The authors have declared that no competing interests exist.
The compound 1,4-dimethylnaphthalene, originally isolated from dormant potatoes, is currently in use as a commercial sprout inhibitor. Growers and processors report a reduction in fungal infections in potatoes treated with DMN resulting in increased yields. To assess the effects of DMN on fungal growth a culture of Fusarium oxysporum was isolated from potato tubers and identified via DNA fingerprinting using the 18ITS ribosomal region. Growth of F. oxysporum was inhibited by 31% after four days of exposure to DMN but overall rate of spore germination was not affected by DMN treatment. The growth of additional fungi, including Alternaria alternata, Aspergillus niger, Epicoccum nigrum, Gnomoniopsis smithogilvyi, Phoma medicaginis, and Pythium ultimum was inhibited by DMN as was suppression of sporulation in A. niger. These results suggest that DMN is fungistatic at the application levels examined.
Harvested potatoes are often treated with sprout regulators to prolong storage. The synthetic compound isopropyl-(N-3-chloro-phenyl)-carbamate (Chlorpropham) is widely used to prevent sprouting on stored potatoes. However, the inability to use Chlorpropham on potato tubers used as seed, and concerns about residue levels on tubers used for food, has created an interest in finding alternative compounds
Essential oils derived from plant material, in many instances containing a complex mixture of hydrophobic molecules, have been shown to have antimicrobial and antifungal activity
Potato tubers (
The fungal strains
Fungal cultures were plated onto PDA plates and placed in 9.5 Liter BBL GasPak chamber (http://www.bd.com/ds/productCenter/ES-Gaspak.asp) containing filter paper strips that had 22.5 ul of 1,4-dimethylnaphthalene spotted on to it. Control chambers contained 22.5 ul of water in place of DMN. The chambers were placed at 22 ºC and incubated for two days. The rate of application of DMN was selected based on previous experiments with potatoes that have shown that this application rate results in a DMN residue level on potatoes similar to that found in commercial storage units
Fungal spores from Fusarium oxysporum and Aspergillus niger were isolated using a modification of the procedure of Espinel-Ingroff and Kerkering
Cultures of A. alternata, A. niger, E. nigrum, G. smithogilvyi, P. medicaginis, and P. ultimum were washed with 1 to 3 ml of sterile water and liquid was placed in a sterile tube and diluted 1:100 with water. Following dilution, 100 ul sample was spread on to PDA and then treated in a GasPak chamber as described for Fusarium oxysporum. Cultures were photographed after two days of exposure to DMN or the water control.
Cultures of Alternaria alternata were allowed to grow across the surface of a plate containing PDA. Using a sterile scalpel mycelial mats of about 0.5 cm were placed on to fresh PDA plates. One set of plates was transferred to a to GasPak chamber for DMN for 24 hrs and another segment of the same culture was placed in a GasPak chamber and exposed to water. Sporulation was assessed after 24 hrs.
Species identification of the isolated strain of Fusarium was based on DNA sequence of the ITS and NL amplified regions. The ITS region revealed similarity to Fusarium oxysporum while the NL regions demonstrated sequence similarity to F. oxysporum sp. lycopersici (
|
|
|
|
|
---|---|---|---|---|
|
18 ITS Forward | Alternaria alternata isolate UFSMLB09 internal transcribed spacer 1, partial sequence; 5.8S ribosomal RNA gene and internal transcribed spacer 2, complete sequence; and large subunit ribosomal RNA gene, partial sequence | 5.00E-107 | MH819177.1 |
18 ITS Reverse | Alternaria alternata isolate ALT2 small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and large subunit ribosomal RNA gene, partial sequence | 4.00E-113 | MH578598.1 | |
|
18 ITS Forward | Epicoccum nigrum strain CCTU118 internal transcribed spacer 1, partial sequence; 5.8S ribosomal RNA gene, complete sequence; and internal transcribed spacer 2, partial sequence | 3.00E-97 | MH_758719.1 |
18 ITS Reverse | Epicoccum nigrum isolate Au-V-1.1 small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1 and 5.8S ribosomal RNA gene, complete sequence; and internal transcribed spacer 2, partial sequence | 1.00E-96 | MF_475942.1 | |
|
ITFS Forward | Fusarium oxysporum small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1 and 5.8S ribosomal RNA gene, complete sequence; and internal transcribed spacer 2, partial sequence | 2.00E-101 | MG252282.1 |
ITFS Reverse | Fusarium oxysporum isolate DSM 106834 small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and large subunit ribosomal RNA gene, partial sequence | 2.00E-106 | MH575293.1 | |
NL1 | Fusarium oxysporum f. sp. lycopersici strain CBS 758.68 large subunit ribosomal RNA gene, partial sequence | 0.00E+00 | MH878407.1 | |
NL2 | Fusarium oxysporum f. sp. lycopersici strain CBS 130307 large subunit ribosomal RNA gene, partial sequence | 0.00E+00 | MH877312.1 | |
|
18 ITS Forward | Gnomoniopsis smithogilvyi isolate THDAS 2016_1120A 18S ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and 28S ribosomal RNA gene, partial sequence | 9.00E-119 | KY_695232.1 |
18 ITS Reverse | Gnomoniopsis smithogilvyi culture-collection ICMP:14040 18S ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and 28S ribosomal RNA gene, partial sequence | 1.00E-126 | KC_145868.1 | |
|
18 ITS Forward | Phoma medicaginis strain 7_ITS1F_E06_17Q 18S ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and 28S ribosomal RNA gene, partial sequence | 5.00E-100 | KF_293990.1 |
18 ITS Reverse | Phoma medicaginis strain 7_ITS4_E05_17Q 18S ribosomal RNA gene, partial sequence; internal transcribed spacer 1 and 5.8S ribosomal RNA gene, complete sequence; and internal transcribed spacer 2, partial sequence | 2.00E-103 | KF_293988.1 |
The Fusarium oxysporum isolate, when grown in the presence of DMN for four days, exhibited a decrease in growth (
Treatment of Alternaria alternata, Aspergillus niger, E. nigrum, G. smithogilvyi, P. medicaginis, and P. ultimum with DMN resulted in suppression of mycelial growth (
Treatment of
DMN, a compound that is utilized as a sprout regulator in the potato industry, has fungistatic activity against species in the Ascomycota and Oomycota. At the dosage utilized in the potato industry to prevent sprouting, DMN does not inhibit germination of fungal spores nor kill fungal cultures. However, DMN inhibited mycelial growth of all fungal species tested and decreased the rate of sporulation in A. alternaria. Thus, anecdotal evidence where growers and processors find a reduction in fungal disease in potato sheds is supported by the fungistatic effects of DMN.
The work was supported in part by grants from the 1,4-Group of Meridian, ID and by the Penn State Behrend Undergraduate Research program.