Studies on existing and new isolates of Cryptophlebia leucotreta granulovirus (CrleGV) on Thaumatotibia leucotreta populations from a range of geographic regions in South Africa
- Authors: Opoku-Debrah, John Kwadwo
- Date: 2012
- Subjects: Cryptophlebia leucotreta Cryptophlebia leucotreta -- South Africa Cryptophlebia leucotreta -- Biological control Cryptophlebia leucotreta -- Life cycles Baculoviruses Lepidoptera -- Biological control Tortricidae -- Biological control Microbial insecticides Pests -- Integrated control
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:5778 , http://hdl.handle.net/10962/d1005466
- Description: Baculoviruses are arthropod-specific DNA viruses that are highly virulent to most lepidopteran insects. Their host specificity and compatibility with IPM programmes has enabled their usage as safe microbial insecticides (biopesticides). Two baculovirus-based biopesticides, Cryptogran and Cryptex, which have been formulated with Cryptophlebia leucotreta granulovirus (CrleGV) have been registered for the control of false codling moth (FCM), Thaumatotibia (=Cryptophlebia) leucotreta (Meyrick) (Lepidoptera: Tortricidae) in South Africa and have been successfully incorporated into IPM programmes. However, several studies have indicated that insects can develop resistance to baculovirus-based biopesticide as was shown with field populations of codling moth (CM), Cydia pomonella (L.), which developed resistance to the biopesticide Cydia pomonella granulovirus (CpGV-M) in Europe. Other studies have shown that, under laboratory conditions, FCM populations differ in their susceptibility to Cryptogran and Cryptex. In order to investigate difference in susceptibility as well as protect against any future resistance by FCM to Cryptogran and Cryptex, a search for novel CrleGV-SA isolates from diseased insects from different geographic regions in South Africa was performed. Six geographic populations (Addo, Citrusdal, Marble Hall, Nelspruit, Baths and Mixed colonies) of FCM were established and maintained in the laboratory. Studies on the comparative biological performance based on pupal mass, female fecundity, egg hatch, pupal survival, adult eclosion and duration of life cycle of the Addo, Citrusdal, Marble Hall, Nelspruit and Mixed colonies revealed a low biological performance for the Citrusdal colony. This was attributed to the fact that FCM populations found in the Citrusdal area are not indigenous and may have been introduced from a very limited gene pool from another region. When insects from five colonies, excluding the Baths colony, were subjected to stress by overcrowding , a latent baculovirus resident in the Addo, Nelspruit, Citrusdal, Marble Hall and Mixed colonies was brought into an overt lethal state. Transmission electron micrographs revealed the presence of GV occlusion bodies (OBs) in diseased insects. DNA profiles obtained by single restriction endonuclease analysis of viral genomic DNA using BamH 1, Sa/1, Xba1 , Pst1, Xh01 , Kpn1, Hindlll and EcoR1 revealed five CrleGV-SA isolates latent within the insect populations. The new isolates were named CrleGV-SA Ado, CrleGV-SA Cit, CrleGV-SA Mbl, CrleGVSA Nels and CrleGV-SA Mix isolates. The novelty of the five CrleGV-SA isolates was confirmed by the presence of unique submolar bands, indicating that each isolate was genetically different. PCR amplification and sequencing of the granulin and egt genes from the five isolates revealed several single nucleotide polymorph isms (SNPs) which, in some cases, resulted in amino acid substitutions. DNA profiles from RFLPs, as well as phylogenetic analysis based on granulin and egt sequencing showed the presence of two CrleGV-SA genome types for the CrleGV-SA isolate. Cryptex and CrleGV-SA Ado, CrleGV-SA Cit, CrleGV-SA Mbl and CrleGV-SA Mix were placed as members of Group one CrleGV-SA, and Cryptogran and CrleGV-SA Nels isolate were placed into Group two CrleGV-SA. In droplet feeding bioassays, the median survival time (STso) for neonate larvae inoculated with Group one and two CrleGV-SA were determined to range from 80 - 88 hours (3.33 - 3.67 days), for all five colonies. LDso values for Group one and two CrleGV-SA against neonates from the Addo, Citrusdal, Marble Hall, Nelspruit and Mixed colonies varied between some populations and ranged from 0.80 - 3.12 OBs per larva, indicating some level of variation in host susceptibility. This is the first study reporting the existence of genetically distinct CrleGV baculovirus isolates infecting FCM in different geographical areas of South Africa. The results of this study have broad-ranging implications for our understanding of baculovirus-host interactions and for the application of baculovirus basedbiopesticides.
- Full Text:
- Date Issued: 2012
- Authors: Opoku-Debrah, John Kwadwo
- Date: 2012
- Subjects: Cryptophlebia leucotreta Cryptophlebia leucotreta -- South Africa Cryptophlebia leucotreta -- Biological control Cryptophlebia leucotreta -- Life cycles Baculoviruses Lepidoptera -- Biological control Tortricidae -- Biological control Microbial insecticides Pests -- Integrated control
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:5778 , http://hdl.handle.net/10962/d1005466
- Description: Baculoviruses are arthropod-specific DNA viruses that are highly virulent to most lepidopteran insects. Their host specificity and compatibility with IPM programmes has enabled their usage as safe microbial insecticides (biopesticides). Two baculovirus-based biopesticides, Cryptogran and Cryptex, which have been formulated with Cryptophlebia leucotreta granulovirus (CrleGV) have been registered for the control of false codling moth (FCM), Thaumatotibia (=Cryptophlebia) leucotreta (Meyrick) (Lepidoptera: Tortricidae) in South Africa and have been successfully incorporated into IPM programmes. However, several studies have indicated that insects can develop resistance to baculovirus-based biopesticide as was shown with field populations of codling moth (CM), Cydia pomonella (L.), which developed resistance to the biopesticide Cydia pomonella granulovirus (CpGV-M) in Europe. Other studies have shown that, under laboratory conditions, FCM populations differ in their susceptibility to Cryptogran and Cryptex. In order to investigate difference in susceptibility as well as protect against any future resistance by FCM to Cryptogran and Cryptex, a search for novel CrleGV-SA isolates from diseased insects from different geographic regions in South Africa was performed. Six geographic populations (Addo, Citrusdal, Marble Hall, Nelspruit, Baths and Mixed colonies) of FCM were established and maintained in the laboratory. Studies on the comparative biological performance based on pupal mass, female fecundity, egg hatch, pupal survival, adult eclosion and duration of life cycle of the Addo, Citrusdal, Marble Hall, Nelspruit and Mixed colonies revealed a low biological performance for the Citrusdal colony. This was attributed to the fact that FCM populations found in the Citrusdal area are not indigenous and may have been introduced from a very limited gene pool from another region. When insects from five colonies, excluding the Baths colony, were subjected to stress by overcrowding , a latent baculovirus resident in the Addo, Nelspruit, Citrusdal, Marble Hall and Mixed colonies was brought into an overt lethal state. Transmission electron micrographs revealed the presence of GV occlusion bodies (OBs) in diseased insects. DNA profiles obtained by single restriction endonuclease analysis of viral genomic DNA using BamH 1, Sa/1, Xba1 , Pst1, Xh01 , Kpn1, Hindlll and EcoR1 revealed five CrleGV-SA isolates latent within the insect populations. The new isolates were named CrleGV-SA Ado, CrleGV-SA Cit, CrleGV-SA Mbl, CrleGVSA Nels and CrleGV-SA Mix isolates. The novelty of the five CrleGV-SA isolates was confirmed by the presence of unique submolar bands, indicating that each isolate was genetically different. PCR amplification and sequencing of the granulin and egt genes from the five isolates revealed several single nucleotide polymorph isms (SNPs) which, in some cases, resulted in amino acid substitutions. DNA profiles from RFLPs, as well as phylogenetic analysis based on granulin and egt sequencing showed the presence of two CrleGV-SA genome types for the CrleGV-SA isolate. Cryptex and CrleGV-SA Ado, CrleGV-SA Cit, CrleGV-SA Mbl and CrleGV-SA Mix were placed as members of Group one CrleGV-SA, and Cryptogran and CrleGV-SA Nels isolate were placed into Group two CrleGV-SA. In droplet feeding bioassays, the median survival time (STso) for neonate larvae inoculated with Group one and two CrleGV-SA were determined to range from 80 - 88 hours (3.33 - 3.67 days), for all five colonies. LDso values for Group one and two CrleGV-SA against neonates from the Addo, Citrusdal, Marble Hall, Nelspruit and Mixed colonies varied between some populations and ranged from 0.80 - 3.12 OBs per larva, indicating some level of variation in host susceptibility. This is the first study reporting the existence of genetically distinct CrleGV baculovirus isolates infecting FCM in different geographical areas of South Africa. The results of this study have broad-ranging implications for our understanding of baculovirus-host interactions and for the application of baculovirus basedbiopesticides.
- Full Text:
- Date Issued: 2012
Geographic variation in the susceptibility of false colding Moth, Thaumatotibia Leucotreta, populations to a granulovirus (CrleGV-SA)
- Authors: Opoku-Debrah, John Kwadwo
- Date: 2008
- Subjects: Cryptophlebia leucotreta , Cryptophlebia leucotreta -- Control , Pests -- Biological control , Citrus -- Diseases and pests
- Language: English
- Type: Thesis , Masters , MTech
- Identifier: vital:10586 , http://hdl.handle.net/10948/984 , Cryptophlebia leucotreta , Cryptophlebia leucotreta -- Control , Pests -- Biological control , Citrus -- Diseases and pests
- Description: The false codling moth (FCM), Thaumatotibia (=Cryptophlebia) leucotreta (Meyrick) (Lepidoptera: Tortricidae) is a serious pest of citrus and other crops in Sub-Saharan Africa. The introduction of the Cryptophlebia leucotreta granulovirus (CrleGV-SA) Cryptogran and Cryptex (biopesticides) has proven to be very effective in the control of FCM. However, markedly lower susceptibility of some codling moth (CM), Cydia pomonella (L.) populations to Cydia pomonella granulovirus (CpGV-M), another granulovirus product used in the control of CM’s in Europe have been reported. Genetic differences between FCM populations in South Africa have also been established. It is therefore possible that differences in the susceptibility of these geographically distinct FCM populations to CrleGV-SA might also exist. To investigate this phenomenon, a benchmark for pathogenecity was established. In continuation of previous work with Cryptogran against the 1st and 5th instar FCM larvae, dose-response relationships were established for all five larval instars of FCM. In surface dose-response bioassays, the LC50 values for the 2nd, 3rd and 4th instars were calculated to be 4.516 x 104, 1.662 x 105 and 2.205 x 106 occlusion bodies (OBs)/ml, respectively. The LC90 values for the 2nd, 3rd and 4th instars were calculated to be 4.287 x 106, 9.992 x 106 and 1.661 x 108 OBs/ml, respectively. Susceptibility to CrleGV-SA was found to decline with larval stage and increase with time of exposure. The protocol was used in guiding bioassays with field collected FCM larvae. Laboratory assays conducted with Cryptogran (at 1.661 x 108 OBs/ml) against field collected FCM larvae from Addo, Kirkwood, Citrusdal and Clanwilliam as well as a standard laboratory colony, showed a significant difference in pathogenecity in only one case. This significant difference was observed between 5th instars from the Addo colony and 5th instars from the other populations. Four geographically distinct FCM colonies from Addo, Citrusdal, Marble Hall and Nelspruit were also established. Since Cryptogran and Cryptex are always targeted against 1st instar FCM larvae in the field, further comparative laboratory assays were conducted with the Addo colony and an old laboratory colony. Cryptogran was significantly more pathogenic than Cryptex against both the Addo and the old colony. However, a high level of heterogeneity was observed in responses within each population.
- Full Text:
- Date Issued: 2008
- Authors: Opoku-Debrah, John Kwadwo
- Date: 2008
- Subjects: Cryptophlebia leucotreta , Cryptophlebia leucotreta -- Control , Pests -- Biological control , Citrus -- Diseases and pests
- Language: English
- Type: Thesis , Masters , MTech
- Identifier: vital:10586 , http://hdl.handle.net/10948/984 , Cryptophlebia leucotreta , Cryptophlebia leucotreta -- Control , Pests -- Biological control , Citrus -- Diseases and pests
- Description: The false codling moth (FCM), Thaumatotibia (=Cryptophlebia) leucotreta (Meyrick) (Lepidoptera: Tortricidae) is a serious pest of citrus and other crops in Sub-Saharan Africa. The introduction of the Cryptophlebia leucotreta granulovirus (CrleGV-SA) Cryptogran and Cryptex (biopesticides) has proven to be very effective in the control of FCM. However, markedly lower susceptibility of some codling moth (CM), Cydia pomonella (L.) populations to Cydia pomonella granulovirus (CpGV-M), another granulovirus product used in the control of CM’s in Europe have been reported. Genetic differences between FCM populations in South Africa have also been established. It is therefore possible that differences in the susceptibility of these geographically distinct FCM populations to CrleGV-SA might also exist. To investigate this phenomenon, a benchmark for pathogenecity was established. In continuation of previous work with Cryptogran against the 1st and 5th instar FCM larvae, dose-response relationships were established for all five larval instars of FCM. In surface dose-response bioassays, the LC50 values for the 2nd, 3rd and 4th instars were calculated to be 4.516 x 104, 1.662 x 105 and 2.205 x 106 occlusion bodies (OBs)/ml, respectively. The LC90 values for the 2nd, 3rd and 4th instars were calculated to be 4.287 x 106, 9.992 x 106 and 1.661 x 108 OBs/ml, respectively. Susceptibility to CrleGV-SA was found to decline with larval stage and increase with time of exposure. The protocol was used in guiding bioassays with field collected FCM larvae. Laboratory assays conducted with Cryptogran (at 1.661 x 108 OBs/ml) against field collected FCM larvae from Addo, Kirkwood, Citrusdal and Clanwilliam as well as a standard laboratory colony, showed a significant difference in pathogenecity in only one case. This significant difference was observed between 5th instars from the Addo colony and 5th instars from the other populations. Four geographically distinct FCM colonies from Addo, Citrusdal, Marble Hall and Nelspruit were also established. Since Cryptogran and Cryptex are always targeted against 1st instar FCM larvae in the field, further comparative laboratory assays were conducted with the Addo colony and an old laboratory colony. Cryptogran was significantly more pathogenic than Cryptex against both the Addo and the old colony. However, a high level of heterogeneity was observed in responses within each population.
- Full Text:
- Date Issued: 2008
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