Epidemiological and Molecular Characteristics of HIV-1 Infection in a Sample of Men Who Have Sex With Men in Brazil: Phylogeography of Major Subtype B and F1 Transmission Clusters.

This study describes human immunodeficiency virus 1 (HIV-1) prevalence, associated factors, viral genetic diversity, transmitted drug resistance (TDR), and acquired drug resistance mutations (DRM) among a population of 522 men who have sex with men (MSM) recruited by the respondent-driven sampling (RDS) method, in Goiânia city, the capital of the State of Goiás, Central-Western Brazil. All serum samples were tested using a four-generation enzyme-linked immunosorbent assay (ELISA), and reactive samples were confirmed by immunoblotting. Plasma RNA or proviral DNA was extracted, and partial polymerase (pol) gene including the protease/reverse transcriptase (PR/RT) region was amplified and sequenced. HIV-1 subtypes were identified by phylogenetic inference and by bootscan analysis. The time and location of the ancestral strains that originated the transmission clusters were estimated by a Bayesian phylogeographic approach. TDR and DRM were identified using the Stanford databases. Overall, HIV-1 prevalence was 17.6% (95% CI: 12.6-23.5). Self-declared black skin color, receptive anal intercourse, sex with drug user partner, and history of sexually transmitted infections were factors associated with HIV-1 infection. Of 105 HIV-1-positive samples, 78 (74.3%) were sequenced and subtyped as B (65.4%), F1 (20.5%), C (3.8%), and BF1 (10.3%). Most HIV-1 subtype B sequences (67%; 34 out of 51) branched within 12 monophyletic clusters of variable sizes, which probably arose in the State of Goiás between the 1980s and 2010s. Most subtype F1 sequences (n = 14, 88%) branched in a single monophyletic cluster that probably arose in Goiás around the late 1990s. Among 78 samples sequenced, three were from patients under antiretroviral therapy (ART); two presented DRM. Among 75 ART-naïve patients, TDR was identified in 13 (17.3%; CI 95%: 9.6-27.8). Resistance mutations to non-nucleoside reverse transcriptase inhibitors (NNRTI) predominated (14.7%), followed by nucleoside reverse transcriptase inhibitor (NRTI) mutations (5.3%) and protease inhibitor (PI) mutations (1.3%). This study shows a high prevalence of HIV-1 associated with sexual risk behaviors, high rate of TDR, and high genetic diversity driven by the local expansion of different subtype B and F1 strains. These findings can contribute to the understanding about the dissemination and epidemiological and molecular characteristics of HIV-1 among the population of MSM living away from the epicenter of epidemics in Brazil.

Homogenous HIV-1 subtype B from the Brazilian Amazon with infrequent diverse BF1 recombinants, subtypes F1 and C among blood donors.

In the last decade a growing HIV/AIDS epidemic with increased incidence and AIDS-related mortality has been reported in Northern Brazil from which molecular data are scarce. Also, apparently healthy, adult blood donors, recently diagnosed with HIV-1 represent important sentinel populations for molecular studies. This cross-sectional study describes HIV-1 subtypes in blood donors from three reference public blood centers located in three States in Northern Brazil. HIV-1 pol sequencing (protease/PR, reverse transcriptase/RT) was performed on plasma samples of HIV-1 positive donors from HEMOAM, Manaus, Amazonas (n = 198), HEMERON, Porto Velho, Rondônia (n = 20) and HEMORAIMA, Boa Vista, Roraima (n = 9) collected from 2011-2017. HIV-1 subtypes were identified by REGA, phylogenetic inference; recombinant viruses were characterized by SIMPLOT. Young, single, males predominated, around half was first-time donors. Syphilis co-infection was detected in 17% (39 out of 227), 8% (18 out of 227) was anti-HBc positive. Subtype B represented ≥ 90% in Amazonas, Rondônia and Roraima, subtype C (3.1%) was found in Amazonas and Rondônia; subtype F1 (0.9%) and BF1 recombinants (5.3%) were only detected in Amazonas. Subtype B sequences from Amazonas (n = 179), Rondônia (n = 18) and Roraima (n = 9) were combined with viral strains representative of the BPANDEMIC (n = 300) and BCARIBBEAN/BCAR (n = 200) lineages. The BPANDEMIC lineage predominated (78%) although BCAR lineages were frequent in Roraima (56%) and Amazonas (22%). Subtype C and subtype F1 sequences identified here clustered within Brazilian CBR and F1BR lineages, respectively. Twelve BF1 mosaics showed 11 different recombination profiles: six were singleton unique-recombinant-forms/URFs, one displays a CRF28/29_BF-like recombinant pattern and the remaining four BF1 isolates branched with other Brazilian BF1 viruses previously described and may represent putative new CRF_BF1 from Northern Brazil. Our study shows a highly homogeneous molecular pattern with prevalent subtype B, followed by BF1, and sporadic subtype C and F1 in blood donors from the Northern region. Surveillance studies are important to monitor HIV-1 diversity which can reveal patterns of viral dissemination, especially in a highly endemic, remote and geographically isolated region as Northern Brazil.

Detection of human immunodeficiency virus Type 1 phylogenetic clusters with multidrug resistance mutations among 2011 to 2017 blood donors from the highly endemic Northern Brazilian Amazon.

BACKGROUND: This study describes transmitted drug resistance (TDR) in blood donors diagnosed with human immunodeficiency virus Type 1 (HIV-1) infection from 2011 to 2017 in three reference public blood centers from the Northern Brazilian Amazon. STUDY DESIGN AND METHODS: This was a cross-sectional study on HIV-positive blood donors from HEMOAM, Manaus, Amazonas, AM (n = 198); HEMERON, Porto Velho, Rondônia, RO (n = 20); and HEMORAIMA, Boa Vista, Roraima, RR (n = 9). HIV-1 pol sequences (protease, reverse transcriptase) were analyzed for drug resistance mutations (DRMs) using the Calibrated Population Resistance tool (Stanford). TDR/DRM clusters were investigated by phylogenetic analysis after removing positions associated with drug resistance of Subtype B sequences from untreated and treated subjects from Northern Brazil. RESULTS: Transmitted drug resistance/DRM in blood donors was 11% (25 of 227), all of them from HEMOAM. Most blood donors with TDR/DRM had multiple and similar DRMs. Nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations predominated (10.1%), followed by nucleoside reverse transcriptase inhibitor (NRTI) mutations (5.3%) and protease inhibitor mutations (0.4%). Dual-class NNRTI/NRTI mutations represented 4.8%. Three highly supported Subtype B monophyletic clades mostly composed by individuals from Amazonas with TDR/DRM mutations were identified. The largest transmission cluster contained 10 sequences, eight from HEMOAM and two sequences described previously (one from a treated subject from Amazonas and the other one from Roraima). This cluster was characterized by NRTI (D67N, T69D, T215S/F/L, K219Q) and NNRTI (K101H, K103 N, G190A) mutations. The other two transmission clades comprised only three and two sequences from HEMOAM sharing the E138A NNRTI mutation. CONCLUSIONS: The identification of transmission clusters of multidrug-resistant viruses in blood donors from Amazonas highlight the need of continued monitoring of TDR/DRM and the importance of pretreatment genotyping in the highly endemic Amazonas state.

Leprosy survey among rural communities and wild armadillos from Amazonas state, Northern Brazil.

There is evidence that in southern US, leprosy is a zoonosis infecting wild Dasypus novemcinctus armadillos but the extent of this finding is unknown. This ecological study investigated leprosy in rural communities and in wild armadillos from the Brazilian Amazon. The study area was the Mamiá Lake of Coari municipality, Amazonas State, Northern region, a hyper endemic leprosy area where residents live on subsistence farming, fishing and armadillo hunting and its meat intake are frequent. The leprosy survey was conducted in sixteen communities by a visiting team of specialists. Local partakers provided wild armadillos to investigate M. leprae infection. Volunteers had complete dermato-neurological examination by a dermatologist with expertise in leprosy diagnosis, suspect skin lesions were biopsied for histopathology (Hematoxylin-eosin/HE, Fite-Faraco/FF staining); slit skin smears were collected. Armadillos' tissue fragments (skins, spleens, livers, lymph nodes, adrenal glands, others) were prepared for histopathology (HE/FF) and for M. leprae repetitive element-RLEP-qPCR. Among 176 volunteers, six new indeterminate leprosy cases were identified (incidence = 3.4%). Suspect skin sections and slit skin smears were negative for bacilli. Twelve wild D. novemcinctus were investigated (48 specimens/96 slides) and histopathological features of M. leprae infection were not found, except for one skin presenting unspecific inflammatory infiltrate suggestive of indeterminate leprosy. Possible traumatic neuroma, granuloma with epithelioid and Langhans cells, foreign-body granuloma were also identified. Granulomatous/non-granulomatous dermatitides were periodic-acid-Schiff/PAS negative for fungus. M. leprae-RLEP-qPCR was negative in all armadillos' tissues; no bacillus was found in histopathology. Our survey in rural communities confirmed the high endemicity for leprosy while one armadillo was compatible with paucibacillary M. leprae infection. At least in the highly endemic rural area of Coari, in the Brazilian Amazon region where infectious sources from untreated multibacillary leprosy are abundant, M. leprae infected armadillos may not represent a major source of infection nor a significant public health concern.

Leprosy survey among rural communities and wild armadillos from Amazonas state, Northern Brazil

There is evidence that in southern US, leprosy is a zoonosis infecting wild Dasypus novemcinctus armadillos but the extent of this finding is unknown. This ecological study investigated leprosy in rural communities and in wild armadillos from the Brazilian Amazon. The study area was the Mamia´ Lake of Coari municipality, Amazonas State, Northern region, a hyper endemic leprosy area where residents live on subsistence farming, fishing and armadillo hunting and its meat intake are frequent. The leprosy survey was conducted in sixteen communities by a visiting team of specialists. Local partakers provided wild armadillos to investigate M. leprae infection. Volunteers had complete dermato-neurological examination by a dermatologist with expertise in leprosy diagnosis, suspect skin lesions were biopsied for histopathology (Hematoxylin-eosin/HE, Fite-Faraco/FF staining); slit skin smears were collected. Armadillos' tissue fragments (skins, spleens, livers, lymph nodes, adrenal glands, others) were prepared for histopathology (HE/FF) and for M. leprae repetitive elementRLEP-qPCR. Among 176 volunteers, six new indeterminate leprosy cases were identified (incidence = 3.4%). Suspect skin sections and slit skin smears were negative for bacilli. Twelve wild D. novemcinctus were investigated (48 specimens/96 slides) and histopathological features of M. leprae infection were not found, except for one skin presenting unspecific inflammatory infiltrate suggestive of indeterminate leprosy. Possible traumatic neuroma, granuloma with epithelioid and Langhans cells, foreign-body granuloma were also identified. Granulomatous/non-granulomatous dermatitides were periodic-acid-Schiff/ PAS negative for fungus. M. leprae-RLEP-qPCR was negative in all armadillos' tissues; no bacillus was found in histopathology. Our survey in rural communities confirmed the high endemicity for leprosy while one armadillo was compatible with paucibacillary M. leprae infection. At least in the highly endemic rural area of Coari, in the Brazilian Amazon region where infectious sources from untreated multibacillary leprosy are abundant, M. leprae infected armadillos may not represent a major source of infection nor a significant public health concern.

Characterization of HIV-1 CRF90_BF1 and putative novel CRFs_BF1 in Central West, North and Northeast Brazilian regions.

The Brazilian AIDS epidemic has been characterized by an increasing rate of BF1 recombinants and so far eight circulating recombinant forms/CRFs_BF1 have been described countrywide. In this study, pol sequences (protease/PR, reverse transcriptase/RT) of 87 BF1 mosaic isolates identified among 828 patients living in six Brazilian States from three geographic regions (Central West, North, Northeast) were analyzed. Phylogenetic and bootscan analyses were performed to investigate the evolutionary relationship and mosaic structure of BF1 isolates. Those analyses showed that 20.7% of mosaics (18 out of 87) were CRFs-like isolates, mostly represented by CRF28/CRF29_BF-like viruses (14 out of 18). We also identified five highly supported clusters that together comprise 42 out of 87 (48.3%) BF1 sequences, each cluster containing at least five sequences sharing a similar mosaic structure, suggesting possible new unidentified CRFs_BF1. The divergence time of these five potential new CRFs_BF1 clusters was estimated using a Bayesian approach and indicate that they probably originated between the middle 1980s and the middle 1990s. DNA was extracted from whole blood and four overlapping fragments were amplified by PCR providing full/near full length genomes (FLG/NFLG) and partial genomes. Eleven HIV-1 isolates from Cluster # 5 identified in epidemiologically unlinked individuals living in Central West and North regions provided FLG/NFLG/partial genome sequences with identical mosaic structure. These viruses differ from any known CRF_BF1 reported to date and were named CRF90_BF1 by the Los Alamos National Laboratory. This is the 9th CRF_BF1 described in Brazil and the first one identified in Central West and North regions. Our results highlight the importance of continued molecular screening and surveillance studies, especially of full genome sequences to understand the evolutionary dynamics of the HIV-1 epidemic in a country of continental dimensions as Brazil.

Clinical and laboratory features of HTLV-I asymptomatic carriers and patients with HTLV-I-associated myelopathy/tropical spastic paraparesis from the Brazilian Amazon.

Clinical and laboratory parameters including blood and cerebrospinal fluid (CSF) neopterin were investigated in human-T-lymphotropic-virus-type-I associated-myelopathy/tropical-spastic-paraparesis-HAM/TSP and in HTLV-I carriers. HAM/TSP (n = 11, 2 males/9 females, median age = 48 years), recently diagnosed HTLV-I carriers (n = 21, 15 females/6 males, median age = 44 years), healthy individuals (n = 20, 10 males/10 females, median age = 34.6 years) from the Brazilian Amazon (Manaus, Amazonas State) were investigated. Neopterin was measured (IBL ELISA Neopterin, Germany) in serum samples of all the participants, in CSF of 9 HAM/TSP patients as well as in 6 carriers. In HAM/TSP patients, CSF cell counts, protein and glucose were measured, the Osame's motor-disability-score/OMDS was determined, and brain/spinal cord magnetic-resonance-imaging (MRI) was performed. HAM/TSP patients had normal CSF glucose, leukocyte counts; and normal protein levels predominated. Brain-MRI showed white-matter lesions in 7 out of 11 HAM/TSP patients. OMDS varied from 2-8: 9 were able to walk, 2 were wheel-chair-users. The median serum neopterin concentration in HAM/TSP patients was 6.6 nmol/ L; min. 2.8- max. 12.5 nmol/ L); was lower in carriers (4.3 nmol/L; min. 2.7- max. 7.2 nmol/ L) as well as in healthy participants (4.7 nmol/ L; min. 2.7- max. 8.0 nmol/ L) (p < 0.05). CSF neopterin concentrations in HAM/TSP patients were higher than in serum samples, and higher compared to carriers (p < 0.05). Carriers had similar serum-CSF neopterin concentrations compared to healthy participants. Variable clinical and laboratory profiles were seen in HAM/TSP patients, however our results support the neopterin measurement as a potential biomarker of disease activity.

Antigen-specific secretion of IFNγ and CXCL10 in whole blood assay detects Mycobacterium leprae infection but does not discriminate asymptomatic infection from symptomatic leprosy.

To advance toward a whole blood assay (WBA)-based test capable of facilitating the diagnosis of paucibacillary (PB) leprosy, we evaluated a prototype in-tube WBA using combinations of Mycobacterium leprae antigens. Blood was collected from newly diagnosed untreated PB (n=38), multibacillary (MB) (n=30), healthy household contacts (HHC) of MB (n=27), and endemic controls (n=61) residing in Goiânia and Fortaleza, Brazil. Blood was incubated with M. leprae cell sonicate, recombinant proteins (46f+LID-1; ML0276+LID-1), or controls (phosphate-buffered saline, phytohemagglutinin, M. tuberculosis purified protein derivative). Antigen-specific IFNγ production was observed in 71-84% and 55% of PB and HHC, respectively. Antigen-specific CXCL10 levels were similarly assessed to determine if, unlike IFNγ, CXCL10 could differentiate PB from HHC with repeated exposure/asymptomatic M. leprae infection. The CXCL10 levels induced in response to M. leprae antigens could not, however, differentiate PB from HHC. Despite these limitations, the WBAs reported here still represent important tools for assessing M. leprae infection rates and evaluating the impact of control measures.

Clinical and laboratory features of HTLV-I asymptomatic carriers and patients with HTLV-I-associated myelopathy/tropical spastic paraparesis from the Brazilian Amazon

ABSTRACT Clinical and laboratory parameters including blood and cerebrospinal fluid (CSF) neopterin were investigated in human-T-lymphotropic-virus-type-I associated-myelopathy/tropical-spastic-paraparesis-HAM/TSP and in HTLV-I carriers. HAM/TSP (n = 11, 2 males/9 females, median age = 48 years), recently diagnosed HTLV-I carriers (n = 21, 15 females/6 males, median age = 44 years), healthy individuals (n = 20, 10 males/10 females, median age = 34.6 years) from the Brazilian Amazon (Manaus, Amazonas State) were investigated. Neopterin was measured (IBL ELISA Neopterin, Germany) in serum samples of all the participants, in CSF of 9 HAM/TSP patients as well as in 6 carriers. In HAM/TSP patients, CSF cell counts, protein and glucose were measured, the Osame’s motor-disability-score/OMDS was determined, and brain/spinal cord magnetic-resonance-imaging (MRI) was performed. HAM/TSP patients had normal CSF glucose, leukocyte counts; and normal protein levels predominated. Brain-MRI showed white-matter lesions in 7 out of 11 HAM/TSP patients. OMDS varied from 2-8: 9 were able to walk, 2 were wheel-chair-users. The median serum neopterin concentration in HAM/TSP patients was 6.6 nmol/ L; min. 2.8- max. 12.5 nmol/ L); was lower in carriers (4.3 nmol/L; min. 2.7- max. 7.2 nmol/ L) as well as in healthy participants (4.7 nmol/ L; min. 2.7- max. 8.0 nmol/ L) (p < 0.05). CSF neopterin concentrations in HAM/TSP patients were higher than in serum samples, and higher compared to carriers (p < 0.05). Carriers had similar serum-CSF neopterin concentrations compared to healthy participants. Variable clinical and laboratory profiles were seen in HAM/TSP patients, however our results support the neopterin measurement as a potential biomarker of disease activity.

Application of Mycobacterium Leprae-specific cellular and serological tests for the differential diagnosis of leprosy from confounding dermatoses.

Mycobacterium leprae-specific serological and cell-mediated-immunity/CMI test were evaluated for the differential diagnosis of multibacillary/MB, and paucibacillary/PB leprosy from other dermatoses. Whole-blood assay/WBA/IFNγ stimulated with LID-1 antigen and ELISA tests for IgG to LID-1 and IgM to PGL-I were performed. WBA/LID-1/IFNγ production was observed in 72% PB, 11% MB leprosy, 38% dermatoses, 40% healthy endemic controls/EC. The receiver operating curve/ROC for WBA/LID-1 in PB versus other dermatoses showed 72.5% sensitivity, 61.5% specificity and an area-under-the-curve/AUC=0.75; 74% positive predictive value/PPV, 59% negative predictive value/NPV. Anti PGL-I serology was positive in 67% MB, 8% PB leprosy, 6% of other dermatoses; its sensitivity for MB=66%, specificity=93%, AUC=0.89; PPV=91%, NPV=72%. Anti-LID-1 serology was positive in 87% MB, 7% PB leprosy, all other participants were seronegative; 87.5% sensitivity for MB, 100% specificity, AUC=0.97; PPV=100%, NPV=88%. In highly endemic areas anti-LID-1/PGL-I serology and WBA/LID-1-represent useful tools for the differential diagnosis of leprosy from other confounding dermatoses.

HIV-1 infection and pregnancy in young women in Brazil: socioeconomic and drug resistance profiles in a cross-sectional study.

OBJECTIVES: To describe socioeconomic and antiretroviral (ARV) drug resistance profiles among young pregnant women infected with HIV-1. SETTING: A public health antenatal programme responsible for screening ∼90 000 pregnant women per year for nine different infectious diseases in Central Western Brazil. PARTICIPANTS: 96 young pregnant women (15-24 years) infected with HIV-1. PRIMARY AND SECONDARY OUTCOME MEASURES: Standard interviews and blood samples were taken at the time of recruitment, at the first medical appointment after confirmation of diagnosis of HIV-1 infection, and before ARV prophylaxis initiation. Clinical and laboratory data were retrieved from medical files. HIV-1 pol gene sequences (entire protease/PR, partial reverse transcriptase/RT) were obtained from plasma RNA. ARV resistance mutations (CPR/Stanford HIV-1; International AIDS Society-USA databases) were identified. RESULTS: The median age was 21 years; most reported <8 years education; 73% were recently diagnosed. Approximately 20% (19/96) presented late for antenatal care (after 26 gestational weeks), while 49% reported ≥2 previous pregnancies. Possible heterosexual transmission by an HIV-1 infected partner (17%) and commercial sex work (2%) were reported. The median of CD4 cell count was 526 cells/mm(3); the median viral load was: 10 056 copies/mL in ARV-naïve (48/96) patients and 5881 copies/mL in ARV-exposed (48/96) patients. Two probable seroconversion cases during pregnancy were identified in adolescents. One mother-to-child transmission case (1.0%) was observed. Transmitted drug resistance among ARV-naïve patients was 9.3% (CI 95% 3.3% to 19.6%); secondary drug resistance among ARV-exposed patients was 12.5% (CI 95% 4.7% to 25.6%). CONCLUSIONS: Despite high access to antenatal care, the low socioeconomic-educational profiles seen in these young HIV-1-infected women highlight the necessity of improved public health educational and preventive strategies regarding HIV infection and early unplanned pregnancy.

Incident and long-term HIV-1 infection among pregnant women in Brazil: Transmitted drug resistance and mother-to-child transmission.

Primary infection, seroconversion, and transmitted drug resistance (TDR) during pregnancy may influence the risk of mother-to-child-transmission (MTCT) of HIV-1 infection. This study estimated recent seroconversion, TDR rates, HIV-1 subtypes and pregnancy outcomes among 95 recently diagnosed, antiretroviral (ARV)-naïve pregnant women recruited during antenatal care in central western Brazil. Recent seroconversion was defined by BED-capture enzyme immunoassay (<155 days) and ambiguous nucleotides base calls (<1 year) in pol sequences (protease-PR and reverse transcriptase-RT regions). TDR was evaluated by the Calibrated Population Resistance tool. HIV-1 subtypes were defined by REGA and phylogenetic analyses. The median age of participants was 25 years; the median gestational age at diagnosis was 20.5 weeks. Based on serology and sequence polymorphism, recent infection was identified in 11.6% (11/95) and, 9 of them (82%), probably seroconverted during pregnancy; one MTCT case was observed among them. Three cases of stillbirth were observed among chronic infected patients (3.6%; 3/84). Moderate rate of TDR was observed (9/90, 10%, CI95% 4.7-18.1%). Subtype B was 60% (54/90), 13.3% (12/90) was subtype C, 6.7% (6/90) was subtype F1. Recombinant B(PR) /F1(RT) and F1(PR) /B(RT) viruses comprised 15.5% (14/90); B(PR) /C(RT) mosaics represented 4.4% (4/90). Seroconversion during pregnancy, late presentation to antenatal care and moderate TDR identified in this study represent significant challenges for the MTCT elimination. J. Med. Virol. 88:1936-1943, 2016. © 2016 Wiley Periodicals, Inc.

Multibacillary leprosy patients with high and persistent serum antibodies to leprosy IDRI diagnostic-1/LID-1: higher susceptibility to develop type 2 reactions.

Leprosy inflammatory episodes [type 1 (T1R) and type 2 (T2R) reactions] represent the major cause of irreversible nerve damage. Leprosy serology is known to be influenced by the patient's bacterial index (BI) with higher positivity in multibacillary patients (MB) and specific multidrug therapy (MDT) reduces antibody production. This study evaluated by ELISA antibody responses to leprosy Infectious Disease Research Institute diagnostic-1 (LID-1) fusion protein and phenolic glycolipid I (PGL-I) in 100 paired serum samples of 50 MB patients collected in the presence/absence of reactions and in nonreactional patients before/after MDT. Patients who presented T2R had a median BI of 3+, while MB patients with T1R and nonreactional patients had median BI of 2.5+ (p > 0.05). Anti-LID-1 and anti-PGL-I antibodies declined in patients diagnosed during T1R (p < 0.05). Anti-LID-1 levels waned in MB with T2R at diagnosis and nonreactional MB patients (p < 0.05). Higher anti-LID-1 levels were seen in patients with T2R at diagnosis (vs. patients with T1R at diagnosis, p = 0.008; vs. nonreactional patients, p = 0.020) and in patients with T2R during MDT (vs. nonreactional MB, p = 0.020). In MB patients, high and persistent anti-LID-1 antibody levels might be a useful tool for clinicians to predict which patients are more susceptible to develop leprosy T2R.

Multibacillary leprosy patients with high and persistent serum antibodies to leprosy IDRI diagnostic-1/LID-1: higher susceptibility to develop type 2 reactions

Leprosy inflammatory episodes [type 1 (T1R) and type 2 (T2R) reactions] represent the major cause of irreversible nerve damage. Leprosy serology is known to be influenced by the patient’s bacterial index (BI) with higher positivity in multibacillary patients (MB) and specific multidrug therapy (MDT) reduces antibody production. This study evaluated by ELISA antibody responses to leprosy Infectious Disease Research Institute diagnostic-1 (LID-1) fusion protein and phenolic glycolipid I (PGL-I) in 100 paired serum samples of 50 MB patients collected in the presence/absence of reactions and in nonreactional patients before/after MDT. Patients who presented T2R had a median BI of 3+, while MB patients with T1R and nonreactional patients had median BI of 2.5+ (p > 0.05). Anti-LID-1 and anti-PGL-I antibodies declined in patients diagnosed during T1R (p < 0.05). Anti-LID-1 levels waned in MB with T2R at diagnosis and nonreactional MB patients (p < 0.05). Higher anti-LID-1 levels were seen in patients with T2R at diagnosis (vs. patients with T1R at diagnosis, p = 0.008; vs. nonreactional patients, p = 0.020) and in patients with T2R during MDT (vs. nonreactional MB, p = 0.020). In MB patients, high and persistent anti-LID-1 antibody levels might be a useful tool for clinicians to predict which patients are more susceptible to develop leprosy T2R.

Alterations to antigen-specific immune responses before and after multidrug therapy of leprosy.

This study evaluated the impact of leprosy multidrug therapy (MDT) on cell-mediated immunity (CMI) and antibody responses at diagnosis in untreated paucibacillary (PB) (n=15) and multibacillary (MB) patients (n=15) using a panel of Mycobacterium leprae recombinant antigens (rMLs) (CMI: 46f, ML0276, ML2055, leprosy IDRI diagnostic 1 [LID-1], and ML2629, as negative control; serology: LID-1, 46f, 92f, and 33f, as negative control, and phenolic glycolipid I [PGL-I]) and at 2 time points after MDT (PB: 8-20months; MB: 4-22months). At diagnosis, PB patients produced interferon gamma (IFNγ), and MB patients exhibited low/absent response. Shortly after MDT, IFNγ production in PB patients declined except for LID-1; MB patients produced IFNγ to LID-1. Almost 2years after MDT, IFNγ levels declined in PB and MB patients. Most untreated PB patients were seronegative to PGL-I and rML, remaining so after MDT. Most untreated MB patients were seropositive to all antigens, and IgG to rMLs declined after MDT. Reduction in antigen-specific CMI in PB and in antibody response in MB patients may help monitor MDT effectiveness.

HIV-1 transmitted drug resistance and genetic diversity among patients from Piauí State, Northeast Brazil.

HIV-1 transmitted-drug-resistance and genetic diversity are dynamic and may differ in distinct locations/risk groups. In Brazil, increased AIDS incidence and related mortality have been detected in the Northeast region, differently from the epicenter in the Southeast. This cross-sectional study describes transmitted-dru- resistance and HIV-1 subtypes in protease/PR and reverse transcriptase/RT regions among antiretroviral naïve patients from Piauí State, Northeast Brazil. Among 96 patients recruited 89 (92.7%) had HIV-1 PR/RT regions sequenced: 44 females and 45 males, 22 self-declared as men who have sex with men. Transmitted-drug-resistance was investigated by CPR tool (Stanford HIV-1 Drug Resistance/SDRM). HIV-1 subtypes were assigned by REGA and phylogenetic inference. Overall, transmitted-drug-resistance rate was 11.2% (10/89; CI 95%: 5.8-19.1%); 22.7% among men who have sex with men (5/22; CI 95%: 8.8-43.4%), 10% in heterosexual men (2/20; CI 95%: 1.7-29.3%) and 6.8% in women (3/44; CI 95%: 1.8-17.4%). Singleton mutations to protease-inhibitor/PI, nucleoside-reverse-transcriptase-inhibitor/NRTI or non-nucleoside-reverse-transcriptase-inhibitor/NNRTI predominated (8/10): PI mutations (M46L, V82F, L90M); NRTI mutations (M41L, D67N) and NNRTI mutations (K103N/S). Dual class resistance mutations to NRTI and NNRTI were observed: T215L (NRTI), Y188L (NNRTI) and T215N (NRTI), F227L (NNRTI). Subtype B prevailed (86.6%; 77/89), followed by subtype F1 (1.1%, 1/89) and subtype C (1.1%, 1/89). B/F1 and B/C intersubtype recombinants represented 11.2% (10/89). In Piauí State extensive testing of incidence and transmitted-drug-resistance in all populations with risk behaviors may help control AIDS epidemic locally.

Molecular epidemiology of hepatitis B and hepatitis delta viruses circulating in the Western Amazon region, North Brazil.

BACKGROUND: Hepatitis B virus (HBV) and hepatitis D virus (HDV) represent important public health problems in the Western Amazon region with reported cases of fulminant hepatitis. This cross sectional study describes HBV and HDV genotypes circulating in the Brazilian Amazon region. METHODS: HBsAg positive individuals (n = 224) were recruited in Manaus/Amazonas State (130 blood donors from the Hematology and Hemotherapy Foundation from Amazonas/HEMOAM; 60 subjects from outpatient clinic) and in Eirunepe city (n = 34) from 2003-2009. Most participants (n = 153) lived in Manaus, 63 were from 20 remote isolated municipalities, 8 lived outside Amazonas State. Genotyping was based on PCR products: HBV genotype A-F specific primers, restricted length polymorphism for HDV. HDV isolates were directly sequenced (delta antigen 405 nucleotide fragment) and phylogenetic analysis performed (MEGA; neighbor-joining, Kimura's two parameter). RESULTS: Most participants were young adult males and HBV mono-infection predominated (70.5%, 158/224). Among blood donors, outpatient subjects and individuals from Eirunepe, HBV/A prevailed followed by HBV/D and F (p > 0.05). HBV/A was more frequent in blood donors (p < 0.05). HBV-HDV coinfection rate was 8.5% in blood donors (11/130), 65.0% (39/60) in outpatient subjects and 47.0% (16/34) in individuals from Eirunepe. Compared to blood donors, coinfection was higher in outpatient subjects (65.0% versus 8.5%; RR = 5.0; CI 3.4-7.9; p < 0.0001) and in subjects from Eirunepe (47.0% versus 8.5%; RR = 5.5; CI 3.0-9.9; p < 0.0001). HBV-HDV coinfection rates were higher in patients from highly endemic remote cities. Only HDV genotype 3 was detected, HBV/F-HDV/3 predominated (20/38; 52.7%), followed by HBV/A-HDV/3 (31.6%; 12/38) and HBV/D-HDV/3 (15.8%; 6/38). CONCLUSIONS: The description of HBV and HDV genotypes circulating in the western Amazon can contribute to a better understanding of their relevance on the regional epidemics. These infections are highly endemic in the Amazon where their control is challenged by its vast territorial dimension with small, hard-to-reach municipalities dispersed into the jungle and populated by diverse ethnic groups.

Polymorphisms in the HIV-1 gp41 env gene, natural resistance to enfuvirtide (T-20) and pol resistance among pregnant Brazilian women.

The selective pressure of antiretroviral drugs (ARVs) targeting HIV-1 pol can promote drug resistance mutations in other genomic regions, such as env. Drug resistance among women should be monitored to avoid horizontal and mother-to-child transmission. To describe natural resistance to T-20 (enfuvirtide), gp41 env polymorphisms, mutations in pol and HIV-1 subtypes, 124 pregnant women were recruited. For 98 patients, the gp41 env, protease (PR) and reverse transcriptase (RT) fragments were sequenced. The patients were ARV naïve (n = 30), taking mother-to-child transmission prophylaxis (n = 50), or being treated with highly active ARV therapy/HAART (n = 18). The Stanford and IAS/USA databases and other sources were used to analyze PR/RT, gp41 env resistance mutations. The HIV-1 genetic diversity was analyzed by REGA/phylogenetic analyses. The patients' median age was 25 years (range, 16-42), 18.4% had AIDS. The frequency of natural resistance to T-20 (N42D, L44M, and R46M-low-impact mutations) was 6.1% (6/98); 20.4% (20/98) had compensatory mutations in HR2. The prevalence of transmitted drug resistance in the pol was 13.3% (4/30), and the prevalence of secondary drug resistance was 33.3% (6/18). Two patients were infected with multidrug resistant/MDR viruses. The analysis of HIV-1 subtypes (PR/RT/gp41) revealed that 61.2% (60/98) were subtype B, 12.2% (12/98) were subtype C, 4.1% (4/98) were subtype F1, and 22.4% (22/98) were possible recombinants (BF1 = 20.4%; BC = 2%). Natural resistance to T-20 was not associated with pol resistance or previous ARV use. The high rate of secondary resistance, including MDR, indicates that the number of women that may need T-20 salvage therapy may be higher than anticipated.

Increasing heterosexual transmission of HIV-1 subtype C in Inland Central Western Brazil.

The molecular epidemiology of HIV-1 in Brazil is complex and heterogeneous because several subtypes co-circulate with some important regional differences. This study evaluated HIV-1 subtypes amongst pregnant women living in the metropolitan area and in the interior cities from central western Brazil. From June 2008 to June 2010, 86.9% of confirmed cases of HIV-1 infection amongst pregnant women (172 out of 198 cases) were recruited in Goiania/Goias state. The HIV-1 pol gene was sequenced after nested-PCR. HIV-1 subtypes were assigned by REGA, phylogenetic, and bootscan analyses. The median age of participants was 26 years (15-41 years range); 58.7% of participants were diagnosed during prenatal care and 51.7% of participants came from >50 interior cities within Goias state. Amongst the 131 HIV-1 pol sequences, 64.9% were subtype B, 13.0% were BF1 recombinant, 11.4% were subtype C, 7.6% were subtype F1, and 2.3% were BC recombinant. According to the HIV-1 diagnosis date (1994-2010), a significant increase in subtype C and a decrease of BF1 mosaics were observed over time. All subtype C patients lived in interior cities where the highest prevalence of subtype C outside southern Brazil was observed (18.4%). Phylogenetic analysis revealed multiple independent introductions of the Brazilian subtype C clade from the southern/southeastern regions of Brazil. The HIV-1 epidemic in women from central western Brazil infected by the heterosexual route is characterized by an unexpectedly high prevalence of subtype C viruses highly related to those circulating in southern/southeastern Brazil. These findings highlight the importance of molecular surveillance programs outside large metropolitan regions in Brazil.

HIV-1 mother-to-child transmission and drug resistance among Brazilian pregnant women with high access to diagnosis and prophylactic measures.

BACKGROUND: A high-coverage public health prenatal program (70,000 women/year) from central western Brazil/Goias State has represented a unique opportunity for the early diagnosis of HIV-1 and implementation of strategies to prevent mother-to-child transmission (MTCT). OBJECTIVES: To investigate MTCT among a prospective cohort of HIV-1 infected mothers/exposed infants. STUDY DESIGN: 142 mothers/their 149 infants (2008-2010) were investigated regarding maternal viral load, CD4(+)cell counts, HIV-1 pol sequences; infants' HIV-1 RNA tests (30/120 days), sequential anti-HIV-1/2 serology. HIV-1 subtypes were assigned by REGA. Transmitted drug resistance was identified by the Calibrated Population Resistance tool, secondary resistance by Stanford HIV-1 Drug Resistance/International AIDS Society databases. RESULTS: Mothers (median age=24 years; 25/142 adolescents) were diagnosed during prenatal care (2008-2010) or previously (1994-2007). Recent cases were younger, mostly asymptomatic. Undetectable viremia and MTCT prophylaxis predominated in formerly diagnosed mothers. Recent cases had higher subtype C prevalence. One naive patient had transmitted resistance; ten antiretroviral-experienced patients had secondary resistance: 6 from MTCT prophylaxis, 4 under HAART. Late disclosure of diagnosis, vaginal delivery, breastfeeding, lack of oral zidovudine were observed in the three MTCT cases (3/149; 2.01%). Two of three infected infants harbored subtype C; infected infants/mothers did not have drug resistance mutations. Two of the transmitting-mothers had viremia <1000 copies/ml. Among exposed-uninfected infants the median time to seroreversion was 12 months. CONCLUSIONS: In this study delayed disclosure of diagnosis, partial/no preventive measures, drug resistance among asymptomatic women under prophylaxis and MTCT in low viremic mothers raise concerns. The expansion of subtype C infection corroborates surveillance of HIV-1 diversity in this region.