Conditions: Antibiotic Resistant Infection Nosocomial Infection Pathogen Transmission Nutrition Disorders Critical Illness Sepsis
Keywords: Microbiome Antibiotic resistance Intensive care unit Sepsis Nutrition Pathogen colonization
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 94
Healthy volunteers: no

Brief summary:
Normal gut bacteria prevent colonization and subsequent infection with MDR organisms (MDROs) through competition for resources and other mechanisms. During critical illness, this function of the microbiome is lost and there are no current treatments to restore it. Preliminary data indicates that the prebiotic fiber inulin is safe and may alter the gastrointestinal microbiome to improve gut barrier function, decrease colonization with MDROs, and reduce downstream risk for intensive care unit (ICU)-acquired MDR infections. However, the impact of inulin during critical illness is unknown. This double-blind, randomized clinical trial will test inulin for the prevention of antibiotic resistant infections in the ICU.

The trial's specific aims are to determine (1) the feasibility, tolerability, and safety of inulin in the intensive care unit; (2) the impact of inulin on gut colonization with antibiotic-resistant pathogens; and (2A/exploratory) the impact of inulin on ICU-acquired antibiotic-resistant infections.

Detailed description:
The proposed trial hypothesizes that inulin maintains short-chain fatty acid (SCFA)-producing colonic anaerobes and that these bacteria are protective against multi-drug resistant organism (MDRO) colonization and subsequent MDR infection. Inulin, a vegetable-derived non-digestible polysaccharide is well established as the key nutrient source for SCFA-producing bacteria. Previous human studies have shown that (1) inulin increases levels of SCFA producers and SCFAs and (2) that this increase correlates with improved colonic mucosal integrity and resistance to MDR pathogens. In animal studies, inulin improves survival after pathogen challenge or injection with lipopolysaccharide. The overall aim of this clinical trial is to determine whether inulin improves gut colonization resistance against antibiotic-resistant pathogens and therefore prevents antibiotic-resistant infections in the setting of critical illness. To accomplish this, 90 critically ill adults who are receiving broad-spectrum antibiotics will be blindly randomized 1:1:1 to receive placebo, inulin 8 g twice daily, or inulin 16 g twice daily for a minimum of 7 days, with bedside follow-up extending to 30 days or hospital discharge.

Arms:
arm 1: Critically ill adults who are receiving broad-spectrum antibiotics will also receive inulin oral suspension (16g twice daily) for a minimum of 7 days. arm 2: Critically ill adults who are receiving broad-spectrum antibiotics will also receive inulin oral suspension (8g twice daily) for a minimum of 7 days. arm 3: Critically ill adults who are receiving broad-spectrum antibiotics will also receive placebo oral suspension for a minimum of 7 days.

Interventions: intervention 1: Inulin Oral Suspension intervention 2: Placebo Oral Suspension intervention 3: Broad-spectrum antibiotics
Intervention descriptions: intervention 1: Inulin powder, derived from chicory root and re-suspended in 250cc water Dosage will be either 8g twice a day or 16g twice a day - dissolved in 250cc sterile water, given oral or via enteric tube intervention 2: 250cc sterile water alone, given twice daily a sweetener is added to the water to create identical flavor intervention 3: Standard of care treatment for infections

Protocol document:
Prot_SAP_001.pdf: 
Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 1 of 37 
 
 
 
 
 
 
CLINICAL TRIAL PROTOCOL 
 
 
 
 
 
 
 
Prebiotic Inulin to Limit Antimicrobial-Resistant Infections During Critical Illness:  
A Phase 2 Clinical Trial 
 
 
 
 
 
 
Draft Date: February 24, 2020 
 
Version 5 
 
 
 
 
 
 
 
Columbia IRB: AAAS2576 
FDA IND: 
 
DoD PRMRP: PR181960 
DoD HRPO: Log Number E00684.1a 
ClinicalTrials.gov: NCT03865706 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 2 of 37 
 
1. Table of Contents 
 
1. Table of contents 
Page 2 
2. Protocol synopsis 
Page 3 
     2.1. Study diagram 
Page 4 
3. Administrative information 
Page 5 
4. Funding 
Page 5 
5. Background and rationale 
Page 5 
6. Previous data 
Page 6 
7. Study objectives 
Page 10 
8. Study design 
Page 11 
9. Intervention 
Page 11 
10. Outcomes 
Page 13 
11. Study population 
Page 16 
12. Enrollment strategy 
Page 17 
13. Study assessments 
Page 19 
14. Data collection and management 
Page 23 
15. Safety and adverse events 
Page 23 
16. Termination and completion 
Page 25 
17. Statistical methods 
Page 25 
18. Publication policy 
Page 27 
Appendices 
 
Appendix A: References 
Page 28 
Appendix B: Abbreviations and terms 
Page 36 
 
 
 
 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 3 of 37 
 
2. Protocol synopsis 
Title 
Prebiotic Inulin to Limit Antimicrobial-Resistant Infections During Critical 
Illness: A Phase 2 Clinical Trial 
Short title 
Inulin in the ICU 
Design 
Randomized, double-blind, placebo-controlled trial in the ICU 
Outcomes 
Primary outcome: 
Change in relative abundance of SCFA producers through ICU Day 3 
Secondary outcome: 
MDRO colonization status on ICU Day 3 
Additional outcomes: 
1. Feasibility 
2. Rates of pre-specified AEs 
3. Antibiotic resistance gene fraction on ICU Day 3 
4. Culture-proven MDR infections through ICU Day 30 
5. Fecal SCFA levels 
6. Proportion of goal calories through ICU Days 3 and 7 
7. Length of ICU stay 
8. Length of hospital stay 
9. Mortality through ICU Day 90 
10. Self-assessed quality of life at 6 months 
11. Times series data including ICU Day 7 outcomes 
Intervention 
1:1:1 allocation to inulin 32 g/day, inulin 16 g/day, or placebo dissolved in 
250 cc sterile water and given in 2 daily doses for 7 days or until 
death/hospital discharge  
Sample size 
90 critically ill patients meeting Sepsis-3 criteria 
Eligibility criteria Inclusion criteria: 
1. Medical ICU 
2. Age ≥ 18 years old 
3. Sepsis, defined according to the Sepsis-3 (2016) consensus as a 
known or suspected infection with a SOFA score of ≥2 points 
above baseline  
4. Broad-spectrum antibiotics, received within the last 24 hours or 
ordered and pending administration 
5. Able to complete enrollment within 24 hours of ICU admission for 
administration of the intervention within 48 hours of ICU admission 
Exclusion criteria: 
1. Inability to receive oral or enteric fluids 
2. Inulin allergy 
3. Hyponatremia (serum sodium ≤128 mEq/L)  
4. Immunosuppression, defined as CD4 count <200 or absolute 
neutrophil count <500 or reasonable expectation of either during the 
study intervention period 
5. Surgery involving the intestinal lumen within 30 days or known 
intestinal strictures 
6. Do Not Resuscitate (DNR) or Do Not Intubate (DNI) status, or “no 
escalation of care” orders 
7. Lack capacity for consent and no appropriate Legally Authorized 
Representative (LAR). 
 
 
 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
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2.1 Study diagram 
 
 
 
 
 
 
 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 5 of 37 
 
3. Administrative information 
 
Primary 
Investigator 
Daniel E. Freedberg 
def2004@cumc.columbia.edu 
Columbia University Medical Center 
Division of Digestive and Liver Diseases 
622 West 168th Street, P&S Building 3-401, New York, NY 10032 
Research office: (212) 342-3302, Cell: (857) 998-9370 
Co-Investigators 
All at Columbia University Medical Center 
 
Julian Abrams: ja660@cumc.columbia.edu 
David Chong: dhc5@cumc.columbia.edu 
Zhezhen Jin: zj7@columbia.edu  
Anne-Catrin Uhlemann: au2110@@cumc.columbia.edu 
Data and Safety 
Monitoring 
Officer 
Daniel Brodie 
hdb5@cumc.columbia.edu 
Columbia University Medical Center 
Vivian and Seymour Milstein Family Heart Center 
173 Fort Washington Avenue, 4-005A, New York, NY 10032 
Phone: (212)305-9817 
Other Key 
Personnel 
Howard Andrews, Columbia University Data Coordinating Center, 
hfa1@columbia.edu 
Research Nurse to be hired 
 
4. Funding 
This project is funded by a Department of Defense Peer-Reviewed Medical Research Program 
Clinical Trial Award (PR181960) recommended for funding on February 20, 2019. 
 
4.1 Role of funding bodies 
The study will be designed and conducted, and the results analyzed, presented, and published 
by the investigators independent of the funding bodies. 
 
5. Background and rationale for study 
 
5.1 Background 
Multidrug resistant (MDR) bacterial infections are a leading cause of morbidity, death, and 
healthcare-associated costs.1-3  Nowhere is this more important than in the intensive care unit 
(ICU), where over half of all infections are MDR4 and where sepsis from MDR bacteria confers 
10-20% absolute increased mortality.5-7  
 
 
This project hypothesizes that antibiotic resistance in the ICU is a problem of the human 
gastrointestinal microbiome, which is the critical reservoir for the bacteria and plasmids that 
encode antibiotic resistance genes.8-10  The normal gut microbiota prevents colonization and 
subsequent infection with MDR organisms through competition for scarce resources and other 
mechanisms.11  In the setting of critical illness, normal colonization resistance is lost and there 
are no currently available therapies to restore it.12-15 
 
Specific gut bacteria contribute to colonization resistance.  ICU patients have low levels of 
butyrate and other short chain fatty acids (SCFAs) which are derived from dietary fiber by 
anaerobic bacteria primarily within the Clostridial Clusters IV and XIVa (e.g., Faecalibacterium 
prausnitzii).  SCFAs moderate colonic inflammation,16 enlarge the pool of regulatory T cells,17 and 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
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contribute to the thickness of the colonic mucus layer and pathogen resistance.18  During critical 
illness, levels of SCFA-producing bacteria and SCFAs are low, and these levels further decline 
with antibiotic treatment.  Loss of SCFA-producing bacteria correlates with an increase in 
colonization with vancomycin-resistant Enterococcus and other antibiotic-resistant pathogens.  
We believe that preservation of SCFA-producing bacteria during critical illness is of major 
importance and can prevent the proliferation of colonizing organisms with antimicrobial 
resistance.   
 
The prebiotic fiber inulin is a vegetable-derived polysaccharide that restores gastrointestinal 
colonization resistance against MDROs.  SCFAs cannot be easily given to humans due to oral 
intolerability and rapid metabolism,19,20 but inulin promotes growth of SCFA producers to raise 
SCFA levels.  In animals, inulin-type fibers improve survival after pathogen challenge21,22 even in 
the face of antibiotics.18,23-25  In a randomized trial of 30 outpatients, inulin increased fecal levels 
of SCFA producers by 35-90%, with a 3-fold decrease of serum markers of bacterial translocation 
(e.g., lipopolysaccharide) relative to placebo.26  Inulin is safe in ambulatory patients27-35 with 
biologic effects seen at 16 g/day.26  In the ICU, inulin-type fibers have been given up to 36 g/day 
without adverse events36  and in our ICU cohort we have observed 30 g/day dietary fiber without 
intolerance.  This trial will test whether inulin supplementation has similar effects in the ICU as it 
does in animal models and as it appears to in healthy volunteers: will it result in higher levels of 
SCFA-producing Clostridia, decreased gastrointestinal antibiotic resistance and lower risk for 
MDR infections? 
 
5.2 Trial purpose and design 
The purpose of this Phase 2 trial is to determine the feasibility and safety of inulin for the 
prevention of antibiotic resistant infections or colonization in critically ill patients.  We will perform 
a double-blind, randomized controlled trial of inulin in the ICU with three arms (placebo, inulin 8 g 
twice daily, and inulin 16 g twice daily).  Subjects will be treated with inulin or placebo for 7 days 
with bedside follow-up for 30 days or hospital discharge and with long-term telephone follow-up 
for 6 months. 
 
6. Previous data 
Our observational ICU data supports the importance of SCFA producers in the ICU.  In animals, 
inulin protects against sepsis.  Inulin increases SCFA producers in ambulatory studies and inulin-
like fibers are safe in the ICU setting. 
 
6.1 Evidence supporting the importance of SCFA producers in the ICU 
We have assembled two large prospective cohorts now totaling over 500 ICU patients who have 
contributed rectal swabs and other data.37-39  Results from these cohorts confirm the importance 
of SCFA producers in protecting against pathogen colonization and infection in the ICU.   
 
 
Cohort #1.40  A single rectal swab was performed on medical ICU admission for consecutive 
patients.  Swabs were cultured for VRE and residual DNA was extracted for 16S rRNA gene 
sequencing; patients were then followed for up to 30 days for death or culture-proven 
infections.  In 9 months, 301 patients were enrolled of whom 91 (30%) were VRE colonized.  
Thirty-day mortality was 25% and an overlapping 21% patients developed culture-proven 
MDR infections. 
 
Cohort #1 demonstrated that the bacteria causing infections in the ICU are present as gut 
colonizers at the time of admission.  We tested whether colonizing MDROs predicted subsequent 
death or culture-proven infection.  Presence of the colonizing organism at ICU admission was 
almost always associated with subsequent organism-specific infection, even after adjusting for 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
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acute severity illness as captured by the Simplified 
Acute Physiology Score-3 (SAPS-3)41,42 and other 
relevant clinical variables.  This was true for VRE 
(aHR 5.85, 95% CI 2.25-15.2, 16 infections), C. 
difficile (aHR 19.0, 95% CI 4.40-81.8, 9 infections), 
Pseudomonas (aHR 2.37, 95% CI 1.04-5.43, 28 
infections), and Klebsiella spp. (aHR 3.81, 95% CI 
1.10-13.2, 28 infections).  To interrogate the process 
by which MDRO colonization may lead to infection, 
we also examined MDRO domination defined as 
≥30% 16S reads corresponding to a potential 
pathogen.43,44  Because this cohort contained VRE 
culture data, we were able to test whether patients 
who had Enterococcus domination were in fact 
colonized by VRE.  Enterococcus domination (15% 
of the cohort) and VRE colonization (30%) both 
predicted all-cause infection (Figure 1).  When 30-day mortality was examined as a stand-alone 
outcome, VRE colonization/domination predicted 40% increased mortality, after adjusting for 
SAPS-3 score and other factors.   
 
Next, we sought to determine the relationship between SCFA producers and VRE based on 
culture at the time of ICU admission.  Only VRE was cultured within this cohort, so we did not 
have the ability to assess colonization with other MDROs using culture (e.g., Gram negative 
bacteria).  Based on the 16S sequencing results, SCFA producers were classified as absent 
(<.01% relative abundance), low (.01% to 1%), or high (>1%) after Haak et al.45  High levels of 
SCFA producers were associated with lower prevalence of VRE colonization based on culture: 
40% when SCFA producers were absent, 27% when they were low, and 22% when they were 
high (p<.01).  Results were similar for Enterococcus domination. 
 
 
Cohort #2.37,39  This cohort was designed to examine dynamic changes in the ICU 
microbiome, focusing on SCFA producers and MDROs.  Swabs were performed in duplicate 
at ICU admission and 72 hours later (±4 hours).  Swabs were 
then (1) cultured for MRSA, VRE, and MDR Gram negatives 
and (2) 16S sequenced.  In 3 years, 230 patients out of a 
target of 300 have been enrolled with 126 sequenced to date. 
 
Cohort #2 demonstrated that SCFA-producing bacteria associate 
with preserved colonization resistance.  Within cohort #2, we first 
performed an unbiased analysis using LEfSe46 to ask which 
bacterial taxa changed most within individuals, comparing their 
admission and Day 3 samples.  14 taxa were identified as having 
significantly changed at a false-discovery adjusted p<.05.  Ten 
taxa decreased over these first 72 hours of which 6 were SCFA 
producers.  These changes were substantial in magnitude (e.g., 
8-fold for F. prausnitzii). 
 
We wanted to know how SCFA producers associated with MDRO 
colonization.  SCFA producers were again classified as absent, 
low, or high based on sequencing.  MDROs were classified as 
present vs absent based on culture results.  At both ICU 
admission and 72 hours later, there was a stepwise relationship 
Figure 1.  VRE colonization/domination was 
associated with death or culture-proven 
infection in the ICU.  VRE colonization or 
domination was assessed at the time of ICU 
admission and patients were followed for all-cause 
culture-proven infection. 
Figure 2.  Higher levels of SCFA-
producing 
bacteria 
associated 
with lower colonizing MDROs, 
both at the time of ICU admission 
and 72 hours later.  Lines are 95% 
confidence estimates.  


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 8 of 37 
 
where higher SCFA producers was associated with 
decreased MDRO colonization (Figure 2, previous 
page).  Comparing high to absent SCFA levels, the 
difference in rates of MDRO colonization was over 
25%. 
 
Last, we sought to retrospectively assess the impact 
of dietary fiber on SCFA producers and MDROs.  
Observed fiber intake was low overall with a mean 
of 13.4 g over 72 hours, but some patients did 
receive large amounts of fiber (maximum 30 g/24 
hours).  There was no intolerance of fiber based on 
documentation of abdominal distension, diarrhea, or 
nausea/vomiting.  Fiber intake was classified into 
tertiles and SCFA producers were examined as a 
continuous measure based on 16S sequencing.  At 
ICU admission, baseline relative abundance of 
SCFA producers was similar across fiber intake 
groups.  After 72 hours, higher fiber intake was 
associated with higher levels of SCFA producers 
(Figure 3).  Higher fiber was also associated with 
less Enterococcus (median abundance .08% high 
fiber vs .35% no fiber, p=.03).  This data is observational so there were baseline patient 
differences between those who did and did not receive high dietary fiber.  To account for this, we 
built a multivariable model adjusted for acute severity of illness, receipt of antibiotics, and total 
caloric intake.  In this model, there was a 0.30% median increase in SCFA producers after 72 
hours per additional 10g of fiber intake (p<0.01).  This suggests that dietary fiber has the potential 
to impact levels of SCFA producers, even during critical illness and with receipt of broad-spectrum 
antibiotics (78% of the cohort). 
 
6.2 Inulin in animal models 
Animal models show protective effects of inulin or similar compounds, which appear to be 
mediated by an increase in SCFA producers.47,48  Importantly, these studies demonstrate that 
inulin is likely to be effective even when given concurrently with antibiotics.  Animal models include 
cecal ligation and puncture (CLP),21 lipopolysaccharide (LPS) injection,22 challenge with  
Citrobacter rodentium18 or pathogenic Escherichia coli strains,47 and Salmonella typhimurium 
challenge with23 or without antibiotics.24,25  All models have their strengths and weaknesses, so it 
is remarkable that results align well across them.  Inulin and similar fructo-oligosaccharides (FOS) 
seem to confer protective effects through multiple mechanisms all of which likely depend on SCFA 
fermenters.  First, they maintain the protective colonic mucus layer.  Inulin fermenters activate 
carbohydrate metabolism pathways and switch fuel utilization from dietary fiber to colonic mucus 
when inulin-like fibers are unavailable, causing thinning of the colonic mucus layer.18,23  Second, 
the bacteria nourished by inulin have direct antagonistic effects on pathogens25 by making 
protonated SCFAs which diffuse across Gram negative cell walls to delay pathogen growth.24  
Luminal acidification by SCFAs also drives broad microbiome shifts towards relatively pH-tolerant, 
Gram positive obligate anaerobes.49,50  Third, SCFA producers induce host immunity.  When 
SCFA-producing anaerobe levels increase during sepsis, there is induction of T regulatory cells 
(Tregs), reduction in activated dendritic cells, and downstream decreased DNA binding of NF-kB 
for an overall reduction in pro-inflammatory cytokines and reduced T cell anergy.22  In these 
animal studies, the protective effect is large, with dramatic differences in outcomes including 
weight loss, colitis severity, and mortality. 
 
Figure 3. Higher fiber intake associated with 
higher levels of SCFA producers in the ICU.
Fiber intake was classified into tertiles, with relative 
abundance of SCFA producers classified based on 
16S sequencing. 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
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6.3. Inulin in the ambulatory setting 
Inulin has been studied in over 400 ambulatory patients where it has shown excellent safety and 
tolerability.27-35  No serious adverse events have been reported in trials using doses up to 34 g/day 
inulin51 or 55 g/day mixed fiber,16 with self-resolving bloating the main side effect.  Three trials 
specifically examined outcomes related to the ability of inulin to prevent colonization or infection.  
First, Dewulf et al. randomized 30 patients  to inulin 16 g/day vs placebo; the inulin group had a 
doubling of F. prausnitzii and other SCFA-producers and concurrent decreases in serum CRP 
and LPS concentration.26  Second, Lecerf et al. randomized 60 volunteers 1:1:1 to placebo, a 
combination xylo-oligosaccharide (XOS), or inulin-XOS.52  The inulin group had the highest post-
treatment SCFA levels and butyrate levels, the lowest circulating LPS, and decreased gene 
expression for the pro-inflammatory cytokines IL-1β, TNFα, and IL-12.  Third, Miguez et al. found 
that an inulin-like fructo-oligosaccharides (FOS) preserved gut microbiome diversity after broad-
spectrum antibiotics, mitigated loss of SCFA producers, and increased butyrate levels 3-fold.53  
Table 1 lists selected studies of inulin in the ambulatory setting, focusing on those with results 
most relevant to this project. 
 
Table 1. Selected ambulatory trials testing inulin alone or in combination 
Author 
Subjects 
Intervention 
Effect 
AEs 
Kruse51 
8 healthy 
volunteers 
Inulin 34 g/day 
Increased Bifidobacteria, 
unchanged SCFA levels 
Self-resolving bloating 
and flatulence. 
Dewulf26 
30 obese 
women 
inulin 16 g/day (50% 
FOS enriched) x 3 
months 
Increased Bifidobacteria 
and F. prausnitzii and 
decreased LPS. 
Self-resolving bloating 
and flatulence. 
Lecerf52 
60 healthy 
volunteers 
inulin 3g/day plus 
1g/day XOS 
Increased fecal SCFAs, 
decreased LPS, 
attenuated LPS-induced 
serum changes in IL-1b 
and IL-13. 
Self-resolving bloating 
and flatulence, small 
decrease in general 
well-being. 
Azpiroz27 
36 with IBS 
inulin 8g/day x 4 
weeks 
Increased Bifidobacteria. 
None reported. 
Canfora29 
44 obese pre-
diabetic adults 
inulin 15g/day x 12 
weeks 
Increased Bifidobacteria. 
None reported. 
Garcia-Peris30 
31 adult 
women with 
gynecological 
cancer 
inulin 12g/day x 4 
weeks (50% FOS 
enriched) 
Increased in 
Lactobacillus and 
Bifidobacteria. 
Self-resolving bloating. 
Holscher31 
29 healthy 
adults 
inulin 7.5g/day x 21 
days (agave-
derived) 
Increased Bifidobacteria. 
None reported. 
Petry32 
32 obese 
female adults 
inulin 20 g/day x 4 
weeks 
Increased Bifidobacteria. 
None reported. 
Rahat-
Rozenbloom54 
25 healthy 
obese or lean 
adults 
inulin 24 g/day x 3 
days 
Increased serum SCFAs. 
None reported. 
Ramirez-Farias33 
12 healthy 
adults 
inulin 10g/day x 16 
days 
Increased Bifidobacteria 
and F. prausnitzii, no 
change in fecal SCFAs. 
None reported. 
Vandeputte35 
42 healthy 
adults 
inulin 12g/day x 4 
weeks 
Increased Bifidobacteria. 
None reported. 
 
6.4 Inulin-like fibers in the ICU 
Inulin has not been studied in the ICU but data can be extrapolated from ICU trials of similar 
fructo-oligosaccharides (FOS).  To date, there are no reports of serious adverse effects or major 
intolerance with over 1,000 patients studied.55  Doses have been used up to 36 g/day for 5 
weeks.36  This emerging safety data is robust enough that consideration of supplemental fiber has 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
Page 10 of 37 
 
been incorporated into relevant ICU nutrition guidelines.56  
Data on the effects of inulin-like FOS on infections in the 
ICU is scarce but promising.  Three double-blind, 
randomized ICU trials tested a commercial synbiotic 
containing Lactobacillus spp. and mixed soluble prebiotic 
fiber including some inulin (up to 20 g/day total).57-59  In 
these studies, the overall rates of ICU-acquired infections 
were 16-27% lower in the group receiving prebiotic fiber, 
although MDR infections were not separately examined.  
In an open-label trial of mixed fiber up to 24 g/day in 30 
critically ill adults, the median hospital length of stay was 
5 days shorter in the fiber group and the overall rate of 
complications, including feeding-related complications 
was lower.60  This provides important reassurance given the concerns occasionally raised 
regarding prebiotics in patients with altered small bowel or colonic motility.61,62  Table 2 lists 
selected studies of fiber supplementation in the ICU, focusing on those with results most relevant 
to this project 
 
Table 2.  Selected ICU-based trials testing inulin or similar fibers, ordered by decreasing amounts of fiber. 
Author 
Subjects 
Intervention 
Effect 
AEs 
O'Keefe36 
13 ICU patients 
(most with 
pancreatitis) 
32 g/day wheat dextrin up to 5 
weeks 
Higher fecal SCFA 
producers and SCFAs. 
None reported. 
Karakan60 
30 ICU patients 
with pancreatitis 
24 g/day mixed soluble and 
insoluble fiber (open label) 
Shorter hospital stays 
and lower complications. 
Self-resolving 
bloating. 
Knight58 
259 ICU 
patients on 
ventilation 
20 g/day mixed fiber synbiotic 
including 5 g/day inulin 
Trend towards lower 
rates of VAP. 
None reported. 
Kotzampassi57 
65 ICU patients 
with SIRS 
20 g/day mixed fiber synbiotic 
including 5 g/day inulin 
Fewer infections and less 
diarrhea. 
None reported. 
Majid63 
22 adults in the 
ICU 
inulin 7g/day 
No difference in 
Bifidobacteria and lower 
F. prausnitzii. 
None reported. 
Spapen64 
25 ICU patients 
with septic 
shock 
22 g/L guar fiber enteral 
feeding, dose individualized 
Less diarrhea. 
None reported. 
Spindler-
Vesel59 
113 ICU trauma 
patients 
17 g/day average mixed 
soluble fiber-synbiotic 
Fewer infections. 
Higher gastric 
residuals. 
 
6.5 Summary of previous experience with inulin 
Our prospective ICU cohort data provides real-world data supporting the hypothesis that inulin, 
by nourishing SCFA producers, will lead to decreased pathogen colonization and infection in the 
ICU.  Animal studies show that inulin confers protection across various pathogen challenge 
models and sepsis models including LPS injection and cecal ligation-and-puncture.  Ambulatory 
studies of inulin demonstrate safety and generally show increases in SCFA producers related to 
inulin.  Few ICU studies have tested inulin, but there is ample ICU experience with inulin-like fibers 
that provides reassurance that high doses of fiber are safe even in the context of sepsis and 
hypotension in the ICU. 
 
7. Study objectives 
The short-term goal of the study is to test inulin for the prevention of MDRO colonization and 
infection in the intensive care unit.  The long-term goal of this research is to develop antibiotic-
free strategies that prevent or ameliorate ICU-acquired infections. 
 


Inulin in the ICU 
Protocol Version 5 – February 24, 2020 
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8. Study Design 
This will be a double-blind, randomized, placebo-
controlled trial with two doses. 
 
9. Intervention 
Block randomization (blocks of six) will be performed with 
1:1:1 random computer allocation of subjects into the 
three study arms.  In the two inulin arms, chicory root-
derived inulin (Cargill Inc., Wayzata, MN) will be diluted in 
250 ml of sterile water and given at 8 or 16 g twice daily 
(Figure 4).  The placebo arm will receive the same 
volume of sterile water twice daily, with the intervention 
given for 7 days or until death or hospital discharge.  Prior 
studies suggest that these doses are likely to be safe and 
sufficient to confer benefit,26,36 even in the setting of 
critical illness.57,58,63,65  Subjects, investigators, and the 
treating ICU team will be blinded to the intervention arm, 
with blinding managed by the CUMC Research Pharmacy 
which will provide labeled, de-identified, semi-opaque 
fluid bags containing inulin/placebo in accordance with 
the U.S. Code of Federal Regulations governing labelling 
of new investigational drugs (21CFR 312.6).  The intervention will be taken orally or given with 
enteric feeding by nursing staff overseen by the study team, with the first dose administered within 
6 hours of ICU admission.  The time course for the intervention and remaining study assessments 
is shown below in Table 3. 
 
Table 3.  Time course of intervention and main assessments during the study. 
 
Inulin-------------> 
Monitoring--------------------> 
Type of 
Assessment 
Day 
0 
Day 
3 
Day 
7 
Day 
14 
Day 
30 
3  
months 
6 
months 
Rectal swab 
X 
X 
X 
X 
X 
 
 
Whole stool 
X 
X 
X 
X 
X 
 
 
Clinical data 
X 
X 
X 
X 
X 
X 
X 
Telephone follow-up 
 
 
 
 
 
X 
X 
 
 
9.1. Storage, accountability, and dispensing 
The study medication will be delivered to the CUMC Research Pharmacy as a powder via 
Swanson/Cargill protected from moisture and light and accessible only to authorized individuals.  
The medication will be stored at 20-25ºC in powder form and reconstituted before dispensation.  
Medication labels will be blinded.  The Research Pharmacy will maintain a record of the inventory 
and disposition of all medication and placebo.  This receipt record will include from whom the 
medication was received, to whom it was distributed, the date, quantity, and lot number.  The 
study medication has the following characteristics (Table 4). 
 
 
Figure 4.  Chemical structure of inulin.  Inulin 
is a naturally occurring polysaccharide derived 
from chicory root, agave, garlic, and other 
vegetable sources; it is a long chain 
carbohydrate of fructose polymers with β(1,2) 
glycosidic linkages which are indigestible by 
human enzymes but fermented by colonic 
anaerobic bacteria into short-chain fatty acids 
(chemical formula: C6nH10n+2O5n+1).  


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Table 4: Study medication characteristics. 
Product name: 
Inulin powder 
Manufacturer: 
Cargill Inc., 15407 McGinty Rd W, Wayzata MN 55391-2365 
Distributor: 
Swanson Health, 4075 40th Ave SW, Fargo, ND 58104-3912 
Appearance: 
White powder 
Flavor: 
Neutral to sweet 
Bulk density: 
0.6 g/mL 
Solubility: 
>100 g/L 
Moisture: 
3.41% 
Dry stability: 
4 years 
 
9.2. Rationale for the selection of duration and dose.   
 
9.2.1. Rationale for duration.  Inulin will be given for 7 days with the primary outcomes adjudicated 
after 3 days.  The Day 7 data will be used to test whether an effect can be extended or whether 
a delayed effect is present.  In our cohort data, 84/126 (67%) new infections occurred within 3 
days and 97/126 (77%) within 7 days.38  This shows that any ICU-based intervention seeking to 
prevent infections must work quickly if it is to be effective.  Meanwhile, our preliminary cohort data 
suggests that fiber can have an impact even within this short timeframe (Figure 3).  A final 
assessment will be made on the day of hospital discharge in subjects who are to be discharged 
prior to Day 14.   
 
9.2.2. Rationale for dose.  In the ambulatory setting, inulin impacts SCFA producer levels at doses 
of 12 to 16 g/day, with biologic effects relevant for ICU patients (e.g., decreases in serum LPS 
and CRP).26,66  Approximately 16 g/day therefore represents the minimum dose at which an effect 
might be seen; this was selected for the low dose arm of the study.  However, patients in these 
ambulatory studies were not critically ill and did not receive antibiotics.  It is possible that inulin 
will have lesser effects in the ICU compared to the ambulatory setting, and logical to test higher 
doses.  To select a maximum study dose, we referenced our own experience with dietary fiber in 
the prospective cohort and published tolerability data from ICU studies of inulin-like prebiotic fiber.  
In our prospective ICU cohort, 30 g/day was the maximum observed dose of dietary fiber, and 
was not associated with any intolerance.  In published literature, O’Keefe et al. reported tolerance 
up to a maximum of 36 g/day wheat dextran in the ICU.36  We therefore selected 32 g/day for the 
high dose study arm because it lies between these values and is exactly double a dose with 
proven biologic activity. 
 
9.3. Regulatory 
No subjects will be enrolled until after review and approval by the U.S. Army Medical Research 
and Materiel Command (USAMRMC) Office of Research Protections (ORP), Human Research 
Protection Office (HRPO).  The PI holds FDA IND 
 (FDA date: September 4, 2018) for 
inulin for the prevention of MDRO colonization/infection in the ICU or other at-risk settings.  Safety 
and adverse effects will be overseen by the PI in conjunction with Daniel Brodie, the Director of 
Medical Intensive Care Units at our institution, who will serve as the Data Safety Monitoring Officer 
(DSMO) and will convene an appropriate Data Safety Monitoring Board (DSMB) for the study.  
Before initiating research, the study will be registered on clinicaltrials.gov. 
 
9.4. Documentation of administration of the intervention 
The study team will work with the treating ICU team to ensure that the study drug is delivered in 
a timely manner.  The time and dose of each medication administration will be documented at 
each study assessment by referring to the documentation of medication administration within the 


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electronic medical record (EMR).  Patients will be enrolled with a goal of administering the 
intervention within 24 hours of ICU admission and it will be considered a protocol deviation if the 
intervention is administered more than 48 hours from ICU admission. 
 
10. Outcomes 
Primary and secondary outcomes focus on the comparison of baseline to Day 3 samples.  This 
project will also gather samples at ICU Days 7, 14, and 30.  These additional samples will be used 
to construct longitudinal time series data and test for differences between groups that may be 
slower to emerge (e.g., baseline vs Day 7 comparisons). 
 
10.1. Primary outcome: Relative abundance of SCFA producers through ICU Day 3.  The within-
individual change in the relative abundance of SCFA producers will be determined by 16S rRNA 
gene sequencing of rectal swabs and compared between intervention groups.  From extracted 
rectal swab DNA, polymerase chain reaction (PCR) will be performed on a Bio-Rad T100 Thermal 
Cycler (Bio-Rad, Hercules, CA) to amplify the V4 hypervariable region of the 16S ribosomal RNA 
gene with primers derived from Klindworth et al.67 appended with overhang sequences for 
compatibility with Illumina index and sequencing adapters (Illumina, San Diego, CA).  The relative 
abundance of SCFA producers levels will be determined as the sum total across the bacterial 
species known to ferment the SCFA butyrate and subtracted within each individual to obtain the 
change (Day 3 minus baseline).39,45,68-70   
 
10.2. Secondary outcome: MDRO colonization status on ICU Day 3.  MDRO colonization status 
will be based on Day 3 rectal swab cultures, classified categorically and compared between 
intervention groups.  MDROs will be defined as methicillin-resistant Staphylococcus aureus 
(MRSA), vancomycin-resistant Enterococcus (VRE), and MDR Gram negative bacteria71 (defined 
as Gram negatives with in vitro non-susceptibility for third-generation cephalosporins).72  Based 
on culture results, Day 3 MDRO colonization status will be classified categorically and compared 
between groups.   
 
10.3. Additional outcomes: 
10.3.1. Feasibility.  Descriptive demographic and clinical data will be compared across study 
groups to assess adequacy of randomization.  Summary statistics (count data) will be 
recorded for patients across all 3 intervention arms to assess feasibility as (1) the enrollment 
rate (enrolled)/(enrolled+refusals) over each 12 month period; (2) protocol adherence 
(proportion of randomized subjects who receive 1 or more doses of the allocated intervention); 
and (3) fidelity of the intervention (defined as the proportion of high/low inulin allocated vs 
received and also categorically as receipt of 90% or more allocated doses of inulin across 
groups).  Feasibility goals are as stated in Aim 1. 
 
10.3.2. Rates of pre-specified AEs.  There will be active ascertainment of AEs in the 
following categories, which will be compared between intervention groups.  This will be done 
through the end of the intervention period (ICU Day 7) and also through Day 30/discharge to 
assess for delayed AEs.  
 
 
Stool frequency, measured by subject or nursing report by the number of spontaneous 
stools over the 24 hours preceding each assessment; 
 
 
Stool consistency, measured by subject or nursing report as an ordinal variable (0-4) 
over the 24 hours preceding each assessment; 
 


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 
Abdominal bloating, measured by subject or nursing report as an ordinal variable (0-3) 
over the 24 hours preceding each assessment. 
 
 
Electrolyte abnormalities, measured by examining for differences in median values, 
minimum/maximum values, and proportions of values outside the normal range.  This 
will be done for glucose, calcium, magnesium, phosphate, potassium, and sodium. 
 
 
Nausea, vomiting, obstruction, and abdominal pain.  These will be recorded as present 
or absent based on bedside assessment of the subject or based on the nursing 
impression. 
 
 
Culture-proven infections, which will be considered both an AE and an outcome (see 
10.3.4). 
 
10.3.3. Antibiotic resistance gene fraction on ICU Day 3.  DNA will be extracted from rectal 
swabs (MoBio PowerFecal, Carlsbad, CA) and assessed using multiplex quantitative PCR 
(Qiagen Cat. No. 330261, Valencia, CA) for 87 common antibiotic resistance genes.73  Within-
individual differences in multiplex qPCR results will be compared from baseline to Day 3 both 
as summed changes and within relevant gene categories (e.g., β-lactamase resistance 
genes) and compared between intervention groups. 
 
10.3.4. Culture-proven MDR infections through ICU Day 30.  Antibiotic-resistant infections will 
be defined as those showing all of the following: (1) site-specific symptoms or signs of 
infection, (2) initiation of targeted antimicrobials by the treating ICU team, and (3) molecular 
diagnostics or cultures showing an MDRO as adapted from the CDC and NHSN74 and 
previously used by our group (Table 4).40  Rates of infections will be compared between 
intervention groups as a sum total, and separately for 3 distinct periods: the intervention period 
of Day 0 to 7, the Day 7 to 30 period, and the Day 30 to 6 month period. 
 
Table 4.  Criteria used to adjudicate culture-proven MDR infections for.  Criteria 
based on CDC/NHSN guidelines. 
Criteria 
Operationalization 
Example 
1. Site-specific 
symptoms or 
signs of infection. 
 Pulmonary: 
documentation 
of 
new 
or 
worsening cough or imaging consistent with 
an infection 
 Urinary: fever, dysuria, increased urinary 
frequency, flank pain, mental status changes, 
or hypotension and clinical exclusion of other 
causes for infection. 
 Bloodstream: 
clinical 
evidence 
of 
a 
bloodstream 
infection 
(fever, 
chills, 
or 
hypotension) without alternative sources for 
infection 
 Stool: documentation of new loose stools or 
frequent stools ≥3 per 24 hours 
 Wound: appropriate surgical site incision or 
wound with discharge, warmth, or erythema. 
Cough and a 
chest X-ray 
showing new 
infiltrate. 
2. Positive 
cultures or other 
diagnostics with 
an MDRO. 
 Pulmonary: quantitative or semi-quantitative 
sputum or bronchoscopic cultures 
 Urinary: ≥105 CFU/mL growth. 
 Bloodstream: 
growth 
of 
a 
recognized 
bloodstream pathogen from blood but not 
another site or growth of a NHSN commensal 
from blood but not another site. 
Sputum 
culture 
growing 
>105 CFU/ml 
of K. 
pneumoniae 
resistant to 
ceftriaxone. 


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 Stool: positive PCR for the C. difficile toxin B 
gene (Cepheid or BioFire) or positive toxin B 
EIA 
 Wound: growth of aseptically obtained 
organisms. 
3. Initiation of 
targeted 
antibiotics within 
24 hours. 
Antibiotics showing non-susceptibility using 
standard CLSI breakpoints or appropriate 
empiric antibiotics initiated at the time of clinical 
suspicion/testing and continued for ≥3 days or 
initiated within 24 hours following the availability 
of diagnostic testing showing a culture-proven 
infection. 
Initiation of 
ceftriaxone 
when the X-
ray was 
ordered; 
later 
changed to 
meropenem. 
 
10.3.5. Fecal SCFA levels.  The SCFAs butyrate, acetate, and propionate will be measured 
from whole stools on a 6490 triple quadrupole mass spectrometer and compared between 
intervention groups using the first stool sample produced after the Day 3 assessment.  
Patients who fail to produce a stool from Day 3 to 7 will be excluded from this analysis. 
 
10.3.6. Proportion of goal calories through ICU Days 3 and 7.  Proportion of goal calories 
consumed will be defined by manually extracting data from electronic nursing flow sheets. 
Intake during the intervention be calculated by multiplying enteral or oral feeding caloric 
content by the flow rate or percentage of meal consumption.  A patient-specific calorie target 
will be calculated by a registered clinical dietician using the Mifflin-St. Jeor equation and the 
Penn State 2003b and 2010 equations, taking into consideration age, body mass index, 
ventilation status, and comorbidities.75  The proportion of goal calories consumed will then be 
calculated as observed nutritional intake divided by calorie target.  Day 3 and Day 7 
proportions will be compared between intervention groups.  
 
10.3.7. Length of ICU stay.  Length of ICU stay through Day 30 will be compared between 
treatment groups, with Fine and Grey methods used to adjust for death as a competing risk.  
Length of ICU stay will be measured from the first to the last set of recorded ICU vital signs. 
 
10.3.8. Length of hospital stay.  Similarly, length of hospital stay through Day 30 will be 
compared between treatment groups, again adjusting for death as a competing risk.  Length 
of hospital stay will be measured from the first set of recorded ICU vital signs to the last set of 
hospital recorded vital signs. 
 
10.3.9. Mortality through ICU Day 90.  Death data will be extracted from the hospital EMR 
which immediately captures in-hospital death and receives monthly mortality uodates from the 
National social security death index.  Kaplan-Meier methods will be used to compared 30- and 
90-day mortality between treatment groups. 
 
10.3.10. Self-assessed quality of life at 6 months.  Subjects or their caregivers will complete 
the EQ-5D-5L76-78 and Katz ADL questionnaires79,80 to test their quality of life at 3- and 6 
months post ICU admission.  Composite data will be compared between intervention groups. 
 
10.3.11. Time series data including Day 7 outcomes.  Time series data will be built and 
visualized for all relevant outcomes.  We will explore the possibility of a delayed effect by 
testing for differences between groups from baseline to Day 7.  If there is an observed effect, 
the Day 14 and Day 30 data will allow us to assess its durability.  
 


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11. Study population 
 
11.1. Study setting 
Patients will be enrolled from medical ICUs affiliated with Columbia University Medical Center.  
These ICUs include but are not limited to the following locations, all of which are in the general 
vicinity of Columbia University’s medical campus:  
ICU short 
designation 
Full name 
Hospital building, 
floor location  
Street address 
MICU-A 
Medical ICU-A 
Milstein, 4th floor 
177 Fort Washington Ave, NY, NY 
MICU-B 
Medical ICU-B 
Milstein, 4th floor 
177 Fort Washington Ave, NY, NY 
NICU 
Neurological 
ICU 
Milstein, 8th floor 
177 Fort Washington Ave, NY, NY 
AICU 
Allen ICU 
Allen Pavilion, 2nd floor 41 Broadway, NY, NY 
 
The study will enroll 90 consecutive adult ICU patients who meet criteria for sepsis, are receiving 
appropriate antibiotics, and can complete a bedside assessment within 4 hours of ICU admission 
and receive the trial intervention within 6 hours of ICU admission.  A replacement strategy will be 
used.  If subjects are randomized but fail to receive the intervention or complete the Day 3 
assessment, an additional subject will be enrolled to take their place with a final target enrollment 
of 30:30:30 patients who complete a minimum of 3 days of assessments.  Specific inclusion and 
exclusion criteria are as follows: 
 
11.2. Inclusion criteria 
 
 Medical ICU.  Subjects will be newly admitted to one of two CUMC medical ICUs during the 
study enrollment period; 
 
 Age.  Subjects will be eighteen or more years old at the time of ICU admission; 
 
 Sepsis.  Subjects will meet criteria for sepsis with life-threatening organ dysfunction caused 
by infection (Sepsis-3 third international consensus definition, 2016).81  According to this 
definition, life-threatening organ dysfunction is operationalized as an increase in the 
Sequential [Sepsis-Related] Organ Failure Assessment (SOFA)82 score of ≥2 points above 
baseline.  Sepsis-3 criteria also require patients to have a known or suspected infection, 
defined as sampling of blood or any other body fluid for infection with concurrent 
administration of appropriate antibiotics (see Table 4);81    
 
 Appropriate antibiotics.  Subjects will receive appropriate antibiotics, which will be 
operationalized as (1) among subjects with positive culture data, the antibiotics within classes 
where the organism is susceptible based on standard Clinical and Laboratory Standards 
Institute (CLSI) cut-offs72; or (2) among subjects receiving empiric antibiotics (i.e., with 
negative cultures), antibiotics within classes with significant anaerobic activity.  This will 
include β-lactam/β-lactamase inhibitor combination antibiotics, cephalosporins (generation 2 
or greater), fluoroquinolones, lincosamides (clindamycin), metronidazole, and monobactams 
(a.g., meropenem).  Vancomycin monotherapy is intentionally excluded.  Antibiotics must be 
received within 24 hours before ICU admission or be actively ordered at the time of bedside 
screening;    
 


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 Prompt intervention.  Subjects will complete study enrollment within 4 hours of ICU admission 
so that the intervention can be administered within 6 hours of ICU admission.  The first set of 
recorded ICU vital signs will define the time of ICU admission.   
 
11.3. Exclusion criteria  
 Inability to receive oral or enteric fluids.  Patients will be excluded if they are nil per os or are 
unable to take fluid by mouth or delivered via enteric nasal or oral tube because they would 
be unable to receive the study intervention; 
 
 Inulin allergy.  Patients will be excluded for safety reasons if they have a known chicory allergy.  
Serious chicory allergies are exceedingly rare although mild respiratory symptoms and IgE-
mediated food allergies have been reported;83 
 
 Hyponatremia.  Patients will be excluded for hyponatremia with serum sodium ≤128 mEq/L 
because the intervention will be delivered in 500 ml of free water daily.  If subjects develop 
hyponatremia (serum sodium serum sodium 125-128 mEq/L) after enrollment and 
randomization which persists at the time the subject is due to receive the study intervention, 
the study team will confer with the treating ICU team regarding whether the intervention should 
be deferred or further concentrated in order to minimize free water.  If subjects develop severe 
hyponatremia (serum sodium serum sodium ≤124 mEq/L) after enrollment and randomization, 
the intervention will be withheld until 2 consecutive serum sodium values are within normal 
limits; 
 
 Immunosuppression.  Patients will be excluded if they have immunosuppression, as CD4 
count <200 or absolute neutrophil count <500 or reasonable expectation of either during the 
study intervention period.  We believe that the inulin works by modifying the gut bacteria.  
Excluding patients who lack the ability to mount an immune response improves trial 
homogeneity; 
 
 Surgery involving the intestinal lumen.  Patients will be excluded for surgeries involving the 
intestinal lumen within 30 days.  The most common side effects from inulin are subjective 
bloating and abdominal distension.26,28,30,52  Although this bloating appears to be mild and self-
limited, patients with recent bowel surgeries may be at greatest risk for such symptoms and 
will therefore be excluded; 
 
 DNR or DNI status.  Such patients have limited treatment goals and will be excluded to 
minimize heterogeneity; 
 
 No LAR.  Patients will be excluded if they lack capacity for consent and have no appropriate 
Legally Authorized Representative (LAR) to act as surrogate decision-maker. 
  
12. Enrollment strategy 
Patients will be identified at the time of ICU bed allocation and screened for inclusion/exclusion 
criteria.  After consultation with the treating ICU team, subjects who meet screening criteria will 
have the eligibility evaluation completed at the ICU bedside.  This will be done after first obtaining 
agreement from the medical ICU attending of record that the patient/surrogate can be approached 
for the study.  We anticipate that >50% of patients will lack decision-making capacity and will 
accept consent from a surrogate in these situations, as previously described by our group.37  
Issues related to altered capacity, privacy and time for decision-making, consent of LARs, and 
ongoing consent are discussed in detail in the Human Subjects and Recruitment and Safety 


Inulin in the ICU 
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Procedures section.  We expect that patients will usually be initially enrolled with surrogate 
consent.  Patients who are on study will then be reassessed for capacity at each interaction, with 
the final determination of capacity made by the treating ICU team.  When subjects regain capacity, 
they will make their own determination of whether or not to continue in the study.  If subjects do 
not regain capacity by the time of hospital discharge, surrogate consent will be accepted as final.  
This approach, which we have used before,37-39 preserves complete patient autonomy while 
recognizing that the target population for the study is unlikely to have decision-making capacity 
at the time of ICU admission. 
 
Compliance statement regarding consent 
This research will meet the requirements of 10 U.S. Code 980 for experimental human subjects 
research that “(1) the informed consent of the subject is obtained in advance; or (2) in the case of 
research intended to be beneficial to the subject, the informed consent may be obtained from a 
legal representative of the subject.” 
 
Retention 
Potential loss of patients after randomization will be from death/discharge prior to ICU Day 3, or 
withdrawal of consent (e.g., after initial surrogate consent).  In our ongoing ICU cohort studies, 
there has been 11.3% mortality within 3 days and a 0% Day 3 discharge rate.  Additional patients 
will be enrolled for patients who do not complete the Day 3 assessment due to death or discharge.  
Thus, we expect to enroll 102 patients to obtain 90 who are able to donate a Day 3 sample and 
therefore can included in the intent-to-treat analysis. 
 
12.1. Randomization and blinding 
Randomization and data entry will take place through a secure, web-based system managed by 
the CUMC Data Coordinating Center (DCC) headed by Howard Andrews.  This password-
protected system is based on the Research Electronic Data Capture (REDCap) platform.  The 
Research Nurse will use paper clinical research forms (CRFs) to document the initial screening 
process.  When this information is entered into REDCap, the system will automatically generate 
a subject identifier and randomization code using 1:1:1 randomization in randomly permuted 
blocks of 6.  This randomization code will be electronically communicated to the research 
pharmacy along with a separate key which will allow them to identify the random assignment.  
The Columbia Research Pharmacy will prepare the study intervention in indistinguishable semi-
opaque fluid bags.  Although inulin is readily soluble at the study doses, the opacity of these bags 
will eliminate any possibility that a small amount of precipitant might reveal the study assignment 
to the ICU nurses who deliver the intervention.  Five drops of non-caloric flavoring syrup 
(McKesson Supply, Yonkers, NY, Product #570727) will be placed into the reconstituted 
inulin/placebo to prevent any possibility of accidental unblinding based on taste in patients who 
are alert.  The patient, treating ICU team including the ICU nurse, and study team will remain 
blinded to each assignment until the completion of the trial, with the DCC preparing unblinded 
safety reports for the DSMO at pre-specified intervals.   
 
12.1.1. Clinical need for unblinding 
It seems unlikely that clinical need for unblinding will arise.  If this occurs, the PI will coordinate 
with the DCC to facilitate unblinding for the treating ICU team.  In any case of unblinding, data 
collection and follow-up will be maintained and the patient analyzed per-protocol.    
 
12.2. Delivery of the intervention 
The treating medical team will order the study intervention as a non-formulary medication to be 
delivered by ICU nurses, as they would any other medication, and the study team will work to 
ensure prompt delivery of the intervention with a goal of having it delivered within 24 hours from 


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the first set of ICU vital signs.  Delivery of the intervention will be managed by ICU nurses who 
are not part of the study team by encouraging the subject to drink the liquid formulation or, in 
subjects who are not taking oral medications, by attaching the intervention to the patient’s oral or 
nasogastric tube.  The time of dosing will be recorded electronically within the patient EMR and 
documented by study staff on the CRFs and within REDCap.   
 
12.3. Premature cessation of the intervention 
Efforts will be made to ensure that all subjects who are randomized receive the study intervention.  
The initial medication order will specify that the study intervention is to be given q12h for a total 
of 14 doses.  The study intervention will be stopped under the following conditions: 
 
 
Death; 
 
 
Discharge from the hospital; 
 
 
Request to withdraw from the study by the patient or surrogate.  This request can be made 
at any time, without providing a reason.  Consent to continue data collection and retain 
samples already collected will be sought in these situations; 
 
 
Definite clinical indication or contraindication becomes apparent.  If the treating ICU team 
feels at any time that the patient needs or should not have fiber supplementation, the 
intervention will be immediately halted.  The patients will remain in the study for follow-up 
and the intervention will be held as long as the indication/contraindication remains.  In 
general, if the indication/contraindication resolves within the 7 day timeframe of the 
intervention, the intervention will be resumed.  If the intervention is withheld for 
hyponatremia, the intervention will be resumed only once 2 consecutive serum sodium 
values are within normal limits; 
 
 
Adverse or serious adverse event related to the study intervention.  Such events will 
constitute unanticipated problems.  The patient will remain in the study for follow-up, but 
no further interventions will be administered. 
 
13. Study assessments 
The total study duration will be no more than 30 days of bedside assessments and no more than 
6 months of telephone follow-up, both measured from the first recorded set of ICU vital signs. 
Samples will be gathered and bedside assessments made at the following intervals after ICU 
admission or until hospital discharge/death: baseline, 3 days, 7 days, 14 days, and 30 days.  
During study assessments, data will be combined from the following sources: (1) samples 
collected by the study team, (2) bedside assessments, and (3) data from the EMR. 
 
13.1 Sample collection and storage 
Sample collection is focused around rectal swabs because the timing of swabs (as opposed to 
whole stools) can be precisely dictated and because rectal colonization with MDR bacteria best 
reflects overall MDR colonization status84-88 and risk for subsequent infection.89  Samples are 
described below. 
 
 Rectal swabs.  At each study assessment, deep flocked nylon swabs of the rectum will be 
performed in duplicate to allow independent processing of swabs for 16S rRNA gene 
sequencing/multiplex qPCR and for bacterial culture of MDROs.  Swabs will be inserted 5 cm 
rectally with patients in the left lateral decubitus position and rotated 5 times, with adequate 


Inulin in the ICU 
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sampling verified by looking for fecal staining.  Swabs to be sequenced will be flash-frozen at 
-80ºC for long-term cold storage90 and batched extraction;91 swabs to be cultured will be 
transported on soy broth with 20% glycerol (Becton Dickinson, Franklin Lakes, NJ) for culture 
or, when culture cannot be performed immediately, for storage overnight at 4º C and culture 
the following morning.92 
 
 Whole stools.  Nursing staff will collect the first spontaneous whole stool subsequent to each 
study assessment.  4 x 8 ml aliquots of stool will be flash-frozen and stored at -80 º C. 
 
 Oral swabs.  Two cotton-tipped oral swabs will be used to gather a composite of the oral flora 
by brushing the following consecutive locations x 10 each: left cheek, right cheek, hard palate, 
tongue, floor of the mouth, and maxillary gingivae (same as the Human Microbiome Project).93  
Swabs will be flash frozen at -80 º C. 
 
 Urine.  ICU nurses will withdraw 2 x 8 ml of urine using standard technique from the urinary 
catheter sampling port.  In patients who lack a urinary catheter, clean catch urine samples will 
be accepted and aliquoted as above. 
 
 Blood.  Residual blood from clinical samples drawn immediately prior to each study 
assessment will be retrieved by the study team.  For each subject, one 6 mL serum-separator 
and one 8 mL EDTA tube will be retrieved.  Buffy coat will be separated by centrifuge and 
with 1-2 mL aliquots generated for plasma/buffy coat and stored at -80 º C.  This strategy 
spares subjects the need for an additional blood draw.   
 
Oral swabs, urine, and blood will be stored for future testing.  The rationale for collecting these 
additional samples is that they are non-invasive, can be easily collected, and will provide 
opportunities for future studies seeking to explore additional markers or mechanisms (e.g., urinary 
3-indoxyl sulfate for gut microbiome disruption94 or micro-RNAs in sepsis).95  All sample storage 
will be in the PI’s -80ºC freezer unit (P&S-8) except for the fresh rectal swab used for culture 
which will be stored within the Uhlemann lab (P&S-9).  Assays performed from rectal swabs and 
whole stools (SCFAs) are described below. 
 
13.2. Sample assays 
The assays described below will be performed from rectal swabs or whole stools (for fecal SCFA 
levels).  Residual sample material will be retained in -80ºC storage for the duration of the study.  
 
 
16S rRNA gene sequencing.  16S rRNA gene sequencing will be performed to determine 
the primary outcome.  From extracted rectal swab DNA, polymerase chain reaction 
(PCR) will be performed on a Bio-Rad T100 Thermal Cycler (Bio-Rad, Hercules, CA) to 
amplify the V4 hypervariable region of the 16S ribosomal RNA gene with primers derived 
from Klindworth et al.67 appended with overhang sequences for compatibility with Illumina 
index and sequencing adapters (Illumina, San Diego, CA).  Samples will be purified with 
Agencourt AMPure XP beads (Beckman Coulter, Jersey City, NY) and quantified using 
the Quant-iT broad range dsDNA Assay Kit (Thermo Fisher Scientific, Fair Lawn, NJ). 
Libraries will be normalized, pooled, and denatured in preparation for cluster generation 
and sequencing, which will be performed on the Illumina MiSeq 300PE.  
 
 
Bacterial culture.  Aerobic culture will be performed to determine the secondary outcome.  
Culture has been selected for the primary outcome because culture identifies viable 
bacteria, remains the most common clinical method for testing for MDRO colonization in 


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the ICU, and best predicts longterm risk for organism-specific infection.96-99  Additional 
analyses will utilize alternative methods for classifying MDROs.  Cultures will identify 
MRSA, VRE, and MDR Gram negatives, which cause 94% of all MDR bacterial infections 
in the ICU.6  Rectal swabs will be gently vortexed to dissociate DNA from the swabs into 
the broth media.  Media will then be split into equal aliquots and plated under a laminar 
flow hood for MRSA (Spectra-MRSA, Remel), VRE (Spectra-VRE, Remel), and ESBL or 
Klebsiella pneumonia carbapenemase (KPC)-producing bacteria (Uhlemann lab).  In 
addition, we will plate cultures on Gram-negative selective agar (MacConkey II agar, 
Becton Dickinson) directly and at three serial dilutions for semi-quantitation (10-2, 10-4, and 
10-6).  While the main analysis will be based on a categorical classification of MDROs 
(present vs absent), an exploratory analysis will be conducted using semi-quantitative 
culture data.  Preliminary data show that these dilutions will yield a countable number of 
colonies.  After 18-24 hours of aerobic growth, selective plates will be classified as positive 
vs negative for growth according to the media characteristics.  The colony-forming units 
(CFUs) on the non-selective plates for Gram positives/negatives will be counted to provide 
relative quantification of each MDRO based on the total CFU counts from the non-
selective plates.  If multiple non-selective plates show growth, the plate with 10-100 
colonies will be used for CFU counting with results normalized to the 10-2 plate.  From 
these plates, organisms will be identified using the VITEK 2 system (bioMerieux) and AST-
N010/020 cards with confirmatory testing as needed.  Resistant isolates will be stored for 
whole genome sequencing analyses. 
 
 
Quantitative PCR.  Quantitative PCR for resistance genes will be performed as an 
additional outcome.  DNA will be extracted from rectal swabs (MoBio PowerFecal, 
Carlsbad, CA) and assessed using multiplex quantitative PCR (Qiagen Cat. No. 330261, 
Valencia, CA) for 87 common antibiotic resistance genes.73  In this assay, 250 ng of 
template genomic DNA will be added to each reaction on a 96-well plate which will be run 
on a StepOnePLus RT-PCR machine using the following cycling conditions: initial 
denaturation for 10 min at 95º C, cycling 45 x 15 sec at 95º C denaturing followed by 2 
min at 60º C annealing and extension.  Controls will detect the presence of bacterial DNA, 
PCR inhibitors and background (no template control (NTC)).  PCR cycle thresholds (CT) 
of <35 will be considered positive for the presence of any given gene.  Adequate reactions 
will be determined by CT values of  <29 for pan-bacterial reference genes (e.g., 16S rRNA, 
gyrA, recA, rpoB)100 and positive PCR controls (PPC) CT values of  <24.  Linearity and 
sensitivity for this PCR have been determined using synthetic templates over a 6-log serial 
dilution ranging from 1 to 1 million copies, with results verified across multiple antibiotic 
resistance genes using pyrosequencing.73 
 
 
Fecal SCFA levels.  Fecal SCFAs levels will be assayed as an additional outcome.  Whole 
stools will be aliquoted and diluted with 80% ethanol to be agitated at 4°C.  After 
centrifugation, 20 µl of supernatant from the homogenized samples will be mixed with 20 
µl of 200 mM N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (1-EDC HCl) 
in 5% pyridine and 40 uL of 100 mM 2-Nitrophenylhydrazine (2-NPH) in 80% acetonitrile 
(ACN) with 50 mM HCl.  After a brief incubation, ACN will be added to the solution which 
will be injected onto a 6490 triple quadrupole mass spectrometer (Agilent, Santa Clara, 
CA) and tested for concentrations of the SCFAs butyrate, acetate, and propionate. 
 
13.3. Bedside assessments including data gathered from the EMR   
During each study assessment, a study-specific Case Report Form (CRF) will be used to record 
the following data: origin for admission (classified as from the hospital floor, operating room, 
emergency room, or outside ICU transfer based on the most recent location prior to CUMC ICU 


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admission); the Glasgow Coma Scale (GCS);101 the use of life support (classified as present vs 
absent) including the use of mechanical ventilation, hemodialysis, or additional life-support 
devices such as extra-corporeal membrane oxygen (ECMO) or an intra-atrial balloon pump 
(IABP); and the presence or absence of intravenous lines or catheters including a central venous 
catheter, urinary catheter, arterial catheter, nasal or oral gastric tube, or rectal tube.  Using 
duplicate data entry, anonymized CRF data will be transferred into the secure REDCap database.  
Additional clinical variables will be entered directly into REDCap from the EMR.  These variables 
will include laboratory analyses, all of the component variables of the Sequential Organ Failure 
Assessment (SOFA) score,82 measures related to nutritional intake, and ICU interventions 
including administration of vasopressors, proton pump inhibitors, and antibiotics (with the name 
of the antibiotic and route of administration recorded). 
 
13.3.1. Tolerability 
Patients who are alert will be asked regarding abdominal bloating which will be graded on a 4-
point Likert scale (0=none, 1=a little, 2=a moderate amount, 3=a lot/severe).  When patients are 
not alert, this will be graded as “unable.”  ICU nurses will be asked to grade the average 
consistency of stools within each preceding 24 hours on a 5-point scale (0=like water, 1=like 
porridge, 2=loose but with chunks, 3=formed, soft, 4=formed, hard).  This information will also be 
recorded from the EMR into REDCap as a separate entry.  When subjects have no stools within 
the preceding 24 hours, this will be recorded as “no stools.”  The frequency of stools within the 
preceding 24 hours will be recorded as an integer at each study assessment.  Again, this will be 
done based both on the nurse’s in-person report and separately based on EMR documentation. 
 
ICU nurses will be additionally asked regarding the presence or absence of the following 
symptoms during the 24 hour period prior to each study assessment: nausea, vomiting, bowel 
obstruction, and abdominal pain.  This data will be recorded categorically on the CRFs/REDCap 
and will appear on DSMO reports. 
 
13.3.2. Nutritional intake 
Information related to nutritional intake will be gathered from the EMR including goal calories, 
actual calories consumed, and the amount of dietary fiber consumed in grams.  All ICU patients 
at our institution receive a complete clinical nutritional assessment with calculation of goal calories 
based on the Penn State Equation102 with the modified equation used for individuals >60 years 
old or those with BMI >30 as described by Frankenfeld et al. in 2009.103,104  Actual calories and 
grams of fiber consumed will be based on the specific components of the tube feeds received or, 
among patients on an ad lib diet, the food consumed.  The nutritional components of all CUMC 
diets are completely specified in terms of calories, fat, protein, carbohydrate, and fiber content.  
To calculate calories and fiber consumed, the study team will refer to the proportion of each meal 
consumed as recorded by ICU nurses and to the volume of tube feeds delivered which is captured 
hourly in the EMR.  In those receiving tube feedings, the interval number of tube feeding 
interruptions will be recorded at each assessment or recorded as 0 in those receiving ad lib diets. 
 
13.3.3. Long-term telephone follow-up 
Telephone interviews will be conducted with subjects or their surrogates 3 months after ICU 
admission and again 6 months after ICU admission.  In the event that subjects are still 
hospitalized, these interviews will be conducted at the bedside.  Vital status will be ascertained 
and, to minimize loss of follow-up information, we will interview subjects or surrogates to 
determine (1) discharge location type (classified as home, short-term rehabilitation, or long-term 
rehabilitation including skilled care facilities), (2) hospital admission to non-CUMC facilities after 
ICU discharge, (3) physician office visits after ICU discharge, and (4) infections or other adverse 
events after ICU discharge.  If there are non-CUMC hospital admissions or other healthcare 


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interactions, permission will be requested to obtain relevant records to determine the nature of 
infections and to assess for potential medium- to long-term AEs related to the study intervention.  
Among surviving subjects, the EQ-5D-5L (EuroQol 2009) will be used to determine health status 
across the domains of mobility, self-care, activity levels, pain, and anxiety/depression.76,77  Use of 
this highly validated instrument will facilitate standardization and generalizability of results and will 
provide preliminary data for post-ICU discharge monitoring for later phase trials.  This instrument 
can be applied via telephone in 5 minutes and was recommended in 2017 for post-ICU 
assessment of overall well-being after a three-round modified Delphi process involving clinical 
researchers from over 16 countries, patients/caregivers, clinicians, and research funders.78  EQ-
5D-5L responses have the potential to vary depending on whether answers are provided on 
patients versus surrogates,105 and therefore we will additionally assess post-ICU disability using 
Katz’ Index of Independence in ADLs (Activities of Daily Living).79  Although focused on functional 
status rather than well-being, Katz’ ADLs has been validated as an important patient-centered 
outcome across dozens of populations and unlike the EQ-5D-5L has high agreement between 
patients and caregivers.80 
 
14. Data collection and management 
The Columbia DCC will oversee use of a custom-designed, REDCap-based clinical trial data 
management system.  REDCap is a widely used web-based relational data system developed at 
Vanderbilt University with NIH support to collect research information in 21CRFPart11, FISMA 
and HIPAA-compliant environments.  REDCap functionality that will be used in this trial include 
randomization, subject scheduling, ad hoc reporting, data verification and audit trails.  Paper 
CRFs will be used to collect bedside data, while readily available selected electronic data (e.g., 
laboratory values) will either be directly “pulled” from Columbia clinical servers and merged into 
the REDCap database or manually entered into REDCap.  REDCap ‘Events’ will be specified 
corresponding to the study assessment timepoints: baseline, Days 3, 7, 14, 30, 3 months, and 6 
months.  Data of each type will be associated in the system with the appropriate time period. 
REDCap “Anytime” functionality will be used to capture adverse events and other incidents that 
can occur at any time, i.e. events that are not associated with fixed time periods.  The REDCap 
system has built-in data verification aspects that will be utilized for the trial including minimum and 
maximum ranges and other logical checks, and automatic checks for completion of individual 
items and forms. 
 
14.1. Data security within REDCap 
The servers on which DCC REDCap project data are stored are located in a physically and 
electronically secure environment maintained by institutional IT staff on the CUMC campus.  The 
DCC confirms project data access privileges on a monthly basis, and REDCap requires strong 
passwords, which must be changed periodically.  In this trial, to ensure data security, explicit 
identifiers needed for recruitment and scheduling (names, addresses, phone numbers, email 
addresses, medical record numbers) will be maintained separately and will not be stored in the 
REDCap database. 
 
14.2. Record retention 
All electronic and paper records will be retained for a minimum of 6 years from the trial end date 
in accordance with New York State law and meeting or exceeding Federal minimums.  Consent 
forms, CRFs, and other relevant paper documents will be moved to secure longterm paper 
storage once the trial has been closed and the data locked.  Record retention will be prolonged 
beyond 6 years as necessary if required by the funding agency. 
 
15. Safety and adverse events (AEs) 


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Adverse health events, laboratory irregularities, and even death are expected in ICU patients with 
sepsis and it would be difficult or impossible to accurately document and label all adverse health 
events.  This is a common problem faced by ICU trials.  The solution recommended by academic 
critical care research groups (e.g., Cook et al.) is to pre-specify AEs of interest for careful 
monitoring.  In keeping with this approach, this study will produce detailed reports of these AEs 
for the DSMC at standard intervals while only unanticipated problems that also meet criteria for 
serious adverse events (SAEs) will be reported to the DSMC in real time.106  AEs will be reported 
for all individuals who have received one or more doses of the intervention.  This study has pre-
specified AEs in the following categories. 
 
 
Bloating: graded 0-3 based on patient report: patient mean and maximum values will be 
reported; 
 
 
Stool consistency and frequency: graded 0-4 for consistency and with an integer denoting 
frequency over 24 hours based on nursing report.  For each patient, mean and maximum 
values will be reported; 
 
 
Electrolyte imbalances: For each patient, the minimum and maximum values for serum 
calcium, magnesium, phosphate, potassium, and sodium will be reported.  This will be 
done for the 7 days of the intervention period and separately from Day 7 through Day 
30/discharge; 
 
 
Gastrointestinal AEs: the gastrointestinal AEs of nausea, vomiting, bowel obstruction, and 
abdominal pain will be recorded as present or absent for each assessment and separated 
from other AEs to improve early pattern recognition by the DSMO;   
 
 
Infections: Infections are both an AE and an outcome and will be adjudicated as previously 
described.  For each infection, the date of the infection, suspected site, organism, and 
antibiotics received will be recorded.  Infections will be further classified as MDR vs non-
MDR using the criteria previously described; 
 
 
Death: death will be gathered from the hospital EMR, which inerfaces with the social 
security death index. 
 
15.1. Serious adverse events 
Federal Regulation 62 Oct. 7, 1997 defines SAEs as those events occurring at any dose which: 
 
 
Result in death; 
 
 
Are life-threatening, meaning the patient was at risk for death at the time of the event; 
 
 
Require hospitalization or prolong hospitalization; 
 
 
Results in significant disability or incapacity; 
 
 
Is a congenital abnormality/birth defect; 
 
 
Or does not meet these criteria but is thought by the investigator to be unusual and 
potentially serious. 
 


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SAEs in this trial will be AEs meeting the above Fed. Reg. 62 definition. 
 
15.2. Adjudicating and reporting adverse events as unanticipated problems 
Unanticipated problems (UPs) will be any unforeseen medical event thought likely related to the 
study intervention.  Such events will be monitored for in real time.  UPs that do not meet criteria 
for seriousness (same as SAE criteria) will be reported to the DSMO and IRB at pre-specified 
intervals.  UPs that meet criteria for seriousness will be reported within 24 hours from our 
notification of the event.  This reporting will include: 
 
 
Contact of the DSMO; 
 
 
Contact of the Columbia IRB; 
 
 
Fax or email scan containing when the event occurred, the name of the reporter, callback 
information, the protocol number, and a description of the event; 
 
 
Availability of the PI to provide follow-up information regarding the event. 
 
The proposed research is pending review by the Columbia University IRB (IRB #AAAS2576) and 
will be reviewed by the U.S. Army Medical Research and Materiel Command (USAMRMC) Office 
of Research Protections (ORP), Human Research Protection Office (HRPO) prior to 
implementation.  Prior to initiation of research, the study will be registered on clinicaltrials.gov.    
 
Safety and adverse effects will be overseen by the PI in conjunction with the Data and Safety 
Monitoring Officer (DSMO).  Daniel Brodie, the Director of the Medical Intensive Care Units at 
our institution will serve as the trial DSMO and will convene an appropriate Data and Safety 
Monitoring Committee (DSMC) to oversee interim study results.  The primary responsibility of the 
DSMC will be to review interim data in terms of safety and make recommendations whether the 
study should be changed, halted, or terminated.  Full details of the DSMC procedures and 
processes as well as additional details regarding AE and SAE reporting are in the DSMP. 
 
15.2.1. Compliance statement regarding research monitoring 
The DSMB under Dr. Brodie will function as a Research Monitor and is responsible to oversee 
the safety of the research and report observations/findings to the IRB or a designated institutional 
official. The Research Monitor will review all unanticipated problems involving risks to subjects or 
others associated with the protocol and provide an independent report of the event to the IRB. 
The Research Monitor may discuss the research protocol with the investigators; shall have 
authority to stop a research protocol in progress, remove individual human subjects from a 
research protocol, and take whatever steps are necessary to protect the safety and well-being of 
human subjects until the IRB can assess the monitor’s report; and shall have the responsibility to 
promptly report their observations and findings to the IRB or other designated official and the 
HRPO. 
 
15.3. Unblinding for adverse events 
All possible efforts will be made to retain blinding for the duration of the trial.  However, if a treating 
clinician feels that blinding must be removed in order to assess a possible adverse reaction quickly 
and correctly, the PI will facilitate this process by working through the DCC. 
 
16. Study termination and completion 


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The study may be terminated at any time for safety concerns by the PI, DSMC, IRB, or other 
appropriate regulatory bodies.  Otherwise the study will continue until 90 patients are enrolled 
who complete the Day 3 assessment and will be considered complete when all study 
assessments and analyses have been concluded. 
 
17. Statistical methods 
 
17.1. Overall plan for statistical analysis 
After enrollment is complete, the REDCap dataset will undergo DCC cross-checks for quality and 
accuracy.  The PI will review blinded data to identify errors which, if they exist, will be rectified by 
reference to paper CRFs or to the EMR.  After quality checks are complete, the REDCap dataset 
will be closed and locked.  The biostatistician will then perform an intent-to-treat analysis to 
determine the primary and secondary outcomes.  The DSMO will have complete access both to 
the raw data and to the analytic code used by the biostatistician and will nominate a qualified 
monitor who will review this work for rigor.  Once this is complete, the primary and secondary 
outcomes will be finalized.  At this point, the locked data will be made available to the PI and other 
investigators for additional analyses.  For all analyses, alpha <0.05 will be considered statistically 
significant and two-sided testing will be performed.  When death is a competing risk, Fine and 
Gray regression analyses will be performed. 
 
17.2. Determination of the primary and secondary outcomes 
 
Primary outcome: within-individual change in relative abundance of SCFA producers.  16S rRNA 
gene sequencing data will be processed using the QIIME platform with appropriate rarification 
and filtering.107  The primary analysis will be intent-to-treat, restricted to the subjects who complete 
the Day 3 assessment and contribute data.  Within-individual change in SCFA producers will be 
calculated as Day 3 minus baseline relative abundance and an ANOVA or Kruskal-Wallis (KW) 
test (if skewed data) will be used to assess for differences across the three groups.  If there is no 
significant difference between the low- and high-dose inulin arms, these arms will be combined 
for comparison against placebo in the final analysis using a two-sample t-test or rank-sum test.  
Although the primary analysis will be with the Day 3 samples, unadjusted and adjusted 
longitudinal data analysis will be performed using the generalized estimating equation approach 
with working independence correlation structure. 
 
Secondary outcome: MDRO colonization.  Subjects who culture positive from rectal swabs from 
Day 3 for MRSA, VRE, or a Gram negative with non-susceptibility to a third generation 
cephalosporin will be classified as MDRO colonized and all other subjects as not MDRO 
colonized.  The proportion of patients who are MDRO colonized on Day 3 will be compared 
between intervention groups using a chi-squared test or Fisher’s test.  For this analysis, patients 
in the inulin arms will be combined if testing shows no significant difference between these inulin 
arms.  Because baseline colonization status is likely to be an important predictor of Day 3 
colonization status, we will test whether baseline MDRO colonization rates differ between 
intervention groups.  If such a difference is observed, we will reanalyze Day 3 MDRO colonization 
with stratification according to baseline MDRO colonization (baseline MDRO colonized vs not 
colonized).  Exploratory analyses will examine Day 7 MDRO colonization status and semi-
quantitative culture data using ANOVA or Kruskal-Wallis testing.  The difference in rates of Day 
3 MDRO colonization will be taken as the effect size estimate for future trials.   
 
17.3. Power calculation and sample size 
 


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Primary outcome: within-individual change in relative abundance of SCFA producers.  The study 
is powered to detect a difference in SCFA producer levels that is equal to or greater than the 
observed difference in SCFA producer levels from our prospective cohort (6.6% for high vs 3.7% 
low dietary fiber).  The rationale for this is that inulin must be at least as impactful as mixed dietary 
fiber to be a viable therapeutic.  The proposed sample size of 30:30:30 will yield 80% power to 
detect a minimum difference of 0.73 standard deviations (SD) comparing any one inulin arm to 
the placebo arm using a two-sided t-test with 0.05 significance level and assuming equal variance.  
If the inulin arms have similar changes in SCFA producer levels, they will be combined and the 
study would have 80% power to detect a minimum 0.63 SD for inulin compared to placebo at the 
.05 significance level.  This sample size will also have 80% power to detect 0.33 SD differences 
among the 3 means versus the alternative of equal means using an F-test at the .05 significance 
level. 
 
Secondary outcome: MDRO colonization.  Power calculations were performed using the observed 
42% rate of MDRO colonization in our prospective ICU cohort and 1:1:1 allocation to the 
intervention.  Because this is an early phase study conducted with the expectation that promising 
results will be followed up by definitive larger studies, sample size calculations are based on one-
sided testing with alpha .10 using a t-test based on the binomial distribution.  Under these 
assumptions, a sample size of 90 patients will yield 80% power to detect a 20.4% or less rate of 
MDRO colonization on Day 7 with inulin, assuming that the two inulin arms are combined.  If the 
inulin arms are not combined, there will be 80% power for a 17.3% or less rate of Day 7 MDRO 
colonization comparing any one inulin intervention arm vs placebo.  The two-sided 95% 
confidence interval for the effect size estimate within the combined inulin arms would be 7.1 to 
26.6% for an estimate centered on 15% and 10.8 to 32.3% for an estimate centered on 20%. 
 
17.4. Interim analyses 
Interim analyses will be performed for safety, and the DSMC will have the ability to halt the trial 
at any time.  Full DSMP reports will be produced after 10, 20, 30, and 45 patients are randomized 
and at least semi-annually based on the date of enrollment of the first patient.  Toxicity monitoring 
will be performed using a frequentist approach and targeting an excess rate of toxic death of no 
more than 5%.  Toxicity will be monitored for each arm at the same 4 time points.  Sequential 
boundaries will be calculated using the Pocock method.  If the number of toxicities differs in any 
one study by 3, 4, 5, or 7 patients corresponding to the assessments above, enrollment will be 
halted and toxicities will be further evaluated to assess if it is appropriate to terminate the study.  
This stopping boundary yields a probability of early stopping of at most 0.05 when the dose-
limiting toxicity is equal to 0.05.  Toxic death will be adjudicated by the DSMC.  There are no plans 
for futility monitoring because of the relatively small size of the proposed trial, non-invasiveness 
of the samples collected, and the potential for longitudinal study results to add significant 
mechanistic understanding to MDRO colonization in the ICU. 
 
18. Publication policy 
Study results will be disseminated as widely as possible by publication and presentation.  The PI 
is ultimately responsible for publications arising from the study.  Funding sources will be 
recognized. 
 
 
 
 
 


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Appendix B: Abbreviations and terms 
 
ADLs 
activities of daily living 
AE 
adverse event 
ANOVA 
analysis of variance 
APACHE 
acute physiology and chronic health evaluation 
ASPEN 
American Society for Parenteral and Enteral Nutrition 
BMI 
body mass index 
CDC 
Centers for Disease Control and Prevention 
CDI 
Clostridium difficile infection 
CFU 
colony-forming units 
CLP 
cecal ligation and puncture 
CRF 
case report form 
CUMC 
Columbia University Medical Center 
DNI 
Do not intubate 
DNR 
Do not resuscitate 
DCC 
Data Coordinating Center 
DSMC 
data and safety monitoring committee 
DSMO 
data and safety monitoring officer 
ECMO 
extra-corporeal membrane oxygen 
EDTA 
ethylenediamineteatraacetic acid 
EMR 
electronic medical record 
EQ-5D-5L 
EuroQol 5-dimension 5-level questionnaire 
ER 
emergency room 
ESBL 
extended-spectrum beta-lactamase 
FDA 
Food and Drug Administration 
GCS 
Glasgow coma scale 
GI 
gastrointestinal 
GMP 
good manufacturing practices 
GRAS 
generally recognized as safe 
HRPO 
human research protection office 
IABP 
intra-atrial balloon pump 
ICD 
International classification of diseases 
ICU 
intensive care unit 
IND 
investigational new drug 
IQR 
interquartile range 
IRB 
institutional review board 
KPC 
Klebsiella pneumonia carbapenemase 
LAR 
legally authorized representative 
LDA 
least discriminant analysis 
LEfSe 
linear discriminant analysis effect size 


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LPS 
lipo-polysaccharide 
MDR 
multidrug resistant 
MDRO 
multidrug resistant organism 
MRSA 
methicillin-resistant Staphylococcus aureus 
NHSN 
National Health and Safety Network 
OR 
operating room 
ORP 
Office of Research Protections 
OTU 
operational taxonomic unit 
PCR 
polymerase chain reaction 
PI 
primary investigator 
PICRUSt 
phylogenetic investigation of communities by reconstruction of unobserved states 
PPIs 
proton pump inhibitors 
PRMRP 
peer-reviewed military research projects 
QIIME 
Quantitative Insights Into Microbial Ecology 
qPCR 
quantitative polymerase chain reaction 
REDCap 
Research Electronic Data Capture 
rRNA 
ribosomal ribonucleic acid 
RT-PCR 
real-time polymerase chain reaction 
SAE 
serious adverse event 
SCFA 
short-chain fatty acids 
SD 
standard deviation 
SAPS3 
simplified acute physiology score-3 
SIRS 
systemic inflammatory response syndrome 
SOFA 
sequential organ failure assessment 
TNF 
tumor necrosis factor 
USAMRMC 
U.S. Army Medical Research and Materiel Command 
VRE 
vancomycin-resistant Enterococcus 
WGS 
whole genome sequencing