G. Mazzarello

Recognition of Antigenic Clusters of Candida albicans by T Lymphocytes from Human Immunodeficiency Virus-Infected Persons

The fine specificity of the cellular immune response to Candida albicans (i.e., recognition of different antigenic components) between normal controls and human immunodeficiency virus-infected patients in various stages of disease was compared. C. albicans-specific T cells, enriched by antigen stimulation and interleukin-2 expansion, were challenged with antigenic fractions of different molecular weight obtained by SDS-gel fractionation of C. albicans extracts in the presence of autologous mononuclear cells as antigen-presenting cells. Proliferative responses showed similar patterns of reactivity between controls and category A and B seropositive subjects. Category C patients with concurrent C. albicans infections did not give rise to C. albicans-specific T cell lines, confirming the T cell defect. Patients without clinically evident C. albicans infection had a low but broad reactivity pattern of C. albicans-specific T cells. These results suggest that depletion of C. albicans-specific T cells, independent of their fine specificity, occurs along with disease progression.

Latent tuberculosis infection and associated risk factors among undergraduate healthcare students in Italy: a cross-sectional study

BackgroundThe screening of both healthcare workers and students attending teaching hospitals for latent tuberculosis infection (LTBI) is recommended in hospitals of many countries with a low-incidence of TB, including Italy, as a fundamental tool of tuberculosis (TB) control programs. The aim of the study was to estimate the prevalence of LTBI and evaluate the main risk-factors associated with this condition in a cohort of healthcare Italian students.MethodsIn a cross-sectional study, performed between January and May 2012, 881 undergraduate students attending the Medical, Nursing, Pediatric Nursing and Midwifery Schools of the University of Genoa, trained at the IRCCS San Martino-IST Teaching Hospital of Genoa, were actively called to undergo the Tuberculin Skin Test (TST). All the TST positive cases were also tested with an Interferon-Gamma Release Assay (IGRA) to confirm the diagnosis of LTBI. A standardized questionnaire was collected for risk-assessment analysis.ResultsSeven hundred and thirty-three (83.2%) subjects underwent TST testing. The prevalence of TST positives was 1.4%, and in 4 (0.5%) out of 10 TST positive cases LTBI diagnosis was confirmed by IGRA. No difference in the prevalence of subjects who tested positive to TST emerged between pre-clinical (n = 138) and clinical (n = 595) students. No statistically significant association between TST positivity and age, gender, and BCG vaccination was observed. The main independent variable associated with TST positivity was to be born in a country with a high TB incidence (i.e., ≥20 cases per 100,000 population) (adjusted OR 102.80, 95% CI 18.09-584.04, p < 0.001).ConclusionsThe prevalence of LTBI among healthcare students resulted very low. The only significant association between TST positivity and potential risk factors was to be born in high TB incidence areas. In countries with a low incidence of TB, the screening programs of healthcare students before clinical training can be useful for the early identification and treatment of the sporadic cases of LTBI.

Role of serum free light chains in predicting HIV‐associated non‐Hodgkin lymphoma and Hodgkin's lymphoma and its correlation with antiretroviral therapy

A nested case‐control study was performed within the Italian cohort of naïve to antiretroviral human immunodeficiency virus (HIV) patients (ICONA) cohort to evaluate the role of serum free light chains (sFLC) in predicting non‐Hodgkins lymphoma (NHL) and Hodgkin lymphoma (HL) in HIV‐infected individuals. Of 6513 participants, 86 patients developed lymphoma and 46 of these (NHL, 30; HL, 16) were included in this analysis having stored prediagnostic blood. A total of 46 serum case samples matched 1:1 to lymphoma‐free serum control samples were assayed for κ and λ sFLC levels and compared by using conditional logistic regression. Because the polyclonal nature of free light chains (FLCs) was the focus of our study, we introduced the k + λ sum as the measurement of choice and as the primary variable studied. κ + λ sFLC values were significantly higher in patient with lymphoma than in controls, especially when considering samples stored 0–2‐year period before the lymphoma diagnosis. In the multivariable analysis, the elevation of sFLC predicted the risk of lymphoma independently of CD4 count, (odd ratio of 16.85 for k + λ sFLC >2‐fold upper normal limit (UNL) vs. normal value). A significant reduction in the risk of lymphoma (odd ratio of 0.07 in model with k + λ sFLC) was found in people with low sFLC and undetectable HIV viremia lasting more than 6 months. Our analysis indicates that an elevated polyclonal sFLC is a strong and sensitive predictor of the risk of developing lymphomas, and it is an easy to measure biomarker that merits consideration for introduction in routine clinical practice in people with HIV. Am. J. Hematol. 2012.

Therapeutical aspects and outcome of HIV/HCV coinfected patients treated with pegylated interferon plus ribavirin in an Italian cohort.

Background:One-third of HIV-infected individuals suffer from chronic hepatitis C virus infection (HCV) in Europe. Recommendations from HCV–HIV International Panel advise current treatment with pegylated interferon plus ribavirin. We assessed the impact of interferon and ribavirin combination in 43 patients between 2002 and 2006.Patients and Methods:All coinfected patients treated for HCV during the 5-year period were included in retrospective data collection. CD4+ T-lymphocyte count, HAART discontinuation, reasons for treatment interruption and factors correlated to sustained virological response (SVR) were monitored.Results:The mean age was 41 ± 6.7 years; the risk factor for coinfection was intravenous drug abuse in 32/43 (74%). The baseline CD4+ T-lymphocytes cell count was > 500 in 51% (22/43). Genotype 3a represented 51% (22/43); 37% were on HAART at baseline (16/43) and half of patients showed high HCV RNA levels (> 800,000 IU/ml). High rates of treatment discontinuation were observed (27/43, 63%), caused by voluntary interruptions in 52% (14/27) and virological failure in 26% (7/27). The overall population had an SVR of 30%; genotypes 3a and 1 had SVR of 38% and 24%, respectively. The SVR was significantly lower in three groups: high HCV RNA viral load (χ2 = 6, p < 0.0025), CD4+ T-lymphocyte historical nadir <350 cells/mm3 (χ2 = 3.26, p < 0.01) and genotype 1 with high viral load (χ2 = 4.8, p < 0.005).Conclusions:Although factors such as HCV viral load rates and genotype 1 have been confirmed to threaten the response to therapy, we observed a significant response rate when patients had a history of CD4+ T-lymphocyte nadir >350 per mm3. The high dropout rates due to voluntary discontinuations complicated the patients’ case management.

Possible Malignant Transformation of Benign Lymphoepithelial Parotid Lesions in Human Immunodeficiency Virus-Infected Patients: Report of Three Cases

Benign lymphoepithelial parotid lesions (BLL) are intraparotid pathological changes that are commonly thought to be an early manifestation of human immunodeficiency virus (HIV) infection. It is not well known whether BLL may undergo malignant transformation into B cell lymphoma and may therefore be a sort of precancerous lesion. We report 3 cases of possible malignant transformation of BLL in HIV-infected patients.

Latent Tuberculosis Infection among a Large Cohort of Medical Students at a Teaching Hospital in Italy

The surveillance of latent tuberculosis infection (LTBI) in both healthcare workers and healthcare students is considered fundamental for tuberculosis (TB) prevention. The aim of the present study was to estimate LTBI prevalence and evaluate potential risk-factors associated with this condition in a large cohort of medical students in Italy. In a cross-sectional study, performed between March and December 2012, 1511 eligible subjects attending the Medical School of the University of Genoa, trained at the IRCCS San Martino-IST Teaching Hospital of Genoa, were actively called to undergo the tuberculin skin test (TST). All the TST positive cases were confirmed with an interferon-gamma release assay (IGRA). A standardized questionnaire was collected for multivariate risk analysis. A total of 1302 (86.2%) students underwent TST testing and completed the questionnaire. Eleven subjects (0.8%) resulted TST positive and LTBI diagnosis was confirmed in 2 (0.1%) cases. Professional exposure to active TB patients (OR 21.7, 95% CI 2.9–160.2; P value 0.003) and previous BCG immunization (OR 28.3, 95% CI 3.0–265.1; P value 0.003) are independently associated with TST positivity. Despite the low prevalence of LTBI among Italian medical students, an occupational risk of TB infection still exists in countries with low circulation of Mycobacterium tuberculosis.

Serum IgG antibodies to human herpesvirus-6 (HHV-6) do not predict the progression of HIV disease to AIDS

Objectives: To evaluate if different levels of human herpesvirus 6 (HHV-6) antibodies can predict HIV disease progression. Design: Longitudinal study of individuals with a documented date of HIV seroconversion. Setting: Clinical centers located throughout Italy. Patients: Individuals who serconverted for HIV between 1983 and 1995 in Italy. Methods: Sera were tested for IgG antibodies to HHV-6 using a commercial enzyme immunoassay. A serum sample with an optical density (OD) ≥ 242 (i.e. the mean value of 10 negative controls+ 4×standard deviation) was considered as HHV-6 positive; the progression of HIV disease was evaluated estimating the relative hazards (RH) of AIDS (by Cox models) for individuals with higher levels vs. lower levels of HHV-6 antibodies or considering levels of antibodies based on 10% increase of the distribution (deciles). Rates of CD4 decline fitting linear regression were also estimated. Results: A total of 381 persons were followed for a median time of 4 years (range: 0.15–9 years) following the date of collection of the serum sample. The median OD value of HHV-6 antibodies was 306, with an interquartile range of 241–440 and a range of 48–2330. A slight inverse correlation was found between HHV-6 antibody levels and age of the individual at the time of serum collection (Spearman rank correlation coefficient, −0.16; p = 0.0013). No association was found between HHV-6 and CD4 level or between HHV-6 and CD8 level at the date of serum collection. The unadjusted RH of progression to AIDS was 0.63 (95% CI: 0.42–0.96) for HHV-6 positive individuals vs. HHV-6 negative; when adjusting for possible confounders (CD4, age, pre-AIDS HIV-related pathologies at the date of sera collection, and previous anti-herpes treatment), the RH of AIDS increased to 0.80 (95% CI: 0.51–1.23). No particular association with HIV disease progression was found when using the deciles of the distribution of HHV-6 antibodies. The median CD4 cell loss was 5.0 × 106 cells/l per month among HHV-6 positive individuals and 5.7 × 106 cells/l per month among the others. Conclusions: The presence of high levels of HHV-6 antibodies does not seem to predict the clinical or immunologic progression of HIV disease.

Quality of life of people living with HIV, preliminary results from IANUA (Investigation on Antiretroviral Therapy) study

The introduction of combined antiretroviral treatment (cART) has reduced HIV‐associated morbidity and mortality, and changed the patients’ perspective of life. As a result, Health Related Quality of Life (HRQOL) has become a crucial clinical issue.

Detection of apoptotic T lymphocytes in peripheral blood of human immunodeficiency virus (HIV)‐infected subjects by Apostain

Apoptosis has been indicated as a mechanism of T cell depletion in HIV-infected subjects and useful in monitoring disease progression. We investigated for the presence of apoptotic T lymphocytes in 130 HIV subjects in various stages of disease by the newly developed cell permeant DNA dye Apostain. Blood was collected in EDTA, lysed in buffered ammonium chloride, fixed in freshly prepared 1% paraformaldehyde and stored in aliquots at -80 degrees C. Samples were thawed and double stained with FITC conjugated-CD3 monoclonal antibody and Apostain. Flow cytometry was then performed and T cells gated on a CD3 versus side scatter dot plot. Normal samples treated in the same manner served to establish the boundary separating non-apoptotic from apoptotic cells. There was no statistically significant association between the proportion of subjects with detectable apoptotic cells and CDC clinical categories A, B and C at the time of admission to the study, although a trend toward a lower apoptotic rate in category A (A= 29%, B=40% and C=41%) was noticed. Conversely, CDC T cell categories 2 and 3 contained significantly higher proportions of Apostain positive patients (1=6%, 2=32% and 3=49%, P=0.072, by chi(2) test). Most importantly, Apostain test identified subjects at risk of disease progression during a 3.5-7 months follow-up in CDC category B and 2 (P=0.008 and P=0.0003, by Fishers exact test, respectively). A similar, albeit not statistically significant trend was observed also in the other categories. Not requiring extensive manipulation of fresh samples nor cumbersome culture techniques, Apostain test appears suitable for identifying HIV subjects at higher risk of disease progression in clinical settings.

Anti-V3 loop spectrotype in HIV-infected individuals during zidovudine therapy.

SummaryIn order to investigate the role played by zidovudine (ZDV) as immune modulator, particularly on B-cell response, the anti-V3 loop spectrotype in 115 sera from 26 HIV-infected individuals was evaluated, prior to and during treatment with ZDV, by isoelectric focusing and reverse blotting (IEF-RB), a technique useful for indirectly measuring the activity and the number of B-cell clones. All 18 patients showing seroreactivity by IEFRB displayed a clear oligoclonal banding pattern, with no change in the spectrotype (i.e. new bands), in sequential analysis over the course of therapy. Only minor changes in band intensity were found, without any correlation with ZDV treatment or CD4+ cell count. In addition, among the sera reactive in spectrotypic analysis, the percentage of those with p24 antigen positivity was significantly lower than those with no detectable p24 antigen (19.8% vs 80.2%, respectively, p=<0.0001, Fishers exact test). In conclusion, it could not be demonstrated by IEFRB that there was any effect of ZDV on the activity and the number of anti-V3 specific B-cell clones. This data is in line with previous studies showing the constancy of anti-gp120 antibody spectrotype over the long course of the disease.ZusammenfassungDer Anti-V3-Loop-Spektrotyp wurden in 115 Seren von 26 HIV-Infizierten vor und nach Zidovudin-Therapie bestimmt, um die Rolle von Zidovudin als Immunmodulator, vor allem der B-Zell-Antwort, zu prüfen. Als Methoden wurden isoelektrische Fokussierung und reverses Blotting (IEFRB) eingesetzt, die sich für die indirekte Bestimmung der Aktivität und der Zahl der B-Zell-Klone als brauchbar erwiesen haben. Bei allen 18 seroreaktiven Patienten stellten sich eindeutige oligoklonale Bandenmuster dar. In Verlaufsanalysen während der Therapie war keine Änderung im Spektrotyp, d. h. keine neuen Banden, zu erkennen. Die Bandenintensität änderte sich nur wenig, wobei keine Beziehung zur Zidovudin-Therapie oder den CD4+ Zellzahlen bestand. In den Spektrotyp-positiven Seren war nur bei einem kleinen Anteil auch p24-Antigen nachzuweisen (19,8% positive gegen 80,2% p-24-Antigen negative Seren; p≤0,0001, Fishers exakter Test). Zusammenfassend fand sich mit IEFRB kein Einfluß von Zidovudin auf die Aktivität und die Zahl anti-V3-spezifischer B-Zell-Klone. Dies stimmt mit den Ergebnissen früherer Studien überein, in denen Konstanz des anti-gp120-Antikörper-Spektrotyps im Langzeitverlauf der Erkrankung festgestellt wurde.In order to investigate the role played by zidovudine (ZDV) as immune modulator, particularly on B-cell response, the anti-V3 loop spectrotype in 115 sera from 26 HIV-infected individuals was evaluated, prior to and during treatment with ZDV, by isoelectric focusing and reverse blotting (IEF-RB), a technique useful for indirectly measuring the activity and the number of B-cell clones. All 18 patients showing seroreactivity by IEFRB displayed a clear oligoclonal banding pattern, with no change in the spectrotype (i.e. new bands), in sequential analysis over the course of therapy. Only minor changes in band intensity were found, without any correlation with ZDV treatment or CD4+ cell count. In addition, among the sera reactive in spectrotypic analysis, the percentage of those with p24 antigen positivity was significantly lower than those with no detectable p24 antigen (19.8% vs 80.2%, respectively, p=<0.0001, Fishers exact test). In conclusion, it could not be demonstrated by IEFRB that there was any effect of ZDV on the activity and the number of anti-V3 specific B-cell clones. This data is in line with previous studies showing the constancy of anti-gp120 antibody spectrotype over the long course of the disease. Der Anti-V3-Loop-Spektrotyp wurden in 115 Seren von 26 HIV-Infizierten vor und nach Zidovudin-Therapie bestimmt, um die Rolle von Zidovudin als Immunmodulator, vor allem der B-Zell-Antwort, zu prüfen. Als Methoden wurden isoelektrische Fokussierung und reverses Blotting (IEFRB) eingesetzt, die sich für die indirekte Bestimmung der Aktivität und der Zahl der B-Zell-Klone als brauchbar erwiesen haben. Bei allen 18 seroreaktiven Patienten stellten sich eindeutige oligoklonale Bandenmuster dar. In Verlaufsanalysen während der Therapie war keine Änderung im Spektrotyp, d. h. keine neuen Banden, zu erkennen. Die Bandenintensität änderte sich nur wenig, wobei keine Beziehung zur Zidovudin-Therapie oder den CD4+ Zellzahlen bestand. In den Spektrotyp-positiven Seren war nur bei einem kleinen Anteil auch p24-Antigen nachzuweisen (19,8% positive gegen 80,2% p-24-Antigen negative Seren; p≤0,0001, Fishers exakter Test). Zusammenfassend fand sich mit IEFRB kein Einfluß von Zidovudin auf die Aktivität und die Zahl anti-V3-spezifischer B-Zell-Klone. Dies stimmt mit den Ergebnissen früherer Studien überein, in denen Konstanz des anti-gp120-Antikörper-Spektrotyps im Langzeitverlauf der Erkrankung festgestellt wurde.