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Mycoplasma iowae is a worldwide spread and economically important avian pathogen that mostly infects turkeys. Currently, multi-locus sequence typing (MLST) serves as the gold standard method for strain identification in M. iowae. However, additional robust genotyping methods are required to effectively monitor M. iowae infections and conduct epidemiological investigations. The first aim of this study was to develop genotyping assays with high resolution, that specifically target M. iowae, namely a multiple-locus variable number of tandem-repeats analysis (MLVA) and a core genome multi-locus sequence typing (cgMLST) schema. The second aim was the determination of relationships among a diverse selection of M. iowae strains and clinical isolates with a previous and the newly developed assays. The MLVA was designed based on the analyses of tandem-repeat (TR) regions in the six serotype reference strains (I, J, K, N, Q and R). The cgMLST schema was developed based on the coding sequences (CDSs) common in 95% of the examined 99 isolates. The samples were submitted for a previously published MLST assay for comparison with the developed methods. Out of 94 TR regions identified, 17 alleles were selected for further evaluation by PCR. Finally, seven alleles were chosen to establish the MLVA assay. Additionally, whole genome sequence analyses identified a total of 676 CDSs shared by 95% of the isolates, all of which were included into the developed cgMLST schema. The MLVA discriminated 19 distinct genotypes (GT), while with the cgMLST assay 79 sequence types (ST) could be determined with Simpson's diversity indices of 0.810 (MLVA) and 0.989 (cgMLST). The applied assays consistently identified the same main clusters among the diverse selection of isolates, thereby demonstrating their suitability for various genetic analyses and their ability to yield congruent results., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Mycoplasma anserisalpingitidis is a waterfowl colonizing mycoplasma, mainly found in geese. In this study, we compared the whole genomes of five atypical M. anserisalpingitidis strains originating from China, Vietnam and Hungary, with the rest of the collection. Common methods used in the description of species are genomic analyses like the analysis of 16 S - intergenic transcribed spacer (ITS) - 23 S rRNA, of housekeeping genes, of the average nucleotide identity (ANI) and average amino acid identity (AAI) and phenotypic analyses like testing the growth inhibition and the growth parameters of the strains. The atypical strains showed notable genomic differences in all of the genetic analyses: on average ANI and AAI 95% (M. anserisalpingitidis ANI Minimum: 92.45, Maximum: 95.10; AAI Minimum: 93.34, Maximum: 96.37). The atypical strains formed a separate branch among the M. anserisalpingitidis strains in all phylogenetic studies. The small genome size and possibly higher mutation rate of the M. anserisalpingitidis species likely contributed to the observed genetic difference. Based on genetic analyses, the studied strains clearly represent a new genotype of M. anserisalpingitidis. The atypical strains showed slower growth in the medium containing fructose and three of the atypical strains showed diminished growth in the inhibition test. However, no definitive geno-phenotype associations were found regarding the fructose metabolism pathway in the atypical strains. The atypical strains are potentially at an early stage of speciation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Mycoplasma anserisalpingitidis infection is associated with the inflammation of the genital tract and cloaca, embryo lethality and decreased egg production in geese, leading to serious economic losses. This bacterium has so far been described in Europe and Asia. There is no commercially available vaccine against M. anserisalpingitidis, thus treatment of waterfowl mycoplasmosis relies mainly on antimicrobial therapy. However, M. anserisalpingitidis isolates with decreased susceptibility to macrolides and lincomycin have been reported before. The minimal inhibitory concentration (MIC) values of tilmicosin, tylosin, tylvalosin and lincomycin were determined against 82 M. anserisalpingitidis isolates originating from Hungary, Poland, China and Vietnam. Whole-genome sequence analyses revealed two mutations in the 23S rRNA coding regions and one mutation in the 50S ribosomal protein L22 coding gene possibly correlating with decreased susceptibility to the examined antibiotics. Mismatch amplification mutation assays coupled with melt analysis (melt-MAMAs) were designed to detect the nucleotide substitutions. This study is the first to describe resistance-related mutations in the goose pathogen M. anserisalpingitidis. The developed molecular assays support targeted antibiotic usage, hence their use may help to reduce the development and spread of antibiotic resistance., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Mycoplasma anserisalpingitidis infection is associated with the inflammation of the genital tract and cloaca, embryo lethality, and decreased egg production in geese, leading to serious economic losses. M. anserisalpingitidis has been detected mainly in Central and Eastern Europe, especially in Hungary, but the pathogen was identified recently in China, predicting it's worldwide occurrence. In this study, a novel multilocus sequence typing (MLST) scheme was developed to analyse phylogenetic relationships between M. anserisalpingitidis field isolates and clinical specimens originating from different geographical locations. Five loci (atpG, fusA, pgiB, plsY, and uvrA) were selected for the final MLST study. The examined 89 M. anserisalpingitidis samples yielded 76 unique sequence types with a 0.994 Simpson's index of diversity. The samples were originated from Hungary, Poland, Ukraine, China, and Vietnam. Phylogenetic analysis revealed the existence of three distinct clades (A-C) and six subclades within clade C. Generally, samples originating from the same geographical locations or livestock integration clustered together. Isolates in clade A showed the closest relationships to the M. anatis outgroup due to sequence similarity of the plsY locus. The highest genetic distance was observed in 5C among the subclades of clade C, containing the Asian and some Hungarian field isolates. The developed MLST assay revealed high diversity of the investigated M. anserisalpingitidis samples. The method proved to be a valuable and cost-effective tool for sequence typing of this waterfowl Mycoplasma species, enabling the better understanding of its phylogeny and providing a robust assay for future molecular epidemiological investigations., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia, a chronic respiratory disease, causing significant economic losses. Results from the 2015-2016 MycoPath pan-European antimicrobial susceptibility monitoring survey of M. hyopneumoniae are presented. In total, 147 M. hyopneumoniae porcine isolates from Belgium, France, Germany, Great Britain, Hungary, Italy, and Spain were tested. One isolate per farm was retained from pigs that had not been recently treated with antimicrobial agents. The minimal inhibitory concentration (MIC) of 13 antimicrobial agents was determined in a central laboratory using a broth microdilution method, with Friis Medium, incubated at 35 ± 1 °C for 5-12 days. M. hyopneumoniae NCTC 10110 was used as Quality Control. MIC 50 /MIC 90 (mg/L) values were: enrofloxacin 0.06/1; marbofloxacin 0.06/2; spiramycin 0.06/0.25; tulathromycin ≤0.001/0.004; gamithromycin 0.06/0.5; tylosin 0.016/0.06; tilmicosin 0.06/0.5; florfenicol 0.5/1; doxycycline 0.25/1; oxytetracycline 0.25/2; lincomycin 0.06/0.25; tiamulin 0.016/0.06 and valnemulin ≤0.001/0.004. Compared with the data from 2010 to 2012 MycoPath study (50 isolates), MIC 50/90 results were similar and the majority were within ± two dilution steps, except for the MIC 50 of oxytetracycline which is more than two dilution steps higher in the present study. Between-country comparisons show some differences in the MIC values for the fluoroquinolones, tulathromycin and tylosin, but the limited sample size per country precludes performing meaningful country comparisons for several countries. Standardized laboratory methods and interpretive criteria for MIC testing of veterinary mycoplasmas are clearly needed; there are currently no clinical breakpoints available to facilitate data interpretation and correlation of MICs with in vivo efficacy., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Mycoplasma synoviae (n = 26) and M. gallisepticum (n = 11) isolates were gained from 164 clinical samples collected from China, India, Indonesia, Malaysia, Philippines, Republic of Korea and Thailand. Most isolates were from commercial chicken production systems. A method of filtering (0.45 μm) samples immediately after collection was convenient allowing over a week for transit to the laboratory. Minimum inhibitory concentrations (MICs) were characterized by a broth microdilution method to enrofloxacin, difloxacin, oxytetracycline, chlortetracycline, doxycycline, tylosin, tilmicosin, tylvalosin, tiamulin, florfenicol, lincomycin, spectinomycin and lincomycin and spectinomycin combination (1:2). Increased MICs to various antimicrobials were seen in different isolates but appeared largely unrelated to the antimicrobial treatment histories. Overall, the results were similar to other MIC surveys around the world. Generally, low MICs to tetracyclines, tiamulin and tylvalosin were observed. Increased tilmicosin MICs were observed in both M. synoviae and M. gallisepticum isolates (≥64 μg/ml MIC 90 values) and this was seen in all isolates with high tylosin MICs. Increases in lincomycin MICs were mostly associated with increases in tilmicosin MICs. The results also suggested that antimicrobial use after mycoplasma vaccination may interfere with vaccine strain persistence and efficacy (field strains were more commonly observed in flocks that had treatments after vaccination) and this area warrants more investigation. The study shows that isolation and MIC determination can be done from remote locations and suggests that this may provide information that will allow more effective use of antimicrobials or other methods of control of avian mycoplasma in chickens (e.g. live vaccines) and therefore more responsible use of antimicrobials from a one health perspective., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Mycoplasma hyorhinis is a swine pathogen bacterium, which causes significant economic losses. The infection spreads through direct contact between the animals. Powerful genotyping methods like PCR based multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) are necessary to monitor the infections and to conduct epidemiological investigations; hence supporting the control of the disease. The aims of the present study were to examine M. hyorhinis isolates originating mainly from Hungary with MLST and MLVA developed in the study, and to compare the results of the two typing methods. To characterize 39 M. hyorhinis isolates and the type strain (NCTC 10,130), six house-keeping genes were selected for MLST and six tandem-repeat regions were chosen for MLVA. We were able to differentiate 31 sequence types and 37 genotypes within the 40 analyzed isolates by the MLST and the MLVA, respectively. With the combination of the two newly developed assays all examined isolates were distinguished with the exception of the ones originating from the same animal. The developed MLST assay provided a robust and high resolution phylogenetic tree, while the MLVA system is suitable for the differentiation of closely related isolates from the same farm, hence the assay is appropriate for epidemiologic studies., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Mycoplasma synoviae is one of the economically most significant avian Mycoplasma species. It can cause great financial losses to the poultry industry by inducing respiratory diseases, infectious synovitis, or eggshell apex abnormalities. There are different approaches to control M. synoviae infection. Although antimicrobial therapy cannot replace long-term solutions, like eradication and vaccination, this strategy can be effective in the short term, as adequate antibiotic treatment can relieve economic losses through the attenuation of clinical signs and reduction of transmission. Using broth microdilution method, minimal inhibitory concentration (MIC) values to fourteen antibiotics related to eight antimicrobial groups were determined in 96 M. synoviae strains. Whole genome sequencing and sequence analysis revealed mutations potentially associated with decreased susceptibility to fluoroquinolones, macrolides and lincomycin. Molecular markers responsible for the high MICs to fluoroquinolones were found in the gyrA, gyrB, parC and parE genes. Besides, single nucleotide polymorphisms identified in genes encoding the 23S rRNA were found to be responsible for high MICs to the 50S inhibitor macrolides and lincomycin, while amino acid change in the 50S ribosomal protein L22 could be associated with decreased susceptibility to macrolides. The revealed mutations can contribute to the extension of knowledge about the genetic background of antibiotic resistance in M. synoviae. Moreover, the explored potentially resistance-related mutations may serve as targets for molecular biological assays providing data of antibiotic susceptibility prior to the laborious and time-consuming isolation of M. synoviae strains., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Mycoplasma hyopneumoniae is the etiologic agent of porcine enzootic pneumonia, a contagious respiratory disease, causing significant economic losses worldwide. Antibiotic treatment is commonly utilised in the pig industry to control M. hyopneumoniae infection. Since the conventional antibiotic susceptibility test is time-consuming, taking up to weeks' period, antibiotics are usually empirically chosen. Certain single nucleotide polymorphisms in the parC (C239A/T, G250A) and gyrA (G242C, C247 T, A260 G) genes show correlation with decreased fluoroquinolone susceptibility by the change of the target site. Furthermore, the nucleotide alteration A2059 G in the 23S rRNA sequence correlates with significantly decreased macrolide and lincosamide susceptibility of M. hyopneumoniae. Mismatch amplification mutation assays (MAMA) and high resolution melt (HRM) analysis, capable to detect the mentioned resistance markers, were developed in the present study, in order to provide susceptibility data in a considerably shorter time than the conventional methods. The results of the MAMA and HRM assays were congruent with the results of the conventional antibiotic susceptibility method of the tested M. hyopneumoniae field isolates. The sensitivity of the MAMAs was 10 3 -10 4 copy numbers, while that of the HRM assay was 10 5 -10 6 copy numbers. To the best of our knowledge this was the first time that MAMA and HRM assays were developed for the rapid detection of decreased fluoroquinolone, macrolide or lincosamide susceptibility in M. hyopneumoniae strains., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Mycoplasma bovis is an important respiratory pathogen of cattle across Europe and is included in the MycoPath pan-European antimicrobial susceptibility monitoring programme. M. bovis strains (232) were isolated from cattle, not recently treated with antimicrobials, at diverse geographical locations in France, Great Britain, Hungary, Italy and Spain during 2014 to 2016. Only one isolate per farm and per outbreak was retained. For each isolate, the MICs of ten antimicrobials were determined in a central laboratory using a broth microdilution method with modified Eaton's medium and incubation at 35 °C ± 1 °C for 24 ± 6 h. MIC 50 /MIC 90 (mg/L) values for the 232 strains were: danofloxacin 0.25/1; enrofloxacin 0.5/8; marbofloxacin 1/4; gamithromycin >64/>64; spiramycin 8/16; tilmicosin >64/>64; tulathromycin >64/>64; tylosin 64/>64; florfenicol 4/8; oxytetracycline 8/32. Minor between-country differences in the MIC 90 values were observed for the fluoroquinolones, spiramycin and oxytetracycline, whilst the MIC values for the other compounds were similar. Spain and Italy had the higher MIC 90 values for the fluoroquinolones. Compared with the 2010-2012 study (156 isolates) results are similar, with an overall MIC 50 increase of at most one doubling dilution for enrofloxacin, spiramycin, tylosin, florfenicol and oxytetracycline. In contrast, the MIC 90 value for oxytetracycline decreased from >64 to 32 mg/L. Standardized laboratory methods and interpretive criteria for MIC testing of veterinary mycoplasmas are clearly needed; there are currently no clinical breakpoints available to facilitate data interpretation and correlation of MICs with in vivo efficacy., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Mycoplasma gallisepticum causes chronic respiratory disease and reproductive disorders in many bird species, resulting in considerable economic losses to the poultry industry. Maintenance of M. gallisepticum-free flocks is the most adequate method to control infection. To this end, monitoring systems and vaccination programs with live vaccine strains are applied worldwide. There is strong demand for efficient epidemiological investigation tools to distinguish M. gallisepticum strains in order to control disease. Up to now, multilocus sequence typing (MLST) has been regarded as gold standard for genotyping bacteria due to its good reproducibility and high discriminatory power. The aim of this study was to develop an MLST assay which can determine phylogenetic distances between M. gallisepticum strains. After analysing more than 30 housekeeping genes, six loci (atpG, dnaA, fusA, rpoB, ruvB, uvrA) were selected for the MLST assay due to their genomic location and high diversity. Examination of 130 M. gallisepticum strains with this MLST method yielded 57 unique sequence types (STs) with a 0.96 Simpson's index of diversity. Considering the large number of STs and high diversity index, this MLST method was found to be appropriate to discriminate M. gallisepticum strains. In addition, the developed method was shown to be suitable for epidemiological investigations, as it confirmed linkage between related strains from outbreaks in different farms. Besides, MLST also suggested high impact of extensive international trade on the spread of different M. gallisepticum strains. Furthermore this method can be used for differentiation among vaccine and field strains., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Mycoplasma hyorhinis is a common pathogen of swine causing mainly polyserositis and arthritis, but it has also been implicated as a cause of pneumonia. The economic losses due to M. hyorhinis infection could be reduced by antibiotic treatment. The aim of this study was to determine minimal inhibitory concentrations (MIC) of antibiotics potentially used to combat M. hyorhinis in swine production. Thirty-eight Hungarian M. hyorhinis strains isolated between 2014 and 2017 were examined by microbroth dilution tests for fifteen antimicrobial agents. Low MIC values of tetracyclines (MIC 50 0.078 μg/ml for doxycycline, ≤0.25 μg/ml for oxytetracycline) and pleuromutilins (MIC 50 0.156 μg/ml for tiamulin, ≤0.039 μg/ml for valnemulin) were detected against all strains. Fluoroquinolones (MIC 50 0.625 μg/ml), gentamicin (MIC 50 1 μg/ml) and florfenicol (MIC 50 2 μg/ml) inhibited the growth of Hungarian isolates at moderate MIC values. Most of the strains were inhibited by spectinomycin with low or moderate MIC values (MIC 50 4 μg/ml) except one strain (>64 μg/ml). Numerous isolates showed decreased susceptibility to macrolides and lincomycin (MIC 90 >64 for tylosin, tilmicosin, tulathromycin, gamithromycin, lincomycin, 8 μg/ml for tylvalosin). This study serves as evidence for the increasing resistance to macrolides and lincomycin in mycoplasmas, and also reports the occurrence of strains with extremely high MIC values to spectinomycin thus emphasizes the importance of the prudent use of antibiotics. Based on our results, tetracyclines and pleuromutilins are the most active compounds in vitro against the Hungarian M. hyorhinis strains., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Control of one of the most important avian mycoplasmas, Mycoplasma synoviae, and tracing the spread of the infection can be challenging as the pathogen is transmissible by both horizontal and vertical routes, and it can be disseminated through long distances via the hatching eggs, day-old chicks or pullets during intensive international trade. Genetic information provided by molecular typing methods support control programmes and epizootiologic studies. The aims of the present study were to develop a multi-locus variable number of tandem-repeats analysis (MLVA) method for the typing of M. synoviae isolates and to evaluate the currently used molecular typing methods which are applicable directly on clinical samples. Tandem repeat (TR) regions were selected from the whole genome sequence of the M. synoviae type strain (WVU1853) to characterise the genetic diversity of 86 M. synoviae strains originating from 15 countries. The strains were also submitted to multi-locus sequence typing (MLST) assays, vlhA gene sequence analysis and to assays designed to differentiate live vaccine strains from field strains. The developed MLVA involves the examination of seven TR regions and provides similar genetic resolution as the tested MLST assays by identifying 35 genotypes among the tested strains. Differentiation of the live vaccine strains from field strains was also successful with the developed assay. The provided MLVA method proved to be a highly discriminative, rapid and cost-effective alternative typing technique for the genetic characterisation of M. synoviae and it is also suitable for the complementation of live vaccine strain differentiating assays in ambiguous cases., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Mycoplasma hyopneumoniae is a swine pathogen bacterium, causing significant economic losses worldwide. Epidemiological investigations based on molecular typing methods support the prevention and eradication strategies for the control of M. hyopneumoniae, through tracing the spreading of the pathogen. The present study describes the genotyping of 44 M. hyopneumoniae strains isolated from Hungarian, Czech and Slovakian porcine lung samples by multi-locus sequence typing (MLST), multiple-locus variable-number tandem repeat analysis (MLVA) and analysing gene p146, and the evaluation of the used methods. The resolution of the three-gene (adk, rpoB, tpiA) and the seven-gene (efp, metG, pgiB, recA, adk, rpoB, tpiA) based MLST systems was identical with 27 sequence types. MLVA utilising loci P97-RR1 and Locus1 extended with the serine repeat numbers of gene p146 showed the highest resolution power among the studied methods differentiating 40 genotypes. The independent analysis of gene p146 revealed 31 different types among the isolates. High variability of M. hyopneumoniae strains was detected by the used typing methods. The results confirmed that utilization of the minimal MLST is suitable for phylogenetic analyses of M. hyopneumoniae strains. The MLVA method extended with the evaluation of serine repeat numbers of gene p146 is adequate for the resolution of genetic relationships within MLST groups. Examination of the p146 gene is suitable to complement both MLST and MLVA methods in order to refine closer genetic relationships., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Determining the antibiotic susceptibility profile of Mycoplasma bovis isolates in vitro provides the basis for the appropriate choice of antibiotics in the therapy. Traditionally, the antibiotic susceptibility examination of mycoplasmas is technically demanding, time-consuming and rarely performed in diagnostic laboratories. The aim of the present study was to develop rapid molecular assays to determine mutations responsible for elevated minimal inhibitory concentrations (MICs) to fluoroquinolones, tetracyclines, aminocyclitols, macrolides, lincosamides, phenicols and pleuromutilins in M. bovis. The nine mismatch amplification mutation assays (MAMA) and seven high resolution melt (HRM) tests designed in the present study enable the simultaneous detection of these genetic markers. The sensitivity of the assays varied between 10 2 -10 5 copy numbers/reaction. Cross-reactions with other mycoplasmas occurring in cattle were detected in assays targeting universal regions (e.g. 16S rRNA). Nevertheless, results of the novel method were in accordance with sequence and MICs data of the M. bovis pure cultures. Also, the tests of clinical samples containing high amount of M. bovis DNA were congruent even in the presence of other Mycoplasma spp. The presented method is highly cost-effective and can provide an antibiogram to 12 antibiotics in approximately 3-4 days when previous isolation of M. bovis is applied. In order to assure the proper identification of the genetic markers at issue, the regions examined by the MAMA and HRM tests are overlapping. In conclusion, the developed assays have potential to be used in routine diagnostics for the detection of antibiotic susceptibility in M. bovis., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Camel brucellosis is a widespread zoonotic disease in camel-rearing countries caused by Brucella melitensis and Brucella abortus. The aim of this study was the first genetic analysis of B. melitensis strains isolated from dromedary camels (Camelus dromedarius) using multiple-locus variable-number tandem repeat analysis (MLVA). MLVA 16 and its MLVA 8 and MLVA11 subsets were used to determine the genotypes of 15 B. melitensis isolates from dromedary camels (11 strains) and other host species (4 strains) from the United Arab Emirates and the results were then compared to B. melitensis MLVA genotypes from other parts of the world. Five, including two novel genotypes were identified with MLVA 8. MLVA 16 further discriminated these five genotypes to ten variants. The eleven camel isolates clustered into four main genetic groups within the East-Mediterranean and African clades and this clustering correlated with the geographic origin of the hosts (United Arab Emirates, Kingdom of Saudi Arabia and Sudan) and the date of their isolation. The camel strains were also genetically related to strains isolated from wild and domestic ruminants from their close habitat or from other parts of the world. Although limited number of strains were analysed, based on our data imported animals from foreign countries, local small ruminants and wildlife species are hypothesized to be the main sources of camel brucellosis in the United Arab Emirates. MLVA was successfully applied to determine the epidemiological links between the different camel B. melitensis infections in the United Arab Emirates and it can be a beneficial tool in future disease control programs., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Porcine brucellosis generally manifests as disorders in reproductive organs potentially leading to serious losses in the swine industry. Brucella suis biovar 2 is endemic in European wild boar (Sus scrofa) and hare (Lepus europeus, Lepus capensis) populations, thus these species may play a significant role in disease spread and serve as potential sources of infection for domestic pigs. The aim of this study was an epidemiologic analysis of porcine brucellosis in Hungary and a comparative analysis of B. suis bv. 2 strains from Europe using multiple-locus variable-number tandem repeat analysis (MLVA). MLVA-16 and its MLVA-11 subset were used to determine the genotypes of 68 B. suis bv. 2 isolates from Hungary and results were then compared to European MLVA genotypes. The analyses indicated relatively high genetic diversity of B. suis bv. 2 in Hungary. Strains isolated from hares and wild boars from Hungary showed substantial genetic divergence, suggesting separate lineages in each host and no instances of cross species infections. The closest relatives of strains from Hungarian wild boars and domestic pigs were mainly in the isolates from German and Croatian boars and pigs. The assessment of the European MLVA genotypes of wild boar isolates generally showed clustering based on geographic origin. The hare strains were relatively closely related to one another and did not cluster based on geographic origin. The limited relationships between geographic origin and genotype in isolates from hares might be the result of cross-border live animal translocation. The results could also suggest that certain B. suis strains are more adapted to hares. Across Europe, isolates from domestic pigs were closely related to isolates originating from both hares and wild boars, supporting the idea that wild animals are a source of brucellosis in domestic pigs., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Blood samples were collected from 100 shepherd dogs, 12 hunting dogs and 14 stray dogs (apparently healthy) in southern Hungary to screen for the presence of emerging tick-borne pathogens. Based on real-time PCR results, 14 dogs (11%) had single or dual haemoplasma infection, and a same number of samples were positive for Anaplasma phagocytophilum. In one sample Coxiella burnetii was molecularly identified, and 20.3% of dogs seroconverted to the Q fever agent. Rickettsaemia (sensu stricto) was also detected in one animal. This is the first molecular evidence of autochthonous infection of dogs with the above pathogens in Hungary. The relatively high prevalence of haemoplasma and anaplasma infection among non-pet dogs is suggestive of a prolonged carrier status and bacteraemia of these animals rendering them epidemiologically significant as potential reservoirs and sentinels for tick-borne infections., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Histophilus somni (former name: Haemophilus somnus) is a Gram-negative, facultative pathogen bacterium that colonises the mucous membranes of cattle and sheep, however it was also described in American bison and bighorn sheep. It can cause local or generalised diseases and asymptomatic carriers can also occur. The presence and the etiological role of this microorganism have not been confirmed in any other domesticated species yet. The purpose of this study was to prove the presence of H. somni in goats by bacterial isolation. Nasal, vaginal or praeputial swab samples were collected from 205 goats in 10 flocks. H. somni strains were isolated from 2 out of 10 flocks; in one flock 10 H. somni strains were isolated from the genital mucosa of 17 goats, while a single H. somni strain was cultured from a vagina of 26 animals in the other flock. Partial amplification and sequencing of the 16S rRNA gene of three H. somni strains verified the identification. The comparative examination of carbon source metabolism using the Biolog Microstation ID System (Biolog, Ca) showed a close relationship of the caprine strains, while they were less related to H. somni type strain CCUG-36157 of bovine origin. H. somni strains were isolated only in the oestrus season from goat flocks with sheep contact. This is the first paper on isolation of H. somni from goats.