SARS-CoV-2 laboratory surveillance during the first year of the COVID-19 pandemic in southern Brazil

ABSTRACT Background: Brazil has one of the highest numbers of COVID-19 cases and deaths. Rio Grande do Sul (RS) in southern Brazil is one of the leading states in terms of case numbers. As part of the national public health network, the State Central Laboratory (LACEN-RS) changed its routine in 2020 to focus on the diagnosis of COVID-19. This study evaluated the laboratory surveillance of COVID-19 suspected cases analyzed at the LACEN-RS in 2020. Methods: Viral detection was performed using RT-qPCR in samples from patients with respiratory infection who met the study criteria. Viral RNA was isolated using commercial manual kits or automated extractors, and SARS-CoV-2 RT-qPCR was performed using the Bio-Manguinhos/Rio de Janeiro, IBMP/Paraná, or Allplex 2019-nCoV assay. In total, 360 representative SARS-CoV-2 samples were sequenced using the Illumina platform. Results: In total, 31,197 of 107,578 (positivity rate = 29%) tested positive for SARS-CoV-2. The number of RT-qPCR tests performed per month followed the COVID-19 epidemic curve observed for the state, with peaks in July-August and December. Females accounted for 63% of the samples, whereas the positivity rate was higher among males (33.1% males vs. 26.5% females). The positivity rate was higher in adults aged 50-79 years compared to the overall positivity rate. The majority of cases were observed in the capital, Porto Alegre, and the metropolitan region. Ten distinct lineages were identified, with B.1.1.28, B.1.1.33, and P.2 being the most frequent. Conclusions: Here, we describe laboratory surveillance of COVID-19 to identify priorities for epidemiological surveillance actions in RS.

Molecular dynamics of the COVID-19 pandemic in Espirito Santo (Brazil) and border States

ABSTRACT This study represents the first overview of the epidemiological dynamics of SARS-CoV-2 in Espirito Santo (ES) State, Brazil, filling in knowledge on this topic, observing data collected in the State, and aiming at understanding the epidemiological dynamics of the virus in ES, as well as its possible routes of transmission and dissemination. . Our results highlight that, so far, nine lineages have been identified with ES State. The B.1.1.33 lineage was the first with the highest occurrence in ES, remaining predominant until September 2020. The second predominant lineage was Gamma, representing 45% of the samples. The Delta lineage appears on the State scene, proving to be the next dominant lineage. This research allowed us to understand how the lineages advanced and were distributed in the State, which is important for future work, also making it possible to guide sanitary control measures. Data analyses were made through the GISAID database for ES State showed that the pandemic in the State has been evolving dynamically with lineage replacements over the months since the first notification.

Community-genotype methicillin-resistant Staphylococcus aureus skin and soft tissue infections in Latin America: a systematic review

ABSTRACT Background: Community-genotype methicillin-resistant Staphylococcus aureus (CG-MRSA) emerged in the 1990s as a global community pathogen primarily involved in skin and soft tissue infections (SSTIs) and pneumonia. To date, the CG-MRSA SSTI burden in Latin America (LA) has not been assessed. Objective: The main objective of this study was to report the rate and genotypes of community-genotype methicillin-resistant Staphylococcus aureus (CG-MRSA) causing community-onset skin and soft tissue infections (CO-SSTIs) in LA over the last two decades. In addition, this research determined relevant data related to SSTIs due to CG-MRSA, including risk factors, other invasive diseases, and mortality. Data sources: Relevant literature was searched and extracted from five major databases: Embase, PubMed, LILACS, SciELO, and Web of Science. Methods: A systematic review was performed, and a narrative review was constructed. Results: An analysis of 11 studies identified epidemiological data across LA, with Argentina presenting the highest percentage of SSTIs caused by CG-MRSA (88%). Other countries had rates of CG-MRSA infection ranging from 0 to 51%. Brazil had one of the lowest rates of CG-MRSA SSTI (4.5-25%). In Argentina, being younger than 50 years of age and having purulent lesions were predictive factors for CG-MRSA CO-SSTIs. In addition, the predominant genetic lineages in LA belonged to sequence types 8, 30, and 5 (ST8, ST30, and ST5). Conclusion: There are significant regional differences in the rates of CG-MRSA causing CO-SSTIs. It is not possible to conclude whether or not CG-MRSA CO-SSTIs resulted in more severe SSTI presentations or in a higher mortality rate.

5S and 5W2H Tools Applied to Research Laboratories: Experience from Instituto Carlos Chagas - FIOCRUZ/PR for Cell Culture Practices

Abstract Good cell culture practices are a set of technical and management tools recommended for application in research and service laboratories to guarantee the traceability and reproducibility of in vitro experiments. However, most research laboratories do not have a structured quality assurance system and have difficulties organizing their workflows or even priorities in implementing acceptable laboratory practices. In this study, we applied management and quality assurance tools to define the steps necessary to implement acceptable laboratory practices in the multiuser laboratory of cell culture and establish a cell bank at the Carlos Chagas Institute FIOCRUZ-PR. Our team applied the 5W2H and 5S tools for initial diagnosis and established an action plan to implement and manage the laboratory over two years. Thereby, we defined the scope of laboratory activities, including the demand for establishing a cell bank, the supply of cell lines to internal users, user training, and quality control tests. We also mapped the main activities, establishing their flows and all the necessary documentation to ensure traceability and reproducibility. Additionally, the laboratory was organized in compliance with the 5S principles. In conclusion, using management tools, such as the 5W2H and 5S methods, is a simple and affordable method to organize and manage a cell culture laboratory and can be applied to other research laboratories.

Influenza A and B in a cohort of outpatient children and adolescent with influenza like-illness during two consecutive influenza seasons

ABSTRACT Introduction Influenza is an important cause of morbimortality worldwide. Although people at the extremes of age have a greater risk of complications, influenza has been more frequently investigated in the elderly than in children, and inpatients than outpatients. Yearly vaccination with trivalent or quadrivalent vaccines is the main strategy to control influenza. Objectives Determine the clinical and molecular characteristics of influenza A and B infections in children and adolescents with influenza-like illness (ILI). Methods: A cohort of outpatient children and adolescents with ILI was followed for 20 months. Influenza was diagnosed with commercial multiplex PCR platforms. Results: 179 patients had 277 episodes of ILI, being 79 episodes of influenza A and 20 episodes of influenza B. Influenza A and B cases were mild and had similar presentation. Phylogenetic tree of influenza B viruses showed that 91.6% belonged to the B/Yamagata lineage, which is not included in trivalent vaccines. Conclusions: Influenza A and B are often detected in children and adolescents with ILI episodes, with similar and mild presentation in outpatients. The mismatch between the circulating influenza viruses and the trivalent vaccine offered in Brazil may have contributed to the high frequency of influenza A and B in this population.

Genetic variability of populations of the white-eared opossum, Didelphis albiventris Lund 1840 (Didelphimorphia; Didelphidae) in Brazil / Variabilidade genética de populações de gambá de orelha branca, Didelphis albiventris Lund 1840 (Didelphimorphia; Didelphidae) no Brasil

Abstract Didelphis albiventris are found throughout Northeast and Central Brazil to central-southern Uruguay and it was subject of few studies in a population level. Given this, the present study investigated the genetic variability of the species using the mitochondrial molecular marker cytochrome oxidase c subunit I. We analyzed samples from the different biomes within three Brazilian regions: Northeast (Caatinga , Cerrado, and Atlantic Forest), Southeast (Cerrado , Atlantic Forest, Cerrado/Atlantic Forest, and Cerrado/Caatinga ecotones) and South (Pampa and Atlantic Forest). Software BAPs retrieved five distinct demes: dm 1, dm 2, and dm 5 that occurs in South, Northeast and Southeast regions respectively and the dm 3 and dm 4 are wide distributed in Northeast and Southeast. Population analysis performed with AMOVA, haplotype network and Mantel test estimated the veracity of the demes. The FST shows structuring for the five demes, with dm 1 (South region) isolated from the others, however the other analysis showed the Northeast/Southeast demes (dm 2-5) united, diagnosing gene flow between them, mainly at the transitional zones, in areas as far away as areas with similar latitude interval (Southeast vs South) that was not detected gene flow. In the haplotype network, the mutational steps was conclusive in split dm1 from dm 2-5 with 15 mutational steps and the Mantel test was moderated, which is explained by genetic similarity despite the great geographic distances (Northeast/Southeast). Thus, our analysis recognized two different lineages (South and Northeast/Southeast) and indicate that the biomes were not decisive in their isolation. The sharing of demes at the transitional zones and in areas with high latitudinal intervals reflects a recent ancestral polymorphism for D. albiventris. The plasticity in the occupation of the space by this species contributes in its wide dispersion capability, that is, geographical distribution. Our results revealed important implications for the management of D. albiventris in these transitional zones areas where demes were shared.

Resumo Didelphis albiventris é encontrada em todo o Nordeste e região central do Brasil até o centro-sul do Uruguai e foi alvo de poucos estudos em nível populacional. Dessa forma, o presente estudo, investiga a variabilidade genética da espécie usando o marcador molecular citocromo c oxidase subunidade I. Analisou-se amostras de diferentes biomas de três regiões brasileiras: Nordeste (Caatinga, Cerrado e Floresta Atlântica), Sudeste (Cerrado, Floresta Atlântica, ecótonos Cerrado/Floresta Atlântica e Cerrado/Caatinga) e Sul (Pampa e Floresta Atlântica). O software BAPs recuperou cinco demes distintos: dm 1, dm 2 e dm 5, que ocorrem nas regiões Sul, Nordeste e Sudeste, respectivamente, e os dm 3 e dm 4, que são amplamente distribuído no Nordeste e Sudeste. Análises populacionais realizadas com AMOVA, rede de haplótipo e teste de Mantel estimaram a veracidade das demes. O FST mostrou estruturação para as cinco demes, com dm 1 (região Sul) isolada das demais, entretanto as outras análises mostraram as demes Nordeste/Sudeste (dm 2-5) unidos, diagnosticando fluxo gênico entre elas, principalmente em zonas de transição, em áreas tão distante quanto áreas com similar intervalo de latitude (Sudeste e Sul), onde não foram detectado fluxo gênico. Na rede de haplótipo, os passos mutacionais foram conclusivos em separar dm 1 do dm 2-5 com 15 passos mutacionais, e o teste de Mantel foi moderado, o que é explicado pela similaridade genética apesar da grande distância geográfica (Nordeste/Sudeste). Assim, duas linhagens diferentes (Sul e Sudeste/Nordeste) foram encontradas, indicando que os biomas não foram decisivos em seus isolamentos. Os compartilhamentos das demes, em zonas de transição e em áreas com elevados intervalos de latitude, refletem um polimorfismo ancestral recente para D. albiventris. A plasticidade na ocupação do espaço por esta espécie contribui em sua ampla capacidade de dispersão, ou seja, distribuição geográfica. Nossos resultados revelam importantes implicações para o manejo de D. albiventris nessas áreas de zonas de transição, onde as demes são compartilhadas.

Current strategy for local- to global-level molecular epidemiological characterisation of global antimicrobial resistance surveillance system pathogens

The prime goal of molecular epidemiology is to identify the origin and evolution of pathogens, which can potentially influence the public health worldwide. Traditional methods provide limited information which is not sufficient for outbreak investigation and studying transmission dynamics. The recent advancement of next-generation sequencing had a major impact on molecular epidemiological studies. Currently, whole-genome sequencing (WGS) has become the gold standard typing method, especially for clinically significant pathogens. Here, we aimed to describe the application of appropriate molecular typing methods for global antimicrobial resistance surveillance system pathogens based on the level of discrimination and epidemiological settings. This shows that sequence-based methods such as multi-locus sequence typing (MLST) are widely used due to cost-effectiveness and database accessibility. However, WGS is the only method of choice for studying Escherichia coli and Shigella spp. WGS is shown to have higher discrimination than other methods in typing Klebsiella pneumoniae, Acinetobacter baumannii and Salmonella spp. due to its changing accessory genome content. For Gram positives such as Streptococcus pneumoniae, WGS would be preferable to understand the evolution of the strains. Similarly, for Staphylococcus aureus, combination of MLST, staphylococcal protein A or SCCmec typing along with WGS could be the choice for epidemiological typing of hospital- and community-acquired strains. This review highlights that combinations of different typing methods should be used to get complete information since no one standalone method is sufficient to study the varying genome diversity.

Efficient derivation of extended pluripotent stem cells from NOD-scid Il2rg mice

Recently we have established a new culture condition enabling the derivation of extended pluripotent stem (EPS) cells, which, compared to conventional pluripotent stem cells, possess superior developmental potential and germline competence. However, it remains unclear whether this condition permits derivation of EPS cells from mouse strains that are refractory or non-permissive to pluripotent cell establishment. Here, we show that EPS cells can be robustly generated from non-permissive NOD-scid Il2rg mice through de novo derivation from blastocysts. Furthermore, these cells can also be efficiently generated by chemical reprogramming from embryonic NOD-scid Il2rg fibroblasts. NOD-scid Il2rg EPS cells can be expanded for more than 20 passages with genomic stability and can be genetically modified through gene targeting. Notably, these cells contribute to both embryonic and extraembryonic lineages in vivo. More importantly, they can produce chimeras and integrate into the E13.5 genital ridge. Our study demonstrates the feasibility of generating EPS cells from refractory mouse strains, which could potentially be a general strategy for deriving mouse pluripotent cells. The generation of NOD-scid Il2rg EPS cell lines permits sophisticated genetic modification in NOD-scid Il2rg mice, which may greatly advance the optimization of humanized mouse models for biomedical applications.

Methicillin-resistant Staphylococcus aureus isolated from an intensive care unit in Minas Gerais, Brazil, over a six-year period

ABSTRACT To characterize methicillin-resistant Staphylococcus aureus isolates from an intensive care unit of a tertiary-care teaching hospital, between 2005 and 2010. A total of 45 isolates were recovered from patients admitted to the intensive care unit in the study period. Resistance rates higher than 80% were found for clindamycin (100%), erythromycin (100%), levofloxacin (100%), azithromycin (97.7%), rifampin (88.8%), and gentamycin (86.6%). The SCCmec typing revealed that the isolates harbored the types III (66.7%), II (17.8%), IV (4.4%), and I (2.2%). Four (8.9%) isolates carried non-typeable cassettes. Most (66.7%) of the isolates were related to the Brazilian endemic clone from CC8/SCCmec III, which was prevalent (89.3%) between 2005 and 2007, while the USA100/CC5/SCCmec II lineage emerged in 2007 and was more frequent in the last few years. The study showed high rates of antimicrobial resistance among methicillin-resistant S. aureus isolates and the replacement of Brazilian clone, a well-established hospital lineage, by the USA100 in the late 2000s, at the intensive care unit under study.

Molecular variants of Grapevine rupestris stem pitting-associated virus infecting grapevines (Vitis spp.) in Brazil / Variantes moleculares de Grapevine rupestris stem pitting-associated virus infectando videiras (Vitis spp.) no Brasil

ABSTRACT: Grapevine rupestris stem pitting-associated virus (GRSPaV) is one of the most common viruses of grapevine. It is involved in the graft-transmissible disease rupestris stem pitting of the rugose wood complex. The objective of the research was to perform the molecular characterization of the coat protein (CP) gene of sixteen Brazilian GRSPaV isolates aiming to determine the occurrence of molecular variants (strains) of this virus. Nine grapevine samples were evaluated, from which dsRNA was extracted. Nucleotide sequences were generated by Next generation sequencing (NGS). Fifteen complete sequences of the GRSPaV CP gene were obtained and phylogenetically analyzed. Multiple alignments of the sequences showed identities of nucleotides ranging from 82% to 99%, suggesting high variability among the CPs of Brazilian isolates. The study revealed that genetic variability of GRSPaV comprising three molecular variants is also present in Brazilian grapevine genotypes.

RESUMO: O GRSPaV é um dos vírus mais comuns da videira. Está associado à doença transmissível por enxertia denominada caneluras de rupestris que compõe o complexo do lenho rugoso. O objetivo do trabalho foi realizar a caracterização molecular do gene da proteína capsidial (CP) de 16 isolados brasileiros de GRSPaV visando determinar a ocorrência de variantes moleculares desse vírus. Nove amostras de videira foram avaliadas das quais foi extraído dsRNA. As sequências de nucleotídeos foram geradas pelo sequenciamento de nova geração (NGS). Quinze sequências completas do gene CP de GRSPaV foram obtidas e filogeneticamente analisadas. Os alinhamentos múltiplos entre as sequências mostraram identidades de nucleotídeos variando de 82% a 99%, sugerindo alta variabilidade entre as CPs de isolados brasileiros. O estudo revelou que a variabilidade genética de GRSPaV compreendendo três variantes moleculares também está presente nos genótipos de videira no Brasil.

Molecular analyses reveal the occurrence of three new sympatric lineages of velvet worms (Onychophora: Peripatidae) in the eastern Amazon basin

Abstract In the present study, we investigated the possible existence of new lineages of peripatids through comparisons between known Neotropical species and specimens obtained from two locations in Pará, a state in eastern Brazilian Amazonia using a molecular approach based on sequences of the mtDNA genes COI, 16Sr RNA, and 18S RNA. The analyses included also sequences of Asian and African taxa for a more systematic understanding of the phylogenetic relationships within the group. The analysis of the COI, 16S rRNA and 18S RNA sequences permitted the identification of three distinct lineages (A, B and C) based on two different phylogenetic approaches (Bayesian methods and ML). The three lineages presented here are completely distinct from all other peripatid taxa so far defined by molecular data. The presence of specimens of three independent onychophoran lineages occurring in sympatry in the Amazon basin was confirmed in all the analyses, providing consistent support for the phylogenies presented in this study. These findings reinforce the importance of the Amazon region in the diversification of Neotropical peripatids, and indicate that onychophoran diversity is much greater than previously thought, given that the number of taxa found at a single site was equivalent to the total number of allopatric species described for the entire region.

Genetic variability of populations of the white-eared opossum, Didelphis albiventris Lund 1840 (Didelphimorphia; Didelphidae) in Brazil

Abstract Didelphis albiventris are found throughout Northeast and Central Brazil to central-southern Uruguay and it was subject of few studies in a population level. Given this, the present study investigated the genetic variability of the species using the mitochondrial molecular marker cytochrome oxidase c subunit I. We analyzed samples from the different biomes within three Brazilian regions: Northeast (Caatinga , Cerrado, and Atlantic Forest), Southeast (Cerrado , Atlantic Forest, Cerrado/Atlantic Forest, and Cerrado/Caatinga ecotones) and South (Pampa and Atlantic Forest). Software BAPs retrieved five distinct demes: dm 1, dm 2, and dm 5 that occurs in South, Northeast and Southeast regions respectively and the dm 3 and dm 4 are wide distributed in Northeast and Southeast. Population analysis performed with AMOVA, haplotype network and Mantel test estimated the veracity of the demes. The FST shows structuring for the five demes, with dm 1 (South region) isolated from the others, however the other analysis showed the Northeast/Southeast demes (dm 2-5) united, diagnosing gene flow between them, mainly at the transitional zones, in areas as far away as areas with similar latitude interval (Southeast vs South) that was not detected gene flow. In the haplotype network, the mutational steps was conclusive in split dm1 from dm 2-5 with 15 mutational steps and the Mantel test was moderated, which is explained by genetic similarity despite the great geographic distances (Northeast/Southeast). Thus, our analysis recognized two different lineages (South and Northeast/Southeast) and indicate that the biomes were not decisive in their isolation. The sharing of demes at the transitional zones and in areas with high latitudinal intervals reflects a recent ancestral polymorphism for D. albiventris. The plasticity in the occupation of the space by this species contributes in its wide dispersion capability, that is, geographical distribution. Our results revealed important implications for the management of D. albiventris in these transitional zones areas where demes were shared.

Resumo Didelphis albiventris é encontrada em todo o Nordeste e região central do Brasil até o centro-sul do Uruguai e foi alvo de poucos estudos em nível populacional. Dessa forma, o presente estudo, investiga a variabilidade genética da espécie usando o marcador molecular citocromo c oxidase subunidade I. Analisou-se amostras de diferentes biomas de três regiões brasileiras: Nordeste (Caatinga, Cerrado e Floresta Atlântica), Sudeste (Cerrado, Floresta Atlântica, ecótonos Cerrado/Floresta Atlântica e Cerrado/Caatinga) e Sul (Pampa e Floresta Atlântica). O software BAPs recuperou cinco demes distintos: dm 1, dm 2 e dm 5, que ocorrem nas regiões Sul, Nordeste e Sudeste, respectivamente, e os dm 3 e dm 4, que são amplamente distribuído no Nordeste e Sudeste. Análises populacionais realizadas com AMOVA, rede de haplótipo e teste de Mantel estimaram a veracidade das demes. O FST mostrou estruturação para as cinco demes, com dm 1 (região Sul) isolada das demais, entretanto as outras análises mostraram as demes Nordeste/Sudeste (dm 2-5) unidos, diagnosticando fluxo gênico entre elas, principalmente em zonas de transição, em áreas tão distante quanto áreas com similar intervalo de latitude (Sudeste e Sul), onde não foram detectado fluxo gênico. Na rede de haplótipo, os passos mutacionais foram conclusivos em separar dm 1 do dm 2-5 com 15 passos mutacionais, e o teste de Mantel foi moderado, o que é explicado pela similaridade genética apesar da grande distância geográfica (Nordeste/Sudeste). Assim, duas linhagens diferentes (Sul e Sudeste/Nordeste) foram encontradas, indicando que os biomas não foram decisivos em seus isolamentos. Os compartilhamentos das demes, em zonas de transição e em áreas com elevados intervalos de latitude, refletem um polimorfismo ancestral recente para D. albiventris. A plasticidade na ocupação do espaço por esta espécie contribui em sua ampla capacidade de dispersão, ou seja, distribuição geográfica. Nossos resultados revelam importantes implicações para o manejo de D. albiventris nessas áreas de zonas de transição, onde as demes são compartilhadas.

The carriage of the serine-aspartate repeat protein-encodingsdr genes among Staphylococcus aureuslineages

ABSTRACTThe serine-aspartate repeat proteins (Sdr) are members of a family of surface proteins and contribute to the pathogenicity of Staphylococcus aureus. Among 288 S. aureus isolates including 158 and 130 associated with skin and soft tissue infections and bloodstream infection, respectively; 275 (95.5%) were positive for at least one of threesdr genes tested. The positivity rates for sdrC, sdrD, and sdrE among S. aureusisolates were 87.8% (253/288), 63.9% (184/288), and 68.1% (196/288), respectively. 224 (77.8%) of 288 isolates were concomitantly positive for two or three sdr genes. There was an association between carriage ofsdrE and methicillin-resistant S. aureus(MRSA) isolates, while the carriage rates of sdrC andsdrD in MRSA isolates were similar to those in methicillin-sensitive S. aureus (MSSA) isolates. The prevalence of co-existence of sdrC and sdrE among MRSA isolates was significantly higher than that among MSSA isolates (p < 0.05). All ST1, ST5, ST7, and ST25 isolates were positive for sdrD. While all ST121 and ST398 isolates were negative for sdrD. All ST59 and ST88 isolates were positive forsdrE. All ST1 isolates were concomitantly positive forsdrC and sdrD. Concomitant carriage ofsdrC, sdrD, and sdrE was found among all ST5, 75.0% (9/12) of ST1, 69.2% (9/13) of ST6, 78.6% (11/14) of ST25, and 90.9% (20/22) of ST88 isolates. sdrD was linked to CC5, CC7 and CC88 isolates, especially CC88 isolates. There was a strong association between the presence of sdrE and CC59, CC88, and CC5 isolates. A significant correlation between concomitant carriage of sdrC, sdrD, and sdrE and CC88 isolates was found.sdrC-positive, sdrD-positive andsdrE-negative gene profile was significantly associated with CC7 clone. There was an association between sdrC-positive,sdrD-negative, and sdrE-positive gene profile and CC59 isolates. A correlation between sdrC-positive,sdrD-negative, and sdrE-negative gene profile and CC121 clone was found. More CC59 isolates carriedsdrC-negative, sdrD-negative, andsdrE-positive gene profile relative to other four CCs isolates. All ST1 and ST5, 95.2% (20/21) of ST188 and 95.2% (20/21) of ST630 isolates were positive for sdrC. Taken together, our investigation indicated that different S. aureus lineages were associated with specific patterns of carriage of sdr genes.

Detection of influenza B lineages from 2001 to 2013 in a tertiary hospital in the city of São Paulo, Brazil

Two antigenically distinct lineages of influenza B viruses, the Victoria-like and Yamagata-like strains, currently circulate among humans. Surveillance from United States of America and Europe over the last 10 years showed that the chance of a correct matching between vaccine and the circulating lineage had been 50%. We investigated influenza B infection in different patient groups (asymptomatic, general community, with comorbidities and hospitalised) attended at a tertiary hospital in the city of São Paulo, Brazil between 2001-2013. All samples were screened for influenza B virus by one-step real-time reverse transcription-polymerase chain reaction. From 2,992 respiratory samples collected, 114 (3.8%) tested positive for influenza B. Teenagers (13-18 years) presented the highest rate of 18.5% (odds ratio 22.87, 95% confidence interval 2.90-180.66, p < 0.001). One hundred nine samples could be characterised: 50 were Yamagata-like and 59 were Victoria-like strains. Mismatching between the vaccine and predominant circulating strain was observed in 2002 and 2013 seasons. Based on data collected during a period of 12 years, we found that influenza B was more frequent in teenagers. Co-circulation of both lineages and mismatch with the vaccine strain can occur. Our data highlighted the importance of quadrivalent vaccines and future analysis of the age groups included in vaccination programs.

Adaptabilidade e estabilidade fenotipica de genótipos de soja em Porto Alegre do Norte, MT / Fentopic adaptability and stability in soybean genotypes in Porto Alegre do Norte, state of Mato Grosso

Estudos de adaptabilidade e estabilidade são de grande importância para os programas de melhoramento de plantas, tornando possível a identificação de cultivares com comportamento previsível frente às variações ambientais. O objetivo deste trabalho foi avaliar, com base na produtividade de grãos, a adaptabilidade e a estabilidade de genótipos de soja de ciclo tardio, cultivados em três anos consecutivos em Porto Alegre do Norte-MT. Os experimentos foram conduzidos na Fazenda Piraguassu, pertencente ao Grupo Itaquere, no município de Porto Alegre do Norte-MT. O delineamento experimental foi o de blocos completos casualizados com três repetições, envolvendo 25 linhagens e quatro cultivares de soja (BRS Garantia, UFUS Impacta, UFUS Xavante e MSOY 8914). Os métodos de Eberhart e Russel (1966) e Lin Binns modificado por Carneiro (1998) permitiram identificar as linhagens UFU-1 e UFU-14 adaptadas a ambiente favorável, alta estabilidade e elevadas médias de produtividade de grãos. Todas as metodologias propostas foram concordantes em destacar a UFU- 16, devido ao maior desempenho produtivo, à adaptação a ambiente favorável, entretanto com baixa estabilidade.

Studies on adaptability and stability are very important in plant improving programs, because they allow identifying cultivars with predictable behavior upon environmental changes. This search aims to evaluate adaptability and productive stability in soybean genotypes [Glycine max (L.) Merrill], in Porto Alegre do Norte, state of Mato Grosso, Brazil. The experiment were carried out in Paraguassu Farm which belongs to Itaquere Group. The experimental design was a randomized block design with three replications, involving 25 lines and four commercial cultivars of soybean (BRS-Garantia, UFUS Impacta, UFUS Xavante e MSOY 8914). Assays were performed in harvests of 2008/2009, 2009/2010 and 2010/2011 in Porto Alegre do Norte-MT. In order to evaluate adaptability and stability, we used the following methods: Eberhart & Russell (1966), Lin & Binns modified by Carneiro (1998), centroid (ROCHA et al., 2005) and Wricke (1965). All these methodologies agreed in highlighting UFU- 16, due to its best productive performance, adaptation to favorable environment, however, it was considered low stability. According to the methods Eberhart & Russell (1966), Lin & Binns modified by Carneiro (1998), UFU-1 and UFU4 lineages showed high grain productivity average, high stability and were classified as adapted to favorable environment. However, both methodologies Lin & Binns modified by Carneiro (1998) and Eberhart & Russell (1966) agreed in classifying the cultivars concerning to adaptability and stability.

Electroforesis en Gel de Campo Pulsado (PFGE) para la diferenciación molecular de Listeria monocytogenes / Pulsed-field gel electrophoresis (PFGE) for the molecular differentiation of Listeria monocytogenes

En Colombia no es obligatoria la notificación deL. monocytogenes en alimentos, pero se vigilan los alimentos dealto riesgo. Clínicamente se reportan como microorganismoGram-positivo sólo cuando causan meningitis. L. monocytogeneses un patógeno intracelular, transmitido por alimentos, letalpara humanos y animales, que causa Listeriosis; enfermedadque genera varios brotes en el mundo, con pérdidashumanas y económicas. Pocos trabajos en Colombia hanlogrado identificar y serotipificar molecularmente losaislamientos, lo que sólo permite distribuir teóricamentelos serotipos en linajes. Esta revisión se limita a mostrarcaracterísticas del patógeno, su importancia en salud públicay en la industria de alimentos, generalidades de la PFGECHEF;identificando el protocolo estandarizado de trabajoy las enzimas de restricción adecuadas para cortar el ADN.Se encontró que la combinación de enzimas XbaI-AscI,seguida de ApaI es la que ofrece mejores resultados en ladiferenciación de los aislamientos; agrupándolos por linajes;mostrando variaciones intra-serotipo y que en varios paíseslatinoamericanos se analizan los resultados a través dePulseNet, lo que garantiza la comparación de los patronesde PFGE en igualdad de condiciones...

The reporting of L. monocytogenes in food in Colombia is not a mandatory; however, foods consideredhigh-risk are monitored, and the organism is only reported clinically as Gram-positive when it causesmeningitis. L. monocytogenes is a foodborne, intracellular, pathogen which causes listeriosis, a disease lethalto humans and animals. Outbreaks of this disease worldwide can bring about human and economiclosses. Only a few studies in Colombia have been able to identify and molecularly serotype isolatesallowing only the theoretical distribution of serotypes by lineage. This review explains the characteristicsof the pathogen, its importance in public health and in the food industry, and provides an overview ofPFGE-CHEF; identifying the standard work protocol and the appropriate restriction enzymes to cutDNA. We found that the enzyme combination, XbaI-AscI, followed by ApaI offers the best results todifferentiate isolates, by grouping them by lineages, and displaying intra-serotype variations. Additionally,we found that in several Latin American countries the results are analyzed using PulseNet; this ensuresthe comparison of PFGE patterns in equivalent conditions...

Na Colômbia não há uma notificação compulsóriade L. monocytogenes em alimentos, mas alimentos de altorisco são monitorados. Clinicamente, são relatados comoorganismos Gram-positivos apenas quando eles causammeningite. L. monocytogenes é um patógeno intracelularde origem alimentar, letal para seres humanos e animais,que causa a listeriose, que gera surtos em todo o mundo,com perdas humanas e econômicas. Poucos trabalhos naColômbia identificaram e sorotipificaram molecularmenteos isolados, que só permite a distribuição de sorotiposteoricamente em linhagens. Esta avaliação é limitada amostrar características do patógeno, sua importância nasaúde pública e na indústria de alimentos, e uma visãogeral do PFGE-CHEF; identificar o protocolo-padrão detrabalho e enzimas de restrição apropriadas para cortar oADN. Verificou-se que a combinação de enzimas XbaIAscI,seguido por ApaI representa a combinação de enzimasque ofereceu melhores resultados na diferenciação dosisolados, agrupando-a por linhagens, mostrando a variaçãointra-serotipo e que, em muitos países da América Latina,os resultados são analisados através PulseNet, que asseguraa comparação de padrões de PFGE em igualdade decondições...

Expression of CD44 in mouse diethylnitrosamine (DEN)-induced hepatic tumors as well as in DEN-treated embryo stem cells and derived hepatic lineage cells / 동물의과학연구지

To clarify the role of stem cells in hepatocarcinogenesis, CD44 expression was investigated in mouse livers as well as embryonic cell lineages treated with diethylnitrosamine (DEN). Liver tumors induced by DEN were analyzed by immunohistochemisty for CD44. Liver tissues were sampled at 6, 24, and 48 hr after treatment with saline or DEN. Mouse embryonic stem cells (ESCs), hepatic progenitor cells (HPCs), and hepatocyte like cells (HCs), representing 0, 22, and 40 days of differentiation, respectively, were treated with DEN at four doses (0, 1, 5, and 15 mM, respectively) for 24 hr, after which CD44 expression levels were examined by relative quantitative real-time PCR. CD44 expression was weakly detected in tumor cells as well as in some hepatocytes surrounding the tumor cells. However, CD44 expression was not detected in liver tissue treated with DEN at early time points. The CD44 mRNA expression level was significantly different among cells treated with 5 mM DEN at day 22 (P<0.01) as well as 1, 5, and 15 mM DEN at day 40 (P<0.01) compared with control. Taken together, CD44 expression slightly increased in mouse DEN-induced tumors. Furthermore, expression of CD44 in embryonic cell lineages treated with various doses of DEN significantly differed among embryo stem cells and derived hepatic lineage cells. This suggests that CD44 expression may be modulated in the progeny of stem cells during their differentiation toward hepatocytes, and its expression may increase in the tumor stage but not during early carcinogenesis.

Assessing the genetic diversity and ancestry of Hartón del Valle cattle using mitochondrial DNA / Diversidad genética y ancestría del ganado Hartón del Valle mediante ADN mitocondrial / Estimação da ancestralidade e da diversidade genética no gado Hartón del Valle utilizando DNA mitocondrial

The Hartón del Valle(HV)cattleisa Criollo breed of Spanish descent that has adapted to the conditionsof the Cauca valley (Colombia). This breed is currently listed as “vulnerable” because its population has dramatically declined in the last two years. Prompted by the uncertain future of this breed and its importance as a potential resource of valuable genes. Objective: this study addressed the diversity, genetic structure and ancestry of HV cattle using mitochondrial DNA (mtDNA). Methods: a 350-bp fragment was amplified and sequenced from 72 HV animals in 9 separate farms and from animals of Brahman and Holstein breeds. To analyze the breed's ancestry, the sequences were compared with 560 sequences available in the GenBank, representing 50 Bos taurus and Bos indicus breeds. Results: in accordance with the Spanish origin of the HV breed, there was a high representation of European mtDNA (91.7%) and a low representation of African (5.5%) and Middle Eastern mtDNA (2.7%). The average haplotype diversity was 0.65 ± 0.05. The farm with the oldest ancestry was the only population in which three mitochondrial lineages were observed; unfortunately, it was recently depopulated. Proximity was observed between HV and two Columbian breeds, the Romosinuano and Costeño con Cuernos. A comparative analysis with the sequences deposited in the GenBank from numerous breeds revealed the presence of 37 haplotypes, seven of which were unique to HV. Conclusions: the following Iberian breeds were found to be most closely related to HV: Tudanca, Rubia Gallega, Negra Serrana, Murciana, Pajuna, Avileña, Garvonesa and Mertolenga. Phylogenetic analysis confirmed the Iberian ancestry and some African influence on this Latin American Criollo breed.

El Hartón del Valle (HV),es una raza adaptada a las condiciones del Valle del Cauca(Colombia) que está catalogada como “vulnerable” y cuya población ha descendido drásticamente en los últimos dos años. Objetivo: debido al futuro incierto y a la importancia como recurso potencial en la seguridad alimentaria regional se abordó el estudio de la diversidad, la estructura genética y la ancestría del HV, mediante ADN mitocondrial (ADNmt). Métodos: se amplificó y secuenció un fragmento de 350 pb en 72 animales HV, provenientes de nueve poblaciones y de controles de las razas Brahman y Holstein. Para analizar la ancestría las secuencias fueron comparadas con 560 secuencias de 50 razas Bos taurus y Bos indicus, depositadas en el GenBank. Resultados: de acuerdo con su origen español, se encontró una marcada influencia del ADNmt europeo (91.7%) y baja participación de taurinos de origen africano (5.5%) y del cercano Oriente (2.7%). La diversidad haplotípica promedio fue 0.65 ± 0.05. El hato más antiguo fue el único que mostró los tres linajes mitocondriales, sin embargo, este ha sido liquidado recientemente. Se observó proximidad entre el HV con las razas colombianas Romosinuano y Costeño con Cuernos. La comparación con las secuencias depositadas en el GenBank con diferentes razas reveló la presencia de 37 haplotipos, de los cuales siete fueron únicos en el HV. Conclusiones: las razas ibéricas más cercanas al HV fueron: Tudanca, Rubia Gallega, Negra Serrana, Murciana, Pajuna, Avileña, Garvonesa y Mertolenga. El árbol filogenético confirmó la ancestralidad ibérica y la influencia africana en las razas criollas de América Latina.

O Hartón del Valle (HV) é uma raça adaptada às condições da região pertencente ao departamento do Valle del Cauca (Colombia). A raça está catalogada como vulnerável já que sua população tem sido reduzida drasticamente nos últimos dois anos. Objetivo: devido a incerteza no futuro da raça e a sua importancia como recurso potencial na segurança alimentaria regional, abordou-se o estudo da diversidade, a estrutura genética e a ancestralidade do gado HV, utilizando DNA mitocondrial (mtDNA). Métodos: para isto, se amplificou e sequenciou um fragmento de 350 pares de bases em 72 animais provenientes de nove populações e também de controles das raças Brahman e Holandesa. Para analizar a ancestralidade das sequências, estas foram comparadas com outras 560 de 50 raças Bos taurus y Bos indicus depositadas no genebank. Resultados: de acordo com sua origem espanhola, foi encontrada uma marcada influência do DNA mitocondrial europeu (91.7%), e baixa participação de taurinos de origem africana (5.5%) e do Oriente Próximo (2.7%). A média da diversidade haplotípica foi de 0.65 ± 0.05. A população de gado HV mais antiga foi a única que apresentou as três linhagens mitocondriais encontradas; embora, esta população foi terminada recentemente. Observou-se também proximidade do gado HV com as raças colombianas da região Caribe, Romosinuano e Costeño con Cuernos. A comparação com as sequências depositadas no genebank com diferentes raças revelou a presença de 37 haplotipos, dos quais sete foram únicos no HV. Conclusões: as raças ibéricas mais aproximadas do HV foram: Tudanca, Rubia Gallega, Negra Serrana, Murciana, Pajuna, Avileña Garvonesa e Mertolenga. A árvore filogenética confirmou sua ancestralidade ibérica e a influência africana nas raças crioulas da América Latina.

Induction of phagocytic activity and nitric-oxide production in natural populations of Trypanosoma Cruzi I and II from the state of Paraná, Brazil / Indução da atividade fagocitária e produção de óxido nítrico numa população natural de Trypanosoma cruzi I e II do Estado do Paraná, Brasil

Twelve strains of Trypanosoma cruzi isolated from wild reservoirs, triatomines, and chronic chagasic patients in the state of Paraná, southern Brazil, and classified as T. cruzi I and II, were used to test the correlation between genetic and biological diversity. The Phagocytic Index (PI) and nitric-oxide (NO) production in vitro were used as biological parameters. The PI of the T. cruzi I and II strains did not differ significantly, nor did the PI of the T. cruzi strains isolated from humans, triatomines, or wild reservoirs. There was a statistical difference in the inhibition of NO production between T. cruzi I and II and between parasites isolated from humans and the strains isolated from triatomines and wild reservoirs, but there was no correlation between genetics and biology when the strains were analyzed independently of the lineages or hosts from which the strains were isolated. There were significant correlations for Randomly Amplified Polymorphic Deoxyribonucleic acid (RAPD) and biological parameters for T. cruzi I and II, and for humans or wild reservoirs when the lineages or hosts were considered individually.

Doze cepas de Trypanosoma cruzi isoladas de reservatórios silvestres, triatomíneos e de pacientes chagásicos crônicos do Estado do Paraná, Brasil, classificadas como Tc I e II foram usadas para avaliar a correlação entre genética e diversidade biológica. Índice fagocítico (IF) e produção de óxido nítrico (ON) in vitro foram os parâmetros biológicos utilizados. O IF de cepas T. cruzi I e II não diferiram significativamente assim como o IF de cepas isoladas de humanos, triatomíneos ou de reservatórios silvestres. Há diferença estatística na inibição da produção de ON entre T. cruzi I e II e entre parasitos isolados de humanos e de cepas isoladas de triatomíneos e reservatórios silvestres, mas não foi observada correlação entre genética e biologia quando as cepas foram analisadas independentemente da linhagem ou hospedeiros das quais elas foram isoladas. Observou-se correlação significativa para amplificação aleatória do DNA polimórfico e parâmetros biológicos de Tc I ou II e para os seres humanos ou reservatório silvestre quando linhagens ou hospedeiros são consideradas separadamente.

Analysis of paternal lineages in Brazilian and African populations

The present-day Brazilian population is a consequence of the admixture of various peoples of very different origins, namely, Amerindians, Europeans and Africans. The proportion of each genetic contribution is known to be very heterogeneous throughout the country. The aim of the present study was to compare the male lineages present in two distinct Brazilian populations, as well as to evaluate the African contribution to their male genetic substrate. Thus, two Brazilian population samples from Manaus (State of Amazon) and Ribeirão Preto (State of São Paulo) and three African samples from Guinea Bissau, Angola and Mozambique were typed for a set of nine Y chromosome specific STRs. The data were compared with those from African, Amerindian and European populations. By using Y-STR haplotype information, low genetic distances were found between the Manaus and Ribeirão Preto populations, as well as between these and others from Iberia. Likewise, no significant distances were observed between any of the African samples from Angola, Mozambique and Guinea Bissau. Highly significant Rst values were found between both Brazilian samples and all the African and Amerindian populations. The absence of a significant Sub-Saharan African male component resulting from the slave trade, and the low frequency in Amerindian ancestry Y-lineages in the Manaus and Ribeirão Preto population samples are in accordance with the accentuated gender asymmetry in admixture processes that has been systematically reported in colonial South American populations.