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de-la-Rosa-Martinez D, Bobadilla Del Valle M, Esteban-Kenel V, Zinser Peniche P, Ponce De León Garduño A, Cornejo Juárez P, Sánchez Cruz MN, Camacho-Ortiz A, Vilar-Compte D. Molecular characterization and genotyping of isolates from cancer patients with Clostridioides difficile infection or asymptomatic colonization. J Med Microbiol 2023; 72. [PMID: 37624363 DOI: 10.1099/jmm.0.001748] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 08/26/2023] Open
Abstract
Introduction. Cancer patients with Clostridioides difficile infection (CDI) are at a higher risk for adverse outcomes. In addition, a high prevalence of Clostridioides difficile asymptomatic colonization (CDAC) has been reported in this vulnerable population.Gap Statement. The molecular characteristics and potential role of CDAC in healthcare-related transmission in the cancer population have been poorly explored.Aim. We aimed to compare the molecular and genotypic characteristics of C. difficile isolates from cancer patients with CDAC and CDI.Method. We conducted a prospective cohort study of cancer patients with CDAC or CDI from a referral centre. Molecular characterization, typification and tcdC gene expression of isolates were performed.Results. The hospital-onset and community-onset healthcare facility-associated CDI rates were 4.5 cases/10 000 patient-days and 1.4 cases/1 000 admissions during the study period. Fifty-one C. difficile strains were isolated: 37 (72 %) and 14 (28 %) from patients with CDI or CDAC, respectively. All isolates from symptomatic patients were tcdA+/tcdB+, and four (10 %) were ctdA+/ctdB+. In the CDAC group, 10 (71 %) isolates were toxigenic, and none were ctdA+/ctdB+. The Δ18 in-frame tcdC deletion and two transition mutations were found in five isolates. After bacterial typing, 60 % of toxigenic isolates from asymptomatic carriers were clonal to those from patients with C. difficile-associated diarrhoea. No NAP1/027/BI strains were detected.Conclusions. We found a clonal association between C. difficile isolates from patients with CDAC and CDI. Studies are needed to evaluate the potential role of asymptomatic carriers in the dynamics of nosocomial transmission to support infection control measures and reduce the burden of CDI in high-risk groups.
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Affiliation(s)
- Daniel de-la-Rosa-Martinez
- Plan de Estudios Combinados en Medicina (PECEM), Faculty of Medicine, Universidad Nacional Autonoma de Mexico, México City, Mexico
- Departament of Infectious Diseases, Instituto Nacional de Cancerología, Mexico City, Mexico
| | - Miriam Bobadilla Del Valle
- Laboratory of Clinical Microbiology, Departament of Infectious Diseases, Instituto Nacional de Ciencias Medicas y Nutrición, Mexico City, Mexico
| | - Veronica Esteban-Kenel
- Laboratory of Clinical Microbiology, Departament of Infectious Diseases, Instituto Nacional de Ciencias Medicas y Nutrición, Mexico City, Mexico
| | - Paola Zinser Peniche
- Departament of Infectious Diseases, Instituto Nacional de Cancerología, Mexico City, Mexico
| | - Alfredo Ponce De León Garduño
- Laboratory of Clinical Microbiology, Departament of Infectious Diseases, Instituto Nacional de Ciencias Medicas y Nutrición, Mexico City, Mexico
| | | | - María Nancy Sánchez Cruz
- Laboratory of Clinical Microbiology, Departament of Infectious Diseases, Instituto Nacional de Ciencias Medicas y Nutrición, Mexico City, Mexico
| | - Adrian Camacho-Ortiz
- Department of Infectious Diseases, Department of Internal Medicine, Hospital Universitario Dr. Jose Eleuterio Gonzalez, Universidad Autónoma de Nuevo León, Monterrey, Mexico
| | - Diana Vilar-Compte
- Departament of Infectious Diseases, Instituto Nacional de Cancerología, Mexico City, Mexico
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Bocchetti M, Ferraro MG, Melisi F, Grisolia P, Scrima M, Cossu AM, Yau TO. Overview of current detection methods and microRNA potential in Clostridioides difficile infection screening. World J Gastroenterol 2023; 29:3385-3399. [PMID: 37389232 PMCID: PMC10303512 DOI: 10.3748/wjg.v29.i22.3385] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/25/2023] [Revised: 03/23/2023] [Accepted: 05/04/2023] [Indexed: 06/06/2023] Open
Abstract
Clostridioides difficile (formerly called Clostridium difficile, C. difficile) infection (CDI) is listed as an urgent threat on the 2019 antibiotic resistance threats report in the United States by the Centers for Disease Control and Prevention. Early detection and appropriate disease management appear to be essential. Meanwhile, although the majority of cases are hospital-acquired CDI, community-acquired CDI cases are also on the rise, and this vulnerability is not limited to immunocompromised patients. Gastrointestinal treatments and/or gastrointestinal tract surgeries may be required for patients diagnosed with digestive diseases. Such treatments could suppress or interfere with the patient's immune system and disrupt gut flora homeostasis, creating a suitable microecosystem for C. difficile overgrowth. Currently, stool-based non-invasive screening is the first-line approach to CDI diagnosis, but the accuracy is varied due to different clinical microbiology detection methods; therefore, improving reliability is clearly required. In this review, we briefly summarised the life cycle and toxicity of C. difficile, and we examined existing diagnostic approaches with an emphasis on novel biomarkers such as microRNAs. These biomarkers can be easily detected through non-invasive liquid biopsy and can yield crucial information about ongoing pathological phenomena, particularly in CDI.
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Affiliation(s)
- Marco Bocchetti
- Department of Precision Medicine, University of Campania “Luigi Vanvitelli,” Naples 80138, Italy
- Department of Molecular Oncology, Precision Medicine Laboratory and COVID19 Laboratory, Biogem Scarl, Ariano Irpino 83031, Italy
| | - Maria Grazia Ferraro
- School of Infection and Immunity, College of Medical Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, United Kingdom
- Department of Pharmacy, School of Medicine and Surgery, University of Naples “Federico II,” Naples 80131, Italy
| | - Federica Melisi
- Department of Precision Medicine, University of Campania “Luigi Vanvitelli,” Naples 80138, Italy
- Department of Molecular Oncology, Precision Medicine Laboratory and COVID19 Laboratory, Biogem Scarl, Ariano Irpino 83031, Italy
| | - Piera Grisolia
- Department of Precision Medicine, University of Campania “Luigi Vanvitelli,” Naples 80138, Italy
- Department of Molecular Oncology, Precision Medicine Laboratory and COVID19 Laboratory, Biogem Scarl, Ariano Irpino 83031, Italy
| | - Marianna Scrima
- Department of Molecular Oncology, Precision Medicine Laboratory and COVID19 Laboratory, Biogem Scarl, Ariano Irpino 83031, Italy
| | - Alessia Maria Cossu
- Department of Precision Medicine, University of Campania “Luigi Vanvitelli,” Naples 80138, Italy
- Department of Molecular Oncology, Precision Medicine Laboratory and COVID19 Laboratory, Biogem Scarl, Ariano Irpino 83031, Italy
| | - Tung On Yau
- School of Science and Technology, Nottingham Trent University, Nottingham NG11 8NS, United Kingdom
- Department of Rural Land Use, Scotland’s Rural College, Aberdeen AB21 9YA, Scotland, United Kingdom
- Department of Health Science, University of the People, Pasadena, CA 9110112, United States
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Clinical evaluation of a non-purified direct molecular assay for the detection of Clostridioides difficile toxin genes in stool specimens. PLoS One 2020; 15:e0234119. [PMID: 32492051 PMCID: PMC7269250 DOI: 10.1371/journal.pone.0234119] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2020] [Accepted: 05/19/2020] [Indexed: 12/22/2022] Open
Abstract
Recently, a new rapid assay for the detection of tcdB gene of Clostridioides difficile was developed using the GENECUBE. The assay can directly detect the tcdB gene from stool samples without a purification in approximately 35 minutes with a few minutes of preparation process. We performed a prospective comparative study of the performance of the assay at eight institutions in Japan. Fresh residual stool samples (Bristol stool scale ≥5) were used and comparisons were performed with the BD MAX Cdiff assay and toxigenic cultures. For the evaluation of 383 stool samples compared with the BD MAX Cdiff assay, the sensitivity, and specificity of the two assays was 99.0% (379/383), 98.1% (52/53), 99.1% (327/330), respectively. In the comparison with toxigenic culture, the total, sensitivity, and specificity were 96.6% (370/383), 85.0% (51/60), and 98.8% (319/323), respectively. The current investigation indicated the GENECUBE Clostridioides difficile assay has equivalent performance with the BD MAX Cdiff assay for the detection of tcdB gene of C. difficile.
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Martins JP, Felgueiras M, Santos R. The reference method influence on the sensitivity of the Clostridium difficile enzyme immunoassays: A meta analysis. J Microbiol Methods 2020; 173:105912. [PMID: 32278778 DOI: 10.1016/j.mimet.2020.105912] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2019] [Revised: 03/22/2020] [Accepted: 03/31/2020] [Indexed: 11/16/2022]
Abstract
The use of enzyme immunoassays to screen for toxins A and B produced by Clostridium difficile is a common procedure in algorithms designed for its detection. Moreover, the absence of a unique test capable of providing reliable results at low cost motivates a great discussion about which algorithm is the best. Thus, several studies have evaluated the performance of these enzyme immunoassays. However, all fail to provide sufficient explanations for the different behaviours observed in different studies that evaluate the same index test against a common reference method. Our main goal was to find out which factors affect the sensitivity of these assays, since the specificity is very close to 1. In this research, we verified that sensitivity increases with the prevalence rate and with the proportion of reported cases of onset diarrhea. Therefore, its use is advisable for high prevalence rates (e.g. in an epidemic setting). As far as reference methods are concerned, nucleic acid amplification tests can be used as a reference method, with a performance similar to the well-accepted toxigenic culture. The method chosen for toxigenicity screening in a toxigenic culture also seems to affect the evaluation performance of tests and should be better studied in the future.
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Affiliation(s)
- João Paulo Martins
- ESTG, Polytechnic Institute of Leiria, Campus 2, Morro do Lena Alto do Vieiro, Apartado 4163, 2411-901 Leiria, Portugal; CEAUL - Centre of Statistics and its Applications, Faculdade de Ciências da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal.
| | - Miguel Felgueiras
- ESTG, Polytechnic Institute of Leiria, Campus 2, Morro do Lena Alto do Vieiro, Apartado 4163, 2411-901 Leiria, Portugal; CARME, Polytechnic Institute of Leiria, Campus 2, Morro do Lena Alto do Vieiro, Apartado 4163, 2411-901 Leiria, Portugal; CEAUL - Centre of Statistics and its Applications, Faculdade de Ciências da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal
| | - Rui Santos
- ESTG, Polytechnic Institute of Leiria, Campus 2, Morro do Lena Alto do Vieiro, Apartado 4163, 2411-901 Leiria, Portugal; CEAUL - Centre of Statistics and its Applications, Faculdade de Ciências da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal
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Carroll KC, Mizusawa M. Laboratory Tests for the Diagnosis of Clostridium difficile. Clin Colon Rectal Surg 2020; 33:73-81. [PMID: 32104159 PMCID: PMC7042017 DOI: 10.1055/s-0039-3400476] [Citation(s) in RCA: 31] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Clostridium (reclassified as " Clostridioides ") difficile is an anaerobic, gram-positive bacterium that causes significant disease through elaboration of two potent toxins in patients whose normal gut microbiota has been altered through antimicrobial or chemotherapeutic agents (dysbiosis). The optimum method of laboratory diagnosis is still somewhat controversial. Recent practice guidelines published by professional societies recommend a two-step approach beginning with a test for glutamate dehydrogenase (GDH), followed by a toxin test and/or a nucleic acid test. Alternatively, in institutions where established clinical algorithms guide testing, a nucleic acid test alone is acceptable. Nucleic acid tests are the methods of choice in approximately 50% of laboratories in the United States. These tests are considered as the most sensitive methods for detection of C. difficile in stool and are the least specific. Because of the lower specificity with nucleic acid tests, some clinicians believe that toxin enzyme immunoassays are better predictors of disease, despite their known poor performance in certain patient populations. This review will discuss the advantages and disadvantages of the currently available test methods for the diagnosis of C. difficile with a brief mention of some novel assays that are currently in clinical trials.
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Affiliation(s)
- Karen C. Carroll
- Division of Medical Microbiology, Department of Pathology, the Johns Hopkins University School of Medicine, Baltimore, Maryland
- Address for correspondence Karen C. Carroll, MD Division of Medical Microbiology, Department of Pathology, the Johns Hopkins University School of MedicineMeyer B1-193, 600 North Wolfe Street, Baltimore MD 21287
| | - Masako Mizusawa
- Section of Infectious Diseases, Department of Internal Medicine, University of Missouri, Kansas City, Missouri
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Karp J, Edman-Wallér J, Toepfer M, Lundqvist A, Jacobsson G. Clostridioides difficile incidence related to in-hospital cephalosporin use: a tale of two highly comparable hospitals. J Antimicrob Chemother 2020; 74:182-189. [PMID: 30358837 DOI: 10.1093/jac/dky408] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2018] [Accepted: 09/07/2018] [Indexed: 11/14/2022] Open
Abstract
Background Antibiotic treatment is a well-known risk factor for healthcare facility-associated Clostridioides (Clostridium) difficile infection (HCF-CDI). Antibiotic stewardship programmes (ASPs) targeting high-risk antibiotics have been shown to decrease HCF-CDI incidence. HCF-CDI incidence is high in Nordic countries despite relatively low antibiotic use in hospital. Objectives To determine if HCF-CDI incidence was modified by a hospital ASP that restricted cephalosporin use. Methods The effects of an ASP on HCF-CDI incidence were evaluated in a two-centre setting using a retrospective design. We exploited a strategy of both individual case ascertainment based on chart reviews and aggregated data from the hospitals. Cases were attributed to the antibiotics given prior to disease onset, in proportion to the number of DDDs used. Three periods were studied: 2007 (before the ASP), 2012 and 2015. Results At the ASP hospital, cephalosporin use decreased by 87% and the number of HCF-CDI/1000 hospital admissions decreased significantly from 2.25 (2007) to 1.16 (2015) (P = 0.0014). The corresponding results at the non-ASP hospital showed a non-significant increase from 2.09 to 2.38. A high number of cases could be attributed to cephalosporins at both hospitals. The increased use of other broad-spectrum antibiotics, e.g. piperacillin/tazobactam, at the ASP hospital was not associated with offsetting increases in attributable HCF-CDI cases. Conclusions Decreased use of cephalosporins is an effective strategy to decrease HCF-CDI incidence over time in a setting with high incidence and low antibiotic use.
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Affiliation(s)
- Johan Karp
- Department of Infectious Diseases, Skaraborg Hospital, Lövängsvägen, Skövde, Sweden.,Center for Antibiotic Resistance Research (CARe), Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Göteborg, Sweden
| | - Jon Edman-Wallér
- Center for Antibiotic Resistance Research (CARe), Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Göteborg, Sweden.,Department of Hospital Infection Control, Södra Älvsborg Hospital, Brämhultsvägen, Borås, Sweden
| | - Michael Toepfer
- Clinical Microbiology, Unilabs AB, Rådhusgatan 6, Skövde, Sweden
| | - Anders Lundqvist
- Department of Infectious Diseases, Södra Älvsborg Hospital, Brämhultsvägen 53, Borås, Sweden
| | - Gunnar Jacobsson
- Department of Infectious Diseases, Skaraborg Hospital, Lövängsvägen, Skövde, Sweden.,Center for Antibiotic Resistance Research (CARe), Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Göteborg, Sweden
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Kraft CS, Parrott JS, Cornish NE, Rubinstein ML, Weissfeld AS, McNult P, Nachamkin I, Humphries RM, Kirn TJ, Dien Bard J, Lutgring JD, Gullett JC, Bittencourt CE, Benson S, Bobenchik AM, Sautter RL, Baselski V, Atlas MC, Marlowe EM, Miller NS, Fischer M, Richter SS, Gilligan P, Snyder JW. A Laboratory Medicine Best Practices Systematic Review and Meta-analysis of Nucleic Acid Amplification Tests (NAATs) and Algorithms Including NAATs for the Diagnosis of Clostridioides ( Clostridium) difficile in Adults. Clin Microbiol Rev 2019; 32:32/3/e00032-18. [PMID: 31142497 PMCID: PMC6589859 DOI: 10.1128/cmr.00032-18] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
The evidence base for the optimal laboratory diagnosis of Clostridioides (Clostridium) difficile in adults is currently unresolved due to the uncertain performance characteristics and various combinations of tests. This systematic review evaluates the diagnostic accuracy of laboratory testing algorithms that include nucleic acid amplification tests (NAATs) to detect the presence of C. difficile The systematic review and meta-analysis included eligible studies (those that had PICO [population, intervention, comparison, outcome] elements) that assessed the diagnostic accuracy of NAAT alone or following glutamate dehydrogenase (GDH) enzyme immunoassays (EIAs) or GDH EIAs plus C. difficile toxin EIAs (toxin). The diagnostic yield of NAAT for repeat testing after an initial negative result was also assessed. Two hundred thirty-eight studies met inclusion criteria. Seventy-two of these studies had sufficient data for meta-analysis. The strength of evidence ranged from high to insufficient. The uses of NAAT only, GDH-positive EIA followed by NAAT, and GDH-positive/toxin-negative EIA followed by NAAT are all recommended as American Society for Microbiology (ASM) best practices for the detection of the C. difficile toxin gene or organism. Meta-analysis of published evidence supports the use of testing algorithms that use NAAT alone or in combination with GDH or GDH plus toxin EIA to detect the presence of C. difficile in adults. There is insufficient evidence to recommend against repeat testing of the sample using NAAT after an initial negative result due to a lack of evidence of harm (i.e., financial, length of stay, or delay of treatment) as specified by the Laboratory Medicine Best Practices (LMBP) systematic review method in making such an assessment. Findings from this systematic review provide clarity to diagnostic testing strategies and highlight gaps, such as low numbers of GDH/toxin/PCR studies, in existing evidence on diagnostic performance, which can be used to guide future clinical research studies.
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Affiliation(s)
| | - J Scott Parrott
- Department of Interdisciplinary Studies, School of Health Professions, Rutgers University, Newark, New Jersey, USA
- Department of Epidemiology, School of Public Health, Rutgers University, Piscataway, New Jersey, USA
| | - Nancy E Cornish
- Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | | | | | - Peggy McNult
- American Society for Microbiology, Washington, DC, USA
| | - Irving Nachamkin
- Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | | | - Thomas J Kirn
- Department of Interdisciplinary Studies, School of Health Professions, Rutgers University, Newark, New Jersey, USA
- Department of Epidemiology, School of Public Health, Rutgers University, Piscataway, New Jersey, USA
| | - Jennifer Dien Bard
- Children's Hospital Los Angeles, Los Angeles, California, USA
- Keck School of Medicine, University of Southern California, Los Angeles, California, USA
| | | | - Jonathan C Gullett
- Kaiser Permanente (Southern California Permanente Medical Group) Regional Reference Laboratories, Greater Los Angeles, Los Angeles, California, USA
| | | | - Susan Benson
- PathWest Laboratory Medicine, Perth, Western Australia, Australia
- University of Western Australia, Perth, Western Australia, Australia
| | - April M Bobenchik
- Rhode Island Hospital/Lifespan Academic Medical Center, Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA
| | | | - Vickie Baselski
- University of Tennessee Health Science Center, Memphis, Tennessee, USA
| | - Michel C Atlas
- Kornhauser Health Sciences Library, University of Louisville, Louisville, Kentucky, USA
| | | | - Nancy S Miller
- Boston Medical Center, Boston, Massachusetts, USA
- Boston University School of Medicine, Boston, Massachusetts, USA
| | | | | | - Peter Gilligan
- University of North Carolina School of Medicine, Chapel Hill, North Carolina, USA
| | - James W Snyder
- Kornhauser Health Sciences Library, University of Louisville, Louisville, Kentucky, USA
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Clinical relevance of enteropathogen co-infections in preschool children-a population-based repeated cross-sectional study. Clin Microbiol Infect 2018; 25:1039.e7-1039.e13. [PMID: 30553029 DOI: 10.1016/j.cmi.2018.11.029] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2018] [Revised: 11/12/2018] [Accepted: 11/30/2018] [Indexed: 01/13/2023]
Abstract
OBJECTIVES This study aimed to (i) determine risk factors for enteropathogen co-infections, (ii) determine whether enteropathogen co-infections influence gastroenteritis risk, and (iii) determine whether enteropathogen co-infection occurred randomly in preschool children. METHODS A monthly-repeated cross-sectional survey in Dutch children aged 0-48 months was conducted during October 2012 to October 2014. A total of 981 stool samples were collected along with questionnaires collecting data on gastrointestinal symptoms and potential risk factors; 822 samples were successfully tested for 19 enteropathogens using real-time multiplex PCRs. Logistic regression analysis assessed co-infections in relation to gastroenteritis and potential risk factors. RESULTS In all, 598/822 (72.7%) stool samples tested positive for at least one enteropathogen, of which 290 (48.5%) were positive for two or more enteropathogens. Risk factors for two or more enteropathogen co-infections were young age (<12 months, OR 1.9, 95% CI 1.1-3.3; 13-36 months, OR 1.7, 95% CI 1.1-2.5, versus 37-48 months), day-care attendance (OR 1.8, 95% CI 1.3-2.5), households with three or more children versus those with one child (OR 1.7, 95% CI 1.1-2.8). Stool samples collected in spring less often had two or more enteropathogens versus summer (OR 0.4, 95% CI 0.2-0.7). Food allergy was a risk factor for three or more enteropathogen co-infections (OR 3.2, 95% CI 1.1-8.9). The frequency of co-infection was higher than expected for norovirus GI/norovirus GII, Clostridium difficile/norovirus GI, C. difficile/rotavirus, astrovirus/Dientamoeba fragilis, atypical enteropathogenic Escherichia coli/adenovirus, typical enteropathogenic E. coli/adenovirus, and enteroaggregative E. coli/astrovirus. No co-infection was associated with increased gastroenteritis risk. CONCLUSIONS Risk factors for enteropathogen co-infections were identified and specific enteropathogens co-occurred significantly more often than expected by chance. Enteropathogen co-infections were not associated with increased gastroenteritis risk, calling into question their clinical relevance in preschool children.
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Usacheva EA, Jin JP, Peterson LR. Host response to Clostridium difficile infection: Diagnostics and detection. J Glob Antimicrob Resist 2016; 7:93-101. [PMID: 27693863 PMCID: PMC5124533 DOI: 10.1016/j.jgar.2016.08.002] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2016] [Revised: 07/29/2016] [Accepted: 08/08/2016] [Indexed: 02/08/2023] Open
Abstract
Clostridium difficile infection (CDI) is a significant healthcare concern worldwide, and C. difficile is recognised as the most frequent aetiological agent of infectious healthcare-associated diarrhoea in hospitalised adult patients. The clinical manifestation of CDI varies from self-limited diarrhoea to life-threatening colitis. Such a broad disease spectrum can be explained by the impact of host factors. Currently, a complex CDI aetiology is widely accepted, acknowledging the interaction between bacteria and the host. C. difficile strains producing clostridial toxins A and B are considered toxigenic and can cause disease; those not producing the toxins are non-pathogenic. A person colonised with a toxigenic strain will not necessarily develop CDI. It is imperative to recognise patients with active disease from those only colonised with this pathogen and to implement appropriate treatment. This can be achieved by diagnostics that rely on host factors specific to CDI. This review will focus on major aspects of CDI pathogenesis and molecular mechanisms, describing host factors in disease progression and assessment of the host response in order to facilitate the development of CDI-specific diagnostics.
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Affiliation(s)
- Elena A Usacheva
- Infectious Disease Research, NorthShore University HealthSystem, 2650 Ridge Ave., Evanston, IL 60201, USA; University of Chicago Pritzker School of Medicine, Chicago, IL, USA.
| | - Jian-P Jin
- Department of Physiology, Wayne State University School of Medicine, Detroit, MI 48201, USA
| | - Lance R Peterson
- Infectious Disease Research, NorthShore University HealthSystem, 2650 Ridge Ave., Evanston, IL 60201, USA; University of Chicago Pritzker School of Medicine, Chicago, IL, USA
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Crobach MJT, Planche T, Eckert C, Barbut F, Terveer EM, Dekkers OM, Wilcox MH, Kuijper EJ. European Society of Clinical Microbiology and Infectious Diseases: update of the diagnostic guidance document for Clostridium difficile infection. Clin Microbiol Infect 2016; 22 Suppl 4:S63-81. [PMID: 27460910 DOI: 10.1016/j.cmi.2016.03.010] [Citation(s) in RCA: 387] [Impact Index Per Article: 43.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2015] [Revised: 03/02/2016] [Accepted: 03/10/2016] [Indexed: 12/14/2022]
Abstract
In 2009 the first European Society of Clinical Microbiology and Infectious Diseases (ESCMID) guideline for diagnosing Clostridium difficile infection (CDI) was launched. Since then newer tests for diagnosing CDI have become available, especially nucleic acid amplification tests. The main objectives of this update of the guidance document are to summarize the currently available evidence concerning laboratory diagnosis of CDI and to formulate and revise recommendations to optimize CDI testing. This update is essential to improve the diagnosis of CDI and to improve uniformity in CDI diagnosis for surveillance purposes among Europe. An electronic search for literature concerning the laboratory diagnosis of CDI was performed. Studies evaluating a commercial laboratory test compared to a reference test were also included in a meta-analysis. The commercial tests that were evaluated included enzyme immunoassays (EIAs) detecting glutamate dehydrogenase, EIAs detecting toxins A and B and nucleic acid amplification tests. Recommendations were formulated by an executive committee, and the strength of recommendations and quality of evidence were graded using the Grades of Recommendation Assessment, Development and Evaluation (GRADE) system. No single commercial test can be used as a stand-alone test for diagnosing CDI as a result of inadequate positive predictive values at low CDI prevalence. Therefore, the use of a two-step algorithm is recommended. Samples without free toxin detected by toxins A and B EIA but with positive glutamate dehydrogenase EIA, nucleic acid amplification test or toxigenic culture results need clinical evaluation to discern CDI from asymptomatic carriage.
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Affiliation(s)
- M J T Crobach
- Department of Medical Microbiology, Centre for Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands
| | - T Planche
- Department of Medical Microbiology, St. George's Hospital, London, UK
| | - C Eckert
- National Reference Laboratory for Clostridium difficile, Paris, France
| | - F Barbut
- National Reference Laboratory for Clostridium difficile, Paris, France
| | - E M Terveer
- Department of Medical Microbiology, Centre for Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands
| | - O M Dekkers
- Departments of Clinical Epidemiology and Internal Medicine, Leiden University Medical Center, Leiden, The Netherlands; Department of Clinical Epidemiology, Aarhus University, Aarhus, Denmark
| | - M H Wilcox
- Department of Microbiology, Leeds Teaching Hospitals & University of Leeds, Leeds, UK
| | - E J Kuijper
- Department of Medical Microbiology, Centre for Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands.
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Angione SL, Croote D, Leung JW, Mermel LA, Tripathi A. Single fluorophore melting curve analysis for detection of hypervirulent Clostridium difficile. J Med Microbiol 2015; 65:62-70. [PMID: 26516039 DOI: 10.1099/jmm.0.000199] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
This study demonstrates a novel detection assay able to identify and subtype strains of Clostridium difficile. Primers carefully designed for melting curve analysis amplify DNA from three C. difficile genes, tcdB, tcdC and cdtB, during quantitative (q)PCR. The tcdB gene allows for confirmation of organism presence, whilst the tcdC and cdtB genes allow for differentiation of virulence status, as deletions in the tcdC gene and the concurrent presence of the cdtB gene, which produces binary toxin, are associated with hypervirulence. Following qPCR, subtyping is then achieved by automated, inline melting curve analysis using only a single intercalating dye and verified by microchip electrophoresis. This assay represents a novel means of distinguishing between toxigenic and hypervirulent C. difficile strains NAP1/027/BI and 078 ribotype, which are highly prevalent hypervirulent strains in humans. This methodology can help rapidly detect and identify C. difficile strains that impose a significant health and economic burden in hospitals and other healthcare settings.
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Affiliation(s)
- Stephanie L Angione
- Center for Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA
| | - Derek Croote
- Center for Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA
| | - Joshua W Leung
- Center for Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA
| | - Leonard A Mermel
- Division of Infectious Diseases, Department of Medicine, Rhode Island Hospital, , Providence, RI, USA.,Warren Alpert Medical School of Brown University, Providence, RI, USA
| | - Anubhav Tripathi
- Center for Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA
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12
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Diagnostic yield of repeat sampling with immunoassay, real-time PCR, and toxigenic culture for the detection of toxigenic Clostridium difficile in an epidemic and a non-epidemic setting. Eur J Clin Microbiol Infect Dis 2015; 34:2325-30. [PMID: 26377204 PMCID: PMC4655006 DOI: 10.1007/s10096-015-2484-9] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2015] [Accepted: 08/27/2015] [Indexed: 12/18/2022]
Abstract
Current international guidelines lack definite conclusions regarding repeat stool sampling for the detection of toxigenic Clostridium difficile. We assessed the value of repeat sampling and compared the diagnostic yield in an epidemic to a non-epidemic setting. Consecutive fecal samples obtained during two time frames were analyzed using direct stool immunoassay toxin testing (enzyme immunoassay [EIA]), direct stool real-time PCR toxin gene testing, and toxigenic culture. Samples collected within 7 days of the initial sample were considered repeat tests. In the epidemic setting 989 patients were analyzed, and in the non-epidemic setting 1,015. In the epidemic setting 204 patients had two or more specimens included for analysis and in the non-epidemic setting 287 patients. In the epidemic setting 136 samples yielded a positive results, either by EIA or toxigenic culture; of these, 108 were positive according to EIA and 123 according to toxigenic culture. In the first test round 98 (90.7%, 95% CI 85.3 to 96.2), 114 (92.7%, 88.1 to 97.3), and 126 (92.6%, 88.3 to 97.0) positives were detected. Subsequent test rounds yielded 10 (9.3%, 3.8 to 14.7), 9 (7.3%, 2.7 to 11.9), and 10 (7.4%, 3.0 to 11.7) extra positives. In the non-epidemic setting EIA, toxigenic culture and PCR detected 33, 66, and 83 positives. The three tests combined 93 detected positives. In the first test round 30 (90.9%, 81.1 to 100.7), 63 (95.5%, 90.4 to 110.5), 76 (91.6%, 85.6 to 97.5), and 87 (93.5%, 88.6 to 98.5) positives were detected. Subsequent test rounds yielded 3 (9.1%, -0.7 to 18.9), 3 (4.5%, -0.5 to 9.6), 7 (8.4%, 2.5 to 14.4), and 6 (6.5%, 1.5 to 11.4) extra positives. In conclusion, repeat testing resulted in 4.5% to 9.3% extra positives. No significant difference between the settings studied could be demonstrated. Repeat sampling and multimodality testing may be chosen in an outbreak situation to detect all cases, effectively controlling nosocomial spread.
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13
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Routine disc diffusion antimicrobial susceptibility testing of Clostridium difficile and association with PCR ribotype 027. Eur J Clin Microbiol Infect Dis 2015; 34:2243-6. [PMID: 26319148 DOI: 10.1007/s10096-015-2475-x] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2015] [Accepted: 08/20/2015] [Indexed: 01/05/2023]
Abstract
Reduced susceptibility to metronidazole and vancomycin in Clostridium difficile has been reported, which emphasises the need for simple antimicrobial susceptibility testing methods. The aim of this study was to apply a published disc diffusion method and zone diameter breakpoint correlates to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) epidemiological minimum inhibitory concentration (MIC) cut-off values in a routine setting. Metronidazole and vancomycin zone diameters from 2702 isolates were recorded. Fifteen isolates had a metronidazole zone diameter below the published breakpoint (<23 mm) and five isolates had a vancomycin zone diameter below the published breakpoint (<19 mm), most of which were polymerase chain reaction (PCR) ribotype 027. The total number of PCR ribotype 027 was 29 (1.1 %). Overall, C. difficile PCR ribotype 027 isolates had smaller zone diameters than non-027 isolates. The disc diffusion method is very simple and inexpensive, and the published zone diameter breakpoints will detect C. difficile isolates with reduced susceptibility to metronidazole and vancomycin.
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14
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Diagnosis of Clostridium difficile: real-time PCR detection of toxin genes in faecal samples is more sensitive compared to toxigenic culture. Eur J Clin Microbiol Infect Dis 2014; 34:727-36. [DOI: 10.1007/s10096-014-2284-7] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2014] [Accepted: 11/10/2014] [Indexed: 02/08/2023]
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15
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Enserink R, Scholts R, Bruijning-Verhagen P, Duizer E, Vennema H, de Boer R, Kortbeek T, Roelfsema J, Smit H, Kooistra-Smid M, van Pelt W. High detection rates of enteropathogens in asymptomatic children attending day care. PLoS One 2014; 9:e89496. [PMID: 24586825 PMCID: PMC3933542 DOI: 10.1371/journal.pone.0089496] [Citation(s) in RCA: 64] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2013] [Accepted: 01/21/2014] [Indexed: 12/18/2022] Open
Abstract
Background Gastroenteritis morbidity is high among children under the age of four, especially amongst those who attend day care. Objective To determine the prevalence of a range of enteropathogens in the intestinal flora of children attending day care and to relate their occurrence with characteristics of the sampled child and the sampling season. Methods We performed three years of enteropathogen surveillance in a network of 29 child day care centers in the Netherlands. The centers were instructed to take one fecal sample from ten randomly chosen children each month, regardless of gastrointestinal symptoms at time of sampling. All samples were analyzed for the molecular detection of 16 enteropathogenic bacteria, parasites and viruses by real-time multiplex PCR. Results Enteropathogens were detected in 78.0% of the 5197 fecal samples. Of the total, 95.4% of samples were obtained from children who had no gastroenteritis symptoms at time of sampling. Bacterial enteropathogens were detected most often (most prevalent EPEC, 19.9%), followed by parasitic enteropathogens (most prevalent: D. fragilis, 22.1%) and viral enteropathogens (most prevalent: norovirus, 9.5%). 4.6% of samples related to children that experienced symptoms of gastroenteritis at time of sampling. Only rotavirus and norovirus were significantly associated with gastroenteritis among day care attendees. Conclusions Our study indicates that asymptomatic infections with enteropathogens in day care attendees are not a rare event and that gastroenteritis caused by infections with these enteropathogens is only one expression of their presence.
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Affiliation(s)
- Remko Enserink
- Center for Infectious Disease Control (Epidemiology and Surveillance Unit), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
- Julius Center for Health Sciences and Primary Care, University Medical Center Utrecht, Utrecht, The Netherlands
- * E-mail:
| | - Rianne Scholts
- Laboratory for Infectious Diseases, Department of Research and Development, Groningen, The Netherlands
| | - Patricia Bruijning-Verhagen
- Center for Infectious Disease Control (Epidemiology and Surveillance Unit), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
- Julius Center for Health Sciences and Primary Care, University Medical Center Utrecht, Utrecht, The Netherlands
| | - Erwin Duizer
- Center for Infectious Disease Control (Laboratory for Infectious Diseases and Perinatal Screening), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
| | - Harry Vennema
- Center for Infectious Disease Control (Laboratory for Infectious Diseases and Perinatal Screening), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
| | - Richard de Boer
- Laboratory for Infectious Diseases, Department of Research and Development, Groningen, The Netherlands
| | - Titia Kortbeek
- Center for Infectious Disease Control (Laboratory for Infectious Diseases and Perinatal Screening), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
| | - Jeroen Roelfsema
- Center for Infectious Disease Control (Laboratory for Infectious Diseases and Perinatal Screening), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
| | - Henriette Smit
- Julius Center for Health Sciences and Primary Care, University Medical Center Utrecht, Utrecht, The Netherlands
| | - Mirjam Kooistra-Smid
- Laboratory for Infectious Diseases, Department of Research and Development, Groningen, The Netherlands
- Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands
| | - Wilfrid van Pelt
- Center for Infectious Disease Control (Epidemiology and Surveillance Unit), National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
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Premarket evaluations of the IMDx C. difficile for Abbott m2000 Assay and the BD Max Cdiff Assay. J Clin Microbiol 2014; 52:1423-8. [PMID: 24554744 DOI: 10.1128/jcm.03293-13] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Clostridium difficile-associated diarrhea is a well-recognized complication of antibiotic use. Historically, diagnosing C. difficile has been difficult, as antigen assays are insensitive and culture-based methods require several days to yield results. Nucleic acid amplification tests (NAATs) are quickly becoming the standard of care. We compared the performance of two automated investigational/research use only (IUO/RUO) NAATs for the detection of C. difficile toxin genes, the IMDx C. difficile for Abbott m2000 Assay (IMDx) and the BD Max Cdiff Assay (Max). A prospective analysis of 111 stool specimens received in the laboratory for C. difficile testing by the laboratory's test of record (TOR), the BD GeneOhm Cdiff Assay, and a retrospective analysis of 88 specimens previously determined to be positive for C. difficile were included in the study. One prospective specimen was excluded due to loss to follow-up discrepancy analysis. Of the remaining 198 specimens, 90 were positive by all three methods, 9 were positive by TOR and Max, and 3 were positive by TOR only. One negative specimen was initially inhibitory by Max. The remaining 95 specimens were negative by all methods. Toxigenic C. difficile culture was performed on the 12 discrepant samples. True C. difficile-positive status was defined as either positive by all three amplification assays or positive by toxigenic culture. Based on this definition, the sensitivity and specificity were 96.9% and 95% for Max and 92.8% and 100% for IMDx. In summary, both highly automated systems demonstrated excellent performance, and each has individual benefits, which will ensure that they will both have a niche in clinical laboratories.
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Pallis A, Jazayeri J, Ward P, Dimovski K, Svobodova S. Rapid detection of Clostridium difficile toxins from stool samples using real-time multiplex PCR. J Med Microbiol 2013; 62:1350-1356. [DOI: 10.1099/jmm.0.058339-0] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023] Open
Abstract
In this study, a total of 650 stool samples were tested to show that our method is capable of detecting four Clostridium difficile genes; tcdA, tcdB, encoding toxin A (TcdA) and toxin B (TcdB), and the binary toxin C. difficile transferase genes (cdtA and/or cdtB) encoding CDT toxin. Besides detecting the targeted C. difficile genes, our method can be used to detect the presence of any inhibitory components in the PCR. This assay, combined with a selective culture medium, such as the chromID™ C. difficile, can be applied directly for screening C. difficile-associated disease. The PCR-based assay developed here is rapid (4 h per 21 stool samples) and accurate in diagnosing C. difficile infection, 100 % assay sensitivity and negative predictive value (NPV) were obtained. However, the assay specificity of 99.1 % and positive predictive value (PPV) of 94.9 % were slightly lower than the optimal value of 100 %. The assay protocol outlined here can be used as a rapid screening tool to assist infection control units and in managing infected patients by reducing the number of patients requiring isolation and extended hospitalization. Rapid detection can prevent unnecessary antibiotic therapy and potentially reduce the spread of infection by emerging hypervirulent C. difficile strains.
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Affiliation(s)
- Ann Pallis
- Molecular Diagnostic and Microbiology Laboratory, Austin Pathology, Melbourne, VIC 3084, Australia
| | - Jalal Jazayeri
- School of Biomedical Sciences, Charles Sturt University, Boorooma Street, Wagga Wagga, NSW 2678, Australia
| | - Peter Ward
- Molecular Diagnostic and Microbiology Laboratory, Austin Pathology, Melbourne, VIC 3084, Australia
| | | | - Suzanne Svobodova
- Molecular Diagnostic and Microbiology Laboratory, Austin Pathology, Melbourne, VIC 3084, Australia
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18
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Abstract
In recent years, quantitative real-time PCR tests have been extensively developed in clinical microbiology laboratories for routine diagnosis of infectious diseases, particularly bacterial diseases. This molecular tool is well-suited for the rapid detection of bacteria directly in clinical specimens, allowing early, sensitive and specific laboratory confirmation of related diseases. It is particularly suitable for the diagnosis of infections caused by fastidious growth species, and the number of these pathogens has increased recently. This method also allows a rapid assessment of the presence of antibiotic resistance genes or gene mutations. Although this genetic approach is not always predictive of phenotypic resistances, in specific situations it may help to optimize the therapeutic management of patients. Finally, an approach combining the detection of pathogens, their mechanisms of antibiotic resistance, their virulence factors and bacterial load in clinical samples could lead to profound changes in the care of these infected patients.
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Affiliation(s)
- Max Maurin
- Laboratoire de Bactériologie, Département des Agents Infectieux, Institut de Biologie et Pathologie, CHU de Grenoble, Université Joseph Fourier Grenoble 1, France.
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19
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O'Horo JC, Jones A, Sternke M, Harper C, Safdar N. Molecular techniques for diagnosis of Clostridium difficile infection: systematic review and meta-analysis. Mayo Clin Proc 2012; 87:643-51. [PMID: 22766084 PMCID: PMC3538482 DOI: 10.1016/j.mayocp.2012.02.024] [Citation(s) in RCA: 59] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/23/2011] [Revised: 02/07/2012] [Accepted: 02/29/2012] [Indexed: 12/16/2022]
Abstract
OBJECTIVE To assess the usefulness of 2 rapid molecular diagnostic techniques, polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP), in Clostridium difficile infection (CDI). METHODS We conducted a systematic review and meta-analysis to evaluate the accuracy of PCR and LAMP in diagnosis of CDI, including studies that used toxigenic culture or cytotoxicity assay as reference standard. RESULTS A search of PubMed and CinAHL medical databases yielded 25 PCR studies, including 11,801 samples that met inclusion criteria and 6 heterogeneous studies that evaluated LAMP. With toxigenic culture as a standard, pooled sensitivity was 0.92 (95% confidence interval [CI], 0.91-0.94); specificity, 0.94 (95% CI, 0.94-0.95); and diagnostic odds ratio, 378 (95% CI, 260-547). With cytotoxicity as a standard, pooled sensitivity was 0.87 (95% CI, 0.84-0.90); specificity, 0.97 (95% CI, 0.97-0.98); and diagnostic odds ratio, 370 (95% CI, 226-606). CONCLUSION Polymerase chain reaction is a highly accurate test for identifying CDI. Heterogeneity in LAMP studies did not allow meta-analysis; however, further research into this promising method is warranted.
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Key Words
- cdi, clostridium difficile infection
- ci, confidence interval
- dor, diagnostic odds ratio
- fn, false-negative
- fp, false-positive
- lamp, loop-mediated isothermal amplification
- lr+, positive likelihood ratio
- lr−, negative likelihood ratio
- npv, negative predictive value
- pcr, polymerase chain reaction
- ppv, positive predictive value
- prisma, preferred reporting items for systematic meta-analysis
- sroc, summary receiver operating curve
- tn, true-negative
- tp, true-positive
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Affiliation(s)
- John C. O'Horo
- Department of Graduate Medical Education, Aurora UW Medical Group, Milwaukee, WI
| | - Amy Jones
- University of Wisconsin School of Medicine and Public Health, Madison, WI
| | - Matthew Sternke
- University of Wisconsin School of Medicine and Public Health, Madison, WI
| | - Christopher Harper
- University of Wisconsin School of Medicine and Public Health, Madison, WI
| | - Nasia Safdar
- William S. Middleton Memorial Veterans Hospital, Madison, WI
- University of Wisconsin School of Medicine and Public Health, Madison, WI
- Section of Infectious Diseases, Department of Medicine, Madison, WI
- Correspondence: Address to Nasia Safdar, MD, PhD, Section of Infectious Diseases, Department of Medicine, MFCB 5221, University of Wisconsin Hospital and Clinics, Madison, WI 53705
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Precise manipulation of the Clostridium difficile chromosome reveals a lack of association between the tcdC genotype and toxin production. Appl Environ Microbiol 2012; 78:4683-90. [PMID: 22522680 DOI: 10.1128/aem.00249-12] [Citation(s) in RCA: 168] [Impact Index Per Article: 12.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
Clostridium difficile causes a potentially fatal diarrheal disease through the production of its principal virulence factors, toxin A and toxin B. The tcdC gene is thought to encode a negative regulator of toxin production. Therefore, increased toxin production, and hence increased virulence, is often inferred in strains with an aberrant tcdC genotype. This report describes the first allele exchange system for precise genetic manipulation of C. difficile, using the codA gene of Escherichia coli as a heterologous counterselection marker. It was used to systematically restore the Δ117 frameshift mutation and the 18-nucleotide deletion that occur naturally in the tcdC gene of C. difficile R20291 (PCR ribotype 027). In addition, the naturally intact tcdC gene of C. difficile 630 (PCR ribotype 012) was deleted and then subsequently restored with a silent nucleotide substitution, or "watermark," so the resulting strain was distinguishable from the wild type. Intriguingly, there was no association between the tcdC genotype and toxin production in either C. difficile R20291 or C. difficile 630. Therefore, an aberrant tcdC genotype does not provide a broadly applicable rationale for the perceived notion that PCR ribotype 027 strains are "high-level" toxin producers. This may well explain why several studies have reported that an aberrant tcdC gene does not predict increased toxin production or, indeed, increased virulence.
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21
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Walter Zea J, Lina Salazar C. Enfermedad asociada a Clostridium difficile: prevalencia y diagnóstico por laboratorio. INFECTIO 2012. [DOI: 10.1016/s0123-9392(12)70016-4] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
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Multiplex PCR method for detection of Clostridium difficile tcdA, tcdB, cdtA, and cdtB and internal in-frame deletion of tcdC. J Clin Microbiol 2011; 49:4299-300. [PMID: 21976756 DOI: 10.1128/jcm.05161-11] [Citation(s) in RCA: 62] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
A multiplex PCR method was developed for the detection of Clostridium difficile toxin genes tcdA, tcdB, ctdA, and cdtB and the major in-frame deletion types (18, 39, and 54 bp) of tcdC. The method has high specificity for PCR ribotype 027 and may identify other C. difficile strains of clinical and epidemiological importance.
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Deshpande A, Pasupuleti V, Rolston DD, Jain A, Deshpande N, Pant C, Hernandez AV. Diagnostic Accuracy of Real-time Polymerase Chain Reaction in Detection of Clostridium difficile in the Stool Samples of Patients With Suspected Clostridium difficile Infection: A Meta-Analysis. Clin Infect Dis 2011; 53:e81-90. [DOI: 10.1093/cid/cir505] [Citation(s) in RCA: 107] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
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Kim H, Jeong SH, Kim M, Lee Y, Lee K. Detection of Clostridium difficile toxin A/B genes by multiplex real-time PCR for the diagnosis of C. difficile infection. J Med Microbiol 2011; 61:274-277. [PMID: 21959205 DOI: 10.1099/jmm.0.035618-0] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023] Open
Abstract
Toxigenic Clostridium difficile culture is considered to be the standard diagnostic method for the detection of C. difficile infection (CDI). Culture methods are time-consuming and although enzyme immunoassay is rapid and easy to use, it has low sensitivity. In the present study, the AdvanSure CD real-time (RT)-PCR kit (LG Life Sciences) was evaluated for its ability to detect C. difficile toxin A (tcdA) and B (tcdB) genes, simultaneously. A total of 127 fresh diarrhoeal stool specimens, submitted to the clinical microbiology laboratory for C. difficile culture, were tested. C. difficile toxins and toxin genes were detected with a VIDAS C. difficile A&B (VIDAS-CDAB) enzyme-linked fluorescent immunoassay (ELFA) and the AdvanSure RT-PCR kit, respectively, according to the manufacturers' instructions. Their performance was compared with a standard toxigenic culture method as a reference. The sensitivity, specificity and positive and negative predictive values using the AdvanSure RT-PCR kit were 100 %, 98.3 %, 84.6 % and 100 %, respectively, while those of the VIDAS-CDAB system were 63.6 %, 100 %, 100 % and 96.6 %, respectively. Four tcdA(+)/tcdB(+) strains of C. difficile were detected with the AdvanSure RT-PCR kit, which offers comparable sensitivity and specificity to the reference method with a turnaround time of ~3 hours.
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Affiliation(s)
- Heejung Kim
- Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752, Republic of Korea
| | - Seok Hoon Jeong
- Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752, Republic of Korea
| | - Myungsook Kim
- Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752, Republic of Korea
| | - Yangsoon Lee
- Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752, Republic of Korea
| | - Kyungwon Lee
- Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752, Republic of Korea
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25
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Hoegh AM, Nielsen JB, Lester A, Friis-Møller A, Schønning K. A multiplex, internally controlled real-time PCR assay for detection of toxigenic Clostridium difficile and identification of hypervirulent strain 027/ST-1. Eur J Clin Microbiol Infect Dis 2011; 31:1073-9. [DOI: 10.1007/s10096-011-1409-5] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2011] [Accepted: 08/27/2011] [Indexed: 02/04/2023]
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26
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Friesema IHM, Boer RF, Duizer E, Kortbeek LM, Notermans DW, Norbruis OF, Bezemer DDL, Heerbeek H, Andel RNJ, Enk JG, Fraaij PLA, Koopmans MPG, Kooistra-Smid AMD, Duynhoven YTHP. Etiology of acute gastroenteritis in children requiring hospitalization in the Netherlands. Eur J Clin Microbiol Infect Dis 2011; 31:405-15. [DOI: 10.1007/s10096-011-1320-0] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2011] [Accepted: 06/08/2011] [Indexed: 12/01/2022]
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27
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Persson S, de Boer RF, Kooistra-Smid AM, Olsen KE. Five commercial DNA extraction systems tested and compared on a stool sample collection. Diagn Microbiol Infect Dis 2011; 69:240-4. [DOI: 10.1016/j.diagmicrobio.2010.09.023] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2010] [Revised: 09/15/2010] [Accepted: 09/29/2010] [Indexed: 10/18/2022]
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Abstract
PURPOSE OF REVIEW This review summarizes the most recent epidemiological data and advances in research into the pathogenesis, diagnosis and treatment of Clostridium difficile infection (CDI). RECENT FINDINGS The epidemiology of CDI has changed with the emergence of hypervirulent strains. CDI rates have increased in the community, in children and in patients with inflammatory bowel disease. Although the North American pulsed-field gel electrophoresis type 1, restriction endonuclease analysis group BI, PCR ribotype 027 (NAP1/BI/027) strain remains prevalent in North America, surveillance suggests that it is decreasing in Europe. A similar strain, PCR ribotype 078, is emerging which is associated with community-associated CDI and has been isolated in animals and food products. The Society for Healthcare Epidemiology of America and the Infectious Diseases Society of America have published new guidelines on the epidemiology, diagnosis, treatment, infection control and environmental management of C. difficile. Several novel therapies for CDI are at different stages of development. There have been promising trial results with fidaxomicin, a novel antibiotic for the treatment of CDI and monoclonal antibodies against toxins A and B, which have been shown to significantly reduce CDI recurrence rates. SUMMARY Major advances have been made in our understanding of the spread and pathogenesis of C. difficile and new treatment options are becoming available.
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Comparison of real-time PCR techniques to cytotoxigenic culture methods for diagnosing Clostridium difficile infection. J Clin Microbiol 2010; 49:227-31. [PMID: 20980562 DOI: 10.1128/jcm.01743-10] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
In the past decade, the incidence of Clostridium difficile infections (CDI) with a more severe course has increased in Europe and North America. Assays that are capable of rapidly diagnosing CDI are essential. Two real-time PCRs (LUMC and LvI) targeting C. difficile toxin genes (tcdB, and tcdA and tcdB, respectively) were compared with the BD GeneOhm PCR (targeting the tcdB gene), using cytotoxigenic culture as a gold standard. In addition, a real-time PCR targeting the tcdC frameshift mutation at position 117 (Δ117 PCR) was evaluated for detecting toxigenic C. difficile and the presence of PCR ribotype 027 in stool samples. In total, 526 diarrheal samples were prospectively collected and included in the study. Compared with those for cytotoxigenic culture, sensitivity, specificity, positive predicted value (PPV), and negative predicted value (NPV) were for PCR LUMC 96.0%, 88.0%, 66.0%, and 98.9%, for PCR LvI 100.0%, 89.4%, 69.7%, and 100.0%, for PCR Δ117 98.0%, 90.7%, 71.9%, and 99.5%, and for PCR BD GeneOhm 88.3%, 96.9%, 86.5%, and 97.4%. Compared to those with feces samples cultured positive for C. difficile type 027, the sensitivity, specificity, PPV, and NPV of the Δ117 PCR were 95.2%, 96.2%, 87.0%, and 98.7%. We conclude that all real-time PCRs can be applied as a first screening test in an algorithm for diagnosing CDI. However, the low PPVs hinder the use of the assays as stand-alone tests. Furthermore, the Δ117 PCR may provide valuable information for minimizing the spread of the epidemic C. difficile PCR ribotype 027.
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