Alphitobius diaperinus Panzer (Insecta, Coleoptera) in a single house of a broiler production facility as a potential source of pathogenic bacteria for broilers and humans.

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.

New Delhi metallo-ß-lactamase (NDM-1)-producing Klebsiella pneumoniae of sequence type ST11: first identification in a hospital of central Italy.

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-ß-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 blaNDM-1 , blaSHV and blaTEM , blaCTX-M , blaKPC , blaVIM , blaIMP , blaGES and blaOXA-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 blaNDM-1 gene. Among the multiple resistance-associated determinants tested, the isolate, which was assigned to the sequence type ST11, only harboured blaSHV and blaTEM 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.

The Clean Care Contest: promoting hand hygiene among healthcare and medical students.

INTRODUCTION: Despite continuing efforts, compliance rates and knowledge of best practices in hand hygiene remain disappointing. Recognizing that conventional educational tools seem out of touch with young people and that the med and messages contents need refreshing, the Italian Study Group of Hospital Hygiene of the Italian Society of Hygiene, Preventive Medicine and Public Health devised a novel approach to promote the creation of innovative educational tools for improving knowledge of, and compliance with, hand hygiene rules among healthcare and medical students. METHODS: A contest in creating educational material on hand hygiene practices involved university students of nursing and medicine, and of other healthcare degrees. Students from the universities of the GISIO network were invited to create educational material (e.g., videos, posters, presentations, leaflets, and screensavers) to be presented by May 5th 2019 during the World Hand Hygiene Day / Save Lives: Clean Your Hands Global Annual Initiative of the World Health Organization). A local and a national winners were awarded. RESULTS: Three different local and national contests were performed during 2016, 2017 and 2018. During the three-year period, more than 270 educational tools have been developed: 130 (48%) were judged useful for hand hygiene promotion campaigns. The most frequent projects participating in the contests were videos (39%), posters (29%), leaflets (14%), and others (18%) submitted by more than 1,500 students of nursing (40%), medicine (31%), dentistry (7%), and of other healthcare courses in 14 universities. Products were evaluated by a local committee and, subsequently, local winners represented their University in a national contest. CONCLUSIONS: The contest provided a framework for the creation of innovative and potentially effective educational tools via an engaging approach that leveraged student creativity. Given the need to improve compliance rates, this study suggests that new ways can be advantageously explored to teach hand hygiene procedures and increase awareness of the importance of their consistent use among healthcare and medical students.

Results of a survey of procedures for cleaning and disinfecting ambulances.

BACKGROUND: In 2014, the Italian Study Group of Hospital Hygiene of the Italian Society of Hygiene, Preventive Medicine and Public Health (GISIO-SItI), in collaboration with the National Association of Medical Hospital Managers (ANMDO), conducted a survey on the availability of procedures for cleaning and disinfecting ambulances in order to assess the practices in use. METHODS: An online questionnaire was prepared through the Survey Monkey® platform and a web link access was sent to a convenience sample of ANMDO doctors working in healthcare management in public and private healthcare facilities. RESULTS: Ninety-six questionnaires were collected (26% response rate). In 73% of cases there was a procedure for cleaning and disinfecting ambulances, which had been produced at a company level (67%) and involved various professionals. In 21% of cases the procedure had been prepared in expectation of an epidemic or following an epidemic (5%). The recommendations had been presented to the staff (90%), in 28% of cases through training events with verification of the knowledge acquired. Monitoring of the implementation of the procedure is planned in the majority of cases (88%), mainly through direct observation (92%). In 67% of cases the tender specifications for ambulance services did not include a section dedicated to cleaning and disinfection and, in the absence of a procedure, this was provided by the hospital in only 51% of case. CONCLUSION: This survey represented a first step towards the development of guidelines for standardising procedures and providing indications useful for their evaluation and monitoring their implementation.

Hospital hygiene in Italy: the GISIO-SItI survey.

BACKGROUND: In Italy there are no rules concerning the establishment of a hospital hygiene structure in hospitals and other healthcare settings, and the hospital organization plans vary widely. The aim of the survey, carried out by the Italian Study Group of Hospital Hygiene of the Italian Society of Hygiene, Preventive medicine and Public health, was to evaluate the presence in the hospital organization plan of a structure referred to as Hospital hygiene, or including in its denomination the words "hygiene" or "hospital hygiene", the activities carried out, the relation to other areas, like patient safety, the type and quantity of professionals involved, the strengths and the critical aspects. METHODS: A semi-structured questionnaire was administered to Healthcare Trusts representing all Italian Regions through the members of the above Study Group. RESULTS: 35 Trusts, 13 in Northern, 8 in Central, 14 in Southern Italy (including Sicily and Sardinia), completed the questionnaire. In 19 Trusts (54.3%) a structure whose denomination included the words "hospital hygiene" or "hygiene" was present. The activities related to the management of infectious risk were most represented, carried out autonomously or in collaboration, but many other activities were covered. In all hospitals the activities of the Hospital Hygiene Unit inter-linked with those of the clinical risk, with different forms of collaboration. CONCLUSION: This survey, even though on a limited sample, provided a picture of hospital hygiene at a national level, showing a considerable heterogeneity and highlighting critical issues but also strengths. It is essential to share organizational and management models that enhance and promote hospital hygiene, to ensure the appropriateness of healthcare practices offered in a safe and comfortable environment to patients, operators, and visitors.

Procedures in endoscope reprocessing and monitoring: an Italian survey.

BACKGROUND: The high diffusion of endoscopes worldwide and the need for effective reprocessing methods requested the development of guidelines and implementation of surveillance procedures at local level. STUDY DESIGN: In order to collect data on everyday's practice and adherence to available guidelines, endoscopy units from different public institutions were surveyed using a dedicated questionnaire. METHODS: Between July and November 2015 a survey was carried in 12 main hospitals from 10 different Italian regions, involving 22 endoscopy units. The state of the art of national and international guidelines was investigated to compare the protocols adopted at local level. RESULTS: In all the surveyed hospitals, the reprocessing activity is based on pre-established protocols in adherence with principal guidelines. Enzymatic detergents, which are recommended by the international guidelines, are used in 55.6% of units and peracetic acid is currently the most widely used chemical disinfectant. Discrepancies were observed in the application of periodic quality controls. CONCLUSION: Updated guidelines are generally applied in reprocessing practice. Quality controls may represent a critical issue to improve effectiveness and surveillance. The whole of acquired data can promote a positive trend towards the application of best practices.

A multicentre study on epidemiology and prevention of needle stick injuries among students of nursing schools.

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.

Knowledge, experiences, and attitudes toward Mantoux test among medical and health professional students in Italy: a cross-sectional study.

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.

High resolution melting analysis for the characterization of lineage II Listeria monocytogenes serovars 1/2a and 1/2c based on single nucleotide polymorphisms identification within the Listeria Pathogenicity Island-1 and inlAB operon: a novel approach for epidemiological surveillance.

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 (Tm ) 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 Tm 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.

Definition of criteria and indicators for the prevention of Healthcare-Associated Infections (HAIs) in hospitals for the purposes of Italian institutional accreditation and performance monitoring.

BACKGROUND: Healthcare-associated infections (HAIs) are an important issue in terms of quality of care. HAIs impact patient safety by contributing to higher rates of preventable mortality and prolonged hospitalizations. In Italy, analysis of the currently available accreditation systems shows a substantial heterogeneity of approaches for the prevention and surveillance of HAIs in hospitals. The aim of the present study is to develop and propose the use of a synthetic assessment tool that could be implemented homogenously throughout the nation. METHODS: An analysis of nine international and of the 21 Italian regional accreditation systems was conducted in order to identify requirements and indicators implemented for HAI prevention and control. Two relevant reviews on this topic were further analyzed to identify additional evidence-based criteria. The project team evaluated all the requirements and indicators with consensus meeting methodology, then those applicable to the Italian context were grouped into a set of "focus areas". RESULTS: The analysis of international systems and Italian regional accreditation manuals led to the identification respectively of 19 and 14 main requirements, with relevant heterogeneity in their application. Additional evidence-based criteria were included from the reviews analysis. From the consensus among the project team members all the standards were compared and 20 different thematic areas were identified, with a total of 96 requirements and indicators for preventing and monitoring HAIs. CONCLUSIONS: The study reveals a great heterogeneity in the definition of accreditation criteria between the Italian regions. The introduction of a uniform, synthetic assessment instrument, based on the review of national and international standards, may serve as a self-assessment tool to evaluate the achievement of a minimum standards set for HAIs prevention and control in healthcare facilities. This may be used as an assessment tool by the Italian institutional accreditation system, also useful to reduce regional disparities.

High Resolution Melting as a rapid, reliable, accurate and cost-effective emerging tool for genotyping pathogenic bacteria and enhancing molecular epidemiological surveillance: a comprehensive review of the literature.

INTRODUCTION: Rapid, reliable and accurate molecular typing methods are essential for outbreaks detection and infectious diseases control, for monitoring the evolution and dynamics of microbial populations, and for effective epidemiological surveillance. The introduction of a novel method based on the analysis of melting temperature of amplified products, known as High Resolution Melting (HRM) since 2002, has found applications in epidemiological studies, either for identification of bacterial species or molecular typing, as well as an extensive and increasing use in many research fields. HRM method is based on the use of saturating third generation dyes, advanced real-time PCR platforms, and bioinformatics tools. OBJECTIVE: To describe, by a comphrehensive review of the literature, the use, application and usefulness of HRM for the genotyping of bacterial pathogens in the context of epidemiological surveillance and public health. MATERIAL AND METHODS: A literature search was carried out during July-August 2016, by consulting the biomedical databases PubMed/Medline, Scopus, EMBASE, and ISI Web of Science without limits. The search strategy was performed according to the following keywords: high resolution melting analysis and bacteria and genotyping or molecular typing. All the articles evaluating the application of HRM for bacterial pathogen genotyping were selected and reviewed, taking into account the objective of each study, the rationale explaining the use of this technology, and the main results obtained in comparison with gold standards and/or alternative methods, when available. RESULTS: HRM method was extensively used for molecular typing of both Gram-positive and Gram-negative bacterial pathogens, representing a versatile genetic tool: a) to evaluate genetic diversity and subtype at species/subspecies level, based also on allele discrimination/identification and mutation screening; b) to recognize phylogenetic groupings (lineage, sublineage, subgroups); c) to identify antimicrobial resistance; d) to detect and screen for mutations related to drug-resistance; e) to discriminate gene isoforms. HRM method showed, in almost all instances, excellent typeability and discriminatory power, with high concordance of typing results obtained with gold standards or comparable methods. Conversely, for the evaluation of genetic determinants associated to antibiotic-resistance or for screening of associated mutations in key gene fragments, the sensitivity and specificity was not optimal, because the targeted amplicons did not encompass all the crucial mutations. CONCLUSIONS: Despite the recent introduction of sequencing-based methods, the HRM method deserves consideration in research fields of infectious diseases, being characterized by low cost, rapidity, flexibility and versatility. However, there are some limitations to HRM assays development, which should be carefully considered. The most common application of HRM for bacterial typing is related to Single Nucleotide Polymorphism (SNP)-based genotyping with the analysis of gene fragments within the multilocus sequence typing (MLST) loci, following an approach termed mini-MLST or Minim typing. Although the resolving power is not totally correspondent to MLST, the Simpson's Index of Diversity provided by HRM method typically >0.95. Furthermore, the cost of this approach is less than MLST, enabling low cost surveillance and rapid response for outbreak control. Hence, the potential of HRM technology can strongly facilitate routine research and diagnostics in the epidemiological studies, as well as advance and streamline the genetic characterization of bacterial pathogens.

[Molecular epidemiology of infectious diseases: analytical methods and results interpretation]. / Epidemiologia molecolare delle malattie infettive:metodi di analisi ed interpretazione dei risultati .

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.

[Vibrio spp. infections of clinical significance and implication for public health]. / Infezioni da Vibrio spp. di interesse clinico e problematiche di Sanitå Pubblica.

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.

Microbiological and toxicological quality of dried herbs.

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.

Typing of Listeria monocytogenes strains isolated in Italy by inlA gene characterization and evaluation of a new cost-effective approach to antisera selection for serotyping.

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.

[Alcohol consumption, pregnancy and fetal alcohol syndrome: implications in public health and preventive strategies]. / Alcolismo, gravidanza e fetopatia alcolica: implicazioni in Sanità Pubblica e strategie preventive.

Alcohol consumption during pregnancy is a significant public health problem and is an established cause of serious birth defects and developmental delay collectively described as fetal alcohol syndrome (FAS). FAS is caused by congenital alcohol-induced damages and is a cause of mental retardation. It is characterised by facial abnormalities and growth deficiency. Infants affected by the syndrome show intellectual impairment, and difficulties in learning, memory, problem-solving, and attention as well as experiencing additional problems with mental health and social interactions. However, an absence of the characteristic facial defects and growth deficiency may result in a failure to identify children with prenatal alcohol exposure, which can further present as alcohol-related neurodevelopment disorder (ARND) or alcohol-related birth defects (ARBD). Estimates of prevalence of FAS in U.S.A. range between 0.3 to 2.2 per 1,000 live births, but much higher rates occurring in some communities. Harmonisation of the methodology used for epidemiological studies, with research activities that establishes real baseline prevalence of FAS and identification of women who are at highest risk of bearing a FAS-affected child, are an essential prerequisite to prevention. In addition, it is essential to assess different FAS preventive approaches through carefully controlled studies. Universal, selected, and indicated preventive strategies have been identified, targeting different kind of populations. Since FAS and other adverse effects of drinking during pregnancy are theoretically completely preventable, it is vital to make more efforts to improve the application of the most appropriate interventions. Although in Italy alcohol consumption has constantly increased, mainly amongst the young (including women of childbearing age), knowledge on FAS and alcohol-related effects is completely lacking. Because of the high cost for care of individuals with this syndrome, it is essential to apply appropriate interventions to prevent this problem.

[Prevalence of Listeria spp. in dairy farm and evaluation of antibiotic-resistance of isolates]. / Prevalenza di Listeria spp. in aziende produttrici di latte e valutazione dell'antibiotico-resistenza dei ceppi isolati.

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.

[Detection of Salmonella, Listeria spp., Vibrio spp., and Yersinia enterocolitica in frozen seafood and comparison with enumeration for faecal indicators: implication for public health]. / Ricerca di Salmonella, Listeria spp., Vibrio spp. e Yersinia enterocolitica in prodotti ittici congelati e surgelati del commercio e correlazione con gli indici di contaminazione fecale: implicazioni in Sanità Pubblica.

Infections transmitted through consumption of contaminated seafood is a significant source of human morbidity. The aim of this study was to compare the detection of Salmonella, Listeria, Vibrio, and Yersinia enterocolitica in frozen seafood with results from enumeration of conventional faecal indicators. A total of 213 crustaceans or molluscs were purchased from local vendors in Italy: 74% were harvested in Italy, 25% from other European countries and 1% from outside Europe. Listeria spp. was isolated from 20% of samples, Vibrio spp. from 11%, Salmonella from 3% and Y. enterocolitica from 1%. Listeria species isolated were L. monocytogenes, L. innocua, L. welshimeri, L. ivanovii and L. seeligeri. Vibrio species isolated were V. alginolyticus and V. fluvialis. The most contaminated shellfish for both faecal indicator microrganism and pathogens were hen clams (6% contained Salmonella, 27% Listeria spp. and 3% Y. enterocolitica), while from 27% of shrimps Vibrio spp. was recovered. Higher levels of faecal indicators were recovered from samples harvested outside Europe, and 66% of samples harvested in Thailand were contaminated from Salmonella. Significant differences were found in the levels of contamination of seafoods depending upon the freezing regime, but there was a limited association between presence of potential pathogens (particularly Vibrio spp.) and conventional faecal indicators. Hence, we suggest reconsideration of current legal parameters to evaluate microbiological quality of seafood.

Detection of Cryptosporidium parvum oocysts in experimentally contaminated lettuce using filtration, immunomagnetic separation, light microscopy, and PCR.

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.