Clinical Evaluation of a Real-Time PCR Assay for Identification of Salmonella, Shigella, Campylobacter (Campylobacter jejuni and C. coli), and Shiga Toxin-Producing Escherichia coli Isolates in Stool Specimens

Buchan, Blake W.; Olson, Wendy; Pezewski, Michael; Marcon, Mario J.; Novicki, Thomas J.; Uphoff, Timothy S.; Chandramohan, Lakshmi; Revell, Paula A.; Ledeboer, Nathan A.

doi:10.1128/jcm.02056-13

Cited by 64 publications

(52 citation statements)

References 26 publications

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“…A review of the literature indicates that the performance of the BD Max EBP assay is similar to those of other multiplex assays developed recently. The sensitivity and specificity of the Prodesse ProGastro SSCS assay compared to culture and EIA determined by one multicenter study of 1,244 specimens were 100% and Ն99.4%, respectively, for all four bacterial targets (9). Early studies of the Luminex xTAG GPP assay demonstrated sensitivities ranging from 83 to 93% for Salmonella spp., 90 to 97% for Campylobacter spp., 93 to 100% for Shigella spp., and 94% to 100% for STEC (6,21).…”

Section: Discussionmentioning

confidence: 99%

“…These systems use a variety of approaches and differ as to the number and type of targets incorporated in the assay and the overall platform design and throughput. The Prodesse ProGastro SSCS assay (Gen-Probe Prodesse, San Diego, CA) detects the same pathogens as those detected by the BD Max EBP assay, but unlike the BD Max assay, the ProGastro SSCS assay requires external extraction on a separate instrument, increasing the hands-on time and technical complexity (9). Other systems have been designed to detect viral and parasitic enteric pathogens as well as additional bacterial targets.…”

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confidence: 99%

“…Other systems have been designed to detect viral and parasitic enteric pathogens as well as additional bacterial targets. The Luminex xTag gastrointestinal pathogen panel (GPP) assay (Luminex Corp., Austin, TX) can detect up to 15 targets (9 bacteria, 3 viruses, and 3 parasites) (6,10), and the FilmArray gastrointestinal (GI) panel (Biofire Diagnostics, Inc., Salt Lake City, UT) contains 23 targets, including 14 bacterial, 5 viral, and 4 parasitic targets (9). The Luminex methodology is a high-throughput assay but also requires an external extraction step.…”

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Multicenter Evaluation of the BD Max Enteric Bacterial Panel PCR Assay for Rapid Detection of Salmonella spp., Shigella spp., Campylobacter spp. (C. jejuni and C. coli), and Shiga Toxin 1 and 2 Genes

et al. 2015

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i Diarrhea due to enteric bacterial pathogens causes significant morbidity and mortality in the United States and worldwide. However, bacterial pathogens may be infrequently identified. Currently, culture and enzyme immunoassays (EIAs) are the primary methods used by clinical laboratories to detect enteric bacterial pathogens. We conducted a multicenter evaluation of the BD Max enteric bacterial panel (EBP) PCR assay in comparison to culture for the detection of Salmonella spp., Shigella spp., Campylobacter jejuni, and Campylobacter coli and an EIA for Shiga toxins 1 and 2. A total of 4,242 preserved or unpreserved stool specimens, including 3,457 specimens collected prospectively and 785 frozen, retrospective samples, were evaluated. Compared to culture or EIA, the positive percent agreement (PPA) and negative percent agreement (NPA) values for the BD Max EBP assay for all specimens combined were as follows: 97.1% and 99.2% for Salmonella spp., 99.1% and 99.7% for Shigella spp., 97.2% and 98.4% for C. jejuni and C. coli, and 97.4% and 99.3% for Shiga toxins, respectively. Discrepant results for prospective samples were resolved with alternate PCR assays and bidirectional sequencing of amplicons. Following discrepant analysis, PPA and NPA values were as follows: 97.3% and 99.8% for Salmonella spp., 99.2% and 100% for Shigella spp., 97.5% and 99.0% for C. jejuni and C. coli, and 100% and 99.7% for Shiga toxins, respectively. No differences in detection were observed for samples preserved in Cary-Blair medium and unpreserved samples. In this large, multicenter study, the BD Max EBP assay showed superior sensitivity compared to conventional methods and excellent specificity for the detection of enteric bacterial pathogens in stool specimens. The World Health Organization reports that, worldwide, there are nearly 1.7 billion cases of diarrheal disease every year and that diarrheal disease is the second leading cause of death of children Ͻ5 years old. Each year, diarrhea kills ϳ760,000 children under the age of 5 years, and most importantly, it is preventable and treatable. Diarrhea is also a leading cause of malnutrition in this same age group. Most of this disease is related to unsafe drinking water, inadequate sanitation, and poor hygiene (1, 2).Acute gastroenteritis is caused by a number of bacterial, viral, and parasitic agents (3). In the United States, noroviruses cause most cases of self-limited, acute gastroenteritis. However, Salmonella, Campylobacter, Shigella, and Shiga toxin-producing Escherichia coli (STEC) are the most common diarrheagenic bacteria, and they are usually associated with foodborne illness (4). Furthermore, Shigella is more frequently transmitted from person to person due to the low infectious dose. Importantly, agents of gastroenteritis may not be distinguished clinically. Identifying the cause of diarrhea is important for both the treatment of individual patients and public health intervention through outbreak management (5).Conventional microbiological cultures remain the gold standard...

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Multicenter Evaluation of the BD Max Enteric Bacterial Panel PCR Assay for Rapid Detection of Salmonella spp., Shigella spp., Campylobacter spp. (C. jejuni and C. coli), and Shiga Toxin 1 and 2 Genes

et al. 2015

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“…Our study also provides real-world data in which informed considerations can be made on whether and how Campylobacter case definitions should be modified in the future. Highly sensitive and more specific nucleic acid amplification assays have more recently been approved by the FDA and may be preferable to stool antigen assays as alternatives to culture for detecting Campylobacter in stool (23)(24)(25)(26).…”

Section: Discussionmentioning

confidence: 99%

Multicenter Evaluation of Clinical Diagnostic Methods for Detection and Isolation of Campylobacter spp. from Stool

et al. 2016

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The use of culture-independent diagnostic tests (CIDTs), such as stool antigen tests, as standalone tests for the detection of Campylobacter in stool is increasing. We conducted a prospective, multicenter study to evaluate the performance of stool antigen CIDTs compared to culture and PCR for Campylobacter detection. Between July and October 2010, we tested 2,767 stool specimens from patients with gastrointestinal illness with the following methods: four types of Campylobacter selective media, four commercial stool antigen assays, and a commercial PCR assay. Illnesses from which specimens were positive by one or more culture media or at least one CIDT and PCR were designated "cases." A total of 95 specimens (3.4%) met the case definition. The stool antigen CIDTs ranged from 79.6% to 87.6% in sensitivity, 95.9 to 99.5% in specificity, and 41.3 to 84.3% in positive predictive value. Culture alone detected 80/89 (89.9% sensitivity) Campylobacter jejuni/Campylobacter coli-positive cases. Of the 209 noncases that were positive by at least one CIDT, only one (0.48%) was positive by all four stool antigen tests, and 73% were positive by just one stool antigen test. The questionable relevance of unconfirmed positive stool antigen CIDT results was supported by the finding that noncases were less likely than cases to have gastrointestinal symptoms. Thus, while the tests were convenient to use, the sensitivity, specificity, and positive predictive value of Campylobacter stool antigen tests were highly variable. Given the relatively low incidence of Campylobacter disease and the generally poor diagnostic test characteristics, this study calls into question the use of commercially available stool antigen CIDTs as standalone tests for direct detection of Campylobacter in stool. Campylobacter infection continues to be a major public health problem. Campylobacter jejuni and Campylobacter coli are pathogens transmitted commonly through food, causing an estimated 1.3 million cases of illness per year in the United States (1), and yet diagnosis can be challenging because the organism is difficult to isolate, grow, and identify. Recent reports describing clinical laboratory practices for Campylobacter diagnostics in Pennsylvania (2) and the Foodborne Diseases Active Surveillance Network (FoodNet) sites (3) highlight the wide range of testing practices in use; currently, no best-practice clinical or public health laboratory guidelines exist for laboratory diagnosis of Campylobacter infections. Direct plating onto a Campylobacter selective medium, followed by incubation at 42°C under microaerobic conditions for 72 h, has long been considered the "gold standard" for diagnosis (4).The use of culture-independent diagnostic tests (CIDTs) for Campylobacter testing on stool samples is increasing, which may have important implications for both patient management and public health surveillance efforts (5). Stool antigen tests to directly detect Campylobacter in fecal samples are fast and generate sameday results, but concerns regarding speci...

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“…Given the low prevalence of STEC infections (1 to 2%) (11)(12)(13), the associated costs for clinical laboratories for personnel, equipment, and reagents are major barriers for implementation of the recommended universal screening of stool for STEC (4,14). To reduce testing costs, we evaluated the feasibility of a pooled nucleic acid amplification test (NAAT) as an approach for low-cost high-throughput screening for stx 1 and stx 2 from stool.…”

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Pooled Nucleic Acid Amplification Test for Screening of Stool Specimens for Shiga Toxin-Producing Escherichia coli

et al. 2016

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c Shiga toxin-producing Escherichia coli (STEC)-associated enteric illness is attributed to O157 and non-O157 serotypes; however, traditional culture-based methods underdetect non-O157 STEC. Labor and cost of consumables are major barriers to implementation of the CDC recommendation to test all stools for both O157 and non-O157 serotypes. We evaluated the feasibility of a pooled nucleic acid amplification test (NAAT) as an approach for screening stool specimens for STEC. For retrospective evaluation, 300 stool specimens were used to create pools of 10 samples each. The sensitivity was 83% for the preenrichment pooling strategy and 100% for the postenrichment pooling strategy compared with those for individual NAAT results. The difference in cycle threshold (C T ) between individual and pooled NAAT results for specimens was significantly lower and more consistent for postenrichment pooling (stx 1 mean ‫؍‬ 3.90, stx 2 mean ‫؍‬ 4.28) than those for preenrichment pooling (excluding undetected specimens; stx 1 mean ‫؍‬ 9.34, stx 2 mean ‫؍‬ 8.96) (P < 0.0013). Cost of consumables and labor time savings of 48 to 81% and 6 to 66%, respectively, were estimated for the testing of 90 specimens by the postenrichment pooled NAAT strategy on the basis of an expected 1 to 2% positivity rate. A 30-day prospective head-to-head clinical trial involving 512 specimens confirmed the sensitivity and labor savings associated with the postenrichment pooled NAAT strategy. The postenrichment pooled NAAT strategy described here is suitable for efficient large-scale surveillance of all STEC serotypes. Comprehensive detection of STEC will result in accurate estimation of STEC burden and, consequently, appropriate public health interventions. Shiga toxin-producing Escherichia coli (STEC) is a major cause of sporadic and outbreak-associated enteric illness worldwide. While more than 100 STEC serotypes can cause illness in humans (1), traditional testing methods focus primarily on the most common serotype, O157:H7, owing to the ease of its detection using culture-based media. These methods underdetect non-O157 serogroups, and as such, their clinical burden is not well understood (2-4). Moreover, recent evidence suggests that infections attributed to non-O157 STEC may be more prevalent than those attributed to O157 (1, 5), with common serogroups being O26, O45, O103, O111, O121, and O145 in the United States (6, 7) and in other countries (1). Although non-O157 STEC serotypes are generally associated with milder disease than O157 STEC, infections caused by non-O157 STEC can also lead to hemolytic uremic syndrome (1) and have been associated with major outbreaks, most notably a 2011 outbreak in Germany that involved 3,816 cases and 54 deaths (8). Enhanced surveillance evidence is needed to determine the true burden of non-O157 STEC for public health investigations of potential exposure sources. Such efforts are ongoing in Canada (9) but require improved and comprehensive screening methods.The Centers for Disease Control and Prevention (CDC) recom...

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Clinical Evaluation of a Real-Time PCR Assay for Identification of Salmonella, Shigella, Campylobacter (Campylobacter jejuni and C. coli), and Shiga Toxin-Producing Escherichia coli Isolates in Stool Specimens

Cited by 64 publications

References 26 publications

Multicenter Evaluation of the BD Max Enteric Bacterial Panel PCR Assay for Rapid Detection of Salmonella spp., Shigella spp., Campylobacter spp. (C. jejuni and C. coli), and Shiga Toxin 1 and 2 Genes

Multicenter Evaluation of the BD Max Enteric Bacterial Panel PCR Assay for Rapid Detection of Salmonella spp., Shigella spp., Campylobacter spp. (C. jejuni and C. coli), and Shiga Toxin 1 and 2 Genes

Multicenter Evaluation of Clinical Diagnostic Methods for Detection and Isolation of Campylobacter spp. from Stool

Pooled Nucleic Acid Amplification Test for Screening of Stool Specimens for Shiga Toxin-Producing Escherichia coli

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