The extent of B-cell activation and dysfunction preceding lymphoma development in HIV-positive people.

OBJECTIVES: B-cell dysfunction and activation are thought to contribute to lymphoma development in HIV-positive people; however, the mechanisms are not well understood. We investigated levels of several markers of B-cell dysfunction [free light chain (FLC)-κ, FLC-λ, immunoglobulin G (IgG), IgA, IgM and IgD] prior to lymphoma diagnosis in HIV-positive people. METHODS: A nested matched case-control study was carried out within the EuroSIDA cohort, including 73 HIV-positive people with lymphoma and 143 HIV-positive lymphoma-free controls. Markers of B-cell dysfunction were measured in prospectively stored serial plasma samples collected before the diagnosis of lymphoma (or selection date in controls). Marker levels ≤ 2 and > 2 years prior to diagnosis were investigated. RESULTS: Two-fold higher levels of FLC-κ [odds ratio (OR) 1.84; 95% confidence interval (CI) 1.19, 2.84], FLC-λ (OR 2.15; 95% CI 1.34, 3.46), IgG (OR 3.05; 95% CI 1.41, 6.59) and IgM (OR 1.46; 95% CI 1.01, 2.11) were associated with increased risk of lymphoma > 2 years prior to diagnosis, but not ≤ 2 years prior. Despite significant associations > 2 years prior to diagnosis, the predictive accuracy of each marker was poor, with FLC-λ emerging as the strongest candidate with a c-statistic of 0.67 (95% CI 0.58, 0.76). CONCLUSIONS: FLC-κ, FLC-λ and IgG levels were higher > 2 years before lymphoma diagnosis, suggesting that B-cell dysfunction occurs many years prior to lymphoma development. However, the predictive value of each marker was low and they are unlikely candidates for risk assessment for targeted intervention.

Changing utilization of Stavudine (d4T) in HIV-positive people in 2006-2013 in the EuroSIDA study.

OBJECTIVES: The long-term side effects of stavudine (d4T) led to recommendations in 2009 to phase out use of this drug. We aimed to describe temporal patterns of d4T use across Europe. METHODS: Patients taking combination antiretroviral therapy (cART) in EuroSIDA with follow-up after 1 January 2006 were included in the study. cART was defined as d4T-containing [d4T plus at least two other antiretrovirals (ARVs) from any class] or non-d4T-containing (at least three ARVs from any class, excluding d4T). Poisson regression was used to describe temporal changes in the prevalence of d4T use and factors associated with initiating d4T. RESULTS: A total of 5850 patients receiving cART on 1 January 2006 were included in the current analysis, rising to 7768 patients on January 1 2013. During this time, the prevalence of d4T use fell from 11.2% to 0.7%, with an overall decline of 19% per 6 months [95% confidence interval (CI) 19-20%]. d4T use declined fastest in Northern Europe [26% (95% CI 23-29%) per 6 months], and slowest in Eastern Europe [17% (95% CI 16-19%) per 6 months]. In multivariable Poisson regression models, new d4T initiations decreased by 14% per 6 months [adjusted incidence rate ratio (aIRR) 0.86; 95% CI 0.80-0.91]. Factors associated with initiating d4T were residence in Eastern Europe (aIRR 4.31; 95% CI 2.17-9.98) versus other European regions and HIV RNA > 400 copies/mL (aIRR 3.11; 95% CI 1.60-6.02) versus HIV RNA < 400 copies/mL. CONCLUSIONS: d4T use has declined sharply since 2006 to low levels in most regions; however, a low but persistent level of d4T use remains in Eastern Europe, where new d4T initiations post 2006 are also more common. The reasons for the regional differences may be multifactorial, but it is important to ensure that all clinicians treating HIV-positive patients are aware of the potential harmful effects associated with d4T.

Insights on the genotype distribution among Cryptococcus neoformans and C. gattii Portuguese clinical isolates.

This study provides a comprehensive picture of the C. neoformans/C. gattii molecular types most often associated with human cryptococcosis in Portugal and assesses the impact of C. gattii in these infections. One hundred and twenty-two clinical isolates, from distinct patients, were identified as C. neoformans and genotyped by URA5-RFLP, with the molecular types VNI (45.5 %) and VNIII (30.9 %) being the most commonly found ones. The molecular types VNII (11.4 %) and VNIV (11.4 %) were less abundant. One patient was found to be infected with a VGII isolate. This patient exhibited unusual clinical symptoms of cryptococcosis, reinforcing the suspicion for the presence of a different genotypic pattern, as determined afterwards. This case was detected in 2007 and is the first report of a potential autochthonous C. gattii infection case in Portugal, as the patient revealed no historical record of travelling outside the country.

Clinical relevance of multiple single-nucleotide polymorphisms in Pneumocystis jirovecii Pneumonia: development of a multiplex PCR-single-base-extension methodology.

Pneumocystis jirovecii pneumonia (PcP) is a major cause of respiratory illness in patients with AIDS. The identification of multiple single-nucleotide polymorphisms (SNPs) at three distinct P. jirovecii loci encoding dihydrofolate reductase (DHFR), mitochondrial large-subunit rRNA (mtLSU rRNA), and superoxide dismutase (SOD) was achieved using multiplex-PCR (MPCR) followed by direct sequencing and two single-base extension (SBE) techniques. Four SNPs (DHFR312, mt85, SOD215, and SOD110), correlated previously with parameters of disease, were amplified and genotyped simultaneously. The concordance of results between the standard sequencing technique (direct sequencing) and SBE analysis was 96.9% for the acrylamide gel electrophoresis and 98.4% for the capillary electrophoresis. The cross-genetic analysis established several statistical associations among the SNPs studied: mt85C-SOD110T, SOD110T-SOD215C, and SOD110C-SOD215T. These results were confirmed by cluster analysis. Data showed that among the isolates with low to moderate parasite burden, the highest percentages of DHFR312C, mt85C, SOD110T, and SOD215C were detected, whereas for high parasite burden cases the highest frequencies were observed among isolates with DHFR312T, mt85T, SOD110C, and SOD215T. The polymorphisms studied were shown to be suitable genetic targets potentially correlated with PcP clinical data that can be used as predictors of outcome in further studies to help clinical decision-making in the management of PcP. The MPCR/SBE protocol described for the first time in the present study was shown to be a rapid, highly accurate method for genotyping P. jirovecii SNPs encoded by different loci that could be used for epidemiological studies and as an additional procedure for the prognostic classification and diagnosis of PcP.

Lymphogranuloma venereum serovar L2b in Portugal.

Chlamydia trachomatis, serovar L2, is the causative agent of lymphogranuloma venereum (LGV), which during recent years has been responsible for various outbreaks reported among men who have sex with men (MSM) in Western Europe, America, Canada and Australia. Samples from nine patients with chronic proctitis, seen at a local hospital were sent to us for identification of C. trachomatis serovar L2. The presence of C. trachomatis serovar L DNA was identified by realtime polymerase chain reaction (PCR) in two patients. They both had high positive C. trachomatis antibody titres (>/=10,000) and were found to be infected with serovar L2b by sequencing after amplification of the omp 1 gene by a nested PCR technique. These two individuals met the diagnostic criteria for LGV serovar L2b infection and, to our knowledge, these are the first cases described in Portugal.

Characterization of HIV-2 chimeric viruses unable to use CCR5 and CXCR4 coreceptors.

We have previously shown the existence of primary human immunodeficiency virus type 2 isolates (MIC97 and MJC97) unable to use major coreceptors to entry into peripheral blood mononuclear cells, including CCR5 and CXCR4. We have now created a set of chimeric viruses derived from HIV-2(ROD), to study the contribution of env gene products in chemokine receptors usage and replication kinetics phenotype. The results obtained indicate that an env gene fragment, corresponding to the C1-C4 region of SU glycoprotein of both MIC97 and MJC97, impair efficient utilization of the major HIV coreceptors CCR5 and CXCR4 in phytohemagglutinin-stimulated peripheral blood mononuclear cells by ROD-MIC97 and ROD-MJC97 chimeric viruses. It also disrupts the ability to utilize established coreceptors for viral entry into GHOST-CD4 coreceptor-expressing cell lines. Resistance to CCR5 and CXCR4 inhibitors, as well as the ability to infect Delta32/Delta32ccr5 PBMC, was also observed in recombinant viruses containing the C1-C4 region from either MIC97 or MJC97. We also show that the presence of the TM region of env gene from MIC97 and MJC97 is sufficient to reduce viral replicative kinetics of ROD strain, indicating that this region, despite the presence and contribution of ROD genetic backbone, has an important role in viral progeny production efficiency.

Anorectal lymphogranuloma venereum: the first two confirmed cases in Portugal.

We describe two cases of lymphogranuloma venereum (LGV) in men who have sex with men in Portugal in 2008. These first two confirmed cases of LGV L2b proctitis in Portugal highlight the need for an enhanced surveillance programme in Portugal.

[The acquired immunodeficiency syndrome, hypertrophic myocardiopathy and multivalvular infectious endocarditis. Apropos a clinical case]. / Síndrome de imunodeficiência adquirida, miocardiopatia hipertrófica e endocardite infecciosa multivalvular. A propósito de um caso clínico.

We present a clinical case of a 33 years old young male, gypsy, intravenous drug abuser with heroine and cocaine and AIDS diagnosis. The clinical anamnesis was mainly fever and systolic heart murmur in a clinical scenario of AIDS. The two-dimensional echocardiographic study was clearly diagnostic of an hypertrophic obstructive cardiomyopathy of the left ventricle. This study showed also the presence of multiple vegetations of the mitral, aortic and pulmonic valves in a clinical setting of an acute Streptococcus Viridans infective endocarditis. In this case report we discuss the incidence of this type of multiple cardiac lesions and particularly the presence of this specific pathogenic agent in this high risk group of patients with intravenous drug abuse and systemic immunosuppression. We pointed out the rarity of these findings of left side valvular vegetations associated with this type of cardiomyopathy and the different factors related to infective endocarditis.

[Human immunodeficiency virus type 2 infection]. / Infecção pelo vírus da imunodeficiência humana do tipo 2.

The isolation of a second retrovirus, HIV-2, led to fears that a second AIDS pandemic, similar in scope and magnitude to that caused by HIV-1, might ensue. However, the peculiar biologic properties of HIV-2, namely the lower transmissibility of this virus through both sexual and vertical routes, contributed to a more regionalized distribution of the virus, which became endemic in West Africa. HIV-2 is genetically more closely related to SIV than to HIV-1. When it comes to clinical disease, the spectrum of opportunistic infections and tumors (except for Kaposi sarcoma) are similar to that observed with HIV-1. Controlled longitudinal studies suggest that the rate of progression to advanced HIV related disease and mortality are far lower for HIV-2 than for HIV-1. Understanding how, immunologically and virologically, HIV-2 behaves differently from HIV-1 may provide some insight into the mechanisms governing HIV-1 pathogenesis.

[Cutaneous infection with the cytomegalovirus virus in AIDS patients]. / Infecção cutânea por vírus citomegálico em doentes com SIDA.

Cutaneous cytomegalovirus (CMV) infection has been observed in a variety of nonspecific skin lesions. Because of this fact, its diagnosis is rare and frequently accidental. The presence of the virus has also been observed in apparently normal skin, from both a clinical and histological point of view. In this context, skin biopsy and immunohistochemistry are often the first means of diagnosis of systemic CMV infection. In 180 skin biopsies carried out on HIV patients in the Infectious and Parasitic Diseases Unit, typical histopathological findings of CMV infection in a nonspecific skin lesion were observed in only one patient. Although the patient showed no extracutaneous manifestations at this time, she died soon after this diagnosis. Because of this fact, we review the literature and discuss the difficulties and implications of the diagnosis of cutaneous CMV infection in AIDS patients.

[Verrucous herpes zoster in AIDS patients]. / Herpes zoster verrucoso em doentes com SIDA.

The authors describe a case of disseminated Herpes-Zoster (VZV) in an HIV 1 positive patient with AIDS. Hyperkeratotic characteristics, acyclovir resistance and sensitivity to foscarnet of cutaneous lesions are the most important features of this example. From the casuistics of the department, the authors describe two similar cases and review the medical literature with emphasis on etiopathogenic, diagnostic and therapeutic factors of lesions caused by DNA Virus in immunocompromised hosts.

[Immunologic profile of HIV-2 seropositive African individuals (follow-up)]. / Perfil imunológico de indivíduos africanos HIV-2 seropositivos (follow-up).

In the geographic distribution of HIV-2, it is known that this infection is most prevalent in West Africa. Since 1986 we have studied seropositive and seronegative clusters, in Guinea-Bissau with follow-ups in 1988, 1989, 1990 and 1991. Analysis of the results show the high incidence of this infection. 8.51% of the 4,372 people of the general population studied were seropositive, showing the high predominance of HIV-2 infection. Only 4 cases were exclusively reactive to HIV-1 and a slow evolution of HIV-1 infections. In the seroconversions of HIV-2 infections the antibodies appeared first to the core components and secondly to the surface glycoproteins. Some of the laboratory parameters affected in the evolution of the infection include a gradual increase in immunoglobulins and a decrease in CD4 lymphocytes and in the CD4/CD8 ratio. A comparison of these variations in HIV-2 infected people, with or without cross-reactivity to HIV-1, reveals that they are much more evident in exclusively HIV-2 positive people. This fact can indicate that the variants responsible for the cross-reactions are less pathogenic and phylogenetically less developed.

PIP: In 1986, a total of 4372 persons were included in an HIV-2 seroepidemiological study covering the whole of Guinea-Bissau. An 8.5% incidence of HIV-2 infection was revealed by the enzyme-linked immunosorbent assay (ELISA) confirmed by Western Blot and/or RIPA. Only 4 cases of HIV-1 seropositivity were found. Annual follow-up of 78 seropositive (e died of AIDS) and 320 seronegative individuals in 1988, 1989, 1990, and 1991 was initiated. In the present investigation, a total of 70 individuals were included: 19 were seropositive (6 with double reactivity) and 51 were seronegative. During this period there were 3 seroconversion cases among seronegative persons, and the appearance of double reactivity in 3 previously HIV-2 positive cases. The reexamination of sera before seroconversion indicated reactivity to the core proteins and in 3 cases to GP-160 or GP-140 surface glycoproteins. The average values of immunoglobulins increased, but the beta-2 microglobulin, and cytomegalovirus antibodies (CMV) did not differ in the sera of 1250 decreased patients compared with seronegative persons. In seropositives, a progressive diminution of the CD4 lymphocyte values and of the T4/T8 (CD4/CD8) ratio was observed which was much more evident in HIV-2 monoreactive patients than in double reactive patients. The interpretation of HIV-1 and HIV-2 double reactivity whether the result of a double infection or of an intermediate variant of the virus between HIV-1 and HIV-2 has not been reconciled with these results. These findings indicate that the variants responsible for double reactivity are less pathogenic, less adapted to the human host, and less developed phylogenetically.

Unusual coexistence of opportunistic lung infections in a human immunodeficiency virus positive patient suffering from persistent Pneumocystis jirovecii pneumonia: a case report.

It is well established that HIV patients are at high risk of opportunistic infections (OI), like the ones caused by Pneumocystis jirovecii, a worldwide pathogen implicated in interstitial pneumonia (PcP). We present a case of a newly diagnosed HIV-1 patient with multiple OI, including a persistent form of PcP, an invasive aspergillosis (IA), cytomegalovirus and Mycobacterium xenopi lung infection. We describe the combination of laboratorial screening, surgery and antimicrobial therapy that were crucial for patient recovery.

Pneumocystis jirovecii multilocus genotyping in pooled DNA samples: a new approach for clinical and epidemiological studies.

Specific single-nucleotide polymorphisms (SNPs) are recognized as important DNA sequence variations influencing the pathogenesis of Pneumocystis jirovecii and the clinical outcome of Pneumocystis pneumonia, which is a major worldwide cause of illness among immunocompromised patients. Genotyping platforms for pooled DNA samples are promising methodologies for genetic characterization of infectious organisms. We have developed a new typing strategy for P. jirovecii, which consisted of DNA pools prepared according to clinical data (HIV diagnosis, microscopic and molecular detection of P. jirovecii, parasite burden, clinical diagnosis and follow-up of infection) from individual samples using quantitative real-time PCR followed by multiplex-PCR/single base extension (MPCR/SBE). The frequencies of multiple P. jirovecii SNPs (DHFR312, mt85, SOD215 and SOD110) encoded at three distinct loci, the dihydrofolate reductase (DHFR), the mitochondrial large-subunit rRNA (mtLSU rRNA) and the superoxide dismutase (SOD) loci, were estimated in seven DNA pooled samples, representing a total of 100 individual samples. The studied SNPs were confirmed to be associated with distinct clinical parameters of infection such as parasite burden and follow-up. The MPCR/SBE-DNA pooling methodology, described in the present study, was demonstrated to be a useful high-throughput procedure for large-scale P. jirovecii SNPs screening and a powerful tool for evaluation of clinically relevant SNPs potentially related to parasite burden, clinical diagnosis and follow-up of P. jirovecii infection. In further studies, the candidate SNPs mt85, SOD215 and SOD110 may be used as molecular markers in association with MPCR/SBE-DNA pooling to generate useful information for understanding the patterns and causes of Pneumocystis pneumonia.

Identification of relevant single-nucleotide polymorphisms in Pneumocystis jirovecii: relationship with clinical data.

Pneumocystis jirovecii is a poorly understood pathogen that causes opportunistic pneumonia (Pneumocystis pneumonia (PcP)) in patients with AIDS. The present study was aimed at correlating genetic differences in P. jirovecii isolates and clinical patient data. A description of genetic diversity in P. jirovecii isolates from human immunodeficiency virus-positive patients, based on the identification of multiple single-nucleotide polymorphisms (SNPs) at five distinct loci encoding mitochondrial large-subunit rRNA (mtLSU rRNA), cytochrome b (CYB), superoxide dismutase (SOD), dihydrofolate reductase (DHFR), and dihydropteroate synthase (DHPS), was achieved using PCR with DNA sequencing and restriction fragment length polymorphism analysis. The statistical analysis revealed several interesting correlations among the four most relevant SNPs (mt85, SOD110, SOD215, and DHFR312) and specific clinical parameters: mt85C was associated with undiagnosed or atypical PcP episodes and favourable follow-up; SOD215C was associated with favourable follow-up; and DHFR312T was associated with PcP cases presenting moderate to high parasite burdens. The genotypes mt85C/SOD215C and SOD110T/SOD215C were found to be associated with less virulent P. jirovecii infections, whereas the genotype SOD110T/SOD215T was found to be related to more virulent PcP episodes. The present work demonstrated that potential P. jirovecii haplotypes may be related to the clinical data and outcome of PcP.

Adult-Onset Still's Disease and cytomegalovirus infection.

We present a case of a previously asymptomatic 34-year-old man that presented to the emergency department with two weeks of fever, arthralgia of the wrists and knees and sore throat. He was diagnosed with cytomegalovirus (CMV) mononucleosis. The patient remained symptomatic in the 5 following months. After an extensive workup to exclude other clinical conditions, a liver biopsy was performed and CMV hepatitis was diagnosed. He started valganciclovir therapy. Approximately one year after the initial complaints, the patient remained ill and presented clinical criteria compatible with Adult Onset Still's Disease. The patient had a marked improvement after institution of prednisolone, an effect that has been sustained during the following months.

Coreceptor usage by HIV-1 and HIV-2 primary isolates: the relevance of CCR8 chemokine receptor as an alternative coreceptor.

The human immunodeficiency virus replication cycle begins by sequential interactions between viral envelope glycoproteins with CD4 molecule and a member of the seven-transmembrane, G-protein-coupled, receptors' family (coreceptor). In this report we focused on the contribution of CCR8 as alternative coreceptor for HIV-1 and HIV-2 isolates. We found that this coreceptor was efficiently used not only by HIV-2 but particularly by HIV-1 isolates. We demonstrate that CXCR4 usage, either alone or together with CCR5 and/or CCR8, was more frequently observed in HIV-1 than in HIV-2 isolates. Directly related to this is the finding that the non-usage of CXCR4 is significantly more common in HIV-2 isolates; both features could be associated with the slower disease progression generally observed in HIV-2 infected patients. The ability of some viral isolates to use alternative coreceptors besides CCR5 and CXCR4 could further impact on the efficacy of entry inhibitor therapy and possibly also in HIV pathogenesis.