A multicentre randomised controlled trial evaluating lactobacilli and bifidobacteria in the prevention of antibiotic-associated diarrhoea in older people admitted to hospital: the PLACIDE study protocol

A random allocation sequence using blocks of variable sizes will be produced using SAS PROC PLAN Version 9.1 (SAS Institute Inc., Cary, NC, USA). The sequence will allocate participants to either placebo or probiotic on a 1:1 basis and be stratified by centre to ensure that similar numbers of patients will be allocated to each arm of the study in each centre. The allocation sequence will not be available to members of the research team during participant recruitment and follow-up.

The absence of a scientific rationale for selecting a particular strain and dosage of organisms for specific health outcomes is reflected in the many different probiotic preparations evaluated in the prevention of antibiotic-associated diarrhea [28]. However, the use a combination of different organisms with large numbers of each strain is likely to maximize gut colonization and, thereby, colonization resistance. To minimise the risk of adverse effects such as systemic infection, we decided not to consider S. boulardii or other organisms that are not part of the normal human commensal flora.

Based on evidence of ability to survive passage through the upper gut, adherence to intestinal mucosa, excellent viability at the point of administration, our earlier study [19], and evidence that L. acidophilus neutralises C. difficile toxin in an epithelial cell assay in vitro (SP pers. comms), we selected a probiotic comprised of two strains of L. acidophilus (CUL60, National Collection of Industrial, Food and Marine Bacteria [NCIMB] 30157 and CUL21, NCIMB 30156), Bifidobacterium bifidum (B. bifidum; CUL20, NCIMB 30153) and B. lactis (CUL34, NCIMB 30172). The probiotic is prepared as lyophilised powder in a vegetarian capsule containing a total of 6 x 10¹⁰ organisms/capsule. These organisms are commercially available through BioCare, UK and Pharmax, USA. The placebo is an identical capsule containing inert maltodextrin powder. The dosage is 1 capsule/day taken with food and, where possible, between antibiotic doses, for 21 days.

To comply with labeling requirements, Obsidian Research Ltd., UK will prepare bottles labeled with each unique study number in the series and containing 21 capsules of the appropriate IMP according to the random allocation sequence. The research nurse will allocate each participant sequentially to the next unique study number in the sequence and provide them with the corresponding IMP bottle. The participant will be instructed to take the first dose of the IMP on the day of recruitment. The daily administration of the IMP will be supervised by the research nurses in ABMUHB and by the ward nurses in CDDFT during hospital admission and participants will be encouraged to complete the course once discharged.

The process for acquiring a license for provision of the IMPs was begun in March 2008 and a Medicines and Healthcare products Regulatory Agency (MHRA) inspector notified in April 2008. The MHRA inspected the facility responsible for the packaging and quality control of the IMPs and the license was granted in November 2008. We had underestimated the time required for MHRA approval and this resulted in a delay in starting participant recruitment.

Research nurses will review participants daily during admission to determine the onset, frequency and duration of any diarrhoea and ask about gastrointestinal symptoms (abdominal pain, bloating, flatus, nausea), acceptability and adverse effects of the interventions. After discharge, follow-up will be weekly by telephone call, postal questionnaire or home visit as appropriate. Participants will be provided with contact details for ready telephone access to research staff to notify the onset of diarrhoea or adverse events. All participants will be followed-up for 8 weeks after completing antibiotic treatment to a maximum of 12 weeks from recruitment for those on longer antibiotic courses.

The two outcomes used to generate the sample size, AAD and CDAD, have been identified as co-primary outcomes (Table 3). Diarrhoea is defined as 3 or more loose stools in a 24 h period. We will use the Bristol Stool Form Scale [29] to assist participants in describing stool consistency; types 5-7 will be consistent with diarrhoea.

All participants who develop diarrhoea during the study period will be asked to provide a stool sample and this will be collected during a home visit if required. The cause of diarrhoea will be determined by NHS laboratories according to their usual practice. Stools will be analysed for diarrhoeal pathogens (bacterial culture for Salmonella sp, Shigella sp, Campylobacter, E. Coli 0157; wet film for ova, cysts and parasites) and for C. difficile toxins. In ABMUHB, C. difficile will be identified by an in-house culture assay with further analysis by Meridian Premier Toxin A + B Enzyme Immunoassay (Meridian Bioscience Europe Inc., Cincinnati, USA) if required. In CDDFT, toxins are identified using the mini VIDAS system (C. diff Quik Check, Techlab, Blacksburg, U.S.A.). If a cause of the diarrhoea is not identified, a further stool sample will be collected and tested 2 days later.

For the primary outcome measures, AAD is defined as diarrhoea without pathogens detected on routine laboratory analysis, negative for C. difficile toxin and without alternative cause (e.g. laxative treatment). CDAD is defined as diarrhoea with stools positive for C. difficile A or B toxin. For quality control purposes, C. difficile culture and confirmation by immunoassay will be undertaken in 1 in 5 C. difficile toxin positive stool samples collected in Swansea. In view of the known inadequacies of current methods for the laboratory diagnosis of C. difficile [30], we will also save stool samples at −20°C from participants with diarrhoea for further testing for the glutamate dehydrogenase antigen and also lactoferrin (IBD scan and C. diff Quik Chek Complete; Alere Limited, Stockport, UK). This may result in the diagnosis of CDAD that was not detected by routine methods.

Participants who develop severe diarrhoea will be investigated and managed according to the current practice of their clinicians. We will extract information from the clinical records (e.g. laboratory parameters, findings at sigmoidoscopy, colectomy) to classify the severity of CDAD. At the end of follow-up, participants will be classified as either developing AAD, CDAD, another cause of diarrhoea (e.g. norovirus) or no diarrhoea. Other endpoints have been listed as secondary outcomes (Table 3).

Conservatively, we expected ADD to occur in 20% and CDAD in 4% of participants allocated to the placebo group [2]. To detect a 50% reduction in the frequency CDAD in the probiotic group (i.e. 2% frequency) with 80% power at the 5% significance level, would require 2,478 subjects (1,239 in each group; 1:1 allocation). At the 5% significance level, this number of participants would provide a power of ≫99% to detect a 50% reduction in ADD (i.e. 10% frequency) and a power of 90% to detect a 25% reduction in ADD (i.e. 15% frequency) in the probiotic group. To allow for 10% drop-outs and 10% loss to follow-up due to deaths unrelated to diarrhoea, we plan to recruit 2,974 participants.

Based on data from 2005/6, about 14,000 patients per year aged ≥65 years and exposed to antibiotics were admitted to the study hospitals and there was a total of 963 cases of CDAD. Conservatively, we expect to be able to recruit between 1:9 and 1:10 of eligible patients (approximately 124 patients/month) and undertake recruitment for a total of 24 months. This estimate was supported by a 2-week pilot study of the recruitment process in 23 medical and surgical wards in Morriston Hospital when research nurses visited the wards daily. Out of a total of 253 admissions aged ≥65 years, 166 patients were excluded (152 patients had no current or planned exposure to antibiotics; 12 already had diarrhoea [including 1 CDAD]; 1 active inflammatory bowel disease; 1 reported a previous adverse reaction to a probiotic). Eighty-seven (34.4%) patients were eligible to participate in the pilot study but 8 of these were either confused or not available (in theatre or undergoing investigations). Therefore, the study design was explained to 79 patients (31.2% of total admissions) and 58 (73.4%) stated that they would have agreed to participate in this study.

We will monitor the number of participants reaching study endpoints in each hospital every 3 months. The ethical permission secured for the study included all hospitals within each health authority providing an ability to extend recruitment if needed.

Primary outcomes will be analysed with standard methods for a binary outcome. Risk difference and odds ratio between two groups and their 95% confidence intervals for AAD and CDAD will be estimated. In addition, covariate adjustment analysis that includes the relevant covariates (such as age, gender, specific antibiotic, centre) will be performed. To explore whether prevention strategies should be provided to all patients or just those at high risk of diarrhoea, we will assess probiotic efficacy according to known or potential risk factors for AAD and CDAD as described previously. Sub-group effects will be identified by inspection of interaction terms, and these will be interpreted in light of power relative to main effects and supporting evidence of mechanism [31]. All analyses will be performed using SAS PROC PLAN Version 9.1 (SAS Institute Inc., Cary, NC, USA).

There are few tools that are validated for measuring QoL in older people and none specifically targeted at treatment-induced diarrhoea. We will use the generic measures EQ-5D and the York SF12 [32] to understand the broader health impact related to treatment-induced diarrhoea and facilitate cost-effectiveness analysis.

Health economic analysis will be undertaken from the perspective of the UK NHS. Resources utilised by each participant will include the cost of the probiotics, staff time involved in administering probiotics, treatment of adverse events, assessment of cases of diarrhoea (stool collection and culture/toxin assay, endoscopy) and the costs of dealing with and treating diarrhoea such as laundry, antibiotics, increased hospital stay and co-morbidities. Unit costs will be determined from discussion with relevant clinicians and finance department staff as well as from recognised published information.

Cost differences between the probiotic and placebo group will be determined and used in conjunction with differences in outcomes between groups in undertaking a cost-consequences analysis. Cost per case of diarrhoea averted will be the primary outcome measure. Sub-group analyses will determine the relative cost-effectiveness of preventative strategies in different risk groups. In addition, a cost-utility analysis will be undertaken based on the differences in costs between the two groups and in quality-adjusted life years derived from responses to the EQ-5D questionnaire.

Given the short timescale of the project, there will be no discounting of the costs or benefits. Sensitivity analyses will investigate the robustness of the results to changes in estimated costs and outcomes and probabilistic sensitivity analysis will use bootstrap resampling to determine the probability that preventive strategies are within certain thresholds. The budgetary impact from a UK NHS perspective of adopting a policy of administering a probiotic preparation to prevent or ameliorate AAD and CDAD in people aged 65 years and over admitted to secondary care NHS facilities and receiving oral or intravenous antibiotics will also be assessed.

The funders (the National Institute of Health Research Health Technology Assessment [HTA] Programme) will appoint the chairs and members for a Trial Steering Committee (TSC) and an independent Data Monitoring and Ethics Committee (DMEC). The DMEC will include an independent statistician who will generate and hold the random allocation sequence for the trial. Trial management will be centered in the Clinical Research Unit, Morriston Hospital, Swansea. A Trial Management Committee will include the Principle Investigator, the Project Manager, the CDDFT Site Co-ordinator and CDDFT hospital site leads.

In view of the expected high frequency of adverse events in an elderly population of hospital in-patients, the recording of gastro-intestinal symptoms during follow-up and the excellent safety record of probiotics, only information regarding serious adverse events (SAEs) will be recorded. SAEs will be defined and reported according to GCP guidelines [27], assessed by the local research clinicians involved in the project as to their attribution and reviewed regularly by an Independent Safety Monitor.

Suspected Unexpected Serious Adverse Reactions (SUSARs) will be defined as the development of:

SUSARs will be reported immediately to the Independent Safety Monitor to consider their attribution to the participant’s inclusion in the trial. It was considered that knowledge of a participant’s allocation would not inform the clinical management of an SAE or SUSAR and, therefore, procedures for the immediate unmasking of participant allocation will not be established. If required, unmasking could be undertaken by the Independent Statistician. The Independent Statistician will undertake an unblinded, interim analysis of SAEs and SUSARs in the first 500 participants with complete data and report the findings to the DMEC.