Colistin Resistance Mediated by mcr-1 in ESBL-Producing, Multidrug Resistant Salmonella Infantis in Broiler Chicken Industry, Italy (2016-2017)

Colistin-resistance mediated by mobilisable and plasmid-borne mcr genes has emerged worldwide, threatening the efficacy of colistin, a last resort antibiotic increasingly used for treating human invasive infections by multidrug-resistant or extensively drug-resistant Enterobacteriaceae. In this study, we report the first evidence of mcr-1-mediated colistin resistance in four multidrug resistant (MDR) out of 324 Salmonella infantis from the Italian antimicrobial resistance (AMR) monitoring (2001–2017) in broilers and broiler meat. Two were also Extended Spectrum Beta-Lactamases (ESBL)-producing isolates. Characterization by whole genome sequencing (WGS), located mcr-1.1 on an incX4 plasmid. Phylogenetic analysis of these isolates with selected Italian S. Infantis previously isolated from animals, meat and human clinical cases with unknown epidemiological relationship, demonstrated that ESBL-producing, mcr-1-positive isolates belonged to the emerging pESI-like-positive-ESBL-producing clone described in Italy in 2015

Azithromycin resistance in Escherichia coli and Salmonella from food-producing animals and meat in Europe.

OBJECTIVES To characterize the genetic basis of azithromycin resistance in Escherichia coli and Salmonella collected within the EU harmonized antimicrobial resistance (AMR) surveillance programme in 2014-18 and the Danish AMR surveillance programme in 2016-19. METHODS WGS data of 1007 E. coli [165 azithromycin resistant (MIC > 16 mg/L)] and 269 Salmonella [29 azithromycin resistant (MIC > 16 mg/L)] were screened for acquired macrolide resistance genes and mutations in rplDV, 23S rRNA and acrB genes using ResFinder v4.0, AMRFinder Plus and custom scripts. Genotype-phenotype concordance was determined for all isolates. Transferability of mef(C)-mph(G)-carrying plasmids was assessed by conjugation experiments. RESULTS mph(A), mph(B), mef(B), erm(B) and mef(C)-mph(G) were detected in E. coli and Salmonella, whereas erm(C), erm(42), ere(A) and mph(E)-msr(E) were detected in E. coli only. The presence of macrolide resistance genes, alone or in combination, was concordant with the azithromycin-resistant phenotype in 69% of isolates. Distinct mph(A) operon structures were observed in azithromycin-susceptible (n = 50) and -resistant (n = 136) isolates. mef(C)-mph(G) were detected in porcine and bovine E. coli and in porcine Salmonella enterica serovar Derby and Salmonella enterica 1,4, [5],12:i:-, flanked downstream by ISCR2 or TnAs1 and associated with IncIγ and IncFII plasmids. CONCLUSIONS Diverse azithromycin resistance genes were detected in E. coli and Salmonella from food-producing animals and meat in Europe. Azithromycin resistance genes mef(C)-mph(G) and erm(42) appear to be emerging primarily in porcine E. coli isolates. The identification of distinct mph(A) operon structures in susceptible and resistant isolates increases the predictive power of WGS-based methods for in silico detection of azithromycin resistance in Enterobacterales

Sheep milk yogurt from a short food supply chain: study of the microbiological, chemico-physical and organoleptic parameters in relation to shelf-life

Aim of this work was to analyse some microbiological, chemico-physical and organoleptic parameters of sheep milk yogurt during and after its declared shelf-life. Five samples of a sheep’s milk yogurt of the same lot, collected from a short supply chain ovine dairy farm of the Roman province, were analysed. Declared shelf-life of the product was 30 days. The products were examined at 2, 14, 30, 35 and 40 days from the production date, performing the following microbiological analyses: enumeration of i) colony-forming units characteristic of the yogurt, ii) <em>Enterobacteriaceae</em>, iii) yeasts and/or moulds at 25°C. Microbiological identification was performed by miniature biochemical tests and for the lactic acid bacteria also by PCR. At every test interval, evaluation of organoleptic parameters and pH was also performed. The analysed product maintained an almost constant amount of lactic acid bacteria until the end of the declared shelf-life. Concerning lactic acid bacteria, a 100% concordance of the results observed by using biochemical identification methods and PCR assays was obtained. After 14 days from the production, the presence of yeasts (<em>Candida famata</em>) was revealed, while the presence of moulds was detected after 30 days. <em>Ralstonia picketii</em>, an environmental microorganism, was also isolated. The results obtained in this study indicate that yogurt spoilage is mainly due to the growth of specific microorganisms of spoilage, such as yeasts and moulds

Yoghurt from short supply chain: preliminary study of microbiological and physicochemical characteristics during shelf life

Yoghurt is one of very popular flavorful and healthful dairy product obtained by fermentation of lactic acid bacteria including Lactobacillus delbrueckii bulgaricus and Streptococcus thermophilus. Its production and consumption is growing continuously due to its therapeutic properties beside its high nutritive value. Thirty samples of yoghurt from short supply chain produced in 2 factory localized in Lazio region were analyzed with the aim of determining how certain microbiological and physicochemical characteristics change during their shelf life. Different types of yoghurt were studied: plain (12), fruit (14) and cereal yoghurt (4) produced with cow (8) and goat milk (22).The obtained results show: no presence of Enterobacteria, occasional presence of moulds and a considerable presence of yeasts. On the other hand, all the products analyzed have shown an almost constant amount of lactic acid bacteria during their shelf life. Lactic acid bacteria were identified by a biochemical and polymerase chain reaction assay. The presence of undesired microorganisms like yeasts was found. However, the quality of products was satisfying for the concentration of lactic acid bacteria detected in their shelf life

Preliminary results on detection of Extended-Spectrum Beta-Lactamase (ESBL)-producing Escherichia coli from red foxes, corvids, and waterfowl in the Emilia-Romagna region, Northern Italy

In the context of the One Health approach, antimicrobial resistance (AMR) is considered one of the main challenges. Various studies have reported how wildlife populations may act as sentinels for AMR in the environment, especially in highly anthropized landscape or where zootechnical activities are intensive. Furthermore, predators may acquire AMR bacteria through consumption of prey. Escherichia coli can acquire AMR genes, including those encoding resistance to Highest Priority Critically Important Antimicrobials (HPCIAs) for human medicine as Extended-spectrum β-lactamases (ESBL) genes, thus representing a good bioindicator for AMR (1-3). In the study period January 2020-April 2022, 370 E. coli strains were isolated from faecal samples of 347 wild animals (115 red foxes, 183 corvids, and 49 waterfowl) in the Emilia-Romagna region (Northern Italy) to assess the prevalence of ESBL E. coli faecal shedders. Following Decision (EU) 2020/1729 (4), the isolates were tested for AMR towards 15 antimicrobials, most of them ranked as Critically Important Antimicrobials (CIAs) by the WHO. By MIC test, 31/370 (8.4%) phenotypically ESBL-producing E. coli were detected as resistant to cefotaxime and/or ceftazidime. ESBL E. coli were isolated from red foxes (n= 13), corvids (n= 11) and waterfowl (n= 7). By PCR, phenotypically-resistant E. coli harboured ESBL-plasmidic related genes; in particular, blaSHV, blaTEM and blaCTX genes were carried by 2 (6.5%), 12 (38.7%) and 30 (96.8%) isolates, respectively. The co-presence of all genes was never found, but 38.7% of ESBL-producing E. coli harboured blaTEM and blaCTX –genes simultaneously. Overall, the genotypically-confirmed ESBL E. coli tested by MIC test were found resistant to cefotaxime (100%), ampicillin (96.9%), sulfamethoxazole (87.1%), ceftazidime (87.1%), ciprofloxacin (51.6%), tetracycline (48.4%), trimethoprim (35.5%), nalidixic acid (16.1%), chloramphenicol (16.1%), gentamycin (6.5%), colistin (3.2%). All isolates were susceptible to amikacin, meropenem, tigecycline and azithromycin. The prevalence of ESBL-producing E. coli found in our study shows the potential role of wild animals, especially waterfowl (7/49; 28.6%) and red foxes (13/115; 11.3%), as carriers of MDR isolates. This phenomenon is worsened by co-resistances to CIAs, thus representing a potential public health risk and suggesting the use of wild animals as sentinels for the analysis of the AMR spread in the environment

Occurrence of methicillin-resistant Staphylococcus aureus in dairy cattle herds, related swine farms, and humans in contact with herds

In this study we investigated the circulation of methicillin-resistant Staphylococcus aureus (MRSA) in 2 dairy cattle farms (farm A and B), previously identified as MRSA-positive in bulk tank milk samples, and epidemiologically related to swine farms. Collected specimens included quarter milk samples and nasal swabs from dairy cows, pig nasal swabs collected at both the farm and slaughterhouse level, environmental dust samples, and human nasal swabs from the farms' owners and workers. The prevalence of MRSA was estimated at the herd level by testing quarter milk samples. The prevalence of MRSA was 4.8% (3/63; 95% confidence interval = 0\u201310.2%) and 60% (33/55; 95% confidence interval = 47.05\u201372.95) in farm A and B, respectively. In farm A, MRSA was also isolated from humans, pigs sampled at both farm and slaughterhouse level, and from environmental samples collected at the pig facilities. The dairy cattle facilities of farm A tested negative for MRSA. In farm B, MRSA was isolated from environmental dust samples in both the cattle and pig facilities, whereas nasal swabs collected from cows and from humans tested negative. Sixty-three selected MRSA isolates obtained from different sources in farm A and B were genetically characterized by multilocus sequence typing, spa-typing, ribosomal spacer-PCR, and also tested for the presence of specific virulence genes and for their phenotypical antimicrobial susceptibility by broth microdilution method. Different clonal complex (CC) and spa-types were identified, including CC398, CC97, and CC1, CC already reported in livestock animals in Italy. The MRSA isolates from quarter milk of farm A and B mostly belonged to CC97 and CC398, respectively. Both lineages were also identified in humans in farm A. The CC97 and CC398 quarter milk isolates were also identified as genotype GTBE and GTAF by ribosomal spacer-PCR respectively, belonging to distinct clusters with specific virulence and resistance patterns. The GTBE and GTAF clusters also included swine, environmental, and human isolates from both farms. A high heterogeneity in the genetic and phenotypic profiles was observed in environmental isolates, in particular from farm B. These results demonstrate the possibility of a dynamic sharing and exchange of MRSA lineages or genotypes between different species and farm compartments in mixed-species farms. The risk of transmission between swine and related dairy cattle herds should be considered. Our findings also confirm the zoonotic potential of livestock-associated MRSA and underline the importance of applying biosecurity measures and good hygiene practices to prevent MRSA spread at the farm level and throughout the food production chain

DS1_JVDI_10.1177_1040638718764786 – Supplemental material for Non-toxigenic <i>Corynebacterium ulcerans</i> sequence types 325 and 339 isolated from two dogs with ulcerative lesions in Italy

<p>Supplemental material, DS1_JVDI_10.1177_1040638718764786 for Non-toxigenic <i>Corynebacterium ulcerans</i> sequence types 325 and 339 isolated from two dogs with ulcerative lesions in Italy by Virginia Carfora, Fabia Scarampella, Manuela Iurescia, Valentina Donati, Fiorentino Stravino, Serena Lorenzetti, Erika Menichini, Alessia Franco, Andrea Caprioli, Antonio Battisti in Journal of Veterinary Diagnostic Investigation</p

Detection of <i>Salmonella</i> Reservoirs in Birds of Prey Hosted in an Italian Wildlife Centre: Molecular and Antimicrobial Resistance Characterisation

In the European Union, salmonellosis is one of the most important zoonoses reported. Poultry meat and egg products are the most common food matrices associated with Salmonella presence. Moreover, wild and domestic animals could represent an important reservoir that could favour the direct and indirect transmission of pathogens to humans. Salmonella spp. can infect carnivorous or omnivorous wild birds that regularly ingest food and water exposed to faecal contamination. Birds kept in captivity can act as reservoirs of Salmonella spp. following ingestion of infected prey or feed. In this paper, we describe the isolation of different Salmonella serovars in several species of raptors hosted in aviaries in an Italian wildlife centre and in the raw chicken necks used as their feed but intended for human consumption. Characterisations of strains were carried out by integrating classical methods and whole genome sequencing analysis. The strains of S. bredeney isolated in poultry meat and birds belonged to the same cluster, with some of them being multidrug-resistant (MDR) and carrying the Col(pHAD28) plasmid-borne qnrB19 (fluoro)quinolone resistance gene, thus confirming the source of infection. Differently, the S. infantis found in feed and raptors were all MDR, carried a plasmid of emerging S. infantis (pESI)-like plasmid and belonged to different clusters, possibly suggesting a long-lasting infection or the presence of additional undetected sources. Due to the high risk of fuelling a reservoir of human pathogens, the control and treatment of feed for captive species are crucial