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Domiziana Di Tella, Manuela Tamburro, Giuliana Guerrizio, Incoronata Fanelli, Michela Lucia Sammarco, Giancarlo Ripabelli Department of Medicine and Health Sciences “Vincenzo Tiberio”, University of Molise, Campobasso, ItalyCorrespondence: Giancarlo RipabelliDepartment of Medicine and Health Sciences “Vincenzo Tiberio”, University of Molise, Via De Sanctis, Campobasso 86100, ItalyTel +39 0874 404961Fax +39 0874 404778Email ripab@unimol.itPurpose: Carbapenemases-producing Klebsiella pneumoniae are challenging antimicrobial therapy of hospitalised patients, which is further complicated by colistin resistance. This study describes molecular epidemiological insights into colistin-resistant and carbapenemases-producing clinical K. pneumoniae.Patients and methods: Cultures collected from 26 hospitalised patients during 2014–2017 in the main hospital in Molise Region, central Italy, were characterized. The minimum inhibitory concentration for 19 antibiotics was determined, including carbapenems and colistin. Prevalence of resistance-associated genes was investigated through PCR, detecting blaKPC, blaGES, blaVIM, blaIMP, blaNDM, blaOXA-48, blaCTX-M, blaTEM, blaSHV, and mcr-1,2,3,4,5,6,7,8. The mgrB gene was also analysed in colistin-resistant strains by PCR and sequencing assays. K. pneumoniae were typed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST).Results: Twenty out of 26 K. pneumoniae were phenotypically resistant to carbapenems and 19 were resistant to colistin. All isolates harbored blaKPC, and blaSHV, blaTEM and blaVIM were further the most common resistance-associated genes. In colistin-resistant strains, mcr-1,2,3,4,5,6,7,8 variants were not detected, while mutations and insertion elements in mgrB were observed in 68.4% (n=13) in 31.6% (n=6) isolates, respectively. PFGE revealed 12 clusters and 18 pulsotypes at 85% and 95% cut-off, while the Sequence Types ST512 (n=13, 50%), ST101 (n=10, 38.5%), ST307 (n=2, 7.7%) plus a novel ST were detected using MLST.Conclusion: All K. pneumoniae showed a multidrug-resistant phenotype, particularly to carbapenems and colistin. According to national data, blaKPC was the prevailing carbapenemase, followed by blaVIM, while blaTEM and blaSHV were among the most frequent beta-lactamases. Consistent with previous reports in Italy, ST512 was the most common clone, particularly during 2014–15, whilst ST101 became dominant in 2016–17. Colistin resistance was mainly associated with deleterious mutations and transposon in the mgrB gene. Improvements of surveillance, compliance with infection prevention procedures and antimicrobial stewardship are essential to limit the spread of resistant K. pneumoniae.Keywords: antimicrobial resistance, carbapenems, central Italy, genetic relatedness, hospital infections, mgrB gene

Introduction: Healthcare-associated infections (HAIs) are a major global public health concern, increasing the transmission of drug-resistant infections. This point prevalence survey investigated HAIs occurrence and antimicrobial consumption (AMC) in pre-COVID-19 era in the public hospitals of a region of Central Italy., Methods: Data were collected using the protocol standardised by the European Centre for Disease Prevention and Control., Results: Three-hundred and sixty-four patients were included (59.3% male) in the study. Overall, HAIs prevalence was 6.6% (95%CI 4.4-9.5), ranging from 5.2% to 7.1% within the surveyed hospitals, with at least one infection in 24 patients (ten each in medical and surgical specialties wards, and four in intensive care). Risk factors for HAIs were advanced age, having undergone surgery and wearing invasive devices. At time of the survey, 44.7% (95%CI 39.7-49.9) of patients was under treatment with at least one antibiotic, and AMC varied between 43% and 48% within hospitals. In all hospitals, a prevalence higher than 10% was found for the prescription reasons other than prophylaxis or therapy., Conclusions: The results revealed a HAIs prevalence lower than that estimated compared to the most recent national data, in contrast to higher antimicrobial usage. These findings highlight the need to reinforce hygiene practices and develop bundles for HAIs, as a broad implementation of infection prevention and control practices extensively applied to both hub and spoke hospitals could significantly reduce their occurrence, as well as to implement antimicrobial stewardship for prescriptive appropriateness., Competing Interests: The authors report there are no competing interests to declare., (©2024 Pacini Editore SRL, Pisa, Italy.)

Surgical site infections (SSIs) represent a valid indicator of the healthcare quality. This study described the preliminary results of one-year active surveillance program on colon surgeries in a hospital in Molise region, central Italy. Patients who had undergone colon surgery according to National Healthcare Safety Network were included. Data on intervention, perioperative antibiotic prophylaxis, and SSIs occurrence were collected. Chi-square and Fisher's Exact test were used to evaluate any association between risk factors and SSIs. Sixty-eight patients (mean age 70.6 years) were included, and 44 (64.7%) were males. The most frequent interventions were right (n = 17, 25.0%) and left (n = 15, 22.0%) hemicolectomy. Surgical interventions were largely elective (n = 43, 63.2%) and with laparotomy (n = 56, 82.4%). During hospital stay, 10 (14.7%) SSIs were detected, including five superficial, three deep and two organ/space infections. Three (4.4%) additional SSIs were detected at post-discharge follow-up, for 13 (19.1%; CI95%: 9.7%-28.5%) total cases detected. Metronidazole plus Ceftriaxone (third generation cephalosporin) was the antibiotics combination mostly used (n = 36, 52.9%) for the perioperative antibiotic prophylaxis within 60 minutes of incision. The study underlines the need of improvements of the practices currently adopted, since SSIs could be significantly reduced through a multimodal strategy generating bundles. As third generation cephalosporins may facilitate resistant strains emergence, for perioperative prophylaxis in clean-contaminated interventions with entry into gastrointestinal tract, Cefazolin plus Metronidazole or only second generation cephalosporin are recommended. Due to the large variability of post-intervention antibiotic therapy, antimicrobial stewardship approach is strictly necessary.

This survey investigated on adverse events after vaccination with mRNA BNT162b2 (Comirnaty, Pfizer-BioNTech) vaccine in children aged 5-11 years in central Italy through active surveillance reporting. During December 2021-January 2022, parents of children who undergone vaccination were interviewed using a structured questionnaire. 197 out of 208 contacted parents participated (94.7% response rate), of whom 166 (84.3%) had one child. Of the 229 children, the mean age was 8.9 years, 50.7% were female. 193 (84.3%) had at least one adverse event after the first dose (mean age 9.1 years; 54.4% female), and 146 (73.4%) of 199 after the second (mean age 8.9 years; 54.8% female), which was not administered to 30 children due to previous COVID-19 history. Local symptoms after the first and second dose occurred in 183 (94.8%) and 141 (96.6%) recipients (p = .435), respectively, while systemic reactions in 62 (32.1%) and 34 (23.3%) ( p  = .074). Mild events were reported by 81.7% and 69.8% children after the first and second dose, followed by moderate (3.9% and 10.6%) and severe (1.3% and 0.5%). After each dose, injection site reactions (79.5% and 68.8%) were the most frequent, followed by headache (13.1%) and lymphadenopathy (8.5%) after the first and second dose, respectively. The adverse events were reported to pediatricians only for 5.7% and 3.9% of children and treated for 17.6% and 15.8%. This is the first report about safety profile through active surveillance of mRNA BNT162b2 among children in Italy, revealing temporary and mild-to-moderate symptoms with no serious events after each vaccine dose.

A booster dose after primary COVID-19 vaccination series was considered crucial after the emergence of the B.1.617.2 (Delta) and B.1.1.529 (Omicron) variants. Active surveillance was used to investigate reporting of adverse events post-booster dose of either of the licensed mRNA Comirnaty (Pfizer/BioNTech) or Spikevax (Moderna) vaccines in adult (17 years and older) recipients in central Italy. Eligible participants were enrolled and interviewed via phone using a structured questionnaire. Primary outcomes related to the occurrence of adverse events post-booster were stratified by vaccine, and frequency of local/systemic, mild/moderate/severe events. Of a total of 622 participants interviewed, 554 (89.1%) reported at least one adverse event (88.2% and 92.9% after the Comirnaty or Spikevax vaccine, respectively): 63.4% were female, and 78.5% aged 17 to 64 years, regardless of vaccine. 87.7% and 68.2% of all recipients described at least one local or systemic reaction, respectively: 97.3, 38.6 and 4.7% reported mild, moderate, or severe events, respectively. The most frequent adverse reactions were pain, redness, or swelling at the injection site and fatigue, while malaise and fever significantly occurred after the Comirnaty, and vomiting after the Spikevax booster. Compared to the primary vaccination, lymphadenopathy was more common after the booster (p < 0.001), especially after Comirnaty vaccine. The study findings revealed no serious or unexpected adverse events, and are in agreement with data available on booster dose for both mRNA vaccines. The transient, mild to moderate, and common to very common side reactions reported should be used to reassure potential recipients of the lack of safety concerns., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

This study describes a SARS-CoV-2 outbreak caused by the Delta (B.1.617.2) variant in a nursing home in Central Italy during October-November 2021. Trained interviewers collected data from residents, staff, and administration officers with an agreed informed consent procedure. Thirty-two (44.5%) out of 72 residents (median age 89 years) and six (26.1%) of 23 healthcare workers were found to be infected with SARS-CoV-2. Infections occurred more often among residents with a higher index of independence in daily living activities, suggesting an increased risk for those with more interactions. Twenty-five infected residents (78.1%) received the booster dose of mRNA anti-COVID-19 vaccine > 7 days before SARS-CoV-2 onset. Half of the infected residents had mild symptoms, and only three required hospitalisation, one of whom died from COVID-19 complications. The study underlines the effectiveness of a booster dose in providing a high protection against severe disease and hospitalisation even among vulnerable individuals infected with the Delta variant of concern., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Pest infestation in any stage can lead to a quality reduction in the finished products. This study aimed to detect Escherichia coli, Salmonella spp., Campylobacter spp., and Staphylococcus aureus in Alphitobius diaperinus adults, and in samples from broiler swabs, administered water and feed collected in a single house from a broiler production facility in central Italy. Three samplings were carried out, each collecting ninety adult beetles for microbial detection in the external, faecal and internal content; ten cloacal swab samples; and one sample of both administered feed and water. Microbiological cultures and biochemical identification were performed on suspected cultures and confirmed by species-specific PCRs. A. diaperinus was abundantly found near the windows, under the manger and in the corners of the facility. Salmonella enterica serovar Cholerasuis was found at the external surface of the beetles, while Staphylococcus xylosus and E. coli were in the faecal content. The latter micro-organism together with Staphylococcus lentus, S. xylosus and other staphylococcal species were detected in the internal microbiota. E. coli and Campylobacter spp. were observed in cloacal swabs, and S. xylosus in one feed sample. The study findings support evidence for Salmonella spp. and E. coli, and remark that adherence to sanitation rules and biosecurity procedures are required., (© 2022 The Society for Applied Microbiology.)

Objectives: to provide an outline of the factors mainly influencing severe disease and fatal outcome among Italian COVID-19 patients in the pre-vaccination phase, also describing the impact of the scenarios driven by variants, vaccines, and available therapies., Design: a literature search was carried out for peer-reviewed articles searching for COVID-19 and prognosis, including severe disease and death., Setting and Participants: Italian patients with COVID-19., Main Outcome Measures: the association between risk factors and severe disease and death as the main outcomes was assessed through epidemiological measures, including relative risk, odds ratio, and hazard ratio., Results: advanced age, obesity, overweight, non-0 blood group, and male gender were the factors more associated with severe disease. Fatal outcome mostly correlated with old age, non-0 blood group, and obesity, together with cardiovascular diseases, diabetes, hypertension, cancer, chronic kidney disease, and acute kidney injury., Conclusions: clinical and epidemiological characteristics of the Italian population, integrated with omics data, could be highly valuable to stratify risk of worse prognosis among patients, and to address targeted prevention and treatment interventions.

In this cross-sectional study, adverse events after the first and second dose of BNT162b2 mRNA (Pfizer-BioNTech, Comirnaty) vaccine against coronavirus disease 2019 were investigated among employees of clinics in central Italy. A 42-items questionnaire was administrated to vaccine recipients. Adverse events were classified based on severity and occurrence as reported in the literature. A descriptive/univariate analysis using Chi-square or Fisher's Exact tests was performed. Odds ratio (OR) and 95% confidence intervals were calculated to assess risk factors. 340 individuals (61.5% females; median age 49 years) participated. Adverse events were reported by 279 (82%) and 281 (82.6%) individuals as induced by the first and second dose, respectively. Mild reactions were mainly reported (80.9% and 80.3%), followed by moderate (11.8% and 37.1%) and severe (3.8% and 4.7%). Adverse events were identical to those already described as very common (81.8% and 80.6%), although vaccine-coincidental events not cited in the literature were reported by 6% and 15.6% following each dose. Age ≤ 55 years was a risk factor for any adverse event after each injection (ORs: 2.942 and 2.818), as well as female sex for those mild (ORs: 1.856 and 2.818) and common (ORs: 3.452 and 2.145). Findings were consistent with national reports as most of the adverse events were mild and associated with female sex and young age, while investigations are needed for reactions not described elsewhere. Data are useful to support the vaccine safety profile, also because largely targeted healthcare personnel more skilled than general population in self-diagnosis of health-related issues., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

General Practitioners (GPs) have good knowledge of health status and sociocultural backgrounds of their patients, thus could be decisive in early detection of cases of asbestos-related diseases (ARDs). In this cross-sectional pilot study, we investigated perception and knowledge on asbestos risk amongst a sample of GPs practicing in Molise region, central Italy, who have anonymously completed a 29-items questionnaire specifically developed for this research. Based on GPs' answers, we obtained scores and classified awareness and knowledge into four percentage quartile classes including inadequate (0-24%), poor (25-49%), moderate (50-74%) and high (75-100%), and scarce (0-24%), sufficient (25-49%), good (50-74%) and optimal (75-100%), respectively. Twenty-eight GPs (median age 63 years; 82% male) participated. Perception and knowledge among participants varied from 62 to 84% and from 18 to 42%, respectively, with 65 and 42% showing an adequate level, respectively. Our survey revealed a satisfactory perception and knowledge of general features about this topic, while a poor awareness of detailed aspects, underlining the need of continuing medical education in the field of occupational medicine. The study remarks that criteria for ARDs diagnosis should be better specified, introducing a detailed list to harmonize medical protocols for symptoms identification and patients' examination. Hence, increasing awareness of asbestos risk and ARDs among GPs represents a crucial opportunity, because of their prominent position to promptly take care of patients and provide targeted information., Competing Interests: Conflict of interest statement The authors declare no conflict of interest., (©2021 Pacini Editore SRL, Pisa, Italy.)

This study compares healthcare-associated infections (HAIs) prevalence and antimicrobial consumption (AMC) from surveys conducted in 2016 and 2019 in an Italian hospital for acute care. HAIs prevalence was 7.1% in both surveys, while patients were under antibiotic treatment slightly increased from 2016 to 2019, from 42.6% to 43.7%, respectively. In the survey of 2016, HAIs risk factors were fatal McCabe score and hospitalization longer than two weeks, while CVC and urinary catheter in that conducted in 2019. In both surveys, CVC and peripheral venous catheter resulted associated to AMC. HAIs and AMC remained stable although the hospital undergone a complexity reduction in the second survey, remarking that critical actions are strictly required implementing infection control practices and antimicrobial stewardship.

Background: The epidemic dynamics of COVID-19 in the Molise region, central Italy, has dramatically changed from the beginning of May 2020, which was when infections were reported amongst Romani people. The aims of this study were to describe the characteristics of an outbreak that occurred in the Romani community and the interventions implemented for control. Methods: A retrospective analysis of outbreak data was performed to describe the SARS-CoV-2 transmission dynamics. Results: A young Romani woman was the first case reported and epidemiological investigation established a possible link with the funeral of a deceased member of this community. In total, 150 close contacts within 34 family groups in two cities were traced, and 109 (72.7%) Romani individuals were found to be infected by COVID-19. The patient's median age was 31 years, 58% were female, and the highest (20.2%) incidence occurred in the 0-9 years age group. A total of 26 (23.8%) patients developed typical SARS-CoV-2 symptoms, 15 (57.8%) were hospitalized, and 21 (22.1%) had comorbidities [most commonly hypertension (28.6%) and/or coronary heart diseases (23.8%)]. The outbreak was effectively controlled through compulsory quarantine and enhanced active surveillance. Conclusions: This is the first study providing insight into COVID-19 transmission dynamics among a Romani population living in Italy. These findings support general conclusions about the role of crowded social gatherings in SARS-CoV-2 spread, the high communicability among close contacts and household settings, and the impact of asymptomatic carriers. These features are of relevance to certain Romani customs where family gatherings are a fundamental pillar of their lives. Although difficulties emerged in interacting with Romani people related to cultural drivers, beliefs, and lifestyle, the outbreak management was effective and should be considered as a valuable model applicable to similar incidents occurring in minority populations., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ripabelli, Sammarco, Cannizzaro, Montanaro, Ponzio and Tamburro.)

The development of high-throughput omics technologies represents an unmissable opportunity for evidence-based prevention of adverse effects on human health. However, the applicability and access to multi-omics tests are limited. In Italy, this is due to the rapid increase of knowledge and the high levels of skill and economic investment initially necessary. The fields of human genetics and public health have highlighted the relevance of an implementation strategy at a national level in Italy, including integration in sanitary regulations and governance instruments. In this review, the emerging field of public health genomics is discussed, including the polygenic scores approach, epigenetic modulation, nutrigenomics, and microbiomes implications. Moreover, the Italian state of implementation is presented. The omics sciences have important implications for the prevention of both communicable and noncommunicable diseases, especially because they can be used to assess the health status during the whole course of life. An effective population health gain is possible if omics tools are implemented for each person after a preliminary assessment of effectiveness in the medium to long term.

The emergence of novel resistant markers hampers the efficacy of beta-lactam antibiotics to treat infections caused by micro-organisms carrying such resistances. This study investigated the antimicrobial susceptibility pattern, the carpapenem-associated determinants and the molecular epidemiology of Klebsiella pneumoniae showing a New Delhi (NDM) metallo-β-lactamase phenotype, isolated from a patient admitted to intensive care unit of the main hospital for acute care of Molise region, central Italy. Antimicrobial susceptibility was assessed for nineteen antibiotics by disc diffusion and agar dilution methods. Carbapenem-associated resistance determinants were detected through gene-specific amplifications, targeting bla NDM-1 , bla SHV and bla TEM , bla CTX-M , bla KPC , bla VIM , bla IMP , bla GES and bla OXA-48-lixe . Molecular characterization was carried out through multilocus sequence typing. The strain showed a multidrug resistant profile, and PCR and sequencing confirmed the presence of bla NDM-1 gene. Among the multiple resistance-associated determinants tested, the isolate, which was assigned to the sequence type ST11, only harboured bla SHV and bla TEM genes. This is the first report of NDM-1 variant in the regional healthcare setting for acute patients, raising significant concerns about the increase in the antimicrobials resistance spread through a different mechanism from the endemic KPC carbapenemase, and underlining the circulation of a virulent clone never identified before in this area., (© 2020 The Society for Applied Microbiology.)

Human Papillomavirus (HPV) is among the most common sexually transmitted infections in both females and males across the world that generally do not cause symptoms and are characterized by high rates of clearance. Persistent infections due at least to twelve well-recognized High-Risk (HR) or oncogenic genotypes, although less frequent, can occur, leading to diseases and malignancies, principally cervical cancer. Three vaccination strategies are currently available for preventing certain HR HPVs-associated diseases, infections due to HPV6 and HPV11 low-risk types, as well as for providing cross-protection against non-vaccine genotypes. Nevertheless, the limited vaccine coverage hampers reducing the burden of HPV-related diseases globally. For HR HPV types, especially HPV16 and HPV18, the E6 and E7 oncoproteins are needed for cancer development. As for other tumors, even in cervical cancer, non-coding microRNAs (miRNAs) are involved in posttranscriptional regulation, resulting in aberrant expression profiles. In this study, we provide a summary of the epidemiological background for HPV occurrence and available immunization programs. In addition, we present an overview of the most relevant evidence of miRNAs deregulation in cervical cancer, underlining that targeting these biomolecules could lead to wide translational perspectives, allowing better diagnosis, prognosis and therapeutics, and with valuable applications in the field of prevention. The literature on this topic is rapidly growing, but advanced investigations are required to achieve more consistent findings on the up-regulated and down-regulated miRNAs in cervical carcinogenesis. Because the expression of miRNAs is heterogeneously reported, it may be valuable to assess factors and risks related to individual susceptibility., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

In July 2018, a Klebsiella pneumoniae carbapenemase (KPC)-producing Escherichia coli ST131 was isolated from a patient admitted to the Vascular Surgery Unit of the main hospital of Molise region, Central Italy. Sequencing and alignment with the available sequences revealed that the isolate harbored the KPC-2 variant and TEM-1 beta-lactamase. This observation raises great concerns about the spread of carbapenem resistance in national and local settings with high endemicity level of KPC in K. pneumoniae , and underlines the importance of strengthening a proactive surveillance.

This study aimed to characterize serovar 1/2a, 1/2b, 1/2c and 4b of Listeria monocytogenes cultures based on High Resolution Melting (HRM) profiles, targeting 53 fragments in the region comprising prs, Listeria Pathogenicity Island-1 (LIPI-1) and ldh loci, and 28 fragments of inlAB operon. Fifty L. monocytogenes cultures (28 of lineage I, 22 of lineage II) were tested. Real time PCR and EvaGreen-based HRM assays were performed, and the HRM profile for each amplicon was compared to that of L. monocytogenes EGD-e strain. Considering all fragments tested, 45 HRM profiles were identified (Diversity Index = 99.35). Similarity analysis identified two main clusters: the first consisted of 25 cultures, including all 1/2a and 1/2c strains, except for three isolates from food of serovar 4b; the second group only included 1/2b and 4b isolates. Eighteen out of targeted amplicons showed constant HRM characteristics irrespective of the serovar/lineage, and hlyA displayed the most stable melting behavior. Conversely, thirteen amplicons were specific for 1/2b and 4b isolates, showing major variations within actA, followed by prs, prfA, mpl and plcB genes. A fragment targeting an intragenic region (part of plcA and part of an unknown gene) had a melting profile allowing differentiation between 4b and 1/2b isolates showing different Tm variants. Amplicons of inlA and inlB exhibited the largest intra-strain melting differences, and one inlB fragment could allow discriminating between 4b and 1/2b cultures, as well as between lineages. A greater level of genetic diversity amongst 1/2a cultures compared to 1/2c, 1/2b and 4b isolates was observed, with variations identified in LIPI-1, as well as within inlA and inlB genes. Sequencing analysis of amplicons with differential HRM profile from EGD-e confirmed point mutations. The study findings underlines that HRM-based approach may be useful for bacterial subtyping for epidemiological purposes when sequencing-based methods are not available., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Healthcare associated infections (HAIs) are a major public health concern both in acute hospitals and long-term care facilities (LTCFs), considering the clinical presentations and related costs. The study describes the first regional experience on HAIs and antimicrobials consumption obtained through point prevalence survey (PPS) 2016-2017 in the "Antonio Cardarelli" acute hospital, and in the LTCF of Larino municipality (Healthcare-associated infections and Antimicrobial use in European Long-Term care facilities, HALT-3) of Molise Region, Central Italy, both coordinated by the European Center for Disease Control (ECDC)., Methods: PPS was carried out during 24-26 November 2016, while HALT-3 on 29 June 2017. Standardized protocols and software provided by ECDC were used to collect facilities information, patients/residents, HAIs and antibiotic consumption data., Results: An active HAI was found in 16 (7.1%) of 224 admitted patients in the acute hospital, and 50% occurred in a clinical specialty ward. Bundles, checklist for HAIs prevention and antimicrobial stewardship were lacking. Among 29 LTCF residents the HAIs prevalence was 3.4%. Surveillance programs for HAIs and antimicrobial stewardship strategy were not available., Conclusion: The overall results identified the need and types of interventions to be implemented in both settings, either at organizational or individual level, to improve antibiotic prescription practice and limit risk factors involved in potentially preventable infections., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: Despite the potential presence in many workplaces, biological risk is little known and largely underestimated, except in healthcare environments. Objectives: To evaluate perception, knowledge and behaviors adopted for protection and prevention of biological risk, related also to the occupational activity, among construction and agricultural workers., Methods: Workers were recruited at farms and construction companies of Molise Region, Central Italy, which were visited within the surveillance activities planned by local institutions. A questionnaire was administered to workers to collect sociodemographic and occupational data, and concerning risk perception and knowledge, and awareness of available prevention measures, also related to the work activity., Results: total of 143 workers (73.4% males, 42.9±10.7 years old) were interviewed, and 56.6% were construction workers. Only 24% of workers believed that infectious diseases could be frequent and health threatening in carrying out the work activities; half of them declared to be exposed to biological hazards, but 70% were unworried. An underestimation of the severity of some pathologies, such as listeriosis, leptospirosis and viral hepatitis emerged, and only 70% of workers believed that tetanus is highly or very highly dangerous for health. Furthermore, 33% of workers said to have scarce or insufficient knowledge of biological risk, and 97.2%, 80.4% and 45.5% were aware that a vaccine is available for tetanus, hepatitis B and A, respectively. All workers undergone the tetanus vaccination and 39.2% the anti-hepatitis B., Conclusions: Despite the high probability of exposure to biological agents, the results of the survey revealed a lack of perception and knowledge of the risk, as well as a poor awareness of available preventive measures among construction and agricultural workers. These data point out the need to activate information and training programs on exposure to biological agents and on both emerging and re-emerging infectious diseases potentially related to work activities, and to provide prevention procedures specific for these workplaces., Competing Interests: The authors of this article have no conflict of interests to disclose., (Copyright© by Aracne Editrice, Roma, Italy.)

Aims: A high resolution melting (HRM) assay was developed for characterizing lineage II Listeria monocytogenes based on the amplification and the melting profiles analysis of 81 fragments targeting the region from the prs to ldh loci, including the Listeria Pathogenicity Island-1 (LIPI-1) genes and the inlAB operon., Methods and Results: Real-time PCR and HRM protocols were standardized using 10 replicate assays from L. monocytogenes EGD-e reference strain (serovar 1/2a). Twenty wild-type isolates of serovar 1/2a and two of serovar 1/2c were tested, and differences between EGD-e strain and the wild-type isolates were defined if the melting temperature (T m ) of an amplicon was not within the lower and the upper limits calculated from replicate testing on EGD-e. The analysis revealed 17 and 19 HRM profiles with respect to prs/LIPI-1/ldh and inlAB target regions (Simpson's Index of Diversity 0·979 and 0·983) respectively. The 1/2c cultures showed 98·1% similarity to melting characteristics with EGD-e, whilst 1/2a isolates had the greatest heterogeneity that was related to inlA, inlB and actA genes. Sequencing of amplicons generating different T m values from EGD-e confirmed the presence of point mutations., Conclusions: This method was useful for L. monocytogenes subtyping based on single nucleotide polymorphisms detection through the melting behaviour analysis of main virulence genes., Significance and Impact of the Study: The study underlines the effectiveness of HRM in differentiating L. monocytogenes strains with high discriminatory power, thus rendering it useful for epidemiological surveillance., (© 2018 The Society for Applied Microbiology.)

Background: The World Health Organization's Action Framework for tuberculosis elimination in low-tuberculosis incidence countries includes the screening for active and latent tuberculosis in selected high-risk groups, including health care workers. In this context, medical and health profession students, exposed to nosocomial tuberculosis transmission during training and clinical rotations, are target populations for tuberculosis screening. No updated data are available on tuberculosis screening practice and knowledge of medical and health profession students in Italy., Methods: Within the activities Italian Study Group on Hospital Hygiene of the Italian Society of Hygiene, Preventive Medicine and Public Health, we carried out a multicentre cross-sectional study to assess knowledge, attitude and practices on tuberculosis prevention and control among Medical, Dentistry, Nursing and other health professions' students. Students were enrolled in the study on a voluntary basis and were administered a previously piloted structured questionnaire. Logistic regression models were applied to explore knowledge on tuberculosis prevention by selected socio-demographic variables and University-based tuberculosis prevention practice., Results: Students of seventeen Universities across Italy participated in the study, and 58.2% of them received compulsory tuberculin skin test either at enrollment or while attending clinical practice. A total of 5,209 students filled the questionnaire. 37.7% were medicine and dentistry students (Group 1), 44.9% were nursing students (Group 2) and 17.4% were other health professions' students (Group 3). Age and gender had different distributions by groups, as well as knowledge and practice on tuberculin skin test. 84.4% of the study population (95% CI = 83.3-85.3) was aware of the existence of the tuberculin skin test, 74.4% (95% CI = 73.2-75.6) knew what is the first-level screening test for latent tuberculosis and only 22.5% (95% CI = 21.4-23.6) knew how to proceed after a positive tuberculin skin test result. Overall, knowledge on tuberculosis prevention was higher in Group 2 and lower Group 3, as compared to Group 1., Conclusion: In Italy, the knowledge on tuberculosis screening among University students is generally good. To reduce some of the criticalities found among the different study courses, it would be appropriate to harmonize both the regulations on tuberculosis screening practices for admission to University courses, and the educational activities on the topic of tuberculosis, to be extended to all workers involved in health care setting.

Introduction: Among the health professions with a long period of training, the students of the Nursing Bachelor's Degree are the most exposed to biological risk resulting from accidents, in particular with needles and cutting edges. The aim of the study was to estimate the frequency and the circumstances for the occurrence of needle stick injuries, as a knowledge base for targeted prevention interventions., Methods: The study was carried out between May and July 2017 in 11 Universities in Italy and 1 in Albania (associated with the "Tor Vergata" University of Rome). An anonymous semi-structured questionnaire was proposed to 1st (second semester), 2nd and 3rd year students of Nursing Bachelor's Degree., Results: A total of 2742 questionnaires were collected. The average age of participants was 22.9 years (median 22, range 19-60 years), 73% of whom were females. A total of 381 injuries were reported. Three hundred and sixteen students (11.8%) underwent at least 1 injury (12.7% among females, 9.7% among males); 41 students declared two or more injuries; four students did not report the number of injuries occurred. The first injury occurred, as an average, 17 days after the start of the internship (median 15 days) and, in 25% of the cases, during the first 9 days. The highest percentage of accidents occurred during the first internship (25.3% of the total) and decreased with the progress of the training path. The injuries occurred in 38% of cases during drug preparation, 24% when disposing of sharp devices, 15% while re-capping needles, 13% during blood sampling and 10% in other circumstances. In 51.2% of cases, the needle was not sterile. Among the nursing students who suffered a needle stick injury, 58.1% declared that they had performed the post-exposure prophylaxis. 96% of students stated to be vaccinated against Hepatitis B virus. Amongst the students who had their serological status checked (74%), 18% stated the antibody titre was not protective. 49.8% of students answered to have been trained in advance on the correct procedures to avoid needle stick and cutting edges injuries in each clinical ward attended, 41.2% referred that this occurred only in some wards and 10% in no ward at all., Conclusion: The results of this study show a high percentage of needle stick injuries in students of the Nursing Bachelor's Degree. Therefore, there is a need for careful reflection on the most effective methods of targeted training acquisition of knowledge, skills and behavioural models useful for the exercise of the profession.

Antimicrobial-resistant Klebsiella pneumoniae represent a global public health concern. K. pneumoniae strains isolated during 2010 and 2014-2016 within a single hospital of Molise Region, Central Italy, were analyzed testing antimicrobial susceptibility, clonality by pulsed-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD)-PCR, and prevalence of carbapenem resistance genes by PCR. Forty isolates (23 wild-type in 2010 and 17 non-wild-type in 2014-2016) were collected from hospitalized patients (65.2 ± 18.1 years old, 75% male, 80% from intensive care unit-ICU). K. pneumoniae showed multidrug-resistant profiles and 15 resistotypes were identified (discriminatory power D = 0.88). The 69.6 and 17.4% of isolates in 2010 resulted intermediate and resistant to imipenem, respectively, and 91.3% was sensitive to meropenem, while 88.2% of isolates of 2014-2016 were resistant to both antibiotics. PFGE identified 16 clusters versus 23 by RAPD, 26 pulsotypes versus 33 RAPD patterns (D ≥ 0.97). PFGE separated strains according to isolation period and identified an outbreak occurred in the ICU during December 2014 and January 2015. No strains harbored bla GES , bla IMP , bla NDM-1 , and bla OXA-48 genes, as well as AmpC plasmid-mediated beta-lactamases genes. Only K. pneumoniae isolated during 2014-2016 were bla KPC positive. Prevalence of bla VIM was 87 and 76.5% during 2010 and 2014-2016, respectively. No strains colistin-resistant harbored mcr-1 plasmid-mediated resistance gene. The study findings underline an increased circulation of multidrug-resistant K. pneumoniae within the hospital, and the acquisition of carbapenem resistance mechanism. The implementation of surveillance and molecular characterization of isolates are needed to identify outbreaks, reduce the spread of resistance, and guide empirical therapy.

Aims of the study were to evaluate Human Papillomavirus (HPV) and type-specific prevalence in four anatomical sites in HIV infected men who have sex with men (MSM) compared with HIV uninfected MSM. Participants were recruited among the attendees of Infectious Diseases Clinics in Central Italy. A trained medical practitioner collected by interview sociodemographic data and information on medical history, sexual behavior, and drug use. Swabs from anal canal, oral cavity, urethral mucosa, and coronal sulcus were tested for HPV DNA and genotyping. Ninety MSM were enrolled, 45 subjects within each group. Overall, 48.9% MSM were HPV positive and prevalence was higher in HIV infected men (60.0% vs 37.8%, P = 0.035). HPV at multiple anatomic sites occurred in 59.1% MSM, with 34.1% and 22.7% at two and three sites, respectively. Prevalence of anal, coronal sulcus, oral, and urethral HPV was 96.3%, 37%, 21.6%, and 18.5% in HIV infected MSM, and 70.6%, 70.6%, 29.4%, and 23.5% among HIV uninfected. A similar proportion of HIV infected and uninfected MSM (59.2% and 58.8%) carried at least one high-risk genotype. Prevalence of types covered by nonavalent vaccine was 77.8% in HIV infected compared with 82.3% in HIV uninfected MSM. HPV 58 and 16 were mostly detected in HIV positive (43.7% and 31.2%) and negative MSM (50.0% and 40.0%). HPV detection rate underlined the high vulnerability of MSM to acquire multisite infections, characterized by various genotype combinations. Since nonavalent vaccine could have prevented 80% of HPV infections, study findings support the implementation of vaccination programs among MSM., (© 2017 Wiley Periodicals, Inc.)

Objective: The aim of this study was to evaluate environmental asbestos sources, mesothelioma incidence and mortality, and awareness on asbestos-related diseases (ARDs) by general practitioners (GPs) in Molise Region., Methods: The contaminated sites in three towns were identified by census; mesothelioma incidence (2000 to 2012) and mortality (2003 to 2013) was achieved from local registries; GPs were interviewed on practiced population's exposures and ARDs diagnosis., Results: About 54.3% of visited sites were contaminated (71.2% by friable asbestos) and 38.8% was extremely damaged. Over above time-periods, 32 mesothelioma cases (62.5% males, 25% in people aged 70 to 75 years) and 27 deaths (90% males, 69 ± 10 years, 70.4% pleural mesothelioma) have been reported. A total of 122 GPs were interviewed who had diagnosed 40 mesothelioma and 28 asbestosis cases., Conclusion: There is the need of remediation/removal interventions for contaminated sites and of strategies to increase GPs awareness on asbestos risks for better patients' management.

Background: Biological risk is a significant issue in workplaces due to continuous modification of work organization and condition., Objectives: In this pilot study, perception and knowledge of biological risk, as well as adoption of prevention measures were evaluated among construction and agriculture workers., Methods: A structured questionnaire was administered to workers to collect socio-demographic and employment information, in addition to data on risk perception and knowledge, and adoption of preventive measures, including those related to work activities., Results: Fifty-three workers aged 45.3±9.8 years participated in the study, 81% were male, and 70% were construction workers. Approximately 80% of participants declared that infectious diseases are neither frequent nor health-threatening in their occupational activity. The majority of workers considered some diseases, such as hepatitis A and B, leptospirosis and listeriosis, as posing a low risk to their health. Forty-seven % claimed to be unexposed to biohazards at their workplace, and 72% were not worried about them. Ninety-six % of the workers knew about vaccination for tetanus, but only 74% and 36% knew about the availability of vaccination for Hepatitis A and B, respectively. During lifetime, 94% of the workers received at least one vaccination. All workers had undergone tetanus vaccination, but only few received vaccine for influenza, hepatitis B and tuberculosis., Conclusions: This pilot study in occupational settings revealed a poor perception of biological risk and a limited awareness of the severity of infectious diseases. Major knowledge gaps existed about the available preventive measures. It is essential to promote safety culture at the workplace also with regard to biological hazards. This requires improving information and education, conducting appropriate studies, and prompting health and safety professional actors to focus on occupational infectious disease problems.

Human papillomavirus (HPV) infection and type-specific prevalence at anal, oral, coronal sulcus, and urethral mucosa in fifty HIV positive men having sex with men (MSM) were evaluated; patients were enrolled in a non-metropolitan area of Central Italy. Clinical and socio-demographic information, drug, and sexual behaviors were obtained for each participant. HPV was detected by PCR from an overall of 200 specimens, and genotyping was performed by both Restriction Fragment Length Polymorphism analysis and sequencing. HPV DNA was found in 60.0% (n = 30) of HIV positive MSM, and prevalence was higher at anal canal (n = 28, 56.0%) compared to all the other anatomical sites (χ(2) test P < 0.01) of coronal sulcus (n = 11, 22.0%), oral (n = 8, 16.0%), and urethral mucosa (n = 5, 10.0%). We found 63.3% (n = 19) of MSM with at least one high-risk genotype, and HPV-58 was more frequently detected (n = 9, 47.4%) respect to HPV-16 (n = 6, 31.6%). This is the first report on HPV detected at four anatomical sites involved in sexual practices in HIV positive MSM. We found an unusual distribution of oncogenic genotypes with an exceeding prevalence of HPV-58 respect to HPV-16. Hence, the recently licensed nonavalent vaccine should be suitable to prevent a larger number of infections caused by potentially emerging high-risk genotypes., (© 2015 Wiley Periodicals, Inc.)

In this study, tolerance at sublethal concentration of benzalkonium chloride and transcription levels of mdrL, ladR, lde, sigB and bcrABC genes in Listeria monocytogenes strains were evaluated. Viable cells reduction occurred in 45% of strains and clinical isolates showed lower sensitivity than isolates from foods. An increased transcription of an efflux system encoding gene was found in 60% of strains, and simultaneous mdrL overexpression and ladR underexpression occurred in 30% of isolates. A significant association between reduced benzalkonium chloride activity and both mdrL and sigB overexpression was observed; sigB expression also correlated with both mdrL and ladR genes. The bcrABC gene was only found in six strains, all isolated from foods and sensitive to benzalkonium chloride, and in four strains an underexpression was observed. Disinfection at sublethal concentration was less effective in clinical isolates, and mdrL and sigB expression was significantly affected by disinfection. Further insights are needed to understand the adaptation to benzalkonium chloride and to evaluate whether changes in gene expression could affect the L. monocytogenes virulence traits and persistence in the environment., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Listeria monocytogenes virulence depends on the activity of well-characterized virulence factors. In this study, transcription levels of inlA, inlB, hly, bsh and prfA genes in L. monocytogenes strains, and the ability of invasion into CaCo-2 cells were investigated. Serotyping, multiplex-PCR for serovar identification and restriction fragment analysis of inlA were performed. Transcription levels and invasiveness were evaluated by quantitative reverse-transcription PCR and by in vitro assays, respectively. The isolates were of serovars 1/2a, 4b, 1/2c, 1/2b and 3a. Full-length inlA profiles were found for nine of ten clinical isolates, while five of seven cultures from foods showed truncated profile. The analysis of transcription levels of virulence factors encoding genes demonstrated a substantial inter-strain heterogeneity, with clinical strains showing higher levels for almost all genes than isolates from food. A correlation between transcription levels of inlA and inlB, as well as between bsh and prfA, was observed. Significant differences between clinical strains and food isolates in the invasion of CaCo-2 cells were found. Analysis of gene transcription and invasiveness of human cells suggests different virulence phenotypes among L. monocytogenes populations, and this characterization could be a useful tool for risk assessment purposes and for the development of public health strategies., (© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Molecular typing and fingerprinting of microbial pathogens represent an essential tool for the epidemiological surveillance, outbreak detection and control of infectious diseases. Indeed, epidemiological investigation without genotyping data may not provide comprehensive information to allow the most appropriate interventions; despite this consideration, some barriers still hamper the routine application and interpretation of molecular typing data. In this paper, the most important methods currently used for characterization of pathogenic microorganisms for microbial source tracking and for the identification of clonal relationships among different isolates, are described according to their principles, advantages and limitations. Criteria for their evaluation and guidelines for the correct interpretation of results are also proposed. Molecular typing methods can be grouped into four categories based on different methodological principles, which include the characterization of restriction sites in genomic or plasmid DNA; the amplification of specific genetic targets; the restriction enzyme digestion and the subsequent amplification; sequence analysis. Although the development and the extensive use of molecular typing systems have greatly improved the understanding of the infectious diseases epidemiology, the rapid diversification, partial evaluation and lack of comparative data on the methods have raised significant questions about the selection of the most appropriate typing method, as well as difficulties for the lack of consensus about the interpretation of the results and nomenclature used for interpretation. Several criteria should be considered in order to evaluate the intrinsic performance and practical advantages of a typing system. However none of the available genotyping methods fully meets all these requirements. Therefore, the combined use of different approaches may lead to a more precise characterization and discrimination of isolates than a single method, especially if used in a hierarchical manner. The interpretation of the molecular results differs according to the typing system's characteristics: for example in the restriction fragments-based analysis, the divergences or the similarity percentages among the profiles are evaluated, whilst the differences in terms of number and intensity of bands are analyzed in the amplification-based approaches. Moreover, a correct interpretation of molecular results significantly depends by other critical factors, such as the comprehension of the typing system and data quality, the microbial diversity, and the epidemiological context in which the method is used. The analysis of PFGE data, considered as the "gold standard", is based on the differences of the number and position of bands patterns, although recent recommendations are now available from the Centers for Diseases Control and Prevention (CDC) for a more accurate interpretation, which also include the evaluation of the gel quality, the genetic diversity of the microorganism, the time and geographical scale of an epidemic event. Future advances in the molecular typing technologies indeed will provide rapid methodological improvements, such as a greater degree of automation, better resolution, higher throughput, and a greater availability of dedicated bioinformatics tools. These factors will all contribute to an increasing application of genotyping methods to better understand the epidemiology of infectious diseases, and to implement, along with the strengthened international and interdisciplinary partnerships, more effective control and prevention strategies for Public Health improvements.

Background: The exposure to air pollution has negative effects on human health, increasing the risk of respiratory diseases, such as asthma. Few data are yet available on the epidemiology of childhood asthma in some areas of Italy. The aim of the study was to estimate asthma prevalence and related risk factors in children and adolescents residents around the industrial area of Termoli, Molise region, Central-South Italy., Methods: Prevalence was assessed through the administration of modified ISAAC questionnaires filled out by parents of 89 children and adolescents for the identification of confirmed and probable cases, and by analyzing pediatricians' databases on drug prescriptions for symptoms control and treatment of assisted population in the study area (n = 1,004), compared to a control area (n = 920) with lower industrialization. The association of asthma with risk factors was evaluated by univariate (Chi-square or Fisher's Exact test) and regression logistic analysis., Results: A total of 22 (24.7%) asthmatics were identified, including both confirmed (n = 7; 7.9%) and probable cases (n = 15; 16.8%), most of them (n = 17; 77.3%) resident of Termoli town. All asthma cases were georeferenced based on the residence, however clusters were not found. Using drug prescriptions analysis, a higher prevalence (n = 138; 13.7%) of diagnosed cases was found. Lifetime history of both atopic dermatitis and bronchitis were significantly relateds to asthma cases, as well as an elevated body mass index, whose association is consistent with prevalence data of overweight/obese children living in the study area. Moreover, being resident of the town of Termoli was associated to the occurrence of cases., Conclusions: Although our data indicated a prevalence concordance with previous national studies in pediatric population, a definitive correlation with environmental industrial factors present in the study area was not established. However, asthma outcome was significantly associated to individuals living in the town of Termoli that, despite the industrial/manufacturing activities, is also subjected to a higher environmental pressure due to the presence of toll road, state highway, railroad, and seaport which may cause air pollution from motor vehicle traffic and increase asthma induction. This study provides hitherto unavailable data on asthma in childhood population living in an industrialized area which was never investigated before, could be part of a systematic review or meta-analysis procedure, might suggest significant findings for larger observational studies, and contribute to complete the frame of disease epidemiology in Italy.

Objective: We examined persistence and clearance of human papillomavirus (HPV) infections and risk factors associated with persistence in 79 women based on the results of two sequential tests performed over 12-24 months., Study Design: Between February 2008 and August 2009, 398 women aged 18-63 years were examined for presence of cervical HPV infection by cervical scrape specimen and PCR. Detection was performed using Linear Array (LA) HPV Genotyping Test. All women were interviewed, and a short questionnaire was administered to collect information on socio-demographic characteristics, sexual and reproductive history, smoking habits, oral contraceptive use, history of sexually transmitted diseases, and Chlamydia trachomatis or Mycoplasma spp. infections. Pearson's χ² test was used to verify the association between all independent variables with the response variable., Results: Initially, high risk-HPV (HR-HPV) and low risk-HPV (LR-HPV) infection was detected in 69.6% and 30.4% of the women, respectively, whilst multiple infections occurred in 53.2%. HPV 16 was the most common (20.2%) high-risk type, followed by 52, 31 and 53. At follow-up, HR-HPV infection was detected in 50.6% of the women; among these, 67.5% had persistent infection, while 12.5% acquired other high-risk types, and 20.0% of those positive for LR-HPV at entry had a new HR-HPV infection. Multiple infections were detected in 38.0% of the women. HPV 16 and 31 were the most frequent types, followed by HPV 73. Type-specific HR-HPV persistence was found in 49.1% of women. HPV 31, 39 and 73 were the most frequently persistent types, whilst HPV 16 was the least persistent. No significant age difference between women with persistence or clearance was found. The highest HR-HPV persistence occurred in the 22-27 years old group, whereas clearance increased in women aged 28-33 years. No significant association between persistent HR-HPV infection and oral contraceptive use, smoking habits and history of sexually transmitted disease was detected both at entry and follow-up study. The association between C. trachomatis or Mycoplasma spp. and HPV persistence could not be investigated because of the low detection rate of these microorganisms., Conclusions: The persistence of HR-HPV infection level was similar to that reported elsewhere, and HPV 31, 39 and 73 showed the highest likelihood of persistence, partially in agreement with other studies. The clinical relevance of the low persistence of HPV 16 and other HR-HPV is unknown. Persistent HR-HPV infection in women aged 22-27 years was in agreement with other authors. To the best of our knowledge, this is the first report on persistence of HR-HPV infections in Italy in a general population, although we examined a small sample in a short follow-up time., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

A 5'-exonuclease real-time triplex-PCR assay was developed for serogrouping Listeria monocytogenes, and differentiation from other Listeria spp. The assay was evaluated on 109 Listeria cultures, and results were compared with a previously validated gel-based multiplex-PCR procedure. All L. monocytogenes were correctly classified into four serogroups, including atypical serotype 4b strains, and differentiated from other Listeria species. The assay is a rapid method for categorisation of suspect L. monocytogenes., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Vibrio spp. infections still are a Public Health concern. Vibrio spp. can be found in marine, estuarine, and freshwater environments, and can be able to cause diseases in fish, shellfish, mammals, as well as in humans. Since '80 to date, the number of species within the genus increased from 21 to more than 100. The most important is Vibrio cholerae, the etiological agent of the cholera, responsible of seven pandemics; serotypes O1 and O139 can produce cholera toxin, while serotypes non-O1/non-O139 are generally associated with sporadic cholera cases and extraintestinal infections. Vibrio parahaemolyticus is an important cause of gastroenteritis associated with contaminated seafood consumption, whereas Vibrio vulnificus and V. alginolyticus can be related to wound infections or seafoodborne primary septicemia in immunocompromised patients. Disease prevention is mainly based on the application of proper individual or collective preventive measures.

Fifty Listeria monocytogenes strains were genotyped by sAFLP and PCR products were separated by agarose gel and automated chip-based microfluidic electrophoresis. A high congruency of results was observed comparing the two techniques, although for some cultures a better separation of sAFLP fragments was achieved with microfluidic system, which proved to be a highly reliable and reproducible tool to improve the molecular typing of L. monocytogenes, requiring lower volumes of samples and reducing significantly analysis time., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Pandemic influenza virus has been implicated in serious lower airways illness and death in subjects both with and without underlying medical conditions. Predictive factors for severe disease in healthy individuals have not been identified., Case Report: Severe A(H1N1)-associated pneumonia occurring in a healthy subject without underlying medical conditions sequential to a Clamidophila pneumoniae infection, is reported., Conclusion: A potential synergistic mechanism by which other pathogens could interfere with the clinical course of A(H1N1) infection, is suggested.

Aims: The microbiological and toxicological quality of 51 samples of dried herbs (Melissa officinalis, Salvia officinalis, Malva sylvestris, Matricaria chamomilla, Alchemilla vulgaris and Centaurea cyanus) cultivated in family-managed farms in Molise Region (Italy) was evaluated., Methods and Results: All the samples were analysed by using conventional methods, and for samples preparation, an alternative Washing and Shaking (WaS) protocol was developed to reduce release of antimicrobial compounds. None of the samples were of unsatisfactory quality with respect to aflatoxin B1, and only three samples from Malva sylvestris exceeded the limit of total aflatoxins according to Recommendation 2004/24/EC. The International Commission on Microbiological Specifications for Foods limits for mesophilic bacteria and total coliforms were exceeded in the 29.4 and 3.9% of samples, respectively: 7.8% of samples also exceeded the limit for Escherichia coli established by European Spice Association. When the 'WaS' method was used, higher microbial counts were obtained, especially for A. vulgaris, S. officinalis and M. officinalis., Conclusions: Herbs cultivated in family-managed small agricultural areas showed a good microbiological and toxicological quality, irrespectively of preliminary washing or selection procedures., Significance and Impact of the Study: Herb matrices may contain antimicrobial activity which should be considered when applying the conventional microbiological methods for sample preparation. Alternative preparation protocols may have advantages to reduce antimicrobial effects and should be further evaluated., (© 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.)

The prevalence of virulence and cytolethal distending toxin (CDT) genes and the cytotoxic activity in Vero and HEp-2 cells was estimated in 29 Campylobacter jejuni and 36 Campylobacter coli from foods, animals and humans isolates. All C. jejuni showed flaA, cadF, cdtA, cdtB, cdtC and cdt cluster genes fragments, except for ceuE (86.2%) and cdt genes (93.1%). Amongst C. coli strains, a lower prevalence of ceuE gene (83.3%) was detected than that for cdtA, cdtB, cdtC genes (97.2%), cdt gene cluster (94.4%) and cdt genes (86.1%); whereas flaA and cadF genes were amplified in all isolates. Despite the high prevalence of CDT genes only 8 (27.6%) C. jejuni and 1 (2.8%) C. coli showed evidence for cytotoxin production in HEp-2 cells. However, how CDT positive and CDT negative strains differ in their biological properties remains unknown, but the relative higher prevalence of cytotoxicity in C. jejuni could be consistent with its predominant epidemiological role in human infections., ((c) 2009 Elsevier Ltd. All rights reserved.)

Aims: In this study, 105 Listeria monocytogenes strains isolated from humans, foods and environmental samples were characterized using several typing methods. Moreover, serotyping procedure was evaluated, and a cost-effective methodological approach based on preliminary PCRs screening was proposed., Methods and Results: The isolates were analysed by conventional serotyping, multiplex-PCRs for serogroup and lineage identification and PCR-RFLP of inlA gene to identify potentially noninvasive L. monocytogenes. Among the strains, only the serotypes 1/2a, 1/2c, 1/2b, 4b and 3a were identified. The isolates were classified into serogroups I (58.10%), II (22.85%), III (12.38%) and IV (6.67%). Among clinical strains, lineage I was more represented (68.75%) than lineage II; whereas, lineage II was more associated with food (90.24%) and environmental (85.72%) isolates. Most of food (89.02%) and environmental (85.71%) isolates were classified into truncated InlA profiles, whereas the 93.75% of clinical strains were associated with a complete form of the protein., Conclusion: Molecular techniques were sensitive and specific for classifying strains into serogroup and lineage and in agreement with the serotyping. Moreover, a preliminary PCRs-based screening was proposed to select only the necessary antisera by a flow chart; this methodological approach allows cost saving up to 42%. Our results further suggest the role of InlA protein in human listeriosis, particularly in immunocompetent individuals, and a correlation between truncated protein and serotype., Significance and Impact of the Study: This study further validates molecular methods for L. monocytogenes analysis and proposed a new cost-effective approach for serotyping. It could help to improve a national surveillance network for L. monocytogenes infections in Italy.

We estimated the prevalence of Campylobacter spp. in retail meat (n = 352 samples; 104 chicken, 106 pork, and 142 beef) collected in Campobasso, Italy, comparing two microbiological methods. All the isolates were characterized by biomolecular techniques for epidemiological purposes. Campylobacter isolation was performed by selective culture and membrane filtration methods. Phenotypic and genotypic methods for genus and species identification were evaluated together with antimicrobial resistance and plasmid profiling. Sixty-nine (86.2%) samples were positive by selective culture, 49 (61.2%) by membrane filtration, and 38 (47.5%) by both methods. Only 74 of 80 strains were confirmed as Campylobacter spp. by PCR, and two Campylobacter coli were identified as Campylobacter jejuni. Chicken meat was more frequently contaminated than other meats. Selective culture was more sensitive than membrane filtration (85 versus 66%), and specificity of the methods was 98 and 100%, respectively. Among Campylobacter isolates from chicken meat, 86.5% were multidrug resistant. Resistance to ciprofloxacin (51.3%) and enrofloxacin (52.7%) was lower than to nalidixic acid (71.6%). C. coli strains showed the highest cross-resistance for quinolones (82.6%) and fluoroquinolones (60.9%) as well as a high resistance to tetracycline. Plasmids were isolated from six C. coli and two C. jejuni isolates, but no association was detected between antimicrobial resistance and plasmid DNA carriage. Selective culture is considered as the optimal method for Campylobacter isolation, although it was unable to detect all contaminated samples. Membrane filtration provided more specific results but with low sensitivity. A combination of both techniques may offer better results.

We examined the prevalence of HR- and LR-HPV by Linear Array genotyping test in 299 women aged 18-63 years who consecutively visited Molise Region main hospitals for routine Pap smear between February and August 2008. Ninety women were positive for any HPV (30.1%), and 66 for any HR-HPV (22.1%). The most prevalent HR-HPV types were HPV 16 (22.2% of all women with HPV infection), HPV 53 (14.4%), and HPV 66 (14.4%). HPV infections increased from 15.8% in the 18-20 years group to 50.0% in the 21-23 years group and then decreased to 9.1% in those aged 50 years or more (p=0.008). Multiple HPV infections were observed in 15.7% of the study sample (52.2% of all HPV positive). There is a significantly higher prevalence of multiple infections in 18-32 years group women (24.5%) compared with females aged 33 years or more (6.8%) (p<0.005). Current smokers were at increased risk of HPV infection (44.2% of HPV infections compared with 23.5% in never smokers, and 25.3% of multiple HPV infections compared with 11.3%; p=0.001). HR-HPV infections were higher in women never been pregnant (27.1% compared with 7.7%; p=0.001). Oral contraceptive use was completely unrelated to infection. Among the 122 women who had both cytological examination and HPV results, multiple HR-HPV types were found in 36.8% of those with abnormal cervical findings, and in 13.6% of those with normal cervical findings (p=0.05). The results of the present investigation provide further evidence for the notion that cervical HPV infection is more widespread than previously suggested.

In this study, we compared and validated a nested ABC polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) assay to amplify Cryptosporidium parvum oocyst wall protein (COWP) gene fragment with a previous nested PCR-RFLP method. When RFLP of COWP gene did not provide clear results, we further analysed samples by cloning and sequencing for species and genotype identification. ABC-PCR was performed on DNA extracted from human faecal samples collected in England where Cryptosporidium was previously detected by conventional methods. COWP gene amplification was successful for all specimens; RFLP analyses using both RsaI and AluI enzymes recognised 54/55 restriction profiles. Among these, 40 (72.7%) characteristic patterns were generated for C. parvum and six (10.9%) for Cryptosporidium hominis; four (7.3%) showed species mixtures, while the remaining four samples (7.3%) showed atypical genotypes. Cloning and sequencing of mixtures and atypical genotypes identified one C. parvum, one C. hominis and three Cryptosporidium meleagridis. Indeed, molecular tools improved PCR-RFLP results. In conclusion, this study showed that ABC-PCR-RFLP technique was quick, simple and specific. Cloning and sequencing were helpful in characterising mixtures or previously uncharacterised RFLP patterns. All these epidemiological findings are useful information for any preventive intervention to control disease diffusion.

The study was performed to estimate the prevalence and antimicrobial sensitivities of Listeria spp. in raw milk, feaces end environmental samples isolated from 10 dairy in Molise Region. A total of 454 samples were collected, which comprised 40 raw milk, 40 animal faeces and 374 environmental samples. Listeria monocytogenes was never isolated from raw milk specimens; one was isolated from faeces speciments and two were isolated from environmental samples. All isolates were resistant to two or more of antimicrobial agents tested (cephalotin, ampicillin, tetracycline, co-trimoxazole, erytromicin, clindamycin, gentamicin, oxacillin). One isolate of L. monocytogenes was susceptible to all antimicrobial agents tested except oxacillin. This study indicates that faeces, equipments and environment are important reservoirs of Listeria spp. in dairy farm, and can represent potential source of contamination of raw milk. However, the contamination of milk, and the risk of infection, can be effectively eliminated by pasteurisation process.

Recovery of Cryptosporidium parvum oocysts in a fecal suspension that experimentally contaminated onto lettuce leaves was investigated. Material recovered from the lettuce samples by washing in detergent solutions were concentrated by filtration using the Envirochek Sampling Capsule. Oocysts were concentrated by immunomagnetic separation (IMS) and detected by microscopy following modified Ziehl-Neelsen (MZN) staining. Cryptosporidal DNA was detected using a nested-PCR assay for amplification of a fragment of the Cryptosporidium oocyst wall protein (COWP) gene, which was applied to DNA extracted from both filtrates, and material recovered from MZN stained smears on glass slides after microscopy. No Cryptosporidium were detected by microscopy or by PCR of un-inoculated lettuce leaves. After IMS, means of 0-6.5% of the total numbers of oocysts inoculated were recovered and detected by microscopy. Detection by PCR was less sensitive than microscopy. There was a strong association between successful PCR amplification, the numbers of oocysts detected by microscopy and the numbers of oocysts in the inoculum. This study confirms that C. parvum oocysts can be recovered from contaminated lettuce using filtration and IMS, and detected by microscopy and PCR. However, further developments are required to improve recovery of this parasite.