High-Frequency Conjugation System Facilitates Biofilm Formation and pAMβ1 Transmission by Lactococcus lactis

Abstract

The importance of conjugation as a mechanism to spread biofilm determinants among microbial populations was illustrated with the gram-positive bacterium Lactococcus lactis. Conjugation triggered the enhanced expression of the clumping protein CluA, which is a main biofilm attribute in lactococci. Clumping transconjugants further transmitted the biofilm-forming elements among the lactococcal population at a much higher frequency than the parental nonclumping donor. This cell-clumping-associated high-frequency conjugation system also appeared to serve as an internal enhancer facilitating the dissemination of the broad-host-range drug resistance gene-encoding plasmid pAM␤1 within L. lactis, at frequencies more than 10,000 times higher than those for the nonclumping parental donor strain. The implications of this finding for antibiotic resistance gene dissemination are discussed.Lactococci are important fermentation starter cultures. As commensal organisms, they are widely distributed (19, 37) and can be found coexisting with many other organisms, including pathogens, in natural and food-processing environments, most commonly in the form of mixed-culture ecosystems. However, to date information regarding lactococcal biofilm formation is very limited (30). The contribution of commensal organisms in mixed-culture biofilm development involving pathogens and other risks associated with these biofilms have not been fully explored.Lactococci are susceptible to various gene transfer mechanisms (13). Many important traits, including lactose utilization, the proteolytic system, bacterial phage resistance, and nisin production and immunity, are associated with mobile elements. Of particular interest, a cell-clumping-associated high-frequency conjugal gene transfer system has been reported in two similar settings involving Lactococcus lactis strains ML3 and 712. In both cases, the conjugative elements, i.e., the sex factor in 712 and pRS01 in ML3, mobilized the transfer of the Lac plasmid to the recipient cells by forming plasmid cointegrates. Some of the conjugation progenies exhibited cell autoaggregation. When these clumping cells served as donors in the second round of mating, they transferred the Lac plasmid 10 2 to 10 7 times more efficiently than the original donor strain (1, 10, 40). Divalent ions were required for autoaggregation, and proteinase treatments significantly decreased both cell clumping (Clu ϩ ) and high-frequency conjugation (41). Strain MG1363 is a nonclumping, plasmid-cured derivative of strain 712 with the sex factor retained in the chromosome. A cluA gene, which encoded a putative cell surface protein containing the well-conserved hexapeptide LPXTGE, was cloned from the sex factor. Expression of the CluA protein through upstream fusion of a lactococcal heat shock promoter in MG1363 partially restored cell aggregation (16).It is anticipated that the biofilm environment would be suitable for gene transfer events such as conjugation and transformation. However, a two-way relationship was recently demonst...

Synergistic Effects in Mixed Escherichia coli Biofilms: Conjugative Plasmid Transfer Drives Biofilm Expansion

“…Indeed, in oral biofilms, the best understood multispecies biofilm, a combination of these mechanisms appears to govern coaggregation among resident species leading to sequential development of dental plaque (11). Similarly, it has been shown in studies with Escherichia coli K-12 (9) and Lactococcus lactis (13) that horizontal gene transfer within the biofilm community can introduce additional dynamics due to an enhanced expression of clumping factors during plasmid transmission, which also promotes biofilm formation by their new hosts. In conclusion, information derived from in vitro studies of single-species biofilms will not be directly transferable to consortia containing multiple genetically distinct species or strains.…”

The ABC-Type Multidrug Resistance Transporter LmrCD Is Responsible for an Extrusion-Based Mechanism of Bile Acid Resistance in Lactococcus lactis

“…One of the most prominently upregulated genes is cluA. This gene confers a cell aggregation phenotype in L. lactis (34), which would explain the clumping phenotype observed. A similar role for the product of llmg1399 can be envisaged as it lies in close proximity to cluA and shares high homology with it.…”

“…CluA is a 136-kDa surface-bound protein covalently linked to the cell wall peptidoglycan. This protein is not only responsible for a constitutive aggregation phenotype in L. lactis MG1363 but also linked to high-frequency conjugation and transfer of the sex factor (34,42). The upregulation of the sex factor genes might thus be responsible for a major morphological change such as the clumping of the cells that may provide a certain level of protection to the inner cells in the aggregate to cholate.…”

Biofilm Formation on Stainless Steel by Streptococcus thermophilus UC8547 in Milk Environments Is Mediated by the Proteinase PrtS

“…The frequency of pAM␤1 transfer was substantially higher in the wild-type strain than in its mutants. A relationship between biofilm formation and high frequency of pAM␤1 conjugal transfer was observed in Lactococcus lactis expressing the CluA protein, a binding substance that induces cell aggregation, high conjugal transfer, and biofilm formation (29).…”