Enterobacteriaceae enrichment broth

Of each meat sample, 10–20 g were placed in a sterile blender bag (Seward, Worthing, United Kingdom (UK)), homogenised at a 1:10 ratio in Enterobacteriaceae enrichment (EE) broth (BD, Franklin Lakes, United States), and incubated at 37 °C for 24 hours.
For the detection of ESBL-producing Enterobacterales, one loopful of each of the EE cultures was streaked onto Brilliance ESBL agar plates (Oxoid, Hampshire, UK). Plates were incubated under aerobic conditions at 37 °C for 24 hours. Colonies with different coloration and growth morphology were sub-cultured on Brilliance ESBL agar plates at 37 °C for 24 hours. From each plate, single colonies were picked and sub-cultured on plate count agar (PCA) for 24 hours at 37 °C.
Species were identified using matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS, Bruker Daltonics, Bremen, Germany). Bacterial identification was carried out using the software Flex Control 3.4., the MALDI Biotyper (MBT) Compass database version 4.1.100, and the MBT Compass Library Revision H 2021.

During September 2020, a total of 24 officially registered organic, and 30 conventional dairy farms were visited throughout four cantons in the northwest region of Switzerland. Visits were conducted with the approval of the farmers. Only calves that were born on the respective farm were included in the study. Where available, data on antimicrobial usage (AMU) was recorded. In the current study, AMU included antimicrobial treatment of the calves or feeding of discard milk from cows treated with antibiotics.
The age of the calves ranged from 2 to 120 days. To ensure a noninvasive procedure, fresh feces were collected from pen floors and animal enclosures using plastic bags. A total of 196 samples were collected and placed in cooler boxes for transport and stored at −20°C until processing.
Before microbiological analysis, the samples were thawed at 4°C overnight. A sterile cotton swab of each sample was placed in a sterile blender bag (Seward), homogenized at a 1:10 ratio in Enterobacteriaceae enrichment (EE) broth (BD), and incubated at 37°C for 24 h.