Expression of Pectinase Activity Among Aspergillus Flavus Isolates from Southwestern and Southeastern United States

Abstract

Aspergillus flavus is a widely distributed filamentous fungus that contaminates crops with the potent carcinogen aflatoxin. This species can be divided into S and L strains on the basis of sclerotial morphology. During crop infection, A. flavus can secrete a large array of hydrolytic enzymes. These include pectinase, which aids fungal spread through plant tissues. A survey of pectinase expression by soil isolates derived from different regions of the United States revealed geographic polymorphisms. Strain L isolates from Arizona produced moderate to high levels of a specific pectinase P2c, while S strain isolates produced variable amounts of P2c. In contrast, L strain isolates from southeastern U.S. yielded variable P2c production, while S strain isolates consistently expressed high P2c levels. These results were corroborated by pectinase surveys of additional collections of A. flavus from soil and cottonseed. Expression patterns for P2c and pectinmethylesterase were evaluated for a select number of isolates using an isoelectric focusing technique. Clear zone reactions from the pectinase plate assay corresponded to the presence of P2c, while red ring reactions corresponded to the lack of P2c. Commercial cottonseed infected by S strain isolates frequently contained aflatoxin, even when infected by S strain isolates that did not produce pectinase P2c. Thus, although P2c-lacking isolates have reduced invasiveness, these isolates still have sufficient pathogenicity to cause aflatoxin contamination.

Etiology and management of aflatoxin contamination.

“…This adaptation can be seen in the size and production habit of sclerotia, the variation in hydrolase production Mellon and Cotty, 2004), the distribution by soil type (JaimeGarcia and Cotty, 2006b), and even in seasonality ( Fig. 1; Orum et al, 1997;Bock et al, 2004).…”

“…The divergence in aflatoxin-producing ability is striking, as some isolates produce much higher levels of aflatoxins than others. It is not unusual to find crop components that range in aflatoxin content from < 10 ng/g to several million ng/g (Lee et al, 1990;Mellon and Cotty, 2004). This range partially reflects the diversity present amongst the infecting fungi.…”

Plant cell wall degrading enzymes, pectinase and cellulase, in the digestive system of the red palm weevil, Rhynchophorus ferrugineus (Coleoptera: Curculionidae)

“…After electrophoresis, the gels were washed in citrate buffer (pH 6) containing 2.5% (v/v) Triton X-100 for 45 min and then incubated in citrate buffer (pH 6) for 60 min with gentle agitation. The gels for visualising pectinase were stained with ruthenium red (0.03%) and the bands of pectinase activity appeared as clear areas in the field of the red background of the gel (Mellon & Cotty 2004). In the case of cellulase, the gels were stained with Congo red (0.1%) for 10-15 min at room temperature.…”

Screening and Molecular Identification of New Microbial Strains for Production of Enzymes of Biotechnological Interest

“…In a secondary screening, colonies of different isolates showing zone of clearance on pectin-nutrient agar medium were specified to be the pectinase microorganisms. The majority of researchers have assayed the pectinase activity qualitatively by spotting the isolates on pectin containing culture plates and then analyzing the pectin digestion zone (Mellon and Cotty 2004). The spotting of these xylanopectinolytic isolates on nutrient-agar medium containing LBG gave a zone of hydrolysis.…”