Correlates of independent HIV-1 replication in the CNS and of its control by antiretrovirals.

OBJECTIVE: To investigate in detail factors associated with independent replication of HIV-1 in CNS, and to predict its therapeutic control. METHODS: HIV RNA concentration was measured by PCR in 134 cross-sectional paired plasma and CSF samples from 95 patients infected with HIV-1 with various conditions, and in longitudinal CSF samples from 50 patients on antiretroviral treatment. Monocyte chemotactic protein (MCP)-1 was quantified in CSF by ELISA. RESULTS: High HIV RNA levels either in plasma or in CSF did not correlate with HIV RNA concentration in the paired biologic sample. A high CSF-to-plasma HIV RNA ratio, suggesting independent viral replication in the CNS, was associated with higher CSF viral load and higher CSF MCP-1 levels. Higher MCP-1 levels in the CSF were also associated with neurologic disorders and were not influenced by the use of highly active antiretroviral therapy (HAART). A higher number of antiretroviral drugs with CSF penetration correlated with a more profound CSF HIV-1 load reduction, independently from the use of HAART alone. Virologic suppression in CSF was predicted by a higher number of CSF-penetrating antiretrovirals and by the baseline CSF viral load, whereas lower baseline CD4 counts and higher MCP-1 levels were associated with increased risk of virologic failure. CONCLUSIONS: Quantification of HIV RNA in CSF is clinically useful, particularly in patients with neurologic disorders. CSF penetration of antiretrovirals must be considered when choosing treatments, mainly in patients with higher CSF viral loads, advanced disease, and CNS disorders associated with significant macrophage activation.

Surveillance of anti-tuberculosis drug resistance: results of the 1998/1999 proficiency testing in Italy. SMIRA (Italian Multicentre Study on Antituberculosis Drug Resistance) Study Group.

OBJECTIVE: To determine the accuracy of drug-susceptibility testing (DST) for isoniazid, rifampicin, ethambutol and streptomycin in a provisional network of 22 regional laboratories in Italy. METHODS: Methods, definitions and reference Mycobacterium tuberculosis strains were derived from the WHO/IUATLD Global Project on Anti-tuberculosis Drug Resistance Surveillance. The laboratories were selected based on technical skills required by the project, the number of DST performed annually and geographic localisation. The results (sensitive/resistant strain) were compared with the gold standard (global project results). Sensitivity (ability to detect true resistance), specificity (ability to detect true susceptibility), positive predictive values for resistance and susceptibility, efficiency and reproducibility were calculated in two rounds. RESULTS: Eighteen of 22 laboratories completed the first round of proficiency testing for the four drugs. Sensitivity was 76.6%, specificity 97.2%, predictive value of a resistant test 89.8% and of a susceptible test 86.8%, efficiency 87.8% and reproducibility 92.8%. A second round was performed by all those laboratories that did not achieve > or = 90% agreement with the results of the Global Project. Overall, after the second round, all the parameters except specificity improved, exceeding 90%. CONCLUSIONS: A network of 15 regional laboratories that fulfil the quality criteria for determining the susceptibility of M. tuberculosis to the four primary antituberculosis drugs was established in Italy.

Brief report: disseminated mycobacteriosis caused by drug-resistant Mycobacterium triplex in a human immunodeficiency virus-infected patient during highly active antiretroviral therapy.

Mycobacterium triplex is a novel species that, until now, has been isolated only from limited clinical samples, and its clinical relevance has been largely unknown. In this report, we describe the first case of disseminated disease caused by M. triplex in a human immunodeficiency virus-infected patient.

Discontinuation of primary prophylaxis for Pneumocystis carinii pneumonia and toxoplasmic encephalitis in human immunodeficiency virus type I-infected patients: the changes in opportunistic prophylaxis study.

A multicenter open, randomized, controlled trial was conducted to determine whether primary prophylaxis for Pneumocystis carinii pneumonia and toxoplasmic encephalitis can be discontinued in patients infected with human immunodeficiency virus type 1 (HIV-1) whose CD4+ T cell counts have increased to >200 cells/mm3 (and who have remained at this level for at least 3 months) as a result of highly active antiretroviral therapy (HAART). Patients were randomized to either the discontinuation arm (i.e., those who discontinued prophylaxis; n=355) or to the continuation arm (n=353); the 2 arms of the study were similar in terms of demographic, clinical, and immunovirologic characteristics. During the median follow-ups of 6.4 months (discontinuation arm) and 6.1 months (continuation arm) and with a total of 419 patient-years, no patient developed P. carinii pneumonia or toxoplasmic encephalitis. The results of this study strongly indicate that primary prophylaxis for P. carinii pneumonia and toxoplasmic encephalitis can be safely discontinued in patients whose CD4+ T cell counts increase to >200 cells/mm3 during HAART.

Expression of the novel T cell activation molecule hpH4 in HIV-infected patients: correlation with disease status.

We have described hpH4, a surface glycoprotein selectively expressed by activated T cells and mature thymocytes and displaying weak lateral association with CD4. The hpH4 expression pattern and biochemical features, together with analysis of its tryptic digest by peptide mass searching using MALDI-MS, suggested that it is a novel molecule. The aim of this work was to evaluate the peripheral blood T cell expression of hpH4 in HIV-infected patients and the interplay between HIV gp120 and hpH4, since both molecules interact with CD4. hpH4 expression during HIV-1 infection was evaluated by assessing 55 patients at various disease stages and following up 3 patients with primary infection and 3 patients with AIDS. hpH4 expression displayed a peak in the early phase of primary infection, dropped to control levels in the asymptomatic phase, and was newly expressed, at low levels, as AIDS developed. The expression kinetics were different than those shown by HLA-DR, CD25, and CD38. The most striking findings were the transient hpH4 expression peak displayed in the earliest stage, which was unique for hpH4. Incubation of T cells from normal donors with HIV gp120 induced transient hpH4 expression in resting CD4+ T cells and potentiated the hpH4 lateral association with CD4 in activated T cells. Moreover, hpH4 triggering inhibited gp120-induced death of CD4+ cells. Therefore, H4 expression may be a response to avoid apoptosis induced by HIV products.

Humoral immune responses to multiple antigens of Mycobacterium tuberculosis in tuberculosis patients co-infected with the human immunodeficiency virus.

A panel of ten protein antigens of Mycobacterium tuberculosis was used to evaluate serum antibody responses to tuberculosis in patients co-infected with the human immunodeficiency virus (HIV) and in HIV-infected control individuals without tuberculosis. Most (70%) of the tuberculosis patients had serum reactivity to at least one antigen and maintained the diverse antibody repertoire previously observed in HIV-negative tuberculosis patients.

Human herpesvirus 8 seropositivity and risk of Kaposi's sarcoma and other acquired immunodeficiency syndrome-related diseases.

BACKGROUND: The incidence of Kaposi's sarcoma (KS) is increased severalfold in individuals infected with human immunodeficiency virus-1 (HIV). Human herpesvirus 8 (HHV8) has also been implicated in KS. We investigated several factors that may determine the onset of KS, particularly HHV8 infection in individuals after becoming seropositive for HIV. METHODS: We studied 366 individuals belonging to different HIV-exposure categories (i.e., homosexual activity, intravenous drug use, and heterosexual contact) for whom a negative HIV serologic test and then a positive HIV serologic test were available within a 2-year period. HHV8 antibody testing was performed by use of an immunofluorescence assay on the first serum sample available after the first positive HIV test. Actuarial rates of progression of KS and of other acquired immunodeficiency syndrome (AIDS)-defining diseases were estimated by use of time-to-event statistical methods. All statistical tests were two-sided. RESULTS: Twenty-one of the 366 study participants developed AIDS-related KS, and 83 developed AIDS without KS. One hundred forty (38.3%) participants had detectable anti-HHV8 antibodies. The actuarial progression rate to KS among persons co-infected with HIV/HHV8 was nearly 30% by 10 years after HIV seroconversion. Increasing HHV8 antibody titers increased the risk of developing KS (for seronegative versus highest titer [1:125 serum dilution], adjusted relative hazard [RH] = 51.82; 95% confidence interval [CI] = 6.08-441.33) but not of other AIDS-defining diseases (adjusted RH = 1.14; 95% CI = 0.72-1.80). HHV8-seropositive homosexual men compared with HHV8-seropositive participants from other HIV-exposure categories showed an increased risk of KS that approached statistical significance (adjusted RH = 6.93; 95% CI = 0.88-54.84). CONCLUSIONS: Approximately one third of individuals co-infected with HIV/HHV8 developed KS within 10 years after HIV seroconversion. Progression to KS increased with time after HIV seroconversion. Higher antibody titers to HHV8 appear to be related to faster progression to KS but not to other AIDS-defining diseases.

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

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) > or =242 (i.e. the mean value of 10 negative controls +4x 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 5.0x10(6) cells/l per month among HHV-6 positive individuals and 5.7x10(6) 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.

HHV-8/KSHV is not associated with AIDS-related primary central nervous system lymphoma.

Primary central nervous system lymphoma (PCNSL) is a major complication of the late stages of human immunodeficiency virus (HIV) disease. Epstein Barr virus (EBV) infection is the only genetic lesion consistently associated with this neoplasia. Recently, it has been proposed that the pathogenesis of AIDS-related PCNSL (AIDS-PCNSL) may be associated with infection by human herpesvirus-8/Kaposi's sarcoma associated herpesvirus (HHV-8/KSHV), although at present such association remains controversial. In order to conclusively assess the link between HHV-8/KSHV infection and AIDS-PCNSL, we performed a comprehensive study based on multiple molecular assays on cerebral tissues and cerebrospinal fluid (CSF) as well as specific immunologic assays on patients' sera. A well characterized panel of 33 Italian patients with AIDS-PCNSL and 13 controls with other HIV-related brain focal diseases from the same geographical area was analyzed. No signs of HHV-8/KSHV infection were detected in cerebral tissues by single-step PCR. Cerebral tissues of all AIDS-PCNSL scored negative for HHV-8/KSHV DNA sequences also by nested PCR, with the exception of one single patient who was simultaneously affected by Kaposi's sarcoma. All CSF samples analyzed were consistently devoid of HHV-8/KSHV sequences by molecular assays. By serologic assays, detecting both latent and lytic HHV-8/KSHV antigens, a specific immunoreactivity was observed in 16/33 (48%) AIDS-PCNSL and in 6/13 (46%) controls (P = 0.88). A significant correlation with HHV-8/KSHV serum reactivity was seen with a homosexual route of HIV transmission (P = 0.018), but not with the presence of AIDS-PCNSL. The results of our analysis conclusively assess that HHV-8/KSHV infection is not a feature of AIDS-PCNSL.

Value of combined approach with thallium-201 single-photon emission computed tomography and Epstein-Barr virus DNA polymerase chain reaction in CSF for the diagnosis of AIDS-related primary CNS lymphoma.

PURPOSE: To determine the diagnostic capability of thallium-201 (201Tl) single-photon emission computed tomography (SPECT) combined with Epstein-Barr virus DNA (EBV-DNA) in CSF for the diagnosis of AIDS-related primary CNS lymphoma (PCNSL). PATIENTS AND METHODS: All human immunodeficiency virus (HIV)-infected patients with focal brain lesions observed between June 1996 and March 1998 underwent lumbar puncture and 201Tl SPECT. Each CSF sample was tested with polymerase chain reaction (PCR) for EBV-DNA. RESULTS: Thirty-one patients were included, 13 with PCNSL and 18 with nontumor disorders. In 11 PCNSL patients, EBV-DNA was positive. Thallium-201 uptake ranged from 1.90 to 4.07 in PCNSL cases (mean, 2.77; 95% confidence interval [CI], 2.35 to 3.19) and from 0.91 to 3.38 in nontumor patients (mean, 1.62; 95% CI, 1.30 to 1.94) (P<.0002). Using a lesion/background ratio of 1.95 as cutoff, a negative SPECT was found in one PCNSL case and 16 nonneoplastic cases. A cryptococcoma and a tuberculoma showed highly increased 201Tl uptake. Epstein-Barr virus DNA was never detected in nonneoplastic patients. For PCNSL diagnosis, hyperactive lesions showed 92% sensitivity and 94% negative predictive value (NPV), whereas positive EBV-DNA had 100% specificity and 100% positive predictive value. The presence of increased uptake and/or positive EBV-DNA had 100% sensitivity and 100% NPV. CONCLUSION: Combined SPECT and EBV-DNA showed a very high diagnostic accuracy for AIDS-related PCNSL. Because PCNSL likelihood is extremely high in patients with hyperactive lesions and positive EBV-DNA, brain biopsy could be avoided, and patients could promptly undergo radiotherapy or multimodal therapy. On the contrary, in patients showing hypoactive lesions with negative EBV-DNA, empiric anti-Toxoplasma therapy is indicated. In patients with discordant SPECT/PCR results, brain biopsy seems to be advisable.

Application of molecular methods for detection and transmission analysis of mycobacterium tuberculosis drug resistance in patients attending a reference hospital in Italy.

A molecular analysis of drug-resistant isolates of Mycobacterium tuberculosis was done in a population with a high prevalence of human immunodeficiency virus infection. Seventy-one consecutive isolates were tested for genotypic resistance to isoniazid, rifampicin, streptomycin, and ethambutol by polymerase chain reaction-single strand conformation polymorphism analysis and automated sequencing of target regions. Phenotypic and genotypic resistance to isoniazid, rifampicin, streptomycin, and ethambutol were detected in 23.4%, 11.2%, 7%, and 5.6% of isolates and in 87%, 88%, 40%, and 100% of resistant isolates, respectively. Specificity was 100% for all target regions. When rpoB, katG, and ahpC mutation analysis were combined, 86% of resistant isolates to any drug were identified. No mutations in inhA were found in isoniazid-resistant isolates. Molecular detection of drug resistance, particularly for isoniazid and rifampicin, may represent a sensitive and very specific technique. The strategy of selecting rpoB, katG, and ahpC to quickly identify most resistant isolates, with a relevant saving of resources, is warranted.

Epstein-Barr virus in monitoring the response to therapy of acquired immunodeficiency syndrome-related primary central nervous system lymphoma.

To evaluate the value of Epstein-Barr virus DNA (EBV-DNA) assay in cerebrospinal fluid (CSF) for monitoring the response to treatment in acquired immunodeficiency syndrome-related primary central nervous system lymphoma (AIDS-PCNSL), 9 human immunodeficiency virus-infected patients with biopsy-proven AIDS-PCNSL who underwent multimodal therapy were investigated for EBV-DNA detection in CSF by semiquantitative nested polymerase chain reaction (PCR). Tumoral tissue expression of bcl-6 oncogene protein and of EBV-encoded latent membrane protein (LMP-1) was also investigated. The 2 patients who had a response to chemotherapy showed a reduction of mean EBV-DNA concentration values after chemotherapy and displayed a large noncleaved morphology and a BCL-6+/LMP-1- phenotype. Conversely, the 4 patients with progressive disease after chemotherapy showed increasing mean values of EBV-DNA and displayed an immunoblastic morphology and a BCL-6-/LMP-1+ phenotype. No significant changes were observed for patients with stable disease. EBV-DNA burden reduction was significantly associated with prolonged survival. These results suggest that EBV-DNA monitoring might be helpful in predicting response to chemotherapy and in segregating distinct biological and prognostic categories of AIDS-PCNSL.

A randomized, double-blind trial on the use of a triple combination including nevirapine, a nonnucleoside reverse transcriptase HIV inhibitor, in antiretroviral-naive patients with advanced disease.

The immunologic and virologic activity of nevirapine in combination with two nucleosides (zidovudine [ZDV] and didanosine [ddI]) was evaluated in antiretroviral-naive patients with a CD4 count <200/mm3 or clinical AIDS. In all, 68 patients were enrolled in a 48-week double-blind, placebo-controlled trial. A group of 32 patients received ZDV + ddI + nevirapine, and 36 patients received ZDV + ddI. Primary efficacy parameters were the activity on HIV-1 RNA and on peripheral blood CD4+ cells, with differences between groups analyzed by the Wilcoxon's nonparametric two-sample test. Baseline RNA was high in both treatment groups (median values, 5.8 and 5.7 log10). RNA and CD4 responses were significantly higher with the triple combination (median RNA reductions, 2.69 versus 1.05 log10 at 24 weeks and 1.97 versus 1.20 log10 at 48 weeks; median CD4 increases, 81 versus 64 cells/mm3 at 24 weeks and 101 versus 27 cells/mm3 at 48 weeks). This study demonstrates that a triple combination of ZDV + ddI + nevirapine used as first-line regimen in antiretroviral-naive patients can induce sustained virologic and immunologic response in patients with low CD4 count or a previous diagnosis of AIDS.

Nosocomial bloodstream infections in HIV-infected patients: attributable mortality and extension of hospital stay.

A 3-year prospective matched case-control study was performed to investigate the potential risk factors, prognostic indicators, extension of hospital stay, and attributable mortality of nosocomial bloodstream infections in HIV-infected patients. Matching variables were: age, gender, number of circulating CD4+ T lymphocytes, cause of hospital admission, hospitalization in the same ward within the 6 weeks of diagnosis of the case, and length of stay before the day of infection in the case. Eighty-four cases and 168 matched controls were studied. Nosocomial bloodstream infections complicated about 3 of 1000 hospital days per patient in the study period. With step-wise logistic regression analysis, the most important predictors for developing nosocomial bloodstream infections were: increasing value of Acute Physiology and Chronic Health Evaluation (APACHE II) score (p = .001) and use of central venous catheter (CVC) (p = .002). The excess of hospital stay attributable to nosocomial bloodstream infections was 17 days. The crude mortality rate was 43%. The attributable mortality rate was estimated to be 27% (95% confidence interval [CI] = 13%-48%). The estimated risk ratio for death was 3.91 (95% CI = 2.06-7.44). Multivariate analysis identified two prognostic indicators that were significantly associated with unfavorable outcome of bloodstream infections: number of circulating CD4+ T cells <100/mm3 (p = .002) and APACHE II score >15 (p = .01). Nosocomial bloodstream infections are more common in patients with advanced HIV disease. Important cofactors are high APACHE II score and use of CVC. These infections can cause an excess mortality and significantly prolong the hospital stay of HIV-infected patients.

Changing disease patterns in focal brain lesion-causing disorders in AIDS.

OBJECTIVES: To assess temporal trends of the different disorders causing focal brain lesions (FBL) in HIV-infected patients and to examine the reliability of the U.S. Centers for Disease Control and Prevention (CDC) criteria for presumptive diagnosis of toxoplasmic encephalitis (TE) for the years 1991 to 1996. DESIGN/METHODS: A prospective, monocenter study. Percentages of occurrence of the different FBL-causing disorders for each year were calculated. Temporal trends were analyzed by chi2 test for linear trend and multivariate polytomous nonordinal logistic regression. The positive predictive value (PPV) of the CDC's presumptive criteria for the diagnosis of TE (recent onset of a focal neurologic abnormality consistent in intracranial disease or a reduced level of consciousness, evidence on brain imaging of a lesion having mass effect or the radiographic appearance of which is enhanced by injection of contrast medium, and serum antibody to toxoplasmosis) was calculated using contingency tables for each calendar year. RESULTS: A highly significant decline of the risk of TE and an increase of the probability of patients to take anti-Toxoplasma prophylaxis were observed. A threefold but statistically not significant augmented risk of diagnosing both primary central nervous system lymphoma (PCNSL) and progressive multifocal leucoencephalopathy (PML) has been registered for 1996 compared with 1991. Among FBL showing contrast enhancement, the increased finding of PCNSL over the years studied was significant. The probability of other FBL-causing disorders, such as focal viral encephalitis sustained by cytomegalovirus or herpes simplex virus, increased significantly over the years studied. Multivariate analysis confirmed that the year of diagnosis of FBL had a significant effect on the risk reduction of TE. The PPV of the CDC's criteria for the presumptive diagnosis of TE dropped from 100% for the year 1991 to 39% in the year 1996. A similar result was obtained in calculating the PPV of presumptive criteria only among patients without previous primary prophylaxis. CONCLUSIONS: Because of the significant decrease of TE and the increase of PCNSL empiric anti-Toxoplasma therapy no longer seems appropriate as a first-line approach to all HIV-positive patients with FBL. Especially in the case of a finding of FBL by contrast enhancement, new diagnostic strategies should be employed to identify the underlying disorder rapidly and accurately.

Risk factors, nutritional status, and quality of life in HIV-infected patients with enteric salmonellosis.

AIMS: To define the potential risk factors, prognostic indicators, and quality of life of HIV-infected patients with enteric salmonellosis. METHODS: A five-year matched (1:2) case-control study was performed. Thirty cases and 60 matched controls were studied. RESULTS: Univariate analysis (p < 0.05) identified six risk factors for enteric salmonellosis: 1) increasing value of APACHE II score, 2) altered nutritional status, 3) previous antibiotic therapy, 4) ingestion of "risk" foods, 5) multiple (> or = 2) previous opportunistic infections, 6) stage C of HIV infection. Using the multivariate analysis, the most powerful predictors for developing an enteric salmonellosis were: increasing value of APACHE II score and altered nutritional status. The response to therapy was favourable in all episodes. Five (17%) patients suffered from one or more relapses. Multivariate analysis identified that a low number of circulating CD4+ (< 100/mm3) and a high APACHE II score (> 15) predict an increased risk of relapses. CONCLUSIONS: HIV-associated enteric salmonellosis occurs more frequently in patients with advanced stage of HIV infection and with impaired nutritional status. We stress the need for a prompt and accurate diagnosis and treatment since the possible haematogenous spread and relapses appear reduced under early and prolonged antibiotic therapy.

General epidemiology of tuberculosis.

An outline of the history of tuberculosis is offered for consideration including the present estimated incidence world-wide. The epidemiology of the disease with respect to its etiology is described focusing on the risk of infection, the development of clinical manifestations subsequent to the exposure to the bacillus, and the risk of reactivation. The situation of the epidemiology of tuberculosis in Italy is analyzed based on available information after the closure of TB dispensaries following the introduction of National Health Service in 1978. In last years over 5000 cases of tuberculosis per year have been notified; however what percentage of the real incidence of TB this data represents, is unknown.