Clinical Trial Details
— Status: Terminated
Administrative data
| NCT number |
NCT02045329 |
| Other study ID # |
COMIRB 13-1513 |
| Secondary ID |
Grant funding |
| Status |
Terminated |
| Phase |
|
| First received |
|
| Last updated |
|
| Start date |
October 2013 |
| Est. completion date |
June 5, 2015 |
Study information
| Verified date |
May 2018 |
| Source |
VA Eastern Colorado Health Care System |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Observational
|
Clinical Trial Summary
The normal bacteria that inhabit the human nose, also known as the nasal microbiome, may
serve as a host defense mechanism against colonization and infection by Staphylococcus
aureus, including methicillin resistant S. aureus (MRSA). Mupirocin is a topical
antibacterial agent that may be used to clear nasal colonization with S. aureus, and reduce
risk of S. aureus infection. The impact of mupirocin on the normal nasal microbiome is not
known. We hypothesize that the nasal microbiome is changed by mupirocin. Our study aims to
define the nasal microbiome before and after decolonization therapy with mupirocin.
Description:
A. Outcome Measures: This is essentially a descriptive study. We will define the nasal
microbiota before and after decolonization therapy with mupirocin.
B. Description of Population to be Enrolled: Patients at VA-Denver who are starting a course
of nasal mupirocin therapy for S. aureus decolonization will be enrolled. The common
indications for decolonization therapy are preparation for joint replacement surgery and to
facilitate removing patient from isolation for a prolonged stay on the rehabilitation
service. Written informed consent will be obtained.
C. Study design and research methods: A nasal swab will be obtained just prior to initiation
of mupirocin therapy, within 48 hours of the completion of mupirocin therapy, one week, two
weeks, four weeks, eight weeks, and 16 weeks following the completion of mupirocin therapy.
The swab will be inserted into one nare and rotated for 3 seconds. The procedure will be
repeated on the other nare. The two heads will be refrigerated immediately, and held at 2-5o
C until they can be placed into sterile micro tubes for storage at -80o C. A separate swab
will be passed through the mouth and rubbed over the tonsils and posterior oropharynx , and
stored in the same fashion.
Broad-Range PCR and High throughput DNA Sequencing PCR amplification and sequencing of rRNA
genes will follow our previously published protocol (13). In brief, DNA lysates are subjected
to PCR with pan-bacterial 16S rDNA rDNA primers, which yields libraries of PCR amplicons
representative of all bacteria or fungi in a specimen (14). Triplicate PCR reactions will be
performed and amplicons pooled for each sample. Poisoning controls spiked with bacterial
(e.g. Bacillus subtilis) genomic DNA will be assayed to detect the presence of PCR inhibitors
in template DNA preparations; although not typically a problem with nasal swabs, inhibitory
samples will be purified by ethanol precipitation and then resubmitted for PCR. PCR amplicon
libraries will be sequenced using the high-throughput Illumina MiSeq personal sequencing
platform. which is available through the University of Colorado's Division of Infectious
Diseases. This platform can generate 5-20x106 DNA sequences in a single instrument run with
mean read lengths ~450 nts. The primers used for broad-range PCR include unique barcoded
sequences in order to simultaneously sequence multiple amplicon libraries in a single
instrument run (15). We will construct and sequence libraries from 100 specimens (50
persistent MRSA carriers [cases] and 50 non-carriers [controls]) to a depth of >50,000
high-quality sequencing reads per specimen. Our preliminary study indicates that this depth
of coverage will represent a complete survey of the nares microbiota for each specimen.
Sequence Analysis Microbes present in specimens will be identified through use of the Naïve
Bayesian Classifier Tool (16) of the Ribosomal Database Project (17). To reduce the overall
complexity of the datasets, similar rDNA sequences will be clustered into operational
taxonomic units (OTUs) by clustering sequences based on taxonomic assignments. Data matrices
are then assembled that tabulate the frequency of each OTU in a sample. Sampling coverage for
each amplicon library will be estimated (18-20) and additional sequences screened if coverage
is less than 95%. All sequences will be deposited into GenBank for public use.
Validation of candidate microorganisms Microbes identified through broad-range rDNA analysis
as potentially impacting S. aureus colonization will be further evaluated based on targeted
QPCR measurements. Based on our preliminary results and previously published studies that
suggest their interference with S. aureus growth, both S. epidermidis, (femA) and
Corynebacterium spp., (rpoB PCR) will be enumerated by Q-PCR (Table, below). Primer sets for
Q-PCR assays of novel microbial groups will be designed for detection of rDNA operons (i.e.
16S-ITS-23S genes) through the ARB software package (21). In the case of previously
recognized microbial groups and/or species, PCR primer sets may be identified by literature
search.
