Gene organization and sequences of pediocin AcH/PA-1 production operons in Pediococcus and Lactobacillus plasmids

Abstract

Aims: To determine the locations and sequences of pediocin AcH production genes in Pediococcus parvulus ATO77 from vegetables, Lactobacillus plantarum WHE92 from Muenster cheese, and a lactose-fermenting isolate Pediococcus pentosaceus S34 from buffalo milk. Methods and Results: Plasmid curing, Southern blot hybridization, and DNA sequence analysis indicate that pediocin AcH production genes are encoded by highly similar operons in unique plasmids designated pATO77 from P. parvulus ATO77, pS34 from P. pentosaceus S34, and pWHE92 from Lact. plantarum WHE92. Structure, immunity and secretion system genes are linked together in the operons, and the promoter sequences are the same. The amino acid sequences of the encoded proteins are highly conserved between plasmids. Conclusions: Pediocin AcH production genes are located within a plasmid-borne operon cassette in all lactic acid bacterial strains examined to date. All four genes needed for production are present within a single plasmid in each strain. Significance and Impact of the Study: This is the first demonstration that the expression of a class IIa bacteriocin is directed by a common gene cassette that has been disseminated to unique plasmids in different genera of lactic acid bacteria. These plasmids should be useful for expressing pediocin AcH in Pediococcus and Lactobacillus strains used in food production.

Screening of lactic acid bacteria for bacteriocins by microbiological and PCR methods

“…Class II contains small (<10 kDa), heat-stable bacteriocins that are formed directly, without posttranslational modification. Examples are pediocin from Pediococcus pentosaceus and P. acidilactici [17], and enterocin and plantaricin 423 from Enterococcus faecium [18] and Lactobacillus plantarum 423 [19], respectively. Class III contains large bacteriocins (>30 kDa) that are heat-labile proteins.…”

“…nisin [16]) or on plasmids (e.g. pediocin [17]). Fortunately, both chromosomal and plasmid DNA can be released from bacteria simply by boiling in buffer solution.…”

Characterization of Endogenous Plasmids from Lactobacillus salivarius UCC118

“…The RepA proteins from pSF118-20 and pSF118-44 (LSL_1965 [repA 20 ] and LSL_2000 [repA 44 ], respectively) are 71% identical to each other and are 72% similar to the Rep protein of lactococcal plasmid pCI2000, which is predicted to replicate via a theta replication mechanism (31).…”

“…Distribution of related plasmids with theta-type replicons in L. salivarius strains. The detection of pSF118-20 repA (repA 20 )-and pSF118-44 repA (repA 44 )-related plasmids in 27 strains of L. salivarius was performed by a combination of PFGE and Southern hybridization. Nuclease S1-treated genomic DNA samples of L. salivarius strains were resolved by PFGE (Fig.…”

“…these plasmids may harbor genes encoding resistance to antibiotics (39,54) and metal ions (55), genes encoding bacteriocins (30,44), gene clusters for conjugation (55), genes involved in adherence and biotin metabolism (10), and genes encoding toxin-antitoxin (TA) proteins for plasmid maintenance (53). In addition to encoding such interesting traits, endogenous plasmids are the most commonly used systems to construct genetic tools especially for gene cloning and gene expression purposes (7,52) due to their ability to replicate in the original hosts.…”