Diversity of Staphylococcus aureus associated with mastitis from dairy cows in Rwanda.

OBJECTIVES: The objective of the present study was to examine the diversity of Staphylococcus aureus from mastitis milk samples of cows in Rwanda. METHODS: A total of 1080 quarter milk samples from 279 dairy cows were collected in 80 different farms from all five provinces of Rwanda. In total, 135 S. aureus isolates were obtained and subjected to genotyping (spa typing, DNA microarray, whole-genome sequencing (WGS)), antimicrobial susceptibility testing (AST) and phenotypic profiling by Fourier Transform Infrared (FTIR) spectroscopy (including capsular serotyping). RESULTS: Resistance to penicillin and/or tetracycline was most frequently observed. Ten sequence types (STs) (ST1, ST151, ST152, ST5477, ST700, ST7110, ST7983, ST7984, ST8320, ST97) belonging to seven clonal complexes (CCs) (CC1, CC130, CC152, CC3591, CC3666, CC705, CC97) were detected. The Panton-Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83) and the human and bovine toxic shock syndrome toxin gene tst-1 variants were detected. FTIR-based capsular serotyping showed CC-specific differences. Most CC97 (cap5 allele) isolates were primarily nonencapsulated (82%), whereas isolates of CC3591 and CC3666 (cap8 allele) were mostly encapsulated (86.4% and 57.8%, respectively). Our results underline the widespread global distribution of cattle-adapted CC97. CONCLUSION: The presence of CC3591 and CC3666 in bovine mastitis suggests an important role in cattle health and dairy production in Rwanda. The results of the present study support the need for a rigorous One-Health Surveillance program of the bovine-human interface.

Detection of mcr-1-1 Positive Enteropathogenic Escherichia coli Isolates Associated with Post-Weaning Diarrhoea in an Organic Piglet-Producing Farm in Austria.

Postweaning diarrhoea (PWD) is a frequent multifactorial disease occurring in swine stocks worldwide. Since pathogenic Escherichia (E.) coli play a pivotal role in the pathogenesis of PWD and porcine E. coli are often resistant to different antibiotics, colistin is frequently applied to treat piglets with PWD. However, the application of colistin to livestock has been associated with the emergence of colistin resistance. This case report describes the detection of the colistin resistance gene mcr-1-1 in two E. coli isolated from piglets with PWD in an Austrian organic piglet-producing farm, which was managed by two farmers working as nurses in a hospital. Both mcr-1-positive E. coli were further analysed by Illumina short-read-sequencing, including assemblies and gene prediction. Both isolates belonged to the same clonal type and were positive for eaeH and espX5, which are both virulence genes associated with enteropathogenic E. coli (EPEC). Due to the detection of mcr-1-positive EPEC and based on the results of the antimicrobial resistance testing, the veterinarian decided to apply gentamicin for treatment instead of colistin, leading to improved clinical signs. In addition, after replacing faba beans with whey, PWD was solely observed in 2/10 weaned batches in the consecutive months.

Multilocus sequence typing schemes for the emerging swine pathogen Mycoplasma hyosynoviae.

Mycoplasma (M.) hyosynoviae is a commensal of the upper respiratory tract in swine, which has the potential to spread systemically, usually resulting in arthritis in fattening pigs and gilts. To date, very little is known about the epidemiology of M. hyosynoviae, mainly due to a lack of suitable typing methods. Therefore, this study aimed to develop both a conventional multi locus sequence typing (MLST) and a core genome (cg) MLST scheme. The development of the cgMLST was based on whole genome sequences of 64 strains isolated from pigs and wild boars during routine diagnostics as well as nine publicly available genomes. A cgMLST scheme containing 390 target genes was established using the Ridom© SeqSphere+ software. Using this scheme as a foundation, seven housekeeping genes were selected for conventional MLST based on their capability to reflect genome wide relatedness and subsequently, all 73 strains were typed by applying both methods. Core genome MLST results revealed a high diversity of the studied strain population and less than 100 allele differences between epidemiologically unrelated strains were only detected for four isolates from the US. On the other hand, seven clonal clusters (≤ 12 allele differences) comprising 20 isolates were identified. Comparison of the two typing methods resulted in highly congruent phylogenetic trees and an Adjusted Rand Coefficient of 0.893, while cgMLST showed marginally higher resolution when comparing closely related isolates, indicated by a slightly higher Simpson's ID (0.992) than conventional MLST (Simpson's ID = 0.990). Overall, both methods seem well suited for epidemiological analyses for scientific as well as diagnostic purposes. While MLST is faster and cheaper, cgMLST can be used to further differentiate closely related isolates.

Molecular Characterization of Chimeric Staphylococcus aureus Strains from Waterfowl.

Staphylococcus aureus is a versatile pathogen that does not only occur in humans but also in various wild and domestic animals, including several avian species. When characterizing S. aureus isolates from waterfowl, isolates were identified as atypical CC133 by DNA microarray analysis. They differed from previously sequenced CC133 strains in the presence of the collagen adhesin gene cna; some also showed a different capsule type and a deviant spa type. Thus, they were subjected to whole-genome sequencing. This revealed multiple insertions of large regions of DNA from other S. aureus lineages into a CC133-derived backbone genome. Three distinct strains were identified based on the size and extent of these inserts. One strain comprised two small inserts of foreign DNA up- and downstream of oriC; one of about 7000 nt or 0.25% originated from CC692 and the other, at ca. 38,000 nt or 1.3% slightly larger one was of CC522 provenance. The second strain carried a larger CC692 insert (nearly 257,000 nt or 10% of the strain's genome), and its CC522-derived insert was also larger, at about 53,500 nt or 2% of the genome). The third strain carried an identical CC692-derived region (in which the same mutations were observed as in the second strain), but it had a considerably larger CC522-like insertion of about 167,000 nt or 5.9% of the genome. Both isolates of the first, and two out of four isolates of the second strain also harbored a hemolysin-beta-integrating prophage carrying "bird-specific" virulence factors, ornithine cyclodeaminase D0K6J8 and a putative protease D0K6J9. Furthermore, isolates had two different variants of SCC elements that lacked mecA/mecC genes. These findings highlight the role of horizontal gene transfer in the evolution of S. aureus facilitated by SCC elements, by phages, and by a yet undescribed mechanism for large-scale exchange of core genomic DNA.

Diversity of Staphylococcus aureus isolated from nares of ruminants.

AIMS: To examine the diversity of Staphylococcus aureus isolated from nasal swabs of ruminants in Rwanda. METHODS AND RESULTS: A total of 454 nasal swabs from 203 cows, 170 goats, and 81 sheep were examined for the presence of S. aureus, and 30 S. aureus isolates were detected and characterized pheno- and genotypically. Resistance to penicillin and/or tetracycline was observed. The isolates were assigned to eight different spa types (t21057 (novel), t10103, t18853, t20842, t318, t355, t458, and t9432) belonging to six clonal complexes (CCs) (CC152, CC30, CC3591, CC3666, CC522, and CC97). Panton-Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83), and the human and bovine variants of the toxic shock syndrome toxin gene tst-1 variants were detected. CONCLUSION: These findings demonstrate that the nares of ruminants in Rwanda are colonized with mastitis-associated S. aureus, including lineages that are also carried by humans, underscoring the zoonotic risk, especially for livestock keepers. These results highlight the crucial importance of hygiene measures when handling livestock.

Assessment of microbial quality in poultry drinking water on farms in Austria.

The quality of poultry drinking water has a significant effect on broiler health and performance. This study conducted an analysis of aerobic mesophilic counts (AMC), Enterobacteriaceae (EB), Pseudomonadaceae (PS), and screened for the presence of Campylobacter spp. in water samples collected from a total of 14 farms in Austria, with either a public or private water source. The efficacy of two water line treatment methods was evaluated: a chemical treatment of the water lines with 4.0 ppm ClO2 (T1) and a combined chemical (4.0 ppm active ClO2 and 3.0% peracetic acid) and mechanical treatment (purging of the water lines with a high-pressure air pump; T2). However, both the T1 and T2 treatments failed to reduce the AMC counts below the maximum acceptable microbial limit of 4.0 log10 CFU/ml in water samples. In addition, no significant reduction in EB and PS counts was observed in water samples after either T1 or T2 water line treatment. The water samples showed a high level of microbial diversity with 18 to 26 different genera. The genus Pseudomonas was most frequently isolated across all poultry farms, while Campylobacter jejuni was identified in a single sample collected before water line treatment. Isolate analysis revealed the presence of opportunistic pathogens in water samples both before (T1 43.1%, T2 30.9%) and after (T1 36.3%, T2 33.3%) water line treatment. Opportunistic pathogens belonging to genera including Pseudomonas spp., Stenotrophomonas spp., and Ochrobactrum spp., were most frequently isolated from poultry drinking water. These isolates exhibited multidrug resistance and resistance phenotypes to antimicrobials commonly used in Austrian poultry farms. The findings of this study emphasize the potential risk of exposure to opportunistic pathogens for poultry and personnel, underscoring the importance of efficient water line management.

Bleeding skin lesions in gestating sows of a piglet producing farm in Austria.

BACKGROUND: Stomoxys calcitrans, the stable fly, occurs in pig producing countries worldwide. While in cattle the impact of this blood sucking insect is quite well described, its role in pig production is poorly investigated. Here we describe a case of a massive stable fly overpopulation in the gestation unit of a piglet producing farm in Austria that resulted in bleeding skin lesions in bitten sows. CASE PRESENTATION: In October 2021, the responsible herd veterinarian of the case farm reported of sows in the gestation area presenting with bloody crusts on the whole skin surface of the body and of bleeding skin lesions. 33/55 sows were affected by moderate to severe skin lesions. Reproductive performance decreased during the time of massive stable fly overpopulation. Sows in the gestation unit showed defensive behaviour and at a certain time point resigned and accepted being bitten by stable flies. After controlling the fly population, reproductive performance improved and even exceeded the performance before the massive overgrowth of the stable fly population. CONCLUSIONS: Stable flies are a serious harm to pigs and should be kept in mind for improved animal health and welfare. Knowledge about the determination of Stomoxys calcitrans and early recognition of an increasing stable fly population in pig farming systems followed by proper insect control measures have to be performed to reduce losses caused by this harming insect.

Retrospective Analysis of the Detection of Pathogens Associated with the Porcine Respiratory Disease Complex in Routine Diagnostic Samples from Austrian Swine Stocks.

The diagnostic workup of respiratory disease in pigs is complex due to coinfections and non-infectious causes. The detection of pathogens associated with respiratory disease is a pivotal part of the diagnostic workup for respiratory disease. We aimed to report how frequently certain viruses and bacteria were detected in samples from pigs with respiratory symptoms in the course of routine diagnostic procedures. Altogether, 1975 routine diagnostic samples from pigs in Austrian swine stocks between 2016 and 2021 were analysed. PCR was performed to detect various pathogens, including porcine reproductive and respiratory syndrome virus (PRRSV) (n = 921), influenza A virus (n = 479), porcine circovirus type 2 (PCV2) (n = 518), Mycoplasma (M.) hyopneumoniae (n = 713), Actinobacillus pleuropneumoniae (n = 198), Glaesserella (G.) parasuis (n = 165) and M. hyorhinis (n = 180). M. hyorhinis (55.1%) had the highest detection rate, followed by PCV2 (38.0%) and Streptococcus (S.) suis (30.6%). PRRSV was detected most frequently in a pool of lung, tonsil and tracheobronchial lymph node (36.2%). G. parasuis was isolated more frequently from samples taken after euthanasia compared to field samples. PRRSV-positive samples were more likely to be positive for PCV2 (p = 0.001), M. hyopneumoniae (p = 0.032) and Pasteurella multocida (p < 0.001). M. hyopneumoniae-positive samples were more likely to be positive for P. multocida (p < 0.001) and S. suis (p = 0.046), but less likely for M. hyorhinis (p = 0.004). In conclusion, our data provide evidence that lung samples that were positive for a primary pathogenic agent were more likely to be positive for a secondary pathogenic agent.

A single, episodic event of unilateral/bilateral scrotal swelling in a group of adult boars at an Austrian boar stud.

BACKGROUND: Scrotal swelling is a clinical situation which can be caused by different aetiologies. In this case report, we describe a multi-week episode of unilateral and bilateral scrotal swelling in boars at an Austrian boar stud and its diagnostic work-up. CASE PRESENTATION: In the summer of 2020, the herd veterinarian of an Austrian boar stud reported that over a period of six weeks, five out of 70 boars presented with unilateral severe swelling of the left scrotum and three out of 70 boars with bilateral severe swelling of the left and moderate swelling of the right scrotum, respectively. A complete history was obtained and an on-site evaluation of the facility was done. Five boars were necropsied, and a variety of samples harvested for further diagnostic investigations. Infectious differential diagnoses associated with unilateral swelling of the scrotum or the testis were excluded through serological and tissue testing. In three of the five boars, histopathology revealed complete acute haemorrhagic necrosis of the left testis concurrent with strongly congested blood vessels. Review of the collected information with a group of experts in the field of boar stud management resulted with consensus that, most likely, trauma was the etiologic event causing the clinical signs and pathology. Coincident with discussion of implementing video recording cameras in the boar housing area, no further clinical cases followed. As this case occurred during the first lockdown of the COVID-19 pandemic, we propose that the distress and travelling restrictions may have contributed to frustration among boar stud workers, which was consequently expressed as misbehaviour against boars. CONCLUSIONS: Once all known infectious causes of unilateral swelling of the scrotum were excluded, a critical diagnostic work-up focused on non-infectious causes. Non-infectious causes, such as trauma, need to be carefully evaluated, as it may also include human misbehaviour against boars. Summarizing all findings of this case report, the authors hypothesize that a blunt trauma was the reason for the series of mainly unilateral swelling of the scrota of boars.

Outbreak of Cronobacter turicensis in European brown hares (Lepus europaeus).

This is the first report of acute deaths in five European brown hares (Lepus europaeus) attributed to mucoid and necrotizing typhlocolitis caused by genetically different Cronobacter (C.) turicensis strains in northeastern Austria. As this opportunistic pathogen is mainly known for causing disease in immunocompromised humans and neonates, this previously unrecognized potential for a spill over from a wildlife reservoir to humans warrants further attention.

Emergence of methicillin resistance predates the clinical use of antibiotics.

The discovery of antibiotics more than 80 years ago has led to considerable improvements in human and animal health. Although antibiotic resistance in environmental bacteria is ancient, resistance in human pathogens is thought to be a modern phenomenon that is driven by the clinical use of antibiotics1. Here we show that particular lineages of methicillin-resistant Staphylococcus aureus-a notorious human pathogen-appeared in European hedgehogs in the pre-antibiotic era. Subsequently, these lineages spread within the local hedgehog populations and between hedgehogs and secondary hosts, including livestock and humans. We also demonstrate that the hedgehog dermatophyte Trichophyton erinacei produces two ß-lactam antibiotics that provide a natural selective environment in which methicillin-resistant S. aureus isolates have an advantage over susceptible isolates. Together, these results suggest that methicillin resistance emerged in the pre-antibiotic era as a co-evolutionary adaptation of S. aureus to the colonization of dermatophyte-infected hedgehogs. The evolution of clinically relevant antibiotic-resistance genes in wild animals and the connectivity of natural, agricultural and human ecosystems demonstrate that the use of a One Health approach is critical for our understanding and management of antibiotic resistance, which is one of the biggest threats to global health, food security and development.

Mycoplasma tauri sp. nov. isolated from the bovine genital tract.

Since 2006, a Mycoplasma species unidentifiable to the species level has been regularly isolated from the semen and prepuce of apparently healthy bulls, and occasionally from cattle displaying inflammatory disease of the genital tract. Seven of these Mycoplasma isolates were subjected to a comprehensive taxonomic study. The strains investigated grew well in modified Hayflick's medium and colonies on agar exhibited typical fried egg morphology and produced 'film and spots'. Transmission electron microscopy revealed a cell morphology characteristic of mycoplasmas with spherically shaped cells bounded by a bi-layered cell membrane. The strains studied neither produced acid from sugar carbon sources nor did hydrolyse arginine or urea, and genome annotation indicated that organic acids (pyruvate, lactate) are used as energy sources. Phylogenetic analyses of 16S rRNA gene sequences, the 16S-23S intergenic spacer region, and partial rpoB gene and protein sequences placed the strains within the Mycoplasma (M.) bovis cluster of the Hominis group with M. primatum, M. agalactiae, and M. bovis being their closest relatives. Genomic information including whole-genome similarity metrics (ANIb, ANIm, TETRA, dDDH, AAI) and phylogenomics, proteomic features revealed by matrix-assisted laser desorption ionization time of flight (MALDI-ToF) mass spectrometry as well as serological reactions and polar lipid profiling strongly indicated that the strains examined were representatives of a hitherto unclassified species of genus Mycoplasma, for which the name Mycoplasma tauri sp. nov. with type strain Zaradi2T (=ATCC BAA-1891T = DSM 22451T) is proposed.

Staphylococcus aureus isolates from Eurasian Beavers (Castor fiber) carry a novel phage-borne bicomponent leukocidin related to the Panton-Valentine leukocidin.

Staphylococcus aureus can be a harmless coloniser, but it can also cause severe infections in humans, livestock and wildlife. Regarding the latter, only few studies have been performed and knowledge on virulence factors is insufficient. The aim of the present study was to study S. aureus isolates from deceased wild beavers (Castor fiber). Seventeen isolates from eleven beavers, found in Germany and Austria, were investigated. Antimicrobial and biocide susceptibility tests were performed. Isolates were characterised using S. aureus-specific DNA microarrays, spa typing and whole-genome sequencing. From two isolates, prophages were induced by mitomycin C and studied by transmission electron microscopy. Four isolates belonged to clonal complex (CC) 8, CC12, and CC398. Twelve isolates belonged to CC1956 and one isolate was CC49. The CC49 and CC1956 isolates carried distinct lukF/S genes related to the Panton-Valentine leukocidin (PVL) from human isolates of S. aureus. These genes were located on related, but not identical, Siphovirus prophages. The beavers, from which those isolates originated, suffered from abscesses, purulent organ lesions and necrotising pneumonia, i.e., clinical manifestations resembling symptoms of severe PVL-associated disease in humans. It might thus be assumed that the "Beaver Leukocidin (BVL, lukF/S-BV)"-positive strains are beaver-specific pathogens, and further studies on their clinical role as well as on a possible transmissibility to other species, including humans, are warranted.

The First Bacterial Endocarditis Due to Achromobacter xylosoxidans in a Dog.

Infectious endocarditis (IE) in dogs is often associated with a high mortality rate as diagnostic work-up as well as antibiotic treatment might be challenging. The present case describes bacteremia in a dog caused by Achromobacter xylosoxidans, leading to an infectious endocarditis. Achromobacter xylosoxidans (A. xylosoxidans) is an aerobic Gram-negative rod-shaped bacterium, which has been associated with multiple nosocomial opportunistic diseases in human medicine. One such manifestation of A. xylosoxidans infection is endocarditis. A. xylosoxidans infections are challenging to treat due to the reduced effectiveness of a wide range of antimicrobial agents. To date, only a few case reports of infections with A. xylosoxidans in animals have been described. This is the first case report of A. xylosoxidans endocarditis in a dog. Whole-genome sequencing was performed to determine the sequencing type and to gain more information about this bacterium regarding its intrinsic resistance genes. With this case report, we seek to increase awareness of A. xylosoxidans as an opportunistic nosocomial pathogen in dogs and to provide a short summary regarding the current state of general knowledge and known resistance patterns.

A core genome multilocus sequence typing scheme for Mycoplasma hyorhinis.

Mycoplasma (M.) hyorhinis is a commensal and pathobiont residing in the upper respiratory tract in swine and with the ability to spread systemically, mainly causing polyserositis and polyarthritis in nursery pigs. Since little is known on the epidemiology of M. hyorhinis infection, whole genome sequences of 73 strains isolated from pigs in Austria (n = 71) and Germany (n = 2), that have been isolated from clinically affected pigs during routine diagnostics, and publicly available genomes of eight M. hyorhinis strains were analyzed in the presented study. For this purpose, a core genome multi locus sequence typing (cgMLST) scheme encompassing 453 target genes was developed using the Ridom© SeqSphere + software. Results were compared to two previously described conventional MLST schemes and to a core genome single nucleotide polymorphism (cgSNP) analysis approach. Core genome MLST showed high diversity among the M. hyorhinis strains studied and while certain isolates from one farm or a single animal formed cgMLST clusters (≤ 8 allele differences), no isolates with identical allele profiles were identified. In addition, cgMLST had superior discriminatory power (Simpson's ID = 0.995) over conventional MLST (Simpson's ID = 0.952 and 0.985), while demonstrating a lack of congruence between conventional MLST and genome-wide relationship. Core genome SNP results were highly congruent with cgMLST results but lacked in resolution when comparing closely related isolates. Thus, cgMLST is the most suitable method for epidemiological investigations such as outbreak analysis, and to gain insights into M. hyorhinis population structure.

The First Report of mcr-1-Carrying Escherichia coli Originating from Animals in Serbia.

The aim of this study was continuous monitoring of the presence of mcr-1 to mcr-5 genes in Enterobacterales isolated from cattle, pigs, and domestic poultry at intensive breeding facilities in Northern Vojvodina, Serbia, from 1 January 1 to 1 October 2020. Out of 2167 examined samples, mcr-1 was observed in five E. coli isolates originating from healthy turkeys. Four isolates belonged to the phylogenetic group B1, and one isolate to the phylogenetic group A. Detected E. coli serogenotypes (somatic O and flagellar H antigens) were O8:H25 and O29:H25. Core-genome multi-locus sequence typing (cgMLST) revealed three ST58 isolates clustering together in Clonal Complex (CC) 155 and two singletons of ST641-CC86 and ST410-CC23, respectively. Clonotyping revealed CH4-32 (n = 3), CH6-53 (n = 1) and CH4-24 (n = 1). In all isolates, the mcr-1 gene was located on a large IncX4 replicon type plasmid. Eight virulence-associated genes (VAGs) typical of avian pathogenic E. coli (APEC) (fyuA, fimH, hlyF, iss, ompT, sitA, traT, iroN) were detected in four isolates. These isolates were investigated for susceptibility to four biocides and revealed MIC values of 0.125% for glutardialdehyde, of 0.00003-0.00006% for chlorohexidine, of 4-6% for isopropanol and of 0.001-0.002% for benzalkonium chloride. All obtained MIC values of the tested biocides were comparable to the reference strain, with no indication of possible resistance. This is the first report of mcr-1.1-carrying E. coli from Serbia. Although only samples from turkeys were mcr-positive in this study, continuous monitoring of livestock samples is advised to prevent a spill-over from animals to humans.

Exudative Epidermitis in Combination with Staphylococcal Pyoderma in Suckling Piglets.

A case of generalized exudative epidermitis (EE) is described, which occurred in a very small piglet producing farm in Austria. The antimicrobial treatment prescribed by the herd veterinarian did not improve the clinical problem. Therefore, the University Clinic for Swine intervened in the case. Lab investigations were initiated in which Staphylococcus hyicus (SH) and Staphylococcus aureus (SA), both methicillin-resistant and susceptible strains, could be isolated from the skin of affected piglets. Poor hygiene and management practices were identified as predisposing factors on site. Adaptation of antimicrobial treatment according to results of the in vitro susceptibility testing and the implementation of proper hygiene measures resolved the clinical problem. Here, we describe a fatal coinfection of SH and SA in suckling piglets.

The Pheno- and Genotypic Characterization of Porcine Escherichia coli Isolates.

Escherichia (E.) coli is the main causative pathogen of neonatal and post-weaning diarrhea and edema disease in swine production. There is a significant health concern due to an increasing number of human infections associated with food and/or environmental-borne pathogenic and multidrug-resistant E. coli worldwide. Monitoring the presence of pathogenic and antimicrobial-resistant E. coli isolates is essential for sustainable disease management in livestock and human medicine. A total of 102 E. coli isolates of diseased pigs were characterized by antimicrobial and biocide susceptibility testing. Antimicrobial resistance genes, including mobile colistin resistance genes, were analyzed by PCR and DNA sequencing. The quinolone resistance-determining regions of gyrA and parC in ciprofloxacin-resistant isolates were analyzed. Clonal relatedness was investigated by two-locus sequence typing (CH clonotyping). Phylotyping was performed by the Clermont multiplex PCR method. Virulence determinants were analyzed by customized DNA-based microarray technology developed in this study for fast and economic molecular multiplex typing. Thirty-five isolates were selected for whole-genome sequence-based analysis. Most isolates were resistant to ampicillin and tetracycline. Twenty-one isolates displayed an ESBL phenotype and one isolate an AmpC ß-lactamase-producing phenotype. Three isolates had elevated colistin minimal inhibitory concentrations and carried the mcr-1 gene. Thirty-seven isolates displayed a multi-drug resistance phenotype. The most predominant ß-lactamase gene classes were blaTEM-1 (56%) and blaCTX-M-1 (13.71%). Mutations in QRDR were observed in 14 ciprofloxacin-resistant isolates. CH clonotyping divided all isolates into 51 CH clonotypes. The majority of isolates belonged to phylogroup A. Sixty-four isolates could be assigned to defined pathotypes wherefrom UPEC was predominant. WGS revealed that the most predominant sequence type was ST100, followed by ST10. ST131 was detected twice in our analysis. This study highlights the importance of monitoring antimicrobial resistance and virulence properties of porcine E. coli isolates. This can be achieved by applying reliable, fast, economic and easy to perform technologies such as DNA-based microarray typing. The presence of high-risk pathogenic multi-drug resistant zoonotic clones, as well as those that are resistant to critically important antibiotics for humans, can pose a risk to public health. Improved protocols may be developed in swine farms for preventing infections, as well as the maintenance and distribution of the causative isolates.

Exploring the evolution and epidemiology of European CC1-MRSA-IV: tracking a multidrug-resistant community-associated meticillin-resistant Staphylococcus aureus clone.

This study investigated the evolution and epidemiology of the community-associated and multidrug-resistant Staphylococcus aureus clone European CC1-MRSA-IV. Whole-genome sequences were obtained for 194 European CC1-MRSA-IV isolates (189 of human and 5 of animal origin) from 12 countries, and 10 meticillin-susceptible precursors (from North-Eastern Romania; all of human origin) of the clone. Phylogenetic analysis was performed using a maximum-likelihood approach, a time-measured phylogeny was reconstructed using Bayesian analysis, and in silico microarray genotyping was performed to identify resistance, virulence-associated and SCCmec (staphylococcal cassette chromosome mec) genes. Isolates were typically sequence type 1 (190/204) and spa type t127 (183/204). Bayesian analysis indicated that European CC1-MRSA-IV emerged in approximately 1995 before undergoing rapid expansion in the late 1990s and 2000s, while spreading throughout Europe and into the Middle East. Phylogenetic analysis revealed an unstructured meticillin-resistant S. aureus (MRSA) population, lacking significant geographical or temporal clusters. The MRSA were genotypically multidrug-resistant, consistently encoded seh, and intermittently (34/194) encoded an undisrupted hlb gene with concomitant absence of the lysogenic phage-encoded genes sak and scn. All MRSA also harboured a characteristic ~5350 nt insertion in SCCmec adjacent to orfX. Detailed demographic data from Denmark showed that there, the clone is typically (25/35) found in the community, and often (10/35) among individuals with links to South-Eastern Europe. This study elucidated the evolution and epidemiology of European CC1-MRSA-IV, which emerged from a meticillin-susceptible lineage prevalent in North-Eastern Romania before disseminating rapidly throughout Europe.

Presence of ß-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals.

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant ß-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-ß-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.