Discovery and Barcoding by Analysis of Spliced Leader RNA Gene Sequences of New Isolates of Trypanosomatidae from Heteroptera in Costa Rica and Ecuador

Abstract

Trypanosomatid diversity in Heteroptera was sampled using a culture-independent approach based on amplification and sequencing of Spliced Leader RNA gene repeats from environmental samples. By combining the data collected herein with that of previous work, the prevalence of parasites was found to be 22%-23%. Out of approximately 170 host species investigated nearly 60 were found to harbor trypanosomatids. The parasites found were grouped by cluster analysis into 48 typing units. Most of these were well separated from the known groups and, therefore, likely represent new trypanosomatid species. The sequences for each typing unit serve as barcodes to facilitate their recognition in the future. As the sampled host species represent a minor fraction of potential hosts, the entire trypanosomatid diversity is far greater than described thus far. Investigations of trypanosomatid diversity, host-specificity, and biogeography have become feasible using the approach described herein.

Selective recovery of the cultivation-prone components from mixed trypanosomatid infections: a case of several novel species isolated from Neotropical Heteroptera

“…Cultivation problems were encountered with Leptomonas jaculum, which, in fact, is phylogenetically close to Blastocrithidia triatomae (Kostygov & Frolov, 2007), also a poorly cultivable species (Peng & Wallace, 1981;Reduth et al, 1989). In recent surveys of trypanosomatid diversity, most of the genotyped organisms that were closely related to the reference Blastocrithidia species were also refractory to axenic cultivation (Maslov et al, 2007). Yet, the same problem was also encountered for other trypanosomatid groups analysed, so it is not generally restricted to a particular group or clade (D. A. Maslov, data not shown).…”

“…We have developed a methodology to analyse spliced leader RNA (SL RNA) gene sequences directly from parasites in insect gut samples (Westenberger et al, 2004;Maslov et al, 2007). Such 'environmental' trypanosomatid sequences were obtained from 60 of 170 species of Neotropical Heteroptera investigated, a number representing a relatively small fraction of all potential hosts.…”

Spliced Leader (SL) RNA: análises de genes e regiões intergênicas com aportes na filogenia, taxonomia e genotipagem de <i>Trypanosoma</i> spp. de todas as classes de vertebrados.

“…Abordagens baseadas no SL têm sido avaliadas em uma série de estudos que analisam a diversidade e as relações hospedeiro-parasito de tripanossomatídeos de insetos Heteroptera (Maslov et al, 2007(Maslov et al, , 2013Yurchenko et al, 2009;Votypka et al, 2010Votypka et al, , 2012 e Diptera (Tyc et al, 2013).…”

“…Com o conhecimento gerado por diversos estudos de que cada espécie de tripanossomatídeo pode ser reconhecida pela respectiva sequência do gene SL (Campbell et al, 1997;Fernandes et al, 1997;Serrano et al, 1999;Merzlyak et al, 2001), esse gene passou a ser utilizado para "DNA barcoding", permitindo distinguir espécies de todos os gêneros, inclusive em amostras de tubo digestivo de insetos, sem a necessidade de cultivo dos parasitas (Maslov et al, 2007). Igualmente, este marcador tem sido utilizado para determinar TUs para identificar novas espécies potenciais de tripanossomatídeos (Maslov et al, 2007(Maslov et al, , 2013Yurchenko et al, 2009;Votypka et al, 2010Votypka et al, , 2012, assim como para confirmar as espécies novas de Trypanosoma (Lima et al, 2013(Lima et al, , 2015a.…”

“…Igualmente, este marcador tem sido utilizado para determinar TUs para identificar novas espécies potenciais de tripanossomatídeos (Maslov et al, 2007(Maslov et al, , 2013Yurchenko et al, 2009;Votypka et al, 2010Votypka et al, , 2012, assim como para confirmar as espécies novas de Trypanosoma (Lima et al, 2013(Lima et al, , 2015a.…”