Pulsed-field gel electrophoresis (PFGE) has become the gold standard of molecular methods in epidemiological investigations. In spite of its high resolving power, use of the method has been hampered by inadequate laboratory-to-laboratory reproducibility. In the project described here we have addressed this problem by organizing a multilaboratory effort in which the same bacterial strains (subtype variants of the Iberian and Brazilian methicillin-resistant Staphylococcus aureus —MRSA—clones) were analyzed by twenty investigators in thirteen different laboratories according to an indentical protocol, which is reproduced here in detail. PFGE patterns obtained were analyzed at a central laboratory in order to identify specific technical problems that produced substandard macrorestriction patterns. The results including the specific technical problems and their most likely causes are described in this communication. Also listed are seven major epidemic clones of MRSA which have been characterized by molecular fingerprinting techniques and the prototypes of which have been deposited at the American Type Culture Collection, from where they will be available for interested investigators for the purpose of typing MRSA isolates. It is hoped that this communication will contribute to the improvement of the reproducibility and technical/aesthetic quality of PFGE analysis.

Urinary tract infections (UTIs) are among the most frequent community-acquired infections worldwide.Escherichia coliis the most common UTI pathogen although underlying host factors such as patients’ age and gender may influence prevalence of causative agents. In this study, 61 273 consecutive urine samples received over a 22-month period from outpatients clinics of an urban area of north Italy underwent microbiological culture with subsequent bacterial identification and antimicrobial susceptibility testing of positive samples. A total of 13 820 uropathogens were isolated and their prevalence analyzed according to patient’s gender and age group. OverallEscherichia coliaccounted for 67.6% of all isolates, followed byKlebsiella pneumoniae(8.8%),Enterococcus faecalis(6.3%),Proteus mirabilis(5.2%), andPseudomonas aeruginosa(2.5%). Data stratification according to both age and gender showedE. coliisolation rates to be lower in both males aged ≥60 years (52.2%),E. faecalisandP. aeruginosabeing more prevalent in this group (11.6% and 7.8%, resp.), as well as in those aged ≤14 years (51.3%) in whomP. mirabilisprevalence was found to be as high as 21.2%.Streptococcus agalactiaeoverall prevalence was found to be 2.3% although it was shown to occur most frequently in women aged between 15 and 59 years (4.1%). Susceptibility ofE. colito oral antimicrobial agents was demonstrated to be as follows: fosfomycin (72.9%), trimethoprim/sulfamethoxazole (72.9%), ciprofloxacin (76.8%), ampicillin (48.0%), and amoxicillin/clavulanate (77.5%). In conclusion, both patients’ age and gender are significant factors in determining UTIs etiology; they can increase accuracy in defining the causative uropathogen as well as providing useful guidance to empiric treatment.

Systematic reviews have concluded that hrHPV DNA testing using target-amplification tests is as accurate on vaginal self-samples as on clinician-taken specimens for the detection of cervical precancer. However, insufficient evidence is available for specific HPV assay/self-sample device combinations. The VALHUDES protocol is designed as a diagnostic test accuracy study that aims to compare the clinical sensitivity and specificity of particular hrHPV assay(s) on vaginal self-samples and first-void-urine, collected in agreement with standardized protocols, with hrHPV testing on matched clinician-taken samples. Five hundred enrolled women referred to a colposcopy clinic are invited to collect a first-void urine sample and one or more vaginal self-samples with particular devices before collection of a cervical sample by a clinician. Sample sets are subsequently analysed in a laboratory accredited for HPV testing. Disease verification for all enrolled patients is provided by colposcopy combined with histological assessment of biopsies. A first VALHUDES study has started in Belgium in December 2017 with enrolment from four colposcopy centres. The following assays are foreseen to be evaluated: RealTime High Risk HPV assay (Abbott), cobas-4800 and -6800 (Roche), Onclarity (BD), Xpert HPV (Cepheid) and Anyplex II HPV HR (Seegene). Given empirical evidence that the relative accuracy of HPV-testing on self- vs clinician-samples is robust across clinical settings, the VALHUDES protocol offers a framework for validation of HPV assay/self-sample device combinations that can be translated to a primary screening setting.

Purpose. The purpose of this review was to identify the best solution for rapid and noninvasive diagnosis and long-term monitoring of patients affected by inflammatory gastrointestinal diseases, colon and gastric cancer, obesity in correlation to diet, and breast milk to evaluate exposure to VOCs in women and infants.Methods. This review included 20 previously published eligible studies. VOC analysis has allowed us to highlight differences in lifestyles, intestinal microbiota, and metabolism. New innovative methods have been described that allow the detection and quantification of a broad spectrum of metabolites present in exhaled breath even at very low levels, some of which have been shown to be indicators of pathological conditions.Results. Five studies were analyzed that involved VOC analysis in relation to type of diet. All of them showed that the type of diet can have an impact on metabolites excreted and therefore can be a useful tool in the nutritional studies related to metabolism and health and disease status. Two studies concerned VOC analysis in inflammatory bowel diseases, and the results showed that VOCs can distinguish active disease from remission; VOC profile is clearly different in patients. In particular, C₁₅H₃₀1-pentadecene, 3-methyl-1-butanal, octane, acetic acid, alpha-pinene, and m-cymene are elevated in active ulcerative colitis. Four studies examined VOCs in gastric and colorectal tumors showing a change in metabolic biomarkers of cancer patients compared to the control group. Finally, the study of VOCs in breast milk has improved the understanding of the potential health risks of exposure of children to chemical pollutants.Conclusions. VOC analysis allowed to highlight differences in behavior, lifestyle, and metabolism of individuals. Analytical methods are continuously developed to allow for better detection and quantification of metabolites, thus enabling the detection of a broader spectrum of pathophysiology and disease biomarkers.