Minimally invasive diagnosis of acquired immunodeficiency syndrome-related primary central nervous system lymphoma.

BACKGROUND: The detection of Epstein-Barr virus (EBV)-DNA in cerebrospinal fluid (CSF) by means of the polymerase chain reaction (PCR) has been revealed, in retrospective studies, to be a good marker of primary central nervous system lymphoma (PCNSL) related to acquired immunodeficiency syndrome (AIDS); however, the technique's usefulness in the management of AIDS patients with focal brain lesions is still unknown. We studied the clinical usefulness of testing CSF obtained by lumbar puncture for the presence of EBV-DNA as a minimally invasive approach to the diagnosis of AIDS-PCNSL in patients with focal brain lesions. METHODS: Human immunodeficiency virus (HIV)-infected patients with focal brain lesions, observed prospectively during a 30-month period, underwent lumbar puncture if not contraindicated; otherwise, ventricular CSF was obtained at brain biopsy. The presence of EBV-DNA was determined by means of PCR. RESULTS: We evaluated 122 patients: 42 diagnosed with brain lymphoma and the remaining 80 diagnosed with other brain disorders. Cerebrospinal fluid was collected from 101 patients--by lumbar puncture in 95, including 40 patients with AIDS-PCNSL. The sensitivity and specificity of PCR for EBV-DNA detection in lumbar CSF were 80% (95% confidence interval [CI] = 60.9%-91.6%) and 100% (95% CI = 92.6%-100%), respectively. Lumbar puncture and subsequent assessment of EBV-DNA would have allowed a correct diagnosis in 63.2% (95% CI = 46.0%-77.7%) of patients with AIDS-PCNSL and excluded this diagnosis in 76.3% (95% CI = 65.2%-84.8%) of patients without lymphoma (because EBV-DNA was not detected). CONCLUSIONS: The presence of EBV-DNA in lumbar CSF is a sensitive and highly specific diagnostic marker of AIDS-PCNSL, and EBV-DNA detection in this fluid may allow a minimally invasive diagnosis in a large percentage of patients with brain lymphomas.

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.

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.

The natural history of HIV infection in intravenous drug users: risk of disease progression in a cohort of seroconverters.

A multicentre cohort study was carried out to estimate the incidence of AIDS and HIV-related conditions in newly infected intravenous drug users (IVDU). The enrollment criteria included the identification of the seroconversion time. Two hundred and five subjects entered the study, and were followed for a mean of 26 months. Twelve subjects developed clinical AIDS over a 4-year period. The actuarial incidence of AIDS estimated by Kaplan-Meier survival technique was 17.8% by 4 years since seroconversion. The risk of developing AIDS increased significantly after 24 months from seroconversion. Relatively small figures accounted for the lack of statistical association between the risk factors investigated and the disease status.

PIP: The natural course of HIV infection in 205 intravenous drug users (IVDU) presenting at 11 centers in Italy was followed since 1983. Criteria for entering the study were spontaneous attendance at the clinic from January 1983-April 1988; history of IVDU; dated negative HIV test and a positive test not over 18 months later. 13 of the 205 subjects developed AIDS by 30 April 1988, a total of 447.25 person-years, for an overall incidence of AIDS of 2.9% person-years. The clinical presentation was opportunistic infections in 7, wasting syndrome in 5, and encephalopathy in 1. No Kaposi's sarcoma was seen. The incidence of AIDS was 1.5% in the 1st year; 2.9% after 2 years; 9.9% after 3 years and 17.9% after 42-48 months. The risk of AIDS increased significantly 24 months after seroconversion. 11 others developed AIDS-related complex (ARC) in the 4-year period, an incidence of 3.4% person-years. The minor opportunistic infections seen were zona (4 cases), oral candidiasis (2), and hairy leukoplakia (1). 2 subjects had myelopathy. 87 others developed persistent generalized lymphadenopathy (PGL). 45.8% of the group remained asymptomatic. The subjects mean age was 26, and the sex ratio was 4:1 overall, but 12:1 among the AIDS group. The natural history of AIDS in this series closely resembled other reports on male homosexuals.

Phase II controlled trial of post-exposure immunization with recombinant gp160 versus antiretroviral therapy in asymptomatic HIV-1-infected adults. VaxSyn Protocol Team.

OBJECTIVE: To alter the natural course of HIV-1 infection by inducing or potentiating immune responses to HIV-1 envelope glycoprotein. DESIGN: Multicentre, double-blind, three-arm, placebo-controlled study. SETTING: Outpatients attending clinics in two University Hospitals. PATIENTS: Ninety-nine asymptomatic HIV-1-infected adults with CD4+ T-cell counts > 400 and < 600 x 10(6)/l and no previous antiretroviral therapy were included. INTERVENTIONS: Patients were randomly assigned to three groups treated with: (i) gp160 in alum over a 2-year period in combination with placebo for the full study duration (n = 32); (ii) gp160 in alum over a 2-year period in combination with zidovudine for the full study duration (n = 34); and (iii) alum over a 2-year period in combination with zidovudine for the full study duration (n = 33). RESULTS: Immunotherapy was well tolerated and no significant differences in disease progression were seen in the treatment groups. The majority of patients (85%) receiving gp160 showed persistent lymphoproliferative responses to the immunogen and to a different Env antigen preparation. CD4+ cell count changes in patients receiving zidovudine alone were significantly higher than those seen in patients receiving immunotherapy alone after 1 year of treatment. Zidovudine administration was associated with initial transient reduction of plasma viraemia. CONCLUSIONS: Prolonged immunization with a soluble HIV-1 subunit provided no benefit to asymptomatic HIV-1-infected patients and was inferior to zidovudine monotherapy. Furthermore, immunization with gp160 shortened the duration of the transient viral load reduction induced by zidovudine.

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.

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.

Diagnosis of AIDS-related focal brain lesions: a decision-making analysis based on clinical and neuroradiologic characteristics combined with polymerase chain reaction assays in CSF.

OBJECTIVE: To identify disease patterns in AIDS-related focal brain lesions (FBL) and to design a decision-making strategy for differential diagnosis. DESIGN: Prospective study. Probabilities of CNS disorders were calculated using Bayes' theorem according to clinical variables (mass effect at CT or MRI, Toxoplasma serology, anti-Toxoplasma prophylaxis) and to the results of polymerase chain reaction (PCR) assays. PATIENTS: 136 consecutive HIV-infected patients with a definitive diagnosis of FBL-causing disorder observed from 1991 to 1995 in a single clinical setting. INTERVENTIONS: Patients underwent empiric anti-Toxoplasma therapy. After 3 weeks, patients with progressive/stable disease underwent brain biopsy. In 66 patients Epstein-Barr virus (EBV)-DNA, JC virus (JCV)-DNA, and T gondii-DNA amplification was performed by PCR in CSF. Diagnostic criteria were histopathologic examination of bioptic or autoptic tissue specimens for all disorders and complete/partial resolution of FBL after empiric therapy for toxoplasmic encephalitis (TE). RESULTS: Neuroradiologic characteristics did not discriminate between TE and primary CNS lymphoma (PCNSL). Probability of TE was 0.87 in Toxoplasma-seropositive patients with mass effect who were not receiving anti-Toxoplasma prophylaxis, but only 0.59 if prophylaxis was performed. In seronegative patients with mass effect, the likelihood of PCNSL was 0.74. If EBV-DNA or T gondii-DNA tests were positive, the probability of PCNSL or TE increased to more than 0.96. The absence of T gondii-DNA did not exclude the possibility of a TE diagnosis. Among FBL without mass effect, the probability of progressive multifocal leukoencephalopathy (PML) was 0.81; this increased to 0.99 if JCV-DNA testing was positive. Sensitivity of brain biopsy was 93%, with a perioperative morbidity of 12% and a mortality of 2%. CONCLUSIONS: Due to the low diagnostic capability of clinical variables, PCR amplifications in CSF, especially for EBV-DNA and for JCV-DNA, represent, in most cases, an essential step in the differential diagnosis of AIDS-related FBL. This is particularly true in patients with FBL without mass effect or with mass effect and who are either seronegative or undergoing anti-Toxoplasma prophylaxis. Brain biopsy remains a necessary procedure in EBV-DNA-positive cases and in seronegative patients with FBL displaying a mass effect. Positive JCV-DNA testing may obviate the need for brain biopsy in patients with FBL without mass effect. An advanced diagnostic strategy based on combined clinical criteria and PCR tests may allow rapid and accurate identification of patients for prompt brain biopsy or specific therapy.

A new, striking morphological alteration of P-glycoprotein expression in NK cells from AIDS patients.

Natural killer cells from healthy donors express P-glycoprotein on their surface. This molecule is rearranged during the process of cell-mediated cytotoxicity and it appears to be clustered in the cell-to-cell contact regions. By contrast, in HIV-infected subjects this rearrangement is hindered. These results seem to be associated with cytoskeleton network alterations of the cell-mediated killing process occurring in AIDS patients and can contribute to the comprehension of the mechanisms of the natural killer cell deficiency found in these patients.

P-170 glycoprotein (P-170) is involved in the impairment of natural killer cell-mediated cytotoxicity in HIV+ patients.

In the present study we analyze peripheral blood lymphocytes (PBL) from patients with human immunodeficiency virus (HIV) infection for both phenotypic expression and function of P-glycoprotein (P-170). This transmembrane efflux pump is known to be one of the mechanisms responsible for the multidrug resistance (MDR) in cancer therapy and it is also constitutively expressed in normal PBL. P-170 function, evaluated as Rhodamine 123 (Rh123) efflux in flow cytometry, was found to be significantly reduced in CD16+ natural killer (NK) cells from patients with HIV infection. Interestingly, this reduced efflux significantly correlates with the decreased NK cytotoxicity observed in HIV+ patients, as evaluated against the NK-specific K562 target cell line. These results support a possible role of the P-170-related pump in specific immunological lymphocyte function such as NK cell-mediated cytotoxicity.

Role of polymorphonuclear leukocytes in infection by retroviruses with emphasis on the human immunodeficiency virus.

Neutrophil function is an integral part of the host defense against multiple pathogens. Through phagocytosis and production of toxic substances, these short lived cells aid in the effective elimination of invading microorganisms such as bacterial and fungal targets. Viral infections, and in particular those of the retroviral type, appear to suppress the immune response through direct cytotoxic destruction of immune cells or alteration of the biochemical interactions that are essential for eradicating the foreign agent. In this report, we describe abnormalities of neutrophil number and function consequent to HIV and other retroviral infections. A myriad of mechanisms, either alone or in concert may explain the underlying aberrations.

Diminished interferon gamma production may be the earliest indicator of infection with the human immunodeficiency virus.

The degree of clinical severity in human immunodeficiency virus infected patients, ranging from asymptomatic seropositive subjects to acquired immune deficiency syndrome, as well as in individuals at risk was assessed in relation to: (1) T-cell subset balance and expression of markers of T-cell activation; (2) natural killer activity; and (3) interferon gamma production. A decrease in the CD4/CD8 (helper/suppressor) ratio and an increase in the percentage of CD8+ (suppressor/cytotoxic) cells coexpressing markers of activation (HLA-DR or CD25) were closely correlated with the clinical severity of the human immunodeficiency virus infection. Natural killer activity was significantly impaired in patients with acquired immune deficiency syndrome and acquired immune deficiency syndrome-related complex but normal in asymptomatic seropositive individuals and subjects at risk. Interferon gamma production, either in response to mitogens or the antigens from infectious agents commonly affecting human immunodeficiency virus-positive individuals, was decreased in patients with acquired immune deficiency syndrome or acquired immune deficiency syndrome-related complex, with lesser involvement in human immunodeficiency virus-seropositive subjects or individuals at risk. Four of the six persons in the last group seroconverted during the ten months subsequent to evaluation of their immune status. Since production of interferon gamma was diminished in these patients while other assays of immunity were normal, measurement of this lymphokine may be a useful determinant of infection with the human immunodeficiency virus.

Synthetic peptides corresponding to sequences in HIV envelope gp41 and gp120 enhance in vitro production of interleukin-1 and tumor necrosis factor but depress production of interferon-alpha, interferon-gamma and interleukin-2.

Patients with human immunodeficiency virus (HIV) infections have aberrant production of a number of lymphokines and monokines. Envelope glycoproteins are believed to be important in HIV pathogenesis and may influence the production of these cytokines. Therefore, synthetic peptides corresponding to amino acid sequences 735-752 and 846-860 of glycoprotein gp41 and to amino acid sequence 304-328 of gp120 were investigated for their abilities to affect the production of the following cytokines by normal peripheral blood mononuclear cells in the presence of appropriate inducers: interferon (IFN)-alpha, IFN-gamma, interleukin (IL)-1, IL-2, and tumor necrosis factor (TNF). In contrast to cells and inducers alone (or in the presence of a control peptide), gp41 or gp120 synthetic peptides were able to depress the production of IFN-alpha, IFN-gamma and IL-2. In contrast, these peptides produced an elevation of the production of IL-1 and TNF. The effect of the gp41 peptides was more marked than that of gp120 peptides in most cases. These studies indicate that these HIV envelope glycoproteins may be directly responsible for aberrant lymphokine and monokine production in patients infected with this virus and therefore may be at least partially responsible for the pathogenesis of AIDS.

Transmembrane P-glycoprotein (P-gp/P-170) in HIV infection: analysis of lymphocyte surface expression and drug-unrelated function.

P-glycoprotein (P-gp/P-170), a transmembrane efflux pump known to be one of the mechanisms responsible for multidrug resistance in cancer therapy, is constitutively expressed in several solid human tissues as well as in normal peripheral blood lymphocytes and bone marrow cells. In particular, this molecule has been associated with the transport of perforin and other cytolysins in natural killer (NK) and T cytotoxic lymphocytes. In the present study, we analyzed peripheral blood lymphocytes (PBLs) from controls and HIV+ patients for phenotypic expression and function of the P-gp/P-170 molecule. We found that 90% of all PBL subsets (i.e., CD4+, CD8+, CD56+, and CD19+ cells) expressed surface P-gp/P-170 both in controls and HIV+ patients. However, a significant decrease in CD4+/P-170+ and CD19+/P-170+ cells was observed in HIV+ individuals with respect to controls. PHA and IL-2 stimulation of PBLs was unable to increase the expression of P-gp/P-170 both in controls and HIV+ patients, despite the increased detection of the CD25 molecule. On the other hand, stimulation with anti-CD3 determined a significant increase in lymphocyte P-gp/P-170. The function of P-gp/P-170, assessed by a flow cytometric assay for rhodamine-123 (Rh123) efflux, was significantly reduced in CD16+ NK cells and CD19+ B cells from HIV+ patients. The Rh123 efflux by NK cells correlated (p < 0.01) with the NK cytotoxicity against the 51Cr-labeled K562 cell line. Last, the effect of the antiretroviral drugs AZT, ddI, and ddC on P-gp expression and function was evaluated. The dideoxynucleoside compounds did not inhibit P-gp/P-170 function of normal mononuclear cells in vitro, and did not increase P-gp/P-170 expression in vivo, in patients undergoing antiretroviral therapy with AZT. These findings provide further evidence of a possible involvement of the P-gp/P-170 system in specific immunological lymphocyte functions, and especially in cytotoxic-type functions. In addition, it is possible to suggest, on the basis of our experimental data, that the dideoxynucleoside class of antiretroviral agents does not contribute to the phenotypic and functional alterations related to P-glycoprotein during HIV infection.

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.

Giant neutrophils with increased peroxidase activity. Another evidence of dysgranulopoiesis in AIDS.

Using the automated hematologic analyzer Technicon H6000, which classifies leukocytes by their size and peroxidase activity, the authors have observed in nine patients with full-blown acquired immune deficiency syndrome (AIDS) a consistent increase in peroxidase content of circulating neutrophils. The increase in peroxidase activity was homogeneous in three patients (P less than 0.05). The most striking finding, however, was the occurrence of single abnormal neutrophils with peroxidase activity higher than the major neutrophil population (i.e., HPX [high peroxidase] cells). The importance of this phenomenon was correlated with the clinical status, higher HPX values being found in patients with more advanced disease. These instrumental observations were associated with the morphologic finding of atypical neutrophils, much larger than normal, with irregular nuclei and abundant cytoplasm filled with peroxidase-positive granulations. Such cells represent, in the authors' experience, the most common expression of dysgranulopoiesis in AIDS.

Diagnosis of Pneumocystis carinii pneumonia: specificity and sensitivity of polymerase chain reaction in comparison with immunofluorescence in bronchoalveolar lavage specimens.

DNA amplification by the polymerase chain reaction (PCR) is a promising method for the detection of Pneumocystis carinii in immunosuppressed patients. The sensitivity and specificity of the PCR technique has been assessed in comparison with the immunofluorescence method (IF) on bronchoalveolar lavage fluid (BALF). Results correlated in 43 (78.8%) of 52 cases studied. P. carinii PCR gave positive results with BALF from all 32 patients found to have P. carinii pneumonia (PCP); IF gave positive results with 26 of them. PCR was more sensitive and as specific as IF. However, at the present time, we do not believe that it is clinically useful for detection of P. carinii in BALF samples. P. carinii DNA amplification by PCR should be reserved for testing IF-negative BALF samples from patients judged clinically to have PCP.

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.

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.

Detection of parasite related antigens associated with conglutinin binding immune complexes in patients with Schistosoma haematobium.

An ELISA assay was designed to detect the presence of parasite related antigens associated with circulating immune complexes in patients affected by urinary schistosomiasis. The assay makes use of bovine conglutinin as the immune complex recognition unit and of human anti-Schistosoma antibody as the antigen recognition unit. Using this method we showed that 10 of 15 (67%) patients with a positive polyethylene glycol assay had circulating immune complexes in which parasite antigens could be detected.