Paleogenetics paleoepidemiology and Ascaris sp. (Linnaeus, 1758) and Trichuris sp. (Roederer, 1761)

Abstract

Ascaris lumbricoides e Trichuris trichiura s??o os helmintos intestinais de maior preval??ncia na popula????o mundial e tamb??m no material arqueol??gico. Por??m, na Am??rica do Sul pr??-colombiana, o encontro de ovos de A. lumbricoides ?? raro. Recentemente um estudo de diagn??stico paleoparasitol??gico molecular apontou para um sub-diagn??stico de Ascaris sp. na Am??rica do Sul. No registro arqueol??gico de parasitos intestinais predominam achados de ovos de Trichuris sp. ao inv??s de Ascaris sp. Isto parece contradit??rio, em virtude do n??mero de ovos eliminados por cada parasito. Os objetivos desta pesquisa foram: avaliar marcadores moleculares para o diagn??stico de Ascaris sp. e Trichuris sp. em material moderno pela caracteriza????o molecular destes parasitos; caracterizar geneticamente isolados de s??tios arqueol??gicos sul americanos para verificar a real paleodistribui????o destes parasitos em uma perspectiva paleoepidemiol??gica e compar??-la com a epidemiologia moderna destas infec????es; avaliar os fatores envolvidos na paleodistribui????o encontrada. As amostras foram submetidas ao diagn??stico por microscopia ??ptica, seguida da extra????o do DNA, PCR e sequenciamento nucleot??dico. Na avalia????o dos marcadores moleculares, a regi??o ITS1 de Ascaris sp. apresentou varia????o intra-indiv??duo, o que descartou seu uso com fins taxon??micos e diagn??sticos. A caracteriza????o molecular dos genes mitocondriais cox1 e nad1 de Ascaris sp. mostrou infec????o cruzada de gen??tipos entre as esp??cies humana e su??na, o que denota a necessidade de monitoramento das popula????es avaliadas assim como de outras regi??es brasileiras para que a infec????o n??o venha a se tornar uma zoonose em potencial no Brasil. Foi poss??vel o diagn??stico molecular de Trichuris sp. pelo gene ribossomal 18S DNA. A an??lise paleogen??tica mostrou que h?? subdiagn??stico para ambas as infec????es na Am??rica do Sul pr??-colombiana. Este ?? o primeiro diagn??stico paleoparasitol??gico molecular de T. trichiura em material sul americano. Estes s??o tamb??m os primeiros registros de recupera????o de DNA de parasitos intestinais em material de s??tio arqueol??gico do tipo ???sambaqui??? e tamb??m do per??odo colonial brasileiro. Comparando-se a paleoepidemiologia molecular de Ascaris sp. com a epidemiologia molecular moderna foi poss??vel notar que h?? hapl??tipos antigos que ainda est??o presentes hoje, no entanto a maioria dos hapl??tipos ?? caracter??stica ao material arqueol??gico. Observou-se que h?? hapl??tipos comuns ao Velho e Novo Mundo, contudo, h?? tamb??m especificidades regionais. Os resultados da an??lise gen??tica claramente apontam para uma pobre preserva????o dos ovos no material arqueol??gico, principalmente de Ascaris sp. Os fatores principais envolvidos nessa paleodistribui????o, seriam fatores tafon??micos que proporcionaram a quebra maior de ovos de Ascaris sp. do que de Trichuris sp., e evid??ncias de consumo de plantas verm??fugas pelos povos pr??-hist??ricos, as quais teriam maior a????o sobre Ascaris sp. do que Trichuris sp.Ascaris lumbricoides and Trichuris trichiura are the intestinal helminths with higher prevalence in the world today as it was in the past. However, in pre-Columbian South America the findings of A. lumbricoides eggs are rare. Recently a study of paleoparasitological molecular diagnosis showed a sub-diagnosis of Ascaris sp. in South America. In the archeological material, eggs of Trichuris sp. are more common compared with Ascaris sp. eggs. This is contradictory taking into account the number of eggs eliminated by each parasite. The aims of this research was: to evaluate molecular markers for Ascaris sp. and Trichuris sp. diagnosis in modern material; genetic characterization of the samples South American archeological sites aiming the paleodistribution of these parasites in a paleoepidemiological perspective; compare results with the modern epidemiology of these infections; evaluate the factors involved in paleodistribution. Extraction of DNA, PCR and nucleotide sequencing were performed after microscopy. In the evaluation of the molecular markers Ascaris sp. ITS1 region showed intra-individual variation. Therefore, this region to taxonomical and diagnoses studies was discarded. With the molecular characterization of Ascaris sp. cox1 and nad1 mitochondrial genes it was possible to identify cross infection of genotypes between human and pig hosts. Results showed that surveillance field works in modern populations are necessary to verify the zoonotic potential of this infection in Brazil. The molecular diagnosis of Trichuris sp. by ribossomal 18S DNA gene was possible. The paleogenetic analysis showed that there is subdiagnosis for both infections in pre-Columbian South America. This is the first paleoparasitological molecular record of T. trichiura in South American samples. These are also the first recovery of DNA of intestinal parasites in "sambaqui" archeological site, and also of the Brazilian colonial period. Molecular paleoepidemiology of Ascaris sp. infection compared with modern molecular epidemiology showed that there are ancient haplotypes still present today. However, most of the haplotypes are characteristic of the archaeological material. It was observed that there are common haplotypes both to the Old World and to the New World, but showing regional specificities. The results of the genetic analysis clearly pointed to a poor preservation of eggs in archeological material, mainly of Ascaris sp. Taphonomy may be the main factor involved in paleodistribution, breaking more eggs of Ascaris sp. than Trichuris sp. Evidences of consumption of vermifuge plants by prehistoric groups should also have influence, as some plants should have more efficacy eliminating Ascaris sp. than Trichuris sp