Rezusta 2016
Rezusta 2016
Rezusta 2016
Original article
Summary
In general, it is recommended to incubate dermatophytes cultures for a minimum of 4 weeks. Several aspects of
routine fungal cultures should be evaluated in order to implement appropriate and necessary changes. The aim of
this study was to determine the optimum incubation time for routine dermatophytes cultures, analysing the time
to find first fungal growth by visual observation. We recorded the time when the initial growth was detected for
all dermatophyte isolates during a 4-year period. A total of 5459 dermatophyte cultures were submitted to our
laboratory. From the total cultures, only 16 (1.42%) isolates were recovered over/after 17 days of incubation and
only three dermatophyte species were recovered over 17 days. Fourteen isolates belong to Trichophyton rubrum,
one isolate to Trichophyton mentagrophytes complex and one isolate to Epidermophyton floccosum. We concluded that
an incubation period of 17 days is enough to establish a microbiological diagnosis of dermatophytosis.
Key words: dermatophytes, dermatophytosis, dermatophyte carriage, onychomycosis, incubation time, mycoses.
read twice a week recording the number of days until Table 1 Distribution of incubation time for days when dermato-
dermatophyte growth was observed. On the other phytes were isolated.
hand, they were considered clinically non-relevant if 0–7 days 0–14 days 0–17 days >17 days
they were recovered from patients who had the same
organism isolated within the first 17 days of incuba- 820 1095 1112 1128
72.69% 97.07% 98.758% 100%
tion from the other specimen source.
Dermatophytes were identified by their macro and
micromorphologic, phenotypic characteristics accord-
six isolates we could not justify the slow growth or the
ing with Summerbell and a sequence analysis of the
delay in the identification process (Fig. 1).
internal transcribed spacer (ITS) of the ribosomal DNA
Only three dermatophyte species were recovered
was performed for strains with no reliable identifica-
over 17 days. Fourteen isolates (87.5%) belong to Tri-
tion by phenotypic methods. For this purpose, a DNA
chophyton rubrum, one isolate (6.25%) to Trichophyton
fragment of about 1000 bp was amplified by poly-
mentagrophytes complex and one isolate (6.25%) to Epi-
merase chain reaction (PCR) using universal fungal
dermophyton floccosum. Regarding to the location
primers (ITS4 and ITS5).6 The sequences of the total
where the samples were obtained, 14 (87.5%) isolates
ITS region were subjected to a BLASTn search (com-
recovered over 17 days were from nails and only two
pares nucleotide similarities) of the National Center for
(12.5%) from the skin. Dermatophyte species recov-
Biotechnology database.
ered over 17 days and their location are summarised
Considering that some factors could influence the
in Table 2.
recovery and identification of dermatophytes, we anal-
ysed the used of previous antifungal drugs by the
patients or possible failures based on untrained person- Discussion
nel in the laboratory.
We conducted a critical evaluation of the protocol for
dermatophytes isolation. This study shows that incu-
Results bation times shorter than 17 days are enough time for
dermatophytes to grow from clinical isolates. Although
A total of 5459 dermatophyte cultures were submitted
dermatophyte differentiation could be ready some days
to our laboratory. One thousand one hundred and
twenty-eight (20.66%) of these were positive for der-
matophytes. Eight hundred and twenty (72.69%) iso-
lates were recovered during the first 7 days, 275
isolates (24%) between 7 and 14 days, and 17 isolates
(1.51%) 15 and 17 days. Only 16 (1.42%) isolates
were recovered over/after 17 days of incubation
(Table 1). The proportion of dermatophytes recovered
before 17 days was significantly higher than those
recovered over 17 days. (P < 0.001, X2 test). Consid-
ering only isolates recovered over 17 days, in four
cases were non-relevant due to the same organism
had already been isolated from other of their samples
during the first 17 days. In addition, seven of the iso-
lates recovered over 17 days, had positive direct vision
with KOH. Therefore, in these patients no change in
the therapeutic approach was implemented because
specific antifungal treatment had been prescribed
based on direct vision findings.
From the 12 isolates diagnosed after 17 days con-
sidered clinically relevant, four isolates belong to
patients under antifungal treatment on the specimen
collection date. In other two cases, the mycology
expert was not present during the identification pro- Figure 1 Summary of the culture results and clinical relevance
cess, which can justify the delay. Therefore, only in or possible justification of growth that occur over 17 days.
Table 2 Distribution and location of dermatophytes species perform the procedure. The reduction of the incuba-
recovered over 17 days. tion time to 17 days will make the technique more
No. % Location effective, because the microbiological diagnose is estab-
lished sooner, and more efficient, because it allows to
Trichophyton rubrum 14 87.50 Nail save money.
Trichophyton mentagrophytes complex 1 6.25 Skin
On the basis of these results, we conclude that
Epidermophyton floccosum 1 6.25 Skin
16 100 mycological cultures positive after 17 days are rarely
relevant for patient care, and in most cases, the incu-
bation time could be reduced. Validation of our pro-
later, we emphasise that to shorten the incubation posal in different settings should be done, especially in
time until visual growth is very convenient. Previous fungal reference laboratories, before being established
studies indicate that fungal cultures for the detection as a general recommendation.
of dermatophytes fungi have to be incubated for
4 weeks. The reduction of the incubation time to Conflicts of interest
17 days will make the technique more effective and
more efficient. The authors have no conflict of interest to declare.
Traditional recommendations are based on the slow
growth of some fungal species, notably dimorphic References
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