Determination of Baylisascaris schroederi Infection in Wild Giant Pandas by an Accurate and Sensitive PCR/CE-SSCP Method

Abstract

It has been recognized that other than habitat loss, degradation and fragmentation, the infection of the roundworm Baylisascaris schroederi (B. schroederi) is one of the major causes of death in wild giant pandas. However, the prevalence and intensity of the parasite infection has been inconsistently reported through a method that uses sedimentation-floatation followed by a microscope examination. This method fails to accurately determine infection because there are many bamboo residues and/or few B. schroederi eggs in the examined fecal samples. In the present study, we adopted a method that uses PCR and capillary electrophoresis combined with a single-strand conformation polymorphism analysis (PCR/CE-SSCP) to detect B. schroederi infection in wild giant pandas at a nature reserve, and compared it to the traditional microscope approach. The PCR specifically amplified a single band of 279-bp from both fecal samples and positive controls, which was confirmed by sequence analysis to correspond to the mitochondrial COII gene of B. schroederi. Moreover, it was demonstrated that the amount of genomic DNA was linearly correlated with the peak area of the CE-SSCP analysis. Thus, our adopted method can reliably detect the infectious prevalence and intensity of B. schroederi in wild giant pandas. The prevalence of B. schroederi was found to be 54% in the 91 fecal samples examined, and 48% in the fecal samples of 31 identified individual giant pandas. Infectious intensities of the 91 fecal samples were detected to range from 2.8 to 959.2 units/gram, and from 4.8 to 959.2 units/gram in the fecal samples of the 31 identified giant pandas. For comparison, by using the traditional microscope method, the prevalence of B. schroederi was found to be only 33% in the 91 fecal samples, 32% in the fecal samples of the 31 identified giant pandas, and no reliable infectious intensity was observed.

Update on Baylisascariasis, a Highly Pathogenic Zoonotic Infection

“…The complete mitochondrial genomes of B. procyonis (124), B. schroederi, B. ailuri, and B. transfuga (125) have been published, and mitochondrial and nuclear markers have been developed for B. columnaris (76). Molecular tests have also been developed for environmental investigations and for the diagnostic identification of Baylisascaris DNA in definitive host fecal samples and tissue samples from biopsies and autopsies performed in clinical cases of NLM (128)(129)(130)(131). Specialists are usually required for larval identification, although even their experience is not always sufficient to differentiate B. procyonis from other types of larvae, particularly within the genus Baylisascaris.…”

“…Ideally, molecular tests would provide a rapid identification of B. procyonis, separating it from closely related species, including B. columnaris (76), and would also serve as a tool for epidemiologic studies and sanitary management, considering the close association between raccoons and humans that can lead to the contamination of soil, water, and food with eggs of Baylisascaris (128,129). These tests are also very important for host-based or environmental diagnosis in cases in which captive animals may be at risk (130).…”

Peer Review #2 of "Factors affecting genotyping success in giant panda fecal samples (v0.1)"

“…Such high usage is attributable to the sampling being convenient, random, non-invasive, and non-interfering to the animal's activity, and is facilitated by the abundance of samples (Kohn and Wayne 1997). The research using fecal DNA includes documents on species identification (Dalen et al 2004), sex determination (Huber et al 2002), kinship and paternity (Constable et al 2001), population genetic diversity (Zhang et al 2007), adaptive variation (Wan et al 2006), population genetic structure (Zhu et al 2013), dispersal pattern (Zhan et al 2007), population size (Eggert et al 2003), evolutionary history of species (Chen et al 2013), mating system (Garnier et al 2001), and disease information (Zhang et al 2012).…”

Prokaryotic Expression and Serodiagnostic Potential of Glyceraldehyde-3-Phosphate Dehydrogenase and Thioredoxin Peroxidase from Baylisascaris schroederi

“…A large quantity of parasites may also cause intestinal obstruction of giant pandas, and even death [ 6 , 7 ]. The diagnosis of this disease mainly depends on sedimentation floatation and Polymerase Chain Reaction (PCR) methods to detect the eggs [ 8 , 9 , 10 ]. However, it is a significant challenge to carry out a diagnosis at an early stage of infection using these methods.…”