Use of colony-based bacterial strain typing for tracking the fate of Lactobacillus strains during human consumption.
BACKGROUND
The Lactic Acid Bacteria (LAB) are important components of the healthy gut flora and have been used extensively as probiotics. Understanding the cultivable diversity of LAB before and after probiotic administration, and being able to track the fate of administered probiotic isolates during feeding are important parameters to consider in the design of clinical trials to assess probiotic efficacy. Several methods may be used to identify bacteria at the strain level, however, PCR-based methods such as Random Amplified Polymorphic DNA (RAPD) are particularly suited to rapid analysis. We examined the cultivable diversity of LAB in the human gut before and after feeding with two Lactobacillus strains, and also tracked the fate of these two administered strains using a RAPD technique.


RESULTS
A RAPD typing scheme was developed to genetically type LAB isolates from a wide range of species, and optimised for direct application to bacterial colony growth. A high-throughput strategy for fingerprinting the cultivable diversity of human faeces was developed and used to determine: (i) the initial cultivable LAB strain diversity in the human gut, and (ii) the fate of two Lactobacillus strains (Lactobacillus salivarius NCIMB 30211 and Lactobacillus acidophilus NCIMB 30156) contained within a capsule that was administered in a small-scale human feeding study. The L. salivarius strain was not cultivated from the faeces of any of the 12 volunteers prior to capsule administration, but appeared post-feeding in four. Strains matching the L. acidophilus NCIMB 30156 feeding strain were found in the faeces of three volunteers prior to consumption; after taking the Lactobacillus capsule, 10 of the 12 volunteers were culture positive for this strain. The appearance of both Lactobacillus strains during capsule consumption was statistically significant (p < 0.05).


CONCLUSIONS
We have shown that genetic strain typing of the cultivable human gut microbiota can be evaluated using a high throughput RAPD technique based on single bacterial colonies. Validation of this strategy paves the way for future systematic studies on the fate and efficacy of bacterial probiotics during human clinical trials.
Mahenthiralingam
Eshwar
E
Cardiff School of Biosciences, Cardiff University, Cardiff, Wales CF103TL, UK. MahenthiralingamE@cardiff.ac.uk
Marchbank
Angela
A
Drevinek
Pavel
P
Garaiova
Iveta
I
Plummer
Sue
S
D000284:Administration, Oral; D015373:Bacterial Typing Techniques / Q000379:methods*; D005243:Feces / Q000382:microbiology; D006801:Humans; D007778:Lactobacillus / Q000145:classification* / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D010802:Phylogeny; D019936:Probiotics / Q000008:administration & dosage / Q000032:analysis*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique / Q000379:methods*; D015203:Reproducibility of Results
D012336:RNA, Ribosomal, 16S