Authors
M. R.
Bonde
and
S. E.
Nester
,
USDA-ARS, Foreign Disease-Weed Science Researh Unit, Fort Detrick, MD 21702-5023
;
A.
Khayat
,
Hunt Wesson, Inc., Fullerton, CA 92833
;
J. L.
Smilanick
,
USDA-ARS, Horticultural Crops Research Laboratory, Fresno, CA 93727
; and
R. D.
Frederick
and
N. W.
Schaad
,
USDA-ARS, Foreign Disease-Weed Science Research Unit, Fort Detrick, MD 21702-5023
ABSTRACT
Definitive identification of free teliospores of Tilletia indica, causal agent of Karnal bunt of wheat, requires polymerase chain reaction (PCR)-based diagnostic tests. Since direct PCR amplification from teliospores has not been reliable, teliospores first must be germinated in order to obtain adequate DNA. We have routinely surface-sterilized teliospores for 2 min with 0.4% (vol/vol) sodium hypochlorite (NaOCl) to stimulate germination and produce axenic cultures. However, we observed that some spores were killed even with a 2-min NaOCl treatment, the shortest feasible duration. Decreasing the NaOCl concentration in our study from 0.4% to 0.3 and 0.2%, respectively, increased teliospore germination, but treatment times longer than 2 min still progressively reduced the germination percentages. In testing alternative methods, we found “acidic electrolyzed water” (AEW), generated by electrolysis of a weak solution of sodium chloride, also surface-sterilized and increased the rate of T. indica teliospore germination. In a representative experiment comparing the two methods, NaOCl (0.4%) for 2 min and AEW for 30 min increased germination from 19% (control) to 41 and 54%, respectively, by 7 days after treatment. Because teliospores can be treated with AEW for up to 2 h with little, if any, loss of viability, compared with 1 to 2 min for NaOCl, treatment with AEW has certain advantages over NaOCl for surface sterilizing and increasing germination of teliospores of suspect T. indica.
Additional keywords:
disinfestant,
superoxidized water,
surface sterilization