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Table 1 Sampling scheme and experimental design

From: Reservoirs of antimicrobial resistance genes in retail raw milk

Sampling states Milk type Incubation temperature Incubation time No. of milk brands No. of samples Bacterial plating 16Sa SMSb (HiSeq 4000) SMSb (NovaSeq S4)
California Raw 4 °C, 23 °C 0 h, 2 h, 4 h, 6 h,12hc, 24 h 2 480 (n = 13) (n = 8)
Vat 4 °C, 23 °C 0 h, 2 h, 4 h, 6 h,12 h, 24 h 1 240
HTST 4 °C, 23 °C 0 h, 2 h, 4 h, 6 h,12 h, 24 h 3 720 (n = 11)
UHT 4 °C, 23 °C 0 h, 2 h, 4 h, 6 h,12 h, 24 h 2 480
South Carolina Raw 23 °C 0 h, 24 h 2 24 (n = 4)
Arizona Raw 23 °C 0 h, 24 h 2 30 (n = 4)
Idaho Raw 23 °C 0 h, 24 h 2 36 (n = 4)
Maine Raw 23 °C 0 h 2 24
  1. For samples from California, the incubations at different temperature shared a single start point (0 h). indicates all the collected samples were subjected to a type of analysis unless a number is specified, while ✕ represents the corresponding analysis was not performed. For the HiSeq 4000 run, 13 raw milk samples (6 samples were collected before any incubations and the remaining 7 samples experienced a 24 h incubation at 23 °C) and 11 HTST samples (5 samples were recruited before incubation and the other 6 were incubated at 23 °C for 24 h) were included to determine the resistome variances between milks as well as before and after room temperature incubation. Unequal number of metagenomes were obtained (i.e., 13, 11) as a result of unexpected sequencing failure of certain DNA samples. In the NovaSeq S4 run, 4 raw milk samples after a 24 h incubation at 23 °C from each state (California, South Carolina, Arizona, and Idaho) were used to assess the geographical variance of milk resistomes, and another 4 samples before incubation from the California cohort were included to validate results between sequencing platforms/runs
  2. a16S rRNA gene sequencing
  3. bShotgun metagenomic sequencing
  4. cSamples from 12-h incubation under 4 °C were skipped for bacterial plating