Fecal Calprotectin Level in Differentiating Between Inflammatory and Non-inflammatory Diarrhea in Patients With Nosocomial Diarrhea
Conditions: Diarrhea, Calprotectin
Sponsor: Mahidol University

Fecal calprotectin level in differentiating between inflammatory and non-
inflammatory diarrhea in patients with nosocomial diarrhea  
  
NCT number: not available   
Document date: January 24, 2019  
 
 

Study Protocol with Statistical Analysis Plan 
Backgound/Rationale 
Nosocomial diarrhea is defined as diarrhea that develops after 72 hours of hospitalization. This 
condition is common among hospitalized patients with reported prevalence of 14-21%. The 
causes of nosocomial diarrhea can be categorized into inflammatory and non-inflammatory 
causes. For inflammatory etiologies, the most concern is Clostridium difficile infection. Other 
infections such as cytomegaloviral enterocolitis can also occur because immune function can be 
suppressed during critical illness. Ischemic enterocolitis is another cause in the inflammatory 
group. The non-inflammatory causes include adverse effects of medications and tube-feeding 
diarrhea. Differentiating inflammatory diarrhea from non-inflammatory diarrhea is important. 
Investigations for distinguishing these two conditions are stool C.difficile toxin test and stool 
examination. Because the reported sensitivity of only 60-70% for C. difficile toxin test by EIA 
method, which is most widely used in many centers, and the diagnosis of CMV is based on 
pathological findings, inflammatory diarrhea cannot be excluded completely by the negative stool 
tests. Colonoscopy is still required in some cases. However, although colonoscopy is a low risk 
procedure, the risk may increase if performed in hospitalized patients, who may have many 
comorbidities. A noninvasive test that can better differentiate inflammatory from 
noninflammatory diarrhea could help physicians to perform colonoscopy more appropriately. 
Calprotectin is a protein found in human neutrophils, and it is released during active periods of 
inflammation of intestine. The sensitivity and specificity has been reported at 93% and 96%, 
respectively in differentiating inflammatory bowel disease from irritable bowel syndrome in 
outpatient setting. Hence, its benefit in differentiating inflammatory from noninflammatory 
diarrhea in nosocomial diarrhea is possible, but never been studied. In the present study, we aim 

to study the efficacy of fecal calprotectin in distinguishing inflammatory nosocomial diarrhea 
from non-inflammatory nosocomial diarrhea. Furthermore, the level of fecal calprotectin has been 
reported to be associated with the degree of intestinal inflammation. It could have a role in 
determine prognosis of the patients.  
Study design 
The study is a prospective observational study performed at tertiary care center from June 2019 to 
June 2020.  
Population 
1. Sample size 
In literature review, we found a study comparing fecal calprotectin levels between C. difficile 
associated diarrhea and antibiotic associated diarrhea. The median fecal calprotectin level in 
C.difficile associated diarrhea  was 684.8 (range:203.7 to 1581.0) and the median level in 
antibiotic associated diarrhea was 66.5(range :23.1 to 145.7). Using Mann-Whitney U test with a 
standard normal of 1.96, alpha error of 5%, and power of 80%, the calculated sample size is 
61patients in each group. 
2. Study population 
Inclusion Criteria: 
  1.Ageat least 18 years 
  2.Diarrhea more than 3 bowel movements per day which develops after 72 hours of admission 
Exclusion Criteria: 

  1.Intraabdominal pressure more than 12 mmHg 
  2.Patient on chemotherapy with neutropenia (ANC less than 1,000/mm3) 
Study procedures 
The patients with nosocomial diarrhea who are tested for C.difficile infection and stool 
examination will be recruited. Their leftover stool specimens about 100mg will be collected and 
frozen at -80 celsius degree until the end of study. All collected samples will be measured for 
calprotectin levels.  
The inflammatory diarrhea is defined if  
1) positive for C. difficile toxin or   
2) stool WBC more than 5/HPF or  
3) inflammatory mucosa or ulceration noted on colonoscopy 
The noninflammatory diarrhea is defined if  
1) negative for C. difficile toxin and  
2) no WBC on stool examination and  
3) dramatic and persistent response to diet adjustment  
Or  
4) no mucosal inflammation or ulceration if colonoscopy is performed 
The management and enrollment flow chart is shown in Figure 1.  

 
Data collection process 
All patients are required to consent to the study. Their stool samples will be routinely sent for 
microscopic examination (cell count and parasites) and C.difficile toxin. Fecal calprotectin will 
be kept at -80 celsius degree and will be measured for fecal calprotectin at the end of study. The 
patients baseline characteristics, indication of hospitalization, treatment, and clinical progression 
will be obtained. All patients will be managed by primary physicians who do not know the 
results of fecal calprotectin level.  
This protocol has been approved by Siriraj Institutional Review Board on January 24, 2019. The 
approval number is 718/2561(EC4).  
Outcome Measures 
Primary outcome:  To determine the efficacy of fecal calprotectin in distinguishing inflammatory 
nosocomial diarrhea from non-inflammatory nosocomial diarrhea. 

Secondary outcome: Correlation of fecal calprotectin levels and prognosis of hospitalized patients 
with nosocomial diarrhea. 
The participants with nosocomial diarrhea were tested for C.difficile and stool examination or 
further colonoscopy for definite diagnosis group of diarrhea. Their leftover specimens will be 
kept until the end of the study. Then all collected stool specimens will be measured for 
calprotectin levels. Overall data collected of 147 participants will be completed analyzed about 
August 2020. 
Statistical Analyses 
The continuous data will be reported in mean and standard deviation or median and range as 
appropriate. 
Categorical data will be reported in percentage or frequency. 
Comparision of factors and patient characteristic between inflammatory and non-inflammatory 
diarrhea groups will be analyzed using independent t-test or Wilcoxon Rank Sum test for 
continuous variables, and chi-square test or fisher exact test for categorical variables. 
Fecal calprotectin levels between inflammatory and non-inflammatory diarrhea will be shown in 
receiver operating characteristic curve(ROC). The best cut off will be calculated. 
Correlation of fecal calprotectin levels and treatment outcomes, both univariate and multivariate 
analysis, will be analyzed by logistic regression. 
P value < 0.05 will be considered statistically significant. 
Adverse Event Information 

This study only collects leftover stool samples without any intervention done to the patients. 
Therefore, there will be no additional risks to the patients apart from the standard care.   
Limitations and Caveats 
1.Some participants cannot undergo colonoscopy for definite diagnosis of nosocomial diarrhea 
due to risk of procedures. These patients will be excluded from the study.  
2.Number of participants in two groups were not equal as this is an observational study.  
Certain Agreements   
The principal investigator is not an employee of the sponsor.  
Name or Official title 
Julajak Limsrivilai, MD, MSc  
Organization Name 
Division of Gastroenterology, Department of Internal Medicine, Siriraj Hospital, Mahidol 
University, Bangkok, Thailand 
Department of Microbiology, Siriraj Hospital, Mahidol University, Bangkok, Thailand  
Phone Number: 02-4197281 
Email: alimsrivilai@gmail.com