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Objective: To determine the presence of intestinal coliform bacteria in colicky vs healthy infants. Study design: We isolated coliform strains from faeces and performed quantitative bacterial cultures in 41 colicky and 39 healthy breastfed infants, identified using PCR with species-specific primers, strain-specific Automated Ribotyping and the API-50E kit for Enterobacteriaceae to identify the most frequent strains. Results: Coliform strains were more abundant in colicky infants (median 6.04 log10 CFU/g faeces, range 2.00–8.76) vs controls (median 4.47 log10 CFU/g faeces, range 1.00–8.08) (p = 0.026). Escherichia coli, Klebsiella pneumoniae, K. oxytoca, Enterobacter cloacae, E. aerogenes and Enterococcus faecalis were the predominant species in colicky and healthy infants. The counts of each bacterial species differed between the two groups, and the difference was significant (p = 0.002) for E. coli: median 6.30 log10 CFU/g faeces (range 3.00–8.74) in colicky infants, and median 4.70 log10 CFU/g faeces (range 2.00–5.85) in controls. Conclusions: This is the first study to evaluate the colonization patterns of gas-forming coliforms in colicky infants and healthy controls identified by molecular methods. Coliform bacteria, particularly Escherichia coli, were found to be more abundant in colicky infants. Our data could help to shed light on the cause of infantile colic.

Aims: A molecular methodology based on PCR-associated automated ribotyping was developed to specifically detect the Lactobacillus strains of two probiotic products (an orally administered lyophilized preparation and vaginal tablets) in human faeces and vaginal swabs. Methods and Results: The 16S-23S rDNA sequences and the ribotype profiles of the probiotic lactobacilli were characterized and new species-specific primer sets were designed. The identification of faecal and vaginal lactobacilli isolated from subjects administered with the probiotic products was performed by using PCR with species-specific primers followed by strain-specific automated ribotyping. Conclusions: The PCR-ribotyping identification allowed to study the colonization patterns of the probiotic lactobacilli in the human gut and vagina evidencing the strains with the best survival capability. Significance and Impact of the Study: The proposed molecular method represents a powerful tool of strain-specific identification, useful for differentiating exogenous from indigenous strains in any microbial ecosystem and for rationally choosing probiotic bacteria with the best chance of survival in the host.

Streptococcus thermophilus, Bifidobacterium infantis Y1 and Bifidobacterium breve Y8 strains were identified and enumerated by PCR assay in human fecal samples after intake of the pharmaceutical preparation VSL-3 or yogurt. ThI/ThII primer set, specific for S. thermophilus, was selected testing its specificity against several strains of enterococci, streptococci and other genera colonizing the human intestine. A culture-independent PCR protocol, developed in this study, allowed to directly detect and enumerate S. thermophilus in human feces, excluding culture-based techniques or time consuming DNA isolation and purification procedures. Intestinal persistence of S. thermophilus was studied in feces of 10 healthy subjects given VSL-3 or yogurt. Streptococcal population was detected after 3 days of administration and persisted for 6 days after the treatment suspension. In the same trial, the colonization kinetics of B. infantis Y1 and B. breve Y8 were studied by amplification of colonies with the strain-specific primer sets InfY-BV.L/R and BreY-BV.R/L, showing a host-dependent transient colonization behaviour. PCR analysis of feces from 10 patients affected by inflammatory bowel diseases (IBD) and treated with VSL-3 for 2 months showed a colonization pattern of S. thermophilus, B. infantis Y1 and B. breve Y8 similar to that observed with the healthy subjects. © 2002 Elsevier Science B.V. All rights reserved.

In a clinical trial, 10 patients suffering from irritable bowel syndrome or functional diarrhea were administered the probiotic preparation VSL-3. Preliminary results indicated that administration of VSL-3 improved the clinical picture and changed the composition and biochemistry of fecal microbiota. Titer variations of intestinal bacterial groups were evaluated by culture and PCR techniques. A significant increase in lactobacilli, bifidobacteria and Streptococcus thermophilus was observed as a consequence of probiotic treatment, while enterococci, coliforms, Bacteroides and Clostridium perfringens did not change significantly. The strains Bifidobacterium infantis Y1 and Bifidobacterium breve Y8, included in VSL-3, were specifically detected in feces of patients treated with the probiotic by using strain-specific PCR primers. In addition, fecal β-galactosidase increased and urease activities decreased as a result of changes in the intestinal microbiota induced by VSL-3 administration. © 2001 Éditions scientifiques et médicales Elsevier SAS.

For PCR specific detection of the strains Bifidobacterium longum Y 10, B. infantis Y 1 and B. breve Y 8 used in a new probiotic product (VSL-3), strains-specific rDNA primers have been developed. Spacer regions between the 16S and 23S rRNA genes (ITS) of the three strains were amplified by PCR with conserved primers and the nucleotide sequence of these ITSs were determined. On the basis of their comparison with the rDNA sequences retrieved from GenBank, we designed new primers which specifically recognize the species B. breve and the two strains B. infantis Y 1 and B. breve Y 8. Specificity of these primers was confirmed through the analysis of 60 bifidobacteria strains belonging to the more representative human species. The feasibility of this PCR method was investigated in commercial VSL-3 product and fecal samples collected from 4 patients affected by inflammatory bowel deseases and two healthy subjects before and after the VSL-3 administration. By PCR analysis of different VSL-3 commercial batches we were successful in differentiating and quantifying the strains B. longum Y 10, B. infantis Y 1 and B. breve Y 8. B. infantis Y 1 and B. breve Y 8 could be detected at high concentration in fecal specimens of both patients and subjects treated with the probiotic preparation, showing a different colonization behaviour. Seven days after the VSL-3 treatment suspension, no patients and subjects harbored B. infantis Y 1 and B. breve Y 8, indicating a transient presence of these exogenous strains.