Listeria monocytogenes is a serious pathogen with several serogroups that differ in virulence and distribution among countries. This study determined the serogroups and virulence of 116 L. monocytogenes from sheep milk (n = 52) and goat milk (n = 64) in Jordan by several multiplex PCR. The results showed that the L. monocytogenes isolates belong to five serogroups. Most isolates (37.9%) belonged to serogroup 4b (d, e), followed by serogroup 1/2a (3a) (31.9%), serogroup 1/2b (26.7%), and serogroup 4a/c (2.6%) and serogroup IV (serotypes 1/2c and 3c) (0.9%). All isolates carried the internalin (inlA, inlC, inlJ) and lmo 2672 genes. Additionally, 36.4% and 29.0 % of 4b and 1/2b isolates carried the Listeriolysin S gene (llsX). Therefore, L. monocytogenes in sheep and goat milk are potential public health hazards because they belong to the major listeriosis-causing serogroups (4b, 1/2a, and 1/2b), and carry major virulence genes. Small ruminants’ milk and cheeses should be considered high-risk foods for L. monocytogenes infection. Thus, safe production practices should be monitored and enforced in small dairy shops.