Agriculture Reference
In-Depth Information
used in combination. Overall, the delivery rate was strain-method-specifi c with some
strains yielding higher endophyte populations than others using the same delivery
method. Internalization was also strain-dependent meaning that the endophytes varied
in their ability to infi ltrate the plant. In a similar study comparing fi ve different deliv-
ery methods—seed inoculation, soil drenching, foliar spray, pruned-root dip, and a
combination of seed inoculation and soil drenching—the pruned-root dip method was
most effective in introducing endophytic bacteria into maize (Bressan and Borges
2004). Using seed inoculation as the delivery method,
Bacillus subtilis
— a potential
biological control agent against black rot disease in cabbage—infi ltrated the roots and
moved into the stems and leaves (Wulff and others 2003). With respect to transplanting
systems, mixing bacteria into a soil-free mix before seed sprouting provided an ideal
means to deliver endophytes into plants (Yan and others 2003).
The above methods developed in plant pathology can be similarly adapted to study
internalization of human pathogens in produce. Foodborne bacteria can invade plant
tissue at many points along the farm-to-fork continuum. Methods used to introduce
foodborne endophytic bacteria into plants during preharvest are based on the most
plausible routes of entry, including the seed, soil, irrigation water, and sewage.
Contamination of sprouts typically involves submerging or spot-inoculating seeds with
the target organism, leading to internal colonization during sprouting (Dong and others
2003; Itoh and others 1998; Warriner and others 2003b). For lettuce, inoculated soil
and water are commonly used (Solomon and others 2002; Wachtel and others 2002a).
Stem injection and fl ower brushing have also been used to introduce
Salmonella
and
E. coli
O157:H7 into tomatoes (Ibarra-Sánchez and others 2004; Guo and others
2002b). Root pruning, together with root-knot nematode damage, can also serve as an
inoculation method (Hora and others 2005). In contrast, other methods have been used
to introduce these same organisms into harvested plant material during processing.
Niemira (2007) successfully used vacuum perfusion to force
E. coli
O157:H7 into the
leaf vasculature and apoplast of spinach and lettuce, whereas Hajdock and Warriner
(2007) introduced
Salmonella
and
E. coli
O157:H7 into lettuce via vacuum fi ltration.
Infi ltration by submerging produce in contaminated fl ume water that is colder than the
product will allow bacteria to infi ltrate damaged tissue and natural openings (Burnett
and others 2000; Han and others 2000; Penteado and others 2004; Seo and Frank 1999).
In a thorough study by Lang and others (2004), an attempt was made to standard-
ize the dip, spot, and spray methods used to inoculate lettuce and parsley with
E. coli
O157:H7,
Salmonella,
and
Listeria monocytogenes.
Signifi cantly higher pop-
ulations of
E. coli
O157:H7 and
Salmonella
were recovered using dip as compared
to spot or spray inoculation. Although
E. coli
O157:H7 and
Salmonella
populations
recovered from spot- and spray-inoculated lettuce were not signifi cantly different,
spot-inoculation did yield signifi cantly higher populations than spray inoculation for
parsley. Signifi cantly different numbers of
L. monocytogenes
were also recovered
from inoculated lettuce (dip
spot), indicating that these inoculation methods
are both produce- and bacteria-dependent. However, bacterial internalization was not
assessed in this study.
Finally, two nonmicrobial approaches—namely aqueous dye solutions and fl uores-
cent microbeads—have been used as indicators of bacterial infi ltration into produce.
Immersing the blossom end of intact apples in a red dye solution and a bacterial
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