Molecular typing and antibiotic resistance profiles of Clostridioides difficile strains isolated in Algeria

Abstract

Clostridioides difficile, an anaerobic, Gram-positive, spore-forming bacterial pathogen, is considered as the leading cause of hospital-acquired post-antibiotic diarrhoea. The major virulence factors of C. difficile are two toxins, toxin A and toxin B. C. difficile can either colonize patients without causing clinical manifestations (healthy carriers), or can cause a spectrum of diseases, ranging from mild diarrhoea to severe forms, such as pseudomembranous colitis, colon perforation, toxic megacolon and sepsis, which can lead to death. C. difficile infections (CDIs) frequently occur in hospital settings, and are transmitted by the faecal-oral route, following ingestion of C. difficile spores, via contaminated hands or environments. Hospitalized patients, those receiving antibiotic therapy, or the elderly are particularly at high risk of developing CDI. The high rate of recurrences and the limited number of effective antimicrobial agents complicate both the diagnosis and management of CDI. Since 2003, the incidence and the severity of CDI, with high mortality rates, have been steadily increasing, and have become a major public health problem throughout the world. This changes in the epidemiology and clinical presentations of CDI were linked to the emergence and rapid dissemination of a hypervirulent strain called 027. Between 2016 and 2019, 300 faecal specimens were collected from hospitalized patients with antibiotic-associated diarrhea. C. difficile were cultured on ChromID CDIF, and identified by Matrix Assisted Laser Desorption Ionisation-Time Of Flight mass spectrometry (MALDI-TOF MS). Toxin gene profiles were characterized by multiplex PCR. The isolates were typed by PCR ribotyping and Multilocus Sequence Typing (MLST). Antimicrobial susceptibility was tested by the Disc diffusion and E-test method against a panel of 6 antibiotics. The antibiotic resistance genetic determinants for clindamycin, erythromycin and tetracycline were analysed by PCR, and by sequencing for the quinolones. C. difficile was detected in 18 (6%) of diarrheal patients, and were assigned to 11 different PCR-ribotypes and 12 sequence types: RT085/ST39, FR248/ST259, FR111/ST48, RT017/ST37, RT014/ST2, RT014/ST14, FR247/new ST, RT005/ST6, RT029/ST16, RT039/ST26, RT056/ST34 and RT446/ST58. Three toxin profiles were detected, two toxigenic, A+B+CDT- (33.3%) and A-B+CDT- (11%); and one non-toxigenic, A-B-CDT- (55.5%). The most common ribotypes were the non-toxigenic RT085 (16.7%), followed by the toxigenic RT014 and RT 017 (11.1% each). MLST analysis grouped the isolates into two clades, 1 and 4. Clade 4 was more homogeneous, as it included mainly non-toxigenic isolates. All C. difficile isolates were susceptible to metronidazole, vancomycin and moxifloxacin, whereas 72.2% and 16.6% were resistant to clindamycin and tetracycline, respectively. In conclusion, The prevalence of CDI in this study was comparable to those reported in many studies from Europe, Africa and the middle East. The C. difficile strains circulating in our healthcare settings were diverse and include novel RTs. Overall, we believe that our data provide important information regarding the epidemiology of CDI in Algeria, and emphasizes the need for continued surveillance to detect and prevent the spread of C. difficile. Further larger studies are needed to assess the true extent of CDI in Algeria.