Abstract
BackgroundCurrent molecular target-dependent methods are used to detect only known viruses. However, metagenomics based on next-generation sequencing (NGS) technique is a target-independent assay that enables simultaneous detection and genomic characterisation of all microorganisms present in a sample. In this study, we aimed to develop a metagenomics approach using NGS to identify and characterise viruses in stool samples from infants and children with Acute Gastroenteritis (AGE) in Kuwait.MethodsWe have investigated 84 stool samples from infants and children aged one month to ten years old with signs and symptoms of gastroenteritis who attended Mubarak Al-Kabeer and Al-Amiri hospitals in Kuwait from January to December 2017. A metagenomics approach using NGS to characterise viruses in clinical samples was used. Also, the commercial Real-Time PCR assay was used to detect viruses causing gastroenteritis.ResultsMetagenomics analysis revealed an average of 280,768 reads in which 5% of the reads were derived from viruses. The analysis of viral sequences verified that single infection of human adenovirus was the leading cause of gastroenteritis among infants and children, which was detected in 23.2% of the patients, followed by a mixed infection of human adenovirus and other viruses, which was detected in 20.9% of patients. Also, the newly discovered viruses known to cause gastroenteritis were detected, such as astrovirus MLB2, primate bocaparvovirus-1, Aichivirus A, cardiovirus, parechovirus A, astrovirus VA4, cosavirus-F, and bufavirus-3. Our results showed 71% agreement (k = 0.445, P = 0.000) between multiplex Real-Time PCR, which is used as a routine diagnostic test and metagenomics approach in the detection of viruses causing gastroenteritis in clinical samples.ConclusionDespite the difficulties in sample preparation and analysis process, we showed that metagenomics approach is a powerful and promising tool for the detection and characterisation of different viruses in clinical samples.
Highlights
Current molecular target-dependent methods are used to detect only known viruses
The number of viral reads obtained from metagenomics approach raged from 1 to 1272, and the highest number of reads was for Astrovirus MLB2
The results demonstrated that 51 (60.7%) of the patients were positive for viruses causing gastroenteritis and 33 (39.2%) of the patients were negative
Summary
Metagenomics based on next-generation sequencing (NGS) technique is a target-independent assay that enables simultaneous detection and genomic characterisation of all microorganisms present in a sample. The traditional methods for virus discovery such as filtration, tissue culture, electron microscopy and serology were powerful techniques for the detection of viruses. Due to their limitations, the traditional techniques were replaced by molecular techniques such as polymerase chain reaction (PCR) and DNA sequencing (Sanger method) [1]. These targetdependant molecular techniques had the credits in the discovery of many viruses, many new and novel human viruses are not yet revealed [2]. Virologists were the first to use the metagenomics approach to detect viruses, causing different diseases in humans such as respiratory tract infections, acute flaccid paralysis in children, and gastroenteritis [8, 11, 12]
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