Distribution and characteristics of Listeria monocytogenes isolates from surface waters of the South Nation River watershed, Ontario, Canada

Abstract

Listeria monocytogenes is a facultative intracellular pathogen thought to be widely distributed in the environment. We investigated the prevalence and characteristics of L. monocytogenes isolates from surface waters derived from catchments within the South Nation River watershed (Ontario, Canada). This watershed is dominated by urban and rural development, livestock and crop production, and wildlife habitats. From June to November 2005, a total of 314 surface water samples were collected biweekly from 22 discrete sampling sites characterized by various upstream land uses. Presumptive Listeria spp. were isolated using a selective enrichment and isolation procedure, and 75 L. monocytogenes isolates were identified based on colony morphology, hemolytic activity, and amplification of three pathogenicity genes: iap, inlA, and hlyA. Thirty-two of 314 (10%) surface water samples were positive for the presence of L. monocytogenes, but detection ranged between 0 and 27% depending on the sampling date. Isolates belonging to serovar group 1/2a, 3a (50%) and group 4b, 4d, 4e (32%) were dominant. L. monocytogenes populations were resolved into 13 EcoRI ribotypes and 21 ApaI and 21 AscI pulsotypes. These had Simpson indexes of discrimination of up to 0.885. Lineage I-related isolates were dominant (61%) during the summer, whereas lineage II isolates were dominant (77%) in the fall. Isolates were, on average, resistant to 6.1 +/- 2.1 antibiotics out of 17 tested. Half of the L. monocytogenes isolates exhibited potential virulence linked to the production of a functional internalin A, and some isolates were found to be moderately to highly virulent by in vitro Caco-2 plaque formation assay (up to 28% of entry). There was a statistically significant link between the occurrence of L. monocytogenes and proximity to an upstream dairy farm and degree of cropped land. Our data indicate that L. monocytogenes is widespread in the studied catchments, where it could represent a public health issue related to agricultural land use.

FIG. 1.

Location of water sampling sites on subwatersheds located within the broader South Nation River watershed, Ontario, Canada.

FIG. 2.

Example of how nearest upstream distance and first-quartile upstream distance variables were derived for analysis. The land use attribute example given here is agricultural buildings.

FIG. 3.

Flowchart of the procedures for enrichment, isolation, confirmation, and characterization of L. monocytogenes strains.

FIG. 4.

CART-based classification tree derived using (i) L. monocytogenes absence and presence information as target data and (ii) land use variables given in Table 1 as predictor variables. Each tree node represents the total number of observations (N) for each node and the number of nodal observations with either presence or absence. Terminal nodes (points in the tree at which no further data splitting occurs) are symbolized by bold rectangles. P, primary split condition (condition used to generate the given data split in the classification tree); S, surrogate split condition; and C, competitor split condition (a competitor variable split is one that provided the second-highest improvement score in the specific nodal splitting process). Percentages in brackets following the S and C split condition information represent, respectively, improvement scores as percentages of primary split improvement scores for given nodal observations (with the primary nodal split having the highest improvement score that could be produced from the available data).