of macrolide-resistant M. pneumoniae (MRMP), which has a negative impact on treatment outcomes. Hence, in the present study, MRMP was determined in hospitalized children with community-acquired pneumonia (CAP). Real-time PCR detected M. pneumoniae in 41 of 348 clinical samples. Sanger sequencing revealed that none of the isolates were associated with the A2063G or A2064G base mutation, which confers macrolide resistance, in domain V of the 23S rRNA gene. Although MRMP was not observed in children with CAP in our study, healthcare practitioners should be vigilant about the potential risk of MRMP infections.

The detection of Mycoplasma sturni and Mycoplasma moatsii from the choana of a barn swallow (Hirundo rustica): a case report

BackgroundMycoplasmas are found in many different species. Until now 26 avian mycoplasma species have been described, but in the most free ranging bird species the prevalence and significance of Mycoplasma spp. is still unclear.Case presentationIn May 2021 a barn swallow (Hirundo rustica) was brought to a veterinary clinic after it hit a window. As part of the routine exam a choanal swab was taken for mycoplasma culture and for the detection of mycoplasmas using a Mycoplasma-genus-specific Polymerase chain reaction.Six single colony subcultures were obtained by the cultivation. Obtained subcultures were investigated by sequencing the 16S rRNA and the 16S-23S rRNA intergenic transcribed spacer region sequence. The 16S rRNA gene sequence from one subculture had a homology of 99.03% and the 16S-23S rRNA intergenic transcribed spacer region sequence of 100% with the sequence of Mycoplasma sturni.The 16S rRNA gene sequence from the other five subcultures shared a homology of 99.89% and the 16S-23S rRNA intergenic transcribed spacer region sequence of 99.81% with the sequence of Mycoplasma moatsii.ConclusionsAccording to the available literature this is the first report about the detection of M. moatsii, in the respiratory tract of a barn swallow. M. moatsii was previously only found in grivit monkeys (Cercopithecus aethiops), Norway rats (Rattus norvegicus) and a mute swan (Cygnus olor). The role of mycoplasmas in barn swallows is still unknown, especially as in the present case both mycoplasma species do not seem to cause clinical symptoms.

Comparison of Five Commercial Molecular Assays for Mycoplasma Testing of Cellular Therapy Products.

Testing of cellular therapy products for Mycoplasma is a regulatory requirement by the United States Food and Drug Administration (FDA) to ensure the sterility and safety of the product prior to release for patient infusion. The risk of Mycoplasma contamination in cell culture is high. Gold standard testing follows USP 63 which requires a 28-day agar and broth cultivation method that is impractical for short shelf-life biologics. Several commercial molecular platforms have been marketed for faster raw material and product release testing; however, little performance data are available in the literature. In this study, we performed a proof-of-principle analysis to evaluate the performance of five commercial molecular assays, including the MycoSEQ Mycoplasma detection kit (Life Technologies), the MycoTOOL Mycoplasma real-time detection kit (Roche), the VenorGEM qOneStep kit (Minerva Biolabs), the ATCC universal Mycoplasma detection kit, and the Biofire Mycoplasma assay (bioMérieux Industry) using 10 cultured Mollicutes spp., with each at four log-fold dilutions (1,000 CFU/mL to 1 CFU/mL) in biological duplicates with three replicates per condition (n = 6) to assess limit of detection (LOD) and repeatability. Additional testing was performed in the presence of tumor infiltrating lymphocytes (TILs). Based on LOD alone, the Biofire Mycoplasma assay was most sensitive followed by the MycoSEQ and MycoTOOL which were comparable. We showed that not all assays were capable of meeting the ≤10 CFU/mL LOD to replace culture-based methods according to European and Japanese pharmacopeia standards. No assay interference was observed when testing in the presence of TILs.