A review on application of next-generation sequencing methods for profiling of protozoan parasites in water: Current methodologies, challenges, and perspectives

Mthethwa, Nonsikelelo Precios; Amoah, Isaac Dennis; Reddy, Poovendhree; Bux, Faizal; Kumari, Sheena

doi:10.1016/j.mimet.2021.106269

Cited by 18 publications

(11 citation statements)

References 122 publications

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“…Conventional Sanger sequencing is still widely used for typing Cryptosporidium, particularly at the gp60 locus [6,7]. Despite the decreasing cost per base of next-generation sequencing (NGS), metagenomic approaches have been under-utilized for Cryptosporidium detection and typing, but are unparalleled for understanding the extent of mixed infections and transmission dynamics [17][18][19] and in detecting and characterizing the diversity of Cryptosporidium species in water and wastewater [20,21]. Despite the utility and importance of both conventional and NGS typing systems for Cryptosporidium, they are still cost-prohibitive for most low-income countries.…”

Section: Cryptosporidium Detection Methodsmentioning

confidence: 99%

Point of care diagnostics for Cryptosporidium: new and emerging technologies

Hijjawi¹,

Zahedi²,

Ryan³

2022

Current Opinion in Gastroenterology

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Purpose of reviewAlthough Cryptosporidium detection and typing techniques have improved dramatically in recent years, relatively little research has been conducted on point of care (POC) detection and typing tools. Therefore, the main purpose of the present review is to summarize and evaluate recent and emerging POC diagnostic methods for Cryptosporidium spp. Recent findingsMicroscopy techniques such as light-emitting diode fluorescence microscopy with auramine-phenol staining (LED-AP), still have utility for (POC) diagnostics but require fluorescent microscopes and along with immunological-based techniques, suffer from lack of specificity and sensitivity. Molecular detection and typing tools offer higher sensitivity, specificity and speciation, but are currently too expensive for routine POC diagnostics. Isothermal amplification methods such as loop-mediated isothermal amplification (LAMP) or recombinase polymerase amplification (RPA) including a commercially available LAMP kit have been developed for Cryptosporidium but are prone to false positives. Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas diagnostic technologies (CRISPRDx) have recently been combined with isothermal amplification to increase its specificity and sensitivity for detection and typing. Other emerging technologies including amplification-free CRISPR detection methods are currently being developed for Cryptosporidium using a smartphone to read the results. SummaryMany challenges are still exist in the development of POC diagnostics for Cryptosporidium. The ideal POC tool would be able to concentrate the pathogen prior to detection and typing, which is complicated and research in this area is still very limited. In the short-term, CRISPR-powered isothermal amplification lateral flow tools offer the best opportunity for POC Cryptosporidium species and subtype detection, with a fully integrated autonomous biosensor for the long-term goal.

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Section: Cryptosporidium Detection Methodsmentioning

confidence: 99%

Point of care diagnostics for Cryptosporidium: new and emerging technologies

Hijjawi¹,

Zahedi²,

Ryan³

2022

Current Opinion in Gastroenterology

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“…Additionally, some difficulties are expected while performing DNA extraction from (oo)cysts, and there is a limited parasite database availability for the development of a data analysis pipeline. Thus, the fundamental needs for NGS application include standardized protocols for the profiling of intestinal parasites in highly complex environments, such as stool samples, and well-established datasets for metagenomics analysis [ 52 ].…”

Section: The Establishment and Expansion Of Molecular Methodsmentioning

confidence: 99%

Diagnostic Methods of Common Intestinal Protozoa: Current and Future Immunological and Molecular Methods

et al. 2022

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Intestinal protozoan infection is a persisting public health problem affecting the populations of developing countries in the tropical and subtropical regions. The diagnosis of intestinal protozoa remains a challenge especially in developing countries due to a shortage of laboratory facilities, limited health funding, and the remoteness of communities. Despite still being widely used, conventional diagnoses using microscopy and staining methods pose important limitations, particularly due to their low sensitivities and specificities. The selection of diagnostic methods needs to be carefully considered based on the objective of examination, availability of resources, and the expected parasite to be found. In this review, we describe various immunodiagnosis and molecular diagnostic methods for intestinal protozoa infection, including their advantages, disadvantages, and suitability for different settings, with a focus on Entamoeba histolytica, Giardia duodenalis, and Cryptosporidium spp.

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“…Reading and decoding genetic material with sequencing technology reveals the diversity and functioning of biological communities. Over the past two decades, next generation sequencing (NGS) technology has been applied to a wide range of water research topics such as aquatic ecology and resource management [1], water treatment [2,3], waterborne disease [4], pollution source tracking [5], and pollution remediation [6]. Nowadays, secondgeneration technologies like MiSeq from Illumina (San Diego, CA, USA) enable the rapid, parallel sequencing of millions to billions of short DNA sequence reads, while thirdgeneration technologies like the PacBio Sequel system (Pacific Biosciences, Menlo Park, CA, USA) can also sequence long DNA fragments [7].…”

Section: Introductionmentioning

confidence: 99%

MinION Nanopore Sequencing Accelerates Progress towards Ubiquitous Genetics in Water Research

Werner

Acharya

Blackburn

et al. 2022

Water

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In 2014, Oxford Nanopore Technologies (ONT) introduced an affordable and portable sequencer called MinION. We reviewed emerging applications in water research and assessed progress made with this platform towards ubiquitous genetics. With >99% savings in upfront costs as compared to conventional platforms, the MinION put sequencing capacity into the hands of many researchers and enabled novel applications with diverse remits, including in countries without universal access to safe water and sanitation. However, to realize the MinION’s fabled portability, all the auxiliary equipment items for biomass concentration, genetic material extraction, cleanup, quantification, and sequencing library preparation also need to be lightweight and affordable. Only a few studies demonstrated fully portable workflows by using the MinION onboard a diving vessel, an oceanographic research ship, and at sewage treatment works. Lower nanopore sequencing read accuracy as compared to alternative platforms currently hinders MinION applications beyond research, and inclusion of positive and negative controls should become standard practice. ONT’s EPI2ME platform is a major step towards user-friendly bioinformatics. However, no consensus has yet emerged regarding the most appropriate bioinformatic pipeline, which hinders intercomparison of study results. Processing, storing, and interpreting large data sets remains a major challenge for ubiquitous genetics and democratizing sequencing applications.

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A review on application of next-generation sequencing methods for profiling of protozoan parasites in water: Current methodologies, challenges, and perspectives

Cited by 18 publications

References 122 publications

Point of care diagnostics for Cryptosporidium: new and emerging technologies

Point of care diagnostics for Cryptosporidium: new and emerging technologies

Diagnostic Methods of Common Intestinal Protozoa: Current and Future Immunological and Molecular Methods

MinION Nanopore Sequencing Accelerates Progress towards Ubiquitous Genetics in Water Research

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