Proteomics is particularly suitable for characterising human pathogens with high life cycle complexity, such as fungi. Protein content and expression levels may be affected by growth states and life cycle morphs and correlate to species and strain variation. Identification and typing of fungi by conventional methods are often difficult, time-consuming and frequently, for unusual species, inconclusive. Proteomic phenotypes from MALDI-TOF MS were employed as analytical and typing expression profiling of yeast, yeast-like species and strain variants in order to achieve a microbial proteomics population study. Spectra from 303 clinical isolates were generated and processed by standard pattern matching with a MALDI-TOF Biotyper (MT). Identifications (IDs) were compared to a reference biochemical-based system (Vitek-2) and, when discordant, MT IDs were verified with genotyping IDs, obtained by sequencing the 25-28S rRNA hypervariable D2 region. Spectra were converted into virtual gel-like formats, and hierarchical clustering analysis was performed for 274 Candida profiles to investigate species and strain typing correlation. MT provided 257/303 IDs consistent with Vitek-2 ones. However, amongst 26/303 discordant MT IDs, only 5 appeared "true". No MT identification was achieved for 20/303 isolates for incompleteness of database species variants. Candida spectra clustering agreed with identified species and topology of Candida albicans and Candida parapsilosis specific dendrograms. MT IDs show a high analytical performance and profiling heterogeneity which seems to complement or even outclass existing typing tools. This variability reflects the high biological complexity of yeasts and may be properly exploited to provide epidemiological tracing and infection dispersion patterns.

In VLBW neonates, a serum PCT value >2.4 ng/ml prompts early empirical antibiotic therapy, while in normal-birth-weight infants, a PCT value ≤2.4 ng/ml carries a low risk of missing an NS.

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Salmonella enterica is a common cause of gastrointestinal illness in Italy. S. Typhimurium accounts for approximately 40% of isolates, and most of these strains belong to the phage type DT104. We describe the investigation of an outbreak of S. Typhimurium DT104A, a subtype never observed before in Italy, which occurred in Rome during spring 2004. We conducted a matched case control study between 24 July and 9 September 2004. Controls were matched for age and area of residence. Each case had between one and four controls. Odds of exposure to potential risk factors and vehicles for the outbreak were compared between cases and controls. A multivariate analysis was conducted to estimate adjusted Odds Ratios. Sixty-three cases of S. Typhimurium DT 104A infection with onset between 1 April and 5 May 2004 were identified. Sixty-one were residents of Rome and two were residents of a neighbouring region. Twenty-six cases (43%) were enrolled in the study. Their median age was 7.5 years. Fourteen of 26 cases and 16 of 62 controls had eaten pork salami (OR= 25.5; 95% CI 1.6- 416.8). No food samples were available for testing. In northern Italy, two months prior to the outbreak, the veterinary surveillance system identified the first isolation of S. Typhimurium DT104A in a pig isolate. Both human and pig isolates showed indistinguishable PFGE patterns. It was not possible to trace the pig after the sample was taken at slaughter. The epidemiological evidence on the implication of pork salami in this outbreak suggests that pork products can also be a vehicle for salmonella in Italy and underlines the importance of good manufacturing practices for ready-to-eat foods. This investigation highlights the value of laboratory-based surveillance in identifying community-wide outbreaks of uncommon pathogens. It also underlines the need to improve surveillance timeliness, for promptly detecting outbreaks, undergoing field investigation, and implementing control measures. Moreover, our study shows the usefulness of integrated human and animal surveillance in tracing the possible source of infection.

Background: Holder pasteurization (HoP) is the recommended method of pasteurization for donor human milk (DHM). The aim of the present study was to compare nutritional and microbiological impact on DHM of a new technique of pasteurization based on technical changes of HoP. Methods: We analyzed milk samples from 25 donors. Each sample, derived from one breast milk expression, was subdivided into three aliquots according to pasteurization: The first was not pasteurized, the second pasteurized by HoP, and the third was pasteurized by modified HoP (MHoP). Each aliquot was assessed as to its microbiological and nutritional profile. Nutritional profile included calcium and triglycerides concentrations detected by spectrophotometry and amino acid levels assessed by high-performance liquid chromatography (HPLC). Results: Triglycerides were significantly lower in pasteurized, by both methods, than in not pasteurized aliquots, while calcium and amino acids concentration were similar. Microbiological profile did not differ between HoP and MHoP aliquots. Conclusions: HoP and MHoP seem to have similar efficacy in preserving some nutritional characteristics of DHM and to confer similar microbiological safety. MHoP is time-saving and potentially costs-effective when compared to HoP, and it is; therefore, potentially of more interest from a practical point of view. Further studies are needed to confirm these findings.

Resistance to antibiotics frequently used to treat Helicobacter pylori infection is one of the most important factors in treatment failure. The aims of this study were to determine the accuracy of culture and other invasive and non-invasive tests for diagnosis of H. pylori infection in children, using histological examination as the gold standard, and to evaluate rates of resistance to metronidazole, clarithromycin and amoxicillin. Between March 2003 and December 2004, gastric biopsies from 215 children were tested by histological examination, a rapid urease test and culture. In addition, stool samples were obtained from all children for H. pylori antigen testing. Sixty-four (30%) patients were positive for H. pylori by histological examination. The H. pylori fecal antigen test revealed a sensitivity of 57.8%, a specificity of 93.4%, a positive predictive value of 78.7% and a negative predictive value of 83.9%; rapid urease test had a sensitivity of 70.3%, a specificity of 94.7%, and positive and negative predictive values of 83.3% and 88.2%; culture had a sensitivity of 90.6%, a specificity of 100%, and positive and negative predictive values of 100% and 96.2%. All positive cultures were tested for susceptibility to amoxicillin, metronidazole and clarithromycin by the E-test method. Resistance rates to clarithromycin and metronidazole were 20.7% and 27.6% respectively; no resistance to amoxicillin was observed. This study confirms the place of histological examination as the definitive test for H. pylori, and shows the importance of H. pylori culture in allowing evaluation of antibiotic resistance profiles in pediatric patients.

SummaryBackground and aims: Blood culture (BC) results are essential to guide antimicrobial chemotherapy for patients with sepsis. However, BC is a time-consuming exam, which can take several days. Reducing BCs turn around time (TAT) could impact on multiple outcome parameters and TAT monitoring is an important tool for measurement of microbiology laboratory performance. The aim of this study was to provide an overview of BC TATs among Italian microbiology laboratories.Materials and methods: Five laboratories collected and recorded, for a month period, date and time of the BC processing events. Cumulative TATs were analysed using the GraphPad software.Results: Participating laboratories reported data from 302 sepsis episodes. The median time from when the BC system produced a positive signal until Gram-stain results were reported was 7.6 hours. A rapid molecular identification and antimicrobial susceptibility testing (AST) was performed in 26.5% of BCs. Mean TAT for identification report was significantly lower when a molecular approach was adopted (12 vs. 28.7 hours, P<0.001). Similarly, results of the molecular AST were obtained more than 24 hours in advance compared with phenotypic AST (mean 13.2 vs. 47.6, P<0.001). TATs from BC positivity of laboratories opened 7 days/week were not significantly lower than those of laboratories opened 6 days/week.Conclusions: BC is a time-consuming exam, however, molecular identification and AST methods can drastically reduce time to results. The lack of difference between TATs observed for laboratories working 7 days/week and 6 days/week, coupled with a high rate of BCs turning positive during the night enable to conclude that the most urgent measure to reduce TATs is the expansion of laboratory regular duty hours. IntroductionSepsis is a severe disease associated with high morbidity and mortality (7). Although conflicting data exist, early administration of appropriate empirical antimicrobial therapy, coupled with supportive treatment, have been shown to have an important impact on outcome (6,9,10,16