Gut microbiota characterisation in obese patients before and after bariatric surgery.

Intestinal microbiota analysis of obese patients after bariatric surgery showed that Proteobacteria decreased after laparoscopic sleeve gastrectomy (SG), while it increased after laparoscopic gastric bypass (LGB). Comparing to normal weight (NW) patients, obese patients that were selected for SG showed an almost equal amount of Firmicutes and Bacteroidetes and the ratio was not affected by the surgery. Obese patients before LGB showed a predominance of Bacteroidetes, whose amount regained a relative abundance similar to NW patients after surgery. Obese patients before LGB showed the predominance of Bacteroides, which decreased after surgery in favour of Prevotella, a bacterium associated with a healthy diet. The bacteria detected at the highest percentages belonged to biofilm forming species. In conclusion, in this study, we found that the characterization of the gut microbial communities and the modality of mucosal colonisation have a central role as markers for the clinical management of obesity and promote the maintenance of good health and the weight loss.

A Novel Panel of Serum Biomarkers for MPM Diagnosis.

Exposure to asbestos is the main cause of malignant pleural mesothelioma (MPM), a highly aggressive cancer of the pleura. Since the only tools for early detection are based on radiological tests, some authors focused on serum markers (i.e., mesothelin). The aim of this study was the evaluation of new serum biomarkers to be used individually or in combination, in order to improve the outcome of patients whose disease would be diagnosed at an earlier stage. Serum and plasma were available from 43 subjects previously exposed to asbestos and 27 MPM patients, all being epithelioid type. All the new markers found differentially expressed in MPM and healthy subjects, by proteomic and genomic approaches, have been validated in the serum by the use of specific ELISA. The combined approach, using tools of genomics and proteomics, is found to be highly innovative for this type of disease and led to the identification of new serum markers in the diagnosis of MPM. These results, if confirmed in a larger series, may have a strong impact in this area, because early detection of this cancer in people at high risk could significantly improve the course of the disease and the clinical approach to an individualized therapy.

Detection of Malawi polyomavirus sequences in secondary lymphoid tissues from Italian healthy children: a transient site of infection.

BACKGROUND: The novel Malawi polyomavirus (MWPyV) was initially detected in stool specimens from healthy children and children with gastrointestinal symptoms, mostly diarrhea, indicating that MWPyV might play a role in human gastroenteric diseases. Recently, MWPyV sequences were additionally identified in respiratory secretions from both healthy and acutely ill children suggesting that MWPyV may have a tropism for different human tissues. This study was designed to investigate the possible sites of latency/persistence for MWPyV in a cohort of healthy Italian children. METHODS: Specimens (n° 500) of tonsils, adenoids, blood, urines and feces, from 200 healthy and immunocompetent children (age range: 1-15 years) were tested for the amplification of the MWPyV LT antigen sequence by quantitative real-time PCR. Samples (n° 80) of blood and urines from 40 age-matched children with autoimmune diseases, were screened for comparison. Polyomaviruses JC/BK and Epstein-Barr Virus (EBV) were also tested as markers of infection in all samples using the same molecular technique. RESULTS: In our series of healthy children, MWPyV was detected only in the lymphoid tissues showing a prevalence of 6 % in tonsils and 1 % in adenoids, although with a low viral load. No JCPyV or BKPyV co-infection was found in MWPyV positive samples, while EBV showed a similar percentage of both in tonsils and adenoids (38 and 37 %). Conversely, no MWPyV DNA was detected in stool from babies with gastroenteric syndrome. With regards to autoimmune children, neither MWPyV nor BKPyV were detected in blood, while JCPyV viremia was observed in 15 % (6/40) of children treated with Infliximab. Urinary BKPyV shedding was observed in 12.5 % (5/40) while JCPyV in 100 % of the samples. CONCLUSIONS: The detection of MWPyV sequences in tonsils and adenoids of healthy children suggests that secondary lymphoid tissues can harbour MWPyV probably as transient sites of persistence rather than actual sites of latency.

Chemokines involved in the early inflammatory response and in pro-tumoral activity in asbestos-exposed workers from an Italian coastal area with territorial clusters of pleural malignant mesothelioma.

OBJECTIVES: Immune mediators are likely to be relevant for the biological response to asbestos exposure. The aim of this study was to investigate the association between immune mediators involved in inflammation, cell survival and angiogenesis, and asbestos-related diseases in workers from a coastal area of North-East Italy with a high incidence of pleural malignant mesothelioma (PMM). MATERIALS AND METHODS: A selected custom set of 12 soluble mediators was evaluated with a Luminex platform in sera, pleural fluid and mesothelioma biopsies from 123 asbestos-exposed workers (38 free from pleural-pulmonary disorders, 46 with non-malignant asbestos diseases, 39 with PMM) and in sera from 33 healthy controls from the same territorial area. RESULTS: Increased immune mediator concentrations were observed in the sera of the asbestos-exposed workers compared to controls for human fibroblast growth factor (FGF-b), vascular endothelial growth factor (VEGF), CCL5 (RANTES), CXCL10 (IP-10), CLEC11A (SCGF-b), CCL27 (CTACK), CCL11 (EOTAXIN), IL-5 and IL-6 (p<0.001). The chemokines IP-10 and RANTES were associated with the severity of asbestos-related diseases. In the workers with PMM, the immune proteins secreted by mesothelioma biopsies showed detectable levels of RANTES, VEGF, and IP-10. In the same workers with PMM, a significant relationship between serum and pleural fluid concentrations was found for RANTES alone. CONCLUSIONS: Occupational exposure to asbestos seems to drive the production of specific growth factors dually involved in the early inflammatory response and in pro-tumoral activity before clinical evidence of related disorders, suggesting that their over-expression may precede the onset of asbestos-related diseases. These findings suggest that some chemokines may have a prognostic role in the progression of asbestos-related diseases and could be used for the health surveillance of either workers with an occupational history of asbestos exposure or patients affected by non-malignant asbestos-related diseases.

Epidemiological and molecular assessment of a measles outbreak in a highly vaccinated population of northeast Italy.

Two distinct measles outbreaks, unrelated from the epidemiological point of view but caused by genetically related strains, occurred in the Friuli Venezia Giulia region of northeastern Italy. Forty-two cases were reported during the period April-May 2008. In the first outbreak the index case was a teacher who introduced the virus into the Pordenone area, involving eight adolescents and young adults. The other concomitant outbreak occurred in the city of Trieste with 33 cases. The containment of the epidemics can be explained by the high MMR vaccine coverage in an area where the first dose was delivered to 93·4% and the second dose to 88·3% of the target children. Phylogenetic analysis of 14 measles virus strains showed that they belonged to a unique D4 genotype indistinguishable from the MVs/Enfield.GBR/14.07 strain, probably introduced from areas (i.e. Piedmont and Germany) where this genotype was present or had recently caused a large epidemic.

Molecular modeling and inhibition of phospholipase A2 by polyhydroxy phenolic compounds.

Phospholipases A(2) are enzymes responsible for the hydrolysis of membrane phospholipids that release arachidonic acid, which serves as substrate for pro-inflammatory mediators, such as prostaglandins and leucotriens. The design of specific inhibitors for PLA(2) might help in the development of new anti-inflammatory drugs. Polyhydroxy phenolic compounds, such as flavonoids, vitamin E, rosmarinic acid and aristolochic acid, are able to inhibit PLA(2) from different sources. Herein, we have studied the kinetic behavior and the capacity of inhibiting edema formation induced by PLA(2) of five different polyhydroxy phenolic compounds (two phenolic derivatives and three acetophenone hydroxylated derivatives) extracted from the venom of Crotalus adamanteus. The results showed that compounds 1,3-dihydroxy benzene, 1,3,5-trihydroxy benzene and 2,4,6-trihydroxy acetophenone were the most efficient in the inhibition of the enzymatic activity and edema induction by PLA(2). It was also verified that the number of hydroxyls in each molecule is not a limiting factor for the inhibition capacity of these compounds. Molecular modeling studies indicated that the most active compounds are linked to the amino acid Asp 49 and that they destabilize the coordination of the calcium atom, which is essential to the catalytic activity. The study of potential surfaces showed that there are conditions in which the potential values must be adequate for enzyme complex formation with polyhydroxy phenolic compounds. When the potential over the hydroxyl surfaces is very high, formation of stable complexes does not occur and the enzyme does not act intensely. These results might be helpful in the design of a drug that specifically inhibits PLA(2).

Human herpes virus 6 in archival cardiac tissues from children with idiopathic dilated cardiomyopathy or congenital heart disease.

OBJECTIVE: To explore the possible role of human herpes virus 6 (HHV-6) in cardiac disorders in childhood in a retrospective study on archival specimens of explanted hearts. METHODS: 16 children (median age at transplantation 11.0 years) with idiopathic dilated cardiomyopathy (DCM) and 19 children (median age at transplantation 1.0 year) with congenital heart disease (CHD), previously found to be negative for other cardiotropic viruses such as enteroviruses, adenovirus, parvovirus B19, cytomegalovirus and Epstein-Barr virus, were tested for HHV-6 by quantitative real-time PCR and by genotyping. In addition, HHV-7/8 infection was investigated by qualitative PCR. RESULTS: HHV-6 B variant was detected in 11 of 35 samples (31.4%) with a mean viral load of 3.1 x 102 copies/microg of DNA. When assessed by heart disorder, the prevalence was different in the two groups (43.7% in DCM and 21% in CHD) while the mean viral loads were similar. In a logistic multivariate analysis HHV-6 was independently associated with DCM, taking CHD as reference and adjusting for age (best estimate: OR = 6.94; 95% CI 1.00 to 49.85; p = 0.05). CONCLUSIONS: Although the clinical significance of the results is unknown, HHV-6 B genome is frequently detected in explanted hearts from children with DCM and to a lesser extent with CHD, thus adding evidence for HHV-6 cardiac involvement.

The association of chronic hepatitis B and myopathy.

In two patients with chronic hepatitis B and myopathy, muscle biopsy showed necrosis and scarce inflammatory infiltrates. CD8+ cells surrounded some non-necrotic fibers. Hepatitis B virus (HBV) DNA and antigens were found inside intact muscle fibers. Major histocompatibility complex class I antigens were coexpressed with viral antigens. In one patient, symptoms improved during antiviral therapy. HBV can infect muscle fibers and an immune-mediated response to viral antigens may cause muscle injury.

HPV genotyping and HLA II analysis in a pedigree study of pediatric RRP: preliminary results.

OBJECTIVE: This preliminary pedigree study aims at the evaluation of HPV infection and HLA class II alleles as predictive markers in pediatric RRP. METHODS: We investigated for HPV genotyping and HLA class II polymorphisms all the components of family nucleus where we detected a child born to an HPV infected mother and suffering from RRP. RESULTS: HPV 11 was detected both in the laryngeal biopsies of two of the three affected babies and in the cervical smear of their mothers. The third child was positive for HPV 6 while his mother harboured a double HPV 6-16 infection. In one family, the HLA-DQB1*0501 allele exerted its protective role. The HLA-DQB1*0301 allele, commonly associated to a high grade of cervical neoplasia and HPV infection, was present in homozygous in one mother and her child. The same allele was found, though in a heterozygous form, in the third patient too. CONCLUSION: Our report is the first attempt to use the pedigree study for the evaluation of HLA class II alleles and HPV infection related to pediatric RRP. This approach could identify genetic markers that may influence disease predisposition and the severity of HPV infection.

HBV, HCV, and TTV detection by in situ polymerase chain reaction could reveal occult infection in hepatocellular carcinoma: comparison with blood markers.

OBJECTIVE: To report a retrospective analysis on the presence of hepatitis B virus (HBV), hepatitis C virus (HCV), and transfusion transmitted virus (TTV) sequences in formalin fixed, paraffin embedded liver biopsies from eight patients with hepatocellular carcinoma, in comparison with blood markers. METHODS: A direct in situ polymerase chain reaction (PCR) technique was developed for the detection and localisation of genomic signals in the liver tissue. Conventional serological and molecular methods were used for blood evaluation. RESULTS: In situ PCR showed the presence of one of the three viruses (four HCV, two HBV, and one TTV) in seven of the eight patients. In addition, a co-infection with HBV and HCV was detected in one patient. HCV and HBV sequences were located in the cytoplasm and the nucleus, respectively. When compared with blood markers, these findings were compatible with one occult HBV and two occult HCV infections. CONCLUSIONS: These findings provide further evidence for occult HBV and HCV infections in cancerous tissues from patients with hepatocellular carcinomas. In situ PCR could be an additional tool for evaluating the viral aetiology of hepatocellular carcinoma alongside conventional diagnostic procedures.

[Vertical transmission of HGV and outcome of the infected babies]. / Trasmissione verticale di HGV e storia naturale dell'infezione nei neonati.

To assess the risk of HGV mother-to-infant transmission and the clinical outcome of infected babies, we investigated 103 mother-infant couples and followed-up the infected children for 4-72 months. Twenty (19.4%) mothers were HGV-RNA positive and transmission occurred in ten (50%) babies; only one child acquired HGV and HCV infection. Maternal factors, such as history of intravenous drug use, HCV-RNA positivity, HIV coinfection, type of delivery and type of feeding were not related to HGV transmission. One HGV infected baby showed a mild hepatitis when he was also infected by Cytomegalovirus. Two babies cleared HGV within the first year of life. The HGV transmission rate is elevated but HGV infection seems to be benign, at least in a short-term follow-up.

[HHV-6, new perspectives]. / HHV-6: nuovi ruoli per un patogeno antico.

HHV-6 is the etiological agent of Exanthema subitum, and its role in human infection is well known. Recently, molecular diagnostics tools showed for HHV-6 new pathogenetic features and new clinical implication. The present paper highlights recent knowledge on HHV-6 infection and presents a number of results concerning HHV-6 infection in children who had undergone BMT and concerning the roles of endothelial cells as viral reservoir.

Three-year (1999-2002) of epidemiological and virological surveillance of influenza in north-east Italy.

The results of the epidemiological and virological surveillance of influenza performed during the 1999/2000, 2000/2001 and 2001/2002 seasons in the northeastern Italy were presented and the relationship between age-specific morbidity rates and circulating strains were discussed.The epidemiological findings pointed out a change in age distribution. During the 1999/2000 season, characterized by a circulation of viruses antigenically close to the vaccine strain, a similar incidence rate in the 0-14 and 15-64-year-old groups was observed, while during the 2001/ 2002 winter the virus infected mostly children. During 2001/2002 season, B type viruses predominated with at least three distinguishable molecular variants. In particular, B/Victoria/2/87-like viruses re-emerged after more than a decade, and the antibodies elicited by the vaccine strain and by the strains circulating in previous seasons were poor or not protecting. The accumulation of susceptible subjects in young age group during the 1990s, due to the lack of circulation of B/Victoria/2/87-like viruses, was responsible for the unusual morbidity in the 0-14 year group. No circulation of B/Victoria/2/87-like viruses was observed in > 64-year-old group during 2001/2002 epidemic, probably due to a long-lasting immunity against viruses belonging to this lineage.

Seroprevalence of HTLV-I and HTLV-II infection among immigrants in northern Italy.

To assess the prevalence of human T cell leukemia virus type I (HTLV-I) and 2 (HTLV-II) infection and the associated risk factors among immigrants living in Northern Italy, we surveyed 3017 open-population subjects from three geographical areas and 371 prisoners. In the open population, the overall prevalence was 0.3% for HTLV-I and 0.1% for HTLV-II, while among prisoners, HTLV-I and HTLV-II infection were detected in 1.4 and 0.8% of subjects, respectively. HTLV-I prevalence was higher in subjects with multiple sexual partners or sexually transmitted diseases. This association was significant in the open-population group and close to significance in prisoners. Multivariate analysis showed that human immunodeficiency virus (HIV) seropositivity remained significantly associated with HTLV-I infection in both targeted populations (OR: 11.2 in the open population; OR: 9.9 among prisoners), whereas sexual exposure was associated with HTLV-I seropositivity only for prisoners (OR: 14.3). No independent variable was related to HTLV-II infection.

In situ polymerase chain reaction detection of transfusion-transmitted virus in liver biopsy.

The potential role of transfusion-transmitted virus (TTV) infection in determining liver damage is poorly understood and no information exists about TTV replication within hepatocytes. In this study, we assess TTV in situ PCR in liver tissue. Twenty-one patients with different degrees of liver damage were studied by both serum TTV-DNA detection and in situ TTV PCR analysis and extractive PCR in liver biopsy paraffin sections (FFPE). Extractive PCR and in situ PCR detected TTV-DNA both in serum and liver tissue of five patients. The presence of TTV in serum matched with that found in the liver and TTV sequences were never found independently in liver or serum. Four out of five TTV-DNA-positive patients have not other known cause of liver damage while in one a coinfection from HCV was observed. Our data indicate that in situ PCR appears to be a reliable tool for the detection of TTV-DNA in FFPE, and may help detecting unknown origin of liver damage.

Direct in situ PCR allows rapid and sensitive detection of high risk human papillomavirus in cytologic specimens and formalin-fixed paraffin tissues by fluorescent labelling.

We developed a rapid, sensitive and robust high risk human papillomavirus (HR HPV) detection protocol based on direct in situ PCR technology and fluorochrome-modified nucleotides on cytologic specimens (cell smears) and on HPV infected tissues (CIN III). Reproducible results on both cytologic specimens and paraffin-embedded tissues were obtained, providing a powerful tool for clinical investigation on HR HPV infection. Quantitative PCR performed on the same tissue sections adjacent to those used for in situ techniques allowed us to establish the sensitivity of our methods, able to detect rare copies (about 15 in our paraffin-embedded tissues) of HPV.

[Typification of HPV in clinical practice]. / Tipizzazione dell'HPV nella pratica clinica.

OBJECTIVE: To improve the sensitivity of cervical carcinoma screening and to determine the optimal management with an ASCUS Pap result we evaluated the effectiveness of combining thin layer cytologic slides (ThinPrep) and HPV DNA testing. METHODS: A total of 170 women were studied with conventional Pap smears, liquid based cytology, HPV testing and colposcopy with eventual histologic evaluation. RESULTS: The ThinPrep method yielded 12.5% more high grade lesions than did the conventional smears (and more severe diagnoses as compared to the conventional smears). HPV prevalence was significantly associated with disease status. Of 30 patients with ASCUS, HPV testing detected 100% of high grade lesions and 67% of low grade. If colposcopy had been limited to HPV+ women, 47% of case would have been spared. CONCLUSIONS: Liquid based cell collection improves sensitivity for the detection of disease. For women with ASCUS cytology, HPV DNA testing of residual specimen can identify the majority of high risk cases using a single sample.

Concomitant therapy with subcutaneous interleukin-2 and zidovudine plus didanosine in patients with early stage HIV infection.

A phase II study was performed to evaluate the feasibility and activity of subcutaneous (SC) interleukin-2 (IL-2) administration plus zidovudine (ZDV) and didanosine (ddI) in patients with early stage HIV infection. Between October 1995 and October 1996, 12 patients completed 6 cycles of the following scheduled therapy: ZDV plus ddI and SC self-administration of 6 mIU of IL-2 at days 1 to 5 and 8 to 12 of a 28-day cycle for a total of 6 cycles (24 weeks). After 6 cycles, patients received only ZDV plus ddI and they were observed up for an additional 24 weeks. Our schedule was well tolerated as an outpatient regimen and led to a significant elevation in CD4 count, which lasted for 24 weeks after the end of IL-2 therapy. Moreover, CD4/CD25, as well as CD4/CD45RO and CD4/CD45RA, cell levels were significantly increased at the end of the therapy and remained significantly elevated after 24 weeks. During the 6 cycles, HIV-associated viremia was significantly decreased and, accordingly, we observed a significant decline of proviral DNA in peripheral blood mononuclear cells (PBMCs). During follow-up, 10 of 12 treated patients continued to show levels of HIV-related viremia <500 copies/ml. Our results demonstrated that IL-2 and ZDV plus ddI is a well tolerated and effective therapy for patients with HIV in early stages of the disease.