Identification of microorganisms for the production of carbohydrolytic enzymes is extremely important
given the increased demand for these enzymes in many industries. To this end, dye–polysaccharide interactions
which provide a visual indication of polymer hydrolysis (clear zones or halos) have been used for
decades. For the detection of extracellular cellulase or xylanase activity many laboratories use Gram’s
iodine as the chromogenic dye, as it is a more rapid initial screening method compared to the use of other
dyes. Here, we compared Gram’s iodine and Congo red as indicators of polysaccharide hydrolysis. We
attempted to detect cellulase activity using carboxymethylcellulose, and xylanase activity using birchwood
xylan, in fourteen uncharacterized bacteria isolated from wood chips. Our results indicate that
Gram’s iodine may lead to identification of false positives in a typical screening protocol and that Congo
red allows for avoidance of such pitfall. Congo red allowed detection of cellulase activity from live microbial
colonies but not Gram’s iodine. To confirm this, detection of enzymatic activity was also assessed
using cell-free enzyme preparations. Congo red was found to be reliable in detecting cellulase activity
with isolated enzymes preparations. Under the same conditions, neither of these dyes detected xylanase
activity, despite independent evidence of xylanase activity for one of the preparations. We detected
xylanase activity for this particular enzyme preparation using a coloured derivative of xylan (Remazol
Brillant Blue R-xylan adduct) that respond to xylan hydrolysis. Our results suggest that methods that rely
on interactions between a dye (Congo red or Gram’s iodine) and a polymeric substrate (carboxymethylcellulose
or birchwood xylan) for indirect detection of hydrolysis may require the use of relevant controls
and independent confirmation of enzymatic activities.