Cryptosporidium parvum among Coprolites from La Cueva de los Muertos Chiquitos (600â•fi800 CE), Rio Zape Valley, Durango, Mexico

In the present study, 90 coprolites from La Cueva de los Muertos Chiquitos (CMC) were subjected to enzyme-linked immunosorbent assay (ELISA) tests for 3 diarrhea-inducing protozoan parasites, Entamoeba histolytica, Giardia duodenalis, and Cryptosporidium parvum, to determine whether these parasites were present among the people who utilized this cave 1,200–1,400 yr ago. These people, the Loma San Gabriel, developed as a culture out of the Archaic Los Caracoles population and lived throughout much of present-day Durango and Zacatecas in Mexico. The Loma San Gabriel persisted through a mixed subsistence strategy of hunting-gathering and agricultural production. The results of ELISA testing were negative for both E. histolytica and G. duodenalis across all coprolites. A total of 66/90 (~73% prevalence) coprolites tested positive or likely positive for C. parvum. The high prevalence of C. parvum among CMC coprolites contributes to our growing knowledge of the pathoecology among the Loma San Gabriel who utilized CMC. Herein, we report the successful recovery of C. parvum coproantigens from prehistoric coprolites. The recovery of these coproantigens demonstrates the existence of C. parvum in Mesoamerica before European contact in the 1400s. Using enzyme-linked immunosorbent assays (ELISAs) as a means of examining archaeological materials for evidence of diarrhea-inducing protozoan parasites has become increasingly more routine among archaeoparasitological researchers (Allison et al., 1999; Gonçalves et al., 2002, 2003, 2004, 2005; Le Bailly and Bouchet, 2006; Le Bailly et al., 2006, 2008, 2014; Mitchell et al., 2008; Friás et al., 2013; Yeh et al., 2014, 2015). Much of the existing work has focused on the recovery of protozoan parasite antigens from latrine sediments (Gonçalves et al., 2004; Le Bailly et al., 2008, 2014, Mitchell et al., 2008; Yeh et al., 2014, 2015). In addition to sediments, Allison et al. (1999) used ELISAs to test coprolites from the intestinal tracts of mummies for Cryptosporidium sp. and Giardia sp. antigens. Another study tested coprolites for Giardia duodenalis antigens and found 4/84 to be positive (Gonçalves et al., 2002). Gonçalves et al. (2004) examined 22 coprolites from Brazil, Chile, and Sudan using an ELISA kit for detecting Entamoeba histolytica antigens and did not find any positive samples. The present study represents an effort to analyze a large number of coprolites (n1⁄4 90) from a single archaeological site for the presence of 3 common diarrhea-inducing species of protozoans. It is reasonable that the occurrence of these parasites contributed to prehistoric childhood mortality rates in ways similar to their significance in modern human populations of developing countries. Presently, approximately 15–31% of child deaths are attributed to diarrheal diseases, rates that are often higher in less developed nations (Walker et al., 2012; Pop et al., 2014). Immunodiagnositic methods offer the promise of increased sensitivity and specificity for the detection of diarrhea-inducing protozoan parasites preserved in fecal samples from prehistoric human populations where microscopic evidence of cysts may be lacking because of infrequent shedding or degradation. The present study utilized commercially available ELISA kits for the detection of E. histolytica, G. duodenalis, and Cryptosporidium parvum coproantigens in preserved fecal samples to better understand the pathoecology of parasitism among the Loma San Gabriel people residing in a cave known as La Cueva de los Muertos Chiquitos (CMC). Studies that explore the parasites of these coprolites yield insights into the pathoecology of the Loma San Gabriel who utilized CMC (Jiménez et al., 2012) and provide data for understanding how parasitism may have affected their daily lives. The Rio Zape Valley, a region approximately 18 km southeast of Guanacevı́ in Durango, Mexico, is home to several caves and rock shelters utilized by the Loma San Gabriel. The region illustrates a transition zone between the northernmost edge of Mesoamerica and the greater American Southwest (Kelley, 1956, 1971; Brooks and Brooks, 1980). CMC itself was used year round as a temporary habitation by the Loma San Gabriel between 1,200 and 1,400 yr ago (Brooks et al., 1962; Foster, 1986). This cave housed an abundance of botanical artifacts, the skeletons of 7 children (ages several months to 5 yr at the time of death), and nearly 500 coprolites, all sealed beneath adobe floors (Brooks et al., 1962; Brooks and Brooks, 1978, 1980; Phillips, 1989). Previous analyses of coprolites reported excellent preservation of parasitic helminth eggs (Jiménez et al., 2012) and bacterial DNA (Tito et al., 2012). The people who utilized CMC subsisted using a mixed strategy of agricultural production and huntinggathering, both of which fluctuated seasonally (Brooks et al., 1962; Foster, 1984; Hammerl et al., 2015). Previous work at CMC focused on helminth infections (Jiménez et al., 2012). From this work we infer that community sanitation was poor by modern standards. Artifacts from CMC (e.g., bones from fish, rodents, and other mammals) demonstrate that there was potential for the transmission of zoonotic parasites. Our understanding of the Loma San Gabriel culture suggests multiple potential pathways for parasite transmission. Studies have shown that those utilizing CMC had living and nutritional behavior patterns that likely perpetuated the life cycles of parasites associated with poor sanitation. As these people modified their environments for food production and took up residence in fecal-contaminated shelters, they inadvertently promoted the transmission of diarrhea-inducing pathogens. The presence of diarrhea is reflected in the morphologies of some CMC coprolites that showed evidence of diarrheal events (Jiménez et al., 2012). Some of these events may have resulted from infections of diarrhea-inducing protozoan parasites (Fig. 1; Table I). Received 22 October 2015; revised 15 April 2016; accepted 20 April 2016.