Parasitism of free-ranging Neotropical primates: examining parasite-host and parasite-parasite relationships

Parasites are infectious agents that require resources from host organisms to complete all or part of their lifecycles. It is customary for wild animals to acquire and maintain multiple parasitic infections during their lifetime. The effects of parasites on hosts vary across demographic and environmental variables, and in relation to each other. Moreover, the propensity for a host animal to acquire an infection can be influenced by other host species in the area that are susceptible to the same parasites. This dissertation describes and explores the natural parasite assemblage of a free-ranging community of nonhuman primates in the Peruvian Amazon rainforest. A mark-recapture program of sympatric saddleback (Leontocebus weddelli) and emperor (Saguinus imperator) tamarins (Callitrichidae) was implemented from June through August, 2012 – 2015, to collect repeat blood and fecal samples from known individuals. Seven other primates on site, belonging to the Pitheciidae, Cebidae, Atelidae, and Aotidae, were followed in 2014 and 2015 for noninvasive collection of fecal samples. Microscopy and molecular techniques were used to assess the degree of parasite–host specificity in gastrointestinal and blood parasites. I also explored individual differences in infection status engendered by various host factors, and tested for non-random associations of co-occurrence between multiple parasites. Patterns of infection were analyzed using multifactorial statistical models. In total, I collected 250 blood samples from 134 known tamarins, 208 fecal samples from 105 known tamarins, and 64 fecal samples distributed across the remaining hosts. Temporal variation of multiple parasite infections confirmed the necessity of a multi-year study to evaluate parasite-host relationships in this system. Individual age was a common predictor of parasite infection, and co-infections were significant predictors for blood parasites. No gastrointestinal parasites appeared to be host specific, but significant differences in prevalence were observed across hosts. I also report the presence of a natural, potentially zoonotic malarial parasite that for first time is demonstrated to chronically infect its nonhuman primate host. Collectively these data demonstrate the importance and need for broad spectrum and long-term screening of parasites from wildlife communities

Detection of neopterin in the urine of captive and wild platyrrhines

Background: Non-invasive biomarkers can facilitate health assessments in wild primate populations by reducing the need for direct access to animals. Neopterin is a biomarker that is a product of the cell-mediated immune response, with high levels being indicative of poor survival expectations in some cases. The measurement of urinary neopterin concentration (UNC) has been validated as a method for monitoring cell-mediated immune system activation in multiple catarrhine species, but to date there is no study testing its utility in the urine of platyrrhine species. In this study, we collected urine samples across three platyrrhine families including small captive populations of Leontopithecus rosalia and Pithecia pithecia, and larger wild populations of Leontocebus weddelli, Saguinus imperator, Alouatta seniculus, and Plecturocebus toppini, to evaluate a commercial enzyme-linked immunosorbent assay (ELISA) for the measurement of urinary neopterin in platyrrhines. Results: Our results revealed measured UNC fell within the sensitivity range of the assay in all urine samples collected from captive and wild platyrrhine study species via commercial ELISA, and results from several dilutions met expectations. We found significant differences in the mean UNC across all study species. Most notably, we observed higher UNC in the wild population of L. weddelli which is known to have two filarial nematode infections compared to S. imperator, which only have one. Conclusion: Our study confirms that neopterin is measurable via commercial ELISA in urine collected from captive and wild individuals of six genera of platyrrhines across three different families. These findings promote the future utility of UNC as a promising biomarker for field primatologists conducting research in Latin America to non-invasively evaluate cell-mediated immune system activation from urine

Real-time genomics for One Health

The ongoing degradation of natural systems and other environmental changes has put our society at a crossroad with respect to our future relationship with our planet. While the concept of One Health describes how human health is inextricably linked with environmental health, many of these complex interdependencies are still not well-understood. Here, we describe how the advent of real-time genomic analyses can benefit One Health and how it can enable timely, in-depth ecosystem health assessments. We introduce nanopore sequencing as the only disruptive technology that currently allows for real-time genomic analyses and that is already being used worldwide to improve the accessibility and versatility of genomic sequencing. We showcase real-time genomic studies on zoonotic disease, food security, environmental microbiome, emerging pathogens, and their antimicrobial resistances, and on environmental health itself – from genomic resource creation for wildlife conservation to the monitoring of biodiversity, invasive species, and wildlife trafficking. We stress why equitable access to real-time genomics in the context of One Health will be paramount and discuss related practical, legal, and ethical limitations

Terrestrial behavior in titi monkeys (Callicebus, Cheracebus, and Plecturocebus) : potential correlates, patterns, and differences between genera

For arboreal primates, ground use may increase dispersal opportunities, tolerance to habitat change, access to ground-based resources, and resilience to human disturbances, and so has conservation implications. We collated published and unpublished data from 86 studies across 65 localities to assess titi monkey (Callicebinae) terrestriality. We examined whether the frequency of terrestrial activity correlated with study duration (a proxy for sampling effort), rainfall level (a proxy for food availability seasonality), and forest height (a proxy for vertical niche dimension). Terrestrial activity was recorded frequently for Callicebus and Plecturocebus spp., but rarely for Cheracebus spp. Terrestrial resting, anti-predator behavior, geophagy, and playing frequencies in Callicebus and Plecturocebus spp., but feeding and moving differed. Callicebus spp. often ate or searched for new leaves terrestrially. Plecturocebus spp. descended primarily to ingest terrestrial invertebrates and soil. Study duration correlated positively and rainfall level negatively with terrestrial activity. Though differences in sampling effort and methods limited comparisons and interpretation, overall, titi monkeys commonly engaged in a variety of terrestrial activities. Terrestrial behavior in Callicebus and Plecturocebus capacities may bolster resistance to habitat fragmentation. However, it is uncertain if the low frequency of terrestriality recorded for Cheracebus spp. is a genus-specific trait associated with a more basal phylogenetic position, or because studies of this genus occurred in pristine habitats. Observations of terrestrial behavior increased with increasing sampling effort and decreasing food availability. Overall, we found a high frequency of terrestrial behavior in titi monkeys, unlike that observed in other pitheciids

Chronic Plasmodium brasilianum infections in wild Peruvian tamarins.

There is an increased interest in potential zoonotic malarias. To date, Plasmodium malariae that infects humans remains indistinguishable from Plasmodium brasilianum, which is widespread among New World primates. Distributed throughout tropical Central and South America, the Callitrichidae are small arboreal primates in which detection of natural Plasmodium infection has been extremely rare. Most prior screening efforts have been limited to small samples, the use of low-probability detection methods, or both. Rarely have screening efforts implemented a longitudinal sampling design. Through an annual mark-recapture program of two sympatric callitrichids, the emperor (Saguinus imperator) and saddleback (Saguinus fuscicollis) tamarins, whole blood samples were screened for Plasmodium by microscopy and nested PCR of the cytochrome b gene across four consecutive years (2012-2015). Following the first field season, approximately 50% of the samples collected each subsequent year were from recaptured individuals. In particular, out of 245 samples from 129 individuals, 11 samples from 6 individuals were positive for Plasmodium, and all but one of these infections was found in S. imperator. Importantly, the cytochrome b sequences were 100% identical to former isolates of P. malariae from humans and P. brasilianum from Saimiri sp. Chronic infections were detected as evidenced by repeated infections (7) from two individuals across the 4-year study period. Furthermore, 4 of the 5 infected emperor tamarins were part of a single group spanning the entire study period. Overall, the low prevalence reported here is consistent with previous findings. This study identifies two new natural hosts for P. brasilianum and provides evidence in support of chronic infections in wildlife populations. Given that callitrichids are often found in mixed-species associations with other primates and can be resilient to human-disturbed environments, they could contribute to the maintenance of P. malariae populations if future work provides entomological and epidemiological evidence indicating human zoonotic infections

Portable sequencing as a teaching tool in conservation and biodiversity research.

As biodiversity loss continues to accelerate, there is a critical need for education and biomonitoring across the globe. Portable technologies allow for in situ molecular biodiversity monitoring that has been historically out of reach for many researchers in habitat nations. In the realm of education, portable tools such as DNA sequencers facilitate in situ hands-on training in real-time sequencing and interpretation techniques. Here, we provide step-by-step protocols as a blueprint for a terrestrial conservation genetics field training program that uses low-cost, portable devices to conduct genomics-based training directly in biodiverse habitat countries

Temporal and demographic blood parasite dynamics in two free-ranging neotropical primates

Parasite-host relationships are influenced by several factors intrinsic to hosts, such as social standing, group membership, sex, and age. However, in wild populations, temporal variation in parasite distributions and concomitant infections can alter these patterns. We used microscropy and molecular methods to screen for naturally occurring haemoparasitic infections in two Neotropical primate host populations, the saddleback (Leontocebus weddelli) and emperor (Saguinus imperator) tamarin, in the lowland tropical rainforests of southeastern Peru. Repeat sampling was conducted from known individuals over a three-year period to test for parasite-host and parasite-parasite associations. Three parasites were detected in L. weddelli including Trypanosoma minasense, Mansonella mariae, and Dipetalonema spp., while S. imperator only hosted the latter two. Temporal variation in prevalence was observed in T. minasense and Dipetalonema spp., confirming the necessity of a multi-year study to evaluate parasite-host relationships in this system. Although callitrichids display a distinct reproductive dominance hierarchy, characterized by single breeding females that typically mate polyandrously and can suppress the reproduction of subdominant females, logistic models did not identify sex or breeding status as determining factors in the presence of these parasites. However, age class had a positive effect on infection with M. mariae and T. minasense, and adults demonstrated higher parasite species richness than juveniles or sub-adults across both species. Body weight had a positive effect on the presence of Dipetalonema spp. The inclusion of co-infection variables in statistical models of parasite presence/absence data improved model fit for two of three parasites. This study verifies the importance and need for broad spectrum and long-term screening of parasite assemblages of natural host populations

Cytochrome b phylogeny with new <i>Plasmodium</i> isolates.

<p>One <i>Plasmodium</i> isolate per year from this study has been included in the phylogeny with infected animals indicated by their unique animal ID numbers. <i>Plasmodium</i> isolates from humans are indicated in red and a squirrel monkey isolate from Perú is in blue. For each sequence, host species, sample locations, and GenBank accession numbers are provided. Emperor tamarin photo reprinted from <a href="https://fieldprojects.org/" target="_blank">https://fieldprojects.org</a> under a CC BY license, with permission from Ishaan Raghunandan, original copyright [2014].</p

Nested-PCR screening results.

<p>Nested-PCR screening results.</p

Field site and surrounding area.

<p>Spatial data layers on intact forest coverage were obtained from the Intact Forest Landscapes project [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184504#pone.0184504.ref038" target="_blank">38</a>].</p