Little is known about the role of lineage of strains of Clostridioides difficile (CD) on the clinical presentation of CD infection (CDI) in Latin America, especially regarding the treatment response. We conducted a multicenter, prospective study to investigate the predictive factors and treatment outcomes of CDI in hospitalized patients and to performed phenotypical and molecular characterization of CD strains. A total of 361 diarrheic patients at 5 hospitals from different regions of the country were enrolled. All stool samples were tested for glutamate dehydrogenase (GDH), toxins A and B, and toxin genes using a nucleic acid amplification test (NAAT). Specimens were cultured and susceptibility profile and whole-genome sequencing (WGS) were performed. CDI positivity was 15% (56/377). Predictive factors for CDI were prior use of meropenem (OR 4.09, 95% CI 2.097–7.095; p<0.001), mucus in stools (OR 3.29; 95% CI 1.406–7.722; p=0.006) and neutrophil left-shift with >20% of bands (OR 3.77; 95% IC 1.280–11.120; p=0.016). Overall mortality was 19%, with no deaths attributed to CDI. Oral metronidazole was used in 74% of cases, with 85% of cure and 14% of recurrence. A total of 35 CD isolates were recovered, all of them susceptible to metronidazole and vancomycin. The WGS revealed 17 different STs, six of which were novel. ST42 was the most common ST and hypervirulent strains were not found. Severe CDI were caused by ST42, ST5, ST8, ST48, ST33 and a novel ST667. The ermB gene was more frequently found in isolates of ST42 (p=0.004).