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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. [ABSTRACT FROM AUTHOR]

Candidemia has been considered a persistent public health problem with great impact on hospital costs and high mortality. We aimed to evaluate the epidemiology and prognostic factors of candidemia in a tertiary hospital in Northeast Brazil from January 2011 to December 2016. Demographic and clinical data of patients were retrospectively obtained from medical records and antifungal susceptibility profiling was performed using the broth microdilution method. A total of 68 episodes of candidemia were evaluated. We found an average incidence of 2.23 episodes /1000 admissions and a 30-day mortality rate of 55.9%. The most prevalent species were Candida albicans (35.3%), Candida tropicalis (27.4%), Candida parapsilosis (21.6%) and Candida glabrata (11.8%). Higher mortality rates were observed in cases of candidemia due to C. albicans (61.1%) and C. glabrata (100%), especially when compared to C. parapsilosis (27.3%). Univariate analysis revealed some variables which significantly increased the probability of death: older age (P = 0.022; odds ratio [OR] = 1.041), severe sepsis (P < 0.001; OR = 8.571), septic shock (P = 0.035; OR = 3.792), hypotension (P = 0.003; OR = 9.120), neutrophilia (P = 0.046; OR = 3.080), thrombocytopenia (P = 0.002; OR = 6.800), mechanical ventilation (P = 0.009; OR = 8.167) and greater number of surgeries (P = 0.037; OR = 1.920). Multivariate analysis showed that older age (P = 0.040; OR = 1.055), severe sepsis (P = 0.009; OR = 9.872) and hypotension (P = 0.031; OR = 21.042) were independently associated with worse prognosis. There was no resistance to amphotericin B, micafungin or itraconazole and a low rate of resistance to fluconazole (5.1%). However, 20.5% of the Candida isolates were susceptible dose-dependent (SDD) to fluconazole and 7.7% to itraconazole. In conclusion, our results could assist in the adoption of strategies to stratify patients at higher risk for developing candidemia and worse prognosis, in addition to improve antifungal management. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) in Brazil is a neglected, vector-borne, tropical parasitic disease that is responsible for several thousand human deaths every year. The transmission route involves sand flies becoming infected after feeding on infected reservoir host, mainly dogs, and then transmitting the Leishmania infantum parasites while feeding on humans. A major component of the VL control effort is the identification and euthanasia of infected dogs to remove them as a source of infection. A rapid, non-invasive, point-of-care device able to differentiate between the odours of infected and uninfected dogs may contribute towards the accurate diagnosis of canine VL. Methodology/Principal findings: We analysed the headspace volatile chemicals from the hair of two groups of dogs collected in 2017 and 2018 using a bench-top eNose volatile organic chemical analyser. The dogs were categorised as infected or uninfected by PCR analysis of blood samples taken by venepuncture and the number of parasites per ml of blood was calculated for each dog by qPCR analysis. We demonstrated using a robust clustering analysis that the eNose data could be discriminated into infected and uninfected categories with specificity >94% and sensitivity >97%. The eNose device and data analysis were sufficiently sensitive to be able to identify infected dogs even when the Leishmania population in the circulating blood was very low. Conclusions/Significance: The study illustrates the potential of the eNose to rapidly and accurately identify dogs infected with Le. infantum. Future improvements to eNose analyser sensor sensitivity, sampling methodology and portability suggest that this approach could significantly improve the diagnosis of VL infected dogs in Brazil with additional potential for effective diagnosis of VL in humans as well as for the diagnosis of other parasitic diseases. [ABSTRACT FROM AUTHOR]

Background: Visceral Leishmaniasis in humans presents with fever, anemia, and splenomegaly and can be lethal if not treated. Nevertheless, the majority of Leishmania infantum-infected individuals does not manifest symptoms and remain so provided they are not immunosuppressed. In this work, the performance of different tests was evaluated to detect asymptomatic individuals who were living in Teresina, Piauí state, Brazil, an endemic area for VL. Methodology: L. infantum-specific antibodies were detected by ELISA and two different rapid immunochromatographic (IC) diagnostic tests, Kalazar Detect and OnSite, and parasitic loads were detected by real time PCR [qPCR]. Additionally, we measured levels of the biomarkers monokine induced by IFN-γ (MIG) and IFN-γ-induced protein 10 (IP-10) before and after stimulation of whole blood with soluble Leishmania antigen [SLA]. Principal findings: Kalazar Detect and OnSite detected, respectively, 76% and 64% of patients presenting with active Visceral Leishmaniasis; 50% and 57% of patients remained positive in these tests, respectively, after treatment. Of the healthy participants in the study who were living in the endemic area, only 1.7% were positive with both of the IC tests. On the other hand, reactivity in ELISA tests revealed that 13% of these individuals presented asymptomatic infections; among VL patients, 84% presenting with active disease were reactive in ELISA, and after treatment, 55.5% were seropositive. L. infantum DNA was present in the blood of 37.9% of infected individuals living in the endemic area, while IP-10 and MIG biomarkers were detected in 26.7% of them. The greatest concordance of positivity occurred between ELISA and qPCR. Conclusion: The association of different techniques can detect asymptomatic infections, however, more research is necessary to develop ideal biomarkers that are simple to use in the clinic and in field studies in areas endemic for Visceral Leishmaniasis. [ABSTRACT FROM AUTHOR]

Detection and sequencing of chikungunya virus (CHIKV) genome was performed using a combination of a modified reverse transcription loop-mediated isothermal amplification (RT-LAMP) method and a MinION sequencer. We developed the protocol for drying all the reagents for the RT-LAMP in a single reaction tube. Using this system, the CHIKV genome was effectively amplified under isothermal conditions, and used as a template for MinION sequencing with a laptop computer. Our in-house RT-LAMP method and MinION sequencing system were also validated with RNAs and serum samples from recent outbreaks of CHIKV patients in Brazil. The obtained sequence data confirmed the CHIKV outbreaks and identified the genotype. In summary, our established inexpensive on-site genome detection and sequencing system is applicable for both diagnosis of CHIKV infected patients and genotyping of the CHIKV virus in future outbreak in remote areas. [ABSTRACT FROM AUTHOR]

The study of host associations of mosquitoes (Diptera, Culicidae) provides valuable information to assist in our understanding of a variety of related issues, from their life-history to the entomological surveillance of pathogens. In this study, we identified and characterized mosquito blood meals from both urban and forested areas in the city of Paranaguá, state of Paraná, Brazil, by analyzing the amplification of host DNA ingested by mosquitoes under different storage conditions and digestion levels. Host DNA preservation was evaluated in fresh blood meals according to storage duration (30 to 180 days) and temperature (-20°C / -80°C) and, in digested blood, according the degree of digestion classified on the Sella scale. Molecular analysis of blood meals was based on DNA extraction and amplification of a fragment of the mitochondrial COI gene. We determined that, up to180 days of storage, the evaluated temperatures did not influence the preservation of fresh blood meals DNA, whereas the amplification success was increasingly reduced over the course of the digestion process. The species Anopheles cruzii, Aedes fluviatilis, Aedes scapularis, Psorophora ferox, Culex quinquefasciatus, Culex mollis, and Culex intrincatus, together with specimens representing four subgenera and one genus of Culicidae [Ae. (Ochlerotatus), Cx. (Culex), Cx. (Melanoconion), Cx. (Microculex), and Limatus, respectively] had their blood meals identified. Their diverse host use was evidenced by the identification of 19 species of vertebrate host, namely two amphibians, three mammals and 14 birds. Birds were the most commonly identified host in blood meals. These results not only show the diversity of mosquito hosts, but also underscore the challenges involved in monitoring arboviruses of public health importance, given potential combinations of host use for each mosquito species. [ABSTRACT FROM AUTHOR]

Candida albicans caused 44% of the overall candidemia episodes from 2006 to 2010 in our university tertiary care hospital. As different antifungal agents are used in therapy and also immunocompromised patients receive fluconazole prophylaxis in our institution, this study aimed to perform an antifungal susceptibility surveillance with the C.albicans bloodstream isolates and to characterize the fluconazole resistance in 2 non-blood C.albicans isolates by sequencing ERG11 gene. The study included 147 C. albicans bloodstream samples and 2 fluconazole resistant isolates: one from oral cavity (LIF 12560 fluconazole MIC: 8μg/mL) and one from esophageal cavity (LIF-E10 fluconazole MIC: 64μg/mL) of two different patients previously treated with oral fluconazole. The in vitro antifungal susceptibility to amphotericin B (AMB), 5-flucytosine (5FC), fluconazole (FLC), itraconazole (ITC), voriconazole (VRC), caspofungin (CASP) was performed by broth microdilution methodology recommended by the Clinical and Laboratory Standards Institute documents (M27-A3 and M27-S4, CLSI). All blood isolates were classified as susceptible according to CLSI guidelines for all evaluated antifungal agents (MIC range: 0,125–1.00 μg/mL for AMB, ≤0.125–1.00 μg/mL for 5FC, ≤0.125–0.5 μg/mL for FLC, ≤0.015–0.125 μg/mL for ITC, ≤0.015–0.06 μg/mL for VRC and ≤0.015–0.125 μg/mL for CASP). In this study, we also amplified and sequenced the ERG11 gene of LIF 12560 and LIF-E10 C.albicans isolates. Six mutations encoding distinct amino acid substitutions were found (E116D, T128K, E266D, A298V, G448V and G464S) and these mutations were previously described as associated with fluconazole resistance. Despite the large consumption of antifungals in our institution, resistant blood isolates were not found over the trial period. Further studies should be conducted, but it may be that the very prolonged direct contact with the oral antifungal agent administered to the patient from which was isolated LIF E-10, may have contributed to the development of resistance. [ABSTRACT FROM AUTHOR]

Aims: To investigate the hypothesis that the change from the female predominance of diabetes mellitus to a standard of equality or even male preponderance can already be observed in Brazilian mortality statistics. Methods: Data on deaths for which diabetes mellitus was listed as the underlying cause were obtained from the Brazilian Mortality Information System for the years 1980 to 2012. The mortality data were also analyzed according to the multiple causes of death approach from 2001 to 2012. The population data came from the Brazilian Institute of Geography and Statistics. The mortality rates were standardized to the world population. We used a log-linear joinpoint regression to evaluate trends in age-standardized mortality rates (ASMR). Results: From 1980 to 2012, we found a marked increment in the diabetes ASMR among Brazilian men and a less sharp increase in the rate among women, with the latter period (2003–2012) showing a slight decrease among women, though it was not statistically significant. Conclusions: The results of this study suggest that diabetes mellitus in Brazil has changed from a pattern of higher mortality among women compared to men to equality or even male predominance. [ABSTRACT FROM AUTHOR]

Background: Efforts have been made to establish sensitive diagnostic tools for malaria screening in blood banks in order to detect malaria asymptomatic carriers. Microscopy, the malaria reference test in Brazil, is time consuming and its sensitivity depends on microscopist experience. Although molecular tools are available, some aspects need to be considered for large-scale screening: accuracy and robustness for detecting low parasitemia, affordability for application to large number of samples and flexibility to perform on individual or pooled samples. Methodology: In this retrospective study, we evaluated four molecular assays for detection of malaria parasites in a set of 56 samples previously evaluated by expert microscopy. In addition, we evaluated the effect of pooling samples on the sensitivity and specificity of the molecular assays. A well-characterized cultured sample with 1 parasite/μL was included in all the tests evaluated. DNA was extracted with QIAamp DNA Blood Mini Kit and eluted in 50 μL to concentrate the DNA. Pools were assembled with 10 samples each. Molecular protocols targeting 18S rRNA, included one qPCR genus specific (Lima-genus), one duplex qPCR genus/Pf (PET-genus, PET-Pf) and one duplex qPCR specie-specific (Rougemont: Roug-Pf/Pv and Roug-Pm/Po). Additionally a nested PCR protocol specie-specific was used (Snou-Pf, Snou-Pv, Snou-Pm and Snou-Po). Results: The limit of detection was 3.5 p/μL and 0.35p/μl for the PET-genus and Lima-genus assays, respectively. Considering the positive (n = 13) and negative (n = 39) unpooled individual samples according to microscopy, the sensitivity of the two genus qPCR assays was 76.9% (Lima-genus) and 72.7% (PET-genus). The Lima-genus and PET-genus showed both sensitivity of 86.7% in the pooled samples. The genus protocols yielded similar results (Kappa value of 1.000) in both individual and pooled samples. Conclusions: Efforts should be made to improve performance of molecular tests to enable the detection of low-density parasitemia if these tests are to be utilized for blood transfusion screening. [ABSTRACT FROM AUTHOR]