Selection for improved virulence of Cryptophlebia peltastica nucleopolyhedrovirus (CrpeNPV) to False Codling Moth, Thaumatotibia leucotreta, by serial passage through a heterologous host
- Authors: Iita, Petrus Paulus
- Date: 2021-04
- Subjects: Cryptophlebia leucotreta -- Biological control , Biological pest control agents , Citrus -- Diseases and pests , Baculoviruses , Cryptophlebia peltastica nucleopolyhedrovirus (CrpeNPV)
- Language: English
- Type: thesis , text , Masters , MSc
- Identifier: http://hdl.handle.net/10962/178180 , vital:42918
- Description: The false codling moth (FCM), Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae) is endemic to southern Africa, and strongly associated with citrus. As South African citrus production is mainly for export to foreign markets, the market access risk due to the phytosanitary status of this pest is considerable and its control is therefore imperative. Various control measures as part of a rigorous integrated pest management (IPM) programme targeted against T. leucotreta have been effective at suppressing the pest in citrus, but there is still a growing need for continued improvement of the programme and augmentation of the available control options. Of these control options, biological control, particularly the use of Cryptophlebia leucotreta granulovirus (CrleGV-SA), is a key component of IPM in citrus orchards and it has been very successful at reducing T. leucotreta populations in the field for almost two decades. There is however, a growing need for more baculovirus variants with an improved virulence against T. leucotreta for a more efficient pest management system. The newly identified insect virus, Cryptophlebia peltastica nucleopolyhedrovirus (CrpeNPV) offers a unique opportunity for an additional biopesticide in IPM for control of T. leucotreta in the field. This study aimed to conduct serial passaging of CrpeNPV through a heterologous host, T. leucotreta, in order to determine the potential for improved virulence or speed of kill against it. In order to select for a variant of CrpeNPV with improved virulence against T. leucotreta, a high dose (LC90) of the virus OBs was used to perform 12 serial passages through T. leucotreta larvae in surface-dose bioassays. Whole genome sequencing and analysis of the passaged virus, along with restriction endonuclease profiling in silico was performed to determine if the genetic identity of the virus had changed during serial passage, in relation to the original virus. These analyses indicated that the dominant genotype of CrpeNPV was maintained following 12 serial passages through the heterologous host. The biological activity of the passaged virus, along with the original virus was evaluated against neonate T. leucotreta in surface-dose bioassays and compared. Results from dose-response bioassays showed that the virulence of CrpeNPV did not improve after 12 serial passages. The LC50 values of the passaged virus and the original virus were estimated at 1.96 × 104 and 1.58 × 104 OBs/ml, respectively, whereas the LC90 values were estimated at 3.46 × 104 OBs/ml for the passaged virus and 3.68 × 104 for the original virus. Similarly, the results from time-response bioassays showed that the speed of kill of CrpeNPV did not improve after 12 serial passages. The LT50 values of the passaged virus and the original virus were 88.44 hours (3 days and 16 hours) and 83.74 hours (3 days and 12 hours), respectively, whereas the LT90 values were 115 hours (4 days 19 hours) for the passaged virus and 102 hours (4 days 6 hours) for the original virus. The virulence and speed of kill of the passaged virus decreased significantly, in relation to the original virus. When the full genome of the passaged virus was sequenced and analysed, only a few SNPs were detected in the viral genome, in comparison to the original virus. No detectable difference in REN digestion patterns were observed following REN analysis of gDNA of the passaged virus with several restriction enzymes in silico. The results for this study suggest that CrpeNPV may already be optimally suited to the heterologous host as it persists under these conditions without significant changes to the genome. These results have positive implications for the genetic integrity of CrpeNPV as a potential biocontrol agent in the field. This study is the first to report the virulence selection of CrpeNPV by serial passage through a heterologous host, and also the first to record bioassay data in terms of dose response (or lethal concentration) against T. leucotreta second instars. The data obtained have added to the knowledge about interactions between CrpeNPV and its heterologous host, and may be fundamental to continued investigation into the effect of serial passage on pathogenicity and genetic diversity of CrpeNPV. , Thesis (MSc) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
- Date Issued: 2021-04
- Authors: Iita, Petrus Paulus
- Date: 2021-04
- Subjects: Cryptophlebia leucotreta -- Biological control , Biological pest control agents , Citrus -- Diseases and pests , Baculoviruses , Cryptophlebia peltastica nucleopolyhedrovirus (CrpeNPV)
- Language: English
- Type: thesis , text , Masters , MSc
- Identifier: http://hdl.handle.net/10962/178180 , vital:42918
- Description: The false codling moth (FCM), Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae) is endemic to southern Africa, and strongly associated with citrus. As South African citrus production is mainly for export to foreign markets, the market access risk due to the phytosanitary status of this pest is considerable and its control is therefore imperative. Various control measures as part of a rigorous integrated pest management (IPM) programme targeted against T. leucotreta have been effective at suppressing the pest in citrus, but there is still a growing need for continued improvement of the programme and augmentation of the available control options. Of these control options, biological control, particularly the use of Cryptophlebia leucotreta granulovirus (CrleGV-SA), is a key component of IPM in citrus orchards and it has been very successful at reducing T. leucotreta populations in the field for almost two decades. There is however, a growing need for more baculovirus variants with an improved virulence against T. leucotreta for a more efficient pest management system. The newly identified insect virus, Cryptophlebia peltastica nucleopolyhedrovirus (CrpeNPV) offers a unique opportunity for an additional biopesticide in IPM for control of T. leucotreta in the field. This study aimed to conduct serial passaging of CrpeNPV through a heterologous host, T. leucotreta, in order to determine the potential for improved virulence or speed of kill against it. In order to select for a variant of CrpeNPV with improved virulence against T. leucotreta, a high dose (LC90) of the virus OBs was used to perform 12 serial passages through T. leucotreta larvae in surface-dose bioassays. Whole genome sequencing and analysis of the passaged virus, along with restriction endonuclease profiling in silico was performed to determine if the genetic identity of the virus had changed during serial passage, in relation to the original virus. These analyses indicated that the dominant genotype of CrpeNPV was maintained following 12 serial passages through the heterologous host. The biological activity of the passaged virus, along with the original virus was evaluated against neonate T. leucotreta in surface-dose bioassays and compared. Results from dose-response bioassays showed that the virulence of CrpeNPV did not improve after 12 serial passages. The LC50 values of the passaged virus and the original virus were estimated at 1.96 × 104 and 1.58 × 104 OBs/ml, respectively, whereas the LC90 values were estimated at 3.46 × 104 OBs/ml for the passaged virus and 3.68 × 104 for the original virus. Similarly, the results from time-response bioassays showed that the speed of kill of CrpeNPV did not improve after 12 serial passages. The LT50 values of the passaged virus and the original virus were 88.44 hours (3 days and 16 hours) and 83.74 hours (3 days and 12 hours), respectively, whereas the LT90 values were 115 hours (4 days 19 hours) for the passaged virus and 102 hours (4 days 6 hours) for the original virus. The virulence and speed of kill of the passaged virus decreased significantly, in relation to the original virus. When the full genome of the passaged virus was sequenced and analysed, only a few SNPs were detected in the viral genome, in comparison to the original virus. No detectable difference in REN digestion patterns were observed following REN analysis of gDNA of the passaged virus with several restriction enzymes in silico. The results for this study suggest that CrpeNPV may already be optimally suited to the heterologous host as it persists under these conditions without significant changes to the genome. These results have positive implications for the genetic integrity of CrpeNPV as a potential biocontrol agent in the field. This study is the first to report the virulence selection of CrpeNPV by serial passage through a heterologous host, and also the first to record bioassay data in terms of dose response (or lethal concentration) against T. leucotreta second instars. The data obtained have added to the knowledge about interactions between CrpeNPV and its heterologous host, and may be fundamental to continued investigation into the effect of serial passage on pathogenicity and genetic diversity of CrpeNPV. , Thesis (MSc) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
- Date Issued: 2021-04
Genetic and biological characterisation of a novel South African Cydia pomonella granulovirus (CpGV-SA) isolate
- Motsoeneng, Boitumelo Madika
- Authors: Motsoeneng, Boitumelo Madika
- Date: 2016
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:20503 , http://hdl.handle.net/10962/d1021266
- Description: The codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae), is the primary pest of pome fruit cultivated worldwide. The control of this insect pest has been dependent on the frequent use of broad-spectrum chemical pesticides, which has led to the development of resistance in pest populations and negative effects on human health and the environment. The Betabaculovirus of C. pomonella has successfully been applied as a biological control agent in integrated pest management (IPM) programmes for the suppression of pest populations worldwide. Previously, all Cydia pomonella granulovirus (CpGV) biopesticides were based on a Mexican isolate (CpGV-M) and although these products are highly efficient at controlling C. pomonella, resistance cases have been reported across Europe. The identification of novel CpGV isolates as additional or alternative control agents to manage resistance is therefore necessary. This study aimed to genetically and biologically characterise a novel South African C. pomonella granulovirus isolate and to test its virulence against neonate larvae. Based on the morphology of the occlusion bodies observed using transmission electron microscopy, granuloviruses were recovered from diseased and dead larvae collected from an orchard in South Africa where no virus applications had been made. DNA was extracted and the identification of the isolated granulovirus was achieved through the PCR amplification and sequencing of the lef-8, lef-9, granulin and egt genes. Submission of the gene sequences to BLAST revealed high percentage identities to sequences from various CpGV isolates, resulting in the naming of the isolate in this study as the South African Cydia pomonella granulovirus (CpGV-SA) isolate. Phylogenetic analysis based on the single nucleotide polymorphisms (SNPs) detected in the lef-8, lef-9 and granulin nucleotide sequences grouped the South African isolate with CpGV-E2 (genome type B) and CpGV-S (genome type E). The CpGV-SA isolate was further genetically characterised by restriction endonuclease analysis and complete sequencing of the genomic DNA. Differences were observed for the BamHI, EcoRI, PstI and XhoI profiles of CpGV-SA in comparison to the respective profiles generated for CpGV-M extracted from a biopesticide, Carpovirusine® (Arysta Lifescience, France). Several genetic variations between the complete genome sequence of CpGV-SA and the reference isolate, CpGV-M1, as well as a recent genome submission of CpGV-M, both representing genome type A were observed. The complete genome analysis confirmed that CpGV-SA is genetically different from the Mexican CpGV isolate, used in thedevelopment of most biopesticides. In silico restriction profiles of the genome sequence obtained for CpGV-SA and genome sequences of genetically different CpGV isolates originating from Mexico (M1 and M), England (E2), Canada (S) and Iran (I12 and I07), available on the NCBI’s GenBank database confirmed that CpGV-SA is of mixed genotypes. Furthermore, the South African isolate shared the single common difference found in the pe38 gene of resistance overcoming isolates, which was the absence of an internal 24 nucleotide repeat present in CpGV-M1. In addition to the common difference, SNPs detected in the pe38 gene grouped the isolate with the CpGV-S isolate, suggesting that the CpGV-SA isolate is predominantly of genome type E. To determine the biological activity of CpGV-SA against neonate C. pomonella larvae, surface bioassays were conducted alongside CpGV-M (Carpovirusine®) bioassays. The LC50 and LC90 values for the South African isolate were 1.6 × 103 and 1.2 × 105 OBs/ml respectively. The LT50 was determined to be 135 hours. These values were similar to the values obtained for CpGV-M (Carpovirusine®). The results in this study suggest that a novel South African CpGV isolate of mixed genotypes, potentially able to overcome resistance in C. pomonella, with biological activity similar to CpGV-M (Carpovirusine®) and important for the control of C. pomonella was recovered. The CpGV-SA isolate could therefore potentially be developed into a biopesticide for use in resistance management strategies against C. pomonella populations in South Africa.
- Full Text:
- Date Issued: 2016
- Authors: Motsoeneng, Boitumelo Madika
- Date: 2016
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:20503 , http://hdl.handle.net/10962/d1021266
- Description: The codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae), is the primary pest of pome fruit cultivated worldwide. The control of this insect pest has been dependent on the frequent use of broad-spectrum chemical pesticides, which has led to the development of resistance in pest populations and negative effects on human health and the environment. The Betabaculovirus of C. pomonella has successfully been applied as a biological control agent in integrated pest management (IPM) programmes for the suppression of pest populations worldwide. Previously, all Cydia pomonella granulovirus (CpGV) biopesticides were based on a Mexican isolate (CpGV-M) and although these products are highly efficient at controlling C. pomonella, resistance cases have been reported across Europe. The identification of novel CpGV isolates as additional or alternative control agents to manage resistance is therefore necessary. This study aimed to genetically and biologically characterise a novel South African C. pomonella granulovirus isolate and to test its virulence against neonate larvae. Based on the morphology of the occlusion bodies observed using transmission electron microscopy, granuloviruses were recovered from diseased and dead larvae collected from an orchard in South Africa where no virus applications had been made. DNA was extracted and the identification of the isolated granulovirus was achieved through the PCR amplification and sequencing of the lef-8, lef-9, granulin and egt genes. Submission of the gene sequences to BLAST revealed high percentage identities to sequences from various CpGV isolates, resulting in the naming of the isolate in this study as the South African Cydia pomonella granulovirus (CpGV-SA) isolate. Phylogenetic analysis based on the single nucleotide polymorphisms (SNPs) detected in the lef-8, lef-9 and granulin nucleotide sequences grouped the South African isolate with CpGV-E2 (genome type B) and CpGV-S (genome type E). The CpGV-SA isolate was further genetically characterised by restriction endonuclease analysis and complete sequencing of the genomic DNA. Differences were observed for the BamHI, EcoRI, PstI and XhoI profiles of CpGV-SA in comparison to the respective profiles generated for CpGV-M extracted from a biopesticide, Carpovirusine® (Arysta Lifescience, France). Several genetic variations between the complete genome sequence of CpGV-SA and the reference isolate, CpGV-M1, as well as a recent genome submission of CpGV-M, both representing genome type A were observed. The complete genome analysis confirmed that CpGV-SA is genetically different from the Mexican CpGV isolate, used in thedevelopment of most biopesticides. In silico restriction profiles of the genome sequence obtained for CpGV-SA and genome sequences of genetically different CpGV isolates originating from Mexico (M1 and M), England (E2), Canada (S) and Iran (I12 and I07), available on the NCBI’s GenBank database confirmed that CpGV-SA is of mixed genotypes. Furthermore, the South African isolate shared the single common difference found in the pe38 gene of resistance overcoming isolates, which was the absence of an internal 24 nucleotide repeat present in CpGV-M1. In addition to the common difference, SNPs detected in the pe38 gene grouped the isolate with the CpGV-S isolate, suggesting that the CpGV-SA isolate is predominantly of genome type E. To determine the biological activity of CpGV-SA against neonate C. pomonella larvae, surface bioassays were conducted alongside CpGV-M (Carpovirusine®) bioassays. The LC50 and LC90 values for the South African isolate were 1.6 × 103 and 1.2 × 105 OBs/ml respectively. The LT50 was determined to be 135 hours. These values were similar to the values obtained for CpGV-M (Carpovirusine®). The results in this study suggest that a novel South African CpGV isolate of mixed genotypes, potentially able to overcome resistance in C. pomonella, with biological activity similar to CpGV-M (Carpovirusine®) and important for the control of C. pomonella was recovered. The CpGV-SA isolate could therefore potentially be developed into a biopesticide for use in resistance management strategies against C. pomonella populations in South Africa.
- Full Text:
- Date Issued: 2016
Genetic and biological characterisation of a novel South African Plutella xylostella granulovirus (PlxyGV) isolate
- Authors: Abdulkadir, Fatima
- Date: 2014
- Subjects: Diamondback moth , Diamondback moth -- Control -- South Africa , Plutellidae -- Control -- South Africa , Baculoviruses , Cruciferae -- Diseases and pests -- South Africa
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4113 , http://hdl.handle.net/10962/d1013059
- Description: The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), is an important pest of cruciferous crops worldwide. The prolonged use of synthetic chemical insecticides as a primary means of control has resulted in the development of resistance in pest populations. In addition, the pest has also evolved resistance to the bacterial insecticidal protein of Bacillus thuringiensis which is also widely used as a method of control. Baculoviruses are considered as effective alternatives to conventional methods of control when incorporated into integrated pest management (IPM) programmes. These viruses target the larval stages of insects, are generally host-specific and are safe for use in the environment. This study aimed to isolate a baculovirus from a laboratory-reared P. xylostella colony, characterise it genetically and then evaluate its virulence against neonate and fourth instar larvae. A laboratory colony of P. xylostella was established using pupae and asymptomatic larvae collected from a cabbage plantation outside Grahamstown in the Eastern Cape province of South Africa. The colony flourished in the laboratory due to prime conditions and availability of food. The duration of development from egg to adult was determined by observation and imaging of the various life stages. The mean developmental time from egg to adult was observed to be 14.59 ± 0.21 days. The population of the insects increased rapidly in number leading to overcrowding of the insect colony, and hence appearance of larvae with viral symptoms. Occlusion bodies (OBs) were extracted from symptomatic larval cadavers and purified by glycerol gradient centrifugation. Analysis of the purified OBs by transmission electron microscopy revealed the presence of a granulovirus which was named PlxyGV-SA. The virus isolate was genetically characterised by restriction endonuclease analysis of the genomic DNA, and PCR amplification and sequencing of selected viral genes. The complete genome sequence of a Japanese P. xylostella granulovirus isolate, PlxyGV-Japan, has been deposited on the GenBank database providing a reference strain for comparison with DNA profiles and selected gene sequences of PlxyGV-SA. BLAST analysis of the granulin gene confirmed the isolation of a novel South African PlxyGV isolate. Comparison of the restriction profiles of PlxyGV-SA with profiles of PlxyGV-Japan and other documented PlxyGV profiles obtained by agarose gel electrophoresis revealed that PlxyGV-SA is a genetically distinct isolate. The data obtained from the sequencing and alignment of ecdysteroid UDP-glucosyltransferase (egt), late expression factor 8 (lef-8) and late expression factor 9 (lef-9) genes with those of PlxyGV-Japan also showed that PlxyGV-SA is a genetically different isolate. In order to determine the biological activity of PlxyGV-SA against neonate and fourth instar P. xylostella larvae, surface dose bioassays were conducted. The median lethal concentration of the virus required to kill 50% (LC₅₀) and 90% (LC₉₀) of the larvae was estimated by feeding insects with a range of doses. In addition, the time to kill 50% of the larvae (LT₅₀) was determined by feeding insects with the LC₉₀ concentration. Larval mortality was monitored daily until pupation. The data obtained from the dose response assays were subjected to probit analysis using Proban statistical software. The time response was determined using GraphPad Prism software (version 6.0). The LC₅₀ and LC₉₀ values for the neonate larvae were 3.56 × 10⁵ and 1.14 × 10⁷ OBs/ml respectively. The LT₅₀ was determined to be 104 hours. The neonate larvae were found to be more susceptible to infection than the fourth instar larvae with the same virus concentration. The concentrations used for the neonate larvae assay did not have a significant effect on the fourth instar as no mortality was recorded. This is the first study to describe a novel South African PlxyGV isolate and the results suggest that PlxyGV-SA has significant potential for development as an effective biopesticide for the control of P. xylostella in the field.
- Full Text:
- Date Issued: 2014
- Authors: Abdulkadir, Fatima
- Date: 2014
- Subjects: Diamondback moth , Diamondback moth -- Control -- South Africa , Plutellidae -- Control -- South Africa , Baculoviruses , Cruciferae -- Diseases and pests -- South Africa
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4113 , http://hdl.handle.net/10962/d1013059
- Description: The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), is an important pest of cruciferous crops worldwide. The prolonged use of synthetic chemical insecticides as a primary means of control has resulted in the development of resistance in pest populations. In addition, the pest has also evolved resistance to the bacterial insecticidal protein of Bacillus thuringiensis which is also widely used as a method of control. Baculoviruses are considered as effective alternatives to conventional methods of control when incorporated into integrated pest management (IPM) programmes. These viruses target the larval stages of insects, are generally host-specific and are safe for use in the environment. This study aimed to isolate a baculovirus from a laboratory-reared P. xylostella colony, characterise it genetically and then evaluate its virulence against neonate and fourth instar larvae. A laboratory colony of P. xylostella was established using pupae and asymptomatic larvae collected from a cabbage plantation outside Grahamstown in the Eastern Cape province of South Africa. The colony flourished in the laboratory due to prime conditions and availability of food. The duration of development from egg to adult was determined by observation and imaging of the various life stages. The mean developmental time from egg to adult was observed to be 14.59 ± 0.21 days. The population of the insects increased rapidly in number leading to overcrowding of the insect colony, and hence appearance of larvae with viral symptoms. Occlusion bodies (OBs) were extracted from symptomatic larval cadavers and purified by glycerol gradient centrifugation. Analysis of the purified OBs by transmission electron microscopy revealed the presence of a granulovirus which was named PlxyGV-SA. The virus isolate was genetically characterised by restriction endonuclease analysis of the genomic DNA, and PCR amplification and sequencing of selected viral genes. The complete genome sequence of a Japanese P. xylostella granulovirus isolate, PlxyGV-Japan, has been deposited on the GenBank database providing a reference strain for comparison with DNA profiles and selected gene sequences of PlxyGV-SA. BLAST analysis of the granulin gene confirmed the isolation of a novel South African PlxyGV isolate. Comparison of the restriction profiles of PlxyGV-SA with profiles of PlxyGV-Japan and other documented PlxyGV profiles obtained by agarose gel electrophoresis revealed that PlxyGV-SA is a genetically distinct isolate. The data obtained from the sequencing and alignment of ecdysteroid UDP-glucosyltransferase (egt), late expression factor 8 (lef-8) and late expression factor 9 (lef-9) genes with those of PlxyGV-Japan also showed that PlxyGV-SA is a genetically different isolate. In order to determine the biological activity of PlxyGV-SA against neonate and fourth instar P. xylostella larvae, surface dose bioassays were conducted. The median lethal concentration of the virus required to kill 50% (LC₅₀) and 90% (LC₉₀) of the larvae was estimated by feeding insects with a range of doses. In addition, the time to kill 50% of the larvae (LT₅₀) was determined by feeding insects with the LC₉₀ concentration. Larval mortality was monitored daily until pupation. The data obtained from the dose response assays were subjected to probit analysis using Proban statistical software. The time response was determined using GraphPad Prism software (version 6.0). The LC₅₀ and LC₉₀ values for the neonate larvae were 3.56 × 10⁵ and 1.14 × 10⁷ OBs/ml respectively. The LT₅₀ was determined to be 104 hours. The neonate larvae were found to be more susceptible to infection than the fourth instar larvae with the same virus concentration. The concentrations used for the neonate larvae assay did not have a significant effect on the fourth instar as no mortality was recorded. This is the first study to describe a novel South African PlxyGV isolate and the results suggest that PlxyGV-SA has significant potential for development as an effective biopesticide for the control of P. xylostella in the field.
- Full Text:
- Date Issued: 2014
Development of techniques for the isolation of a granulovirus from potato tuber moth, phthorimaea operculella (Zeller)
- Authors: King, Shirley Anne
- Date: 2011
- Subjects: Potato tuberworm -- Larvae , Agricultural pests -- Biological control , Potato tuberworm , Baculoviruses
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:5910 , http://hdl.handle.net/10962/d1015202
- Description: Phthorimaea operculella, commonly known as the Potato Tuber Moth, is an economically important agricultural pest worldwide. The baculovirus, Phthorimaea operculella granulovirus (PhoGV) has been considered as a means of control alternative to chemical control because of its host specificity and harmless impact on other organisms and ecosystems. An isolate of PhoGV obtained from a South African PTM population would be beneficial in the production of a biopesticide, which is not yet available. An efficient and cost-effective rearing method would be advantageous for potential commercial production. Commercial table and seed potato plantations and storage facilities located in Patensie, Bathurst, Howick and Ivanhoe were surveyed for PTM infestations. Patensie was the only site where milky discoloured larvae were found, a potential symptom of PhoGV infection. TEM analysis revealed no virus in these samples. Since no virus was found in the field-collected samples, PTM insects were collected to initiate rearing in the laboratory. PTM was raised by three different methods in the laboratory. A cost/benefit analysis, survival rate, fertility and sex ratio were recorded for each rearing method. Rearing method one was deemed unsuccessful for efficient commercial rearing, as survival percentage and fertility were low. Rearing methods two and three had high survival rates and high fertility, and were efficient and less labour intensive than rearing method one. Rearing method three was the most productive technique, but for commercial production rearing method two was considered the most manageable and efficient. The sex ratio was 1:1 for all three cultures. The cost analysis revealed that rearing methods two and three were less expensive than rearing method one because less labour was required to monitor insects. The success of rearing PTM for 19 months will enable these cultures to be up-scaled to a large production facility for mass rearing. Virus was not found in the field surveys or in laboratory cultures, therefore chemical, temperature, humidity and carbon dioxide stressors were used in an attempt to initiate a baculoviral infection. Symptoms were exhibited in larvae subjected to chemical, temperature and humidity treatments, but these were confirmed by TEM analysis not to be a result of PhoGV infection. The success of rearing PTM in the laboratory suggests that the method could be used in the commercial rearing of the insects in a large mass-rearing facility. The data obtained from induction protocols have allowed for better understanding for future induction for PhoGV and other baculoviruses in other insect species. The failure to isolate a South African PhoGV strain for developing a biopesticide against PTM has motivated further studies in obtaining a baculovirus in order for South Africa to develop a commercial product against this pest.
- Full Text:
- Date Issued: 2011
- Authors: King, Shirley Anne
- Date: 2011
- Subjects: Potato tuberworm -- Larvae , Agricultural pests -- Biological control , Potato tuberworm , Baculoviruses
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:5910 , http://hdl.handle.net/10962/d1015202
- Description: Phthorimaea operculella, commonly known as the Potato Tuber Moth, is an economically important agricultural pest worldwide. The baculovirus, Phthorimaea operculella granulovirus (PhoGV) has been considered as a means of control alternative to chemical control because of its host specificity and harmless impact on other organisms and ecosystems. An isolate of PhoGV obtained from a South African PTM population would be beneficial in the production of a biopesticide, which is not yet available. An efficient and cost-effective rearing method would be advantageous for potential commercial production. Commercial table and seed potato plantations and storage facilities located in Patensie, Bathurst, Howick and Ivanhoe were surveyed for PTM infestations. Patensie was the only site where milky discoloured larvae were found, a potential symptom of PhoGV infection. TEM analysis revealed no virus in these samples. Since no virus was found in the field-collected samples, PTM insects were collected to initiate rearing in the laboratory. PTM was raised by three different methods in the laboratory. A cost/benefit analysis, survival rate, fertility and sex ratio were recorded for each rearing method. Rearing method one was deemed unsuccessful for efficient commercial rearing, as survival percentage and fertility were low. Rearing methods two and three had high survival rates and high fertility, and were efficient and less labour intensive than rearing method one. Rearing method three was the most productive technique, but for commercial production rearing method two was considered the most manageable and efficient. The sex ratio was 1:1 for all three cultures. The cost analysis revealed that rearing methods two and three were less expensive than rearing method one because less labour was required to monitor insects. The success of rearing PTM for 19 months will enable these cultures to be up-scaled to a large production facility for mass rearing. Virus was not found in the field surveys or in laboratory cultures, therefore chemical, temperature, humidity and carbon dioxide stressors were used in an attempt to initiate a baculoviral infection. Symptoms were exhibited in larvae subjected to chemical, temperature and humidity treatments, but these were confirmed by TEM analysis not to be a result of PhoGV infection. The success of rearing PTM in the laboratory suggests that the method could be used in the commercial rearing of the insects in a large mass-rearing facility. The data obtained from induction protocols have allowed for better understanding for future induction for PhoGV and other baculoviruses in other insect species. The failure to isolate a South African PhoGV strain for developing a biopesticide against PTM has motivated further studies in obtaining a baculovirus in order for South Africa to develop a commercial product against this pest.
- Full Text:
- Date Issued: 2011
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