Quantification of citrate lyase by enzyme-linked immunosorbent assay for determining the population of Lactococcus lactis subsp. Lactis biovar diacetylactis in pure and mixed cultures

Abstract

Citrate lyase production by Lactococcus lactis subsp. lactis biovar diacetylactis DRC2 was quantified by an enzyme-linked immunosorbent assay (ELISA). The citrate lyase reached a concentration equivalent to 41 ± 4 μg/ml purified citrate lyase in pure culture. When the strain DRC2, grown in mixed culture with L. lactis subsp. cremoris AM2, represented around 70% (DC culture) or 30% (CD culture) of the total initial population, the level of citrate lyase decreased to 21 ± 7 μg/ml and 4.5 ± 1.5 μg/ml respectively. The maximum bacterial concentration of strain DRC2 in pure culture reached 2.6 × 109 cfu/ml and decreased to 1.5 (± 0.2) × 109 cfu/ml and 0.5 (± 0.3) × 109 cfu/ml in DC and CD mixed cultures respectively. In mixed cultures, the proportion of the strain DRC2 was 8.5 ± 5.0% lower at the end of the fermentation than immediately after inoculation, thus showing that this strain was clearly inhibited. However, the maximum rate of citrate consumption was the same during pure DRC2 culture and CD mixed culture (2.5 ± 0.3 mmol/h) and slightly highre in DC culture (3.07 mmol/h). The maximum rate of acidification was 0.37 ± 0.04 pH unit/h regardless of the culture. A good correlation was obtained between the population of the strain DRC2 and the citrate lyase concentration determined by ELISA but no relationship was found between citrate consumption and citrate lyase synthesis. Therefore an ELISA test of this kind can be used to monitor the growth of L. lactis subsp. lactis biovar diacetylactis in mixed cultures.