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The Role of Multiplex Realtime PCR in Detecting Acute Diarrhoea Pathogens

13 Tháng 06, 2026

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The Role of Multiplex Realtime PCR in Detecting Acute Diarrhoea Pathogens

Van Hung Pham^1,2*, Quang Duy Ha^1,2, Duy Khanh Tran^1,2, Huong Thien Pham^1,2,3, Anh Hong Pham ^1,2,4
¹Vietnam Research and Development Institute of Clinical Microbiology, Ho Chi Minh City 70000, Vietnam; ²NK-Biotek Co.; ³International University,Vietnam National University, Ho Chi Minh City 70000, Vietnam; ⁴Pham Ngoc Thach University of Medicine, Ho Chi Minh City 70000, Vietnam

BACKGROUND

Gastrointestinal infections are primary causes of acute diarrhea. Detecting and identifying the micro-organism pathogens is often challenging due to the limitations of classical microbiological methods and the vast diversity of pathogens, each requiring distinct conditions and diagnostic methods that many laboratories cannot accommodate.

AIMS OF THE STUDY

This study aims to apply Multiplex Real-time PCRdeveloped by Vietnam Research and Development Institute of Clinical Microbiology to simultaneously detect 18 viruses, 21 bacteria, and 11 parasites, therefore provides clinicians with rapid and accurate diagnostic support for timely treatment.

MATERIALS AND METHODS

Stool samples from patients clinically diagnosed with acute diarrhea were collected and transported from the hospitals to the laboratory of NK-Biotek company supported by Viet Nam Research and Development Institute of Clinical Microbiology. Right after receiving, the DNA/RNA of the sample was extracted using the NKDNA/RNAprep MAGBEAD kit, and the NKDiarrhea Multiplex Real-time PCR (MPL-rPCR) kit was subsequently performed to detect the pathogens existed in the patients’ stool. This method had applied in some studies in Viet Nam [1,2,3,4], and in this study we want to analyze the result of this method to detect diarrhea pathogens from a large number of stool samples collected from patients hospitalized by acute diarrhea.

RESULTS

A total 1,992 samples were analyzed, with the majority of cases belonging to the 0–4 age group (66.7%). The overall positivity rate reached 63.5% (1,265/1,992 cases). Notably, single-pathogen infections were identified in 48.4% of cases, while multi-pathogen co-infections were: 15.1%. Among the 1,265 positive cases, viruses were the most frequent contributors (795 cases), followed by bacteria (685) and parasites (95). Rotavirus Group A (31.4%), Adenovirus (16.0%), and Norovirus GII (14.6%) were the most prevalent pathogens among the viral group. In the bacterial group, diarrheagenic E. coli (DEC) dominated at 20.9%, followed by Clostridium difficile (10.9%) and Salmonella (10.5%). In parasitological group, Blastocystis was the leading parasite agent (44.2%).

DISCUSSIONS AND CONCLUSION

Detecting the causative microorganisms of diarrhea is a challenge in clinical microbiology laboratories because many pathogens are beyond the laboratory’s capabilities. Therefore, the application of MPL-rPCR for multi-pathogens detection is a necessary solution that needs to be implemented. This study showed a high detection rate of pathogens, including viral pathogens such as rotavirus and norovirus. In addition, bacterial and parasitic pathogens that would be undetectable with conventional microbiology methods, such as DEC and Campylobacter, were also detected with the developed MPL-rPCR.

KEY WORDS

Acute Diarrhea Pathogens; MPL real-time PCR; Diarrheagenic viral pathogens; Diarrheagenic bacterial pathogens; Diarrheagenic parasitological pathogens

REFERENCES

1. Tran KQ, Nguyen UTP, Le LTT, Nguyen NTY, Nguyen HHT. (2025). Bacterial etiology of acute diarrhea in Vietnamese children under five: A cross-sectional study using real-time polymerase chain reaction. Acta Biomed. 2025;96(6):17159. doi:10.23750/abm. v96i6.171592. Khai Quang Tran, Hung Hoang Tuan Nguyen, Van Hung Pham, Nghia Quang Bui, Tho Kieu Anh Pham, Toan Hoang Ngo and Phuong Minh Nguyen. (2024). A Cross-sectional Study on the Role of Rotavirus and Microbial Co-infection in Children with Acute Diarrhea in Vietnam. Arch Pediatr Infect Dis. 2024 January; 12(1): e140509. doi: https://doi.org/10.5812/apid-140509
3. Khai Quang Tran, Phuong Minh Nguyen, Uyen Thi Phuong Nguyen, Hung Hoang Tuan Nguyen, Nghia Quang Bui. (2025). High Frequency of Viral Etiology and Antibiotic Utilization Patterns in Pediatric Acute Diarrhea: A Cross-Sectional Study at a Tertiary Hospital in Vietnam. Turkish Archives of Pediatrics. 2025;60(5):536-542. DOI: 10.5152/TurkArchPediatr.2025.25146
4. Thanh Minh Ngoc Bui, Tri Minh Bui, Phuc Le Hoang, Van Hung Pham, Tuan Anh Nguyen. (2026). Diarrheal pathogens in children under five: a cross-sectional study from Children’s Hospital 1, Vietnam. Journal of Tropical Pediatrics, 2026, Vol, 72, Issue 2. https://doi.org/10.1093/tropej/fmag016.

Đào tạo & nghiên cứu

The Role of Multiplex Realtime PCR in Detecting Acute Diarrhoea Pathogens

The Role of Multiplex Realtime PCR in Detecting Acute Diarrhoea Pathogens

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