Among the L. lactis strains tested, i.e., of both subspecies
lactis and cremoris, 11 were shown to produce putrescine
from agmatine. Such a capacity had already been
reported for some L. lactis strains [17], and putrescine-producing
L. lactis can be found in large numbers in cheeses
with high putrescine concentrations [55]. Not all the L. lactis
strains tested were able to produce putrescine, although
the capacity to produce it from agmatine has been described
as a species-level trait [17]. Traditionally, BA-producing
pathways have been thought horizontally acquired [44,
48]. The present L. lactis strains unable to synthesize
putrescine may have lost this capacity during their use in
the dairy environment. Putrescine would negatively affect
acidification and/or final flavor, and such putrescine-producing
strains would have been avoided [17]. The loss of
this capacity seems to have occurred in two ways: (1) via
the loss of the AGDI pathway genes, as shown for strains
of L. lactis subsp. cremoris, and (2) the inactivation of the
cluster by an insertion element (IS) in L. lactis subsp. lactis
strains [17]. To differentiate between putrescine and nonputrescine
producers, a specific PCR test is available [17]
in which non-putrescine-producing L. lactis subsp. cremoris
strains do not render a PCR band, while L. lactis subsp.
lactis non-putrescine-producing strains do, although the
amplification product is 1 kb larger than expected due to
the presence of an IS element. In the present work, none
of the non-putrescine-producing strains of L. lactis subsp.
cremoris was associated with any positive amplification,
while those of L. lactis subsp. lactis rendered the expected
enlarged amplicon (Fig. 1).
Variation in the efficiency of putrescine production was
observed among the producing strains of L. lactis; this
allowed their classification as ‘strong’ or ‘medium putrescine
producers’ (Table 2). Variation in the capacity to produce
putrescine from agmatine has been described before
among L. lactis subsp. cremoris strains [31]. In the present
work, however, the greatest variation was observed among
the L. lactis subsp. lactis producers (Table 2).
One of the most effective measures for reducing the
presence of BA in dairy products is the use of starter cultures
that have been properly tested and selected as non-BA
producers [22]. The present results show that both culturedependent
and culture-independent methods are appropriate
for ruling out BA-producing strains for use as starters
and adjunct cultures. The culture-independent methods
based on PCR testing detected not only BA producers but
also non-producer strains that possessed genes involved in
BA production; these pose a risk since they might be spread
by horizontal gene transfer.
There is increasing interest in the use of autochthonous
LAB strains in starter and adjunct cultures for producing
dairy products with PGI status [8, 57]. Strains intended for
use in their manufacture should be systematically monitored
for BA production capacity to avoid the accumulation
of these toxins and thus produce safer dairy products.