Performance of four commercial real-time PCR assays for the detection of bacterial enteric pathogens in clinical samples

Berenger, Byron M.; Chui, Linda; Ferrato, Christina; Lloyd, Tracie; Li, Vincent; Pillai, Dylan R.

doi:10.1016/j.ijid.2021.10.035

Cited by 11 publications

(9 citation statements)

References 38 publications

(57 reference statements)

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“…A poor performance of Y. enterocolitica detection and lack of non- Y. enterocolitica detection was demonstrated by assessing four commercially available real-time PCR systems, including the BD MAX™ system ( 30 ). The poor agreement observed in the study of the four PCR systems for detection of Y. enterocolitica might be explained by known heterogeneity between strains and different choices of chromosomal target genes such as ail , for detection of pathogenic Y. enterocolitica , and yst B, which is also present in most BT1A strains ( 31 , 32 ).…”

Section: Discussionmentioning

confidence: 99%

“…A poor performance of Y. enterocolitica detection and lack of non-Y. enterocolitica detection was demonstrated by assessing four commercially available real-time PCR systems, including the BD MAX TM system (30). The poor agreement observed in the study Analysis of sequence data of Y. enterocolitica O:3/BT4 from sample C2 derived from Ion Torrent and Illumina showed that virulence factors involved in the pathogenicity of Y. enterocolitica, YadA, VirF, and the Yops, carried on a plasmid, were present in the first sequencing data from Ion Torrent and absent in the later sequencing performed using the Illumina platform.…”

Section: Discussionmentioning

confidence: 99%

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One Health surveillance—A cross-sectoral detection, characterization, and notification of foodborne pathogens

Lahti

Karamehmedovic

Riedel

et al. 2023

Front. Public Health

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IntroductionSeveral Proficiency Test (PT) or External Quality Assessment (EQA) schemes are currently available for assessing the ability of laboratories to detect and characterize enteropathogenic bacteria, but they are usually targeting one sector, covering either public health, food safety or animal health. In addition to sector-specific PTs/EQAs for detection, cross-sectoral panels would be useful for assessment of the capacity to detect and characterize foodborne pathogens in a One Health (OH) perspective and further improving food safety and interpretation of cross-sectoral surveillance data. The aims of the study were to assess the cross-sectoral capability of European public health, animal health and food safety laboratories to detect, characterize and notify findings of the foodborne pathogens Campylobacter spp., Salmonella spp. and Yersinia enterocolitica, and to develop recommendations for future cross-sectoral PTs and EQAs within OH. The PT/EQA scheme developed within this study consisted of a test panel of five samples, designed to represent a theoretical outbreak scenario.MethodsA total of 15 laboratories from animal health, public health and food safety sectors were enrolled in eight countries: Denmark, France, Italy, the Netherlands, Poland, Spain, Sweden, and the United Kingdom. The laboratories analyzed the samples according to the methods used in the laboratory and reported the target organisms at species level, and if applicable, serovar for Salmonella and bioserotype for Yersinia.ResultsAll 15 laboratories analyzed the samples for Salmonella, 13 for Campylobacter and 11 for Yersinia. Analytical errors were predominately false negative results. One sample (S. Stockholm and Y. enterocolitica O:3/BT4) with lower concentrations of target organisms was especially challenging, resulting in six out of seven false negative results. These findings were associated with laboratories using smaller sample sizes and not using enrichment methods. Detection of Salmonella was most commonly mandatory to notify within the three sectors in the eight countries participating in the pilot whereas findings of Campylobacter and Y. enterocolitica were notifiable from human samples, but less commonly from animal and food samples.DiscussionThe results of the pilot PT/EQA conducted in this study confirmed the possibility to apply a cross-sectoral approach for assessment of the joint OH capacity to detect and characterize foodborne pathogens.

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Section: Discussionmentioning

confidence: 99%

“…A poor performance of Y. enterocolitica detection and lack of non-Y. enterocolitica detection was demonstrated by assessing four commercially available real-time PCR systems, including the BD MAX TM system (30). The poor agreement observed in the study Analysis of sequence data of Y. enterocolitica O:3/BT4 from sample C2 derived from Ion Torrent and Illumina showed that virulence factors involved in the pathogenicity of Y. enterocolitica, YadA, VirF, and the Yops, carried on a plasmid, were present in the first sequencing data from Ion Torrent and absent in the later sequencing performed using the Illumina platform.…”

Section: Discussionmentioning

confidence: 99%

One Health surveillance—A cross-sectoral detection, characterization, and notification of foodborne pathogens

Lahti

Karamehmedovic

Riedel

et al. 2023

Front. Public Health

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“…Not all on-site applications reported provide the required low detection limit as demonstrated by our method since reducing the complexity of the method can compromise the detection sensitivity. Additionally, several commercial methods currently exist for the detection of STEC, ranging from nucleic acid-based techniques to immunoassays [37][38][39]. Our described strategy is similarly able to detect both toxins in a manner that is specific.…”

Section: Discussionmentioning

confidence: 99%

Isothermal Amplification and Lateral Flow Nucleic Acid Test for the Detection of Shiga Toxin-Producing Bacteria for Food Monitoring

et al. 2022

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Foodborne bacteria have persisted as a significant threat to public health and to the food and agriculture industry. Due to the widespread impact of these pathogens, there has been a push for the development of strategies that can rapidly detect foodborne bacteria on-site. Shiga toxin-producing E. coli strains (such as E. coli O157:H7, E. coli O121, and E. coli O26) from contaminated food have been a major concern. They carry genes stx1 and/or stx2 that produce two toxins, Shiga toxin 1 and Shiga toxin 2, which are virulent proteins. In this work, we demonstrate the development of a rapid test based on an isothermal recombinase polymerase amplification reaction for two Shiga toxin genes in a single reaction. Results of the amplification reaction are visualized simultaneously for both Shiga toxins on a single lateral flow paper strip. This strategy targets the DNA encoding Shiga toxin 1 and 2, allowing for broad detection of any Shiga toxin-producing bacterial species. From sample to answer, this method can achieve results in approximately 35 min with a detection limit of 10 CFU/mL. This strategy is sensitive and selective, detecting only Shiga toxin-producing bacteria. There was no interference observed from non-pathogenic or pathogenic non-Shiga toxin-producing bacteria. A detection limit of 10 CFU/mL for Shiga toxin-producing E. coli was also obtained in a food matrix. This strategy is advantageous as it allows for timely identification of Shiga toxin-related contamination for quick initial food contamination assessments.

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“…The stx2f subtype is the most divergent (by nucleotide and amino acid sequence) of the known Stx2 subtypes [30], showing very low homology with other subtypes. Assay evaluation studies have shown that certain in-house and commercial PCR assays, and immunoassays targeting gastrointestinal pathogens including STEC, do not detect stx2f [29,[31][32][33][34], leading to under reporting of this variant. Given the uncertainty as to whether E. coli encoding stx2f can cause severe clinical outcomes and the limitations of the current diagnostic assays, we aimed to analyse the laboratory data linked to isolates of stx2f-positive E. coli in the UKHSA archive, review the clinical outcome data and assess the public health burden of strains of Stx2f-producing E. coli in England.…”

Section: Introductionmentioning

confidence: 99%

Escherichia coli encoding Shiga toxin subtype Stx2f causing human infections in England, 2015–2022

Den Ouden,

Greig,

Rodwell

et al. 2023

Journal of Medical Microbiology

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Introduction. Shiga toxin-producing Escherichia coli (STEC) belong to a diverse group of gastrointestinal pathogens defined by the presence of Shiga toxin genes (stx) of which there are at least ten subtypes (Stx1a-Stx1d and Stx2a-Stx2g). Gap Statement. Initially thought to be associated with mild symptoms, more recently STEC encoding stx2f have been isolated from cases of haemolytic uraemic syndrome (HUS) and the clinical significance and public health burden require further investigation. Aim. We analysed clinical outcomes and genome-sequencing data linked to patients infected with STEC encoding-stx2f in England to assess the risk to public health. Methodology. One hundred and twelve E. coli (n=58 isolates encoded stx2f; n=54 isolates E. coli belonging to CC122 or CC722 that had eae but were negative for stx) isolated from patients' faecal specimens between 2015 and 2022 were genome sequenced and linked to epidemiological and clinical outcome data. All isolates were investigated for the presence of virulence genes and a maximum-likelihood phylogeny of isolates belonging to CC122 and CC722 was constructed. Results. There were 52 cases infected with STEC harbouring stx2f between 2015 and 2022, with the majority identified in 2022. Most cases resided in the North of England (n=39/52, 75 %), were female (n=31, 59.6 %) and/or aged five and under (n=29, 55.8 %). Clinical outcome data were available for 40/52 cases (76.9 %) and 7/40(17.5 %) were diagnosed with STEC-HUS. In the two most common clonal complexes, CC122 and CC722, the presence of the stx2f-encoding prophage correlated with the presence of additional virulence genes, astA, bfpA and cdt, located on an 85kbp IncFIB plasmid. Conclusions. Certain serotypes of E. coli harbouring stx2f cause severe clinical outcomes, including STEC-HUS. Public health advice and possible interventions are limited, as little is known about the animal and environmental reservoirs and transmission routes. We recommend more comprehensive and standardized collection of microbiological and epidemiological data, and routine sharing of sequencing data between public health agencies worldwide.

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Performance of four commercial real-time PCR assays for the detection of bacterial enteric pathogens in clinical samples

Cited by 11 publications

References 38 publications

One Health surveillance—A cross-sectoral detection, characterization, and notification of foodborne pathogens

One Health surveillance—A cross-sectoral detection, characterization, and notification of foodborne pathogens

Isothermal Amplification and Lateral Flow Nucleic Acid Test for the Detection of Shiga Toxin-Producing Bacteria for Food Monitoring

Escherichia coli encoding Shiga toxin subtype Stx2f causing human infections in England, 2015–2022

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