Circulation of Shiga Toxin-Producing Escherichia coli Phylogenetic Group B1 Strains Between Calve Stable Manure and Pasture Land With Grazing Heifers

Abstract

Escherichia coli strains carrying Shiga toxins 1 and 2 (stx 1 and stx 2), intimin (eae), and hemolysin (ehxA) production genes were found in grass shoot, rhizosphere soil, and stable manure samples from a small-scale cattle farm located at the center of Netherlands, using cultivation-dependent and-independent microbiological detection techniques. Pasture land with grazing heifers in the first year of sampling in 2014 and without grazing cattle in 2015 was physically separated from the stable that housed rose calves during both years. Manure from the stable was applied to pasture via injection into soil once per year in early spring. Among a variety of 35 phylogenetic distinctly related E. coli strains, one large group consisting of 21 closely resembling E. coli O150:H2 (18), O98:H21 (2), and O84:H2 (1) strains, all belonging to phylogenetic group B1 and carrying all screened virulence traits, was found present on grass shoots (10), rhizosphere soil (3), and stable manure (8) in 2014, but not anymore in 2015 when grazing heifers were absent. Presence and absence of these strains, obtained via enrichments, were confirmed via molecular detection using PCR-NALFIA in all ecosystems in both years. We propose that this group of Shiga toxin-producing E. coli phylogenetic group B1 strains was originally introduced via stable manure injection into the pasture. Upon grazing, these potential pathogens proliferated in the intestinal track systems of the heifers resulting in defecation with higher loads of the STEC strain onto the grass cover. The STEC strain was further smeared over the field via the hooves of the heifers resulting in augmentation of the potential pathogen in the pasture in 2014, whereas in 2015, in the absence of heifers, no augmentation occurred and only a more diverse group of potentially mild virulent E. coli phylogenetic group A and B1 strains, indigenous to pasture plants, remained present. Via this model, it was postulated that human pathogens can circulate between plants and farm animals, using the plant as an alternative ecosystem. These data indicate that grazed pasture must be considered as a potential carrier of human pathogenic E. coli strains and possibly also of other pathogens.

The Role of Pea (Pisum sativum) Seeds in Transmission of Entero-Aggregative Escherichia coli to Growing Plants

“…Two other strains were used for comparison in pea plant colonization: extended spectrum beta lactamase E. coli strain 0611 and Shiga toxin producing E. coli O150:H2 strain N112 (Table 1). Strain 0611 was obtained from morning glory leaves imported as culinary herb from Thailand [34], whereas strain N112 was obtained from grass plants taken from a meadow land located in the neighbourhood of Nijkerk, the Netherlands [23]. All E. coli strains were grown overnight under shaking (180 RPM) at 37 • C in Luria Bertani broth (LB: Tryptone, 10 g; yeast extract, 5 g; NaCl, 10 g; dissolved in one L water and standard autoclaved for 20 min at 121 • C).…”

“…Escherichia coli genomes are flexible, allowing strains easily to adapt to local reigning circumstances [19][20][21]. Therefore, it should be accounted for that E. coli strains present in soil ecosystems are distinct from the ones in animals [22,23]. Human pathogens taxonomically related to E. coli, such as Salmonella enterica, also can colonize plants and even can communicate with plant cells [7,24,25].…”

“…Individual purple stained colonies, indicative for strain 55989, at the last samplings in 2015 and 2016 (respectively, at 85 and 83 DAS) were streaked to purity onto BECSA and single colonies were grown overnight in LB at 37 • C. Resulting dense cultures were either mixed with sterile glycerol to a final concentration of 20% for storage at -70 • C, or cells from one mL culture suspensions were pelleted by centrifugation after which DNA was extracted from resulting cell pellets for whole genome sequencing, using the Illumina MiSeq platform. Comparison of obtained isolates with strain 55989 and other E. coli strains, as described in van Overbeek et al [23], was made on the basis of DNA sequences of 10 cellular household genes, i.e., adenylate kinase (adk), classII fumarate hydratase (fumC), glycerol kinase (glpK), DNA gyrase subunit B (gyrB), 3-isopropylmalate dehydrogenase (icd), diaminopimelate decarboxylase (lysA), malate/lactate/ureidoglycolate dehydrogenase (mdh), methionine-tRNA ligase (metG), adeylosuccinate synthetase (purA), and DNA recombination/repair protein (recA).…”

Bacteria Broadly-Resistant to Last Resort Antibiotics Detected in Commercial Chicken Farms

“…To evaluate whether CRE and ESBL-producing E. coli detected in this study can be pathogenic, we tested our isolates using E. coli phylogenetic typing, a common technique utilized to sort E. coli isolates into groups that differ in ecological niches, life-history characteristics, and propensity to cause disease [ 65 ]. Namely, four main E. coli phylotypes (A, B1, B2, and D) are extensively used for classification [ 66 , 67 , 68 ]. We identified A, B2, and D groups in all maturity stages in both environments.…”

Transmission of Escherichia coli from Manure to Root Zones of Field-Grown Lettuce and Leek Plants

“…Factors related to leaf axis (adaxial or abaxial sides), age and surface topography as well as local environmental circumstances, such as nutrient availability and temperature, were shown to play decisive roles in plant colonization by E. coli [18]. The root zone of plants is the habitat that is commonly favored by E. coli, as demonstrated for cabbage [20], lamb's lettuce [19], lettuce [5,6,21,22] pea [15], radish [5] and grass plants [23]. Escherichia coli can accumulate on/inside roots and in rhizosphere soil (together named the 'plant root zone') when plants come into contact with contaminated soil.…”