Research Article: Validation of use of the miniPCR thermocycler for Ebola and Zika virus detection

Date Published: May 9, 2019

Publisher: Public Library of Science

Author(s): Everardo González-González, Jackelin Lizeth Mendoza-Ramos, Sara Cristina Pedroza, Aimé Alexandra Cuellar-Monterrubio, Alan Roberto Márquez-Ipiña, Daniel Lira-Serhan, Grissel Trujillo-de Santiago, Mario Moisés Alvarez, Rashid Ansumana.

http://doi.org/10.1371/journal.pone.0215642

Abstract

The development of point-of-care (POC) diagnostic systems has received well-deserved attention in recent years in the scientific literature, and many experimental systems show great promise in real settings. However, in the case of an epidemic emergency (or a natural disaster), the first line of response should be based on commercially available and validated resources. Here, we compare the performance and ease of use of the miniPCR, a recently commercially available compact and portable PCR device, and a conventional thermocycler for the diagnostics of viral nucleic acids. We used both thermocyclers to detect and amplify Ebola and Zika DNA sequences of different lengths (in the range of 91 to 300 nucleotides) at different concentrations (in the range of ~50 to 4.0 x 108 DNA copies). Our results suggest that the performance of both thermocyclers is quite similar. Moreover, the portability, ease of use, and reproducibility of the miniPCR makes it a reliable alternative for point-of-care nucleic acid detection and amplification.

Partial Text

The development of cost-efficient diagnostic point of care (POC) systems for the opportune diagnosis of infectious diseases is a research niche of high relevance [1,2]. The recent pandemic/epidemic episodes associated with viral diseases (e.g., influenza epi-centered in México in 2009 [3,4], Ebola in West Africa in 2013–2015 [5–7], and Zika in Latin America and Southeast Asia in 2016 [8–10]) are tangible and cruel reminders of the need for portable, low-cost, and easy-to-use diagnostic systems that can effectively address epidemic episodes in remote or underprivileged areas [5,9,11,12].

Previous epidemic episodes (for example, those related to EBOV in West Africa in 2014–2016) have proven that an actual emergency does not provide the required timeframe for testing new strategies. This is true for new therapies as well as for diagnosis. In the case of epidemic emergencies (or natural disasters), the first line of response must be based on commercially available and validated resources. Here, we compare the performance of a commercially available portable PCR unit (aimed at several markets, including POC applications) versus that of a conventional, regular-sized PCR thermocycler. The comparison was made in terms of the ability of both units to identify and amplify different synthetically designed genetic sequences of EVOB and ZIKV under the same set of experimental protocols and using the same materials and reagents (see Materials and Methods).

The challenge of POC detection of viral threats is of paramount importance, particularly in underdeveloped regions and in emergency situations (i.e., natural disasters or epidemic outbreaks). In the context of an emergency, time is very limited (as are other resources) to do research or develop new technologies; therefore, the use of commercially available and tested technologies is an obvious first countermeasure. Our research extends the validation of the miniPCR technology to the as yet unexplored topic of detection of Ebola and Zika, and we have validated six sets of primers for Ebola and one set for Zika detection. Our results suggest that the capacity of selective amplification in a conventional thermocycler and in a miniPCR is essentially the same. The use of the miniPCR is intuitive and simple; the user can easily follow the advance of the iterative temperature cycling using a laptop. Despite its compact size, the miniPCR allows a full amplification protocol to be performed in a similar time as in a conventional thermocycler (a difference of only six minutes in a 30-cycle protocol). The mini-PCR thermocycler exhibits the essential attributes of a POC system: (a) the use of small volumes, (b) low capital cost, (c) portability, (d) and a fast, accurate, and selective response. Therefore, this already commercially available and simple nucleic acid amplification system has great potential for use in remote or underprivileged areas, in the case of natural disasters, on the battlefield, or during epidemic emergencies.

 

Source:

http://doi.org/10.1371/journal.pone.0215642

 

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