Interstrain interactions between bacteria isolated from vacuum-packaged refrigerated beef

Abstract

The formation of bacterial spoilage communities in food is influenced by both extrinsic and intrinsic environmental factors. Although many reports describe how these factors affect bacterial growth, much less is known about interactions among bacteria, which may influence community structure. This study investigated interactions among representative species of bacteria isolated from vacuum-packaged (VP) beef. Thirty-nine effectors and 20 target isolates were selected, representing 10 bacterial genera: Carnobacterium, Pseudomonas, Hafnia, Serratia, Yersinia, Rahnella, Brochothrix, Bacillus, Leuconostoc, and Staphylococcus. The influence of live effectors on growth of target isolates was measured by spot-lawn agar assay and also in liquid culture medium broth using live targets and effector cell-free supernatants. Inhibition on agar was quantified by diameter of inhibition zone and in broth by measuring detection time, growth rate, and maximum population density. A number of interactions were observed, with 28.6% of isolates inhibiting and 4.2% promoting growth. The majority of Pseudomonas isolates antagonized growth of approximately one-half of target isolates. Two Bacillus spp. each inhibited 16 targets. Among lactic acid bacteria (LAB), Carnobacterium maltaromaticum inhibited a wider range of isolates compared to other LAB. The majority of effector isolates enhancing target isolate growth were Gram-negative, including Pseudomonas spp. and Enterobacteriaceae. These findings markedly improve the understanding of potential interactions among spoilage bacteria, possibly leading to more mechanistic descriptions of bacterial community formation in VP beef and other foods.

FIG 1

Representative growth inhibition as determined by spot-lawn assay. Inhibition of target isolates was determined to be at four levels, ++++, +++, ++, and +, corresponding to D ≥ 4 mm, 2 mm ≤ D < 4 mm, 0.5 mm < D < 2 mm and 0 < D ≤ 0.5 mm, respectively.

FIG 2

Representative growth inhibition and promotion by CFS-broth assay.

FIG 3

Interactions among effector and target isolates. (A) Inhibition, spot-lawn assay; (B) inhibition, CFS assay; (C) promotion, CFS assay. Symbols: ◊, target; ○, effector; □, isolate tested as both target and effector. a → b = a inhibited (A and B) or promoted (C) b. Thick to thin black (solid and dashed) arrows indicate “++++,” “+++,” “++” and “+” inhibition, respectively. Medium and thin green arrows indicate “++” and “+” growth promotion, respectively. Dashed and solid black arrows indicate diffuse and clear inhibition zones, respectively, in panel A. In panels A and B, the size of an effector and target node is, respectively, positively and negatively correlated with the number and level of inhibitions. In panel C, the size of both an effector and target node is positively correlated with the number and level of promotions.