Detection of African Swine Fever Virus in Feed and Feed Mill Environment Following Extended Storage

Table 2

Proportion of qPCR positive and main effects of batch and day on cycle threshold (Ct) value and log₁₀ genomic copies/g for feed held in room temperature storage for ASFV DNA survival after experimental inoculation of swine feed and subsequent feed batch sequencing.

Item^†	Proportion PCR positive	Cycle threshold value^‡	Log10 genomic copies/g^§
Batches of feed
2	82/82	32.1^a	4.9^c
3	58/62	37.7^b	3.3^b
4	37/62	41.8^c	1.5^a
5	37/62	42.3^c	1.6^a
6	26/62	43.2^c	1.0^a
Day
1	115/120	37.9^a	3.5^b
3	17/30	39.8^a,b	2.2^a
7	16/30	39.9^b	2.2^a
14	24/30	38.1^a,b	3.0^a,b
28	18/30	39.6^a,b	2.3^a
60	18/30	40.0^b	2.2^a
90	17/30	39.8^a,b	2.2^a
180	15/30	40.3^b	2.0^a

Note: ^†Swine gestation feed was inoculated with African swine fever virus (ASFV) for a final concentration of 5.6 × 104 TCID50/gram (Batch 2) following an initial priming of the feed manufacturing equipment with ASFV free feed. Four subsequent batches of initially ASFV-free feed were then manufactured (Batches 3–6). On Day 1, 3, 7, 14, 28, 60, 90, and 180 after manufacture following room temperature storage, three samples were mixed with approximately 35 mL of phosphate buffered solution, incubated for 2 hr at room temperature then centrifuged at 1,000 × g for 3 min. Samples were then analyzed using qRT-PCR for detection of the gene encoding for the p72 protein. Analysis of Day 1 feed samples have been reported by Elijah et al. [12] and are included in the current analysis of ASFV detection over time. ^‡Samples that had no detectable ASFV DNA were assigned a Ct value of 45.0. Batch, , SEM = 0.43; Day, SEM for Day 1 = 0.31, otherwise SEM = 0.57. ^§Log₁₀ genomic copies/g feed. Batch, , SEM = 0.19; Day, , SEM for Day 1 = 0.13; otherwise SEM = 0.25. ^a–cMeans within item lacking common superscript differ () using Tukey multiple comparison adjustment.