Immune response and protective efficacy of S9 ribosomal protein of Streptococcus pneumoniae in a model of sepsis.

Vaccination is the most promising strategy to reduce the incidence of pneumococcal infection. Although there are vaccines available, all of them are based on polysaccharide antigens (conjugated or not). In addition to their high cost, those vaccines do not cover all serotypes. To overcome these hindrances, we evaluated the immunogenicity and the protective efficacy of the S9 ribosomal protein of Streptococcus pneumoniae with the aim of developing a protein-based vaccine in the future. The gene encoding the S9 ribosomal protein was cloned in pET21-a expression vector, and the recombinant S9 protein was used to immunize mice. Significantly higher levels of anti-S9 immunoglobulin G were achieved (with predominance of immunoglobulin G1) in comparison with the control. Antibodies elicited against S. pneumoniae protein extract in rabbit recognized the recombinant S9 protein by Western blot, thus demonstrating its immunogenicity. Moreover, mice immunized with recombinant S9 protein and challenged with a virulent strain of S. pneumoniae presented a significant reduction of bacteremia after 24 h of infection as compared with the control. However, in the S9-immunized mice the onset of death was insignificantly delayed, but all of them died by the fourth day postinfection.

Field and classroom initiatives for portable sequence-based monitoring of dengue virus in Brazil.

Brazil experienced a large dengue virus (DENV) epidemic in 2019, highlighting a continuous struggle with effective control and public health preparedness. Using Oxford Nanopore sequencing, we led field and classroom initiatives for the monitoring of DENV in Brazil, generating 227 novel genome sequences of DENV1-2 from 85 municipalities (2015-2019). This equated to an over 50% increase in the number of DENV genomes from Brazil available in public databases. Using both phylogenetic and epidemiological models we retrospectively reconstructed the recent transmission history of DENV1-2. Phylogenetic analysis revealed complex patterns of transmission, with both lineage co-circulation and replacement. We identified two lineages within the DENV2 BR-4 clade, for which we estimated the effective reproduction number and pattern of seasonality. Overall, the surveillance outputs and training initiative described here serve as a proof-of-concept for the utility of real-time portable sequencing for research and local capacity building in the genomic surveillance of emerging viruses.

Biochemical and functional properties of a new l-amino acid oxidase (LAAO) from Micrurus lemniscatus snake venom.

This study reports the purification of ML-LAAO, a new LAAO from the venom of Micrurus lemniscatus snake (ML-V), using size exclusion chromatography. ML-LAAO is a 69-kDa glycoprotein that represents ~2.0% of total venom proteins. This enzyme exhibited optimal activity at pH 8.5, displaying high specificity toward hydrophobic l-amino acids. MALDI TOF/TOF and Blast analysis identified internal segments in ML-LAAO that share high sequence identity with homologous snake venom LAAOs. Western blot analysis on two-dimensional SDS-PAGE of ML-V, using anti-LAAO revealed the presence of ML-LAAO isoforms (pI 6.3-8.9). ML-LAAO blocked aggregation induced by collagen on washed platelets in a rather weak manner, it did not, however, inhibit platelet aggregation induced by ADP on platelet-rich plasma. In addition, this enzyme displayed in vitro antibacterial activity against Staphylococcus aureus (MIC/MBC of 0.39 µg/mL) and in vitro leishmanicidal action against Leishmania amazonensis and L. chagasi (IC50 values of 0.14 and 0.039 µg/mL, respectively). These activities were significantly reduced by catalase, suggesting that hydrogen peroxide production is involved in some way. The data presented here revealed that ML-LAAO has bactericidal and leishmanicidal effects, suggesting that it may have therapeutic potential.

Genomic, epidemiological and digital surveillance of Chikungunya virus in the Brazilian Amazon.

BACKGROUND: Since its first detection in the Caribbean in late 2013, chikungunya virus (CHIKV) has affected 51 countries in the Americas. The CHIKV epidemic in the Americas was caused by the CHIKV-Asian genotype. In August 2014, local transmission of the CHIKV-Asian genotype was detected in the Brazilian Amazon region. However, a distinct lineage, the CHIKV-East-Central-South-America (ECSA)-genotype, was detected nearly simultaneously in Feira de Santana, Bahia state, northeast Brazil. The genomic diversity and the dynamics of CHIKV in the Brazilian Amazon region remains poorly understood despite its importance to better understand the epidemiological spread and public health impact of CHIKV in the country. METHODOLOGY/PRINCIPAL FINDINGS: We report a large CHIKV outbreak (5,928 notified cases between August 2014 and August 2018) in Boa vista municipality, capital city of Roraima's state, located in the Brazilian Amazon region. We generated 20 novel CHIKV-ECSA genomes from the Brazilian Amazon region using MinION portable genome sequencing. Phylogenetic analyses revealed that despite an early introduction of the Asian genotype in 2015 in Roraima, the large CHIKV outbreak in 2017 in Boa Vista was caused by an ECSA-lineage most likely introduced from northeastern Brazil. Epidemiological analyses suggest a basic reproductive number of R0 of 1.66, which translates in an estimated 39 (95% CI: 36 to 45) % of Roraima's population infected with CHIKV-ECSA. Finally, we find a strong association between Google search activity and the local laboratory-confirmed CHIKV cases in Roraima. CONCLUSIONS/SIGNIFICANCE: This study highlights the potential of combining traditional surveillance with portable genome sequencing technologies and digital epidemiology to inform public health surveillance in the Amazon region. Our data reveal a large CHIKV-ECSA outbreak in Boa Vista, limited potential for future CHIKV outbreaks, and indicate a replacement of the Asian genotype by the ECSA genotype in the Amazon region.

Human leptospirosis: occurrence of serovars of Leptospira spp. in the state of Minas Gerais, Brazil, from 2008 to 2012.

BACKGROUND: Leptospirosis is an infectious and acute disease caused by Leptospira spp. that have high epidemic potential. This study verified the main Leptospira spp. serovars detected by MAT from serum of patients with suspicion of leptospirosis from 2008 to 2012 in Minas Gerais State. METHODS: The laboratory received sera from 4654 patients. All serum were screened by IgM-ELISA according to the manufacturer's instructions. Each sample reactive or indeterminate were tested against twenty-four serovars of Leptospira by MAT. RESULTS: In this study, 597 patients were classified as reactive on MAT. Only 301 patients were confirmed by laboratory test. It was not possible confirmation by laboratory diagnosis of 296 patients. Among the samples classified as reactive on MAT, 273 patients exhibited titers bigger than 800 for one or more serovars; seroconversion was detected in 28 cases. Percentage of 85.1% of the samples reactive on MAT corresponded to males, 39.4% corresponded to patients aged between 20 and 39 years old. The most common serovars found were Icterohaemorrhagiae, Andamana, Patoc, Tarassovi, Copenhageni, Hardjo and Australis. Concerning the samples that exhibited titers bigger than 800, serovar Icterohaemorrhagiae was also the most common, followed by Copenhageni, Andamana, Patoc, Tarassovi, Grippotyphosa and Canicola. In this study, 40% of the cases occurred to the metropolitan area, state capital and 34 neighboring towns. CONCLUSION: Our results show the possibly spreading serovars in Minas Gerais State and contribute to knowledge of human leptospirosis, aiming at improving the prevention, control of the disease, as well as the treatment of infected patients.

Human leptospirosis: occurrence of serovars of Leptospira spp. in the state of Minas Gerais, Brazil, from 2008 to 2012

Abstract Background: Leptospirosis is an infectious and acute disease caused by Leptospira spp. that have high epidemic potential. This study verified the main Leptospira spp. serovars detected by MAT from serum of patients with suspicion of leptospirosis from 2008 to 2012 in Minas Gerais State. Methods: The laboratory received sera from 4654 patients. All serum were screened by IgM-ELISA according to the manufacturer's instructions. Each sample reactive or indeterminate were tested against twenty-four serovars of Leptospira by MAT. Results: In this study, 597 patients were classified as reactive on MAT. Only 301 patients were confirmed by laboratory test. It was not possible confirmation by laboratory diagnosis of 296 patients. Among the samples classified as reactive on MAT, 273 patients exhibited titers bigger than 800 for one or more serovars; seroconversion was detected in 28 cases. Percentage of 85.1% of the samples reactive on MAT corresponded to males, 39.4% corresponded to patients aged between 20 and 39 years old. The most common serovars found were Icterohaemorrhagiae, Andamana, Patoc, Tarassovi, Copenhageni, Hardjo and Australis. Concerning the samples that exhibited titers bigger than 800, serovar Icterohaemorrhagiae was also the most common, followed by Copenhageni, Andamana, Patoc, Tarassovi, Grippotyphosa and Canicola. In this study, 40% of the cases occurred to the metropolitan area, state capital and 34 neighboring towns. Conclusion: Our results show the possibly spreading serovars in Minas Gerais State and contribute to knowledge of human leptospirosis, aiming at improving the prevention, control of the disease, as well as the treatment of infected patients.