Clonal dissemination of a toxin-A-negative/toxin-B-positive Clostridium difficile strain from patients with antibiotic-associated diarrhea in Poland.
OBJECTIVE
To determine the incidence of toxin-A-negative/toxin-B-positive Clostridium difficile strains and their genetic relatedness in the feces of patients suffering from antibiotic-associated diarrhea (AAD) in Polish hospitals.


METHODS
C. difficile strains were cultured from patients' stool samples. The present study characterises these strains with respect to their cytopathogenicity on McCoy cells and the absence of toxin A despite a functional toxin B as determined with commercial test kits (Culturette Brand Toxin CD-TCD toxin A test and C. difficile Tox A/B test). In addition, PCR using different primer pairs aiming at non-repeating or repeating regions of the toxin A and B genes were used to confirm the findings. All toxin A(-)B(+) strains were genetically characterised by random amplification of polymorphic DNA (RAPD) analysis, PCR ribotyping and, in part, pulsed-field gel electrophoresis (PFGE) of DNA macrorestriction fragments.


RESULTS
We here present the presence of 17 toxin A(-)B(+) strains among 159 C. difficile strains (11%) isolated from fecal samples from 413 patients with antibiotic-associated diarrhea. All 17 strains possessed the toxin B gene, demonstrated a cytopathogenic effect on the McCoy cells, and were positive in the Tox A/B test. Molecular typing of these 17 C. difficile strains revealed that 7 of 17 (41%) toxin A(-)/B(+) C. difficile strains could not be discriminated. It appeared that these strains had a genotype that could not be distinguished from that of a Japanese control strain.


CONCLUSIONS
Our observations imply that a particular genotype of toxin A(-)B(+) C. difficile has spread extensively, not only in Poland but possibly even worldwide.
Pituch
H
H
Department of Medical Microbiology, Center of Biostructure Research, The Medical University of Warsaw, Warsaw, Poland. hanna.pituch@ib.amwaw.edu.pl
van den Braak
N
N
van Leeuwen
W
W
van Belkum
A
A
Martirosian
G
G
Obuch-Woszczatyński
P
P
Łuczak
M
M
Meisel-Mikołajczyk
F
F
D000900:Anti-Bacterial Agents / Q000009:adverse effects; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000032:analysis / Q000096:biosynthesis*; D016360:Clostridioides difficile / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D003967:Diarrhea / Q000453:epidemiology / Q000382:microbiology*; D016521:Electrophoresis, Gel, Pulsed-Field; D004761:Enterocolitis, Pseudomembranous / Q000453:epidemiology / Q000382:microbiology*; D004768:Enterotoxins / Q000032:analysis / Q000096:biosynthesis*; D005243:Feces / Q000737:chemistry / Q000382:microbiology; D014644:Genetic Variation; D006801:Humans; D007124:Immunoenzyme Techniques; D015994:Incidence; D011044:Poland / Q000453:epidemiology; D016133:Polymerase Chain Reaction; D021521:Ribotyping
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004768:Enterotoxins; C041348:tcdA protein, Clostridium difficile; C031815:toxB protein, Clostridium difficile