Trends in Parasitology
OpinionQuantitative PCR-Based Diagnosis of Soil-Transmitted Helminth Infections: Faecal or Fickle?
Section snippets
A Timely Diagnosis
The rapid, sensitive, and accurate diagnosis of human helminth infections underpins efforts to control or possibly eliminate disease-causing parasitic worms. For many decades, microscopy has been the most widely used and mainstay tool for detecting, identifying, and enumerating parasites. Advances in molecular biology and genomics provides opportunities for better tests. Of considerable interest is the application of qPCR to diagnose STH infections by testing stool samples; qPCR and its
Cost of qPCR
qPCR is a fairly complex tool and a relatively expensive procedure, requiring established facilities, an adequate supply of reagents, sufficient training for protocol reproducibility or troubleshooting, and local (company, contract, country) support. Overall, cost is decreasing due to increased reproducibility of qPCR results, obviating the need for multiple sampling [5], and reductions in the costs of some reagents [1]. Importantly, evaluating cost as ‘price per test alone’ is misguided, since
Concluding Remarks
It is dispiriting that some significant species of intestinal helminths, known to infect hundreds of millions of people and cause destructive diseases, have not been able to ‘make the cut’ to the list of parasites to be controlled. Helminths such as Strongyloides stercoralis are not included in current intervention agendas because diagnosis has not been standardized; with no egg output, and with highly varied levels of larval output, the intensity of infection cannot be reliably measured [45].
Outstanding Questions
What is needed for qPCR to be a robust, accurate, and reliable method for STH diagnosis and as a measure for STH infection intensity? If all that is measured by qPCR is copies of the target DNA present, how can outputs (Cq-value) be related to intensity of infection?
Is there a need to demonstrate the presence and utility of repetitive elements identified from limited genomic data across species ranges and in perpetuity?
How can identification and choice of molecular markers be improved as
Glossary
- Breakpoint of transmission
- defined by mathematical modelling, the point at which parasite reproduction becomes unsustainable and continuation of the parasite life cycle (transmission) falls to such levels that do not allow recrudescence.
- Inhibition
- any factor than can stall a PCR reaction; matrices like blood, stool, and urine carry a plethora of inhibitors (bacterial debris, bile salts, haeme, humic acids, polysaccharides) that modern nucleic acid extraction kits remove or wash away by
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2022, Food ControlCitation Excerpt :Zhang et al. (2022) developed a triplex TaqMan-based real-time PCR to detect and characterize three E. multilocularis, E. granulosus sensu stricto, and E. shiquicus in canid fecal samples with sensitivity 10 copies plasmid/μl, which regarding the high possibility of contamination of vegetable farmlands with canid feces, could be employed to investigate food and water samples (Zhang et al., 2022). However, beside advantages of real-time PCR for detection of pathogens in different samples, need for expensive instruments, trained technicians, risk of contaminations, and non-specific amplification, particularly due to primer-dimer, have limited applications of this technique for detection of pathogens in food and water resources (Maurer, 2011; Papaiakovou et al., 2019). There are a lot of nucleic acid amplification (NAA) techniques that amplify target genes at isothermal conditions, such as nucleic acid sequence-based amplification (NASBA), self-sustained sequence replication (3SR) (Fahy et al., 1991), strand displacement amplification (SDA) (Walker et al., 1992), multiple displacement amplification (MDA) (Huang et al., 2020), recombinase polymerase amplification (RPA) (Rostron et al., 2019), and LAMP (Wong et al., 2018).