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

BACKGROUND: Mycoplasma 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. RESULTS: In 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. CONCLUSIONS: The 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.

Short communication: Bovine mastitis caused by a multidrug-resistant, mcr-1-positive (colistin-resistant), extended-spectrum ß-lactamase-producing Escherichia coli clone on a Greek dairy farm.

We performed a survey aimed at analyzing milk samples collected from cows with mastitis for the presence of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli. Single-quarter mastitic milk samples obtained from 400 cows in 23 Greek dairy herds with a history of E. coli mastitis were processed for the selective isolation of ESBL-producing E. coli. The antimicrobial susceptibility of the ESBL-producing isolates was analyzed using agar disk diffusion, and minimum inhibitory concentrations of colistin were determined by broth microdilution. We used PCR followed by DNA sequencing to characterize the ß-lactamases and mcr-1 (colistin resistance) genes, and for phylotyping and multilocus sequence typing. We found a total of 89/400 (22.25%) E. coli isolates from 12/23 (52%) farms. Six isolates originating from 6 cows on a single farm were ESBL producers and were resistant to cefquinome, amoxicillin-clavulanic acid, aztreonam, ampicillin, and colistin. Five of these isolates were resistant to trimethoprim-sulfamethoxazole, and 5 to streptomycin. The 6 ESBL producers were mcr-1-positive and carried blaTEM-1 genes; 3 also carried blaCTX-M genes, and 3 carried blaSHV genes. All of the ESBL producers belonged to phylogroup A, multilocus sequence type ST666 (n = 5), or a single locus variant of ST666 (n = 1). To our knowledge, this is the first report of endemic bovine mastitis caused by mcr-1-positive, ESBL-producing E. coli. These results highlight the value of active surveillance of antimicrobial resistance not commonly tested by diagnostic laboratories for the early detection of novel resistant strains.

Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) in bulk tank milk, livestock and dairy-farm personnel in north-central and north-eastern Greece: Prevalence, characterization and genetic relatedness.

Recently, there has been an increased tendency towards raw-milk consumption, which may pose a consumer risk, due to the possible presence of human pathogenic microorganisms, such as Staphylococcus aureus and even methicillin-resistant S. aureus (MRSA). The prevalence of S. aureus and methicillin-resistant S. aureus (MRSA) was investigated in 40 dairy (cattle, sheep and goat) farms in northern Greece. S. aureus and MRSA were detected in 47.8% and 4.1% of the 387 samples (raw milk, farmers and animal samples) tested, respectively. Most (81.3%) of the MRSA isolates harbored the mecA gene, whereas the mecC or Panton-Valentine Leucocidin (PVL) genes were not detected. Seven spa types were identified, with t127 being the most prevalent. Spa type t034 (CC398) was isolated for the first time from livestock in Greece. Staphylococcal enterotoxin genes were detected in 93.8% of the MRSA isolates. The MRSA isolates were genetically diverse and were all capable of biofilm production. Our results confirm the lurking threat of MRSA in raw milk and dairy farms and suggest the need for surveillance programs starting at the farm level.

Pyogranulomatous pneumonia in goats caused by an undescribed Porphyromonas species, "Porphyromonas katsikii".

A yet-undescribed bacterial species, tentatively named "Porphyromonas katsikii," was isolated from individuals of a small goat herd with pyogranulomatous pneumonia during an outbreak of acute respiratory disease. The isolated bacteria grew in the form of black-pigmented colonies after 14 days of incubation under anaerobic conditions at 37°C on a tryptic soy blood agar medium. The bacteria were identified as a yet-undescribed Porphyromonas species by determination of the nucleotide sequence of the rrs 16S rRNA gene, and this species was tentatively named Porphyromonas katsikii. PCR amplification with specific primers for this yet-undescribed species revealed the presence of P. katsikii in the lung tissue of all affected animals, while no PCR signals were evidenced from the lungs of healthy goats or from goats with pasteurellosis caused by Mannheimia haemolytica. These data indicate P. katsikii as the causative agent of acute respiratory distress. P. katsikii is phylogenetically related to Porphyromonas somerae and Porphyromonas levii, which cause pathologies in humans and animals, respectively. P. katsikii was not detected by PCR from samples of the gingival pockets or of the faces of healthy goats.

Isolation and analysis of tetracycline-resistant Mycoplasma agalactiae strains from an infected goat herd in Cyprus - short communication.

A major concern with the use of tetracycline against mycoplasmas is the development of resistance. Infections in small ruminants due to tetracyclineresistant Mycoplasma agalactiae strains are becoming a frequent problem worldwide. In the present paper the detection and analysis of three tetracycline-resistant M. agalactiae strains, isolated from infected goats in Cyprus, are reported. The three field isolates were identified as M. agalactiae by polymerase chain reaction (PCR) showing 98% identity to the M. agalactiae PG2 reference strain. Furthermore, they were found sensitive to tylosin, enrofloxacin, spiramycin and lincomycin. In contrast, they were resistant to tetracycline. None of the putative genes [tet(M), tet(O) and tet(S)] that commonly contribute to high-level resistance to tetracycline could be amplified from their genome. Contrarily, the field isolates were found to carry ISMag1, an insertion sequence related to the IS30 family of mobile elements. Although ISMag1 is widely believed to induce high-frequency chromosomal rearrangements resulting in phenotypic changes of microorganisms, its potential role in tetracycline resistance of mycoplasmas requires further studies.

Stenotrophomonas maltophilia as a causal agent of pyogranulomatous hepatitis in a buffalo (Bubalus bubalis).

A 7-year-old female buffalo (Bubalus bubalis) from a local herd in Serres, northern Greece, was presented to a private veterinary clinic with a chronic loss of appetite for 15 days. The clinical examination revealed high fever (41.5 degrees C), lethargy, yellow discoloration of skin and mucous membranes, an abdomen that appeared to be empty, hyperactive rumen motility, and tachypnea. A biochemical profile revealed an elevated total bilirubin concentration and hepatic enzyme activities, whereas globulin, creatinine, and glucose concentrations were within the reference intervals. The animal received a 12-day course of treatment with intramuscular administration of ampicillin and corticosteroids. However, no significant clinical improvement was achieved, and the buffalo was euthanized. Gross necropsy lesions included serous atrophy of adipose tissue and hepatomegaly. Microscopic lesions included necrotizing pyogranulomatous hepatitis with thrombosis, hemorrhages, edema, and fibrosis. Small, nonpigmented, bacterial colonies were harvested in pure culture from the liver and were confirmed as Stenotrophomonas maltophilia by polymerase chain reaction. The bacterium was sensitive to ciprofloxacin, enrofloxacin, colistin, polymyxin, trimethoprim/sulfamethaxazole, and chloramphenicol. In contrast, resistance to ticarcillin, piperacillin, imipenem, ceftazidime, amikacin, gentamicin, tobramycin, and tetracycline was displayed. The bacterial strain carried the L1 metallo-beta-lactamase (L1) and tet35 genes, which contribute to high-level resistance to beta-lactams and tetracycline, respectively. Although S. maltophilia is widely believed to be a contaminant, the present report suggests that the isolation, identification, and susceptibility testing of this multidrug-resistant bacterium may be of clinical importance in diagnostic samples.

Molecular evidence of Anaplasma phagocytophilumin aborted goat fetuses and placenta.

Anaplasma phagocytophilum, transmitted by Ixodes ticks, is an intracellular pathogen of zoonotic interest. Regarding animals of veterinary importance, infection by this agent has been linked mainly to high fever, neutropenia, reduced milk production, but hemorrhagic diathesis, abortion and impaired spermatogenesis have also sporadically been reported. In Greece, A. phagocytophilum has been detected in dogs, ticks and humans, while so far only A. ovis had been detected in farm animals. Following the occurrence of multiple abortions in two goat farms in Northern Greece, samples were collected from aborted animals. Stomach contents and placental tissue from aborted animals tested positive for A. phagocytophilum by molecular assays and negative for other infectious and parasitic agents. Treatment with oxytetracycline LA stopped the abortions. In tick risk areas clinicians should consider A. phagocytophilum as a cause of abortion in goats.

Isolation and characterization of Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA) from milk of dairy goats under low-input farm management in Greece.

The presence of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) in raw milk is a challenge for veterinarians and public health professionals. In this study, we investigated the presence and clonality of S. aureus and MRSA in milk of individual dairy goats with subclinical mastitis reared under the low-input farming system in Greece and determined the isolates' enterotoxin gene carriage and their ability to form biofilms. S. aureus was isolated from 162 out of the 559 milk samples examined (29 %) and one isolate per S. aureus-positive sample was further characterized. S. aureus isolates were very closely related even among farms of distant geographical regions. Nine S. aureus isolates carried a functional mecA gene and were classified as MRSA. The S. aureus protein A (spa) typing in the MRSA isolates showed that four belonged to spa type t127 (44.4 %), three to t2049 (33.3 %) and two to t7947 (22.2 %). The spa type t7947 is reported for the first time in Greece. The MRSA isolates originated from two very distantly located farms, one located in the island of Skopelos and the other in Central Macedonia. Four of the MRSA isolates carried the staphylococcal enterotoxin genes sea or sec. Most of the isolates (92 % of S. aureus and 77.8 % of the MRSA) possessed moderate or weak biofilm-formation ability. Raw milk from low-input goat herds may serve as a potential vector of antimicrobial-resistant S. aureus to raw-milk consumers.

Clostridioides (Clostridium) Difficile in Food-Producing Animals, Horses and Household Pets: A Comprehensive Review.

Clostridioides (Clostridium) difficile is ubiquitous in the environment and is also considered as a bacterium of great importance in diarrhea-associated disease for humans and different animal species. Food animals and household pets are frequently found positive for toxigenic C. difficile without exposing clinical signs of infection. Humans and animals share common C. difficile ribotypes (RTs) suggesting potential zoonotic transmission. However, the role of animals for the development of human infection due to C. difficile remains unclear. One major public health issue is the existence of asymptomatic animals that carry and shed the bacterium to the environment, and infect individuals or populations, directly or through the food chain. C. difficile ribotype 078 is frequently isolated from food animals and household pets as well as from their environment. Nevertheless, direct evidence for the transmission of this particular ribotype from animals to humans has never been established. This review will summarize the current available data on epidemiology, clinical presentations, risk factors and laboratory diagnosis of C. difficile infection in food animals and household pets, outline potential prevention and control strategies, and also describe the current evidence towards a zoonotic potential of C. difficile infection.

Prevalence, antimicrobial susceptibility and characterization of Staphylococcus aureus and methicillin-resistant Staphylococcus aureus isolated from dairy industries in north-central and north-eastern Greece.

Staphylococcus aureus is an important cause of food intoxication, whereas methicillin-resistant S. aureus (MRSA) constitutes a serious public-health concern due to its ability to colonize and infect humans and animals. S. aureus and MRSA have often been isolated from milk and dairy products. The objectives of this study were to estimate the prevalence and the antimicrobial resistance of S. aureus and MRSA in four Greek dairy industries, to identify virulence factors of MRSA isolates and to describe their genetic diversity, in order to identify possible epidemiological links and evaluate the risk of MRSA dissemination to the community. S. aureus was isolated from 67 out of 305 (22.0%) dairy industry samples (bulk-tank milk, dairy products, employee nasal swabs and equipment/surface swabs). Almost all (99%) of the 227 corresponding S. aureus isolates (approximately 4 isolates per positive sample) were resistant to at least one antimicrobial and 22% were multi-drug resistant (MDR). MRSA were isolated from 11 different samples (3.6%) originating from three of the dairy plants. All MRSA isolates were capable of forming biofilms, while staphylococcal enterotoxin (SE) genes were detected in 91% of the MRSA isolates, with sec being the most frequent. All of the MRSA isolates harbored the mecA gene but the mecC and Pandon-Valentine leucocidin (PVL) genes were not detected. Pulse-Field Gel Electrophoresis (PFGE) analysis showed genetic diversity among the MRSA isolates and indicated clonal circulation in one of the dairy plants. Seven spa types were identified among the MRSA isolates with the most prevalent (t065) isolated only in one dairy plant. Certain spa types (t065, t337 and t3536) were isolated for the first time in Greece. The presence of MDR, biofilm-forming and enterotoxigenic MRSA strains in dairy plant facilities may lead to their dissemination to the community, but also to staphylococcal food poisoning, when conditions are favorable. The study's findings highlight the need for continuous monitoring of the dairy production chain, the need for re-evaluating the implemented cleaning and sanitizing processes and the adoption of preventive strategies in order to minimize public-health risks.

Microbiological quality of indoor and outdoor swimming pools in Greece: investigation of the antibiotic resistance of the bacterial isolates.

During 1997-2005, the microbiological quality and susceptibility of bacterial isolates of swimming pool waters were investigated. A total of 462 water samples were collected from three indoor swimming pools (a teaching pool, a competition public pool, a hydrotherapy pool) and two outdoor swimming pools (a hotel semi-public and a residential private pool) in Northwestern Greece. All water samples were analyzed for the presence of bacteria, protozoa and fungi and susceptibility tests were performed for the bacterial isolates. Sixty-seven percent of the examined water samples conformed to the microbiological standards and 32.9% exceeded at least one of the indicated limits. Out of 107 bacterial isolates, 38 (35.5%) resistant strains were detected. Multi-resistant Pseudomonas alcaligenes, Leuconostoc, and Staphylococcus aureus (isolated from the teaching pool), Staphylococcus wernerii, Chryseobacterium indologenes and Ochrobactrum anthropi (isolated from the competition pool), Pseudomonas aeruginosa, P. fluorescens, Aeromonas hydrophila, Enterobacter cloacae, Klebsiella pneumoniae and S. aureus (isolated from the hydrotherapy pool) and A. hydrophila (isolated from the hotel pool) were detected. The swimming pool with the poorest microbiological quality (THC 500 cfu/ml in 12.1% of the samples, P. aeruginosa counts 1500 cfu/100 ml in 6% of the samples) and the highest prevalence of multi-resistant isolates (73.6%) was the hydrotherapy pool. No Cryptosporidium or Giardia cysts and no Legionella, Mycobacteria and Salmonella were detected, but there were isolations of Candida albicans, Aspergillus spp., Mucor spp., Alternaria spp., Rhizopus spp., Trichophyton spp., and Penicillium spp.

Fly repellency using deltamethrin may reduce intramammary infections of dairy cows under intensive management.

The aim of this study was to assess the possible effect of the fly repellent deltamethrin on the full microbiological profile of the intramammary infections as well as on the somatic cell count in one Greek dairy cattle farm under intensive management, during peak fly season. Fifty five multiparous cows, stabled in the same farm, were randomly divided in three groups; cows of Group A were dressed on their back with deltamethrin, cows of Group B remained untreated within the same pen and cows of Group C remained untreated in a separate pen. Clinical records of the herd showed a history of clinical and subclinical mastitis (especially during spring and early summer) and fly infestation. Deltamethrin reduced fly population, landing on the cows of Group A, facilitating a significant decrease of S. aureus, coagulase negative staplylococci, E. coli intramammary infections and somatic cell count, throughout the study period. Consequently, there is a relationship between fly abundance and transmission of bacteria resulting in intramammary infections and mastitis in dairy cows. The use of the pyrethroid deltamethrin reduced fly abundance and therefore, the possibility of transmitting pathogenic bacteria in the mammary gland.

High efficiency electrotransformation of Lactococcus lactis spp. lactis cells pretreated with lithium acetate and dithiothreitol.

BACKGROUND: A goal for the food industry has always been to improve strains of Lactococcus lactis and stabilize beneficial traits. Genetic engineering is used extensively for manipulating this lactic acid bacterium, while electropolation is the most widely used technique for introducing foreign DNA into cells. The efficiency of electrotransformation depends on the level of electropermealization and pretreatment with chemicals which alter cell wall permeability, resulting in improved transformation efficiencies is rather common practice in bacteria as in yeasts and fungi. In the present study, treatment with lithium acetate (LiAc) and dithiothreitol (DTT) in various combinations was applied to L. lactis spp. lactis cells of the early-log phase prior to electroporation with plasmid pTRKH3 (a 7.8 kb shuttle vector, suitable for cloning into L. lactis). Two strains of L. lactis spp. lactis were used, L. lactis spp. lactis LM0230 and ATCC 11454. To the best of our knowledge these agents have never been used before with L. lactis or other bacteria. RESULTS: Electrotransformation efficiencies of up to 105 transformants per mug DNA have been reported in the literature for L. lactis spp.lactis LM0230. We report here that treatment with LiAc and DDT before electroporation increased transformation efficiency to 225 +/- 52.5 x 107 transformants per mug DNA, while with untreated cells or treated with LiAc alone transformation efficiency approximated 1.2 +/- 0.5 x 105 transformants per mug DNA. Results of the same trend were obtained with L. lactis ATCC 11454, although transformation efficiency of this strain was significantly lower. No difference was found in the survival rate of pretreated cells after electroporation. Transformation efficiency was found to vary directly with cell density and that of 1010 cells/ml resulted in the highest efficiencies. Following electrotransformation of pretreated cells with LiAc and DDT, pTRKH3 stability was examined. Both host-vector systems proved to be reproducible and highly efficient. CONCLUSION: This investigation sought to improve still further transformation efficiencies and to provide a reliable high efficiency transformation system for L. lactis spp. lactis. The applied methodology, tested in two well-known strains, allows the production of large numbers of transformants and the construction of large recombinant libraries.

Serological prevalence of Coxiella burnetii in dairy goats and ewes diagnosed with adverse pregnancy outcomes in Greece.

INTRODUCTION: Coxiella burnetii is an obligatory intracellular bacterial pathogen causing the zoonotic disease Q fever. The most common reservoirs of C. burnetii are wild mammals, birds and ticks. Pregnant domestic ruminants infected with this bacterium are also a major source of human infection. MATERIAL AND METHODS: The serological prevalence of C. burnetii in goats and sheep diagnosed with adverse pregnancy outcomes was assessed by undertaking a survey on 800 dairy goats and 800 dairy ewes reared in four different regions of Greece (Macedonia, Thrace, Thessaly, and Peloponnese). A stratified sampling was carried out, taking also as a criterion the age of the animals. Serum antibodies were analyzed by a commercial ELISA according to the manufacturer's recommendations. RESULTS: Generally, there was a statistically significantly higher serological prevalence of C. burnetii (14.4%) in goats compared to sheep (8%). Serological prevalence was higher in adults (15.5% in goats and 8.5% in sheep) compared to yearlings (7.4% in goats and 4.6% in sheep). The prevalence increased significantly with age only in goats. Finally, all animals reared in Peloponnese had a prevalence significantly higher (21% in goats and 18% in sheep) than animals reared in the other three regions. CONCLUSIONS: To the best of the authors' knowledge, this is the first report that associates C. burnetii with reproductive disturbances of domestic ruminants in Greece. However, considering the importance of coxiellosis for public health, further investigations are required on its epidemiology regarding abortion, premature delivery, stillbirth and weak offspring in small ruminants, as well as in other domestic and wild animal species.

Evaluation of the in-field efficacy of oregano essential oil administration on the control of neonatal diarrhea syndrome in calves.

The objective of this study was to evaluate under field conditions, whether daily administration of oregano essential oil is effective in preventing and/or diminishing the severity of neonatal diarrhea syndrome in calves aged less than 15days. Ninety-one newborn calves from three dairy farms were assigned into two groups; "Eco" group (n=46) calves were drenched with Greek oregano (Origanum vulgare ssp. Hirtum) essential oil (ECODIAR® liquid 5%) at the dose of 12.5mg/kg body weight once per day until the age of 10days. "Conts" group (n=45) calves were left untreated and served as controls. All animals were monitored daily for the incidence of diarrhea until the age of 15days and their fecal score was recorded. Fecal samples were collected on days 3, 6 and 10 for microbiological and parasitological evaluation. Average fecal score throughout the experiment, incidence of diarrhea, duration and severity of diarrhea episodes were significantly lower in Eco group compared to the controls. Daily administration of oregano essential oil in calves for the first 10days of their life effectively diminishes the severity of naturally acquired diarrhea under field conditions and, under certain hygiene practices, possess a preventive effect against neonatal diarrhea syndrome.

Determination of bacteriocin activity with bioassays carried out on solid and liquid substrates: assessing the factor "indicator microorganism".

BACKGROUND: Successful application of growth inhibition techniques for quantitative determination of bacteriocins relies on the sensitivity of the applied indicator microorganism to the bacteriocin to which is exposed. However, information on indicator microorganisms' performance and comparisons in bacteriocin determination with bioassays is almost non-existing in the literature. The aim of the present work was to evaluate the parameter "indicator microorganism" in bioassays carried out on solid -agar diffusion assay- and liquid -turbidometric assay- substrates, applied in the quantification of the most studied bacteriocin nisin. RESULTS: The performance of characterized microorganisms of known sources, belonging to the genera of Lactobacillus, Pediococcus, Micrococcus and Leuconostoc, has been assessed in this work in the assays of plate agar diffusion and turbidometry. Dose responses and sensitivities were examined and compared over a range of assay variables in standard bacteriocin solutions, fermentation broth filtrates and processed food samples. Measurements on inhibition zones produced on agar plates were made by means of digital image analysis. The data produced were analyzed statistically using the ANOVA technique and pairwise comparisons tests. Sensitivity limits and linearity of responses to bacteriocin varied significantly among different test-microorganisms in both applied methods, the lower sensitivity limits depending on both the test-microorganism and the applied method. In both methods, however, only two of the nine tested microorganisms (Lactobacillus curvatus ATCC 51436 and Pediococcus acidilactici ATCC 25740) were sensitive to very low concentrations of the bacteriocin and produced a linear-type of response in all kinds of samples used in this work. In all cases, very low bacteriocin concentrations, e.g. 1 IU/ml nisin, were more accurately determined in the turbidometric assay. CONCLUSION: The present work shows that in growth inhibition techniques used in bacteriocin quantification, the choice of the indicator microorganism is critical. Evaluation of sensitivity levels and type of produced responses showed that they can vary widely among different test-microorganisms and different applied methods, indicating that not all microorganisms can be used successfully as indicators and that measurements of growth inhibition in liquid media produce more reliable results.

Genome Sequences of Two Pseudorabies Virus Strains Isolated in Greece.

Pseudorabies virus (species Suid herpesvirus 1) belongs to the genus Varicellovirus, subfamily Alphaherpesvirinae, family Herpesviridae, and is the causative agent of an acute and frequently fatal disease that affects mainly pigs. Here, we report the genome sequences of two strains of this virus isolated in Greece in 2010.

In vitro susceptibilities of caprine Mycoplasma agalactiae field isolates to six antimicrobial agents using the E test methodology.

The minimum inhibitory concentrations (MICs) of enrofloxacin, ciprofloxacin, spectinomycin, tetracycline, spiramycin and erythromycin against 30 caprine Greek isolates of Mycoplasma agalactiae were determined using E test methodology. The E test strips were placed on Eaton's agar medium without antimicrobials and phenol red. MICs were then read by determining where the growth inhibition zone intersected with the MIC scale on the strip. An MIC value of 8 µg/mL was considered as a guide to mycoplasma resistance. All isolates were sensitive to fluoroquinolones (MIC50, 0.19 g/mL; MIC90, 0.38 µg/mL; highest MIC, 0.5 µg/mL), spectinomycin (MIC50, 0.5 µg/mL; MIC90, 1 µg/mL; highest MIC, 1 µg/mL), and spiramycin (MIC50, 1 µg/mL; MIC90, 1.5 µg/mL; highest MIC, 2 µg/mL). Two strains exhibited resistance to tetracycline (MIC 32 µg/mL) but these were not found to carry any of the tet(M), tet(O), and tet(S) resistance genes. Finally all isolates expressed resistance to erythromycin (MIC50, 128 µg/mL; MIC90, >256 µg/mL).

Ciprofloxacin-resistant Escherichia coli isolated from the intestinal microbiota of goats in Greece in the absence of selective pressure.

The presence of ciprofloxacin resistance commensal Escherichia coli (C-R-Ec) was determined for goats in the absence of selective pressure in Northern and Central Greece. The C-R-Ec was categorized in 3 groups with respect to their phenotypic resistance to other antibiotics as well as the carriage of antibiotic resistance genes. The first group consisted of 7 C-R-Ec that were found also resistant to tetracycline. Among them tet(B) (n = 7), qnr(S) (n = 7), and qnr(B) (n = 3) producers were identified by polymerase chain reaction. The second group consisted of 10 C-R-Ec that were found sensitive to all other antibiotics, and their phenotypic resistance to ciprofloxacin was not attributed to the presence of resistance genes. Finally, the third group consisted of 2 C-R-Ec also resistant to sulfamethoxazole. These strains were not carrying any transferable elements that contribute to resistance either to ciprofloxacin or to sulfamethoxazole. This is the first report of ciprofloxacin-resistant E. coli isolated from goats in Greece.

An outbreak of caprine meningoencephalitis due to Escherichia coli O157:H7.

Five 1-month-old kid goats from a local herd in Kozani (northwest Greece) developed neurological disorders characterized by decreased appetite, ataxia, and head pressing. The animals received a 3-day course of treatment with intramuscular administration of enrofloxacin and ketoprofen. However, no significant clinical improvement was achieved, and 2 kids died. The remaining 3 animals were euthanized, and a necropsy was performed within 1 hr. Macroscopic lesions were confined to the central nervous system, with congestion and petechiae in the meninges. Microscopic lesions in all 3 animals revealed multifocal acute meningoencephalitis characterized by infiltrations composed of mononuclear inflammatory cells, lesser numbers of lymphocytes, and occasionally neutrophils and eosinophils. Additionally, in the kidney, there was multifocal expansion of the glomerular tufts by eosinophilic amorphous material, multifocal interstitial hemorrhages, and multifocal glomerular hypercellularity. The above noted lesions are consisted with an acute ongoing nephropathy indicative of a septicemic-toxemic procedure at its primary stages. Small, gray bacterial colonies, 3-4 mm in diameter, were obtained in pure culture from the brain of all 3 necropsied animals and were confirmed as Escherichia coli O157:H7 by use of phenotypic and genotypic methods. The isolates were sensitive to cefuroxime, ceftazidime, and gentamicin. In contrast, resistance to enrofloxacin, trimethoprim-sulfamethoxazole, and tetracycline was displayed. Additionally the bacterial isolates were found to carry a plasmid that harbored qnrS, sulII, and tetB genes that contribute to high-level resistance to fluoroquinolones, co-trimoxazole, and tetracycline, respectively.