Development and assessment of molecular diagnostic tests for 15 enteropathogens causing childhood diarrhoea: a multicentre study

Liu, Jie; Kabir, Furqan; Manneh, Jainaba; Lertsethtakarn, Paphavee; Begum, Sharmin; Gratz, Jean; Becker, Steve M; Operario, Darwin J.; Taniuchi, Mami; Janaki, Lalitha; Platts-Mills, James A; Haverstick, Doris M.; Kabir, Mamun; Sobuz, Shihab U.; Nakjarung, Kaewkanya; Sakpaisal, Pimmada; Silapong, Sasikorn; Bodhidatta, Ladaporn; Qureshi, Shahida; Kalam, Adil; Saidi, Queen; Swai, Ndealilia; Mujaga, Buliga; Maro, Athanasia; Kwambana-Adams, Brenda; Dione, Michel; Antonio, Martín; Kibiki, Gibson; Mason, Carl J.; Iqbal, Najeeha Talat; Zaidi, Anita K. M.; Houpt, Eric R.

doi:10.1016/s1473-3099(14)70808-4

Cited by 277 publications

(305 citation statements)

References 26 publications

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“…The biological relevance of a pathogen detected at a high CT value (low target copy number) may therefore be questionable, and association with a clinically measurable outcome such as gastroenteritis is more common for lower CT values. 21 For this reason we performed quantitative statistical analysis of CT values in addition to analysis of pathogen prevalence and we found consistent results with both approaches. We were also reliant on molecular biomarkers of EE in stool/plasma and did not perform intestinal biopsies in this asymptomatic population to confirm enteropathy.…”

Section: Discussionmentioning

confidence: 52%

“…20 The presence of bacterial, viral and eukaryotic pathogens in stool samples was assessed through quantitative PCR of extracted DNA and RNA using TaqMan® array cards (TACs). 21 A complete list of pathogen sequence targets is provided in the appendix. Faecal biomarkers of intestinal inflammation, protein-losing enteropathy and immune activation (myeloperoxidase, calprotectin, α1-antitrypsin, neopterin) and plasma biomarkers of microbial translocation (soluble CD14 and endotoxin-core IgG (EndoCAb)) and epithelial damage (intestinal fatty acid binding protein (I-FABP)) were measured using enzyme-linked immunosorbent assays.…”

Section: Laboratory Methodsmentioning

confidence: 99%

See 1 more Smart Citation

The effect of azithromycin on the immunogenicity of oral poliovirus vaccine: a double-blind randomised placebo-controlled trial in seronegative Indian infants

Grassly

Praharaj

Babji

et al. 2016

The Lancet Infectious Diseases

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Section: Discussionmentioning

confidence: 52%

Section: Laboratory Methodsmentioning

confidence: 99%

The effect of azithromycin on the immunogenicity of oral poliovirus vaccine: a double-blind randomised placebo-controlled trial in seronegative Indian infants

Grassly

Praharaj

Babji

et al. 2016

The Lancet Infectious Diseases

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“…Further, we acknowledge that recent evidence shows that detection of bacterial enteropathogens, particularly, Shigella spp., may be increased through the use of molecular diagnostic techniques over conventional bacterial culture. 31 Therefore, we may have underdetected bacterial enteropathogens in our FIGURE 4. Phylogenetic diversity and species richness in diarrhea and recovery stools by age group (whiskers indicate the ranges of phylogenetic diversity or species richness values; *P < 0.01 paired t test).…”

Section: Discussionmentioning

confidence: 96%

Gut Microbiome Composition in Young Nicaraguan Children During Diarrhea Episodes and Recovery

Becker‐Dreps¹,

Allali²,

Monteagudo³

et al. 2015

The American Society of Tropical Medicine and Hygiene

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Abstract. Understanding how the gut microbiota is affected by diarrhea episodes may help explain alterations in intestinal function among children in low-income settings. This study examined the composition of the gut microbiome of Nicaraguan children both during diarrhea episodes and while free of diarrhea for at least 2 months. Relative abundances of bacterial taxa, phylogenetic diversity, and species richness were determined by 16S amplicon sequencing and compared between paired diarrhea and recovery samples. A total of 66 stools were provided by 25 children enrolled in a 1-year cohort study of diarrhea etiologies. Children in our cohort had a mean age of 21.9 months; 64% were breast-fed, and 10% had received an antibiotic during the diarrhea episode. Overall, phylogenetic diversity and species richness did not differ significantly between diarrhea and recovery stools. However, of children who had a bacterial enteropathogen detected in any diarrhea stool, none experienced an increase in phylogenetic diversity in recovery, whereas of those in whom no bacterial enteropathogens were detected in their diarrhea stool(s), 59% experienced an increase in phylogenetic diversity in recovery (P = 0.008). This preliminary study suggests that recovery of the gut microbiota after a diarrhea episode may take longer time than previously thought and may be pathogen specific.

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“…However, because of increased sensitivity, positive detection may be observed in asymptomatic individuals, and other independent methods may be required to confirm active infection (26). Few reports have shown an association between the quantity of pathogens in stool specimens and diarrhea (28)(29)(30). Both qualitative molecular detection and quantification of viral, bacterial, and parasitic enteric pathogens in stool specimens can refine the diagnosis of gastroenteritis and help to ascribe an etiology to coinfected samples.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of a New Device for Simplifying and Standardizing Stool Sample Preparation for Viral Molecular Testing with Limited Hands-On Time

et al. 2016

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Sensitive molecular assays have greatly improved the diagnosis of viral gastroenteritis. However, the proper preparation of stool samples for clinical testing remains an issue. bioMérieux has developed a stool preprocessing device (SPD) that includes a spoon for calibrated sampling and a vial containing buffer, glass beads, and two filters. The resulting stool filtrate is used for nucleic acid extraction. The purpose of this study was to evaluate the performance of the SPD for the quantification of human adenovirus (HAdV) DNA in stool samples collected from hematopoietic stem cell transplant (HSCT) recipients. HAdV DNA was quantified with the Adenovirus R-gene kit. The suitability of the device to reproducibly quantify HAdV DNA in stools using different extraction platforms (easyMAG and QIAsymphony) was determined using archived HAdV-positive stool samples. Coefficients of variation of HAdV DNA quantifications ranged from 1.79% to 1.83%, and no difference in quantification was observed between the two extraction systems. The HAdV DNA limit of quantification using the SPD was 3.75 log 10 copies/g of stool. HAdV DNA quantification using the SPD was then compared to that of the routine preprocessing technique on 75 fresh stool samples collected prospectively from pediatric HSCT recipients at risk for HAdV infections. Thirty-eight samples were HAdV DNA positive with both the SPD and routine preprocessing methods. HAdV DNA loads were on average 1.14-log 10 copies/g of stool higher with the SPD (P < 0.0001) than with routine methods. This new device enabled a standardized preparation of stool samples in <5 min and a reproducible and sensitive quantification of HAdV DNA. The use of the SPD for the detection of other gastrointestinal infections warrants further evaluation. PCR is a very rapid and reliable tool for molecular biologybased diagnosis of a variety of infectious diseases, using a wide range of clinical sample types (1).However, stool samples are among the most difficult clinical samples to process for molecular pathogen detection because of the presence of very potent inhibitors of nucleic acid (NA) amplification that are often coextracted along with pathogen NAs (2-4). Bile salts, hemoglobin, polysaccharides, heme, and bilirubin have been identified as factors that inhibit amplification assays (3, 5-8). Inhibitory effects can be reduced by adding amplification facilitators, such as bovine serum albumin, to the PCR mixture (9) by using thermostable polymerases that are more resistant to PCR inhibition (10), or by using more efficient processes for extracting NAs from stool samples (11). The efficiency of NA extraction and purification has been shown to influence the sensitivity, reproducibility, and accuracy of NA amplification-based target detection (12).Due to the potential presence of amplification inhibitors, sample preprocessing before NA extraction is an essential step to improve amplification-based pathogen detection from stool samples. Currently, stool preparation is not standardized and can be labor-...

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Development and assessment of molecular diagnostic tests for 15 enteropathogens causing childhood diarrhoea: a multicentre study

Cited by 277 publications

References 26 publications

The effect of azithromycin on the immunogenicity of oral poliovirus vaccine: a double-blind randomised placebo-controlled trial in seronegative Indian infants

The effect of azithromycin on the immunogenicity of oral poliovirus vaccine: a double-blind randomised placebo-controlled trial in seronegative Indian infants

Gut Microbiome Composition in Young Nicaraguan Children During Diarrhea Episodes and Recovery

Evaluation of a New Device for Simplifying and Standardizing Stool Sample Preparation for Viral Molecular Testing with Limited Hands-On Time

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