Bioprospecting for entomopathogenic fungi against a foliar citrus pest

Abstract

Historically, pest management was highly dependent on the use of chemical insecticides for the control of agriculturally important pests. However, more recently, key export markets have imposed stringent chemical residue restrictions for citrus export. This deterring factor for chemical use has been coupled with the fact that these economically important pests are experiencing insecticidal resistance. As a result, the use of entomopathogenic fungi (EPF) has been explored as a complementary control option in integrated pest management (IPM) regimes. Previous research in South Africa identified several strains of Beauveria bassiana and Metarhizium spp. (including isolate M. pinghaense FCM Ar 23 B3). Laboratory bioassays evaluating the virulence of these isolates against major pests such as the false codling moth (FCM) (Thaumatotibia leucotreta, Lepidoptera: Tortricidae), citrus thrips (Scirtothrips aurantii, Thysanoptera: Thripidae), and citrus mealybugs (Planococcus citri, Hemiptera: Pseudococcidae) highlighted the potential of these EPF. While field trials targeting FCM with soil-applied treatments yielded encouraging results, foliar applications aimed at controlling citrus thrips and mealybugs showed limited success. These findings highlighted the need to assess the biological traits of the recovered isolates. Varying temperature ranges and humidity levels were found to not hinder the isolates' efficacy in the field. Conidial inactivation induced by ultraviolet (UV) radiation however, was. As these strains were recovered from the soil environment, it stood to reason that EPF isolates recovered from the foliar environment may be more suited for foliar application. Thus, bioprospecting for isolates from the aboveground environment was initiated and was the focal point of this thesis. Following the isolation and identification, the pathogenic ability and virulence, as well as the UV tolerance of these novel strains were established. Of the isolates recovered from the aboveground environment and identified using morphological and molecular techniques, four were B. bassiana (Px LM 4, Ha LM 11, Ha LM 12, Coe 18), one M. anisopliae (Hu LM 14), one Fusarium oxysporum (Pc HV 9), and one Geotrichum candidum yeast (Ha LM 2). The majority were isolated from insect cadavers, but one (Coe 18) was isolated as a foliar endophyte from an organically managed citrus farm in the Eastern Cape. Using standard protocols and conidial doses, the virulence of the recovered isolates was established against a common foliar pest of citrus, citrus mealybug. Isolate FCM Ar 23 B3 was included as a comparative control in this study as the virulence against citrus mealybug has previously been established. The initial screening of the isolates ranged between 15 and 90 % mortality. Isolates Px LM 4 and FCM Ar 23 B3 both induced an average mortality of 90 %. Isolates Ha LM 11, Ha LM 12, Hu LM 14, and Coe 18 caused mortalities greater than 60 % and were further investigated under dose-response assays. Of the six isolates measured for LC50, FCM Ar 23 B3 was the most virulent (5.25 × 105 conidia/ml), followed by Px LM 4 (1.09 × 106 conidia/ml) and Hu LM 14 (1.32 × 106 conidia/ml). The UV susceptibility to simulated sunlight of the six most virulent isolates was investigated. Whilst UV radiation certainly delayed the conidial germination of all the isolates, all the strains isolated from the aboveground environment demonstrated significant initial tolerance to UV radiation compared to the most virulent M. pinghaense FCM Ar 23 B3, which was recovered from the soil environment. Even though the B. bassiana Coe 18, which was recovered as an endophytic EPF, was not the most virulent, it stood out with strong initial UV tolerance and sustained a relatively high germination rate over time, establishing it as the most UV-tolerant isolate. Although formulation for development as a microbial biocontrol programme should not be overlooked for these isolates, the initial UV and sustained tolerance demonstrated by these aboveground isolates warrants further investigation under field conditions.