Laboratory tests for diagnosis of Clostridium difficile infection: Past, present, and future Pruebas de laboratorio para el diagnóstico de la infección por Clostridium difficile: pasado, presente y futuro

Clostridium difficilewas first isolated in 1935 in the normal flora of newborns.1 Since 1978, it has been reported to be a cause of antimicrobial-associated diarrhea, colitis, and pseudomembranous colitis.2 During the late 1970s and early 1980s, several studies concluded that two toxins, toxin A (enterotoxin) and toxin B (cytotoxin), were responsible for the characteristic symptoms of C. difficile infection (CDI).3 Subsequent investigations showed that not all strains of C. difficile expressed these toxins and that toxin-producing strains possessed a 19.6-kb pathogenicity locus containing the tcdA and tcdB genes, which encoded toxin A (308kDa) and toxin B (269kDa), respectively. Therefore, as only toxigenic strains cancausedisease, diagnosis ofCDI shouldbebased on the detection of toxins instead of only on the detection of the bacterium. The first useful method for the diagnosis of CDI was the cell cytotoxicity assay, which was developed at the end of the 1970s.4 This method was based on the detection of cytotoxic activity in stool specimens, which was observed by rounding of cells in tissue culture and neutralization of the activity by C. difficile or C. sordellii antitoxin. The high correlation between the results of this technique and the presence of disease led it to be considered the gold standard for the diagnosis of CDI for many years. During the same period, the first specific and effective selective media containing cycloserine, cefoxitin, fructose, and egg yolk (CCFE) were being developed.5 Subsequent modification of these media—replacement of egg yolk by blood and addition or modification of antimicrobials—improved the yield of C. difficile culture. During the 1980s, the development of enzyme immunoassays (EIAs) capable of detecting C. difficile toxins was a considerable advance in the laboratory diagnosis of CDI.6 The ease of use, speed (minutes to hours), and relatively low cost of these tests compared with the cytotoxicity assay and culture led to their widespread