Comparison of polymerase chain reaction on fresh tissue samples and fecal drops on filter paper for detection of Trypanosoma cruzi in Rhodnius prolixus

Abstract

PCR detection of

Target Product Profile (TPP) for Chagas Disease Point-of-Care Diagnosis and Assessment of Response to Treatment

“…For other settings, more complex technologies such as enzyme-linked immunosorbent assay (ELISA) and PCR might be acceptable. Molecular methods applied to dried blood on filter paper, which has been carried out for triatomine samples [ 28 , 29 ], could enhance detection accuracy in acute and congenital Chagas disease cases. Moreover, serological methods based on low-volume samples are currently under standardization and validation [ 30 ].…”

PCR reveals significantly higher rates of Trypanosoma cruzi infection than microscopy in the Chagas vector, Triatoma infestans: High rates found in Chuquisaca, Bolivia

“…In T. dimidiata , higher rates of infection were revealed when studying rectal samples as compared to those from intestine and stomach; although the difference was not significant, prevalence detected by PCR was higher than for microscopy (Discrepancy = 14.5%) [13]. The use of different primers and sample processing procedures [16], improvements in PCR detection limits or differences in the microscopes, and variation in intensity of T. cruzi infection in T. infestans [13] could all contribute to differences in sensitivity of the two methods. Considering the results of this and other studies, the most sensitive method appears to be using TCZ1 and TCZ2 primers directed to amplify a 188 bp of the 195-bp repetitive nuclear sequence and sampling the rectum of the bug.…”

“…Molecular assays based on the polymerase chain reaction (PCR) have been proposed as an alternative for parasitic detection of T. cruzi infection in vectors [10]. Several primers complementary to the conserved region of the kinetoplast-minicircle part of the mitochondrial DNA have been used in PCR based assays in order to detect this parasite in vertebrate blood samples [8,11,15] and in fecal content of triatomines [16-18]. However, it has been suggested that the technique using the TCZ1 and TCZ2 primers that amplify a 188 bp of a 195-bp repetitive nuclear sequence is the most sensitive compared with methods using S35 and S36 primers to amplify a 330-bp minicircle sequence [10,19].…”

Suitability of a rapid DNA isolation and amplification for detection ofTrypanosoma cruziinTriatoma infestansdry fecal spots collected on filter paper

“…Nevertheless, all negative filter paper samples revealed the 144 bp DNA fragment after addition of T. cruzi DNA thus confirming the inexistence of amplification inhibitor factors what was crucial to rule out the presence of inhibitors, that have already been reported by Breniere et al (1995) who used the same boiling procedure of the present study, and Araújo et al (2002) who used a more complex extraction procedure with phenol-chloroform extractions. Dorn et al (1999;2001) have associated amplification inhibition to the type of biological sample, suggesting that digestive tract samples, especially stomach contents are more prone to inhibit PCR. We decided to increase the number of insects in the second study group (lower parasitemia model) with a proportional reduction of the number of insects in the control group due to the loss of some insects during the first experiment (high parasitemia), and also because all negative triatomines did not generate any amplification during the first part of the study, taken into account that our capacity to breed and maintain triatomines was limited by the laboratory infrastructure.…”