Molecular detection of Mycoplasma gallisepticum in different poultry breeds of Abbottabad and Rawalpindi, Pakistan / Detecção molecular de Mycoplasma gallisepticum em diferentes raças de aves de Abbottabad e Rawalpindi, Paquistão

The poultry sector in Pakistan is contributing mainly in bridging gap between demand and supply for protein. Mycoplasma gallisepticum is an emerging bacterium causing serious problems in poultry industry of Pakistan. A cross-sectional study was conducted to evaluate the M. gallisepticum load in poultry populated regions of Pakistan. Total 600 serum and 600 swab samples were collected, 200 from each broiler, layers and breeders poultry in Rawalpindi and Abbottabad districts. Serum samples were analyzed through ELISA for seroprevalence. Swabs were cultured on Frey’s medium followed by PCR and partial mgc2 gene sequencing. Results of seroprevalence of M. gallisepticum showed that layers (75%, n=150) are more positive as compared to breeders (70%, n=140) and broilers (50%, n=100). Typical colonies of the M. gallisepticum were observed in breeder (26.5%), followed by layer (21%) and broilers (9%). A total of 37.1% (n=42) samples were identified positive through PCR out of total 113 cultured based positive samples. A total of six M. gallisepticum isolates of current study showed 98-99 percent similarity with previously reported isolates on the basis of mgc2 gene partial sequencing. The M. gallisepticum was found highly prevalent in different poultry breads. Results of this study would add into basic data and provide a direction for livestock sector to strengthen a control strategy for mycoplasmosis in poultry farms.

O setor avícola do Paquistão está contribuindo principalmente para preencher a lacuna entre a demanda e a oferta de proteína. Mycoplasma gallisepticum é uma bactéria emergente que causa sérios problemas na indústria avícola do Paquistão. Um estudo transversal foi conduzido para avaliar a carga de M. gallisepticum em regiões de avicultura do Paquistão. Um total de 600 amostras de soro e 600 amostras de esfregaço foi coletado, 200 de cada frango de corte, poedeiras e aves reprodutoras nos distritos de Rawalpindi e Abbottabad. Amostras de soro foram analisadas por ELISA para soroprevalência. As zaragatoas foram cultivadas em meio Frey, seguido de PCR e sequenciação parcial do gene mgc2. Os resultados da soroprevalência de M. gallisepticum mostraram que as poedeiras (75%, n = 150) são mais positivas em comparação com matrizes (70%, n = 140) e frangos de corte (50%, n = 100). Colônias típicas de M. gallisepticum foram observadas em reprodutoras (26,5%), seguidas de poedeiras (21%) e frangos de corte (9%). Um total de 37,1% (n = 42) das amostras foi identificado como positivas por PCR de um total de 113 amostras positivas baseadas em cultura. Um total de seis isolados de M. gallisepticum do estudo atual mostrou 98-99% de similaridade com isolados relatados anteriormente com base no sequenciamento parcial do gene mgc2. O M. gallisepticum foi encontrado com alta prevalência em diferentes pães de aves. Os resultados deste estudo acrescentariam dados básicos e forneceriam orientação para o setor pecuário fortalecer uma estratégia de controle da micoplasmose em granjas avícolas.

Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test.

Contamination of cells/tissues by infectious pathogens (e.g., fungi, viruses, or bacteria, including mycoplasma) is a major problem in cell-based transplantation. In this study, we tested a polymerase chain reaction (PCR) method to provide rapid, simple, and sensitive detection of mycoplasma contamination in laboratory cultures for clinical use. This mycoplasma PCR system covers the Mycoplasma species (spp.) listed for testing in the 17th revision of the Japanese Pharmacopoeia, and we designed it for use in transplantable retinal cells. Here, we analyzed mycoplasma contamination in induced pluripotent stem cell (iPS cell)-derived transplantable retinal pigment epithelium (RPE) cells. In the spike tests to RPE cells with nine species of class Mollicutes bacteria, including seven Mycoplasma spp. and one of each Acholeplasma spp. and Ureaplasma spp., contamination at the concentration of 100 and 10 CFU/mL were detected with 100% probability in all cases, while 1 CFU/mL had a detection rate of 0-75%. DNA prepared from bacteria species other than class Mollicutes species was not detectable, indicating the specificity of this PCR. While iPS cells and iPS-RPE cells established in our laboratory were all negative by this PCR, some of the commercially available cell lines were positive. Cells for transplantation should never have infection, as once pathogens are implanted into the eyes, they can cause severe intraocular inflammation. Thus, it is imperative to monitor for infections in the transplants, although generally, mycoplasma infection is difficult to detect.

Contagious Bovine Pleuropneumonia: A Comprehensive Overview.

Contagious bovine pleuropneumonia (CBPP) is a respiratory disease of cattle that is listed as notifiable by the World Organization for Animal Health. It is endemic in sub-Saharan Africa and causes important productivity losses due to the high mortality and morbidity rates. CBPP is caused by Mycoplasma mycoides subsp. mycoides (Mmm) and is characterized by severe fibrinous bronchopneumonia and pleural effusion during the acute to subacute stages and by pulmonary sequestra in chronic cases. Additional lesions can be detected in the kidneys and in the carpal and tarsal joints of calves. Mmm infection occurs through the inhalation of infected aerosol droplets. After the colonization of bronchioles and alveoli, Mmm invades blood and lymphatic vessels and causes vasculitis. Moreover, Mmm can be occasionally demonstrated in blood and in a variety of other tissues. In the lung, Mmm antigen is commonly detected on bronchiolar and alveolar epithelial cells, in lung phagocytic cells, within the wall of blood and lymphatic vessels, inside necrotic areas, and within tertiary lymphoid follicles. Mmm antigen can also be present in the cytoplasm of macrophages within lymph node sinuses, in the germinal center of lymphoid follicles, in glomerular endothelial cells, and in renal tubules. A complete pathological examination is of great value for a rapid presumptive diagnosis, but laboratory investigations are mandatory for definitive diagnosis. The purpose of this review is to describe the main features of CBPP including the causative agent, history, geographic distribution, epidemiology, clinical course, diagnosis, and control. A special focus is placed on gross and microscopic lesions in order to familiarize veterinarians with the pathology and pathogenesis of CBPP.

Keeping It Clean: The Cell Culture Quality Control Experience at the National Center for Advancing Translational Sciences.

Quality control monitoring of cell lines utilized in biomedical research is of utmost importance and is critical for the reproducibility of data. Two key pitfalls in tissue culture are 1) cell line authenticity and 2) Mycoplasma contamination. As a collaborative research institute, the National Center for Advancing Translational Sciences (NCATS) receives cell lines from a range of commercial and academic sources, which are adapted for high-throughput screening. Here, we describe the implementation of routine NCATS-wide Mycoplasma testing and short tandem repeat (STR) testing for cell lines. Initial testing identified a >10% Mycoplasma contamination rate. While the implementation of systematic testing has not fully suppressed Mycoplasma contamination rates, clearly defined protocols that include the immediate destruction of contaminated cell lines wherever possible has enabled rapid intervention and removal of compromised cell lines. Data for >2000 cell line samples tested over 3 years, and case studies are provided. STR testing of 186 cell lines with established STR profiles revealed only five misidentified cell lines, all of which were received from external labs. The data collected over the 3 years since implementation of this systematic testing demonstrate the importance of continual vigilance for rapid identification of "problem" cell lines, for ensuring reproducible data in translational science research.

Role of Mycoplasma Chaperone DnaK in Cellular Transformation.

Studies of the human microbiome have elucidated an array of complex interactions between prokaryotes and their hosts. However, precise bacterial pathogen-cancer relationships remain largely elusive, although several bacteria, particularly those establishing persistent intra-cellular infections, like mycoplasmas, can alter host cell cycles, affect apoptotic pathways, and stimulate the production of inflammatory substances linked to DNA damage, thus potentially promoting abnormal cell growth and transformation. Consistent with this idea, in vivo experiments in several chemically induced or genetically deficient mouse models showed that germ-free conditions reduce colonic tumor formation. We demonstrate that mycoplasma DnaK, a chaperone protein belonging to the Heath shock protein (Hsp)-70 family, binds Poly-(ADP-ribose) Polymerase (PARP)-1, a protein that plays a critical role in the pathways involved in recognition of DNA damage and repair, and reduces its catalytic activity. It also binds USP10, a key p53 regulator, reducing p53 stability and anti-cancer functions. Finally, we showed that bystander, uninfected cells take up exogenous DnaK-suggesting a possible paracrine function in promoting cellular transformation, over and above direct mycoplasma infection. We propose that mycoplasmas, and perhaps certain other bacteria with closely related DnaK, may have oncogenic activity, mediated through the inhibition of DNA repair and p53 functions, and may be involved in the initiation of some cancers but not necessarily involved nor necessarily even be present in later stages.

Emerging role for Ureaplasma parvum serovar 3: Active infection in women with silent high-risk human papillomavirus and in women with idiopathic infertility.

Recently, there are controversial opinions on the presence of Mycoplasmas/Ureaplasmas as colonizers or pathogens, and on the use of a targeted therapy. This study aimed to characterize Mycoplasmas/Ureaplasmas infections in reproductive age women, including the acquisition of sexually transmitted (ST) pathogens and poor birth outcomes. A total of 646 healthy Italian women fulfilled the inclusion criteria including 521 infertile women, 65 pregnant women, and 60 fertile women with identified risk factors and symptomatic for vaginitis/cervicitis. Multiplex and quantitative molecular techniques and direct automatic DNA sequencing were performed to assess the genome structure of Mycoplasma/Ureaplasma species and ST infected pathogens. Ureaplasma parvum serovar 3 represented the predominant colonizer of the urogenital tract of this series and the unique species significantly associated with ST pathogens coinfection (p < 0.01). U. parvum load >104 bacteria/ml, suggestive of active infection, has been measured only in asymptomatic high-risk human papillomavirus infected women (24.3%) and in 40% of women with idiopathic infertility. To note, 16% of the follicular fluid from these idiopathic women resulted infected with U. parvum. In conclusion, the present study focused the attention on U. parvum serovar 3 as emerging microorganism in sexually active women that may have the benefit of targeted therapy.

Primer aislamiento de Mycoplasma leachii asociado a enfermedad en terneros de tambo en Argentina / First report of Mycoplasma leachii isolation associated with disease in dairy calves in Argentina

There are few reports about the isolation of Mycoplasma species associated with cattle disease in Argentina. In this work we describe the detection of Mycoplasma leachii associated with disease in dairy calves in Santa Fe Province, Argentina. Samples obtained from a 4 day-old dairy calf suffering from polyarthritis and from two other calves, one with arthritis and the other one with a mandibular abscess, were subjected to microbiological culture. Classical culture and generic PCR confirmed the presence of Mycoplasma spp. The spacer region between the 16S and 23S ribosomal RNA gene from the first isolate was amplified and sequenced. The sequence obtained showed 99% identity with M. leachii. A PCR was developed to amplify a specific fragment of the 16S-23S ITS region corresponding to M. leachii, which allowed to identify the isolates associated with disease in calves.

Existen pocos informes acerca del aislamiento de especies de Mycoplasma asociadas con enfermedades del ganado en Argentina. En esta comunicación se describe el aislamiento de Mycoplasma leachii asociado a enfermedad en terneros de tambo en la provincia de Santa Fe, Argentina. Se obtuvieron muestras de un ternero de 4 días de vida con poliartritis, de un ternero con artritis y uno con un absceso mandibular. A partir del cultivo clásico se detectó la presencia de Mycoplasma, lo cual fue confirmado por PCR genérica. Se amplificó y secuenció la región ITS 16S-23S a partir del primer aislamiento, mostrando una identidad del 99% con Mycoplasma leachii. Se desarrolló una PCR para amplificar un fragmento específico de la región ITS 16S-23S correspondiente a M. leachii, que permitió identificar los aislamientos asociados con enfermedad en terneros.

Mycoplasma promotes malignant transformation in vivo, and its DnaK, a bacterial chaperone protein, has broad oncogenic properties.

We isolated a strain of human mycoplasma that promotes lymphomagenesis in SCID mice, pointing to a p53-dependent mechanism similar to lymphomagenesis in uninfected p53-/- SCID mice. Additionally, mycoplasma infection in vitro reduces p53 activity. Immunoprecipitation of p53 in mycoplasma-infected cells identified several mycoplasma proteins, including DnaK, a member of the Hsp70 chaperon family. We focused on DnaK because of its ability to interact with proteins. We demonstrate that mycoplasma DnaK interacts with and reduces the activities of human proteins involved in critical cellular pathways, including DNA-PK and PARP1, which are required for efficient DNA repair, and binds to USP10 (a key p53 regulator), impairing p53-dependent anticancer functions. This also reduced the efficacy of anticancer drugs that depend on p53 to exert their effect. mycoplasma was detected early in the infected mice, but only low copy numbers of mycoplasma DnaK DNA sequences were found in some primary and secondary tumors, pointing toward a hit-and-run/hide mechanism of transformation. Uninfected bystander cells took up exogenous DnaK, suggesting a possible paracrine function in promoting malignant transformation, over and above cells infected with the mycoplasma. Phylogenetic amino acid analysis shows that other bacteria associated with human cancers have similar DnaKs, consistent with a common mechanism of cellular transformation mediated through disruption of DNA-repair mechanisms, as well as p53 dysregulation, that also results in cancer-drug resistance. This suggests that the oncogenic properties of certain bacteria are DnaK-mediated.

Effects of mycoplasma infection on the host organism response via p53/NF-κB signaling.

Mycoplasmas are bacteria lacking the cell wall, which is the major characteristic of this taxonomic class (Mollicutes). Among bacteria, mycoplasmas possess the smallest genome known for free-living organisms. This feature limits the autonomy of bacteria and makes them increasingly susceptible to changes in the host organism. Many mycoplasmas themselves cause pathological changes in the host organism, often complicated by immune disorders. Infection with certain strains of mycoplasma results in the activation of the nuclear factor kappa-light-chain-enhancer of activated B cells, which is the major mediator of the inflammatory response. Furthermore, mycoplasmas can inhibit p53-mediated checkpoint control of cell cycle and apoptosis. Collectively, these properties indicate that mycoplasmas might act as cancer-promoting factors. In this review, we summarize the information known to date on the role of mycoplasmas in the regulation of the host immune response and their functional interactions with p53.

Necrotizing pneumonia caused by refractory Mycoplasma pneumonia pneumonia in children.

BACKGROUND: To investigate the clinical features of necrotizing pneumonia (NP) caused by refractory Mycoplasma pneumoniae pneumonia (RMPP). METHODS: A retrospective observational study was carried out in patients with NP caused by RMPP who were admitted to our hospital from January 2008 to December 2015, and the clinical manifestations, laboratory data, imaging performances, hospital courses and outcomes were analyzed. RESULTS: Twenty-five patients with NP caused by RMPP were collected, with a median age of 5.1 (4.0-7.9) years. The mean duration of fever and hospital stay was 21.0 ± 8.9 and 19.9 ± 9.9 days, respectively. The levels of lactate dehydrogenase (LDH), C-reactive protein, interleukin (IL)-6, IL-10 and interferon-gamma were elevated. Meanwhile, the pleural fluid cell count, LDH and protein were also increased. 80.0% of the patients had pleural effusion; and a high incidence of lobar atelectasis and pulmonary consolidation was found the patients. The mean duration from the onset of symptoms to the discovery of necrotic lesions was 21.0 ± 6.9 days. 80.0% of the patients were administrated corticosteroids, and bronchoalveolar lavage was extracted separately from all patients. Of the 20 patients who presented with pleural effusion, 11 underwent thoracocentesis alone and 2 underwent chest drainage. All patients received prolonged courses of antibiotics (32.2 ± 8.7 days). All patients were dischaged home and recovered without surgical intervention; and chest lesions were resolved or only minimal residual fibrotic changes were residual within 3.0 (2.0-6.0) months. CONCLUSIONS: Necrotizing pneumonia caused by RMPP is severe, however, self-limiting and reversible. Good outcomes can be achieved with appropriate management.

High-sensitivity virus and mycoplasma screening test reveals high prevalence of parvovirus B19 infection in human synovial tissues and bone marrow.

BACKGROUND: Latent microorganism infection is a safety concern for the clinical application of mesenchymal stem cells (MSCs). The aim of this study is to investigate the frequencies and sensitivities of the latent virus and mycoplasma infections in synovium, bone marrow, peripheral blood cells, and blood plasma and cultured synovial MSCs. METHODS: Total DNA and RNA of the synovium (n = 124), bone marrow (n = 123), peripheral blood cells (n = 121), plasma (n = 121), and 14-day cultured synovial MSCs (n = 63) were collected from patients who underwent total knee arthroplasty or anterior ligament reconstruction after written informed consents were obtained. The multiplex polymerase chain reaction (PCR) primers were designed to quantitatively measure the representative genomes of 13 DNA viruses, 6 RNA viruses, and 9 mycoplasmas. Multi-spliced mRNA detection and virus spike test were also performed to demonstrate the sensitivity of synovial MSCs to the candidate pathogens. RESULTS: In synovium and bone marrow, the positive rates of parvovirus B19 genome were significantly higher than in peripheral blood cells (18.7% and 22% vs. 0.8%, respectively). Multi-alignment analysis of amplified and sequenced viral target genes showed the proximity of the parvovirus B19 gene from different tissue in the same patients. Synovial MSCs cultured for 14 days were positive for virus infection only in two patients (2/62 = 3%). Parvovirus B19 multi-spliced mRNAs were not detected in these two samples. Virus spike test demonstrated the sensitivity of synovial MSCs to herpes simplex virus (HSV)1 and cytomegalovirus (CMV), but not to parvovirus B19. CONCLUSION: This study revealed a relatively high incidence of latent parvovirus B19 in synovium and bone marrow tissue.

Mycoplasma infection and hypoxia initiate succinate accumulation and release in the VM-M3 cancer cells.

Succinate is known to act as an inflammatory signal in classically activated macrophages through stabilization of HIF-1α leading to IL-1ß production. Relevant to this, hypoxia is known to drive succinate accumulation and release into the extracellular milieu. The metabolic alterations associated with succinate release during inflammation and under hypoxia are poorly understood. Data are presented showing that Mycoplasma arginini infection of VM-M3 cancer cells enhances the Warburg effect associated with succinate production in mitochondria and eventual release into the extracellular milieu. We investigated how succinate production and release was related to the changes of other soluble metabolites, including itaconate and 2-HG. Furthermore, we found that hypoxia alone could induce succinate release from the VM-M3 cells and that this could occur in the absence of glucose-driven lactate production. Our results elucidate metabolic pathways responsible for succinate accumulation and release in cancer cells, thus identifying potential targets involved in both inflammation and hypoxia. This article is part of a Special Issue entitled 20th European Bioenergetics Conference, edited by László Zimányi and László Tretter.

Simulated acute central Mycoplasma infections in rats induce fever, anorexia, body mass stunting and lethargy but spare memory.

Despite the documented post-infectious neurological complications of a central nervous system (CNS) Mycoplasma infection in humans, very few studies have investigated the acute inflammatory responses and sickness behaviours induced by CNS Mycoplasma infections. We therefore determined the effect of acute central administration of fibroblast-stimulating lipopeptide-1 (FSL-1), derived from Mycoplasma salivarium, and FAM-20 from a more pathogenic species, namely Mycoplasma pneumoniae, on behavioural and inflammatory responses in rats. Male Sprague-Dawley rats had radiotransmitters implanted, intra-abdominally, to measure body temperature and cage activity continuously. After recovery from surgery, rats were conditioned in a fear conditioning task and then immediately received an intra-cisterna magna (i.c.m.) injection of either: (1) FSL-1 (10 or 100µg/5µl) or its vehicle (phosphate-buffered saline, 5µl), or (2) FAM-20 (10 or 100µg/5µl) or its vehicle (dimethyl sulfoxide, 5µl). Body mass and food intake were measured daily. Memory was assessed seven days after injection using fear conditioning tests. A single, i.c.m. injection of either FSL-1 or FAM-20 induced profound, dose-dependent fever, anorexia, lethargy and body mass stunting in rats. Moreover, rats that received an i.c.m. injection of 100µg/5µl FAM-20 had a significant increase in the concentration of IL-1ß in both the hypothalamus and the hippocampus for ~27h after injection. Seven days after FSL-1 or FAM-20 injection, when body mass of rats still was stunted, they maintained their memory for fear of the context and for fear of the tone, despite the increase in hippocampal IL-1ß concentration after FAM-20 administration. Thus, acute simulated CNS Mycoplasma infections caused pronounced sickness responses and brain inflammation in rats, but spared fear memory.

Low-pathogenicity Mycoplasma spp. alter human monocyte and macrophage function and are highly prevalent among patients with ventilator-acquired pneumonia.

BACKGROUND: Ventilator-acquired pneumonia (VAP) remains a significant problem within intensive care units (ICUs). There is a growing recognition of the impact of critical-illness-induced immunoparesis on the pathogenesis of VAP, but the mechanisms remain incompletely understood. We hypothesised that, because of limitations in their routine detection, Mycoplasmataceae are more prevalent among patients with VAP than previously recognised, and that these organisms potentially impair immune cell function. METHODS AND SETTING: 159 patients were recruited from 12 UK ICUs. All patients had suspected VAP and underwent bronchoscopy and bronchoalveolar lavage (BAL). VAP was defined as growth of organisms at >10(4) colony forming units per ml of BAL fluid on conventional culture. Samples were tested for Mycoplasmataceae (Mycoplasma and Ureaplasma spp.) by PCR, and positive samples underwent sequencing for speciation. 36 healthy donors underwent BAL for comparison. Additionally, healthy donor monocytes and macrophages were exposed to Mycoplasma salivarium and their ability to respond to lipopolysaccharide and undertake phagocytosis was assessed. RESULTS: Mycoplasmataceae were found in 49% (95% CI 33% to 65%) of patients with VAP, compared with 14% (95% CI 9% to 25%) of patients without VAP. Patients with sterile BAL fluid had a similar prevalence to healthy donor BAL fluid (10% (95% CI 4% to 20%) vs 8% (95% CI 2% to 22%)). The most common organism identified was M. salivarium. Blood monocytes from healthy volunteers incubated with M. salivarium displayed an impaired TNF-α response to lipopolysaccharide (p=0.0003), as did monocyte-derived macrophages (MDMs) (p=0.024). MDM exposed to M. salivarium demonstrated impaired phagocytosis (p=0.005). DISCUSSION AND CONCLUSIONS: This study demonstrates a high prevalence of Mycoplasmataceae among patients with VAP, with a markedly lower prevalence among patients with suspected VAP in whom subsequent cultures refuted the diagnosis. The most common organism found, M. salivarium, is able to alter the functions of key immune cells. Mycoplasmataceae may contribute to VAP pathogenesis.

Cellular Microbiology of Mycoplasma canis.

Mycoplasma canis can infect many mammalian hosts but is best known as a commensal or opportunistic pathogen of dogs. The unexpected presence of M. canis in brains of dogs with idiopathic meningoencephalitis prompted new in vitro studies to help fill the void of basic knowledge about the organism's candidate virulence factors, the host responses that it elicits, and its potential roles in pathogenesis. Secretion of reactive oxygen species and sialidase varied quantitatively (P < 0.01) among strains of M. canis isolated from canine brain tissue or mucosal surfaces. All strains colonized the surface of canine MDCK epithelial and DH82 histiocyte cells and murine C8-D1A astrocytes. Transit through MDCK and DH82 cells was demonstrated by gentamicin protection assays and three-dimensional immunofluorescence imaging. Strains further varied (P < 0.01) in the extents to which they influenced the secretion of tumor necrosis factor alpha (TNF-α) and the neuroendocrine regulatory peptide endothelin-1 by DH82 cells. Inoculation with M. canis also decreased major histocompatibility complex class II (MHC-II) antigen expression by DH82 cells (P < 0.01), while secretion of gamma interferon (IFN-γ), interleukin-6 (IL-6), interleukin-10 (IL-10), and complement factor H was unaffected. The basis for differences in the responses elicited by these strains was not obvious in their genome sequences. No acute cytopathic effects on any homogeneous cell line, or consistent patterns of M. canis polyvalent antigen distribution in canine meningoencephalitis case brain tissues, were apparent. Thus, while it is not likely a primary neuropathogen, M. canis has the capacity to influence meningoencephalitis through complex interactions within the multicellular and neurochemical in vivo milieu.

Leucyl-tRNA synthetase editing domain functions as a molecular rheostat to control codon ambiguity in Mycoplasma pathogens.

Mycoplasma leucyl-tRNA synthetases (LeuRSs) have been identified in which the connective polypeptide 1 (CP1) amino acid editing domain that clears mischarged tRNAs are missing (Mycoplasma mobile) or highly degenerate (Mycoplasma synoviae). Thus, these enzymes rely on a clearance pathway called pretransfer editing, which hydrolyzes misactivated aminoacyl-adenylate intermediate via a nebulous mechanism that has been controversial for decades. Even as the sole fidelity pathway for clearing amino acid selection errors in the pathogenic M. mobile, pretransfer editing is not robust enough to completely block mischarging of tRNA(Leu), resulting in codon ambiguity and statistical proteins. A high-resolution X-ray crystal structure shows that M. mobile LeuRS structurally overlaps with other LeuRS cores. However, when CP1 domains from different aminoacyl-tRNA synthetases and origins were fused to this common LeuRS core, surprisingly, pretransfer editing was enhanced. It is hypothesized that the CP1 domain evolved as a molecular rheostat to balance multiple functions. These include distal control of specificity and enzyme activity in the ancient canonical core, as well as providing a separate hydrolytic active site for clearing mischarged tRNA.

Haemotropic mycoplasmas: what's their real significance in cats?

PRACTICAL RELEVANCE: The feline haemotropic mycoplasmas ('haemoplasmas') are a group of bacteria that can induce haemolytic anaemia in cats. Mycoplasma haemofelis is the most pathogenic of the species; 'Candidatus Mycoplasma haemominutum' and 'Candidatus Mycoplasma turicensis' are less pathogenic. The natural route of transmission of feline haemoplasma infection has not been confirmed, but fleas are implicated. When disease results, common clinical signs are pallor, lethargy, anorexia, weight loss, depression, dehydration and pyrexia. Treatment with tetracyclines or fluoroquinolones is usually effective at resolving clinical disease, but clearance of infection may not result. GLOBAL IMPORTANCE: The feline haemoplasmas are found worldwide, although prevalence varies geographically. PATIENT GROUP: Older male non-pedigree cats are believed to be at increased risk of haemoplasma infection, although younger cats are possibly more likely to show clinical disease associated with M haemofelis. CLINICAL CHALLENGES: The significance of feline haemoplasma infection is difficult to determine due to the existence of asymptomatic carrier cats and the variable pathogenicity of the haemoplasma species. Polymerase chain reaction (PCR) results should be interpreted in the light of the patient's clinical signs and haematological findings, infecting haemoplasma species and level of haemoplasma DNA present in the blood. Trial antibiotic treatment for haemoplasmosis may be warranted in suspected cases while awaiting PCR results. EVIDENCE BASE: Aspects of feline haemoplasmosis, particularly risk factors, pathogenesis, diagnostic methods and treatment, have been the focus of much recent research. This article draws on the current evidence base with a view to helping clinicians diagnose and manage cases more effectively.

Atividade antibacteriana in vitro de extratos brutos de espécies de Eugenia sp frente a cepas de molicutes / In vitro antibacterial activity of a crude extract of Eugenia species against mollicute strains

A necessidade da busca de novos antimicrobianos e o interesse nas espécies de molicutes vêm aumentandoconstantemente, principalmente pelo fato de serem os molicutes responsáveis pelo desenvolvimento de várias doençasimportantes, como é o caso de Mycoplasma arginini, M. hominis e Ureaplasma urealyticum. Estes dois últimos apresentam envolvimento em infecções do trato urogenital que podem levar à infertilidade e a complicações da gestação. M. arginini é um importante patógeno animal isolado de ovelhas, que causa uma doença respiratória grave chamada de "síndrome da tosse", e, eventualmente, zoonoses no homem. O gênero Eugenia pertence à família Myrtaceae, que compreende cerca de500 espécies de plantas com potencial terapêutico. Neste trabalho foram obtidos extratos brutos das folhas de Eugenia beaurepaireana, E. brasiliensis e E. umbelliflora, e estes foram testados pelo método de microdiluição em caldo frente às cepas de molicutes. Com isso, foi determinada a concentração inibitória mínima dos extratos das plantas, sendo de 1,25 mg/mL para E. beaurepaireana, 2,5 mg/mL para E. brasiliensis, e 5,0 mg/mL para E. umbelliflora contra as três espécies demolicutes igualmente. Dentre os resultados observados, destaca-se principalmente E. beaurepaireana, que apresentou um bom potencial antimicrobiano. Devido às patogenias importantes que estas bactérias causam, e à crescente resistência aosantimicrobianos tradicionais utilizados no tratamento, torna-se importante a busca por novos tipos de antibióticos dentro da biodiversidade brasileira. Existem raros estudos sobre as propriedades farmacológicas das plantas do gênero Eugenia, e nossos resultados são os primeiros relatos sobre sua atividade antimicrobiana frente a espécies de molicutes...

The need for new antimicrobials against mollicutes is constantly increasing, mainly because mollicutes are responsible for the development of several prominent diseases, including Mycoplasma arginini, M. hominis and Ureaplasma urealyticum. The last two are involved in urogenital tract infections that can lead to infertility and gestational complications. Meanwhile, M. arginini is an animal pathogen, isolated from sheep, that causes a serious respiratory disease, known as "coughing syndrome", and possibly zoonoses. The Eugenia genus belongs to the Myrtaceae family, which includes around 500 plant species with therapeutic potential. In this study, crude extracts from the leaves of E. beaurepaireana, E. brasiliensis and E. umbelliflora were obtained and tested by broth microdilution against mollicute strains. The minimal inhibitory concentration determined for the plant extracts was 1.25 mg/mL for E. beaurepaireana, 2.5 mg/mL for E. brasiliensis and 5.0 mg/mL for E. umbelliflora against three species of mollicutes. E. beaurepaireana in particular stands out as having good antimicrobial potential. Due to the serious diseases caused by these pathogens, as well as growing antibiotic resistance to conventional treatment, it is important research new antibiotic candidates derived from the Brazilian ecosystem. There exist very few studies of the pharmacological properties of the Eugenia plant genus and our report is the first to describe its antimicrobial properties against mollicutes...

[Lipid-associated membrane lipopeptides of M. arginini activate NF-kB by interacting with TLR2/1, TLR2/6, and TLR2/CD14].

Various strains of mycoplasmas cause activation of transcriptional factor NF-kB as a result of interaction with different combinations of Toll-like receptors (TLR). It is well known that the MALP-2 protein of M. fermentans activates the NF-kB through interaction with the TLR2/6, lipid-associated membrane lipopeptides (LAMPs) of M. penetrans through the TLR1/2, LAMPs of M. pneumoniae through combinations of Toll-like receptors (TLR2/6 and TLR1/2), and superantigene of M. arthritidis through the TLR2 and TLR4-dependent pathways. In this study, we defined specific Toll-like receptors for LAMPs of M. arginini. For carrying out the research we used cell lines 293-null, 293-hTLR2, 293-hTLR1/2, 293-hTLR2/CD14, 293-hTLR2/6, 293-hTLR4/ CD14-MD2 expressing certain combinations of TLR and their coreceptors. It was shown that LAMPs of M. arginini cause activation of NF-kB interacting with TLR2/1, TLR2/6 and TLR2/ CD14, but not with TLR2 alone or TLR4.