Fabio Franzetti

Molecular Epidemiology Study of Exogenous Reinfection in an Area with a Low Incidence of Tuberculosis

ABSTRACT In geographical areas with a low incidence of tuberculosis, recurrent tuberculosis is generally due to reactivation of the disease. However, the relative contribution of tuberculosis reinfection increases in parallel with the incidence of disease and is likely to depend on the epidemiological context: factors such as the spread of multidrug resistance, human immunodeficiency virus (HIV) infection, and immigration from developing countries could modify disease transmission in areas at low risk for tuberculosis. A molecular epidemiology study was performed in Lombardy, Northern Italy, where the incidence of tuberculosis is 17.5 cases per 100,000 persons. A total of 2,452 cases of culture-confirmed tuberculosis in 2,127 patients were studied. A group of 32 patients (1.5%), each of whom had two episodes of tuberculosis with cure as the outcome of the first episode and with more than 6 months between the two episodes, were studied by means of restriction fragment length polymorphism DNA fingerprinting analysis. For 5 of the 32 patients (16%), the DNA fingerprinting patterns ofMycobacterium tuberculosis strains responsible for the second episode did not match those of the corresponding isolates of the first episode, indicating exogenous reinfection. Two of these patients developed multidrug-resistant tuberculosis during the second episode, and in three cases the isolates belonged to clusters of M. tuberculosis strains spreading in the community. A fourfold-increased risk for reinfection was observed in immigrant patients compared to Italian subjects. In contrast, a higher risk of relapse rather than reinfection was evidenced in HIV-positive subjects and in patients infected with multidrug-resistant tuberculosis. Episodes of tuberculosis reinfection in areas with a low incidence of tuberculosis are rare compared to those in high-incidence geographical regions. In populations that have immigrated from high-risk areas, reinfection may represent a considerable contributor to the rate of recurrent tuberculosis. This finding emphasizes the importance of containing the spread of epidemic strains in close communities, in order to prevent changes in global tuberculosis trends for developed countries.

Comparative analysis of T-cell turnover and homeostatic parameters in HIV-infected patients with discordant immune-virological responses to HAART

Objective:Inadequate CD4 cell count recovery despite full HIV RNA control occurs in 30% of HAART-treated HIV-infected patients. A better understanding of the relationship between T-cell dynamics and the HIV intracellular reservoir in HIV-infected patients failing to recover CD4 cell count following long-term HAART, is required. Methods:In a cross-sectional study T-cell turnover and homeostatic parameters featuring discordant responses were investigated in 27 immunologic non-responders (INR; CD4 count, ≤ 200 cells/μl; HIV RNA, ≤ 50 copies/ml), 15 virological non-responders (VNR; CD4 count, ≥ 350 cells/μl; HIV RNA, ≥ 10 000) and 22 full responders (FR; CD4 count, ≥ 500 cells/μl; HIV RNA, ≤ 50 copies/ml). Results:INR displayed significantly higher activated CD38CD8 than FR (P < 0.05) and was comparable to VNR (P > 0.05). As compared with VNR and FR, INR displayed the highest level of proliferating Ki67CD4 and apoptotic CD4 cells (P < 0.05). VNR presented lower proliferation and apoptosis than FR and INR. INR displayed the lowest levels of naive T cells (P < 0.05) and a predominant memory pattern. Despite the memory/activated/apoptotic phenotype, INR showed a statistically non-significant reduction in T-cell receptor excision circles (TREC) compared to FR (P > 0.05), and substantially heightened interleukin (IL)-7 (P < 0.05), while VNR showed higher naive T-cell counts and TREC. Moreover, the reservoir of infected CD4 cells was increased in INR, with a trend toward highest intracellular HIV DNA within total, naive and memory CD4 cells. Conclusions:The lack of CD4 cell count recovery in INR seems to reflect a highly activated apoptotic T-cell compartment, with elevated IL-7 and thymic impairment. High levels of intracellular HIV-DNA in INR could be strictly involved in the lack of T-cell reconstitution. Immune correlates of an ultimate direction of the response to HAART, could be exploited in clinical practice for the most effective management of discordant patients, to amend immune imbalances and to improve clinical outcome.

An outbreak of multidrug-resistant tuberculosis involving Hiv-infected patients of two hospitals in Milan, Italy

Objective: To describe an outbreak of multidrug‐resistant tuberculosis (MDR‐TB), amongst HIV‐infected patients, spread from one hospital in Milan to another. Design: Descriptive epidemiological investigation and molecular typing. Methods: All cases identified by intensive case‐finding were described in terms of clinical characteristics, previous nosocomial exposure to an infectious MDR‐TB patient, previous stays in other institutional settings where exposure to MDR‐TB could have occurred, and restriction fragment length polymorphism (RFLP) pattern. Results: Between October 1991 and July 1995, 116 cases of MDR‐TB were identified (85 at hospital A and 31 at hospital B). A single case patient, infected at hospital A, introduced the strain into hospital B. Eighty‐two of the 92 strains available for fingerprinting revealed an identical pattern; 10 strains had unique RFLP patterns. Nosocomial exposure to an infectious MDR‐TB patient was ascertained for 39 of the 56 patients with the ‘outbreak’ RFLP strain at hospital A (69.6%) and for 24 of the 26 patients at hospital B (92.3%). The median duration of exposure was 32 days at hospital A and 40 days at hospital B. For eight patients with the outbreak strain, exposure was determined to have probably occurred in other hospitals, in the community or in prison. Conclusions: This is the largest nosocomial outbreak of MDR‐TB reported in Europe. Exposure to MDR‐TB cases in other institutions caring for HIV‐infected patients probably contributed to the spread of this epidemic. Strict control measures should be immediately adopted in order to prevent the spread of TB amongst HIV‐infected patients in institutional settings in Europe.

Spoligotyping and Mycobacterium tuberculosis

Speed of spoligotyping could be a benefit in the clinical setting.

Pseudomonas infections in patients with AIDS and AIDS‐related complex

Abstract. We identified and reviewed retrospectively all the cases of infection by Pseudomonas and related genera in patients with AIDS and AIDS‐related complex (ARC) who were hospitalized at our Institution over a 36‐month period. We recorded 48 episodes of infection in 34 of 355 patients with AIDS, and in two of 73 patients with ARC: 25 pneumonias (9 community‐acquired and 16 of nosocomial origin), 20 urinary tract infections, two soft tissue infections and one sepsis. In 14 of 16 patients with nosocomial pneumonia but in only one of nine patients with community‐acquired pneumonia did we find coexisting opportunistic lung diseases. The following micro‐organisms were isolated: P. aeruginosa in 41 cases. P. fluorescens in three cases. Xanthomonas maltophilia (P. maltophilia) in two cases, P. putida in one case, Comamonas testosteronis (P. testosteronis) and Comamonas acidovorans (P. acidovorans) in one case. Amikacin and ceftazidime, alone or in combination, appear to be the optimal choice of therapy for severe Pseudomonas infections in HIV‐infected patients, although in our study six of 47 isolates were resistant in vitro to amikacin, and nine of 31 isolates were resistant to ceftazidime.

Nosocomial Bacterial Pneumonia in HIV-Infected Patients: Risk Factors for Adverse Outcome and Implications for Rational Empiric Antibiotic Therapy

Background:Nosocomial bacterial pneumonia (NBP) was once considered a common cause of morbidity and mortality among advanced AIDS patients. However, clinical and microbiological characteristics and outcome-associated risk factors in this population are poorly defined.Patients:We conducted a retrospective study of all HIV-infected patients admitted during the period 1988–2002 at the Infectious Diseases Clinic of Milan, Italy, to determine incidence rate and factors affecting mortality of NBP, and to gather clinical and microbiological findings about the condition.Results:We identified 120 episodes of NBP among 4,967 admissions of HIV-infected individuals. A reduction of incidence became evident after the introduction of highly active antiretroviral therapy (HAART). The more common causative agents were Pseudomonas aeruginosa (33%) Staphylococcus aureus (25%) and Streptococcus pneumoniae (21%). Methicillin resistance was frequent among staphylococci (65%). The mortality rate of NBP was 25.8%. Non-statistically significant factors associated with shorter survival were: CD4+ count < 10 cells/ml, concomitant lung neoplasm, and complicated roentgenographic picture. Only one factor was significantly associated with lower survival, both in univariate and multivariate analysis: a methicillin-resistant Staphylococcus serving as an etiologic agent of pneumonia (RR 4.05; 95% CI, 1.076–15.239; p = 0.039).Conclusion:A decline in incidence of NBP in HIV-infected individuals was observed after introduction of HAART. S. aureus and P. aeruginosa were the leading causes of NBP, but frequency of pneumococcal pneumonia was significant. The sole predictor for mortality was methicillin-resistant Staphylococcus as a pneumonia-causing agent.

CD4+ T cell depletion, immune activation and increased production of regulatory T cells in the thymus of HIV-infected individuals.

Mechanisms by which HIV affects the thymus are multiple and only partially known, and the role of thymic dysfunction in HIV/AIDS immunopathogenesis remains poorly understood. To evaluate the effects of HIV infection on intra-thymic precursors of T cells in HIV-infected adults, we conducted a detailed immunophenotypic study of thymic tissue isolated from 7 HIV-infected and 10 HIV-negative adults who were to undergo heart surgery. We found that thymuses of HIV-infected individuals were characterized by a relative depletion of CD4+ single positive T cells and a corresponding enrichment of CD8+ single positive T cells. In addition, thymocytes derived from HIV-infected subjects showed increased levels of activated and proliferating cells. Our analysis also revealed a decreased expression of interleukin-7 receptor in early thymocytes from HIV-infected individuals, along with an increase in this same expression in mature double- and single-positive cells. Frequency of regulatory T cells (CD25+FoxP3+) was significantly increased in HIV-infected thymuses, particularly in priorly-committed CD4 single positive cells. Our data suggest that HIV infection is associated with a complex set of changes in the immunophenotype of thymocytes, including a reduction of intrathymic CD4+ T cell precursors, increased expression of activation markers, changes in the expression pattern of IL-7R and enrichment of T regulatory cells generation.

MYCOBACTERIUM LENTIFLAVUM INFECTION IN IMMUNOCOMPETENT PATIENT

Mycobacterium lentiflavum is a recently described nontuberculous mycobacterium that has mainly clinical importance in young children with cervical lymphadenitis and in immunocompromised patients. We describe a case of chronic pulmonary infection in an immunocompetent patient. Our observation confirms clinical, diagnostic, and treatment difficulties in the management of M. lentiflavum infection.

Clinical diagnosis of mycobacterial diseases versus autopsy findings in 350 patients with AIDS

Three-hundred fifty consecutively autopsied AIDS patients in Milan, Italy, were studied to determine the frequencies of clinical and postmortem diagnoses of mycobacterial diseases, to evaluate the clinical histories of those patients with mycobacterial diseases, and to investigate the reasons for nondiagnosis of mycobacterial diseases during life. Seventy-eight patients (22.3%) had mycobacterial diseases. In 64 cases (18.3%) the diagnosis was made antemortem and in 50 (14.2%) at autopsy; there were 36 cases of concordant clinical and postmortem diagnoses. Nontuberculous mycobacterioses (NTM) were diagnosed in 41 patients (20 clinical/postmortem diagnoses, 11 clinical diagnoses, and 10 postmortem diagnoses), extrapulmonary tuberculosis (TB) in 19 patients (7 clinical/postmortem, 8 clinical, 4 postmortem), and pulmonary TB in 18 patients (9 clinical/postmortem, 9 clinical). Patients with a clinical diagnosis of mycobacteriosis but with no pathological evidence of disease at autopsy were considered to have recovered on the basis of negative culture findings and prolonged antimycobacterial treatment. In Italian patients with AIDS, NTM occurs less frequently and TB more frequently than in American AIDS patients. At least some of the patients reported in this study seemed to have recovered from mycobacterial disease after prolonged treatment. The lack of diagnosis during life can be attributed to aspecific symptoms, a rapidly terminal course, and the presence of concomitant opportunistic diseases.

Interleukin-2 immunotherapy exerts a differential effect on CD4 and CD8 T cell dynamics.

Background: Emerging evidence indicates that CD4 and CD8 T cell recovery is differentially regulated during HIV infection. The hallmark of interleukin-2 (IL-2)-induced immune reconstitution is the selective outgrowth of CD4 through undefined mechanisms. Objective: To delineate the effect of IL-2 on T cell homeostasis by analysing the differential impact of IL-2 immunotherapy on CD4 and CD8 dynamics. Design: A randomized trial of 15 HIV-positive patients, eight receiving IL-2 immunotherapy with highly active antiretroviral therapy (HAART) and seven with HAART alone. Patients were followed for a 48-week period following three IL-2 cycles (overall, 10 weeks in duration). Methods: CD4 and CD8 count, naive and memory immunophenotype, proliferation by Ki67, and CD8+CD38+ activated pattern were measured longitudinally by flow cytometry. Thymic output contribution to both CD4 and CD8 was evaluated by measurement of T cell receptor excision circles (TREC). Wilcoxon test was used to compare results. Results: Compared with changes seen with HAART alone, IL-2 induced a more significant rise in CD4 than CD8 T cell count (P < 0.01), associated with a significant increase in Ki67-proliferating CD4 (P < 0.05), whereas no changes were seen in CD8+Ki67+ (P > 0.05). Furthermore, IL-2 administration was associated with CD4 TREC increase, whereas CD8 TREC remained stable (P > 0.05). Modifications in CD4 and CD8 T cells seen in patients taking only HAART were not associated with changes in CD4 and CD8 TREC. Conclusions: By showing a differential impact on CD4 and CD8 homeostasis, the study suggests that IL-2-associated immune reconstitution results from protean interactions between T cell compartments; this has significant implications for the correct planning of immunotherapeutic strategies.