Spontaneously induced prophages in Lactobacillus gasseri contribute to horizontal gene transfer

Abstract

Lactobacillus gasseri is an endogenous species of the human gastrointestinal tract and vagina. With recent advances in microbial taxonomy, phylogenetics, and genomics, L. gasseri is recognized as an important commensal and is increasingly being used in probiotic formulations. L. gasseri strain ADH is lysogenic and harbors two inducible prophages. In this study, prophage adh was found to spontaneously induce in broth cultures to populations of ∼ 10(7) PFU/ml by stationary phase. The adh prophage-cured ADH derivative NCK102 was found to harbor a new, second inducible phage, vB_Lga_jlb1 (jlb1). Phage jlb1 was sequenced and found to be highly similar to the closely related phage LgaI, which resides as two tandem prophages in the neotype strain L. gasseri ATCC 33323. The common occurrence of multiple prophages in L. gasseri genomes, their propensity for spontaneous induction, and the high degree of homology among phages within multiple species of Lactobacillus suggest that temperate bacteriophages likely contribute to horizontal gene transfer (HGT) in commensal lactobacilli. In this study, the host ranges of phages adh and jlb1 were determined against 16 L. gasseri strains. The transduction range and the rate of spontaneous transduction were investigated in coculture experiments to ascertain the degree to which prophages can promote HGT among a variety of commensal and probiotic lactobacilli. Both adh and jlb1 particles were confirmed to mediate plasmid transfer. As many as ∼10(3) spontaneous transductants/ml were obtained. HGT by transducing phages of commensal lactobacilli may have a significant impact on the evolution of bacteria within the human microbiota.

FIG 1

Mitomycin C (μg/ml) induction of bacteriophage from the ϕadh lysogen NCK100 (A) and from the ϕadh-cured derivative of L. gasseri NCK102 (B). The data shown are representative of several replicate experiments.

FIG 2

Negatively stained TEM images of the temperate phages that reside in L. gasseri NCK100, taken at a magnification of 55,000×. (A) Siphoviridae phage ϕadh. (B) Myoviridae phage jlb1.

FIG 3

(A) Map of the open reading frames and known features of phage jlb1. (B) Alignment of genomes of phages KC5a, jlb1, LgaI, Lj771, and ϕadh (from top to bottom). Similarly colored blocks are regions of local DNA colinearity that are homologous and internally free from genomic rearrangement. Small rectangles below each genome are open reading frames. Colored graphs within the blocks show the similarity profiles of the genome sequences. Areas that are completely white were not aligned and probably contain sequence elements specific to a particular genome.

FIG 4

(A) Growth of the ϕadh and jlb1 lysogen NCK100 in MRS broth under controlled-pH and batch culture conditions. (B) Spontaneous induction of ϕadh in broth medium maintained at various pHs over time. The data shown are representative of several replicate experiments.

FIG 5

Coculture experiment using L. gasseri NCK374 Cm^r (containing pTRK170) and transduction recipient L. gasseri NCK110 (Rif^r and Gen^r at 25 μg/ml and 200 μg/ml, respectively). Every 2 h, samples monitored the PFU/ml of ϕadh, total cell count, and the number of spontaneous transductants/ml (L. gasseri Rif^r Gen^r Cm^r). The data shown are representative of several replicate experiments.