Application of a Multiplex Quantitative PCR to Assess Prevalence and Intensity Of Intestinal Parasite Infections in a Controlled Clinical Trial

Background

Accurate quantitative assessment of infection with soil transmitted helminths and protozoa is key to the interpretation of epidemiologic studies of these parasites, as well as for monitoring large scale treatment efficacy and effectiveness studies. As morbidity and transmission of helminth infections are directly related to both the prevalence and intensity of infection, there is particular need for improved techniques for assessment of infection intensity for both purposes. The current study aimed to evaluate two multiplex PCR assays to determine prevalence and intensity of intestinal parasite infections, and compare them to standard microscopy.

Methodology/Principal Findings

Faecal samples were collected from a total of 680 people, originating from rural communities in Timor-Leste (467 samples) and Cambodia (213 samples). DNA was extracted from stool samples and subject to two multiplex real-time PCR reactions the first targeting: Necator americanus, Ancylostoma spp., Ascaris spp., and Trichuris trichiura; and the second Entamoeba histolytica, Cryptosporidium spp., Giardia. duodenalis, and Strongyloides stercoralis. Samples were also subject to sodium nitrate flotation for identification and quantification of STH eggs, and zinc sulphate centrifugal flotation for detection of protozoan parasites. Higher parasite prevalence was detected by multiplex PCR (hookworms 2.9 times higher, Ascaris 1.2, Giardia 1.6, along with superior polyparasitism detection with this effect magnified as the number of parasites present increased (one: 40.2% vs. 38.1%, two: 30.9% vs. 12.9%, three: 7.6% vs. 0.4%, four: 0.4% vs. 0%). Although, all STH positive samples were low intensity infections by microscopy as defined by WHO guidelines the DNA-load detected by multiplex PCR suggested higher intensity infections.

Conclusions/Significance

Multiplex PCR, in addition to superior sensitivity, enabled more accurate determination of infection intensity for Ascaris, hookworms and Giardia compared to microscopy, especially in samples exhibiting polyparasitism. The superior performance of multiplex PCR to detect polyparasitism and more accurately determine infection intensity suggests that it is a more appropriate technique for use in epidemiologic studies and for monitoring large-scale intervention trials.     

Sub-chronic iron overload triggers oxidative stress development in rat brain: implications for cell protection

This work was aimed to test the hypothesis that sub-chronic administration of iron-dextran (Fe-dextran) (six doses of 50 mg Fe-dextran/kg) to rats triggers a transient oxidative stress in brain and mechanisms of cellular antioxidant defence. After 2 h of administration of the 6th dose, a significant increase of total Fe, the labile Fe pool (LIP), the lipid radical (LR^?)/α-tocopherol (α-T) content ratio were observed, as compared to values in control brain homogenates. The ascorbyl radical (A^?)/ascorbate (AH^?) content ratio and the oxidation rate of 2′,7′-dichlorodihidrofluorescein (DCFH-DA) were significantly higher in Fe-dextran treated rats, as compared to values in brain from control rats after 4 h treatment. An increase in both catalase (CAT) and superoxide dismutase (SOD) activity was observed at 8 and 1–2 h, respectively. No significant changes were detected in the nuclear factor-κB (NF-κB) levels in nuclear extracts from rat brains after 1–8 h of Fe-dextran administration. After 2 h of Fe administration Fe concentration in cortex, striatum and hippocampus was significantly increased as compared to the same areas from control animals. Both, CAT and SOD activities were significantly increased in cortex after Fe administration over control values, without changes in striatum and hippocampus. Taken as a whole, sub-chronic Fe administration enhances the steady state concentration of Fe in the brain LIP that favors the settlement of an initial oxidative stress condition, both at hydrophilic and lipophilic compartments, resulting in cellular protection evidenced by antioxidant enzyme upregulation.     

Death among primates: a critical review of non‐human primate interactions towards their dead and dying

For the past two centuries, non‐human primates have been reported to inspect, protect, retrieve, carry or drag the dead bodies of their conspecifics and, for nearly the same amount of time, sparse scientific attention has been paid to such behaviours. Given that there exists a considerable gap in the fossil and archaeological record concerning how early hominins might have interacted with their dead, extant primates may provide valuable insight into how and in which contexts thanatological behaviours would have occurred. First, we outline a comprehensive history of comparative thanatology in non‐human primates, from the earliest accounts to the present, uncovering the interpretations of previous researchers and their contributions to the field of primate thanatology. Many of the typical behavioural patterns towards the dead seen in the past are consistent with those observed today. Second, we review recent evidence of thanatological responses and organise it into distinct terminologies: direct interactions (physical contact with the corpse) and secondary interactions (guarding the corpse, vigils and visitations). Third, we provide a critical evaluation regarding the form and function of the behavioural and emotional aspects of these responses towards infants and adults, also comparing them with non‐conspecifics. We suggest that thanatological interactions: promote a faster re‐categorisation from living to dead, decrease costly vigilant/caregiving behaviours, are crucial to the management of grieving responses, update position in the group's hierarchy, and accelerate the formation of new social bonds. Fourth, we propose an integrated model of Life‐Death Awareness, whereupon neural circuitry dedicated towards detecting life, i.e. the agency system (animate agency, intentional agency, mentalistic agency) works with a corresponding system that interacts with it on a decision‐making level (animate/inanimate distinction, living/dead discrimination, death awareness). Theoretically, both systems are governed by specific cognitive mechanisms (perceptual categories, associative concepts and high‐order reasoning, respectively). Fifth, we present an evolutionary timeline from rudimentary thanatological responses likely occurring in earlier non‐human primates during the Eocene to the more elaborate mortuary practices attributed to genus Homo throughout the Pleistocene. Finally, we discuss the importance of detailed reports on primate thanatology and propose several empirical avenues to shed further light on this topic. This review expands and builds upon previous attempts to evaluate the body of knowledge on this subject, providing an integrative perspective and bringing together different fields of research to detail the evolutionary, sensory/cognitive, developmental and historical/archaeological aspects of primate thanatology. Considering all these findings and given their cognitive abilities, we argue that non‐human primates are capable of an implicit awareness of death.     

Rapid identification and interpretation of gene–environment associations using the new R.SamBada landscape genomics pipeline

sam βada is a genome–environment association software, designed to search for signatures of local adaptation. However, pre‐ and postprocessing of data can be labour‐intensive, preventing wider uptake of the method. We have now developed R.SamBada, an r ‐package providing a pipeline for landscape genomic analysis based on sam βada , spanning from the retrieval of environmental conditions at sampling locations to gene annotation using the Ensembl genome browser. As a result, R.SamBada standardizes the landscape genomics pipeline and eases the search for candidate genes of local adaptation, enhancing reproducibility of landscape genomic studies. The efficiency and power of the pipeline is illustrated using two examples: sheep populations from Morocco with no evident population structure and Lidia cattle from Spain displaying population substructuring. In both cases, R.SamBada enabled rapid identification and interpretation of candidate genes, which are further discussed in the light of local adaptation. The package is available in the r CRAN package repository and on GitHub (github.com/SolangeD/R.SamBada).