Detection of Plasmodium in faeces of the New World primate Alouatta clamitans

Abstract Plasmodium falciparum and Plasmodium vivax have evolved with host switches between non-human primates (NHPs) and humans. Studies on the infection dynamics of Plasmodium species in NHPs will improve our understanding of the evolution of these parasites; however, such studies are hampered by the difficulty of handling animals in the field. The aim of this study was to detect genomic DNA of Plasmodium species from the faeces of New World monkeys. Faecal samples from 23 Alouatta clamitans from the Centre for Biological Research of Indaial (Santa Catarina, Brazil) were collected. Extracted DNA from faecal samples was used for molecular diagnosis of malaria by nested polymerase chain reaction. One natural infection with Plasmodium simium was identified by amplification of DNA extracted from the faeces of A. clamitans. Extracted DNA from a captive NHP was also used for parasite genotyping. The detection limit of the technique was evaluated in vitro using an artificial mixture of cultured P. falciparum in NHP faeces and determined to be 6.5 parasites/µL. Faecal samples of New World primates can be used to detect malaria infections in field surveys and also to monitor the genetic variability of parasites and dynamics of infection.

Cross-species amplification of microsatellite markers in Mycteria leucocephala Pennant 1769: Molted feathers as successful DNA source.

DNA from molted feathers is being increasingly used for genetic studies on birds. However, the DNA obtained from such non-invasive sources is often not of enough quantity and quality for isolation of new microsatellite markers. The present study examined the potential of shed feathers of near threatened Painted Stork as a source of its DNA for cross-species amplification of microsatellites. Thirty-one shed feathers of varying conditions (‘good’ and ‘deteriorated’) and sizes (‘large’, ‘intermediate’ and ‘small’) collected in a north Indian population were used to isolate DNA by a standard isopropanol method and 11 microsatellite markers already developed in the Wood Stork were screened for amplification. Nine plucked feathers from two dead Painted Storks were also used to compare the DNA yield and amplification success. The DNA yield of feathers varied significantly in relation to the calamus size and condition. Among molted feathers, ‘good’ and ‘large’ samples provided more DNA than ‘deteriorated’ and ‘small’ ones, respectively. ‘Large’ plucked feathers yielded more DNA than ‘large’ molted feathers. DNA was almost degraded in all the samples and ratio of absorbance at 260/280 nm varied from 1.0 to 1.8, indicating impurity in many samples. Independent of DNA yields, all microsatellites were cross-amplified in all kinds of feathers, with >80% success in different feather categories. It is concluded that the shed feathers can be successfully used to isolate DNA in the Painted Stork and for cross-species amplification of microsatellites.

A new world moneey microsatellite (ap74) highly conserved in primates / AP74, un microsatélite de Monos del Nuevo Mundo altamente conservado en Primates

Given their great variability, microsatellites or STRs became the most commonly used genetic markers over the last 15 years. The analysis of these markers requires minimum quantities of DNA, allowing the use of non invasive samples, such as feces or hair. We amplified the microsatellite Ap74 in blood and hair samples in order to analyze the levels of genomic conservation among a wide range of primates including: Lemur catta, Alouatta caraya, Ateles belzebuth, Ateles chamek, Pan troglodytes, Papio sp., and Homo sapiens. In all cases we obtained amplification products that exhibited similar size both in monkeys and human (oscillating between 126 and 176 bp), except in the lemur where the detected fragment presented a size of approximately 1000 bp. The analysis of the nucleotide sequences permitted the evaluation of the molecular modifications experienced during the evolutionary process in primates.

Dado su alta variabilidad, los microsatélites o STR se convirtieron en los marcadores genéticos más ampliamente utilizados en los últimos 15 años. El análisis de estos marcadores requiere una mínima cantidad de ADN, permitiendo el uso de muestras no invasivas, tales como pelos o heces. Con el objetivo de analizar niveles de conservación genómica, amplificamos el microsatélite Ap74 en muestras de pelo y sangre de un amplio rango de primates incluyendo: Lemur catta, Alouatta caraya, Ateles belzebuth, Ateles chamek, Pan troglodytes, Papio sp., y Homo sapiens. En todos los casos obtuvimos productos de amplificación que exhibieron un tamaño similar (oscilando entre 126 y 176 pb), con excepción del lémur, donde el fragmento detectado presentó un tamaño de aproximadamente 1000 pb. El análisis de las secuencias nucleotídicas nos permitió evaluar las modificaciones moleculares ocurridas durante el proceso evolutivo en primates.