CRISPR/Cas system and its application in the diagnosis of animal infectious diseases

Abstract

Infectious diseases are a serious threat to the existence of animals and humans' life. In the 21st century, the emergence and re-emergence of several zoonotic and non-zoonotic global pandemic diseases of socio-economic importance has affected billions of humans and animals. The need for expensive equipment and laboratories, non-availability of on-site testing abilities, with time-consuming and low sensitivity and specificity issues of currently available diagnostic techniques to identify these pathogenic micro-organisms on a large scale highlighted the need for developing cheap, portable environment friendly diagnostic methods. In recent years, these issues have been addressed by clustered regularly interspaced palindromic repeats (CRISPR)-based diagnostic platforms that have transformed the molecular diagnostic field due to their outstanding ultra-sensitive nucleic acid detecting capabilities. In this study, we highlight the types, potential of different Cas proteins, and amplification systems. We also illuminate the application of currently available CRISPR integrated setups on the diagnosis of infectious diseases, majorly in food-producing animals (pigs, ruminants, poultry, and aquaculture), domestic pets (dogs and cats), and diseases of zoonotic importance. We conclude the challenges and future perspectives of using these systems to rapidly diagnose and treat other infectious diseases and also develop control strategies to prevent the spread of pathogenic organisms.

Keywords: CRISPR/Cas systems; applications; biosensor; infectious disease; molecular diagnostics; point‐of‐care.

FIGURE 1

Diagrammatic representation of different Cas proteins and their cleavage abilities. (A) Cas9 requires two domains (HNH and RuvC) and produces blunt ends, (B) dCas9 catalytic activity mutated, (C) Cas12a needs PAM and targets dsDNA showing trans‐cleavage activity, (D) Cas12b requires sgRNA like Cas9 but shows similar cleavage activity like Cas12a, (E) Cas13 mainly targets RNA, and (F) Cas14 do not need any specific PAM and target ssDNA.

FIGURE 2

Graphical depiction of key steps in CRISPR–Cas systems for detecting infectious pathogens; (1) selection of sample, (2) extraction of DNA/RNA, (3) amplification of DNA/RNA product, (4) signal production by Cas proteins, and (5) biosensors to readout these signals.

FIGURE 3

List of infectious pathogens diagnosed by CRISPR system in animals. CRISPR‐based diagnostic platforms are used to diagnose infectious pathogens (viruses, bacteria, parasites, and protozoa) in different animals (aquaculture, ruminants, cats and dogs, poultry, and pigs), and disease of zoonotic potential.

FIGURE 4

CRISPR–Cas systems: versatile applications. CRISPR–Cas systems are transforming multiple fields, from precise gene editing in healthcare to rapid diagnostics and agricultural advancements. Their ability to target and modify DNA makes them powerful tools for both research and real‐world solutions.