Identification and characterization of methicillin-resistant Staphylococcus aureus (MRSA) from Austrian companion animals and horses

The aim of this study was to investigate the antimicrobial resistance, resistance gene patterns and genetic relatedness of a collection of Austrian methicillin-resistant Staphylococcus aureus (MRSA) isolates from companion animals and horses.A total of 89 non-repetitive MRSA isolates collected during routine veterinary microbiological examinations from April 2004 to the end of 2012, and one isolate from 2013 were used for this study. The presence of mecA and other resistance genes was confirmed by PCR. Isolates were genotyped by spa typing, two multiple-locus variable-number tandem repeat analyses (MLVA) analyses, SCCmec typing and multilocus sequence typing (MLST). PCR targeting Panton-Valentine leukocidin (PVL) and detection of staphylococcal enterotoxins (SE), toxic shock syndrome toxin (TSST) was performed using PCR assays. Antimicrobial susceptibility testing was performed.Five sequence types (STs—ST398, ST254, ST22, ST5 and ST1), SCCmec types II, IVa, V, and non-type-abele, 8 spa-types (t003, t011, t036, t127, t386, t1348, and t4450), and two isolates could not be assigned, 21 MLVA-14Orsay types Multiplex-PCR MLVA (mMLVA) displayed 17 different MLVA types.The present study is the most comprehensive dealing with MRSA from Austrian companion animals and horses. The results confirm that MRSA ST398 is present in a wide range of animal species and is predominant especially in horses. In other companion animals it is unclear whether the infections with the different MRSA isolates investigated in the present study truly represents a rare phenomenon or may be an emerging problem in companion animals.     

Methicillin resistant Staphylococcus aureus (MRSA) infection in a dog in the Netherlands

In the Netherlands, methicillin resistant Staphylococcus aureus (MRSA) are regularly isolated from humans. We present the first isolation of MRSA from animal origin in the Netherlands. A coagulase positive staphylococcus was cultured from an infected wound in a Dutch dog that recently underwent surgery abroad. The staphylococcus was resistant to methicillin, ampicillin, amoxycillin + clavulanic acid, cephalexin, erythromycin, lincomycin, tetracycline, gentamicin and enrofloxacin. It was identified as S. aureus by fermentation of mannitol and Martineau-PCR. The presence of mecA was confirmed by PCR.     

Methicillin‐Resistant Staphylococcus aureus (MRSA) ST398 in Pig Farms and Multispecies Farms

During the last few years, methicillin‐resistant Staphylococcus aureus (MRSA) ST398 has been isolated frequently from livestock, especially from pigs and to a lesser extent from cattle and poultry. To gain insight into the distribution of this bacterium in pig farms versus multispecies farms, 30 Belgian farms (10 pig, 10 pig/poultry and 10 pig/cattle farms) were screened for the presence of MRSA. On each farm, 10 nasal swabs were taken from pigs. When present, cattle (n = 10) were sampled in the nares and poultry (n = 10) in the nares, earlobes and cloaca. A selection of the obtained isolates were further characterized using multilocus sequence typing (MLST), spa typing, SCCmec typing, pulsed field gel electrophoresis (PFGE), multiple‐locus variable‐number tandem repeat analysis (MLVA) and antimicrobial susceptibility testing. On 26 of 30 farms, MRSA was isolated from pigs. Furthermore, MRSA was also isolated from poultry and cattle on one pig/poultry and five pig/cattle farms, respectively. All tested MRSA isolates belonged to ST398. Eight spa types (t011, t034, t567, t571, t1451, t2974, t3423 and t5943) were detected, among which t011 predominated. SCCmec cassettes type IVa and V were present in 20% and 72% of the isolates, respectively. When combining the results of the two remaining typing methods, PFGE and MLVA, eighteen genotypes were obtained of which one genotype predominated (56% of the positive farms). All MRSA isolates were resistant to tetracycline. Resistance to trimethoprim, aminoglycosides, macrolides, lincosamides, fluoroquinolones and chloramphenicol was also observed. In conclusion, there was no effect of the farm type on the MRSA status of the pigs. A statistically significant difference was observed when comparing the pig/poultry or the pig/cattle MRSA status on the multispecies farms. Additionally, a wide variety of MRSA ST398 strains was found within certain farms when combining different typing methods.     

Methicillin‐resistant Staphylococcus aureus keratitis in a dog

The purpose of this study is to report a case of methicillin‐resistant Staphylococcus aureus (MRSA) keratitis in a dog. A 7‐year‐old intact male American cocker spaniel that had undergone removal of a nictitating gland was referred for severe ulcerative keratitis. Slit‐lamp examination showed swelling of the eyelid, mucopurulent discharge, conjunctival injection and chemosis, diffuse corneal edema and opacity, and a deep ulcer in central cornea. Gram staining of discharge from the eye demonstrated Gram‐positive cocci. Despite topical ofloxacin, oxytetracycline and polymyxin B ophthalmic solution and intravenous cefazolin, there was no improvement. Cultures revealed MRSA that was sensitive only to chloramphenicol, vancomycin, lincomycin, and clindamycin. The antibiotic regimen was changed to topical and systemic chloramphenicol. After 9 days of treatment, although inflammation started to be resolved, the dog developed nonregenerative anemia. The antimicrobial regimen was changed again to topical and systemic vancomycin. Inflammation continued to improve over the next week. MRSA should be considered a potential organism in infectious keratitis, especially when general antibiotics are not effective. Although topical and systemic chloramphenicol and/or vancomycin are effective for treating MRSA keratitis, vancomycin should only be used when culture and susceptibility results indicate it is appropriate and no other options are available. To our knowledge, this is the first detailed case report of MRSA keratitis in a dog.     

Antimicrobial activity of a bovine myeloid antimicrobial peptide (BMAP‐28) against methicillin‐susceptible and methicillin‐resistant Staphylococcus aureus

A bovine myeloid antimicrobial peptide (BMAP‐28) is a member of the cathelicidin family which is included in the innate immune system of mammals. Recently, there have been many studies about antimicrobial peptides. This study aims to clarify whether BMAP‐28 has bactericidal activity against methicillin‐resistant Staphylococcus aureus (MRSA) and compares its activity against methicillin‐susceptible S. aureus (MSSA) and MRSA. We found that the peptide was effective in killing MRSA (minimal inhibitory concentration (MIC) range; 5–20 µg/mL). It was also revealed that MSSA (MIC range; 1.25–20 µg/mL) had two levels of susceptibility to BMAP‐28. We also examined the effect of BMAP‐28 on bacterial shape to visually show its activity. After exposure to the peptide, both MSSA and MRSA cells showed the morphological changes on their surfaces. Our results indicate that BMAP‐28 is a promising candidate for medicine against drug‐resistant bacteria.     

Detection of methicillin‐resistant Staphylococcus aureus (MRSA) in the microbial flora from the conjunctiva of healthy donkeys from Sicily (Italy)

Objective To describe the bacterial flora present in the normal conjunctiva of donkeys from Sicily (Italy). Animals studied A total of 46 healthy donkeys housed in 3 locations within the territory of Palermo (Sicily, Italy) were studied. Donkeys ranged from 2 to 13 years of age, with a median age of 6 years. Procedures Forty‐six conjunctival swabs were obtained from both eyes of each animal, and specimens were cultured for aerobic bacteria. Furthermore, the antimicrobial activity of methicillin (1 μg) and oxacillin (5 μg) on Staphylococcus spp. isolates was evaluated, and a specific PCR assay, which allows the detection of mecA gene specific for methicillin‐resistant Staphylococcus aureus (MRSA) strains, was performed. Results Forty of 46 (86.9%) donkeys were positive for bacteria. Eighty bacterial isolates, representing 9 bacteria genera, were successfully cultured. The most frequently recovered bacterial genus was Staphylococcus (52/80 isolates; 65%). Several strains (20/80 isolates; 25%) belonging to the Enterobacteriaceae family were also isolated, among which the most frequently isolated genus was Enterobacter (eight isolates). Of the 52 Staphylococcus spp. isolates, 14 (26.9%) strains were oxacillin/methicillin resistant. The mecA gene was detected in 6/52 (11.5%) strains. Conclusions This study contributes to the knowledge about normal ocular flora and MRSA occurrence in donkey farms in Sicily.     

Susceptibility difference between methicillin‐susceptible and methicillin‐resistant Staphylococcus aureus to a bovine myeloid antimicrobial peptide (BMAP‐28)

A bovine myeloid antimicrobial peptide antimicrobial peptide (BMAP‐28) is a member of the cathelicidin family and acts as a component of innate immunity. There are few reports of susceptibility difference of methicillin‐resistant Staphylococcus aureus (MRSA) and susceptible strains (MSSA) against BMAP‐28. This study aims to clarify how a few amino acid substitutions of BMAP‐28 are related to its antimicrobial activity using four analog peptides of BMAP‐28. We also compared cellular fatty acid components of MSSA and MRSA using gas chromatography. We found that a few amino acid substitutions of BMAP‐28 do not change antimicrobial activity. It was also revealed that the percentage of cis‐11‐eicosenoic acid in total detected fatty acids of MRSA was significantly higher than that of MSSA. In addition, the percentage of palmitic acid in total detected fatty acids of MRSA tended to be lower than that of MSSA. Our results will provide new information to deal with the question of differences in bacterial susceptibility against BMAP‐28.     

Prevalence and Molecular Characteristics of Methicillin‐Resistant Staphylococcus aureus (MRSA) in Organic Pig Herds in The Netherlands

The prevalence of the methicillin‐resistant Staphylococcus aureus (MRSA) among conventional pig herds in the Netherlands is high (around 71%). Nevertheless, information about the prevalence of MRSA among organic pig herds is lacking. Here, we report a study on 24 of the 49 organic pig herds in the Netherlands. The prevalence of MRSA positive herds showed to be 21%. The genetic characteristics of the MRSA isolates were similar to MRSA CC398 described in conventional pigs except one exceptional HA‐MRSA CC30 found in one herd, which was presumably caused by human to animal transmission. This resulted in a prevalence of MRSA CC398 in the organic herds of 16.7%.     

A bovine myeloid antimicrobial peptide (BMAP‐28) kills methicillin‐resistant Staphylococcus aureus but promotes adherence of the bacteria

The cathelicidin family is one of the several families of antimicrobial peptides (AMPs). A bovine myeloid antimicrobial peptide (BMAP‐28) belongs to this family. Recently, the emergence of drug‐resistant bacteria such as methicillin‐resistant Staphylococcus aureus (MRSA) has become a big problem. AMPs are expected to be leading compounds of new antibiotics against drug‐resistant bacteria. In this study, we focused on the activity of BMAP‐28 against bacterial cell surfaces. First, we observed morphological change of MRSA caused by BMAP‐28 using a scanning probe microscope. We also studied activities of BMAP‐28 against adherence of S. aureus to fibronectin, collagen type I, collagen type IV. We confirmed whether BMAP‐28 can bind to lipoteichoic acid (LTA) of S. aureus. BMAP‐28 was indicated as damaging the cell surface of MRSA. In a particular range of concentrations, BMAP‐28 promoted adherence of S. aureus against fibronectin and collagens. It was revealed that BMAP‐28 and LTA of S. aureus bound with each other. Our study showed the potential of BMAP‐28 which can damage MRSA and interact with LTA of S. aureus but promote its adherence in some concentrations. This study provides new points of which to take notice when we use AMPs as medicines.     

Endodontic treatment of a mandibular premolar abscess in a horse: A case study

A pulpar abscess of cheek tooth 307 (according to the Triadan numerical system) was diagnosed in a 7‐year‐old female Quarter Horse. History included a painful response to palpation in the mandibular region for approximately 4 weeks. Symptoms included swelling of the mandibular bone with subsequent fistulisation. A complete intraoral examination was performed and no lesions or abnormalities were found. Digital radiographs of the mandibular arcade demonstrated a periapical lesion of cheek tooth 307. Various therapeutic options were considered, including standing oral extraction, retrograde repulsion and endodontic therapy. Together with the client, a decision was made to perform endodontic therapy, with the double aim of preserving the tooth and maintaining normal molar occlusion. This was done under general anaesthesia without apicectomy, and using materials commonly used in the field of human endodontics. Following surgery, radiographs were taken and confirmed the complete sealing of the pulp canals. Recovery was uneventful and no post operative complications were observed. A complete intraoral follow‐up examination was performed one year after surgery, and revealed normal eruption and occlusion of the tooth involved. Radiographic examination confirmed the correct position and integrity of the sealant material, as well as normal perialveolar bone structure. This case report indicates that endodontic therapy, following the above protocol, involving pulpectomy and sealing of the pulp cavity, represents a valid alternative to the more commonly used technique of tooth extraction.     

Cross-sectional study of antimicrobial-resistant bacteria in horses. Part 1: Prevalence of antimicrobial-resistant Escherichia coli and methicillin-resistant Staphylococcus aureus

Reasons for performing study: The increasing prevalence of antimicrobial‐resistant bacteria such as methicillin‐resistant Staphylococcus aureus (MRSA) and antimicrobial‐resistant Escherichia coli represents a significant problem. However, the carriage of such bacteria by horses in the UK has not been well characterised. Objectives: To estimate the prevalence of nasal carriage of MRSA and faecal carriage of antimicrobial‐resistant E. coli amongst horses in the general equine community of the mainland UK. Methods: A cross‐sectional study of horses recruited by 65 randomly selected equine veterinary practices was conducted, with nasal swabs and faecal samples collected. Faecal samples were cultured for antimicrobial‐resistant E. coli. Nasal swabs were cultured for staphylococcal species; methicillin‐resistant isolates identified as S. aureus were characterised by SCCmec and spa gene typing. Multilevel logistic regression models were used to calculate prevalence estimates with adjustment for clustering at practice and premises levels. Spatial variation in risk of antimicrobial resistance was also examined. Results: In total, 650 faecal samples and 678 nasal swabs were collected from 692 horses located on 525 premises. The prevalence of faecal carriage of E. coli with resistance to any antimicrobial was 69.5% (95% CI 65.9–73.1%) and the prevalence of extended‐spectrum β‐lactamase (ESBL)‐producing E. coli was 6.3% (95% CI 4.1–9.6%). The prevalence of nasal carriage of MRSA was 0.6% (95% CI 0.2–1.5%). Spatial analysis indicated variation across the UK for risk of carriage of resistant and multidrug‐resistant (resistant to more than 3 antimicrobial classes) E. coli. Conclusions and potential relevance: Carriage of MRSA by horses in the community appears rare, but the prevalence of antimicrobial‐resistant E. coli (including ESBL‐producing E. coli) is higher. A high prevalence of antimicrobial‐resistant bacteria could have significant health implications for the horse population of the UK.     

Methicillin‐resistant Staphylococcus aureus in Resident Animals of a Long‐term Care Facility

Animals provide benefits to elderly and chronically ill people by decreasing loneliness, increasing social interactions, and improving mental health. As a result, many hospitals and long‐term care facilities allow family pets to visit ill or convalescing patients or support animal‐assisted therapy programs. These include programs that have resident animals in long‐term care facilities. Despite the benefits, there are concerns about disease transmission between pets and patients. Antibiotic‐resistant bacteria, such as methicillin‐resistant Staphylococcus aureus (MRSA), are a recognized problem in healthcare settings leading to refractory infections and potentially life‐threatening illnesses. MRSA has been isolated from numerous animal species, yet few studies are available on the carriage of this pathogen in animals residing in long‐term care facilities. Our objective was to characterize MRSA carriage among resident animals in a long‐term care facility. Methods: To document MRSA colonization, nasal swabs from 12 resident animals (one dogs and 11 cats) of a long‐term care facility were collected weekly for 8 weeks. Staphylococcus isolates were characterized by antimicrobial susceptibility and MRSA isolates were further characterized by pulsed‐field gel electrophoresis (PFGE). PFGE isolate patterns were compared with an existing database of MRSA isolate patterns at the Minnesota Department of Health. Results: Two of 11 cats were colonized with MRSA. MRSA was recovered from five of eight weekly samples in one cat and two of eight weekly samples in the other cat. All isolates were classified as USA100 (healthcare‐associated strains). Discussion: Long‐term care resident animals may acquire MRSA. Clonally related strains were identified over the 8‐week sampling period. It is unclear if pets serve as an on‐going source of infection to their human companions in long‐term care facilities.     

Genome analysis of methicillin‐resistant Staphylococcus aureus isolated from pigs: Detection of the clonal lineage ST398 in Cameroon and South Africa

Food animals are considered reservoirs of methicillin‐resistant Staphylococcus aureus (MRSA) and are implicated in their zoonotic transmission in the farm‐to‐plate continuum. LA‐MRSA has been reported as a zoonotic agent that has the potential to spread to humans and may cause infections in at‐risk groups. In this study, whole genome sequencing was used to describe the genetic environment (resistance mechanisms, virulence factors and mobile genetic elements) and investigate the genetic lineages of MRSA isolates from pigs in Cameroonian and South African abattoirs. During March–October 2016, 288 nasal and rectal pooled samples from 432 pigs as well as nasal and hand swabs from 82 humans were collected. Genomic DNA was sequenced using an Illumina MiSeq platform. Generated reads were de novo‐assembled using the Qiagen CLC Genomics Workbench and SPAdes. The assembled contigs were annotated, and antibiotic resistance genes, virulence factors, plasmids, SCCmec and phage elements were identified with ResFinder, Virulence Finder, PlasmidFinder, SCCmec Finder and PHAST, respectively. Core genome single nucleotide analysis was undertaken to assess clonal relatedness among isolates. A lower MRSA prevalence was observed in pigs in Cameroon (n = 1/13; 0.07%) compared with South Africa (n = 4/22; 18.18%), and none of the workers were colonized by MRSA. Genome analysis identified various antibiotic resistance genes along with six virulence factors in all isolates. All MRSA isolates belonged to the clonal lineage ST398 (spa‐type t011) and harboured the type Vc SCCmec and several plasmids. Our study shows that the livestock‐associated MRSA clonal lineage ST398 is already present in both Cameroon and South Africa and is probably underestimated in the absence of molecular epidemiological studies. It reveals the serious food safety and public health threat associated with this animal strain and underscores the need for interventions to contain this resistant clone.     

Risk factors for methicillin-resistant Staphylococcus aureus (MRSA) infection in dogs and cats: a case-control study

Risk factors for methicillin-resistant Staphylococcus aureus (MRSA) infection in dogs and cats were investigated in an unmatched case-control study. A total of 197 animals from 150 veterinary practices across the United Kingdom was enrolled, including 105 MRSA cases and 92 controls with methicillin-susceptible S. aureus (MSSA) infection. The association of owners and veterinarian staff with the human healthcare sector (HCS) and animal-related characteristics such as signalment, antimicrobial and immunosuppressive therapy, and surgery were evaluated as putative risk factors using logistic regression. We found that significant risk factors for MRSA infection were the number of antimicrobial courses (p = 0.005), number of days admitted to veterinary clinics (p = 0.003) and having received surgical implants (p = 0.001). In addition, the odds of contact with humans which had been ill and admitted to hospital (p = 0.062) were higher in MRSA infected pets than in MSSA controls. The risk factors identified in this study highlight the need to increase vigilance towards identification of companion animal groups at risk and to advocate responsible and judicious use of antimicrobials in small animal practice.     

Isolation,Indentification and Antibiotic Resistance Analysis of Methicillin-resistant Staphylococcus aureus (MRSA) from Raw Cow Milk

In this study,we collected 502 california mastitis test (CMT) positive milk samples from several cattle farms and then the bacteria were isolated and identified from them.After that,we used multiplex-PCR for screening out methicillin-resistant Staphylococcus aureus (MRSA).Then we investigated the MRSA's antibiotics susceptibility to 19 kinds of antibiotics by discdiffusion method.The results showed that 338 strains of Staphylococci including 89 strains of MRSA were detected out from the 452 milk samples with bacteria.Additionally,most MRSA showed high sensitivity to vancomycin,fosfomycin,chloramphenicol and clarithromycin,while resistant to penicillin,polymyxin,spectinomycin and ceftazidime.The results provided certain factual basis and method for guiding the use of antibiotics in bovine mastitis,rapid identification of MRSA strains and prevention or control of MRSA infection.     

Methicillin‐resistant Staphylococcus aureus in urban Norway rat (Rattus norvegicus) populations: Epidemiology and the impacts of kill‐trapping

Urban Norway rat (Rattus norvegicus) populations can carry the bacteria methicillin‐resistant Staphylococcus aureus (MRSA). There are numerous knowledge gaps in the epidemiology of MRSA in these populations that limit understanding of its ecology in urban environments. For example, fecal shedding of MRSA, which may increase environmental contamination, has been reported in other species; however, it is unknown whether Norway rats carry the bacteria rectally. Furthermore, while intermittent MRSA shedding has been shown in other species and may dictate when the risk of transmission is highest, duration of carriage has not been examined for Norway rats. Previous work has shown that lethal animal‐control methods may increase the level of pathogens within reservoir populations, possibly by disrupting ecological patterns. However, the impact of rodent‐control on potentially environmentally acquired pathogens like MRSA has not been tested. Using capture‐mark‐recapture methods in an inner‐city neighborhood in Vancouver, Canada, we show that rats intermittently carry MRSA both in the rectum and oropharynx. By assessing the prevalence of MRSA before and after enacting a pest‐control intervention, we report that kill‐trapping had no impact on the prevalence of carriage of this environmentally‐acquired agent.     

奶牛源MRSA菌株的分离鉴定和耐药性分析

本研究对部分奶牛场采集的502份隐性乳房炎试验（california mastitis test,CMT）检测呈阳性的奶样进行细菌分离、鉴定,运用多重PCR方法筛选出耐甲氧西林金黄色葡萄球菌（methicillin-resistant Staphylococcus aureus,MRSA）,并对分离出的MRSA用纸片扩散法进行药敏试验,检测其对19种抗菌药物的敏感性。结果显示,分离到细菌的奶样有452份,其中检出葡萄球菌338株,MRSA 89株;MRSA分离株主要对万古霉素、磷霉素、氯霉素和克拉霉素敏感,对青霉素、多黏菌素、大观霉素及头孢他啶具有较强的耐药性。本研究为指导奶牛乳房炎用药、MRSA菌株的快速鉴定及制定MRSA感染的防控措施提供参考。     

Subchondral cyst‐like lesions in the talus of four horses

Subchondral cyst‐like lesions (SCLs) of horses have been found in various locations, but SCLs of the talus have been reported rarely. In this report, we describe 4 horses affected with a SCL of the talus. Each SCL was identified using digital radiography, and each horse was treated by intralesional injection of triamcinolone acetonide. Case details, history, findings during clinical and radiographic examination, treatment, and outcome of each horse are discussed.