Bioprospecting for entomopathogenic fungi against a foliar citrus pest
- Authors: Boon, Erin Ashley
- Date: 2025-04-02
- Subjects: Citrus Diseases and pests South Africa , Entomopathogenic fungi , Pests Integrated control , Biological assay , Cryptophlebia leucotreta , Ultraviolet radiation Physiological effect
- Language: English
- Type: Academic theses , Master's theses , text
- Identifier: http://hdl.handle.net/10962/478991 , vital:78247
- Description: 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. , Thesis (MSc) -- Faculty of Science, Zoology and Entomology, 2025
- Full Text:
- Authors: Boon, Erin Ashley
- Date: 2025-04-02
- Subjects: Citrus Diseases and pests South Africa , Entomopathogenic fungi , Pests Integrated control , Biological assay , Cryptophlebia leucotreta , Ultraviolet radiation Physiological effect
- Language: English
- Type: Academic theses , Master's theses , text
- Identifier: http://hdl.handle.net/10962/478991 , vital:78247
- Description: 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. , Thesis (MSc) -- Faculty of Science, Zoology and Entomology, 2025
- Full Text:
Potential Synergism between Entomopathogenic Fungi and Entomopathogenic Nematodes for the control of false codling moth (Thaumatotibia leucotreta)
- Authors: Prinsloo, Samantha Lee
- Date: 2021-10
- Subjects: Cryptophlebia leucotreta , Entomopathogenic fungi , Insect nematodes , Citrus Diseases and pests , Cryptophlebia leucotreta Biological control , Pests Integrated control , Biological pest control agents
- Language: English
- Type: Masters theses , text
- Identifier: http://hdl.handle.net/10962/188832 , vital:44790
- Description: False codling moth, Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae) (FCM), is a major phytosanitary pest of citrus in South Africa. Sufficient control measures for the soil-dwelling life stages of FCM have yet to be identified and owing to restrictions on the use of insecticides, non-chemical control options have been investigated including the use of entomopathogenic fungi (EPF) and entomopathogenic nematodes (EPN). Laboratory and field trials on an indigenous EPF, Metarhizium anisopliae FCM Ar 23 B3, have shown that this isolate is capable of inducing mortality in FCM soil-dwelling life stages. Other agents that have been highlighted as potential controls for soil-dwelling FCM life stages are the EPN species Steinernema yirgalemense 157-C, S. jeffreyense J194 and H. noenieputensis 158-C. This study conducted laboratory bioassays to assess the virulence of these four control agents on fifth instar FCM, in 24-well plates. These results reaffirmed the virulence of the four microbial control agents at their recommended doses of 50 IJs (EPN) and 1×107 conidia/ml (EPF) against fifth instar FCM with 80 to 96% larval mortality recorded. The EPF isolate exhibited the lowest mortality whilst S. yirgalemense induced the greatest mortality. In addition, the lethal concentration (LC) values for each isolate were determined using dose response bioassays. These values were previously unknown for all EPN species and for the EPF isolate based on the methodology used in this study. The LC50 results in order from lowest to highest EPN IJ concentration requirements were 4.38 IJs (S. yirgalemense), 4.47 IJs (S. jeffreyense) and 7.11 IJs (H. noenieputensis). The EPF isolate exhibited an LC50 of 3.42×105 conidia/ml. Lastly, research has shown that the combination of two control agents may increase control of late instar lepidopteran and coleopteran larvae, through synergistic interactions. Thus, the interactions that occurred between the combination of these EPN species with the EPF isolate were determined. This study found that when all three EPN species were combined simultaneously and sequentially with the EPF isolate M. anisopliae FCM AR 23 B3, additive interactions took place with exception of the simultaneous application of S. yirgalemense and H. noenieputensis, with the EPF and S. jeffreyense applied 24 h post EPF application. For the former, a synergistic interaction was found, whilst for the latter two, an antagonistic interaction. Although no strongly synergistic interactions were observed, additive interactions have been shown to reach a synergistic level when certain parameters are changed. Moving forward, a uniform methodology for conducting EPF/EPN interaction experiments has been suggested. It has also been recommended that due to the additive interactions observed in this study, laboratory soil-bioassays and field trials should be carried out for all three EPN species in combination with the EPF isolate. This research will inevitably facilitate the constant knowledge into management strategies for the phytosanitary pest, FCM in South African citrus. , Thesis (MSc) -- Science, Zoology and Entomology, 2021
- Full Text:
- Authors: Prinsloo, Samantha Lee
- Date: 2021-10
- Subjects: Cryptophlebia leucotreta , Entomopathogenic fungi , Insect nematodes , Citrus Diseases and pests , Cryptophlebia leucotreta Biological control , Pests Integrated control , Biological pest control agents
- Language: English
- Type: Masters theses , text
- Identifier: http://hdl.handle.net/10962/188832 , vital:44790
- Description: False codling moth, Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae) (FCM), is a major phytosanitary pest of citrus in South Africa. Sufficient control measures for the soil-dwelling life stages of FCM have yet to be identified and owing to restrictions on the use of insecticides, non-chemical control options have been investigated including the use of entomopathogenic fungi (EPF) and entomopathogenic nematodes (EPN). Laboratory and field trials on an indigenous EPF, Metarhizium anisopliae FCM Ar 23 B3, have shown that this isolate is capable of inducing mortality in FCM soil-dwelling life stages. Other agents that have been highlighted as potential controls for soil-dwelling FCM life stages are the EPN species Steinernema yirgalemense 157-C, S. jeffreyense J194 and H. noenieputensis 158-C. This study conducted laboratory bioassays to assess the virulence of these four control agents on fifth instar FCM, in 24-well plates. These results reaffirmed the virulence of the four microbial control agents at their recommended doses of 50 IJs (EPN) and 1×107 conidia/ml (EPF) against fifth instar FCM with 80 to 96% larval mortality recorded. The EPF isolate exhibited the lowest mortality whilst S. yirgalemense induced the greatest mortality. In addition, the lethal concentration (LC) values for each isolate were determined using dose response bioassays. These values were previously unknown for all EPN species and for the EPF isolate based on the methodology used in this study. The LC50 results in order from lowest to highest EPN IJ concentration requirements were 4.38 IJs (S. yirgalemense), 4.47 IJs (S. jeffreyense) and 7.11 IJs (H. noenieputensis). The EPF isolate exhibited an LC50 of 3.42×105 conidia/ml. Lastly, research has shown that the combination of two control agents may increase control of late instar lepidopteran and coleopteran larvae, through synergistic interactions. Thus, the interactions that occurred between the combination of these EPN species with the EPF isolate were determined. This study found that when all three EPN species were combined simultaneously and sequentially with the EPF isolate M. anisopliae FCM AR 23 B3, additive interactions took place with exception of the simultaneous application of S. yirgalemense and H. noenieputensis, with the EPF and S. jeffreyense applied 24 h post EPF application. For the former, a synergistic interaction was found, whilst for the latter two, an antagonistic interaction. Although no strongly synergistic interactions were observed, additive interactions have been shown to reach a synergistic level when certain parameters are changed. Moving forward, a uniform methodology for conducting EPF/EPN interaction experiments has been suggested. It has also been recommended that due to the additive interactions observed in this study, laboratory soil-bioassays and field trials should be carried out for all three EPN species in combination with the EPF isolate. This research will inevitably facilitate the constant knowledge into management strategies for the phytosanitary pest, FCM in South African citrus. , Thesis (MSc) -- Science, Zoology and Entomology, 2021
- Full Text:
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