Filippa Bonura

Human herpesvirus type 8 DNA sequences in biological samples of HIV-positive and negative individuals in Sicily.

Objective:To evaluate the circulation of a new human herpesvirus (HHV), HHV-8 or Kaposis sarcoma (KS)-associated herpesvirus in a geographical area where a high incidence rate of classical KS was already present before the appearance of the AIDS epidemic. Design and methods:The study was carried out by analysing: (i) bioptic samples from classic, AIDS-associated KS, and controls; (ii) peripheral blood mononuclear cells (PBMC) from classic KS, HIV-positive subjects with and without KS and healthy HIV-negative individuals; (iii) semen samples from heterosexual HIV-positive and HIV-negative individuals affected or not by KS; and (iv) cervical swabs from HIV-negative healthy heterosexual females. All specimens were tested for the presence of HHV-8 DNA sequences by a two-step polymerase chain reaction. Results:Positive results were obtained in 90% of bioptic samples of classic KS and in 100% of AIDS-associated KS. Viral sequences were also present in 50% of PBMC of subjects with classic KS and AIDS-associated KS, in 10% of AIDS patients without the angiosarcoma, and in 11% of healthy HIV-negative individuals. Finally, HHV-8 DNA was detected in 13% of semen of HIV-negative heterosexual individuals and in 10% of AIDS patients without KS. Both PBMC and ejaculates from the same individual gave positive results. No HHV-8 sequences were found in cervical swabs. Conclusions:HHV-8 is widespread in the general population in Sicily since it was detected in PBMC and semen of heterosexual HIV-negative individuals and is not found only in high-risk groups. The viral load appears to be more elevated in a high-risk population and it may be ascribed to a viral reactivation. The higher incidence rates of KS in Sicily compared with northern Italy and other European countries might be related to the presence of HHV-8 in the general population.

Transmission of Drug-Resistant HIV Type 1 Strains in HAART-Naive Patients: A 5-Year Retrospective Study in Sicily, Italy

The transmission of drug-resistant HIV-1 strains might compromise the efficacy of current first-line antiretroviral (ARV) regimens. Between 2004 and 2008, HIV-1 reverse transcriptase (RT) and protease (PR) genes of 108 ARV-naive Sicilian patients were amplified and sequenced to describe the prevalence of ARV resistance mutations among HAART-naive HIV-1-infected individuals. The frequency of transmitted drug resistance mutations (DRAMs) was determined by using genotypic interpretation algorithms. The proportion of HAART-naive HIV-1-infected patients in Sicily increased from 18.4% to 23.5% during 2004-2008. Among naive patients, the overall prevalence of DRAMs was 15.7% [17/108; 95% CI: 9.4-24.0]. DRAMs to nonnucleoside reverse transcriptase inhibitors (nNRTI) were detected most frequently [11/108 (10.2%)], of which K103N was the most prevalent (4.6%), whereas the prevalence of DRAMs was lowest for protease inhibitors (PI) [3/108 (2.8%)]. Drug resistance substitutions associated with two or three drug classes were rarely observed. The prevalence of HIV-1 DRAMs in Sicily was relatively higher than that observed in Italy and other European geographic areas and much higher than in resource-limited countries. However, the possible clinical role played by DRAMs in HAART-naive HIV-1-infected individuals will require further assessment.

Detection of HIV type 1 non-B subtypes in Sicily, Italy

To evaluate the presence of HIV-1 non-B subtypes in Sicily, we sequenced and genotyped HIV-1 PR and RT regions of the pol gene using plasma from 169 HIV-1-infected adult patients. All samples were obtained from a study of antiretroviral-associated resistance mutations resulting in virological failure during highly active antiretroviral therapy (HAART). Eight (4.7%) patients had the non-B HIV-1 subtype including some circulating recombinant forms (CRFs). All of these individuals acquired the infection by heterosexual transmission. The detection of HIV-1 non-B strains was significantly associated with younger age of HIV-1 acquisition. Our findings indicate, for the first time, the presence of HIV-1 non-B subtypes in Sicily in patients who experienced virological failure during HAART, and highlight the need for implementing a network for the epidemiological surveillance of HIV-1 subtypes in Southern Europe.

Differences in Kaposi sarcoma-associated herpesvirus-specific and herpesvirus-non-specific immune responses in classic Kaposi sarcoma cases and matched controls in Sicily

Kaposi sarcoma (KS) might develop because of incompetent immune responses, both non‐specifically and specifically against the KS‐associated herpesvirus (KSHV). Peripheral blood mononuclear cells from 15 classic (non‐AIDS) KS cases, 13 KSHV seropositives (without KS) and 15 KSHV‐seronegative controls were tested for interferon‐γ T‐cell (enzyme‐linked immunospot [Elispot]) responses to KSHV‐latency‐associated nuclear antigen (LANA), KSHV‐K8.1 and CMV/Epstein–Barr virus (EBV) peptide pools. The forearm and thigh of each participant was also tested for delayed‐type hypersensitivity (DTH) against common recall antigens. Groups were compared with Fisher exact test and multinomial logistic regression to calculate odds ratios (OR) and 95% confidence intervals (CI). A KSHV Elispot response was detected in 10 (67%) classic KS cases, 11 (85%) KSHV seropositives (without KS) and two (13%) seronegative controls. All four cases with KSHV‐LANA responses had current KS lesions, whereas five of six cases with KSHV‐K8.1 responses had no lesions (P = 0.048). No case responded to both LANA and K8.1. Compared with the seronegative controls, the risk for classic KS was inversely related to DTH in the thigh (OR 0.71, 95% CI 0.55–0.94, P = 0.01), directly associated with DTH in the forearm (OR 1.35, 95% CI 1.02–1.80, P = 0.04) and tended to be increased fivefold per KSHV Elispot response (OR 5.13, 95% CI 0.86–30.77, P = 0.07). Compared with KSHV seropositives (without KS), the risk for classic KS was reduced fivefold (OR 0.20, CI 0.03–0.77, P = 0.04) per KSHV response. The CMV/EBV Elispot responses were irrelevant. Deficiency of both KSHV‐specific and KSHV‐non‐specific immunity is associated with classic KS. This might clarify why Kaposi sarcoma responds to immune reconstitution. (Cancer Sci 2011; 102: 1769–1773)

Serum rantes levels in HIV-positive individuals and in HIV-negative exposed health-care workers

In 1995, soluble factors that suppressed cellular entry of HIV-1 strains were identified as three chemokines: Rantes, macrophage inflammatory protein lc~ (MiplcQ and Mip113 [1]. More recently, it was established that for entry into target cells HIV-1 requires celt-surface CD4 receptor and additional host cell cofactors. A cofactor required by viruses adapted for growth in transformed T-cell lines, classified as T-tropic or syncytia-inducing (SI) strains, was identified and named fusin [2]. However, fusin does not promote entry of macrophage-tropic viruses, classified as M-tropic or non syncytia inducing (NSI) strains, that represent the most prevalent phenotypic isolates from individuals with acute HIV infection. The cofactor which mediates entry of M-tropic strains is CC-CKR-5, a receptor for the [3-chemokines Rantes, Mipl a and Mip1t 3 [3]. During the virological surveillance of HIV-1 infected persons and AIDS patients, we have isolated and phenotypically characterized several NSI and SI strains of HIV-1 according to Tersmette et al. [4]. Therefore, we carried out a study on serum levels of Rantes and Mip1~ chemokines in 7 HIV-l-seronegative individuals (control group), eight HIV-linfected individuals and nine AIDS patients, whose HIV-1 viral isolates showed the phenotypic characteristics of NSI and SI, respectively. Serum levels of chemokines were also tested in 13 HIV-l-infected individuals and in six longterm non-progressors (LTNP) (more than 10 years of HIV-1 seropositivity and CD4 > 500/mm3), whose primary lymphocyte cultures did not reveal any viral isolation. Moreover, chemokines were measured in serum samples taken at 1, 2, 3, and 5 months

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.

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

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.