scholarly journals Host Immune Markers Distinguish Clostridioides difficile Infection From Asymptomatic Carriage and Non–C. difficile Diarrhea

2019 ◽  
Author(s):  
Ciaran P Kelly ◽  
Xinhua Chen ◽  
David Williams ◽  
Hua Xu ◽  
Christine A Cuddemi ◽  
...  
Keyword(s):  
Adaptive Immunity ◽  
Symptomatic Patient ◽  
Nucleic Acid Amplification ◽  
Toxin A ◽  
Toxin B ◽  
Innate And Adaptive Immunity ◽  
Clostridioides Difficile ◽  
Asymptomatic Carriage ◽  
Positive Stool ◽  
Tnf Α

Abstract Background Recent data indicate that Clostridioides difficile toxin concentrations in stool do not differentiate between C. difficile infection (CDI) and asymptomatic carriage. Thus, we lack a method to distinguish a symptomatic patient with CDI from a colonized patient with diarrhea from another cause. To address this, we evaluated markers of innate and adaptive immunity in adult inpatients with CDI (diagnosed per US guidelines), asymptomatic carriage, or non-CDI diarrhea. Methods CDI-NAAT patients had clinically significant diarrhea and positive nucleic acid amplification testing (NAAT) and received CDI treatment. Carrier-NAAT patients had positive stool NAAT but no diarrhea. NAAT-negative patients (with and without diarrhea) were also enrolled. A panel of cytokines and anti–toxin A and B immunoglobulin (Ig) were measured in serum; calprotectin and anti–toxin B Ig A/G were measured in stool. NAAT-positive stool samples were tested by an ultrasensitive toxin assay (clinical cutoff, 20 pg/mL). Results Median values for interleukin (IL)-4, IL-6, IL-8, IL-10, IL-15, granulocyte colony-stimulating factor (GCSF), MCP-1, tumor necrosis factor α (TNF-α), and IgG anti–toxin A in blood and IgA/G anti–toxin B in stool were significantly higher in CDI patients compared with all other groups (P < .05). Concentration distributions for IL-6, GCSF, TNF-α, and IgG anti–toxin A in blood, as well as IgA and IgG anti–toxin B in stool, separated CDI patients from all other groups. Conclusions Specific markers of innate and adaptive immunity distinguish CDI from all other groups, suggesting potential clinical utility for identifying which NAAT- and toxin-positive patients with diarrhea truly have CDI.

scholarly journals Nucleic Acid Amplification Test Quantitation as Predictor of Toxin Presence in Clostridium difficile Infection

2017 ◽  
Vol 56 (3) ◽  
Author(s):  
M. J. T. Crobach ◽  
N. Duszenko ◽  
E. M. Terveer ◽  
C. M. Verduin ◽  
E. J. Kuijper
Keyword(s):  
Nucleic Acid ◽  
Clostridium Difficile ◽  
Characteristic Curve ◽  
Nucleic Acid Amplification Test ◽  
Nucleic Acid Amplification ◽  
Toxin A ◽  
Toxin B ◽  
Content Type ◽  
Quantitative Results ◽  
Testing Algorithm

ABSTRACT Multistep algorithmic testing in which a sensitive nucleic acid amplification test (NAAT) is followed by a specific toxin A and toxin B enzyme immunoassay (EIA) is among the most accurate methods for Clostridium difficile infection (CDI) diagnosis. The obvious shortcoming of this approach is that multiple tests must be performed to establish a CDI diagnosis, which may delay treatment. Therefore, we sought to determine whether a preliminary diagnosis could be made on the basis of the quantitative results of the first test in algorithmic testing, which provide a measure of organism burden. To do so, we retrospectively analyzed two large collections of samples ( n = 2,669 and n = 1,718) that were submitted to the laboratories of two Dutch hospitals for CDI testing. Both hospitals apply a two-step testing algorithm in which a NAAT is followed by a toxin A/B EIA. Of all samples, 208 and 113 samples, respectively, tested positive by NAAT. Among these NAAT-positive samples, significantly lower mean quantification cycle ( C q ) values were found for patients whose stool eventually tested positive for toxin, compared with patients who tested negative for toxin (mean C q values of 24.4 versus 30.4 and 26.8 versus 32.2; P < 0.001 for both cohorts). Receiver operating characteristic curve analysis was performed to investigate the ability of C q values to predict toxin status and yielded areas under the curve of 0.826 and 0.854. Using the optimal C q cutoff values, prediction of the eventual toxin A/B EIA results was accurate for 78.9% and 80.5% of samples, respectively. In conclusion, C q values can serve as predictors of toxin status but, due to the suboptimal correlation between the two tests, additional toxin testing is still needed.

scholarly journals Steroid Sulfates from Ophiuroids (Brittle Stars): Action on Some Factors of Innate and Adaptive Immunity

2016 ◽  
Vol 11 (6) ◽  
pp. 1934578X1601100
Author(s):  
Anna K Gazha ◽  
Lyudmila A. Ivanushko ◽  
Eleonora V. Levina ◽  
Sergey N. Fedorov ◽  
Tatyana S. Zaporozets ◽  
...  
Keyword(s):  
Adaptive Immunity ◽  
Inflammatory Cytokines ◽  
Brittle Stars ◽  
Innate And Adaptive Immunity ◽  
Functional Activities ◽  
In Vivo Experiments ◽  
Tnf Α ◽  
Pro Inflammatory Cytokines

The action of seven polyhydroxylated sterol mono- and disulfates (1-7), isolated from ophiuroids, on innate and adaptive immunity was examined in in vitro and in vivo experiments. At least, three of them (1, 2 and 4) increased the functional activities of neutrophils, including levels of oxygen-dependent metabolism, adhesive and phagocytic properties, and induced the expression of pro-inflammatory cytokines TNF-α and IL-8. Compound 4 was the most active for enhancing the production of antibody forming cells in the mouse spleen.

scholarly journals Fidaxomicin and OP-1118 Inhibit Clostridium difficile Toxin A- and B-Mediated Inflammatory Responses via Inhibition of NF-κB Activity

2017 ◽  
Vol 62 (1) ◽  
Author(s):  
Hon Wai Koon ◽  
Jiani Wang ◽  
Caroline C. Mussatto ◽  
Christina Ortiz ◽  
Elaine C. Lee ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Human Colon ◽  
Toxin A ◽  
Primary Metabolite ◽  
Necrosis Factor Alpha ◽  
Toxin B ◽  
Content Type ◽  
Tnf Α

ABSTRACTClostridium difficilecauses diarrhea and colitis by releasing toxin A and toxin B. In the human colon, both toxins cause intestinal inflammation and stimulate tumor necrosis factor alpha (TNF-α) expression via the activation of NF-κB. It is well established that the macrolide antibiotic fidaxomicin is associated with reduced relapses ofC. difficileinfection. We showed that fidaxomicin and its primary metabolite OP-1118 significantly inhibited toxin A-mediated intestinal inflammation in micein vivoand toxin A-induced cell roundingin vitro. We aim to determine whether fidaxomicin and OP-1118 possess anti-inflammatory effects against toxin A and toxin B in the human colon and examine the mechanism of this response. We used fresh human colonic explants, NCM460 human colonic epithelial cells, and RAW264.7 mouse macrophages to study the mechanism of the activity of fidaxomicin and OP-1118 against toxin A- and B-mediated cytokine expression and apoptosis. Fidaxomicin and OP-1118 dose-dependently inhibited toxin A- and B-induced TNF-α and interleukin-1β (IL-1β) mRNA expression and histological damage in human colonic explants. Fidaxomicin and OP-1118 inhibited toxin A-mediated NF-κB phosphorylation in human and mouse intestinal mucosae. Fidaxomicin and OP-1118 also inhibited toxin A-mediated NF-κB phosphorylation and TNF-α expression in macrophages, which was reversed by the NF-κB activator phorbol myristate acetate (PMA). Fidaxomicin and OP-1118 prevented toxin A- and B-mediated apoptosis in NCM460 cells, which was reversed by the addition of PMA. PMA reversed the cytoprotective effect of fidaxomicin and OP-1118 in toxin-exposed human colonic explants. Fidaxomicin and OP-1118 inhibitC. difficiletoxin A- and B-mediated inflammatory responses, NF-κB phosphorylation, and tissue damage in the human colon.

scholarly journals 2359. Prospective Feasibility Study for Novel Ultrasensitive Multiplexed Immunoassay for Clostridiodes difficile Toxins A and B

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S812-S813
Author(s):  
Lauren Watson ◽  
Michele L Zimbric ◽  
Catherine Shaughnessy ◽  
Shraddha Kale ◽  
Jeff Debad ◽  
...  
Keyword(s):  
Diagnostic Tests ◽  
Standard Of Care ◽  
Positive Sample ◽  
Nucleic Acid Amplification ◽  
Serum Samples ◽  
Toxin A ◽  
Toxin B ◽  
Diagnostic Assays ◽  
Stool Samples ◽  
Multiplexed Immunoassay

Abstract Background The diagnosis of Clostridiodes difficile infection is challenging. A wide array of diagnostic tests are used in practice; however, each available test has important limitations. We examined the feasibility and analytical performance of a novel ultrasensitive multiplexed immunoassay designed by Meso Scale Diagnostics (MSD) compared with five current diagnostic assays for detection of C. difficile toxin A and B. Methods Stool, serum and urine samples from 44 admitted inpatients were collected within 72 hours of a standard of care nucleic acid amplification test (NAAT) result (23 positive, 21 negative). These specimens underwent five standard diagnostic assays: enzyme immunoassay for toxins A and B (EIA), cytotoxin cell assay, bacterial culture isolation, and two different NAATs to determine presence of viable C. difficile cells, toxins, and toxin-encoding genes (Table 1). The concentration (fg/mL) of toxin A and toxin B in all stool samples was then quantified using MSD’s multiplexed immunoassay (Table 1). Results At least one of the five standard diagnostic tests for C. difficile was positive in 16 of the 23 clinically positive patients. The MSD multiplex immunoassay detected toxin A and/or toxin B in 15 of these 16 samples and quantified low levels of toxin A in one clinically positive sample that was negative for all other tests. In contrast, only 2 of the 16 positive samples were positive by EIA, demonstrating the benefits of the ultrasensitive assay over standard immunoassay methods. All clinically negative specimens were negative in all tests. Toxin detection in urine and serum samples was negligible. In stool samples, the MSD test had an estimated sensitivity of 93% (95% CI: 70–99%) and specificity of 93% (95% CI: 78–98%) compared with the clinically used NAAT. Conclusion The MSD multiplex toxin assay is a feasible test to move forward for further evaluation. Ultimately, future studies should examine the performance of this test compared with standard of care in a prospective randomized trial assessing clinical outcomes. Disclosures All authors: No reported disclosures.

scholarly journals Human Monoclonal Antibodies against Clostridium difficile Toxins A and B Inhibit Inflammatory and Histologic Responses to the Toxins in Human Colon and Peripheral Blood Monocytes

2013 ◽  
Vol 57 (7) ◽  
pp. 3214-3223 ◽  
Author(s):  
Hon Wai Koon ◽  
David Q. Shih ◽  
Tressia C. Hing ◽  
Jun Hwan Yoo ◽  
Samantha Ho ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Clostridium Difficile ◽  
Peripheral Blood ◽  
Blood Monocytes ◽  
Human Monoclonal Antibodies ◽  
Toxin A ◽  
Toxin B ◽  
Peripheral Blood Monocytes ◽  
Content Type ◽  
Tnf Α

ABSTRACTClostridium difficileinfection (CDI) is a common and debilitating nosocomial infection with high morbidity and mortality.C. difficilemediates diarrhea and colitis by releasing two toxins, toxin A and toxin B. Since both toxins stimulate proinflammatory signaling pathways in human colonocytes and both are involved in the pathophysiology of CDI, neutralization of toxin A and B activities may represent an important therapeutic approach against CDI. Recent studies indicated that human monoclonal antibodies (MAbs) against toxins A and B reduce their cytotoxic and secretory activities and prevent CDI in hamsters. Moreover, anti-toxin A and anti-toxin B MAbs together with antibiotics also effectively reduced recurrent CDI in humans. However, whether these MAbs neutralize toxin A- and toxin B-associated immune responses in human colonic mucosa or human peripheral blood monocyte cells (PBMCs) has never been examined. We used fresh human colonic biopsy specimens and peripheral blood monocytes to evaluate the effects of these antibodies against toxin A- and B-associated cytokine release, proinflammatory signaling, and histologic damage. Incubation of anti-toxin A (MK3415) or anti-toxin B (MK6072) MAbs with human PBMCs significantly inhibited toxin A- and toxin B-mediated tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β) expression. MK3415 and MK6072 also diminished toxin A- and toxin B-mediated NF-κB p65 phosphorylation in human monocytes, respectively, and significantly reduced toxin A- and B-induced TNF-α and IL-1β expression as well as histologic damage in human colonic explants. Our results underline the effectiveness of MK3415 and MK6072 in blockingC. difficiletoxin A- and toxin B-mediated inflammatory responses and histologic damage.

scholarly journals 730. Severity Of Clostridioides difficile Infection Based On Toxin Analysis, Acid Suppressant Medications and Antibiotics

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S415-S416
Author(s):  
Gayathri Krishnan ◽  
Richa Parikh ◽  
Anna N Witt ◽  
Kulsum Bano ◽  
Sudeepa Bhattacharyya ◽  
...  
Keyword(s):  
Health Care ◽  
Medical Center ◽  
The United States ◽  
Nucleic Acid Amplification ◽  
Clinical Severity ◽  
Toxin Gene ◽  
Antibiotic Exposure ◽  
Toxin B ◽  
Major Health Problem ◽  
Clostridioides Difficile

Abstract Background Clostridioides difficile (C difficile) infection (CDI) is a major health problem in the United States and despite updated guidelines, the laboratory diagnosis remains vexed. A multistep algorithm is recommended to diagnose CDI that includes antigen, toxin and toxin gene Nucleic Acid Amplification (NAAT) assays. This study was done to assess severity of CDI based on toxin B and NAAT statuses. The other objective was to analyze if antibiotics and PPI/H2B (Proton Pump Inhibitors and H2 blockers) affected severity of CDI. Methods Retrospective analysis of all adult patients admitted to a tertiary medical center with diarrhea and a positive C difficile antigen test from 01/2017- 12/2017. From more than 2000 stool samples submitted to the lab, C diff antigen was positive in 265 patients. 191 were diagnosed with CDI based on the 2-step algorithm. Clinical data was available for 168 patients. Severity of CDI was determined based on published guidelines. Fischer’s exact test was used for statistical analysis. Results The mean age at diagnosis was 55.96. Toxin B was detected in 34% (57/168) patients and Toxin NAAT positive in 66% (111/168) patients. 57% of CDI was health care onset compared to 43% with community onset. 42% (72/168) were classified as severe out of which 40.2% (29) were toxin B positive, and 59.8% (43) were NAAT positive. There were no significant differences in severity of CDI based on toxin B and NAAT status (50.9% vs 38.4%, p=0.14). 46% of cases from community vs 39.6% from hospitals were classified as severe CDI (p=0.415). 72% of cases had antibiotic use in the last 30 days. Use of antibiotics was significantly associated with severe CDI (82% vs 64%, p=0.015). 62.5% (105) patients had history of PPI/H2B use and severity was not significantly associated with its use (p=0.872). Conclusion Our study shows that the presence of toxin did not significantly impact the clinical severity of CDI. The use of antibiotics did not affect the presence of toxin although the total number of CDI cases with previous antibiotic exposure was high. Patients who had recent antibiotic exposure were more likely to have severe clinical presentation. More toxin positive cases were health care onset but the effect was not pronounced. Severity of CDI did not significantly depend on health care onset or on exposure to PPI/H2B. Disclosures Atul Kothari, MD, Ansun Biopharma (Consultant)

scholarly journals Toxin A–Predominant Pathogenic Clostridioides difficile: A Novel Clinical Phenotype

2019 ◽  
Vol 70 (12) ◽  
pp. 2628-2633 ◽  
Author(s):  
Qianyun Lin ◽  
Nira R Pollock ◽  
Alice Banz ◽  
Aude Lantz ◽  
Hua Xu ◽  
...  
Keyword(s):  
Vaccine Development ◽  
Toxin Production ◽  
Toxin A ◽  
Toxin B ◽  
Clostridioides Difficile ◽  
Quantitative Immunoassay ◽  
Clinical Detection ◽  
Stool Samples

Abstract Background Most Clostridioides difficile toxinogenic strains produce both toxins A and B (A+B+), but toxin A–negative, toxin B–positive (A−B+) variants also cause disease. We report the identification of a series of pathogenic clinical C. difficile isolates that produce high amounts of toxin A with low or nondetectable toxin B. Methods An ultrasensitive, quantitative immunoassay was used to measure toxins A and B in stool samples from 187 C. difficile infection (CDI) patients and 44 carriers. Isolates were cultured and assessed for in vitro toxin production and in vivo phenotypes (mouse CDI model). Results There were 7 CDI patients and 6 carriers who had stools with detectable toxin A (TcdA, range 23–17 422 pg/mL; 5.6% of samples overall) but toxin B (TcdB) below the clinical detection limit (&lt;20 pg/mL; median TcdA:B ratio 17.93). Concentrations of toxin A far exceeded B in in vitro cultures of all 12 recovered isolates (median TcdA:B ratio 26). Of 8 toxin A&gt;&gt;B isolates tested in mice, 4 caused diarrhea, and 3 of those 4 caused lethal disease. Ribotyping demonstrated strain diversity. TcdA-predominant samples were also identified at 2 other centers, with similar frequencies (7.5% and 6.8%). Conclusions We report the discovery of clinical pathogenic C. difficile strains that produce high levels of toxin A but minimal or no toxin B. This pattern of toxin production is not rare (&gt;5% of isolates) and is consistently observed in vitro and in vivo in humans and mice. Our study highlights the significance of toxin A in human CDI pathogenesis and has important implications for CDI diagnosis, treatment, and vaccine development.

scholarly journals Detoxification of toxin A and toxin B by copper ion-catalyzed oxidation in production of a toxoid-based vaccine against Clostridioides difficile

2020 ◽  
Vol 160 ◽  
pp. 433-446
Author(s):  
Aria Aminzadeh ◽  
Manish Kumar Tiwari ◽  
Srwa Satar Mamah Mustapha ◽  
Sandra Junquera Navarrete ◽  
Anna Bielecka Henriksen ◽  
...  
Keyword(s):  
Copper Ion ◽  
Toxin A ◽  
Toxin B ◽  
Clostridioides Difficile ◽  
Catalyzed Oxidation
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