Enterobius vermicularis: ancient DNA from North and South American human coprolites

A molecular paleoparasitological diagnostic approach was developed for Enterobius vermicularis. Ancient DNA was extracted from 27 coprolites from archaeological sites in Chile and USA. Enzymatic amplification of human mtDNA sequences confirmed the human origin. We designed primers specific to the E. vermicularis 5S ribosomal RNA spacer region and they allowed reproducible polymerase chain reaction identification of ancient material. We suggested that the paleoparasitological microscopic identification could accompany molecular diagnosis, which also opens the possibility of sequence analysis to understand parasite-host evolution.     

The finding of Enterobius vermicularis eggs in pre-Columbian human coprolites

Enterobius vermicularis eggs were found in human coprolites collected in the archaeological site of Caserones, Tarapaca Valley, Chile, dating from 400 BC to 800 AD. The human pinworm had already been found in other pre-historic archaeological sites in America, and its introduction in this continent is discussed.     

Temporal and Spatial Distribution of Enterobius vermicularis (Nematoda: Oxyuridae) in the Prehistoric Americas

Investigations of Enterobius sp. infection in prehistory have produced a body of data that can be used to evaluate the geographic distribution of infection through time in the Americas. Regional variations in prevalence are evident. In North America, 119 pinworm positive samples were found in 1,112 samples from 28 sites with a prevalence of 10.7%. Almost all of the positive samples came from agricultural sites. From Brazil, 0 pinworm positive samples were found in 325 samples from 7 sites. For the Andes region, 22 pinworm positive samples were found in 411 samples from 26 sites for a prevalence of 5.3%. Detailed analyses of these data defined several trends. First, preagricultural sites less frequently show evidence of infection compared to agricultural populations. This is especially clear in the data from North America, but is also evident in the data from South America. Second, there is an apparent relationship between the commonality of pinworms in coprolites and the manner of constructing villages. These analyses show that ancient parasitism has substantial value in documenting the range of human behaviors that influence parasitic infections.     

First study of fossil rodent middens as source of paleoparasitological evidences (northwestern Patagonia,Argentina)

The present paper reports the first paleoparasitological results obtained from coprolites of fossil rodent middens and demonstrates the potential of rodent middens as a source of paleoparasitological evidences in South America. Ten fossil rodent middens from northwestern Patagonia, Argentina, were studied. Five coprolites of each midden were fully processed, rehydrated, homogenized, subjected to spontaneous sedimentation, and examined through light microscopy. Eight of the 10 examined rodent middens contained parasite eggs. The eggs of parasites were assigned to Heteroxynema (Cavioxyura) viscaciae Sutton & Hugot, 1989 and Helminthoxys sp. (Nematoda: Oxyuridae), Trichuris sp. (Nematoda: Trichuridae) and one unidentified nematode. Fossil rodent middens were assigned to Lagidium viscacia (Caviomorph: Chinchillidae). The excellent preservation of parasite remains in coprolites from fossil rodent middens provided an opportunity to perform paleoparasitological inferences. The results of this papers demonstrates that fossil rodent middens offer an excellent opportunity for the recovery of parasite remains for future paleoparasitological studies in the southwest of South America.     

Molecular paleoparasitological diagnosis of Ascaris sp. from coprolites: new scenery of ascariasis in pre-Colombian South America times

Paleoparasitological studies using microscopy showed that Ascarisand Trichuris trichiura are the human intestinal parasites most found in archaeological sites. However, in pre-Columbian South American archaeological sites, Ascaris is rare. In this work we standardized a molecular methodology for Ascaris diagnosis directly from ancient DNA retrieved from coprolites. Using cythochrome b gene (142 bp) target, ancient DNA sequences were retrieved from South American samples, negative by microscopy. Moreover, the methodology applied was sensitive enough to detect ancient DNA extracted from 30 Ascaris eggs from an European coprolite. These results revealed a new scenery for the paleodistribution of Ascaris in South America.     

The finding of Echinostoma (Trematoda: Digenea) and hookworm eggs in coprolites collected from a Brazilian mummified body dated 600-1,200 years before present

The identification of parasites from ancient cultures expands our list of parasites infective to extant humans. A partially mummified human body from the archeological site of Lapa do Boquete, Minas Gerais State, Brazil, was recently discovered. It was interred between 600 and 1,200 yr ago. Dietary analysis showed that the mummified body was from a society that had a mixed subsistence of agriculture and gathering of wild foods. Coprolites from the body contained numerous helminth eggs. The eggs were identified as those of Echinostoma sp. and hookworm. Hookworm infection in pre-Columbian populations is already established, but this is the first evidence of Echinostoma sp. eggs found in human coprolites. The diagnosis of a true infection, as opposed to false parasitism, is discussed. The possibility of Echinostoma ilocanum infection is discussed, as this is a common species found in humans in the Asiatic region, which could have been introduced in South America in the pre-Columbian period. Alternative possibilities are also considered, including indigenous Brazilian Echinostoma species.     

Enterobius vermicularis eggs discovered in coprolites from a medieval Korean mummy

While the presence of pinworm eggs in archaeological samples has been reported by many researchers in the New World, those have been detected very scarcely in the Old World, especially in East Asian countries. In fact, many parasite species were recovered from the archeological remains in Korea, eggs of Enterobius vermicularis had not been found. Recently, a female mummy buried in the 17th century was discovered in the Joseon tomb from Dangjin-gun, Chungcheongnam-do, Korea. After rehydration process for 12 days, investigations were carried on the luminal surface of the colon. From them, 3 eggs of E. vermicularis were recovered. They were elliptical, transparent with a thin egg shell, 50.3±5.2 μm (length) and 28.2±3.9 μm (width) in size. This is the first discovery of E. vermicularis eggs in East Asia.     

Parasites as probes for prehistoric human migrations?

Host-specific parasites of humans are used to track ancient migrations. Based on archaeoparasitology, it is clear that humans entered the New World at least twice in ancient times. The archaeoparasitology of some intestinal parasites in the New World points to migration routes other than the Bering Land Bridge. Helminths have been found in mummies and coprolites in North and South America. Hookworms (Necator and Ancylostoma), whipworms (Trichuris trichiura) and other helminths require specific conditions for life-cycle completion. They could not survive in the cold climate of the northern region of the Americas. Therefore, humans would have lost some intestinal parasites while crossing Beringia. Evidence is provided here from published data of pre-Columbian sites for the peopling of the Americas through trans-oceanic or costal migrations.     

Paleoparasitological analysis of coprolites from K2, an Iron Age archaeological site in South Africa: the first finding of Dicrocoelium sp. eggs

Until now, Dicrocoelium sp. eggs have only been recorded from European and 1 North American archaeological sites. We present evidence for the first record of Dicrocoelium sp. from an African archaeological site. A paleoparasitological study was conducted on 7 coprolite samples from K2, a Late Iron Age site on the farm Greefswald, in the Northern Province of South Africa. Standard parasitological analysis revealed the presence of Dicrocoelium sp. and Trichuris sp. eggs. Today, the parasite does not occur in this region. Trichurid eggs are a relatively common find in paleoparasitological analysis. The presence of Dicrocoelium sp. provides new clues about the antiquity of this parasite, as well as aspects of ancient environment, climate, and interactions among humans, animals, and parasites.     

Analysis of ancient DNA from coprolites: a perspective with random amplified polymorphic DNA-polymerase chain reaction approach

The aim of this work was to determine approaches that would improve the quality of ancient DNA (aDNA) present in coprolites to enhance the possibility of success in retrieving specific sequence targets. We worked with coprolites from South American archaeological sites in Brazil and Chile dating up to 7,000 years ago. Using established protocols for aDNA extraction we obtained samples showing high degradation as usually happens with this kind of material. The reconstructive polymerization pretreatment was essential to overcome the DNA degradation and the serial dilutions helped with to prevent polymerase chain reaction (PCR) inhibitors. Moreover, the random amplified polymorphic DNA-PCR has been shown to be a reliable technique for further experiments to recover specific aDNA sequences.     

Human intestinal parasites in the past: new findings and a review

Almost all known human specific parasites have been found in ancient feces. A review of the paleoparasitological helminth and intestinal protozoa findings available in the literature is presented. We also report the new paleoparasitologic findings from the examination performed in samples collected in New and Old World archaeological sites. New finds of ancylostomid, Ascaris lumbricoides, Trichuris trichiura, Enterobius vermicularis, Trichostrongylus spp., Diphyllobothrium latum, Hymenolepis nana and Acantocephalan eggs are reported. According to the findings, it is probable that A. lumbricoides was originally a human parasite. Human ancylostomids, A. lumbricoides and T. trichiura, found in the New World in pre-Columbian times, have not been introduced into the Americas by land via Beringia. These parasites could not supported the cold climate of the region. Nomadic prehistoric humans that have crossed the Bering Land Bridge from Asia to the Americas in the last glaciation, probably during generations, would have lost these parasites, which life cycles need warm temperatures in the soil to be transmitted from host to host. Alternative routes are discussed for human parasite introduction into the Americas.     

Scanning electron microscope study of ancient parasite eggs recovered from Korean mummies of the Joseon Dynasty

We have previously shown that parasite eggs have been identified in the coprolites of Korean mummies. These eggs have shed light on parasitic infection patterns in Korean populations living several hundred years ago. We conducted a scanning electron microscopy (SEM) study on ancient Trichuris trichiura, Ascaris lumbricoides, Metagonimus yokogawai, Paragonimus westermani, and Gymnophalloides seoi eggs recovered from Korean mummies of the Joseon Dynasty. We anticipated that the taphonomic conditions of mummification would alter the eggs of certain species but not of others. Our SEM data show that each species of ancient egg exhibited different degrees of preservation. Thus, some of them, for example, M. yokogawai, exhibited a better preservation status than others, suggesting that they should be the first candidates considered when choosing subjects for future paleoparasitological studies.     

Syphacia sp. (Nematoda: Oxyuridae) in coprolites of Kerodon rupestris Wied, 1820 (Rodentia: Caviidae) from 5,300 years BP in northeastern Brazil

We present the results of paleoparasitological analyses in coprolites of Kerodon rupestris, rodent endemic to rocky areas of Brazil's semiarid region. The coprolites were collected from excavations at the archaeological site of Toca dos Coqueiros, in the National Park of Serra da Capivara, southeastern of state of Piauí. Syphacia sp. (Nematoda: Oxyuridae) eggs were identified in coprolites dated at 5,300 ± 50 years before present. This is the first record of the genus Syphacia in rodent coprolites in the Americas.     

Egg positive rate of Enterobius vermicularis and Taenia spp. by cellophane tape method in primary school children in Sivas, Turkey

The aim of the present study was to find out the number of students with enterobiasis and/or taeniasis in primary schools of Sivas. Among the 2,029 students in 6 primary schools, 316 (15.6%) were positive to Enterobius vermicularis eggs and 32 (1.6%) were positive to Taenia spp. eggs by the cellophane tape method. The egg positive rates of E. vermicularis and Taenia spp. ranged from 9.4% to 27.2% and from 0.8% to 2.6% respectively among six schools. The egg positive rate of E. vermicularis was found to be significantly different among these schools (chi2 = 31.96, P < 0.05), whereas there was no significant difference between the schools for Taenia spp. (chi2 = 4.37; P > 0.05). The rate (18.7%) of E. vermicularis in the urban slum regions was higher than the rate (11.5%) in the urban central regions (chi2 = 19.20; P < 0.05). Above results demonstrate that the egg positive rate of E. vermicularis and Taenia spp. was still prevalent among primary school children.     

Trichuris sp. from 1,040 +/- 50-year-old Cervidae coprolites from the archaeological site Furna do Estrago, Pernambuco, Brazil

We present results of the paleoparasitological analysis of Cervidae coprolites that were recovered from the archaeological site Furna do Estrago, Pernambuco, Brazil. Trichuris sp. eggs were recovered from the coprolite samples dated 1,040 ± 50 years before present. This is the first record of Trichuris sp. in semiarid Cervidae, unexpectedly recorded in archaeological material.     

Pre-Columbian origins of Native American dog breeds,with only limited replacement by European dogs,confirmed by mtDNA analysis

Dogs were present in pre-Columbian America, presumably brought by early human migrants from Asia. Studies of free-ranging village/street dogs have indicated almost total replacement of these original dogs by European dogs, but the extent to which Arctic, North and South American breeds are descendants of the original population remains to be assessed. Using a comprehensive phylogeographic analysis, we traced the origin of the mitochondrial DNA lineages for Inuit, Eskimo and Greenland dogs, Alaskan Malamute, Chihuahua, xoloitzcuintli and perro sín pelo del Peru, by comparing to extensive samples of East Asian (n = 984) and European dogs (n = 639), and previously published pre-Columbian sequences. Evidence for a pre-Columbian origin was found for all these breeds, except Alaskan Malamute for which results were ambigous. No European influence was indicated for the Arctic breeds Inuit, Eskimo and Greenland dog, and North/South American breeds had at most 30% European female lineages, suggesting marginal replacement by European dogs. Genetic continuity through time was shown by the sharing of a unique haplotype between the Mexican breed Chihuahua and ancient Mexican samples. We also analysed free-ranging dogs, confirming limited pre-Columbian ancestry overall, but also identifying pockets of remaining populations with high proportion of indigenous ancestry, and we provide the first DNA-based evidence that the Carolina dog, a free-ranging population in the USA, may have an ancient Asian origin.     

Parasitism of prehistoric humans and companion animals from Antelope Cave, Mojave County, northwest Arizona

Previously, we reported a tick recovered from Antelope Cave in extreme northwest Arizona. Further analyses of coprolites from Antelope Cave revealed additional parasitological data from coprolites of both human and canid origin. A second tick was found. This site is the only archaeological locality where ticks have been recovered. We also discovered an acanthocephalan in association with Enterobius vermicularis eggs in the same coprolite. This association shows that the coprolite was deposited by a human. This discovery expands our knowledge of the range of prehistoric acanthocephalan infection. In addition, findings from canid coprolites of Trichuris vulpis are reported. This is the first published discovery of T. vulpis from a North American archaeological context. The close association of dogs with humans at Ancestral Puebloan (Anasazi) sites raises the potential that zoonotic parasites were transferred to the human population. The archaeological occupation is associated with the Ancestral Pueblo culture 1,100 yr ago.     

Ancient Mitochondrial DNA Analyses of Ascaris Eggs Discovered in Coprolites from Joseon Tomb

Analysis of ancient DNA (aDNA) extracted from Ascaris is very important for understanding the phylogenetic lineage of the parasite species. When aDNAs obtained from a Joseon tomb (SN2-19-1) coprolite in which Ascaris eggs were identified were amplified with primers for cytochrome b (cyt b) and 18S small subunit ribosomal RNA (18S rRNA) gene, the outcome exhibited Ascaris specific amplicon bands. By cloning, sequencing, and analysis of the amplified DNA, we obtained information valuable for comprehending genetic lineage of Ascaris prevalent among pre-modern Joseon peoples.     

Egg positive rate of Enterobius vermicularis among preschool children in Cheongju, Chungcheongbuk-do, Korea

In an attempt to determine the prevalence of pinworm infection, the egg positive rate of Enterobius vermicularis was examined using the adhesive cello-tape anal swab method in 1,512 preschool children sampled from a total of 20 kindergartens in Cheongju city, in November to December of 2004 (951 children from 13 kindergartens) and September to October of 2005 (561 from 7 kindergartens). The overall egg positive rate was found to be 7.9% (119/1,512); 9.3% (73/784) for boys and 6.3% (46/728) for girls, respectively. The 5-year age group evidenced the highest egg positive rate (10.9%, 47/430) among the examined age groups. As compared to those reported from previous works (ranged from 9.2 to 26.1%), the prevalence of E. vermicularis in the Cheongju city area is relatively low.