Performance evaluation of gastrointestinal viral ELIte panel multiplex RT-PCR assay for the diagnosis of rotavirus, adenovirus and astrovirus infection.

Rotavirus, adenovirus, norovirus and astrovirus are considered to be among the major causes of sporadic cases and outbreaks of acute gastroenteritis globally. Rapid and accurate identification of enteric viruses is still a challenge for the clinical laboratory. Recently, several molecular platforms for the detection of viral enteric pathogens have become available. In this study, the diagnostic accuracy of InGenius Gastrointestinal Viral (GV) Elite Panel, a newly developed one-step multiplex real-time RT-PCR assay simultaneously detecting rotavirus, adenovirus and astrovirus, was evaluated retrospectively analyzing an archival collection of 128 stool samples of children hospitalized with acute gastroenteritis. The overall sensitivity and specificity for the GV assay was 100% and 96.2% for rotavirus, 96.9% and 100% for astrovirus, 100% and 100% for adenovirus, respectively. The InGenius GV assay showed a high concordance with the reference methods and was able to detect all tested genotypes of rotavirus (including G1, G3, G4, G9 and G12P[8] and G2P[4]), adenovirus and astrovirus (AstV-1 and 2). Studies of considerable sample size are required to determine robust Cycle threshold cut-off values to effectively correlate infection to disease. These preliminary results suggest that InGenius GV assay can be recommended as a valuable method for accurate diagnosis of epidemic and sporadic gastroenteritis.

Waterborne Norovirus outbreak at a seaside resort likely originating from municipal water distribution system failure.

In May 2016 a Norovirus (NoV) gastroenteritis outbreak involved a high school class visiting a seaside resort near Taormina (Mascali, Sicily). Twenty-four students and a teacher were affected and 17 of them showed symptoms on the second day of the journey, while the others got ill within the following 2 days. Symptoms included vomiting, diarrhoea and fever, and 12 students required hospitalisation. Stool samples tested positive for NoV genome by Real-Time polymerase chain reaction assay in all 25 symptomatic subjects. The GII.P2/GII.2 NoV genotype was linked to the outbreak by ORF1/ORF2 sequence analysis. The epidemiological features of the outbreak were consistent with food/waterborne followed by person-to-person and/or vomit transmission. Food consumed at a shared lunch on the first day of the trip was associated to illness and drinking un-bottled tap water was also considered as a risk factor. The analysis of water samples revealed the presence of bacterial indicators of faecal contamination in the water used in the resort as well as in other areas of the municipal water network, linking the NoV gastroenteritis outbreak to tap water pollution from sewage leakage. From a single water sample, an amplicon whose sequence corresponded to the capsid genotype recovered from patients could be obtained.

Performance analysis of two immunochromatographic assays for the diagnosis of rotavirus infection.

Group A rotaviruses (RVAs) are the primary cause of acute gastroenteritis (AGE) in young children worldwide. Several commercial tests including latex agglutination, enzyme-linked assays (ELISA) and immunochromatographic tests (ICT) have been developed for the diagnosis of RVA infection. In the present study, the performance of two commercially available one-step chromatographic immunoassays, CerTest Rotavirus+Adenovirus (Biotec S.L, Zaragoza, Spain) and Vikia Rota-Adeno (bioMerieux SA, Lyon, France) were retrospectively evaluated using Real-time PCR as reference test. Re-testing by Real-time PCR of 2096 stool samples of children hospitalized with AGE previously screened by ICTs (1467 by CerTest and 629 by Vikia) allowed to calculate higher sensitivity for Vikia (94% vs 85% of CerTest) and higher specificity for CerTest (93% vs 89% of Vikia). Accordingly, higher Positive Predictive Values (87% vs 78%) and Positive Likelihood Ratios (12.32 vs 8.8) were found for CerTest and lower Negative Predictive Values (91% vs 97%) and Negative Likelihood Ratios (0.16 vs 0.06) for Vikia. However, both CerTest and Vikia showed a substantial agreement (κ=0.79) with the Real-time PCR. A correlation between false negative results by ICTs and high Cycle Threshold values of Real-time PCR, indicative of low viral load, was observed. False positive results by the two ICT assays were not related to Norovirus, Adenovirus or Astrovirus infections, therefore the risk of cross-reactions was excluded. Both CerTest and VIKIA were able to detect the wide range of RVA genotypes circulating over the study period (including G1P[8], G2P[4], G3, G4, G9 and G12P[8]). The results of the present study showed a satisfactory efficacy of the two diagnostic tests analyzed.

Introduction and prolonged circulation of G12 rotaviruses in Sicily.

Genotype G12 strains are now considered to be the sixth most prevalent human rotaviruses worldwide. In two Sicilian cities, Palermo and Messina, surveillance of rotavirus circulation performed since 1985 and 2009, respectively, did not detect G12 strains until 2012. From 2012 to 2014 rotavirus infection was detected in 29·7% of 1647 stool samples collected from children admitted for acute gastroenteritis to three Sicilian hospitals in Palermo, Messina and Ragusa. In 2012, G12P[8] was first detected in Palermo and then in Messina where it represented the second most frequent genotype (20% prevalence) after G1P[8]. Thereafter, G12 strains continued to circulate in Sicily, showing a marked prevalence in Ragusa (27·8%) in 2013 and in Palermo (21%) and Messina (16·6%) in 2014. All but one of the Sicilian G12 strains carried a P[8] VP4 genotype, whereas the single non-P[8] rotavirus strain was genotyped as G12P[9]. Phylogenetic analysis of the VP7 and VP4 sequences allowed distinction of several genetic lineages and separation of the G12P[8] strains into three cluster combinations. These findings indicate independent introductions of G12 rotavirus strains in Sicily in recent years.

Analysis of early strains of the norovirus pandemic variant GII.4 Sydney 2012 identifies mutations in adaptive sites of the capsid protein.

Global surveillance for norovirus identified in 2012 the emergence of a novel pandemic GII.4 variant, termed Sydney 2012. In Italy, the novel pandemic variant was identified as early as November 2011 but became predominant only in the winter season 2012-2013. Upon sequencing and comparison with strains of global origin, the early Sydney 2012 strains were found to differ from those spreading in 2012-2013 in the capsid (ORF2) putative epitopes B, C and D, segregating into a distinct phylogenetic clade. At least three residues (333, 340 and 393, in epitopes B, C and D, respectively) of the VP1 varied among Sydney 2012 strains of different clades. These findings suggest that the spread of the pandemic variant in Italy during the winter season 2012-2013 was due to the introduction of strains distinct from those circulating at low frequency in the former winter season and that similar strains were also circulating elsewhere worldwide.

Evidence for recombination between pandemic GII.4 norovirus strains New Orleans 2009 and Sydney 2012.

During 2012, a novel pandemic GII.4 norovirus variant, Sydney 2012, emerged worldwide. A signature of the variant was a GII.Pe ORF1, in association with GII.4 Apeldoorn 2008-like ORF2-ORF3 genes. We report the detection of recombinant GII.4 Sydney 2012 strains, possessing the ORF1 gene of the former pandemic variant New Orleans 2009.

Dynamics and molecular evolution of HIV-1 strains in Sicily among antiretroviral naïve patients.

HIV-1 subtype B is the most frequent strain in Sicily. To date, there is no available data about the genetic diversity of HIV-1 viral strains circulating in Sicily among antiretroviral (ARV) naïve subjects and the role of immigration as potential determinant of evolutionary dynamics of HIV-1 molecular epidemiology. For this purpose, HIV-1 polymerase (pol) sequences obtained from 155 ARV naïve individuals from 2004 to 2009 were phylogenetically analysed. The overall rate of HIV-1 non-B infections was 31.0% (n=48/155), increasing from 7.8% in 2004-2006 to 40.9% in 2009, and about one-third were identified as unique recombinant forms. CRF02_AG was the prevalent non-B clade (n=28/48, 58.3%), while subtype C-related strains were responsible for about 30% HIV-1 infections. Non-B viruses strictly associated with heterosexual transmission (85.4%) and were mostly found among immigrants (77.1%). Phylogenetic analysis of non-B sequences found in foreign-born subjects was geographically correlated to the respective country of origin. Moreover, the detection of non-B viral variants in the autochthonous population may support an increasing genetic diversity in Sicily as well as a local circulation of HIV strains also uncommon in our country. In Sicily, HIV-1 epidemic is still mostly attributable to the B subtype. Nevertheless, migration and population movements are progressively introducing novel HIV-1 subtypes causing a continuous increase of HIV-1 molecular dynamic at local level. Molecular surveillance is needed to monitor the genetic evolution of HIV-1 epidemic.

Nationwide surveillance study of human astrovirus infections in an Italian paediatric population.

The study investigated the genetic diversity of human astroviruses (HAstVs) detected in children hospitalized with gastroenteritis in Italy in 2008-2009. A total of 1321 faecal samples were collected in Parma (northern Italy), Bari (southern Italy), and Palermo (Sicily) and screened for the presence of HAstVs. RT-PCR amplification of a portion at the 5'-end of ORF2 allowed the detection of HAstVs in 3·95% of the patients. Four different genotypes (HAstV-1, HAstV-2, HAstV-4, HAstV-5) were found to be circulating during the study period, with HAstV-1 being the predominant type. Interestingly, a novel lineage, proposed as HAstV-2d, was found to have emerged in Parma in 2009. Investigating the genetic variability of HAstVs will be important for understanding the epidemiological trends and evolution of these viruses.

Genetic heterogeneity and recombination in human type 2 astroviruses.

Novel lineages of human astrovirus (HAstV) types 2, 2c, and 2d have been identified. Upon sequencing of the 3' end of the genome, the type 2c and 2d HAstVs were found to be open reading frame 1b (ORF1b)-ORF2 recombinant, with ORF1b being derived from type 3 and type 1 HAstVs, respectively. An ORF2 interlineage recombinant strain, 2c/2b, was also identified.

Surveillance of hospitalised patients with influenza-like illness during pandemic influenza A(H1N1) season in Sicily, April 2009-December 2010.

This paper describes the epidemiology of hospitalised cases with influenza-like illness (ILI) and laboratory-confirmed influenza A cases in Sicily (Italy) during the 2009 influenza pandemic. The first ILI case diagnosed as infected with pandemic influenza A(H1N1)2009 in Sicily was reported in June 2009 and it rapidly became the dominant circulating strain. In the period from 30 April 2009 through 31 December 2010, a total of 2,636 people in Sicily were hospitalised for ILI and 1,193 were laboratory-confirmed for influenza A. Basic demographic and clinical information for all hospitalised patients was collected and population mortality rates (PMRs) and case fatality ratios (CFRs) were calculated. The median age of hospitalised patients infected with pandemic influenza A(H1N1)2009 was significantly lower than that of hospitalised ILI cases in general (18.0 vs. 32.1 years; p<0.0001). Among adults, women were more susceptible than men. The majority of clinical presentations were mild, but 6.6% of hospitalised patients required admission to an intensive care unit, of whom 26.3% had confirmed influenza A. Twenty-four fatal cases were documented. The age group of 45­54 year-olds showed the highest PMRs once hospitalised, while CFRs were higher in elderly patients of 65 years and older. All fatal cases were confirmed as influenza A(H1N1)2009 and most of them had established risk factors for influenza complications.

Chemotherapy-induced cardiotoxicity: role of the conventional echocardiography and the tissue Doppler.

AIM: The cardiotoxicity of anticancer drugs is an emerging problem and only an identification of the early signs of cardiotoxicity by conventional echocardiography and not (tissue Doppler imaging, TDI), will limit and contain the long-term cardiotoxicity effects. The aim of this study was to identify, through conventional echocardiography and TDI, parameters to use as early "signs" of cardiotoxicity. METHODS: A prospective study was performed using patients with breast cancer (72 women, median age 57 ± 12) treated with anticancer drugs (adjuvant chemotherapy). All patients underwent a careful cardiological evaluation before starting treatment (T0) and during follow-up at 3 months (T1), 6 months (T2) and 1 year (T3). Electrocardiography and echocardiography were performed in all patients in these times. Echocardiography evaluation considered the following parameters: systolic and diastolic diameters and volumes, LVEF, MAPSE, TAPSE, E/A TDI (Em, Am, Sm, IVCT, IVRT, ET, TEI index). On the basis of chemotherapy treatment, patients were divided into 5 groups: A=FEC (fluorouracil, epirubicin, cyclophosphamide), B=FEC+trastuzumab, C=trastuzumab, D=FEC+taxotere, E=FEC+taxolo+trastuzumab. RESULTS: A significant reduction in the echo parameters of TDI was observed. TDI appears to offer important advantages over traditional techniques in revealing the presence of early signs of cardiotoxicity. CONCLUSION: The TDI should be utilized to complement conventional echocardiography in the assessment of cardiotoxicity.

Insulin resistance and acute coronary syndrome.

BACKGROUND: Insulin resistance (IR), which can be quantified by HOMA index (fasting glucose X fasting insulin/22.5), is considered the "primum movens" for the development of Metabolic Syndrome. Many authors have suggested that insulin resistance could raise both incidence and mortality of coronary heart disease (CHD). IR is also associated with important predictors of cardiovascular disease, as increased concentration of LDL or triglyceride, decreased concentration of HDL, high systolic blood pressure, visceral obesity. There is accumulating evidence that chronic sub-clinical inflammation, as measured by inflammatory markers as C-reactive protein (CRP) and fibrinogen, is related with insulin resistance. AIM OF THE STUDY: To clarify if insulin resistance would predict cardiovascular disease independently of the other risk factors, such as hypertension, visceral obesity or dyslipidemia, by focusing our attention on the relation between Acute Coronary Syndrome (ACS) and high HOMA index. METHODS: We evaluated glucose and insulin levels at baseline and post-prandial phase, in order to estimate HOMA index in both the conditions; we related the data obtained with the incidence of cardiovascular events, also investigating traditional cardiovascular risk factors. The cohort included 118 patients with a clinical diagnosis of ACS and excluded those with type 1 diabetes, acute inflammatory diseases, hepatic or renal failure, disreactive disorders, autoimmunity and cancer. SUBJECTS: Subjects were followed-up for a period of 1 year, being subdivided in three groups: (1) subjects at elevated HOMA (HOMA > or = 6); (2) subjects at intermediate HOMA (HOMA <6 and > or = 2); (3) subjects at low HOMA (HOMA < or = 2). We considered as end points new cardiovascular events, cerebrovascular events (both TIA and stroke), procedures of revascularization with angioplasty or surgery, cardiovascular death, sudden death. RESULTS: Patients with elevated HOMA have a higher incidence of previous cardio- and cerebrovascular events (p=0.03), myocardial infarction without ST elevation (p=0.005), unstable angina (p=0.01), asymptomatic carotid plaques (p=0.05), depressed systolic function (p=0.05); we found, also, a significant statistic correlation between HOMA index and high levels of CRP, fibrinogen, serum creatinine and TnI. Cardiovascular and cerebrovascular events were registered in 61% of patients with elevated HOMA during the follow up, despite of 25% registered in the control group: so we could consider HOMA index as a negative prognostic variable, also in virtue by the statistic correlation with the inflammatory markers, whose power of prediction is already known. CONCLUSIONS: Beyond traditional cardiovascular risk factors, insulin resistance quantified by HOMA index seems to significantly have an important prognostic role, both in primary and secondary prevention in patients with Acute Coronary Syndrome.

A survey on knowledge and perceptions regarding head lice on a sample of teachers and students in primary schools of north and south of Italy.

High rates of pediculosis are found in every part of the world. The age-range most affected is between 3 and 12 years. No-nit policies are ineffective in preventing infestations. On the other hand, misdiagnosis and overuse of pediculicides, increase resistance to treatment. Lack of information leads people to consider this kind of infestation to be associated to low social classes and immigrants. This research has been implemented to find out about the information level on pediculosis on a sample of students (722) and teachers (408) of some primary schools in northern and southern Italy, and to highlight the role of personal aspects such as age, gender, cultural level, geographical position which may influence this topic and, eventually, allow the use of the correct knowledge in developing appropriate procedures within the school district. Data was obtained through a questionnaire containing 21 multiple choice questions for the teachers and 14 for the students. Standard descriptive statistics were computed. chi2 tests were applied to highlight statistical association among observed variables; test for the difference of two proportions were applied to confirm significant differences among the observed proportions. The level of information for students seems to be, approximately, the same both for northern and southern Italy. There was a slight prevalence of correct answers from southern teachers, probably because the phenomenon of pediculosis has a positive trend of growth in the south. The number of correct answers was, for all, on average about 60.0%; a negative result in itself considering the simplicity of the questions. Knowledge about the biology of the louse was virtually absent. The area of prevention showed lack of information and need for improvement. Most of the teachers believe that there are specific products that can prevent infestation by louse. Most of the teachers have information which does not come from scientific sources. Students receive some short and incomplete information from their parents. Deficiencies in teachers' knowledge indicate that they are inadequately equipped to manage lice infestation. Educational interventions with teachers and families and, as a consequence, with students should be taught at school to allow a correct understanding of the pediculosis, increasing the teachers' competence and, consequently, as soon as the infestation should manifest, a rapid alert of the Health Service so that proper treatment could be provided.

TFPI antigen and activity levels in patients with asymptomatic atherosclerosis and target organ acute and chronic complications.

AIM: In patients with atherosclerosis there is an activation of the tissue factor mediated coagulation cascade. The aim of our study was to ascertain if there is a relationship between increased tissue factor pathway inhibitor (TFPI) antigen and activity plasma levels and atherosclerosis. DESIGN: case-control study. SETTING: Department of Internal Medicine and Cardiovascular Diseases, University of Palermo and Laboratory of Genetics and Molecular Biology, Mayo Clinic, Rochester, MN, USA. PATIENTS: 63 consecutive patients with asymptomatic atherosclerosis or with its acute or chronic complications, and 20 healthy volunteers. MEASUREMENTS: TFPI antigen was detected by an immunoenzimatic assay (Imunobind total TFPI ELISA kit, American Diagnostica Inc., USA). TFPI activity was evaluated by an activity assay (Actichrome TFPI activity assay, American Diagnostica Inc., USA). RESULTS: PATIENTS with chronic (P=0.0001 for TFPI Ag, P=0.006 for TFPI Ac) and acute (P=0.04 for TFPI Ag, P=0.01 for TFPI Ac) vascular disease and with asymptomatic carotid plaque (P=0.0019 for TFPI Ag, P<0.05 for TFPI Ac) had significantly increased TFPI antigen and activity plasma levels vs healthy volunteers. Moreover, patients with chronic vascular disease had higher TFPI Ag levels vs patients with asymptomatic atherosclerosis (P=0.0013). CONCLUSIONS: The originality of our study was the finding of increased TFPI levels, not only in patients with acute complication of atherosclerosis, but also in those with chronic vascular disease and with asymptomatic carotid plaque.

Soluble tumor necrosis factor alpha receptors (sTNF-Rs) in HIV-1-infected intravenous drug users: change in circulating sTNF-R type II level and survival for AIDS patients.

UNLABELLED: This study in intravenous drug users (IVDUs) investigated differences in serum soluble tumor necrosis factor types I and II (sTNFR-I and II) concentrations in HIV-1-infected IVDUs and controls. This study also investigated whether changes of sTNFRs concentration affect the risk of death among patients with AIDS. A cross-sectional study of 54 subjects with AIDS, 47 HIV-seropositive IVDUs, 47 HIV-seronegative IVDUs, and 21 healthy subjects showed that sTNFRs concentration increases from healthy controls to AIDS patients through HIV-seronegative and HIV-seropositive subjects (p < 0.01). sTNFR-I concentration, however, was shown to be similar in HIV-seronegative IVDUs and healthy controls. In the longitudinal study, serum concentration of sTNFRs was determined near AIDS diagnosis in 21 IVDUs and 1 year later (start for the survival study). Cox proportional hazards regression was performed to assess the prognostic value of percent change of sTNFR level alone and in combination with T lymphocyte subsets, HIV-p24 antigenemia and opportunistic infections for death within 240 days. Uni- and multivariate Cox modelling for dichotomised variables according to its median showed an increase of sTNFR-II by at least 30% to be single significant predictor of death: crude relative risk 3.69, p = 0.03; adjusted relative risk 5.67, p = 0.02. Mean survival was 126 days in 11 patients whose sTNFR-II level increased by at least 30%, and 176 days in 10 patients with less change in sTNFR-II (p = 0.02). CONCLUSIONS: sTNFRs concentration is higher in IVDUs than in healthy controls and is highest in AIDS patients. Survival of patients with AIDS is associated with variation in the concentration of sTNFR-II.

Antibodies to human herpes virus type 8 (HHV8) in general population and in individuals at risk for sexually transmitted diseases in Western Sicily.

BACKGROUND: Human herpes virus 8 (HHV8) appears to be the primary aetiologic agent of Kaposi sarcoma (KS). To study the distribution of HHV8, a seroepidemiological study was carried out in western Sicily, where a high incidence rate of classical KS is well documented. METHODS: A total of 970 sera of healthy human immunodeficiency virus (HIV) negative individuals of general population (1-70 years old) and 742 sera of individuals in different risk groups for HIV infection were evaluated by means of an indirect immunofluorescence assay able to detect antibodies to lytic and latent HHV8 antigens. RESULTS: Crude seroprevalence to HHV8 antigens was 11.5% in the general population, and it increased significantly with age from 6% under age 16 to 22% after age 50. Significantly higher HHV8 seroprevalence rates were detected among HIV positive and negative homosexual men (62% and 22%, respectively), men who had sex with prostitutes (40% and 29%, respectively); female prostitutes (42% and 30%, respectively), and clients at a sexually transmitted disease clinic (male: 60% and 33%, respectively, female: 63% and 43%, respectively). In contrast, heterosexual intravenous drug users had seroprevalence rates comparable to those found in the general population. CONCLUSIONS: The results suggest that HHV8 infection is widespread in Western Sicily. The high seroprevalence in individuals with high risk sexual activity point to the role of sexual behaviour in the transmission of the infection in adults, whereas the detection of antibodies in younger population (under 16 years old) is suggestive of a non-sexual route of transmission, probably occurring during childhood by close personal contact.

Serological and virological evidence of non-sexual transmission of human herpesvirus type 8 (HHV8).

To evaluate whether or not human herpesvirus 8 (HHV8) can be transmitted through a non-sexual route a serological survey was carried out in a group of 51 catholic nuns. The seroprevalence rate and the geometrical mean antibody titre to anti-latent HHV8 antigen were similar in nuns and in a group of 60 women, matched by age, in the general population (27 vs. 24%; 1028 vs. 1575, respectively). Moreover, by using nested polymerase chain reaction (PCR), HHV8 DNA sequences were detected in 7 of 16 (43.8%) saliva and peripheral blood mononuclear cells (PBMC) from patients with classical Kaposi's sarcoma (KS) and in 3 out of 7 (42%) AIDS-KS patients. None of 5 HIV positive persons who did not have KS tested positive for HHV8 DNA. HHV8 DNA sequences were also detected in 2 of 12 (17%) saliva and 1 PBMC sample out of 12 healthy HHV8 positive individuals (age range: 30-80 years old). This paper suggests that non-sexual transmission of HHV8 is operating in our geographical setting and saliva may be a potential source of HHV8 spreading in the general population.

Inhibition of human immunodeficiency virus 1 (HIV-1) by variant B of human herpesvirus 6 (HHV-6).

Four HHV-6 strains were initially isolated during attempts to observe HIV-1 replication in cultured primary lymphocytes from 48 patients with AIDS. HHV-6 DNA from each strain was extracted from primary cell cultures and amplified using specific primers in a nested polymerase chain reaction (PCR) assay. All HHV-6 strains were classified as B variants by submitting the PCR products to the digestion of two restriction enzymes (Hind III and Bgl II). Since in primary cultures, the appearance of HHV-6 cytopathic effect was followed by a progressive reduction of HIV-1 replication, we tried to reproduce the observed inhibition in vitro. Two HHV-6 strains, used throughout the experiments, showed their ability to suppress HIV-1 replication when the viruses co-infected CD4+T lymphocyte cultures. While the intrinsic mechanism of this finding still remains unclear, the inhibition of HIV-1 replication was observed only when a high multiplicity of infection (m.o.i.) of HHV-6 and a low m.o.i. of HIV-1 were used in dually infected cell cultures. By using a semiquantitative determination of HIV-1 cDNA by PCR, it appears that the inhibition begins in infected cell cultures and, once established, does not allow any further HIV-1 replication.

Distribution of hepatitis C virus genotypes among intravenous drug users. A ten-year study in Palermo, Sicily.

A cross-sectional study was carried out on the sera of 88 active intravenous drug users (IVDU) collected between 1985 and 1986 to evaluate the circulation of HCV genotypes in Western Sicily. The patients were grouped by age and classified by their HIV status. Genotype 3a (48.8% of all cases) was most frequently detected, followed by genotype 1a (20.4%) and type 1b (17.0%). No significant differences in HCV genotype distribution were observed between HIV positive and negative individuals. Next, the HCV genotype distribution found in sera samples of IDVUs drawn between 1985 and 1986 was studied and divided into three age groups. The genotype distribution in the younger group was then compared with samples collected ten years later, between 1995 and 1996, from young HIV negative IVDU individuals. A different distribution between HCV genotypes 3a and 1a was found with a relative, though not significant, increase in the detection of genotype 1a (38%). Finally, sera from six IVDUs obtained at three different times over a ten-year period were genotyped for HCV. None of the subjects showed any change in the genotype found at the first sampling throughout the ten years. The results suggest that a) genotype 1a and 3a are the most common among IVDUs in Western Sicily, b) concurrent HIV infection does not seem to influence HCV genotype and c) infected IVDUs harbor almost exclusively one genotype.