Chapters authored
Nucleic Acid-based Diagnosis and Epidemiology of Infectious Diseases
In this chapter, the immense contribution of nucleic acid discovery to the diagnosis and molecular epidemiology of pathogenic microorganisms and its relevance for veterinary and human health will be discussed. The development of nucleic acid detection, amplification, and sequencing techniques, principally after the introduction of polymerase chain reaction (PCR), allowed the improvement of different strategies to diagnose and to quantify infectious microbiological agents in a variety of organisms and biological samples. Pos-PCR associated techniques such as fragment enzyme restriction and sequence analysis permit the determination of nucleic acid sequence diversity to detect drug resistance, to associate pathogen genetic markers with disease outcome, and to predict temporal and spatial distribution of microorganisms which can be used to prevent and treat infectious diseases efficiently.
Part of the book: Nucleic Acids
Novel Single Hematophagous Insect RNA Detection Method Supports Its Use as Sentinels to Survey Flaviviruses Circulation
Anthropogenic actions, including deforestation, disorganized urbanization, and globalization, contribute to emergence and reemergence of arboviruses worldwide, where Flavivirus is the most prevalent, and its continuous monitoring can help in preventive control strategies. Thus, the aim of this study was to detect flavivirus RNA in single hematophagous insects, which are used as sentinels. Total RNA was extracted from six Aedes aegypti stored since 2003 and from 100 Culicidae and collected through CDC trap in a public park of a Brazilian Northwest city of São Paulo State. Flavivirus was detected through RT/PCR targeting 230–250 bp of the RNA polymerase coding sequence (NS5). PCR amplicons were sequenced by Sanger method, used in comparative analysis over Basic Local Alignment Search Tool (BLAST) in GenBank, and subjected to Neighbor-Joining phylogenetic analyses. Efficiency of Flavivirus diagnosis was confirmed by detection of Dengue virus serotype 2 in Ae. aegypti. From the 100 collected insects, 19 were positive for Culex flavivirus (CxFV). NS5 partial sequence phylogenetic analysis clustered all CxFV in one branch separated from vertebrate flaviviruses, being applicable to the identification of Flavivirus species. The dipteran RNA extraction methodology described in this work supports detection of flaviviruses in single insects maintained in 80% ethanol, which can be used to constant arbovirus surveillance.
Part of the book: Dengue Fever in a One Health Perspective