Monophasic Variant of Salmonella Typhimurium 4,[5],12:i:- (ACSSuGmTmpSxt Type) Outbreak in Central Italy Linked to the Consumption of a Roasted Pork Product (Porchetta).

The monophasic variant of S. Typhimurium 4,[5],12:i:- (MVST) is the third most commonly reported Salmonella serovar involved in human infections (8.8%) in the EU and ranks after S. Enteritidis (54.6%) and S. Typhimurium (11.4%). In Italy, in contrast, the MVST has achieved peculiar epidemiological and ecological success which has allowed it to be, since 2011, the serovar most frequently isolated from humans. In the summer of 2022, a foodborne outbreak of the MVST involving 63 people occurred in the Marche Region (Central Italy). A common food exposure source among some human cases was a roasted, ready-to-eat (RTE) pork product, porchetta, which is a typical product of Central Italy. This paper describes the results of investigations conducted to clarify this outbreak. The porchetta was produced by a local manufacturing plant and distributed to at least two local retail stores, one of which was the retail outlet for the manufacturing plant. The MVST was isolated from surface samples collected at the porchetta manufacturing plant and at both local retail stores via bacterial analysis, and the porchetta sampled at one store contained the MVST. These data confirm this type of RTE pork product can be a source of Salmonella infection in humans.

Isolation and Characterisation of Human-Derived blaKPC-3-Producing Salmonella enterica Serovar Rissen in 2018.

In this study, we describe a Salmonella enterica serovar (S.) Rissen strain with a reduced susceptibility to meropenem, isolated from a urinary infection in an 89-year-old woman in 2018 during activity surveillance in Italy (Enter-Net Italia). The genomic characteristics, pathogenicity, and antimicrobial resistance mechanisms were investigated via a genomic approach. Antimicrobial susceptibility testing revealed a "susceptible, increased exposure" phenotype to meropenem in the S. Rissen strain (4_29_19). Whole-genome sequencing (WGS) was performed using both the NovaSeq 6000 S4 PE150 XP platform (Illumina, San Diego, CA, USA) and MinION (Oxford Nanopore). The S. Rissen 4_29_19 strain harboured two plasmids: a pKpQIL-like plasmid carrying the blaKPC-3 resistance gene in a Tn4401a transposon (pKPC_4_29_19), and a ColE-like plasmid (p4_4_29_19) without resistance genes, highly prevalent among Enterobacterales. Comparative analysis revealed that the pKPC_4_29_19 plasmid was highly related to the pKpQIL reference plasmid (GU595196), with 57% coverage and 99.96% identity, but lacking a region of about 30 kb, involving the FIIK2 replicon region and the entire transfer locus, causing the loss of its ability to conjugate. To our knowledge, this is the first time that a pKpQIL-like plasmid, carrying blaKPC-3, highly diffused in Klebsiella pneumoniae strains, has been identified in a Salmonella strain in our country. The acquisition of blaKPC genes by Salmonella spp. is extremely rare, and is reported only sporadically. In zoonotic bacteria isolated from humans, the presence of a carbapenem resistance gene carried by mobile genetic elements, usually described in healthcare-associated infection bacteria, represents an important concern for public health.

A Cholera Case Imported from Bangladesh to Italy: Clinico-Epidemiological Management and Molecular Characterization in a Non-Endemic Country.

Despite the number of cholera outbreaks reported worldwide, only a few cases are recorded among returning European travellers. We describe the case of a 41-year-old male, returning to Italy after a stay in Bangladesh, his origin country, who presented with watery diarrhoea. Vibrio cholerae and norovirus were detected in the patient's stools via multiplex PCR methods. Direct microscopy, Gram staining, culture and antibiotic susceptibility tests were performed. The isolates were tested using end-point PCR for the detection of potentially enteropathogenic V. cholera. Serotype and cholera toxins identification were carried out. Whole genome sequencing and bioinformatics analysis were performed, and antimicrobial resistance genes identified. A phylogenetic tree with the most similar genomes of databases previously described was built. Sample of the food brought back by the patient were also collected and analysed. The patient was diagnosed with V. cholerae O1, serotype Inaba, norovirus and SARS-CoV-2 concomitant infection. The isolated V. cholerae strain was found to belong to ST69, encoding for cholera toxin, ctxB7 type and was phylogenetically related to the 2018 outbreak in Dhaka, Bangladesh. Adopting a multidisciplinary approach in a cholera non-endemic country ensured rapid and accurate diagnosis, timely clinical management, and epidemiological investigation at national and international level.

Colistin Resistance Mechanisms in Human Salmonella enterica Strains Isolated by the National Surveillance Enter-Net Italia (2016-2018).

BACKGROUND: A collection of human-epidemiologically unrelated S. enterica strains collected over a 3-year period (2016 to 2018) in Italy by the national surveillance Enter-Net Italia was analysed. METHODS: Antimicrobial susceptibility tests, including the determination of minimal inhibitory concentrations (MICs) for colistin, were performed. Colistin resistant strains were analysed by PCR to detect mobile colistin resistance (mcr) genes. In mcr-negative S. enterica serovar Enteritidis strains, chromosomal mutations potentially involved in colistin resistance were identified by a genomic approach. RESULTS: The prevalence of colistin-resistant S. enterica strains was 7.7%, the majority (87.5%) were S. Enteritidis. mcr genes were identified only in one strain, a S. Typhimurium monophasic variant, positive for both mcr-1.1 and mcr-5.1 genes in an IncHI2 ST4 plasmid. Several chromosomal mutations were identified in the colistin-resistant mcr-negative S. Enteritidis strains in proteins involved in lipopolysaccharide and outer membrane synthesis and modification (RfbN, LolB, ZraR) and in a component of a multidrug efflux pump (MdsC). These mutated proteins were defined as possible candidates for colistin resistance in mcr-negative S. Enteritidis of our collection. CONCLUSIONS: The colistin national surveillance in Salmonella spp. in humans, implemented with genomic-based surveillance, permitted to monitor colistin resistance, determining the prevalence of mcr determinants and the study of new candidate mechanisms for colistin resistance.

A Strong Evidence Outbreak of Salmonella Enteritidis in Central Italy Linked to the Consumption of Contaminated Raw Sheep Milk Cheese.

Salmonellosis is the second most commonly reported gastrointestinal infection in humans after campylobacteriosis, and an important cause of foodborne outbreaks in the EU/EEA. The vast majority (72.4%) of the salmonellosis foodborne outbreaks reported in EU in 2019 were caused by Salmonella Enteritidis, even if their total number due to this serovar decreased. In spring 2020, a foodborne outbreak of S. Enteritidis occurred in the Marche region (Central Italy), involving 85 people. The common exposure source was a cheese, pecorino "primo sale", produced with raw sheep milk. The cheese batches were produced by two local dairies, with a livestock production facility, also including a sheep farm, being part of one dairy. Bacteriological analysis of samples collected allowed the detection of S. Enteritidis in animal faeces, environmental samples, raw-milk bulk tanks and milk taken from single animals. These data confirm that, despite the scarce scientific evidence, S. Enteritidis can infect sheep and be shed into the animals' milk. Hence, this is a real risk for public health when unpasteurized milk is used in production of such cheese. The present paper describes the results of the investigations conducted to clarify this outbreak.

Genetic Profiles and Antimicrobial Resistance Patterns of Salmonella Infantis Strains Isolated in Italy in the Food Chain of Broiler Meat Production.

This work aimed to evaluate the antimicrobial susceptibility of 87 Salmonella Infantis strains isolated in Italy from 2016 to 2019 along the food chain of broiler meat production and in humans and to determine the genetic profiles of the strains in order to establish a possible correlation with the antimicrobial pattern. All isolates were tested by the disk diffusion method to evaluate antimicrobial susceptibility toward sixteen antimicrobials, and the broth microdilution method was used to confirm extended spectrum ß-lactamase (ESBL) production. PCR and pulsed field gel electrophoresis (PFGE) were applied to characterize ESBL-encoding and AmpC ß-lactamase genes and to analyze the S. Infantis strains genetic profiles respectively. S. Infantis isolates showed high prevalence of resistance, in particular toward nalidixic acid (97.7%), tetracycline (96.5%), sulphamethoxazole/trimethoprim (91%) and cefepime (72.4%). The 80.5% of isolates were ESBL, cefotaxime-resistant, carrying the blaCTX-M1 gene. The most prevalent PFGE profile was XbaI.0126 (35.6%). The remaining strains had a genetic homology from 81% to 97% with the XbaI.0126 profile. The strains belonging to these profiles were isolated from different matrices collected along the broiler food chain independently on the year and from the region and there was no correlation between the PFGE profiles and resistance patterns. We found two ESBL-producing S. Infantis strains with the same XbaI.2621 profile isolated from humans and from poultry feces, not yet reported in Italy. Our findings confirmed the diffusion of ESBL-multi drug resistant (MDR) S. Infantis along the broiler food chain and in humans and underlined the importance of continuous monitoring to control and to reduce the prevalence of this bacterium, applying a global One Health approach.

Characterization of CRISPR-Cas Systems in Serratia marcescens Isolated from Rhynchophorus ferrugineus (Olivier, 1790) (Coleoptera: Curculionidae).

The CRISPR-Cas adaptive immune system has been attracting increasing scientific interest for biological functions and biotechnological applications. Data on the Serratia marcescens system are scarce. Here, we report a comprehensive characterisation of CRISPR-Cas systems identified in S. marcescens strains isolated as secondary symbionts of Rhynchophorus ferrugineus, also known as Red Palm Weevil (RPW), one of the most invasive pests of major cultivated palms. Whole genome sequencing was performed on four strains (S1, S5, S8, and S13), which were isolated from the reproductive apparatus of RPWs. Subtypes I-F and I-E were harboured by S5 and S8, respectively. No CRISPR-Cas system was detected in S1 or S13. Two CRISPR arrays (4 and 51 spacers) were detected in S5 and three arrays (11, 31, and 30 spacers) were detected in S8. The CRISPR-Cas systems were located in the genomic region spanning from ybhR to phnP, as if this were the only region where CRISPR-Cas loci were acquired. This was confirmed by analyzing the S. marcescens complete genomes available in the NCBI database. This region defines a genomic hotspot for horizontally acquired genes and/or CRISPR-Cas systems. This study also supplies the first identification of subtype I-E in S. marcescens.

Molecular and Epidemiologic Analysis of Reemergent Salmonella enterica Serovar Napoli, Italy, 2011-2015.

Human infections with Salmonella enterica serovar Napoli are uncommon in Europe. However, these infections represented 5.9% of salmonellosis cases in Italy during 2014-2015. The source of infection is unknown. We analyzed surveillance data and compared strain genetic similarities and found that contaminated vegetables and surface water are probable sources of human infection.

Carbapenemases-producing Klebsiella pneumoniae in hospitals of two regions of Southern Italy.

Carbapenem-resistant Klebsiella pneumoniae infections are reported with increasing frequency elsewhere in the world, representing a worrying phenomenon for global health. In Italy, there are hotspot data on the diffusion and type of carbapenemase-producing Enterobacteriaceae and K. pneumoniae in particular, with very few data coming from Apulia and Basilicata, two regions of Southern Italy. This study was aimed at characterizing by phenotypic and genotypic methods carbapenem-resistant K. pneumoniae isolated from several Hospitals of Apulia and Basilicata, Southern Italy. Antibiotic susceptibility was also evaluated. The relatedness of carbapenemase-producing K. pneumoniae strains was established by pulsed-field gel electrophoresis (PFGE). Among the 150 K. pneumoniae carbapenemase producers, KPC-3 genotype was the most predominant (95%), followed by VIM-1 (5%). No other genotypes were found and no co-presence of two carbapenemase genes was found. A full concordance between results obtained by both the phenotypic and the genotypic tests was observed. All strains were resistant to ß-lactam antibiotics including carbapenems, and among antibiotics tested, only tetracycline and gentamycin showed low percentage of resistance (18% and 15%, respectively). Resistance to colistin was detected in 17.3% of strains studied. The analysis of PFGE profiles of the carbapenemases-positive strains shows that one group (B) of the five (A to E) main groups identified was the most prevalent and detected in almost all the hospitals considered, while the other groups were randomly distributed. Three different sequence types (ST 307, ST 258, and ST 512) were detected with the majority of isolates belonging to the ST 512. Our results demonstrated the wide diffusion of K. pneumoniae KPC-3 in the area considered, the good concordance between phenotypic and genotypic tests. Gentamicin and colistin had a good activity against these strains.

Molecular and Epidemiological Analysis of a Campylobacter jejuni Outbreak in Northern Italy in November 2013.

Campylobacter spp. is the most common gastrointestinal pathogen worldwide with a very low reported incidence in Italy. In November of 2013, local and national public health authorities investigated an outbreak caused by Campylobacter jejuni among children in a kindergarten in Northern Italy. A case was defined as a child who had diarrhea with a laboratory-confirmed diagnosis of C. jejuni between 11 and 30 November. Stool samples from the kindergarten kitchen staff and environmental samples from the kitchen were examined for enteric pathogens. As food leftovers were not available, the menu logbook of the refectory was reviewed to identify a possible source of the outbreak. C. jejuni strains were tested for antimicrobial susceptibility and subtyped by pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). We identified 20 cases among 247 schoolchildren (attack rate = 8%), all who reported having lunch in the kindergarten. The stools from the kitchen staff as well as the environmental samples were negative for enteric pathogens. The identified outbreak strains (n = 5) were sensitive to all of the antimicrobials tested; the first four strains showed an identical PFGE profile, whereas the fifth strain showed a PFGE pattern similarity of 89%. Using MLST, all five strains were assigned to a single sequence type (ST), ST451 (clonal complex, CC21); this was the first identification of ST and the third reported outbreak of C. jejuni in Italy. Molecular typing confirmed that most of the cases belonged to a clonal cluster supporting the hypothesis of a common source; however, the source was not identified. Due to a delayed start of the investigation, it was not possible to perform any microbiological evaluation of the food consumed.

Outbreak of unusual Salmonella enterica serovar Typhimurium monophasic variant 1,4 [5],12:i:-, Italy, June 2013 to September 2014.

Monophasic variant of Salmonella enterica subspecies enterica serovar Typhimurium (monophasic S. Typhimurium), with antigenic structure 1,4,[5],12:i:-, appears to be of increasing importance in Europe. In Italy, monophasic S. Typhimurium represented the third most frequent Salmonella serovar isolated from human cases between 2004 and 2008. From June 2013 to October 2014, a total of 206 human cases of salmonellosis were identified in Abruzzo region (Central Italy). Obtained clinical isolates characterised showed S. Typhimurium 1,4,[5],12:i:- with sole resistance to nalidixic acid, which had never been observed in Italy in monophasic S. Typhimurium, neither in humans nor in animals or foods. Epidemiological, microbiological and environmental investigations were conducted to try to identify the outbreak source. Cases were interviewed using a standardised questionnaire and microbiological tests were performed on human as well as environmental samples, including samples from fruit and vegetables, pigs, and surface water. Investigation results did not identify the final vehicle of human infection, although a link between the human cases and the contamination of irrigation water channels was suggested.

Salmonella enterica Serovar Napoli Infection in Italy from 2000 to 2013: Spatial and Spatio-Temporal Analysis of Cases Distribution and the Effect of Human and Animal Density on the Risk of Infection.

BACKGROUND: Salmonella Napoli is uncommon in Europe. In Italy however, it has been growing in importance since 2000. To date, no risk factors have been identified to account for its rise. This study aims at describing the epidemiology, spatial and spatio-temporal patterns of S. Napoli in Italy from 2000 to 2013, and to explore the role of several environmental correlates, namely urbanization, altitude and number of livestock farms, on the risk of S. Napoli infection among humans. METHOD: Data were obtained from Enter-Net Italy, a network of diagnostic laboratories. The data were aggregated at the municipality level. Descriptive epidemiology, multivariate regression models, spatial and spatio-temporal analyses were performed on the number of cases and incidence rates. RESULTS: S. Napoli showed an expanding trend at the national level, and an increasing number of cases. Compared to the other main serovars in Italy, the risk of S. Napoli infection was higher in the age group <1 year, and lower in the other age groups. Although urbanization and the number of farms were associated with the risk of S. Napoli infection to some extent, their role in the epidemiology of the disease remains inconclusive. S. Napoli cases showed a positive global spatial autocorrelation as well as a significant spatio-temporal interaction. Twenty-four spatial and spatio-temporal clusters were identified, seven purely spatial and 17 spatio-temporal, mainly in north-western Italy. Most of the clusters were in areas characterized by urban and industrial settlements surrounded by agricultural land and an abundance of freshwater bodies. CONCLUSIONS: Our results point to the presence, in a number of areas in Italy, of a Salmonella of public health concern originating in the environment. This highlights the increasing relevance of environmental, non-food-related sources of human exposure to enteric pathogens.

Emergence of Ciprofloxacin-Resistant Salmonella enterica Serovar Typhi in Italy.

In developed countries, typhoid fever is often associated with persons who travel to endemic areas or immigrate from them. Typhoid fever is a systemic infection caused by Salmonella enterica serovar Typhi. Because of the emergence of antimicrobial resistance to standard first-line drugs, fluoroquinolones are the drugs of choice. Resistance to ciprofloxacin by this Salmonella serovar represents an emerging public health issue. Two S. enterica ser. Typhi strains resistant to ciprofloxacin (CIP) were reported to the Italian surveillance system for foodborne and waterborne diseases (EnterNet-Italia) in 2013. The strains were isolated from two Italian tourists upon their arrival from India. A retrospective analysis of 17 other S. enterica ser. Typhi strains isolated in Italy during 2011-2013 was performed to determine their resistance to CIP. For this purpose, we assayed for susceptibility to antimicrobial agents and conducted PCR and nucleotide sequence analyses. Moreover, all strains were typed using pulsed-field gel electrophoresis to evaluate possible clonal relationships. Sixty-eight percent of the S. enterica ser. Typhi strains were resistant to CIP (MICs, 0.125-16 mg/L), and all isolates were negative for determinants of plasmid-mediated quinolone resistance. Analysis of sequences encoding DNA gyrase and topoisomerase IV subunits revealed mutations in gyrA, gyrB, and parC. Thirteen different clonal groups were detected, and the two CIP-resistant strains isolated from the individuals who visited India exhibited the same PFGE pattern. Because of these findings, the emergence of CIP-resistant S. enterica ser. Typhi isolates in Italy deserves attention, and monitoring antibiotic susceptibility is important for efficiently managing cases of typhoid fever.

Diffusion and persistence of multidrug resistant Salmonella Typhimurium strains phage type DT120 in southern Italy.

Sixty-two multidrug resistant Salmonella enterica serovar Typhimurium strains isolated from 255 clinical strains collected in Southern Italy in 2006-2008 were characterised for antimicrobial resistance genes, pulsotype, and phage type. Most strains (83.9%) were resistant to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline (ACSSuT) encoded in 88.5% by the Salmonella genomic island (SGI1) and in 11.5% by the InH-like integron (bla OXA-30-aadA1) and catA1, sul1, and tet(B) genes. STYMXB.0061 (75%) and DT120 (84.6%) were the prevalent pulsotype and phage type identified in these strains, respectively. Five other resistance patterns were found either in single or in a low number of isolates. The pandemic clone DT104 (ACSSuT encoded by SGI1) has been identified in Italy since 1992, while strains DT120 (ACSSuT encoded by SGI1) have never been previously reported in Italy. In Europe, clinical strains DT120 have been reported from sporadic outbreaks linked to the consumption of pork products. However, none of these strains were STYMXB.0061 and SGI1 positive. The prevalent identification and persistence of DT120 isolates would suggest, in Southern Italy, a phage type shifting of the pandemic DT104 clone pulsotype STYMXB.0061. Additionally, these findings raise epidemiological concern about the potential diffusion of these emerging multidrug resistant (SGI linked) DT120 strains.

First isolation of Salmonella enterica serovar Napoli from wild birds in Italy.

Salmonella enterica serovar Napoli (S. Napoli) is an emerging serovar in Italy. It accounts for 2-4% of all serovars isolated from human infections. The zoonotic origin of this serovar is still unknown and this makes difficult to apply any control intervention. We report here the isolation of S. Napoli from a river nightingale (Cettia cetti, Temminck 1820) which represents the first description of this serovar from wild birds. This finding adds knowledge to the ecology of S. Napoli and addresses further studies aimed to assess the epidemiologic link between S. Napoli isolated from wild birds, food, environmental sources and human infections.

Molecular characterization of Salmonella enterica serovar 4,[5],12:i:- DT193 ASSuT strains from two outbreaks in Italy.

Salmonella enterica subsp. enterica serovar 4,[5],12:i:- DT193 is recognized as an emerging monophasic variant of Salmonella Typhimurium in many European countries. Resistance to ampicillin, streptomycin, sulphonamides, and tetracycline (R-type ASSuT) is described as one of the most common profiles of resistance within this clone. Recently, strains presenting such features were isolated from two unrelated outbreaks in Italy. Strains were characterized by pulsed-field gel electrophoresis (PFGE), performed with XbaI, BlnI, and SpeI, and multiple-locus variable-number tandem repeat analysis (MLVA). XbaI-PFGE showed strains related to the two outbreaks as indistinguishable. Conversely, both BlnI-PFGE and MLVA characterized the strains related the two outbreaks as different. XbaI-PFGE identified two profiles, differing by one band, within strains isolated from one of the two outbreaks. Also BlnI-PFGE and MLVA generated different profiles among the strains related to that outbreak. Combining the PFGE profiles obtained by XbaI and BlnI and comparing them with the MLVA profiles, the two methods grouped the same isolates based on identity. Moreover, genomic deletions of the genes included in the operon fljAB, the flanking iroB gene, and the closely located STM2757 gene were investigated. For all strains, the same profile of deletion characterized by the absence of fljA, fljB, and hin genes and the presence of STM2757 and iroB genes was identified. This profile of deletion represents a mixture between two profiles of Salmonella 4,[5],12:i:- described as the "Spanish" and the "U.S." clones. This study demonstrated that although strains of Salmonella 4,[5],12:i:- DT193 ASSuT are highly clonal, minor differences between strains may be seen during the same outbreak by using in parallel PFGE with different restriction enzymes, MLVA, and the analysis of molecular markers related to the operon fljAB. The combination of these different molecular approaches was essential to clarify the epidemiological relationship among the strains.

Resistance genes, phage types and pulsed field gel electrophoresis pulsotypes in Salmonella enterica strains from laying hen farms in southern Italy.

Twenty-four Salmonella enterica isolates (13 serovar Enteritidis and 11 Typhimurium) isolated from 5,600 samples from intensive laying hen farms in Italy in 1998-2007 were characterized for antimicrobial resistance genes, pulsotype and phage type. Most of S. Typhimurium strains were pulsotype STYMXB.0147 (81.8%), phage type DT143 and resistant to sulfamethoxazole encoded by sul2. Two multidrug resistant (MDR) strains were identified. One strain, STYMXB.0061, was resistant to ampicillin (A), chloramphenicol (C), streptomycin (S), sulfamethoxazole (Su) and tetracycline (T) encoded by the Salmonella Genomic Island SGI1. The second MDR strain, STYMXB.0110, was resistant to SSuT encoded by sul1 and sul2, aadA1 and tet(C)-flanked by an IS26 element, respectively. The tet(C) gene has been reported to confer low levels of resistance and it has very rarely been detected in S. Typhimurium from poultry. In the current study, the MIC value (32 µg/mL) was consistent with the breakpoint (≥16 µg/mL) reported for Enterobacteriaceae. Most of the S. Enteritidis strains were resistant to Su (encoded by sul2). One MDR strain (ANxSSuT) was identified. With the exception of nalidixic acid (Nx), the resistances were respectively encoded by bla(TEM), strAB, sul2 and tet(A) harbored by an IncN conjugative plasmid. All isolates were pulsotype SENTXB.0001 with PT14b being the most prevalent identified phage type (57.1%). In Europe, SENTXB.0001 is the predominant PFGE profile from clinical cases and the identification of PT14b has steadily been on the increase since 2001. The findings presented in this study highlight the potential spread of S. Enteritidis phage types PT14b and S. Typhimurium DT143 in a field of particular relevance for zoonoses. Additional, the presence of resistance genes and genetic elements (conjugative plasmid and IS element) underlines the need to assess routinely studies in field, such as poultry farms, relevant fot the public health and suitable for the storage and diffusion of antimicrobial resistance.

Phenotypic and genetic traits of Salmonella enterica subsp. serovar Typhimurium strains causing salmonellosis foci in rabbit farms from Southern Italy in 1999-2003.

In this study, we characterised the Salmonella Typhimurium strains responsible for four outbreaks which occurred in distinct rabbit farms (Southern Italy) from 1999 to 2003. Strains were typed by Pulsed Field Gel Electrophoresis (PFGE) and the genetic basis of antimicrobial resistance was established. A major group of clonally related isolates, pulsotype STYMXB.0061, accounted for three of the salmonellosis foci. Strains were resistant to streptomycin, chloramphenicol, tetracycline, ampicillin and sulphonamides encoded respectively by the aadA2, floR, tetG, blaPSE-1, sul1 gene cluster harboured by a Salmonella Genomic Island 1. The clonally related group of isolates included strains phage type DT104, DT12 or undefined type (NT). The fourth salmonellosis focus was caused by a strain pulsotype STYMXB.0147, resistant to sulphonamides (encoded by sul2) and phage type U302. Results provided first molecular characterisation of S. Typhimurium strains isolated from rabbit farms in Italy and highlighted the presence of the pulsotype STYMXB.0061 even before its wide detection among human clinical isolates collected in Italy in the mid 2000s from clinical cases.

Nucleotide sequence of the chromosomal region conferring multidrug resistance (R-type ASSuT) in Salmonella Typhimurium and monophasic Salmonella Typhimurium strains.

OBJECTIVES: The aim of this study was to sequence the chromosomal region conferring resistance to ampicillin, streptomycin, sulphonamides and tetracycline (R-type ASSuT) in a Salmonella Typhimurium (STM) monophasic strain (4,[5],12:i:-) belonging to the PFGE profile STYMXB.0079. The presence of this resistance region and the analysis of its genetic environment was investigated in a selection of strains. METHODS: A Sau3A1 genomic library was used to determine the nucleotide sequence of the genomic resistance region. PCRs were performed on 10 epidemiologically unrelated Salmonella strains, both STM and monophasic STM, with R-type ASSuT and PFGE profile STYMXB.0079, in order to investigate the presence of the resistance genes, the left and right junctions and the internal regions of the resistance region, as well as the genetic environment. RESULTS: The genomic resistance region consisted of two regions, resistance region 1 (RR1), conferring resistance to ampicillin, streptomycin and sulphonamides, and resistance region 2 (RR2), conferring tetracycline resistance. These resistance regions were both surrounded by IS26 elements and sequence comparative analysis showed 99% sequence identity with a region of plasmid pO111_1 from an Escherichia coli strain. All 10 strains were positive for the four resistance genes, the left and right junctions and the internal regions of RR1 and RR2. Concerning the genetic environment, all the strains lacked the STM1053-1997 and STM2694 genes, while only monophasic STM strains showed deletion of the fljA-fljB operon. CONCLUSIONS: This study describes two resistance regions localized on the bacterial chromosome of a clonal lineage of STM and monophasic STM that are widespread in Italy.