Abstract
We studied formation of non-culturable forms of three bacteriocin (nisin) producing strains of Lactococcus lactis sub sp. lactis: MSU, 729 and F-116 under carbohydrate starvation stress. Two different types of inoculum were applied: A) unwashed cells with culture liquid, B) cells washed twice with normal 0,9%saline. Resulting total numbers of cells were 0.6 1.0×108 cells/ml for both types of inoculum. Population obtained using type A inoculum demonstrated active growth phase within first 1-5 days of incubation (up to 2.4×109 cells/ml) while those obtained using type B inoculum did not grow within that period. Type B population of strain MSU showed phenotypic dissociation that resulted in appearance of micro colonies. After that, we observed active growth phase (up to 5.2×109 cells/ml). Type B cultures of strains 729 and F-116 did not grow during the whole experiment. It was shown that type A population shifted into non-culturability faster than type B. This is due to differences in metabolic strategies and stress sensitivity of these types of population. After 1 year of incubation (383 days) culturability decreased by 3 orders of magnitude for type B (5 orders for type B population of strain MSU) and by 6 orders of magnitude for type A population. We also observed considerable reduction of cell size for type A population of strains 729 and F-116. Studies of bacteriocin activity showed that in type B population cells were up to 78 times more productive compared to those of type A cultures. This phenomenon can be explained by differences in survival strategies of population that use antibacterial potential of bacteriocins for their benefit.
Highlights
Bacterial cells in response to various stressful factors can reversibly lose ability to form colonies on traditional nu-trient media [1,2,3,4,5,6,7,8,9,10,11]
Formation of noncul-turable cells is confirmed for a wide range of microorganisms among which there are pathogenic and opportunistic types for the people [4]
Weshowed that from first days of cultivation the difference between the type A and type B population was evident. The former within 1-5 days after inoculation underwent a phase of active growth and increased in numbers from 0.6 - 1×108 viable cells/ml to 1.5 1.6×109 viable cells/ml depending on the strain. For this variant of experimental conditions the maximum number of bacteria in 1 ml did not depend on quantity of the cells, used as inoculum
Summary
Bacterial cells in response to various stressful factors can reversibly lose ability to form colonies on traditional nu-trient media [1,2,3,4,5,6,7,8,9,10,11]. After a year of observation the percentage of viable cells for type A population of strains of MSU and 729, became close to 100% again. By fifth day of incubation under conditions of carbohydrate starvation 82.1 - 99.6% of cells in population (see Table 4.1), depending on a strain and the variant of experiment did not form colonies.
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