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Real-Time PCR Reveals the Presence of Three Distinct Brachyspira Species in Human Intestinal Spirochaetosis
the effect of fidaxomicin compared with vancomycin in preventing early vs. late CDI reoccurrence. Methods: Adults with CDI symptoms and a positive toxin test received oral fidaxomicin or vancomycin for ten days as part of a phase 3 clinical trial. Recurrence was defined as the reappearance of diarrhea and a positive stool toxin test within 4 weeks following completion of therapy. Multivariate analysis was used to identify patient characteristics at either the beginning or completion of therapy that correlated with recurrence or time to recurrence. Results: Of 518 patients (253 treated with fidaxomicin and 265 with vancomycin), 106 had a recurrence of CDI (20.5%). The distribution of recurrences over time was consistent with two distinct periods: early (within the first 2 weeks after stopping initial therapy; 10.5 reoccurrences/1000 patient days) and late (2 to 4 weeks after stopping initial therapy; 5.0 reoccurrences/1000 patient days). The table summarizes patient characteristics associated with increased incidence of early or late recurrence. Treatment with fidaxomicin was associated with a lower rate of early recurrence (5.9 recurrences/1000 patient days) compared with vancomycin (15.1 recurrences/1000 patient days, p<0.001). There was no difference between treatments in the rate of recurrence during weeks 3 and 4 of follow-up. Conclusions: CDI treatment with fidaxomicin resulted in a lower rate of early recurrence (within 14 days of completing therapy) than vancomycin. This differential effect is likely due to a less deleterious effect of fidaxomicin on normal fecal microflora. Late recurrence is likely related to a new infection and is affected by patient-related factors, not initial treatment choices. Such factors may define a group at high risk of a new infection and could guide prevention measures and treatment choices.
Sa1983
Introduction: Infections with Brachyspira species result in a wide range of intestinal disorders. While there is vast knowledge on the impact of Brachyspira infections in animals, little is known regarding human infections. Prevalence ranges from 1-32% in healthy human subjects and up to 63% in HIV-positive patients. Human intestinal spirochaetosis is thought to be caused by two Brachyspira species: Brachyspira aalborgi and Brachyspira pilosicoli. Diagnosis of human intestinal spirochaetosis is based on histopathology of colon-biopsies and does not allow for definitive species identification. Aim: To design a real-time PCR for the diagnosis and species identification in human intestinal spirochaetosis. Methods: Colon biopsies (n=50) from 25 histopathologically proven positive patients collected between 2001 and 2010 were identified from the archival collection of Tergooiziekenhuizen and the University Medical Centre Utrecht. As negative controls 16 histopathologically spirochaete negative colon-biopsy samples from 12 patients were used. All samples were revised by a trained pathologist to confirm the diagnosis. Biopsy samples were deparafinized and spiked with phocine herpes virus as internal control. DNA was isolated on a Roche MP96 and realtime PCR and SYBR-green melt-curve analysis of the PCR products was performed on a Roche LC 480. Species designation was confirmed by DNA sequence analysis of the PCR fragments. PCR design: Small subunit rRNA sequences from the public databases were downloaded and aligned with the Clone Manager program (Scientific & Educational Software, Cary, North Carolina, USA) in order to design a PCR primerset against a 136 bp region in the 5'end of the rRNA gene that is both specific for, and conserved within, all Brachyspira species. Results: All 50 samples from histopathologically positive patients were positive in the PCR, whereas all 16 negative controls were PCR-negative. SYBR-green based melt-curves of the 50 positive samples revealed the presence of three different Brachyspira species. Sequence analysis confirmed the presence of B. aalborgi (12/25), B. pilosicoli (3/25) and a thus far unreported Brachyspira species (3/25) and seven double infections: 3/25 B. aalborgi with the novel Brachyspira species, 2/25 B. pilosicoli with the novel species and 2/25 B. aalborgi with B. pilosicoli. Conclusions: The described real-time PCR allows for rapid, simple, and sensitive detection of Brachyspira species in routine biopsy materials. To our knowledge this is the first real-time PCR that allows for both detection and immediate species discrimination. In contrast to what is generally assumed we identified not two but three genetically different Brachyspira species in the human colon. In addition, a surprisingly high number of double infections was observed. Sa1984 Early Experience With a Fecal Bacteriotherapy (FB) Program for Recurrent and C-Difficile Infection (CDI) Gilbert M. Wilcox
Sa1982 A Prediction Model of Disease Severity in Clostridium difficile-Associated Disease Gene Y. Im, Rani J. Modayil, Martin Feuerman, Cheng Ting Lin, Anil R. Balani, Douglas S. Katz, James H. Grendell
Early Experience with a Fecal Bacteriotherapy (FB) Program for Recurrent c.difficile infection(CDI). Gilbert M Wilcox, Jennifer Lewis, Elizabeth Siefert, Michael Jones. Introduction: Treatments for recurrent CDI are limited and controversial. Fecal bacteriotherapy (FB) is a known but little used treatment because of concerns about efficacy, patient acceptance, and complications. This study reports on these issues with observations from a new FB program at Maine Medical Center, Portland ,ME (MMC). Rationale: Many cases of severe CDI were seen at MMC during the initial phase (2003-2004) of the epidemic of toxin gene-variant (BI/NAP1) CDI. Our index case for FB was a 60 year old woman with multiple comorbidities seen in 2004 with severe CDI unresponsive to 2 weeks of intensive medical therapy. All antibiotics were discontinued and she was treated with FB after which diarrhea, abdominal pain, fever and leukocytosis quickly resolved without further intervention. This dramatic outcome from FB encouraged interest in this unusual therapy which we have subsequently applied to our cases of recurrent CDI as described in this report. Methods: All patients had at least three episodes of recurrent CDI each developing after control of active disease. Patients were asymptomatic and off all antibiotics 48hr at the time of FB. Stool donors were family members or friends of the patient, and were evaluated for hepatitis, HIV, syphilis, stool pathogens, and CDI. Donors were interviewed about bowel movement patterns and problems with the donation experience. After bowel lavage the patients had colonoscopy exam to the cecum and all findings were recorded. The donated stool specimen was suspended in saline, filtered, and then infused through the colonoscope into the patient's cecum. Patients were followed by telephone interview. Results: 13 patients (9F/4M) had FB done between June 2009 and August 2010; patient age range: 27-93yr (mean 69.4); number of recurrences before FB: 3-30 (mean 6.5); time interval from first CDI to FB: 6-72 months (mean 17.5); number of hospitalizations for CDI: 0-4 (mean 1.6); mucosal inflammation present at FB: 1 of 13 (7.7%); number of recurrences after FB: 0 of 13 (0%); follow-up interval: 2-14 months (mean 8.4); complications: 0 of 13 (0%); patient complaints after FB: 0 of 13 (0%); donor complaints after FB: 2 of 13 (15%) (stress associated with donation). Conclusions: In this small prospective series FB appears to be effective, tolerated well by patients, and without complications. Some issues remain with the donation process. Observations of these difficult to treat patients with CDI are continuing.
Background: The incidence and case-fatality ratio of Clostridium difficile-associated disease (CDAD) have increased, and identifying those at risk for serious complications of CDAD such as colectomy or death is crucial. Several unvalidated severity scoring systems have been published to identify high risk patients. They are limited by complexity of use, subjective variables and low predictive capabilities in part related to suboptimal measurements of CDAD outcomes. The purpose of this study was to examine the correlation of demographic, clinical, laboratory, CT, pharmacologic and inpatient outcome variables with the severity of CDAD and to develop a prediction model of disease severity in CDAD. Methods: We reviewed the records of 396 adult inpatients with positive C. difficile toxin assay from 2005 to 2007. In the cohort undergoing CT, patients were included if the CT was performed within 7 days of initial positive assay. The primary endpoint was inpatient mortality from CDAD. A rigorous independent review determined if mortality was due primarily to CDAD, with a consensus reached in unclear cases. CT data were examined blinded to outcomes. Univariate and multivariate statistical analyses were performed to devise a prediction model of disease severity in CDAD. Results: 218 patients underwent a CT within 7 days of a positive toxin assay and 178 did not. More than 40 variables were examined, out of which the 10 most significant on multivariate analysis were used to construct the prediction model. A 2-variable prediction model was devised consisting of two laboratory variables (OR 8.92 and 8.06, respectively): risk score = 1 x (WBC ≥30 x 103/μL) + 1 x (BUN ≥40 mg/dL), with presence of the variable entered as 1. It was highly predictive of inpatient mortality from CDAD with an AUC of 0.866. An increasing risk score was associated with an increasing probability of observed inpatient mortality from CDAD. A risk score of 0 conferred a 0% probability, a score of 1, a 13% probability and a score of 2, a 50% probability of mortality from CDAD, respectively. By classifying patients with 0 variables as low risk and ≧1 variable as high risk, the sensitivity and specificity of the prediction model was 100% and 62%, respectively. The specificity of the model increased to 82.4% by adding a third CT variable, the presence of moderate and severe pericolonic stranding, with an AUC of 0.894. Conclusions: Using a rigorous methodology in the largest study to date examining the association of CT and clinical variables with outcomes in CDAD, a simple, cheap, universally available, prediction model of disease severity in CDAD was devised. The ability to stratify patients with CDAD into mild or severe disease could identify high risk patients for aggressive treatment and provide a framework upon which future studies of therapeutic efficacy could be based.
Sa1985 Antimicrobial Peptides Are Differentially Expressed in Clostridium difficile Infection Renate Schlottmann, Laura Martin, Andrea Tannapfel, Wolfgang E. Schmidt, Jan-Michel Otte Background & Aims: Clostridium difficile (C. difficile) is the leading cause of nosocomial infectious diarrhea in the USA and Europe. Over the last years C. difficile infection (CDI) exhibits increasing incidence with raising disease severity and mortality as well as outbreaks with more virulent strains. Further understanding of the pathomechanisms involved might allow for the development of urgently needed alternativ preventive and therapeutic strategies.
S-361
AGA Abstracts
AGA Abstracts
Real-Time PCR Reveals the Presence of Three Distinct Brachyspira Species in Human Intestinal Spirochaetosis Laurens J. Westerman, Herbert V. Stel, Marguerite E. Schipper, Leendert J. Bakker, Edwin C. Boel, Jan M. Van den Brande, Peter D. Siersema, Johannes G. Kusters
Sa1983
Introduction: Infections with Brachyspira species result in a wide range of intestinal disorders. While there is vast knowledge on the impact of Brachyspira infections in animals, little is known regarding human infections. Prevalence ranges from 1-32% in healthy human subjects and up to 63% in HIV-positive patients. Human intestinal spirochaetosis is thought to be caused by two Brachyspira species: Brachyspira aalborgi and Brachyspira pilosicoli. Diagnosis of human intestinal spirochaetosis is based on histopathology of colon-biopsies and does not allow for definitive species identification. Aim: To design a real-time PCR for the diagnosis and species identification in human intestinal spirochaetosis. Methods: Colon biopsies (n=50) from 25 histopathologically proven positive patients collected between 2001 and 2010 were identified from the archival collection of Tergooiziekenhuizen and the University Medical Centre Utrecht. As negative controls 16 histopathologically spirochaete negative colon-biopsy samples from 12 patients were used. All samples were revised by a trained pathologist to confirm the diagnosis. Biopsy samples were deparafinized and spiked with phocine herpes virus as internal control. DNA was isolated on a Roche MP96 and realtime PCR and SYBR-green melt-curve analysis of the PCR products was performed on a Roche LC 480. Species designation was confirmed by DNA sequence analysis of the PCR fragments. PCR design: Small subunit rRNA sequences from the public databases were downloaded and aligned with the Clone Manager program (Scientific & Educational Software, Cary, North Carolina, USA) in order to design a PCR primerset against a 136 bp region in the 5'end of the rRNA gene that is both specific for, and conserved within, all Brachyspira species. Results: All 50 samples from histopathologically positive patients were positive in the PCR, whereas all 16 negative controls were PCR-negative. SYBR-green based melt-curves of the 50 positive samples revealed the presence of three different Brachyspira species. Sequence analysis confirmed the presence of B. aalborgi (12/25), B. pilosicoli (3/25) and a thus far unreported Brachyspira species (3/25) and seven double infections: 3/25 B. aalborgi with the novel Brachyspira species, 2/25 B. pilosicoli with the novel species and 2/25 B. aalborgi with B. pilosicoli. Conclusions: The described real-time PCR allows for rapid, simple, and sensitive detection of Brachyspira species in routine biopsy materials. To our knowledge this is the first real-time PCR that allows for both detection and immediate species discrimination. In contrast to what is generally assumed we identified not two but three genetically different Brachyspira species in the human colon. In addition, a surprisingly high number of double infections was observed. Sa1984 Early Experience With a Fecal Bacteriotherapy (FB) Program for Recurrent and C-Difficile Infection (CDI) Gilbert M. Wilcox
Sa1982 A Prediction Model of Disease Severity in Clostridium difficile-Associated Disease Gene Y. Im, Rani J. Modayil, Martin Feuerman, Cheng Ting Lin, Anil R. Balani, Douglas S. Katz, James H. Grendell
Early Experience with a Fecal Bacteriotherapy (FB) Program for Recurrent c.difficile infection(CDI). Gilbert M Wilcox, Jennifer Lewis, Elizabeth Siefert, Michael Jones. Introduction: Treatments for recurrent CDI are limited and controversial. Fecal bacteriotherapy (FB) is a known but little used treatment because of concerns about efficacy, patient acceptance, and complications. This study reports on these issues with observations from a new FB program at Maine Medical Center, Portland ,ME (MMC). Rationale: Many cases of severe CDI were seen at MMC during the initial phase (2003-2004) of the epidemic of toxin gene-variant (BI/NAP1) CDI. Our index case for FB was a 60 year old woman with multiple comorbidities seen in 2004 with severe CDI unresponsive to 2 weeks of intensive medical therapy. All antibiotics were discontinued and she was treated with FB after which diarrhea, abdominal pain, fever and leukocytosis quickly resolved without further intervention. This dramatic outcome from FB encouraged interest in this unusual therapy which we have subsequently applied to our cases of recurrent CDI as described in this report. Methods: All patients had at least three episodes of recurrent CDI each developing after control of active disease. Patients were asymptomatic and off all antibiotics 48hr at the time of FB. Stool donors were family members or friends of the patient, and were evaluated for hepatitis, HIV, syphilis, stool pathogens, and CDI. Donors were interviewed about bowel movement patterns and problems with the donation experience. After bowel lavage the patients had colonoscopy exam to the cecum and all findings were recorded. The donated stool specimen was suspended in saline, filtered, and then infused through the colonoscope into the patient's cecum. Patients were followed by telephone interview. Results: 13 patients (9F/4M) had FB done between June 2009 and August 2010; patient age range: 27-93yr (mean 69.4); number of recurrences before FB: 3-30 (mean 6.5); time interval from first CDI to FB: 6-72 months (mean 17.5); number of hospitalizations for CDI: 0-4 (mean 1.6); mucosal inflammation present at FB: 1 of 13 (7.7%); number of recurrences after FB: 0 of 13 (0%); follow-up interval: 2-14 months (mean 8.4); complications: 0 of 13 (0%); patient complaints after FB: 0 of 13 (0%); donor complaints after FB: 2 of 13 (15%) (stress associated with donation). Conclusions: In this small prospective series FB appears to be effective, tolerated well by patients, and without complications. Some issues remain with the donation process. Observations of these difficult to treat patients with CDI are continuing.
Background: The incidence and case-fatality ratio of Clostridium difficile-associated disease (CDAD) have increased, and identifying those at risk for serious complications of CDAD such as colectomy or death is crucial. Several unvalidated severity scoring systems have been published to identify high risk patients. They are limited by complexity of use, subjective variables and low predictive capabilities in part related to suboptimal measurements of CDAD outcomes. The purpose of this study was to examine the correlation of demographic, clinical, laboratory, CT, pharmacologic and inpatient outcome variables with the severity of CDAD and to develop a prediction model of disease severity in CDAD. Methods: We reviewed the records of 396 adult inpatients with positive C. difficile toxin assay from 2005 to 2007. In the cohort undergoing CT, patients were included if the CT was performed within 7 days of initial positive assay. The primary endpoint was inpatient mortality from CDAD. A rigorous independent review determined if mortality was due primarily to CDAD, with a consensus reached in unclear cases. CT data were examined blinded to outcomes. Univariate and multivariate statistical analyses were performed to devise a prediction model of disease severity in CDAD. Results: 218 patients underwent a CT within 7 days of a positive toxin assay and 178 did not. More than 40 variables were examined, out of which the 10 most significant on multivariate analysis were used to construct the prediction model. A 2-variable prediction model was devised consisting of two laboratory variables (OR 8.92 and 8.06, respectively): risk score = 1 x (WBC ≥30 x 103/μL) + 1 x (BUN ≥40 mg/dL), with presence of the variable entered as 1. It was highly predictive of inpatient mortality from CDAD with an AUC of 0.866. An increasing risk score was associated with an increasing probability of observed inpatient mortality from CDAD. A risk score of 0 conferred a 0% probability, a score of 1, a 13% probability and a score of 2, a 50% probability of mortality from CDAD, respectively. By classifying patients with 0 variables as low risk and ≧1 variable as high risk, the sensitivity and specificity of the prediction model was 100% and 62%, respectively. The specificity of the model increased to 82.4% by adding a third CT variable, the presence of moderate and severe pericolonic stranding, with an AUC of 0.894. Conclusions: Using a rigorous methodology in the largest study to date examining the association of CT and clinical variables with outcomes in CDAD, a simple, cheap, universally available, prediction model of disease severity in CDAD was devised. The ability to stratify patients with CDAD into mild or severe disease could identify high risk patients for aggressive treatment and provide a framework upon which future studies of therapeutic efficacy could be based.
Sa1985 Antimicrobial Peptides Are Differentially Expressed in Clostridium difficile Infection Renate Schlottmann, Laura Martin, Andrea Tannapfel, Wolfgang E. Schmidt, Jan-Michel Otte Background & Aims: Clostridium difficile (C. difficile) is the leading cause of nosocomial infectious diarrhea in the USA and Europe. Over the last years C. difficile infection (CDI) exhibits increasing incidence with raising disease severity and mortality as well as outbreaks with more virulent strains. Further understanding of the pathomechanisms involved might allow for the development of urgently needed alternativ preventive and therapeutic strategies.
S-361
AGA Abstracts
AGA Abstracts
Real-Time PCR Reveals the Presence of Three Distinct Brachyspira Species in Human Intestinal Spirochaetosis Laurens J. Westerman, Herbert V. Stel, Marguerite E. Schipper, Leendert J. Bakker, Edwin C. Boel, Jan M. Van den Brande, Peter D. Siersema, Johannes G. Kusters