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• W2053449919 abstract "Panstrongylus megistus Burmeister 1835 was the first triatomine bug species identified as being responsible for transmitting the agent of Chagas disease. It is still considered a very important vector in the Brazilian Cerrado, a biome consisting of savanna-like grasslands, and attracts the attention of local entomological surveillance agencies. The species has a widespread distribution and is found in tropical forests, the Cerrado and Caatinga, consisting of semi-arid scrubland and thorn forest, in palm trees, rodent and marsupial shelters, tree hollows, and caves (Forattini 1980). In addition, adult P. megistus can colonize anthropogenic ecotopes. The first invasions of anthropogenic landscapes by P. megistus occurred during the Brazilian post-colonial period as a result of the major impact of agricultural development on the natural habitat of this insect (Miles 1976, Dórea at al. 1982). The degree of synanthropy exhibited by P. megistus varies among regions: in Argentina, Bolivia, Paraguay, and Uruguay (Lent and Wygodzinsky 1979) it is exclusively non-synanthropic; from the State of Rio Grande do Sul (in the south of Brazil) to the midwest of the state of São Paulo it is hemi-synanthropic; and in the state of Bahia and the midwest of the state of Minas Gerais it is eusynanthropic (Barbosa et al. 2004). Some authors relate this variation to physiological and ethological differences in P. megistus populations (Dórea et al. 1982, Barbosa et al. 2003, 2004, 2006, Kopp et al. 2009). Molecular markers have proven useful in understanding population genetics and in evolutionary studies of P. megistus and other triatomine species (Barbosa et al. 2003, 2006, Abad-Franch and Monteiro 2005). The mitochondrial gene cytochrome B oxidase (cytb) has provided useful information for investigations into the biogeography and evolutionary rate of triatomine species (Bargues et al. 2002, Mas-Coma and Bargues 2009). In the present study, we analyzed partial cytb sequences of twelve Brazilian P. megistus populations to infer their phylogeographic relationships. P. megistus specimens were obtained from colonies in the Triatominae Insectarium in the Department of Biological Sciences, Faculty of Pharmaceutical Sciences, UNESP, Araraquara, São Paulo, Brazil. The number of generations of each colony was estimated based on a life cycle of 608 days/generation in accordance with Heitzmann-Fontenelle (1980) (Table 1). The colonies were originally collected from the locations shown in Figure 1. Genomic DNA was extracted from individual samples (n=10 for each colony) using the DNeasy Blood and Tissue Kit (Qiagen, Crawley, United Kingdom). A 341 bp fragment of the mitochondrial cytb gene was amplified as described by Lyman et al. (1999) and sequenced using ABI Prism BigDye Terminator Ready Reaction Kit (Perkin Elmer, Foster City, CA) on an ABI/Hitachi PRISM 3100 Genetic Analyzer. The sequences were deposited in the GenBank/EMBL Data Bank, and their accession numbers are shown in Table 1. Because of the high similarity between the cytb sequences found in each colony, the phylogenetic trees were constructed using consensus sequences for each population. Phylogenetic reconstructions were performed with the Neighbor Joining (NJ) algorithm with Kimura 2-parameter distance correction and 1,000 bootstrap replications and with Maximum Likelihood (ML) and the Tamura-Nei distance model with 1,000 bootstrap replications using MEGA 5.0 software (Tamura et al. 2011). A cytb sequence of P. megistus (AF045722) from a colony maintained by the Oswaldo Cruz Institute (Rio de Janeiro, Brazil) for which no information about the origin was available was included in this analysis. Haplotypes were identified using DnaSP v. 5.0 (Librado and Rozas 2009). All phylogenies were rooted with cytb sequences from Triatoma dimidiata (Latreille, 1811) (AF045726) and Rhodnius neglectus Lent, 1954 (AF045716) from Genbank. The phylogenies inferred from the cytb data set used a T92 nucleotide substitution model, the best-fit model based on the lowest Akaike Information Criterion (AIC) score inferred from jModelTest (Posada 2008). The level of genetic structuring was estimated by means of different hierarchical analyses of molecular variance (AMOVA) with Arlequin version 3.5.1.2 (Excoffier and Lischer 2010) The phylogenetic trees obtained by the NJ and ML methods had similar topologies (the ML tree is shown in Figure 2). Sequence divergence ranged from less than 1.0% to almost 3.0 % between the populations, which clustered into two groups. The first group, composed of samples from the states of São Paulo, Minas Gerais, Goiás, Alagoas, and Rio Grande do Norte, showed about 3.0% divergence (Figure 2A), whereas the other samples from the states of Paraná, São Paulo, and Bahia (Figure 2B) exhibited less than 1.0% genetic difference. The same result was observed using AMOVA, which failed to show any significant difference between the groups. Variance was 67.4% among groups and 32.6% among populations within a group. Genetic variability across populations was estimated. Haplotype diversity (Hd) was 0.897 ± 0.067, and eight different haplotypes were found, as shown in the ML tree. Five different haplotypes were found in São Paulo (H2, H3, H4, H6, H7), one of which was shared with the Minas Gerais population (H6) and another of which was shared with the Paraná population (H2). Alagoas and Rio Grande do Norte shared the same haplotype (H5). There was only one haplotype for Bahia (H1) (Figure 2). According to Mas-Coma and Bargues (2009), the genetic distances between species of genus Panstrongylus can vary between 27% and 33% for nuclear genes. Analysis based on the cytb marker to determine genetic distances between triatomines from the same species but different geographic regions indicates that there must be more than 6% difference between them if they are to be considered subspecies. Therefore, the distance between the two main clusters observed in this study corresponds to a within-species polymorphism. Based on these results, we hypothesize that P. megistus disperses passively to different regions of Brazil and that São Paulo is the center where the various populations converge. This distribution reflects the human migrations from rural areas of Brazil to more economically developed urban centers that occurred mainly in the second half of the 20th century, when the state of São Paulo was the primary destination for millions of Brazilians, especially those from the northeast of the country (Brito and Carvalho 2006). The hypothesis of passive dispersal of P. megistus is reinforced by a study based on isoenzyme analysis which showed relationships between populations in Paraná (in the south of Brazil) and in the northeast and southeast of the country (Kopp et al. 2009). A previous study based on morphological analysis of eggs separated P. megistus populations into two groups, one comprising the population in Bahia (northeast Brazil) and the other formed by populations from Santa Catarina and Minas Gerais (south and southeast Brazil) (Barbosa et al. 2003). In the same study, morphometric analysis of the head and thorax of P. megistus showed that southern populations were different from midwestern populations. These differences were also noted when salivary gland proteins were analyzed and are probably due to differences in feeding habits (Barbosa et al. 2006). Here we included other populations from northeast Brazil and different locations in the state of São Paulo. Our results, which agree with those reported by Barbosa et al. (2006), show that the clades formed two distinct groups: the cluster formed by populations from Alagoas (145) and Rio Grande do Norte (145) (in the northeast) together with populations in the southeast, attributed by Barbosa et al. to the characteristics of the vegetation in these regions (Atlantic Forest), and the cluster formed by populations from Bahia (northeast) and populations from Minas Gerais and São Paulo (southeast), regions where vegetation typical of the Cerrado is found. These associations suggest the existence of sympatric populations in the past, favoring a certain degree of similarity between their genetic profiles (Barbosa et al. 2006). Our analysis highlights the importance of the cytb gene as a molecular marker in genetic population, systematics, and taxonomy studies of Triatominae species. However, the results obtained here may have been affected by the genetic homogeneity of the colonies. This homogeneity is caused by the founder effect, which results from inbreeding of populations in colonies or human dwellings, leading to reduced gene polymorphism compared with natural populations (Schofield et al. 1999). According to Heitzmann-Fontenelle (1980), inbreeding weakens the genetic vigor of triatomines, extending the nymph phase, shortening its adult life, and reducing the number of eggs, egg laying, and egg viability. Although colonies can lead to inbreeding and, consequently, low diversity among individuals in the same colony, they can increase genetic isolation between individuals from distinct populations (Fredrickson et al. 2007). In a study by Gómez-Sucerquia et al. (2009), it was shown that the structure of Rhodnius pallescens populations from colonies was very different from those of field populations. However, the results reported by Gómez-Sucerquia et al. (2009) were obtained using microsatellite markers, while our study used a molecular marker, which implies slow evolutionary rates. We therefore assumed that the number of generations of our samples (between 7 and 15) was not enough to generate significant differences between colonies and field populations. Although there are serious concerns regarding the use of colonies and the limited polymorphism associated with them, they are important in research studies as they allow comparative analyses between recently collected populations and populations originated from P. megistus species that have been eradicated in many parts of Brazil. Further studies using field populations of P. megistus are required to provide a more realistic picture of phylogeographic relationships within this species. This study was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP). We thank the Department of Biological Sciences of the Faculty of Pharmaceutical Sciences, UNESP, for providing the specimens. DPV received a Master's scholarship from FAPESP (2010/02960–3)." @default.
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• W2053449919 title "Analysis of mitochondrial cytochrome B oxidase gene suggests passive dispersal of the <i>Panstrongylus megistus</i> population in Brazil" @default.
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