Baicalin attenuated Mycoplasma gallisepticum-induced immune impairment in chicken bursa of fabricius through modulation of autophagy and inhibited inflammation and apoptosis.

BACKGROUND: Mycoplasma gallisepticum (MG) is the primary etiologic agent of chronic respiratory disease in poultry. However, the mechanism underlying MG-induced immune dysregulation in chicken is still elusive. Baicalin shows excellent anti-bacterial, anti-inflammatory, anti-carcinogenic and anti-viral properties. In the present study, the preventive effects of baicalin against immune impairment in chicken bursa of fabricius (BF) were studied in an MG infection model. RESULTS: Histopathological examination showed increased inflammatory cell infiltrations and fragmented nuclei in the model group. Ultrastructural analysis revealed the phenomenon of apoptosis in bursal cells, along with the deformation of mitochondrial membrane and swollen mitochondria in the model group. However, these abnormal morphological changes were partially alleviated by baicalin. Meanwhile, baicalin treatment attenuated the level of proinflammatory cytokines, and suppressed nuclear factor-kappa B expression at both protein and mRNA level. Terminal deoxynucleotidyl transferase-mediated dUTP nick endlabeling assay showed extensive apoptosis in BF in the model group. The mRNA and protein expression levels of apoptosis-related genes were upregulated in BF, while baicalin treatment significantly alleviated apoptosis in BF. In addition, alterations in mRNA and protein expression levels of autophagy-related genes and mitochondrial dynamics proteins were significantly alleviated by baicalin. Moreover, baicalin treatment significantly attenuated MG-induced decrease in CD8+ cells and reduced bacterial load in chicken BF compared to the model group. CONCLUSIONS: These results suggested that baicalin could effectively inhibit MG-induced immune impairment and alleviate inflammatory responses and apoptosis in chicken BF. © 2020 Society of Chemical Industry.

The Enemy Within: How Does a Bacterium Inhibit the Foraging Aptitude and Risk Management Behavior of Allenby's Gerbils?

AbstractMicrobes inhabiting multicellular organisms have complex, often subtle effects on their hosts. Gerbillus andersoni allenbyi are commonly infected with Mycoplasma haemomuris-like bacteria, which may cause mild nutrient (choline, arginine) deficiencies. However, are there more serious ecological consequences of infection, such as effects on foraging aptitudes and risk management? We tested two alternatives: the nutrient compensation hypothesis (does nutrient deficiency induce infected gerbils to make up for the shortfall by foraging more and taking greater risks?) and (2) the lethargy hypothesis (do sick gerbils forage less, and are they compromised in their ability to detect predators or risky microhabitats?). We compared the foraging and risk management behavior of infected and noninfected gerbils. We experimentally infected gerbils with the bacteria, which allowed us to compare between noninfected, acutely infected (peak infection loads), and chronically infected (low infection loads) individuals. Our findings supported the lethargy hypothesis over the nutrient compensation hypothesis. Infected individuals incurred dramatically elevated foraging costs, including less efficient foraging, diminished "quality" of time spent vigilant, and increased owl predation. Interestingly, gerbils that were chronically infected (lower bacteria load) experienced larger ecological costs than acutely infected individuals (i.e., peak infection loads). This suggests that the debilitating effects of infection occur gradually, with a progressive decline in the quality of time gerbils allocated to foraging and managing risk. These increased long-term costs of infection demonstrate how small direct physiological costs of infection can lead to large indirect ecological costs. The indirect ecological costs of this parasite appear to be much greater than the direct physiological costs.

Interaction of Mycoplasma synoviae with chicken synovial sheath cells contributes to macrophage recruitment and inflammation.

Mycoplasma synoviae (MS) is an important avian pathogen causing considerable economic hardship in the poultry industry. A major inflammation caused by MS is synovitis that occurs in the synovial tendon sheath and joint synovium. However, the overall appearance of pathological changes in the tendon sheath and surrounding tissues caused by MS infection at the level of pathological tissue sections was poor. Studies on the role of MS and synovial sheath cells (SSCs) interaction in the development of synovitis have not been carried out. Through histopathological observation, our study found that a major MS-induced pathological change of the tendon sheath synovium was extensive scattered and focal inflammatory cell infiltration of the tendon sheath synovial layer. In vitro research experiments revealed that the CFU numbers of MS adherent and invading SSC, the levels of expression of various pattern recognition receptors, inflammatory cytokines, and chemokines coding genes, such as IL-1ß, IL-6, IL-8, CCL-20, RANTES, MIP-1ß, TLR7, and TLR15 in SSCs, and chemotaxis of macrophages were significantly increased when the multiplicity of infection (MOI) of MS to SSC were increased tenfold. The expression level of IL-12p40 in SSC was significantly higher when the MOIs of MS to SSC were increased by a factor of 100. The interaction between MS and SSC can activate macrophages, which was manifested by a significant increase in the expression of IL-1ß, IL-6, IL-8, CCL-20, RANTES, MIP-1ß, and CXCL-13. This study systematically demonstrated that the interaction of MS with chicken SSC contributes to the inflammatory response caused by the robust expression of related cytokines and macrophage chemotaxis. These findings are helpful in elucidating the molecular mechanism of MS-induced synovitis in chickens.

Baicalin inhibits inflammation caused by coinfection of Mycoplasma gallisepticum and Escherichia coli involving IL-17 signaling pathway.

Coinfection of Mycoplasma gallisepticum (MG) and Escherichia coli (E. coli) is frequently reported in poultry farms. Baicalin possess various pharmacological properties such as anti-inflammatory, anticancer, and antioxidant, etc. However, the protective effects of baicalin against coinfection of MG and E. coli are still elusive. In this study, baicalin (450 mg/kg) treatment was started on day 13 after infection and continued for 5 d. Histopathological examination, qRT-PCR, ELISA, and molecular docking technique were used to evaluate the effects of baicalin on MG and E. coli coinfection in chicken lung and trachea. The results showed that coinfection caused severe lesions in the lung and tracheal tissues. However, baicalin treatment partially alleviated these lesions in coinfection group. Histopathological examination showed the alveolar spaces and mucosal layer thickening was restored and cilia gradually recovered with baicalin treatment compared in coinfection group and MG-infection group. Meanwhile, IL-17 singling pathway-related genes were significantly reduced (P < 0.05) in baicalin treatment group in lung, including IL-17C, TRAF6, NF-κB, CXCL1, CXCL2, MMP1, GM-CSF, and MUC5AC. The activities of cytokines and chemokines (CXCL1, CXCL2, MMP1, GMCSF, and MUC5AC) were decreased significantly (P < 0.05) in baicalin-treated group. The molecular docking of baicalin and NF-κB showed the highest fitness score and interaction. From these results, it has been suggested that baicalin proved effective against coinfection of MG and E. coli in chicken and provided scientific basis for further dose-response and drug-target interaction studies.

Mycoplasma Infection Mediates Sensitivity of Multidrug-Resistant Cell Lines to Tiopronin: A Cautionary Tale.

We previously reported that some, but not all, multidrug-resistant cells that overexpressed various drug-resistance transporters were collaterally sensitive to tiopronin. In recent follow-up studies, we discovered that sensitivity to tiopronin in the original study was mediated by infection of the cells by a human-specific strain of mycoplasma. These results strongly support the need to constantly monitor cells for mycoplasma infection and keep stored samples of all cells that are used for in vitro studies.

A plethora of manifestations following a Mycoplasma pneumoniae infection: a case report.

Mycoplasma pneumoniae infection can present with a plethora of symptoms and result in a systemic vasculitis by activating a cascade of autoimmune reactions. In this case report, a young man without relevant past medical history was admitted to the hospital with diarrhea, abdominal pain and spiking fever. A CT-scan showed terminal ileitis. A 5-day broad spectrum antibiotic treatment (ciprofloxacin/clindamycin) did not result in any clinical improvement. On the contrary, the patient developed a cholestatic hepatitis, bilateral anterior uveitis and a dry cough. Extensive serological testing finally led to the diagnosis of a M. pneumoniae infection by paired serology (≥4-fold rise in IgG titer). In the diagnostic work-up, a PET-CT was performed and showed increased tracer uptake in the carotids and vertebral arteries, suggesting the diagnosis of vasculitis. After start of azithromycin and low-dose corticosteroids (0.5 mg/kg/day), a gradual clinical and biochemical improvement was observed. But subsequently, the patients relapsed and presented with an acute coronary syndrome. Coronary angiography revealed aneurysmatic deformation of the three coronary arteries, leading to the assumption of coronary vasculitis. Clinical improvement was achieved with high-dose corticosteroids (1 mg/kg/day). This case shows that M. pneumoniae is not merely a pulmonary infection, but that its primary symptoms can be diverse and misleading. All clinicians should be aware of its extrapulmonary manifestations.

The dynamics between limited-term and lifelong coinfecting bacterial parasites in wild rodent hosts.

Interactions between coinfecting parasites may take various forms, either direct or indirect, facilitative or competitive, and may be mediated by either bottom-up or top-down mechanisms. Although each form of interaction leads to different evolutionary and ecological outcomes, it is challenging to tease them apart throughout the infection period. To establish the first step towards a mechanistic understanding of the interactions between coinfecting limited-term bacterial parasites and lifelong bacterial parasites, we studied the coinfection of Bartonella sp. (limited-term) and Mycoplasma sp. (lifelong), which commonly co-occur in wild rodents. We infected Bartonella- and Mycoplasma-free rodents with each species, and simultaneously with both, and quantified the infection dynamics and host responses. Bartonella benefited from the interaction; its infection load decreased more slowly in coinfected rodents than in rodents infected with Bartonella alone. There were no indications for bottom-up effects, but coinfected rodents experienced various changes, depending on the infection stage, in their body mass, stress levels and activity pattern, which may further affect bacterial replication and transmission. Interestingly, the infection dynamics and changes in the average coinfected rodent traits were more similar to the chronic effects of Mycoplasma infection, whereas coinfection uniquely impaired the host's physiological and behavioral stability. These results suggest that parasites with distinct life history strategies may interact, and their interaction may be asymmetric, non-additive, multifaceted and dynamic through time. Because multiple, sometimes contrasting, forms of interactions are simultaneously at play and their relative importance alternates throughout the course of infection, the overall outcome may change under different ecological conditions.

Comparison of Finnish meat inspection records and average daily gain for cattle herds differing in Mycoplasma bovis test-status.

Detecting Mycoplasma bovis on cattle farms represents a challenge in the absence of an outbreak or cases of M. bovis mastitis, yet identification of an infection is essential to control the spread of the disease successfully. The objectives of this study were: (1) to determine whether meat inspection records can aid identification of cattle farms supporting M. bovis infection, and (2) to compare the average daily weight gain estimated from carcass weight for cattle originating from farms differing in M. bovis test-status. Meat inspection records were collected from two abattoirs in 2015; 80 677 animals in total. All the dairy and mixed breed cows and bulls used for meat production were categorized according to known M. bovis infection status of the farms from which the cattle were derived; positive, contact or control farms. The associations between animals from different M. bovis categories and lung lesions of bulls and cows (pneumonia and pleuritis), identified during meat inspection, and estimated average daily gain (ADG) of bulls, were investigated. The odds ratios for lung lesions, especially pleuritis, were higher in M. bovis test-positive or contact farms compared with control farms. Additionally, odds ratios for pleuritis were higher among animals from M. bovis test-positive farms and animals from contact slaughtering farms originating from M. bovis-free rearing farms. Bulls originating from M. bovis test-positive farms had higher estimated average daily gain than cattle from control farms. Meat inspection records can be used alongside other methods to detect M. bovis-positive farms where M. bovis causes lung lesions.

Bovine Epithelial in vitro Infection Models for Mycoplasma bovis.

Mycoplasma bovis causes bovine mycoplasmosis. The major clinical manifestations are pneumonia and mastitis. Recently an increase in the severity of mastitis cases was reported in Switzerland. At the molecular level, there is limited understanding of the mechanisms of pathogenicity of M. bovis. Host-pathogen interactions were primarily studied using primary bovine blood cells. Therefore, little is known about the impact of M. bovis on other cell types present in infected tissues. Clear in vitro phenotypes linked to the virulence of M. bovis strains or tissue predilection of specific M. bovis strains have not yet been described. We adapted bovine in vitro systems to investigate infection of epithelial cells with M. bovis using a cell line (MDBK: Madin-Darby bovine kidney cells) and two primary cells (PECT: bovine embryonic turbinate cells and bMec: bovine mammary gland epithelial cells). Two strains isolated before and after the emergence of severe mastitis cases were selected. Strain JF4278 isolated from a cow with mastitis and pneumonia in 2008 and strain L22/93 isolated in 1993 were used to assess the virulence of M. bovis genotypes toward epithelial cells with particular emphasis on mammary gland cells. Our findings indicate that M. bovis is able to adhere to and invade different epithelial cell types. Higher titers of JF4278 than L22/93 were observed in co-cultures with cells. The differences in titers reached between the two strains was more prominent for bMec cells than for MDBK and PECT cells. Moreover, M. bovis strain L22/93 induced apoptosis in MDBK cells and cytotoxicity in PECT cells but not in bMec cells. Dose-dependent variations in proliferation of primary epithelial cells were observed after M. bovis infection. Nevertheless, an indisputable phenotype that could be related to the increased virulence toward mammary gland cells is not obvious.

Mycoplasma genitalium adhesin P110 binds sialic-acid human receptors.

Adhesion of pathogenic bacteria to target cells is a prerequisite for colonization and further infection. The main adhesins of the emerging sexually transmitted pathogen Mycoplasma genitalium, P140 and P110, interact to form a Nap complex anchored to the cell membrane. Herein, we present the crystal structures of the extracellular region of the virulence factor P110 (916 residues) unliganded and in complex with sialic acid oligosaccharides. P110 interacts only with the neuraminic acid moiety of the oligosaccharides and experiments with human cells demonstrate that these interactions are essential for mycoplasma cytadherence. Additionally, structural information provides a deep insight of the P110 antigenic regions undergoing programmed variation to evade the host immune response. These results enlighten the interplay of M. genitalium with human target cells, offering new strategies to control mycoplasma infections.

Mycoplasma hyorhinis reduces sensitivity of human lung carcinoma cells to Nutlin-3 and promotes their malignant phenotype.

PURPOSE: MDM2 inhibitors are promising anticancer agents that induce cell cycle arrest and tumor cells death via p53 reactivation. We examined the influence of Mycoplasma hyorhinis infection on sensitivity of human lung carcinoma cells NCI-H292 to MDM2 inhibitor Nutlin-3. In order to unveil possible mechanisms underlying the revealed effect, we investigated gene expression changes and signal transduction networks activated in NCI-H292 cells in response to mycoplasma infection. METHODS: Sensitivity of NCI-Н292 cells to Nutlin-3 was estimated by resazurin-based cell viability assay. Genome-wide transcriptional profiles of NCI-H292 and NCI-Н292Myc.h cell lines were determined using Illumina Human HT-12 v3 Expression BeadChip. Search for key transcription factors and key node molecules was performed using the geneXplain platform. Ability for anchorage-independent growth was tested by soft agar colony formation assay. RESULTS: NCI-Н292Myc.h cells were shown to be 1.5- and 5.2-fold more resistant to killing by Nutlin-3 at concentrations of 15 and 30 µM than uninfected NCI-Н292 cells (P < 0.05 and P < 0.001, respectively). Transcriptome analysis revealed differential expression of multiple genes involved in cancer progression and metastasis as well as epithelial-mesenchymal transition (EMT). Moreover, we have shown experimentally that NCI-Н292Myc.h cells were more capable of growing and dividing without binding to a substrate. The most likely mechanism explaining the observed changes was found to be TLR4- and IL-1b-mediated activation of NF-κB pathway. CONCLUSIONS: Our results provide evidence that mycoplasma infection is an important factor modulating the effect of MDM2 inhibitors on cancer cells and is able to induce EMT-related changes.

Improvement of semen parameters after antibiotic therapy in asymptomatic infertile men infected with Mycoplasma genitalium.

OBJECTIVES: To elucidate the association between asymptomatic infections caused by Mycoplasma genitalium and male infertility, and evaluate the role of antibiotic therapy in treatment of this failure. METHODS: A total of 165 infertile males having abnormal semen parameters (study group) and 165 healthy fertile men (control group) were included. Semen samples were taken from all participants and after analyzing for semen parameters, undergone real-time PCR, microbial culture, and reactive oxygen species (ROS), as well as total antioxidant capacity (TAC) assays. Infected individuals of study group were treated with antibiotic. One month after the treatment completion, second semen samples were taken and subjected to all the tests mentioned. The data were analyzed using SPSS statistical software, version 22.0. RESULTS: The frequency of M. genitalium was significantly higher in the infertile men compared with the fertile ones (9.7% vs. 1.2%; p = 0.001). Mean cycle threshold (C t) value was lower in infected infertile than infected fertile men (p < 0.001). All semen parameters, except volume, pH, and viscosity, were improved (p < 0.05), most of which reached their normal range; leukocytes in seminal fluid decreased (p = 0.02), the level of TAC was elevated (p = 0.002), and ROS level as well as ROS/TAC ratio reduced after antibiotic treatment (p = 0.03). Wives of seven infected infertile men (43.8%) became pregnant 4 months after the treatment completion. CONCLUSIONS: Asymptomatic infection caused by M. genitalium is correlated with male infertility and antibiotic therapy can improve the semen quality and be used to treat male infertility.

Novel role of Vpmas as major adhesins of Mycoplasma agalactiae mediating differential cell adhesion and invasion of Vpma expression variants.

Mycoplasma agalactiae exhibits antigenic variation by switching the expression of multiple surface lipoproteins called Vpmas. Although implicated to have a significant influence on the pathogenicity, their exact role in pathogen-host interactions has not been investigated so far. Initial attachment to host cells is regarded as one of the most important steps for colonization but this pathogen lacks the typical mycoplasma attachment organelle. The aim of this study was to determine the role of Vpmas in adhesion of M. agalactiae to host cells. 'Phase-Locked' Mutants (PLMs) steadily expressing single well-characterized Vpma lipoproteins served as ideal tools to evaluate the role of each of the six Vpmas in cytadhesion, which was otherwise not possible due to the high-frequency switching of Vpmas in the wildtype strain PG2. Using in vitro adhesion assays with HeLa and sheep mammary epithelial (MECs) and stromal (MSCs) cells, we could demonstrate differences in the adhesion capabilities of each of the six PLMs compared to the wildtype strain. The PLMV mutant expressing VpmaV exhibited the highest adhesion rate, whereas PLMU, which expresses VpmaU showed the lowest adhesion values explaining the reduced in vivo fitness of PLMU in sheep during experimental intramammary and conjunctival infections. Furthermore, adhesion inhibition assays using Vpma-specific polyclonal antisera were performed to confirm the role of Vpmas in M. agalactiae cytadhesion. This led to a significant decrease (p<0.05) in the adhesion percentage of each PLM. Immunofluorescence staining of TX-114 phase proteins extracted from each PLM showed binding of the respective Vpma to HeLa cells and MECs proving the direct role of Vpmas in cytadhesion. Furthermore, as adhesion is a prerequisite for cell invasion, the ability of the six PLMs to invade HeLa cells was also evaluated using the gentamicin protection assay. The results showed a strong correlation between the adhesion rates and invasion frequencies of the individual PLMs. This is the first report that describes a novel function of Vpma proteins in cell adhesion and invasion. Besides the variability of these proteins causing surface antigenic variation, the newly identified phenotypes are likely to play critical roles in the pathogenicity potential of this ruminant pathogen.

Acute necrotising encephalopathy of childhood: A review of two cases.

Acute necrotising encephalopathy of childhood (ANEC) is an uncommon disease with characteristic clinical and imaging findings. We present two cases of ANEC secondary to Respiratory Syncytial Virus (RSV) and mycoplasma infections. An eight-month-old boy presented with features of gastroenteritis but soon developed multiple episodes of seizures. Blood and CSF cultures were negative but nasopharyngeal aspirate immunofluorescence was positive for RSV. A nine-year-old girl presented with abnormal behaviour following two days of prodromal symptoms. Her serological markers implicated mycoplasma (IgM titre 1: 640). CT brain of both patients showed bilateral symmetrical thalamic hypodensities, while MRI revealed more extensive white matter involvements.

[Mixed infections and inflammatory ophthalmic diseases: clinical and laboratory observations]. / Mikst-infektsii i vospalitel'naia oftal'mopatologiia: kliniko-laboratornye nabliudeniia.

In recent years, all medical specialists, including ophthalmologists, have been facing the problem of mixed infections. Recurrent inflammation in the anterior and posterior eye segments is often a result of infection by more than one variety of pathogens. MATERIAL AND METHODS: Over the period 2013-2016, 34 patients (14 men and 20 women) with different inflammatory processes in the eye who appeared DNA-positive for mycoplasmas (Mycoplasma hominis, Ureaplasma urealyticum) and/or chlamydiae (Chlamydia trachomatis) (PCR testing of tear fluid and/or urine) were followed up. All patients were examined for intensive production of herpesvirus, adenovirus, and enterovirus DNA in biological fluids. After being consulted by related specialists, all the patients started local and systemic (antibacterial and antiviral) therapy. In the end of the latter, laboratory tests were repeated. RESULTS: Among all the clinical forms, anterior segment inflammation (i.e. of conjunctiva, cornea, and the anterior vascular tract) prevailed - 76%. In most patients, mycoplasmas and/or chlamydiae formed associations with herpesviruses (n=19; 56%). Bacterial DNA alone (mycoplasma and/or chlamydia) was detected in 12 cases (35%). In 4 cases, mycoplasma and/or chlamydia DNA was detected in tear fluid, in 19 patients - in urine, and in 10 patients - in both secreta. Local and systemic causal treatment enabled resolution of the complaints and symptoms and yielded negative results of follow-up laboratory tests. CONCLUSION: More than a half of the patients demonstrated concomitant viral-bacterial infection (22 cases). The presence of bacterial/viral DNA in biological secreta, as revealed by PCR, reflects the systemic nature of the infection process and, thus, necessitates engagement of related specialists (dermatologists, urologists, gynecologists).

The association between Mycoplasma pneumoniae infection and speech and language impairment: A nationwide population-based study in Taiwan.

Manifestations of Mycoplasma pneumoniae infection can range from self-limiting upper respiratory symptoms to various neurological complications, including speech and language impairment. But an association between Mycoplasma pneumoniae infection and speech and language impairment has not been sufficiently explored. In this study, we aim to investigate the association between Mycoplasma pneumoniae infection and subsequent speech and language impairment in a nationwide population-based sample using Taiwan's National Health Insurance Research Database. We identified 5,406 children with Mycoplasma pneumoniae infection (International Classification of Disease, Revision 9, Clinical Modification code 4830) and compared to 21,624 age-, sex-, urban- and income-matched controls on subsequent speech and language impairment. The mean follow-up interval for all subjects was 6.44 years (standard deviation = 2.42 years); the mean latency period between the initial Mycoplasma pneumoniae infection and presence of speech and language impairment was 1.96 years (standard deviation = 1.64 years). The results showed that Mycoplasma pneumoniae infection was significantly associated with greater incidence of speech and language impairment [hazard ratio (HR) = 1.49, 95% CI: 1.23-1.80]. In addition, significantly increased hazard ratio of subsequent speech and language impairment in the groups younger than 6 years old and no significant difference in the groups over the age of 6 years were found (HR = 1.43, 95% CI:1.09-1.88 for age 0-3 years group; HR = 1.67, 95% CI: 1.25-2.23 for age 4-5 years group; HR = 1.14, 95% CI: 0.54-2.39 for age 6-7 years group; and HR = 0.83, 95% CI:0.23-2.92 for age 8-18 years group). In conclusion, Mycoplasma pneumoniae infection is temporally associated with incident speech and language impairment.

Comprehensive RNA-Seq Profiling to Evaluate the Sheep Mammary Gland Transcriptome in Response to Experimental Mycoplasma agalactiae Infection.

Mycoplasma agalactiae is a worldwide serious pathogen of small ruminants that usually spreads through the mammary route causing acute to subacute mastitis progressing to chronic persistent disease that is hard to eradicate. Knowledge of mechanisms of its pathogenesis and persistence in the mammary gland are still insufficient, especially the host-pathogen interplay that enables it to reside in a chronic subclinical state. This study reports transcriptome profiling of mammary tissue from udders of sheep experimentally infected with M. agalactiae type strain PG2 in comparison with uninfected control animals using Illumina RNA-sequencing (RNA-Seq). Several differentially expressed genes (DEGs) were observed in the infected udders and RT-qPCR analyses of selected DEGs showed their expression profiles to be in agreement with results from RNA-Seq. Gene Ontology (GO) analysis revealed majority of the DEGs to be associated with mycoplasma defense responses that are directly or indirectly involved in host innate and adaptive immune responses. Similar RNA-Seq analyses were also performed with spleen cells of the same sheep to know the specific systemic transcriptome responses. Spleen cells exhibited a comparatively lower number of DEGs suggesting a less prominent host response in this organ. To our knowledge this is the first study that describes host transcriptomics of M. agalactiae infection and the related immune-inflammatory responses. The data provides useful information to further dissect the molecular genetic mechanisms underlying mycoplasma mastitis, which is a prerequisite for designing effective intervention strategies.

Background review for the 2016 European guideline on Mycoplasma genitalium infections.

Mycoplasma genitalium is a cause of 10-35% of non-chlamydial non-gonococcal urethritis in men and in women, and is associated with cervicitis and pelvic inflammatory disease (PID). Transmission of M. genitalium occurs through direct mucosal contact. In women, symptoms include vaginal discharge, dysuria or symptoms of PID - abdominal pain and dyspareunia. In men, urethritis, dysuria and discharge predominates. Asymptomatic infections are frequent. In this review, we present the evidence base for the recommendations in the 2016 European guideline on M. genitalium infections and describe indications for testing, recommended diagnostic methods, treatment and patient management. The guideline was prepared on behalf of the European branch of The International Union against Sexually Transmitted Infections; the European Academy of Dermatology and Venereology; the European Dermatology Forum; the European Society of Clinical Microbiology and Infectious Diseases; the Union of European Medical Specialists. The European Centre for Disease Prevention and Control and the European Office of the World Health Organisation also contributed to their development.

Development of fluorescence expression tools to study host-mycoplasma interactions and validation in two distant mycoplasma clades.

Fluorescence expression tools for stable and innocuous whole mycoplasma cell labelling have been developed. A Tn4001-derivative mini-transposon affording unmarked, stable mutagenesis in mycoplasmas was modified to allow the constitutive, high-level expression of mCherry, mKO2 and mNeonGreen. These tools were used to introduce the respective fluorescent proteins as chromosomal tags in the phylogenetically distant species Mycoplasma mycoides subsp. mycoides and Mycoplasma bovis. The production, selection and characterisation of fluorescent clones were straightforward and resulted in the unprecedented observation of red and green fluorescent mycoplasma colonies in the two species, with no apparent cytotoxicity. Equivalent fluorescence expression levels were quantified by flow cytometry in both species, suggesting that these tools can be broadly applied in mycoplasmas. A macrophage infection assay was performed to assess the usefulness of mNeonGreen-expressing strains for monitoring mycoplasma infections, and notably cell invasion. The presence of fluorescent mycoplasmas inside live phagocytic cells was detected and quantified by flow cytometry and corroborated by confocal microscopy, which allowed the identification of individual mycoplasmas in the cytoplasm of infected cells. The fluorescence expression tools developed in this study are suitable for host-pathogen interaction studies and offer innumerable perspectives for the functional analysis of mycoplasmas both in vitro and in vivo.