TITLE:
Bio-Control of Alternaria alternata during Banana Storage by Purified AFP Using Isoelectric Focusing Technique
AUTHORS:
Hassan Barakat
KEYWORDS:
Antifungal Protein (AFP), Alternaria alternata, Isoelectric Focusing, Antifungal Activity,
JOURNAL NAME:
Food and Nutrition Sciences,
Vol.5 No.15,
August
27,
2014
ABSTRACT:
Interestingly, antifungal protein AFP was purified from Aspergillus giganteus supernatants with
modified isoelectric focusing procedure after adaptation of the secretion
conditions. Subsequently, the antifungal activity as well as the mode of action
against Alternaria alternata was
tested in vitro. Moreover, different
concentrations of AFP were applied to banana fruits for 15 days at 20°C in vivo. Obtained
results illustrated that A. giganteus was able to secrete about 39.78 ± 2.39 mg AFP·l-1 Olson medium. The employed ammonium sulfate
(AS 75%) precipitation procedure followed by dialysis steps yielded about 16 - 22 mg AFP·l-1 culture supernatant
with general mean of 18.67 ± 1.98 mg·l-1. The lost amount of AFP during purification
using AS and 3KDa cut-off dialysis membrane is about 50% thus, purification
procedure must be further improved. Indeed, concluded results from MIC and hyphal extension inhibition test noticed that AFP was
efficiently affected by either growth or hyphae form of A. alternata in vitro. The MIC of AFP against A. alternata was 2μg·ml-1. However,short, thick and highly septated hyphae with damaged constricted apical
regions extruding from condensed mycelium aggregates in treated hyphae compared
to the untreated culture was remarkably shown. The mode of action of in vitro experiment manifested that AFP
was effective to act the fungal cell and permeabilize the cell membrane of A. alternata. Furthermore, the in vivo experiment showed that AFP could reduce
post-harvest decay on banana caused by A.
alternata. AFP at concentration of 15 and 25 μg·ml-1 exhibit Alternaria decayed reduction by 45.45% and 77.27%, respectively.
While no Alternaria decayed area was observed when 50 μg·ml-1 was applied during the storage time. Quantification of DNA by
species-specific PCR could exude a positive correlation between the DNA
amount and decayed area. In conclusion, AFP can be efficiently used as a
bio-preservative agent during storage and handling of banana fruits, and
considered as an excellent biological alternative to combat secondary growth of
filamentous fungi.