In Vitro and In Vivo Inhibitory Effects of Gaseous Chlorine Dioxide Against Diaporthe batatas Isolated from Stored Sweetpotato

Abstract

Chlorine dioxide ($ClO_2$) can be used as an alternative disinfectant for controlling fungal contamination during postharvest storage. In this study, we tested the in vitro and in vivo inhibitory effects of gaseous $ClO_2$ against Diaporthe batatas SP-d1, the causal agent of sweetpotato dry rot. In in vitro tests, spore suspensions of SP-d1 spread on acidified potato dextrose agar were treated with various $ClO_2$ concentrations (1-20 ppm) for 0-60 min. Fungal growth was significantly inhibited at 1 ppm of $ClO_2$ treatment for 30 min, and completely inhibited at 20 ppm. In in vivo tests, spore suspensions were drop-inoculated onto sweetpotato slices, followed by $ClO_2$ treatment with different concentrations and durations. Lesion diameters were not significantly different between the tested $ClO_2$ concentrations; however, lesion diameters significantly decreased upon increasing the exposure time. Similarly, fungal populations decreased at the tested $ClO_2$ concentrations over time. However, the sliced tissue itself hardened after 60-min $ClO_2$ treatments, especially at 20 ppm of $ClO_2$. When sweetpotato roots were dip-inoculated in spore suspensions for 10 min prior to treatment with 20 and 40 ppm of $ClO_2$ for 0-60 min, fungal populations decreased with increasing $ClO_2$ concentrations. Taken together, these results showed that gaseous $ClO_2$ could significantly inhibit D. batatas growth and dry rot development in sweetpotato. Overall, gaseous $ClO_2$ could be used to control this fungal disease during the postharvest storage of sweetpotato.

Fig. 1. Populations of Diaporthe batatas SP-d1 on acidified potato dextrose agar treated with various ClO₂ concentrations (1, 5, 10, and 20 ppm) for 0, 1, 10, 30, and 60 min. Colony-forming units (cfus) were counted 2 days after incubation. Isolate SPd1 (200 μl of 5 × 10³ spores/ml) was spread on APDA before ClO₂ gas treatments. Error bars are the standard deviations of the means (n = 6).

Fig. 2. Photographs of sweetpotato slices drop-inoculated with Diaporthe batatas SP-d1 (10 μl of 5 × 10⁶ spores/ml) following treatments with various ClO₂ concentrations (5, 10, and 20 ppm) for 0, 10, 30, and 60 min. These photographs were taken 10 days after inoculation.

Fig. 3. (A) Lesion diameters and (B) populations of Diaporthe batatas SP-d1 on inoculated slices of sweetpotatoes treated with gaseous chlorine dioxide (ClO₂). (C) Populations of isolate SP-d1 in the surface layers of sweetpotato roots treated with ClO₂ gas. Slices were inoculated with isolate SP-d1 (10 μl of 5 × 10⁶ spores/ml) and then treated with various ClO₂ concentrations (5, 10, and 20 ppm) for 0, 10, 30, and 60 min. Roots were dipped in spore suspension (5 × 10⁶ spores/ml) for 10 min and then treated with different ClO₂ concentrations (20 and 40 ppm) for 0, 30, and 60 min. Different lowercase and uppercase letters on bars (n = 3) are significantly different between time at a given concentration and between concentrations at a given time according to the least significant difference test at P < 0.05, respectively. Repeated experiments are indicated as experiments 1 and 2.

Table 1. Analysis of variance components including the degrees of freedom (df), sum of squares (SS), F ratio, and P value for lesion diameters and fungal populations of Diaporthe batatas SP-d1 on inoculated slices of sweetpotatoes treated with gaseous chlorine dioxide (ClO₂)

Table 2. Analysis of variance components including the degrees of freedom (df), sum of squares (SS), F ratio, and P value for fungal populations of Diaporthe batatas SP-d1 in surface layers of sweetpotato roots treated with gaseous chlorine dioxide (ClO₂)