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LETTERS TO THE EDITOR

Use of a Culture-Independent Gastrointestinal Multiplex PCR Panel during a Shigellosis Outbreak: Considerations for Clinical Laboratories and Public Health

Vittal P. Prakash, Lindsay LeBlanc, Nicole E. Alexander-Scott, Jaclyn Skidmore, Deanna Simmons, Daniela Quilliam, Kimberle C. Chapin
E. Munson, Editor
Vittal P. Prakash
aDepartment of Pathology, Rhode Island Hospital, Providence, Rhode Island, USA
bDepartment of Pathology and Laboratory Medicine, Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA
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Lindsay LeBlanc
aDepartment of Pathology, Rhode Island Hospital, Providence, Rhode Island, USA
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Nicole E. Alexander-Scott
cRhode Island Department of Health, Providence, Rhode Island, USA
dDivision of Pediatric/Adult Infectious Diseases, Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA
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Jaclyn Skidmore
cRhode Island Department of Health, Providence, Rhode Island, USA
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Deanna Simmons
cRhode Island Department of Health, Providence, Rhode Island, USA
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Daniela Quilliam
cRhode Island Department of Health, Providence, Rhode Island, USA
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Kimberle C. Chapin
aDepartment of Pathology, Rhode Island Hospital, Providence, Rhode Island, USA
bDepartment of Pathology and Laboratory Medicine, Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA
eDepartment of Medicine, Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA
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E. Munson
Roles: Editor
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DOI: 10.1128/JCM.03374-14
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LETTER

Shigella is estimated to cause nearly 500,000 infections annually in the United States (1). This estimate for the nationally notifiable disease is much higher than the actual 7,746 laboratory-confirmed cases of Shigella reported to the Centers for Disease Control and Prevention (CDC) by state public health laboratories in 2012 (2). The true incidence of Shigella infections is likely underrecognized due to incorrect preanalytical steps, including specimen collection, transfer into stool preservative medium (e.g., Cary-Blair), and timely transport to the laboratory, as well as limit of detection with culture, or a “viable but non-culturable” (VBNC) state that might render stool culture insensitive (3).

In early July 2013, the Rhode Island Department of Health (RIDOH) noted a significant increase in reports of diarrheal illness, which were subsequently identified as a shigellosis outbreak (4, 5). During this same time, the Rhode Island Hospital (RIH) laboratory was conducting an institutional review board-approved prospective research study evaluating the FilmArray gastrointestinal (GI) panel (BioFire Diagnostics, Salt Lake City, UT). The GI panel is a culture-independent diagnostic test (CIDT) that simultaneously detects multiple bacterial, viral, toxin gene, and parasitic targets, including Shigella/enteroinvasive Escherichia coli (EIEC) using a nested multiplex PCR directly from stool specimens collected in Cary-Blair transport medium. During a short period of the outbreak, specimens tested at the RIH laboratory were analyzed via both conventional stool culture and the FilmArray GI panel. Comparing the methods, a number of potential issues related to timing of pathogen detection and subsequent interpretation and/or reporting of cases to public health were noted. Specifically, the GI panel detected 10 cases of Shigella/EIEC compared to only 6 Shigella sonnei cases identified by culture, a 40% increase in positive cases (Table 1). Stool samples positive for Shigella/EIEC by the GI panel were further confirmed by an independent PCR assay and bidirectional sequencing (6), and while the GI panel does not distinguish between Shigella and EIEC, given the timing of the specimens during the outbreak, these patients were all likely to have Shigella and determined to be true positive cases. However, the public health lab determined only the six culture-positive Shigella isolates as true positive cases based on the current definition for defining an outbreak in the state of Rhode Island. Three other notable and not surprising findings of the FilmArray GI panel were as follows. (i) Shigellosis cases were identified more quickly than conventional culture for most of the cases (hours from onset of testing compared to days) despite the fact that the molecular assay was not always performed the day the specimen was received. (ii) The majority of patient specimens that were negative by culture were from outpatients (as opposed to emergency room [ER] or hospitalized patients) where culture may have been compromised during collection and/or transit to the testing site. (iii) Finally, patients positive for Shigella had additional potential GI pathogens detected. Specifically, in 40% of cases, enteroaggregative E. coli (EAEC), enteropathogenic E. coli (EPEC), Shiga toxin-producing E. coli, norovirus, and sapovirus were identified that would not have been detected by routine methods. Previously noted in international studies in children coinfected with viral and bacterial GI pathogens, more-severe diarrhea may develop than in those monoinfected with either pathogen (7, 8). As CIDTs become more commonly used (9), better understanding of pathogenicity, disease severity, and host response will be required in order to accurately interpret the implications of coinfection patterns. Subsequent to the study, discordant GI panel results compared to culture allowed a dialogue between the clinical laboratory and RIDOH on the issues of CIDT methods and public health reporting. Current Rhode Island public health definitions and methodologies continue to require culture and submission of an isolate for determining pulsed-field gel electrophoresis (PFGE) pattern linkages in an outbreak. Since a number of GI panels are now FDA cleared, CIDTs could be considered a primary screening tool with culture isolation as a reflexive test for positive specimens to address the public health needs to effectively identify disease linkages. To be determined is how clinical and public health laboratories will address the conflicting results and reporting of positive CIDT results with negative culture results for notifiable conditions until other confirmatory methods are routinely available.

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TABLE 1

Testing results of specimens/isolates associated with the shigellosis outbreak

(This work was presented in part at the 114th General Meeting of the American Society for Microbiology, Boston, MA, 17 to 20 May 2014.)

ACKNOWLEDGMENTS

We thank Michael Gosciminski, Adam Miller, and Sharon Mallard at the RIDOH for their assistance with PFGE and epidemiology work for this study. We thank Kim Hyun, RIH, for helping with the FilmArray assay.

This work was supported by a NIH/NIAID grant R01 A1 104593 and Bio-Fire Diagnostics, Inc.

  • Copyright © 2015, American Society for Microbiology. All Rights Reserved.

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Use of a Culture-Independent Gastrointestinal Multiplex PCR Panel during a Shigellosis Outbreak: Considerations for Clinical Laboratories and Public Health
Vittal P. Prakash, Lindsay LeBlanc, Nicole E. Alexander-Scott, Jaclyn Skidmore, Deanna Simmons, Daniela Quilliam, Kimberle C. Chapin
Journal of Clinical Microbiology Feb 2015, 53 (3) 1048-1049; DOI: 10.1128/JCM.03374-14

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Use of a Culture-Independent Gastrointestinal Multiplex PCR Panel during a Shigellosis Outbreak: Considerations for Clinical Laboratories and Public Health
Vittal P. Prakash, Lindsay LeBlanc, Nicole E. Alexander-Scott, Jaclyn Skidmore, Deanna Simmons, Daniela Quilliam, Kimberle C. Chapin
Journal of Clinical Microbiology Feb 2015, 53 (3) 1048-1049; DOI: 10.1128/JCM.03374-14
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