Mycoplasma Agalactiae Research Articles

Predominant Single Stable VpmaV Expression in Strain GM139 and Major Differences with Mycoplasma agalactiae Type Strain PG2.

Simple SummaryMALDI-ToF MS analysis demonstrates major differences between Mycoplasma agalactiae type strain PG2 and strain GM139. Evaluation of the Vpma (variable proteins of M. agalactiae) phenotypic profiles of these two strains reveals that unlike PG2, GM139 expresses a single Vpma, namely VpmaV, and without any visible phase variation. Although the presence of only one Vpma seems to be sufficient for the pathogenesis of the GM139 strain, the absence of phase variation can compromise immune system evasion and spread to other anatomical sites in the host.Although mycoplasmas have a reduced genome and no cell wall, they have important mechanisms for the antigenic variation in surface lipoproteins that modulate their interactions with the host. Mycoplasma agalactiae, the main etiological agent of contagious agalactia, has a multigene family involved in the high-frequency phase variation in surface lipoproteins called variable proteins of M. agalactiae (Vpmas). The Vpma lipoproteins are involved in the immune evasion, colonization, dissemination, and persistence of M. agalactiae in the host. In this paper, we evaluate the Vpma phenotypic profiles of two different strains of M. agalactiae, namely, GM139 and the type strain PG2, to assess possible correlations between Vpma phase variability and the geographic localization, animal origin, and pathogenicity of these two strains. Using monospecific Vpma antibodies against individual Vpmas in immunoblots, we demonstrate that, unlike PG2, which expresses six Vpma proteins with high-frequency phase variation, colonies of GM139 predominantly express VpmaV and do not exhibit any sectoring phenotype for any Vpma. Since VpmaV is one of the most important Vpmas for cell adhesion and invasion, its predominant sole expression in GM139 without high-frequency variation may be the basis of the differential pathogenicity of GM139 and PG2. Additionally, MALDI-ToF MS analysis also demonstrates significant differences between these two strains and their relatedness with other M. agalactiae strains.

Mycoplasma agalactiae ST35: a new sequence type with a minimal accessory genome primarily affecting goats

BackgroundMycoplasma agalactiae, causing agent of contagious agalactia, infects domestic small ruminants such as sheep and goats but also wild Caprinae. M. agalactiae is highly contagious and transmitted through oral, respiratory, and mammary routes spreading rapidly in an infected herd.ResultsIn an outbreak of contagious agalactia in a mixed herd of sheep and goats, 80% of the goats were affected displaying swollen udders and loss of milk production but no other symptom such as kerato-conjunctivitis, arthritis or pulmonary distress commonly associated to contagious agalactia. Surprisingly, none of the sheep grazing on a common pasture and belonging to the same farm as the goats were affected. Whole genome sequencing and analysis of M. agalactiae strain GrTh01 isolated from the outbreak, revealed a previously unknown sequence type, ST35, and a particularly small, genome size of 841′635 bp when compared to others available in public databases. Overall, GrTh01 displayed a reduced accessory genome, with repertoires of gene families encoding variable surface proteins involved in host-adhesion and variable antigenicity being scaled down. GrTh01 was also deprived of Integrative Conjugative Element or prophage, and had a single IS element, suggesting that GrTh01 has a limited capacity to adapt and evolve.ConclusionsThe lack of most of the variable antigens and the Integrative Conjugative Element, both major virulence- and host specificity factors of a M. agalactiae strain isolated from an outbreak affecting particularly goats, indicates the implication of these factors in host specificity. Whole genome sequencing and full assembly of bacterial pathogens provides a most valuable tool for epidemiological and virulence studies of M. agalactiae without experimental infections.

Assessment of animal diseases caused by bacteria resistant to antimicrobials: sheep and goats.

In this opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to the health of sheep and goats have been assessed. The assessment has been performed following a methodology based on information collected by an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play on antimicrobial resistance in clinical isolates of Staphylococcus aureus, Escherichia coli (non‐VTEC), Pseudomonas aeruginosa, Dichelobacter nodosus, Moraxella ovis, Mannheimia haemolytica, Pasteurella multocida, Mycoplasma ovipneumoniae, Mycoplasma agalactiae, Trueperella pyogenes, Streptococcus uberis, Bibersteinia trehalosi, Campylobacter fetus, Mycoplasma mycoides subsp. capri, Mycoplasma capricolum subsp. capricolum, Fusobacterium necrophorum is provided. Among those bacteria, EFSA identified E. coli with ≥ 66% certainty as being the most relevant antimicrobial‐resistant bacteria in sheep and goat in the EU based on the available evidence. The animal health impact of these most relevant bacteria, as well as their eligibility for being listed and categorised within the animal health law framework will be assessed in separate scientific opinions.

Detection of Mycoplasma agalactiae in Ticks (Rhipicephalus bursa) Collected by Sheep and Goats in Sicily (South-Italy), Endemic Area for Contagious Agalactia.

The aim of this preliminary study was to investigate the presence of Mycoplasma agalactiae (Ma) or other Contagious Agalactia (CA) causative organisms, in hard ticks infesting milking sheep and goats in endemic areas for CA in Sicily (South-Italy). Although there is accumulating evidence to support the role of ticks in the transmission of blood-borne haemoplasmas, information regarding their role in the transmission of CA, remains scarce. Ticks (n = 152) were collected from 25 lactating sheep and goats from three farms with previous outbreaks of CA. Microbiological and biomolecular, as well as serological analysis were performed on milk, tick, and serum samples, respectively. Rhipicephalus bursa species predominated, comprising 84.8% of the sampled ticks. Mycoplasma-like colonies were isolated from 5/56 (8.9%) tick pools and were identified as Ma by specific PCR and 16S rRNA gene sequencing. Unexpectedly, the organism was isolated from R. bursa ticks recovered only from animals whose milk tested negative for the pathogen. This preliminary demonstration suggests the potential role for ticks to act as a reservoir for the organisms, with potential involvement in the spread and maintenance of CA. Further work is required to determine the location of the organisms within the body of the ticks and to assess transmission potential.

Targeting Toll-Like Receptor 2: Polarization of Porcine Macrophages by a Mycoplasma-Derived Pam2cys Lipopeptide.

Toll-like receptor 2 (TLR2) ligands are attracting increasing attention as prophylactic and immunotherapeutic agents against pathogens and tumors. We previously observed that a synthetic diacylated lipopeptide based on a surface protein of Mycoplasma agalactiae (Mag-Pam2Cys) strongly activated innate immune cells, including porcine monocyte-derived macrophages (moMΦ). In this study, we utilized confocal microscopy, flow cytometry, multiplex cytokine ELISA, and RT-qPCR to conduct a comprehensive analysis of the effects of scalar doses of Mag-Pam2Cys on porcine moMΦ. We observed enhanced expression of activation markers (MHC class I, MHC class II DR, CD25), increased phagocytotic activity, and release of IL-12 and proinflammatory cytokines. Mag-Pam2Cys also upregulated the gene expression of several IFN-α subtypes, p65, NOS2, and molecules with antimicrobial activities (CD14, beta defensin 1). Overall, our data showed that Mag-Pam2Cys polarized porcine macrophages towards a proinflammatory antimicrobial phenotype. However, Mag-Pam2Cys downregulated the expression of IFN-α3, six TLRs (TLR3, -4, -5, -7, -8, -9), and did not interfere with macrophage polarization induced by the immunosuppressive IL-10, suggesting that the inflammatory activity evoked by Mag-Pam2Cys could be regulated to avoid potentially harmful consequences. We hope that our in vitro results will lay the foundation for the further evaluation of this diacylated lipopeptide as an immunopotentiator in vivo.

Mycoplasma agalactiae: The Sole Cause of Classical Contagious Agalactia?

Simple SummaryFor over thirty years, contagious agalactia has been recognized as a mycoplasma disease affecting small ruminants caused by four different pathogens: Mycoplasma agalactiae, Mycoplasma mycoides subsp. capri, Mycoplasma capricolum subsp. capricolum and Mycoplasma putrefaciens which were previously thought to produce clinically similar diseases. Today, with major advances in diagnosis enabling the rapid identification by molecular methods of causative mycoplasmas from infected flocks, it is time to revisit this issue. In this paper, we discuss and argue the reasons to support Mycoplasma agalactiae infection as the sole cause of contagious agalactia.Contagious agalactia (CA) is suspected when small ruminants show all or several of the following clinical signs: mastitis, arthritis, keratoconjunctivitis and occasionally abortion. It is confirmed following mycoplasma isolation or detection. The historical and major cause is Mycoplasma agalactiae which was first isolated from sheep in 1923. Over the last thirty years, three other mycoplasmas (Mycoplasma mycoides subsp. capri, Mycoplasma capricolum subsp. capricolum and Mycoplasma putrefaciens) have been added to the etiology of CA because they can occasionally cause clinically similar outcomes though nearly always in goats. However, only M. agalactiae is subject to animal disease regulations nationally and internationally. Consequently, it makes little sense to list mycoplasmas other than M. agalactiae as causes of the OIE-listed CA when they are not officially reported by the veterinary authorities and unlikely to be so in the future. Indeed, encouraging countries just to report M. agalactiae may bring about a better understanding of the importance of CA. In conclusion, we recommend that CA should only be diagnosed and confirmed when M. agalactiae is detected either by isolation or molecular methods, and that the other three mycoplasmas be removed from the OIE Manual of Diagnostic Tests and Vaccines in Terrestrial Animals and associated sources.

Detecting Mollicutes by PCR in goats in southwestern Bahia, Brazil.

Brazil has a herd of over 9 million goats, and the northeast of Brazil is home to over 93% of this herd. Caprine mycoplasmosis are widely disseminated worldwide, being highly contagious with high rates of morbidity and mortality, causing considerable economic loss to goat herders. In addition, there has been a lack of research using molecular testing to monitor the health and detect Mollicutes in this herd in Brazil. Therefore, the aim of this study is to associate animal management with the presence of the caprine origin Mollicutes in goats, in the southwest region of the state of Bahia, Brazil. A cross-sectional study was conducted on twelve farms, and statistical analyses were performed to identify associations between the presence of Mollicutes and the management of goats. Molecular testing identified Mollicutes class, Mycoplasma agalactiae (Ma) and M. conjunctivae (Mc), in the samples analyzed. Statistical associations were observed between animals from intensive livestock facilities and the presence of Mollicutes in nasal samples and dairy ranch animals and the presence of Mollicutes in ocular samples and animals from extensive ranching sites and positive results of Mollicutes in genital samples. We conclude that mycoplasmas are present in goat herds in the southwestern region of Bahia, which supports the need for more focused studies of mycoplasmas throughout the country. Our research also demonstrated the presence of two important opportunistic bacteria, Mc and Ma, and, to the best of our knowledge, this is the first time that M. conjunctivae was detected in Brazilian goats by molecular testing.

PK/PD Analysis of Marbofloxacin by Monte Carlo Simulation against Mycoplasmaagalactiae in Plasma and Milk of Lactating Goats after IV, SC and SC-Long Acting Formulations Administration.

Simple SummaryIn some countries like Spain and France, contagious agalactia (CA) is a highly relevant issue. CA is a mycoplasmosis affecting small ruminants and it is associated with a relevant economic impact on dairy. The poor efficacy of vaccines and their inability to prevent disease transmission is conducive to the use of antibiotics to control CA. However, only a few groups of antimicrobial agents are effective against these species, and selecting an adequate antimicrobial agent following the categorization of antibiotics made by the different international organisms (European Medicine Agency, World Health Organization) in veterinary medicine becomes a difficult task. The PK/PD approach is a useful tool to guide veterinarians on the appropriate targets through a rational selection of the best dose regimen of antimicrobial agents. In this study, marbofloxacin pharmacokinetics was studied after three routes of administration with two long-acting formulations. The minimum inhibitory concentrations (MIC) values of Mycoplasma agalactia isolated from goats affected by CA in Spain were calculated. The results show that systemic exposure achieved in lactating goats following these formulations provides rate of drug release that could be adequate to maintain effective plasma concentrations against M. agalactiae. The PK/PD analysis by Monte Carlo simulation showed that a dosage regimen from 8.47 to 11.57 mg/kg every 24 h could effectively treat goats affected by CA.Contagious agalactia is a mycoplasmosis affecting small ruminants that have become an important issue in many countries. However, PK/PD studies of antibiotics to treat this problem in lactating goats affected by Mycoplasma (M.) agalactiae, the main CA-causing mycoplasma are almost non-existent. The aims of this study were to evaluate the plasma and milk disposition of marbofloxacin in lactating goats after intravenous (IV), subcutaneous (SC) and subcutaneous poloxamer P407 formulations with and without carboxy-methylcellulose (SC-P407-CMC and SC-P407) administration. Marbofloxacin concentrations were analysed by the High Performance Liquid Chromatography (HPLC) method. Minimum inhibitory concentrations (MIC) of M. agalactiae field isolates from mastitic goat’s milk were used to calculate surrogate markers of efficacy. Terminal half-lives of marbofloxacin after IV, SC, SC-P407 and SC-P407-CMC administration were 7.12, 6.57, 13.92 and 12.19 h in plasma, and the half-lives of elimination of marbofloxacin in milk were 7.22, 7.16, 9.30 and 7.74 h after IV, SC, SC-P407 and SC-P407-CMC administration, respectively. Marbofloxacin penetration from the blood into the milk was extensive, with Area Under the Curve (AUCmilk/AUCplasma) ratios ranged 1.04–1.23, and maximum concentrations (Cmax-milk/Cmax-plasma) ratios ranged 0.72–1.20. The PK/PD surrogate markers of efficacy fAUC24/MIC and the Monte Carlo simulation show that marbofloxacin ratio (fAUC24/MIC > 125) using a 90% of target attainment rate (TAR) need a dose regimen between 8.4 mg/kg (SC) and 11.57 mg/kg (P407CMC) and should be adequate to treat contagious agalactia in lactating goats.

Identification of conserved Mycoplasma agalactiae surface antigens by immunoproteomics

Contagious agalactia represents one of the most relevant infectious diseases of dairy sheep, with Mycoplasma agalactiae being the primary etiological agent. The early, sensitive, and specific identification of infected animals, as well as the development of efficient prophylactic tools, remain challenging. Here, we present a comprehensive characterization of M. agalactiae antigens focusing on those shared among different isolates. Leveraging on previous proteomic data obtained on individual strains, we adopted a strategy entailing sample pooling to optimize the identification of conserved proteins that induce an immune response. The liposoluble proteins from previously characterized field isolates and the type strain PG2T were enriched by Triton X-114 fractionation, pooled, analysed by one-dimensional (1D) and two-dimensional (2D) electrophoresis, and subjected to western immunoblotting against sheep sera collected during natural infection with M. agalactiae. Immunodominant antigens were identified by Matrix-Assisted Laser Desorption-Time-Of-Flight-Mass Spectrometry (MALDI-TOF-MS). This combined immunoproteomic approach confirmed the role of several known immunogens, including P80, P48, and P40, and most variable surface proteins (Vpmas), and unveiled novel immunodominant, conserved antigens, including MAG_1000, MAG_2220, MAG_1980, phnD, MAG_4740, and MAG_2430. Genomic context, functional prediction, subcellular localization, and invariable expression of these proteins in all isolates suggest their possible involvement in bacterial pathogenicity and metabolism. Moreover, most of the identified antigens elicit a host humoral response since the early stages of infection, persisting for at least 270 days. The immunodominant, conserved antigen panel identified in this work supports the development of effective vaccines and diagnostic tools with higher sensitivity and specificity in all the natural infection stages.

Identification of conserved Mycoplasma agalactiae surface antigens by immunoproteomics

Identification of conserved Mycoplasma agalactiae surface antigens by immunoproteomics

Age and Seasonal Pattern of Contagious Agalactia in Small Ruminants in Ukraine.

IntroductionThe aim of the study was to determine how the spread of contagious agalactia in sheep and goats in the Odesa region depended on the age of the animals and the season.Material and MethodsFrom January 2016 to December 2018, 1,964 ewes and 1,484 nanny goats of different age groups were studied by ELISA for antibodies to Mycoplasma agalactiae.ResultsThe highest incidence of contagious agalactia was registered in one-year-old animals and was 59.7‒83.0%, two-year-old ruminants showed 17.0‒40.3% prevalence, in livestock at the age of 3–4 years no serological evidence of the disease was registered and in ewes and nanny goats older than 5–6 years 1.5–3.6% were infected. The most susceptible were young animals at the age of one-month (11.6‒14.5%). The first peak of the disease was recorded in March‒April (21.0‒26.1%), in the lambing period, which coincided with the beginning of lactation and the suckling period, and the second peak occurred in June–July (28.9‒34.2%), the period of maximum lactation and of manual milking of sheep and goats.ConclusionThe results of serological investigations indicate the circulation of M. agalactiae in small ruminants in the south of Ukraine. To avoid greater dissemination of the pathogen, appropriate measures should be applied and strategies for its control need to be drawn up.

Molecular Identification of Mycoplasma agalactiae in Iran Based on P30 Gene.

Mycoplasma agalactiae (M. agalactiae) is known as the main etiological agent of contagious agalactia (CA). The CA is a disease affecting dairy sheep and goats, the main characteristics of which include keratoconjunctivitis, arthritis, and mastitis. This pathogen results in milk production reduction and suppression, thereby leading to serious economic loss. In the present study, 125 sheep and goat samples were collected from 15 provinces of Iran. Cultural and molecular methods were used for sample characterization. After extracting genomic DNAs using the phenol/chloroform method, the PCR technique was employed to detect Mycoplasma genus in 163bp fragment of 16S rRNA gene (M-PCR) and M. agalactiae in 800bp fragment of conserve and specific P30 lipoprotein gene (P30-PCR) in cultural and clinical samples. Finally, to validate the experimental approach, a 375 bp amplicon of P80 lipoprotein was amplified using the MA-PCR. Out of 125 samples under investigation, 43 cases were positive, and Mycoplasma colonies were observed in the pleuropneumonia-like organisms agar culture. Based on the results of the M-PCR method, 61 specimens (out of 125 samples) were scored positive for Mycoplasma presence. Furthermore, 20 samples were positive according to the P30-PCR data. It should be mentioned that the MA-PCR was performed based on the P80 gene on 125 total samples to furtherverify the results for M.agalactiae detection. Based on the obtained data, P30 and P80 genes were presented and amplified in all Iranian M. agalactiae isolates (n=20). Our results indicated that the P30 gene was conserved and specific to all Iranian M. agalactiae isolates and this new P30-PCR method (as an MA-PCR technique) might be useful in the detection of this pathogen.

Bacterial Conjugation Protocol for Ruminant Mycoplasmas.

In Mycoplasma agalactiae, two simultaneous processes of DNA transfer have been described that require direct cell-to-cell contact and are similar to conjugation. One involves the self-transmission of an integrative conjugative element (ICE) while the second concerns the horizontal transfer of large and small fragments of chromosomal DNA. Here, we describe an optimized conjugation protocol for the horizontal transfer of ICE or chromosomal DNA carrying antibiotic resistance markers (i.e., tetracycline, gentamicin, puromycin) from donor to recipient mycoplasma cells. Calculation of the conjugation frequencies, selection and characterization of transconjugants are detailed. This protocol has been developed with M. agalactiae but has been successfully used for M. bovis and can be adapted to other related mycoplasma species.

An update on joint ill in sheep.

This focus article has been prepared by Vanessa Swinson of the APHA Small Ruminant Expert Group.

Ultrasensitive Electrochemical Impedance Detection of Mycoplasma agalactiae DNA by Low-Cost and Disposable Au-Decorated NiO Nanowall Electrodes.

Nanostructured electrodes detecting bacteria or viruses through DNA hybridization represent a promising method, which may be useful in on-field applications where PCR-based methods are very expensive, time-consuming, and require trained personnel. Indeed, electrochemical sensors combine disposability, fast response, high sensitivity, and portability. Here, a low-cost and high-surface-area electrode, based on Au-decorated NiO nanowalls, demonstrates a highly sensitive PCR-free detection of a real sample of Mycoplasma agalactiae (Ma) DNA. NiO nanowalls, synthesized by aqueous methods, thermal annealing, and Au decoration, by electroless deposition, ensure a high-surface-area platform for successful immobilization of Ma thiolated probe DNA. The morphological, chemical, and electrochemical properties of the electrode were characterized, and a reproducible detection of synthetic Ma DNA was observed and investigated by impedance measurements. Electrochemical impedance spectroscopy (EIS) ascribed the origin of impedance signal to the Ma DNA hybridization with its probe immobilized onto the electrode. The electrode successfully discriminates between DNA extracted from healthy and infected sheep milk, showing the ability to detect Ma DNA in concentrations as low as 53 ± 2 copy number μL-1. The Au-decorated NiO nanowall electrode represents a promising route toward PCR-free, disposable, rapid, and molecular detection.

Novel antigenic proteins of Mycoplasma agalactiae as potential vaccine and serodiagnostic candidates

Contagious agalactia (CA) is a serious disease notifiable to the World Organisation for Animal Health (OIE) causing severe economic losses to sheep and goat producers worldwide. Mycoplasma agalactiae, considered as its main etiological agent, inflicts a variety of symptoms in infected animals, including keratoconjunctivitis, mastitis, arthritis, ankylosis, abortions, stillbirths and granular vulvovaginitis. Despite its significance, developing a successful vaccine remains elusive, mostly due to the lack of knowledge about M. agalactiae’s pathogenicity factors and pathogenic mechanisms, including its "core" antigens. The aim of this study was to identify, characterize and express antigenic proteins of M. agalactiae as potential vaccine candidates. Predicted proteins of type strain PG2 were analyzed using bioinformatic algorithms to assess their cellular localization and to identify their linear and conformational epitopes for B cells. Out of a total of 156 predicted membrane proteins, three were shortlisted as potential antigenic surface proteins, namely [MAG_1560 (WP_011949336.1), MAG_6130 (WP_011949770.1) and P40 (WP_011949418.1)]. These proteins were expressed in recombinant Escherichia coli strains. Purified proteins were evaluated for their antigenicity using Western blot and ELISA using sera of M. agalactiae-naturally infected and non-infected sheep and goats. All 3 proteins were specifically recognized by the tested sera of M. agalactiae-infected animals. Also, specific rabbit antisera raised against each of these 3 proteins confirm their membrane localization using TritonX-114 phase partioning, Western and colony immunoblotting. In conclusion, our study successfully identified P40 (as proof of concept and validation) and two novel antigenic M. agalactiae proteins as potential candidates for developing effective CA vaccines.

A novel chimeric recombinant protein PDHB-P80 of Mycoplasma agalactiae as a potential diagnostic tool.

The aim of this study was to construct, expression of a novel recombinant chimeric protein consisting of Pyruvate dehydrogenase beta subunit (PDHB) and high antigenic region of integral membrane lipoprotein P80 of Mycoplasma agalactiae as a potential diagnostic tool. The full-length sequence of pdhb and a portion of antigenic regions of P80 were selected and analyzed by CLC main workbench 5.5 software. Several linkers and three dimensional structure of PDHB-P80 were compared to the native PDHB and analyzed to select a proper one for expression. The fusion gene sequence was optimized and synthesized in pMAT cloning vector. The synthetic pMAT-pdhb-p80 was digested using Bam HI and Sal I restriction enzymes and ligated into pMAL-p5X expression vector. The pMAL-pdhb-p80 construct was transfected into E.coli BL21 strain cells and expressed protein were purified using amylose resin. and the purified protein was analyzed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. In silico analysis demonstrated that fusion proteins using IgG4 middle hinge (CPSCP) with TM-score of 0.99 showed the higher similarity between three dimensional structure of PDHB before and after fusion with high antigenic region of P80. Successful cloning verified by PCR colony, double digestion and sequence analysis. Besides, SDS-PAGE analysis and Western blotting indicated and confirmed the expression of intact recombinant chimeric protein MBP-PDHB-P80 along with some truncated forms of the recombinant protein. it could be concluded that the fusion construct has a potential for serodiagnostic assay in future studies.

Gangrenous mastitis in dromedary camels in UAE caused by Streptococcus agalactiae

BackgroundMastitis is a disease of economic concern that affects dairy industry worldwide. This study aimed to investigate and identify possible etiologies encountered in an episode of acute gangrenous mastitis in lactating she-camels in Al Dhafra region, Abu Dhabi Emirate, United Arab Emirates (UAE).Beside the routine clinical examination, conventional bacteriological methods were used to isolate and identify possible aerobic/anaerobic bacterial or fungal pathogens from cultured milk samples collected from the mastitic she-camels. Moreover, quantitative real-time polymerase chain reaction (qPCR) was used for the detection of Mycoplasma agalactiae and Mycoplasma bovis strains, and the 16S rRNA gene was sequenced to confirm the isolation. The isolates were also tested for their susceptibility to antimicrobials.ResultsAcute gangrenous mastitis is reported in the dromedary camel herd with about 80% morbidity rate among lactating she-camels exhibited acute, painful hard swelling of affected teat, quarter or entire udder. About 41.7% of the infected animals were stamped out for culling due to complete or partial amputation of udder quarters. Streptococcus agalactiae was the sole isolated organism (6 isolates). The antimicrobial susceptibility testing revealed that, the Streptococcus agalactiae isolates were sensitive to both penicillin and ampicillin. Comparison of the 16S rRNA gene sequencing results by BLASTN confirmed the presence of Streptococcus agalactiae with high confidence (100% identity). Phylogenetic analysis indicated clustering of one isolate (CMAUAE accession number; MN267805.1) with Streptococcus agalactiae that infects multi-hosts including humans, while strains (CMBUAE to CMFUAE with accession numbers; MN267806.1 to MN267810.1 respectively) clustered with Streptococcus agalactiae that infects humans. No Mycoplasma spp was detected by qPCR analysis.ConclusionsIn the present study, the Streptococcus agalactiae was found to be the main cause of acute gangrenous mastitis in dromedary camels in UAE. More research should be done to investigate other possible causes of clinical or subclinical mastitis in dromedary camels in UAE.

Inferring Active Metabolic Pathways from Proteomics and Essentiality Data

SummaryHere, we propose an approach to identify active metabolic pathways by integrating gene essentiality analysis and protein abundance. We use two bacterial species (Mycoplasma pneumoniae and Mycoplasma agalactiae) that share a high gene content similarity yet show significant metabolic differences. First, we build detailed metabolic maps of their carbon metabolism, the most striking difference being the absence of two key enzymes for glucose metabolism in M. agalactiae. We then determine carbon sources that allow growth in M. agalactiae, and we introduce glucose-dependent growth to show the functionality of its remaining glycolytic enzymes. By analyzing gene essentiality and performing quantitative proteomics, we can predict the active metabolic pathways connected to carbon metabolism and show significant differences in use and direction of key pathways despite sharing the large majority of genes. Gene essentiality combined with quantitative proteomics and metabolic maps can be used to determine activity and directionality of metabolic pathways.

Determination of Contagious Agalactia in Sheep and Goats and Investigation of Antibiotic Susceptibility of Mycoplasma spp. Positive Isolates

Antibiotic resistance is one of the most important issues encountered globally in managing infectious diseases and is a potential problem in the treatment of Mycoplasma infections. The aims of this study were to: determine carrier rates in sick and healthy herds of sheep and goats; determine the presence in herds of carrier animals that were clinically asymptomatic for contagious agalactia and use antibiogram tests to investigate the susceptibility to antibiotics of Mycoplasma spp. positive isolates derived from sheep and goat ear swabs. The presence of contagious agalactia was diagnosed by analyzing ear swabs (n = 300, bacterial method, Mycoplasma spp.) and blood serum samples (n = 300, serological method [ELISA], Mycoplasma agalactiae) taken from sheep and goat herds located in Elazýð and Malatya provinces in eastern Turkey. The ELISA tests revealed seropositivity in 10 (3.33%) of 300 samples. In order to determine the most effective antibiotic for disease treatment, antibiogram testing was performed on 87 (29%) positive isolates that had been isolated from swab cultures. We determined tulathromycin (MIC 16 µg/mL) and tiamulin (MIC 16 µg/mL) to be the most effective antibiotics, whereas disease agents were resistant to trimethoprim/sulfamethoxazole and neomycin.