In 2006, 3,531 fecal samples were collected from patients with diarrhea in Henan Province, China. A total of 467 (13.2%) Shigella strains were isolated and serotyped. Seventy-one Shigella flexneri strains were characterized by MIC determination, pulsed-field gel electrophoresis (PFGE), and detection of genes encoding cephalosporin resistance. Most infections were caused by S. flexneri variant X [IV:(7),8] (27.6%), S. sonnei (24.2%), and S. flexneri 2a (20.8%). However, large regional differences were observed. Significantly higher odds (2.0) of females compared to males were infected with S. flexneri 2a. Untypeable S. flexneri (-:6) isolates were absent among males, as were untypeable S. flexneri [I:(7),8] isolates among females. Patient ages ranged from 2 months to 82 years, with 231 subjects (49.7%) <5 years of age. Most of the patients were male (62.1% [n = 290]). Infections peaked in July; week 27 with 38 cases (8.1%). All of the 71 S. flexneri conferred resistance to nalidixic acid; in addition, 21% (n = 15) and 79% (n = 56) were high- and low-level resistant to ciprofloxacin, respectively. Six S. flexneri isolates {serotype 2b [II:7,(8)] and 2b [II:(3),4;7,(8)]} harbored the bla(CTX-M-14) or bla(CTX-M-15) gene. A total of 52 unique XbaI PFGE patterns were observed among the 71 S. flexneri isolates with 11 distinct PFGE clusters. This study revealed a high prevalence of shigellosis with geographical differences in the distribution of serotypes in the distribution of serotypes and also differences in comparisons by gender. A high frequency of resistance, including 100% resistance to ciprofloxacin and resistance to extended-spectrum cephalosporins, was observed. We detected several isolates exhibiting the same PFGE type and MIC profile, indicating multiple undetected outbreaks.