Table 3. Characteristics of the polymerase chain reaction (PCR) primers used in this study for strain characterization.

Primer	Primer sequence (5´ to 3´)	Target	PCR program (30 cycles)a	Product size (bp)	Reference
P1b	CTGAACGGCGATTACGCGAA
P2	CCAGACGATACGATCCAG	eae c	94°C, 30 s; 53°C, 30 s; 72°C, 60 s	917	(17)
P3	CTGGAGTTGTCGATGTT	eae-α	94°C, 30 s; 53°C, 30 s; 72°C, 120 s	1,648	(17)
P4	GTAATTGTGGCACTCC	eae-β	94°C, 30 s; 53°C, 30 s; 72°C, 120 s	1,926	(17)
P5	GCCTCTGACATTGTTAC	eae-γ	94°C, 30 s; 53°C, 30 s; 72°C, 120 s	1,770	(17)
ecsD-lower	TATTTTCAAAAAGAATGATGTC	eae	94°C, 30 s; 56°C, 60 s; 72°C, 150 s	»2,990d	(18)
SK1e	CCCGAATTCGGCACAAGCATAAGC
LP5	AGCTCACTCGTAGATGACGGCAAGCG	eae-ε	94°C, 30 s; 55°C, 60 s; 72°C, 120 s	2,608	(18)
LP6B	TAGTTGTACTCCCCTTATCC	eae-ζ	94°C, 30 s; 53°C, 60 s; 72°C, 150 s	2,430	(18)
LP7	TTTATCCTGCTCCGTTTGCT	eae-ι	94°C, 30 s; 52°C, 60 s; 72°C, 150 s	2,685	(18)
LP8	TAGATGACGGTAAGCGAC	eae-η	94°C, 30 s; 52°C, 60 s; 72°C, 150 s	2,590	(18)
LP10	GGCATTGTTATCTGTTGTCT	eae-κ	94°C, 30 s; 52°C, 60 s; 72°C, 150 s	2,769	(18)
LP11B	GTTGATAACTCCTGATATTTTA	eae-θ	94°C, 30 s; 50°C, 60 s; 72°C, 150 s	2,686	(18)
LPFDF LPFDR	GAACTGTAGATGGGTAC AGCAGGCATAACGCAAG	lpfD	94°C, 60 s; 48°C, 50 s; 72°C, 60 s	798	(19)
Donne-280 Donne-281	CGGAACAGTAGGTTCACCTTC AGTGCCCGTGTTCTTGAACTG	efa1	94°C, 30 s; 50°C, 30 s; 72°C, 120 s	2,226	(20)
EASTOS1 EASTOS2	GCCATCAACACAGTATATCCG CGCGAGTGACGGCTTTGTAG	astA	94°C, 30 s; 50°C, 60 s; 72°C, 90 s	109	(10)
AggRks1 AggRkas2	GTATACACAAAAGAAGGAAGC ACAGAATCGTCAGCATCAGC	aggR	94°C, 30 s; 50°C, 60 s; 72°C, 45 sf	254	(21)

^aBefore the first cycle, the sample was denatured for 10 min at 95°C; after the last cycle, the sample was extended for 7 min at 72°C.
^bPrimer P1 was used as forward primer in all PCR reactions in combination with P2, P3, P4, and escD-lower.
^cConserved region of eae.
^dAmplicon sequenced.
^ePrimer SK1 was used as forward primer in all PCR reactions in combination with LP5, LP6B, LP7, LP8, LP10, and LP11B.
^f35 cycles.