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Predicting Recurrence of C. difficile Colitis Using Bacterial Virulence Factors: Binary Toxin Is the Key

  • 2012 SSAT Plenary Presentation
  • Published:
Journal of Gastrointestinal Surgery Aims and scope

Abstract

Background

Recurrent Clostridium difficile colitis is common, yet the ability to predict recurrence is poorly developed.

Methods

Patients ≥18 years of age treated at our institution for C. difficile of any severity were consecutively enrolled. C. difficile colitis was defined as symptoms of colitis with a positive PCR stool test. Each bacterial isolate was studied for virulence factors: tcdC mutations via PCR; the presence of genes for toxins A, B, and binary toxin using restriction fragment length polymorphism; and identification of ribotype 027 by PCR. Chi-squared tests, t tests, and logistic and linear regression were used to determine which virulence factors predicted recurrence.

Results

Sixty-nine patients (male, 57 %) were studied, with a mean age of 64 ± 13 years. Twenty-one (30 %) patients were initially diagnosed as outpatients. There was no difference (p > 0.05) between virulence factors among inpatients and outpatients. The presence of a binary toxin gene was the single virulence factor independently associated with recurrence (p = 0.02). The combination of a tcdC mutation with binary toxin gene resulted in the highest odds of recurrence (OR, 5.3; 95 % CI, 3.52–6.09).

Conclusion

Binary toxin gene is a predictor of recurrent infection. Its presence may require longer antibiotic regimens in an effort to lower already elevated recurrence rates.

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Authors and Affiliations

  1. Department of Surgery, Penn State College of Medicine, 500 University Drive, H137, Hershey, PA, 17033, USA

    David B. Stewart, Arthur Berg & John Hegarty

Authors
  1. David B. Stewart
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  2. Arthur Berg
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  3. John Hegarty
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Corresponding author

Correspondence to David B. Stewart.

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Discussant

Dr. Sekar Dharmarajan (St. Louis, MO): Thank you Mr. Chairman. While the surgical literature is replete with studies that correlate clinical patient factors with morbidity, need for surgery, and mortality from C. difficile infection, Dr. Stewart and colleagues have taken the novel and thought-provoking approach of characterizing bacterial virulence factors and correlating these with morbidity from C. difficile in the form of recurrent infection. I have three broad areas of comments/questions.

The first surrounds the definition of recurrent C. difficile colitis. The authors define recurrent C. difficile infection as two consecutive positive C. difficile stool samples no closer than 21 days apart. Without a negative intervening sample, how do we know this is recurrent C. difficile infection as opposed to persistent C. difficile infection, as we know that C. difficile is notoriously difficult to eradicate? Specifically, do you have any data on how far apart temporally the positive stool samples were in the patients with recurrent C. difficile? More interestingly, do you have any data on the bacteriology of the recurrent C. difficile isolates to see how they are compared to the original isolate?

The second area is with regard to the broader generalizability or applicability of the study. As the authors state in their manuscript, the prevalence of the binary toxin gene in this study was five- to tenfold higher than that previously reported in the literature. Similarly, ribotype 027, which has been found in previous studies to be hypervirulent, was not associated with any morbidity or mortality in the present study. Is it possible that, while binary toxin is the key to predicting recurrent infection at Hershey, these factors may differ institution to institution?

Finally, the last area is with regard to patient or host factors that certainly must contribute to morbidity and mortality from C. difficile infection. While the present study found no clinical factors that predicted recurrent infection, I wonder if the authors have plans or could comment on studying the interaction of patient genotypes at genetic loci that may render susceptibility to infection with bacterial virulence factors on morbidity from C. difficile. Similarly, as the authors comment in their manuscript, the host microbiome has become an increasingly important area of research in determining outcomes from a variety of disease processes, and I wonder if the authors had any comment on their plans to study this.

Thank you for the opportunity to review your extremely well-written manuscript.

Selection Bias: There is no patient with fulminant C. difficile colitis as defined by need for surgery or death.

Closing Discussant

Dr. David B. Stewart: I would like to thank the SSAT for the opportunity to present our research, and I would also like to express my gratitude to Dr. Dharmarajan for being willing to serve as a discussant for our presentation.

Dr. Dharmarajan’s first point is perhaps the most critical issue which he raised as a discussant. There are no consensus-based definitions, for all research endeavors on the topic, which define recurrent C. difficile colitis (RCDC) in any scientifically meaningful manner. This lack of standardization would potentially allow for an inflated estimate of RCDC, if the time interval between two consecutive CDC episodes was diminished to a great enough degree. In fact, in reviewing the literature which deals with recurrent C. difficile, the reader will encounter both heterogeneity in the definition of recurrence as well as a degree of arbitrariness which includes extremely short time intervals (<10 days) as well as criteria that range from clinical symptoms of diarrhea to tests confirming the presence of toxigenic C. difficile. In our case, it seemed appropriate to set the definition of RCDC at a point at least far enough after the diagnosis of the index episode to allow for the standard course of 10–14 days of antibiotics to be completed. We then allowed for an additional week of time to elapse in order to better ensure that we avoided collapsing recurrence into persistence. While our approach is not universal in acceptance, we believe it is fair and that it has a conceptual appeal given what we have described. Further, our recurrence rates fall into the same general range as those previously published in larger studies.

There are no data available to currently guide the definition of when persistence ends and recurrence begins, and so there are no data available to answer the issue of temporal relationships for persistence or recurrence. In part, this very relevant question will not be answered until we have large C. difficile registries that can provide, at minimum, regional epidemiologic information regarding endemic ribotypes and virulence characteristics. Such resources do not exist in this country, though they do exist in seminal forms in Canada.

The issue related to whether a recurrent episode of CDC is due to a different ribotype is something our group will be focusing on in the future. There are no relevant data on this topic currently available, though the issue is critical. Our group has unpublished data which demonstrate that C. difficile responds to drugs such as proton-pump inhibitors in a ribotype-dependent manner, such that some ribotypes will produce more toxins in the presence of PPIs and some will express toxin genes to a lesser degree. Some are actually unaffected by PPIs. If this is confirmed in larger studies, it will introduce an additional complexity in understanding C. difficile and in how environmental cues, including the drugs we use to eradicate the organism, may lead to different clinical outcomes. There is a biologic basis for such phenomena, and in the same manner that we push for personalized medicine with respect to diseases like cancer, we need personalized microbiology to a much greater degree than is presently available.

The second comment deals with whether those bacterial genetic signatures that were associated with higher rates of recurrence in our study might not be relevant in a different population of C. difficile. Until our findings are prospectively evaluated in a multiregional fashion, then this concern will not be definitively addressed. However, the idea that binary toxin would negatively affect clinical outcomes in CDC has scientific plausibility, given that previous reports have suggested that this is a more potent form of toxin.

Dr. Dharmarajan’s last comment deals with how host factors, including other microorganisms within the gut, may influence the behavior of C. difficile. The issue of the gut microbiome is certainly the hot topic of our day in microbiology, and for good reason. However, our understanding of the microbiome concept is currently so piecemeal that the more we learn, the less we know. There is at least one publication from several years ago which demonstrated a higher rate of RCDC when the human subject was found to harbor certain single nucleotide polymorphisms in the gene for IL-8, though this publication used a definition for recurrence, which may have lent toward artificially higher recurrence rates. Our own group is now using metabolomics to study how different environmental elements within the gut, such as varying levels of electrolytes and micronutrients, might affect transcriptional control of virulence genes in C. difficile. We have also begun obtaining rectal swabs on CDC patients in order to study the mRNA present in the organism at the time the sample is obtained, in an effort to study the “transcriptome” and how differential gene expression is associated with clinical outcomes such as recurrence, fulminant colitis, and death from CDC.

One final comment—these were consecutively enrolled patients which were presented in our study. Though none of our study patients required colectomy or died, there were patients with severe CDC based on the IDSA classification schema. Surgical intervention for CDC, while much more common than in previous eras, is still relatively infrequent, which makes studying this particular patient group more difficult when attempting to obtain informed consent for a stool sample is required and when the patient is critically ill.

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Stewart, D.B., Berg, A. & Hegarty, J. Predicting Recurrence of C. difficile Colitis Using Bacterial Virulence Factors: Binary Toxin Is the Key. J Gastrointest Surg 17, 118–125 (2013). https://doi.org/10.1007/s11605-012-2056-6

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