Annika Posautz

Characterization of methicillin-resistant Staphylococcus spp. carrying the mecC gene, isolated from wildlife

OBJECTIVES A recently identified mecA homologue, mecC, in methicillin-resistant Staphylococcus aureus (MRSA) has been isolated from humans and different animal hosts. The aim of this study was to determine antimicrobial resistance and provide molecular characterization of MRSA and methicillin-resistant non-Staphylococcus aureus staphylococci (MRnSA) isolated from wildlife that carried the gene mecC. METHODS Five S. aureus and one coagulase-negative Staphylococcus isolate displaying phenotypic oxacillin resistance, but not recognized with conventional PCR for mecA, were further characterized by a polyphasic approach. The presence of mecC in all isolates was determined using specific PCR. PCR targeting Panton-Valentine leucocidin (PVL) genes of MRSA was performed. MRSA isolates were genotyped by spa typing and multilocus sequence typing. All isolates were genotyped by staphylococcal cassette chromosome mec (SCCmec) typing. 16S rDNA sequence analysis for MRnSA identification was performed. Antimicrobial susceptibility testing was performed for all isolates. RESULTS All five MRSA isolates contained the mecC gene, were PVL negative, carried SCCmec type XI and belonged to ST130 (where ST stands for sequence type), with spa types t843, t10513 or t3256, or to ST2620, with spa type t4335. The MRnSA isolate, most closely related to Staphylococcus stepanovicii, carried mecA and blaZ genes related to SCCmec XI. MRSA isolates exhibited resistance to the β-lactams only. CONCLUSIONS The MRSA isolates described in this study represent the first detection of mecC-positive MRSA in a European otter (Lutra lutra) and a European brown hare (Lepus europaeus). The MRnSA isolate represents the first isolation of MRnSA from a Eurasian lynx (Lynx lynx).

Immunoblotting for the serodiagnosis of alveolar echinococcosis in alive and dead Eurasian beavers (Castor fiber)

A novel species-specific anti-beaver-IgG-alkaline-phosphatase conjugate was synthesized for the development of a new serological test for echinococcosis in beavers. Two different ELISAs conventionally used for human Echinococcus multilocularis serology (Em18-ELISA and Em2-ELISA) yielded diagnostic sensitivities of 0% and 46%, respectively. In contrast, the subsequently developed immunoblotting assay gave an 85% diagnostic sensitivity (11 out of 13 beavers with alveolar echinococcosis were immunoblotting-positive, i.e. showed reactivity with a specific 21 Mr band), and maximal specificity. In conclusion, this immunoblotting assay should be the method of choice for use in serological studies on E. multilocularis in Eurasian beavers, and the test proved suitable to investigate both animals alive and post-mortem.

Characterization of ESBL- and AmpC-Producing and Fluoroquinolone-Resistant Enterobacteriaceae Isolated from Mouflons ( Ovis orientalis musimon ) in Austria and Germany

The aim of this study was to investigate the presence of β-lactamase producing or fluoroquinolone-resistant members of the family Enterobacteriaceae in European mouflons (Ovis orientalis musimon). The mouflon samples originated from nasal and perineal swabs and/or organ samples in cases of a suspected infection. Only one of the 32 mouflons was tested positive for the presence of Enterobacteriaceae that displayed either an ESBL/AmpC phenotype or were resistant to ciprofloxacin. The positively tested swab originated from a sample of the jejunal mucosa of a four-year old female mouflon. Two different colony morphotypes were identified as Escherichia coli and Klebsiella pneumoniae. These isolates were phenotypically and genotypically characterized in detail by a polyphasic approach. Both isolates were multi-drug resistant. The E. coli isolate belonged to the phylogenetic group B1 and sequence type (ST) 744 and harboured the β-lactamase genes blaCTX-M-15 and blaOXA-1. The K. pneumoniae, identified as ST11, harboured the β-lactamase genes blaSHV-11, blaOXA-1, and blaDHA-1 as well as the plasmid-mediated quinolone resistance (PMQR) gene qnrB55. The present study demonstrates that wild animals can acquire human-derived resistance determinants and such findings may indicate environmental pollution with resistance determinants from other sources.

Health screening of free-ranging European brown hares (Lepus europaeus) on the German North-Sea island Pellworm.

BackgroundA sudden decline of the European brown hare (Lepus europaeus) population in one of the best hunting districts for small game species in northern Germany, the German North-Sea island Pellworm, in the years 2007/08 following marked habitat changes led to the implementation of a thorough health assessment program of the population. 110 animals were collected during the normal hunting season in the years 2010 and 2011. A post-mortem examination and histopathological investigation was performed on all animals. Additionally, routine bacteriology of the small intestine and parasitology were carried out. Sera of hares were tested for European Brown Hare Syndrome (EBHS) by enzyme linked immunosorbent assay, and for Treponema sp. by indirect immunofluorescent test. Additional testing was performed when deemed necessary.ResultsThe most striking result was a shift in the intestinal bacterial flora towards Gram-negative Enterobacteriaceae with a predominance of either Escherichia coli, or Aeromonas sp., or a high-grade double-infection with these two pathogens with subsequent catarrhal enteritis. Additionally, a marked coccidiosis, and varying infestations with the nematode Trichostrongylus retortaeformis were found. The sero-prevalence for EBHS was 78.1%, and for Treponema 43.9%.ConclusionsThe shift and decrease in diversity of the intestinal flora was the main and most consistent result found. In the authors’ opinion the change of the habitat combined with other stressors increased the animals’ sensitivity to ubiquitous bacterial species and parasites which usually would not have such fatal effects.

Bunyaviruses in human, animal and mosquito samples from southeast Austria

In Austria occurrence of 40 different species of mosquitoes belonging to 6 genera has been described. AGES in 2011 initiated a nation-wide mosquitoe-surveillance program to identify invading species. As part of this project, pools of Culex, Aedes, Anopheles and Culisetta species collected in different parts of the country were analyzed by PCR analysis; in addition seroepidemiological testing for bunyaviruses was performed in selected regions. In order to be able to detect also new strains, a broad spectrum CODEHOP (Consensus-Degenerate Hybrid Oligonucleotide Primer) approach was used initially; presumably due to the low sensitivity of degenerated primer designs and due to dilution in insect pools, we did not pick up any new isolates so far using Orthobunyavirus, Phlebovirus or Nairovirus group-specific primers. Two bunyaviruses were found in Culex pipiens from the southern province Carynthia in 2012, detected with non-degenerate multiplex PCR, the sequences highly homologous to Italian Tahynavirus (TAHV) isolates. 245 sera collected for a study of zoonotic infections in hunters, veterinarians, farmers, and abattoir workers (conducted by the Styrian health authorities in 2003), were tested by ELISA tests, using inactivated Crimean Congo virus (CCHF) from Bulgaria as well as recombinant CCHF nucleoprotein produced in baculovirus. In addition, cell culture derived TAHV lysates as well as TAHV infected cells were used as antigens in ELISA and immunofluorescence (IF) under an experimental setup. No seropositivity for CCHF or related nairoviruses was found in these Austrian human sera originating from risk groups for zoonotic infection.

Different population – different prevalence or, what is going on in the European brown hare (Lepus europaeus)

Different population - different prevalence or, what is going on in the European brown hare (Lepus europaeus)