In April 2015, an outbreak of dengue-like illness occurred in Tuparetama, a small city in the northeast region of Brazil; this outbreak was characterized by its fast expansion. An investigation was initiated to identify the viral etiologies and advise the health authorities on implementing control measures to contain the outbreak. This is the first report of this outbreak in the northeast, even though a few cases were documented earlier in a neighboring city.Plasma samples were obtained from 77 suspected dengue patients attending the main hospital in the city. Laboratory assays, such as real-time reverse transcription polymerase chain reaction, virus cDNA sequencing, and enzyme-linked immunosorbent assay, were employed to identify the infecting virus and molecular phylogenetic analysis was performed to define the circulating viral genotypes.RNA of Zika virus (ZIKV) and Dengue virus (DENV) or IgM antibodies (Abs) to DENV or chikungunya (CHIKV) were detected in 40 of the 77 plasma samples (51.9%). DENV was found in 9 patients (11.7%), ZIKV was found in 31 patients (40.2%), CHIKV in 1 patient (1.3%), and coinfection of DENV and ZIKV was detected in 2 patients (2.6%). The phylogenetic analysis of 2 available partial DENV and 14 ZIKV sequences revealed the identities of genotype 1 and the Asiatic lineage, respectively.Consistent with recent reports from the same region, our results showed that the ongoing outbreak is caused by ZIKV, DENV, and CHIKV. This emphasizes the need for a routine and differential diagnosis of arboviruses in patients with dengue-like illness. Coordinated efforts are necessary to contain the outbreak. Continued surveillance will be important to assess the effectiveness of current and future prevention strategies.

Objective: To identify a new circulating recombinant form (CRF) of HIV-1 comprising two circulating subtypes in the southern region in Brazil, subtypes B and C. Methods: A total of 152 HIV-positive patients followed at two hospitals in southern Brazil had their viral pol genes isolated by reverse transcriptase–polymerase chain reaction (PCR) from plasma. PCR products were sequenced and phylogenetically analysed using HIV-1 subtype reference sequences. Six full-length subtype C viruses from Brazil previously described as ‘pure’ strains were included in the analysis. Sequences suggestive of recombination were analysed by bootscanning and phylogenetic analyses of separate fragments. The common ancestry of recombinant strains was evaluated by similarity plot and informative site analyses. Results: HIV-1 subtypes commonly found in Brazil (B, C and F1) were observed. Sixty-two viruses were initially assigned as subtype C, but 15 viruses clustered in a separate internal clade. Pol from two full-length genomes of subtype C viruses grouped together with those samples. Bootscanning analysis showed that all 17 viruses had the same recombinant structure, with a 240 base pair fragment of subtype B in the middle of the reverse transcriptase pol region. Subtype B assignment of this fragment was confirmed by phylogenetic analyses using different methods of tree inference and cluster robustness tests. Mosaics were shown to have a common ancestry. Conclusion: As CRF_BC represents 11% of the HIV-1 viruses circulating in the southern region of the country, which borders several south American countries, the assessment of its spread is of pivotal importance to the HIV/AIDS epidemic in Brazil and Latin America.

Great efforts have been made to increase accessibility of HIV antiretroviral therapy (ART) in low and middle-income countries. The threat of wide-scale emergence of drug resistance could severely hamper ART scale-up efforts. Population-based surveillance of transmitted HIV drug resistance ensures the use of appropriate first-line regimens to maximize efficacy of ART programs where drug options are limited. However, traditional HIV genotyping is extremely expensive, providing a cost barrier to wide-scale and frequent HIV drug resistance surveillance. We have developed a low-cost laboratory-scale next-generation sequencing-based genotyping method to monitor drug resistance. We designed primers specifically to amplify protease and reverse transcriptase from Brazilian HIV subtypes and developed a multiplexing scheme using multiplex identifier tags to minimize cost while providing more robust data than traditional genotyping techniques. Using this approach, we characterized drug resistance from plasma in 81 HIV infected individuals collected in São Paulo, Brazil. We describe the complexities of analyzing next-generation sequencing data and present a simplified open-source workflow to analyze drug resistance data. From this data, we identified drug resistance mutations in 20% of treatment naïve individuals in our cohort, which is similar to frequencies identified using traditional genotyping in Brazilian patient samples. The developed ultra-wide sequencing approach described here allows multiplexing of at least 48 patient samples per sequencing run, 4 times more than the current genotyping method. This method is also 4-fold more sensitive (5% minimal detection frequency vs. 20%) at a cost 3–5× less than the traditional Sanger-based genotyping method. Lastly, by using a benchtop next-generation sequencer (Roche/454 GS Junior), this approach can be more easily implemented in low-resource settings. This data provides proof-of-concept that next-generation HIV drug resistance genotyping is a feasible and low-cost alternative to current genotyping methods and may be particularly beneficial for in-country surveillance of transmitted drug resistance.

Rapid evolution and high intrahost sequence diversity are hallmarks of human and simian immunodeficiency virus (HIV/SIV) infection. Minor viral variants have important implications for drug resistance, receptor tropism, and immune evasion. Here, we used ultradeep pyrosequencing to sequence complete HIV/SIV genomes, detecting variants present at a frequency as low as 1%. This approach provides a more complete characterization of the viral population than is possible with conventional methods, revealing low-level drug resistance and detecting previously hidden changes in the viral population. While this work applies pyrosequencing to immunodeficiency viruses, this approach could be applied to virtually any viral pathogen.

The occurrence of chronic myeloid leukemia in pregnancy is rare and its management poses a clinical challenge for physicians treating these patients. We report a 30-year-old woman with chronic myeloid leukemia who became pregnant twice successfully. Philadelphia-positive CML in its chronic phase was diagnosed at 16 weeks of her first gestation. At that time, she received no treatment throughout her pregnancy. At 38 weeks of gestation, a normal infant was delivered by cesarean section. At six weeks postpartum, the patient underwent imatinib mesylate therapy but she could not tolerate the treatment. The treatment was then changed to nilotinib at 400 mg orally b.i.d. Two years later, she became pregnant again while she was on nilotinib 200 mg b.i.d. The unplanned pregnancy was identified during her 7.4 weeks of gestation. Because the patient elected to continue her pregnancy, nilotinib was stopped immediately, and no further treatment was given until delivery. Neither obstetrical complications nor structural malformations in neonates in both pregnancies were observed. Both babies' growth and development have been normal. Although this experience is limited to a single patient, the success of this patient demonstrates that the management of chronic myeloid leukemia in pregnant women may be individualized based on the relative risks and benefits of the patient and fetus.

Genetic variability is a major feature of human immunodeficiency virus type 1 (HIV-1) and is considered the key factor frustrating efforts to halt the HIV epidemic. A proper understanding of HIV-1 genomic diversity is a fundamental prerequisite for proper epidemiology, genetic diagnosis, and successful drugs and vaccines design. Here, we report on the partial and near full-length genomic (NFLG) variability of HIV-1 isolates from a well-characterized cohort of recently infected patients in São Paul, Brazil. HIV-1 proviral DNA was extracted from the peripheral blood mononuclear cells of 113 participants. The NFLG and partial fragments were determined by overlapping nested PCR and direct sequencing. The data were phylogenetically analyzed. Of the 113 samples (90.3% male; median age 31 years; 79.6% homosexual men) studied, 77 (68.1%) NFLGs and 32 (29.3%) partial fragments were successfully subtyped. Of the successfully subtyped sequences, 88 (80.7%) were subtype B sequences, 12 (11%) BF1 recombinants, 3 (2.8%) subtype C sequences, 2 (1.8%) BC recombinants and subclade F1 each, 1 (0.9%) CRF02 AG, and 1 (0.9%) CRF31 BC. Primary drug resistance mutations were observed in 14/101 (13.9%) of samples, with 5.9% being resistant to protease inhibitors and nucleoside reverse transcriptase inhibitors (NRTI) and 4.9% resistant to non-NRTIs. Predictions of viral tropism were determined for 86 individuals. X4 or X4 dual or mixed-tropic viruses (X4/DM) were seen in 26 (30.2%) of subjects. The proportion of X4 viruses in homosexuals was detected in 19/69 (27.5%). Our results confirm the existence of various HIV-1 subtypes circulating in São Paulo, and indicate that subtype B account for the majority of infections. Antiretroviral (ARV) drug resistance is relatively common among recently infected patients. The proportion of X4 viruses in homosexuals was significantly higher than the proportion seen in other study populations.