Diabrotica undecimpunctata virus 2, a Novel Small RNA Virus Discovered from Southern Corn Rootworm, Diabrotica undecimpunctata howardi Barber (Coleoptera: Chrysomelidae)

The genome of Diabrotica undecimpunctata virus 2 (DuV2), a positive-sense, single-stranded RNA virus identified from the southern corn root- worm transcriptome, comprises 5,313 nucleotides, including a short poly(A) tail. The two open reading frames encode a nonstructural polyprotein (p156) and a putative capsid protein (p25)

Genome Sequence of a Small RNA Virus of the Southern Corn Rootworm, Diabrotica undecimpunctata howardi Barber (Coleoptera: Chrysomelidae)

The genome sequence of a novel small RNA virus, tentatively named Diabrotica undecimpunctata virus 1 (DuV1), was discovered from the transcriptome of the southern corn rootworm, Diabrotica undecimpunctata howardi Barber. DuV1 has a positive-sense, single-stranded RNA genome that encodes a single polyprotein of 3,401 amino acids with limited similarity to other viruses

Avaliação de métodos para inoculação e diagnóstico de vírus de mosaico do pepino (CMV)

The Cucumber mosaic virus (CMV) is a limiting agent in the production and quality of agricultural crops; whose effective inoculation and reliable diagnosis are essential to implement suitable and timely management strategies. Two protocols for CMV inoculation in Nicotiana benthamiana and N. tabacum cv. Xanthi as indicator plants and different virus detection methods were evaluated. Infection of plants with CMV was achieved by mechanical transmission and infiltration. The presence of the virus was confirmed by expression of symptoms, serology (ImmunoStrip®), RT-PCR and by sequencing. The results confirmed the effectiveness of the mechanical transmission. The infected plants showed symptoms associated with the pathogen 15 days after inoculation. The serological test allowed detecting the presence of the pathogen only in N. benthamiana plants, while the RT-PCR test in all evaluated plants. The sequencing results showed high nucleotide identity values with the sequences reported in the NBCI for this virus. Mechanical transmission was the most effective method for CMV inoculation, and molecular analysis techniques were more sensitive and reliable for CMV identification, compared to serological test.El virus del mosaico del pepino (CMV) es un agente limitante en la producción y calidad de cultivos agrícolas; cuya inoculación efectiva y diagnóstico confiable son esenciales para implementar estrategias de manejo adecuadas y oportunas. Se evaluaron dos protocolos para la inoculación del CMV en plantas indicadoras (Nicotiana benthamiana y N. tabacum cv. Xanthi) y diferentes métodos de detección de este virus. La infección de plantas con CMV se logró mediante transmisión mecánica e infiltración. La presencia del virus se confirmó mediante la expresión de síntomas, serología (ImmunoStrip®), RT-PCR y por secuenciación. Los resultados confirmaron la efectividad del método de transmisión mecánica, en el cual, las plantas infectadas presentaron sintomatología asociada con el virus 15 días después de su inoculación. La prueba serológica permitió detectar la presencia del patógeno solo en las plantas de N. benthamiana, mientras que la prueba de RT-PCR en todas las plantas evaluadas. Los resultados de secuenciación mostraron altos valores de identidad nucleotídica con las secuencias reportadas en el NCBI para este virus. La transmisión mecánica fue el método más efectivo para la inoculación del CMV y las técnicas de análisis molecular fueron más sensibles y confiables para su identificación, en comparación con la prueba serológica.O vírus do mosaico do pepino (CMV) é um agente limitante na produção e qualidade das culturas agrícolas, cuja inoculação eficaz e diagnóstico confiável são essenciais para implementar estratégias de manejo adequadas e oportunas. Foram avaliados dois protocolos de inoculação de CMV em plântulas indicadoras (Nicotiana benthamiana e N. tabacum cv. Xanthi) e diferentes métodos de detecção do vírus. A infecção com CMV foi obtida por transmissão mecânica e por infiltração. A presença do vírus foi confirmada pela expressão dos sintomas, serologia (ImmunoStrip®), RT-PCR e sequenciamento. Os resultados confirmaram a eficácia da transmissão mecânica, no qual plântulas infectadas apresentaram sintomas associados ao vírus 15 dias após a inoculação. O teste serológico permitiu detectar a presença do patógeno apenas nas plantas de N. benthamiana, enquanto o teste RT-PCR em todas as plantulas avaliadas. Os resultados do sequenciamento mostraram altos valores de identidade de nucleotídeos com as sequências do vírus no NCBI. A transmissão mecânica foi o método mais eficaz para a inoculação do CMV e as técnicas de análise molecular foram mais sensíveis e confiáveis ​​para identificação do CMV, em comparação com os testes serológicos

Characterization of Western Corn Rootworm (Coleoptera: Chrysomelidae) susceptibility to foliar insecticides in northeast Nebraska

Foliar-applied insecticides are commonly used for adult Western Corn Rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), control in Nebraska but little efficacy data is available. Anecdotal reports of reduced efficacy in areas of northeast Nebraska led to the conduct of this study. Objectives were to (i) evaluate the efficacy of commercial applications of commonly used formulated insecticides (bifenthrin, lambda-cyhalothrin, chlorpyrifos, or tank mixes) for WCR control in 7 northeast Nebraska counties during 2019 and 2020 and (ii) conduct adult WCR concentration-response vial bioassays with bifenthrin, chlorpyrifos, and dimethoate active ingredients on a subset of field populations. Whole plant counts (WPC) were used to measure WCR densities in insecticide-treated and untreated maize fields before and after insecticide application. Field control was excellent with organophosphate/pyrethroid tank mixes as proportional change in mean WPC of treated fields was significantly reduced (\u3e0.90) versus untreated fields where little change in WPC occurred. The exception was one treated Boone County field where proportional reduction in WPC was ≤0.78. Bioassays revealed LC50s and resistance ratios of most populations exposed to bifenthrin and dimethoate were not significantly different than the susceptible control. Most populations exhibited a low level of chlorpyrifos resistance when compared to the susceptible control. Field and lab data suggest the local onset of practical WCR field-evolved resistance to bifenthrin in Boone County and chlorpyrifos in Boone and Colfax counties. Results of this study will increase our understanding of WCR resistance evolution, serve as a comprehensive baseline for future research, and inform WCR management programs

Transcriptome and gene expression analysis of three developmental stages of the coffee berry borer, Hypothenemus hampei

Coffee production is a global industry valued at approximately 173 billion US dollars. One of the main challenges facing coffee production is the management of the coffee berry borer (CBB), Hypothenemus hampei, which is considered the primary arthropod pest of coffee worldwide. Current control strategies are inefficient for CBB management. Although biotechnological alternatives, including RNA interference (RNAi), have been proposed in recent years to control insect pests, characterizing the genetics of the target pest is essential for the successful application of these emerging technologies. In this study, we employed RNA-seq to obtain the transcriptome of three developmental stages of the CBB (larva, female and male) to increase our understanding of the CBB life cycle in relation to molecular features. The CBB transcriptome was sequenced using Illumina Hiseq and assembled de novo. Differential gene expression analysis was performed across the developmental stages. The final assembly produced 29,434 unigenes, of which 4,664 transcripts were differentially expressed. Genes linked to crucial physiological functions, such as digestion and detoxification, were determined to be tightly regulated between the reproductive and nonreproductive stages of CBB. The data obtained in this study help to elucidate the critical roles that several genes play as regulatory elements in CBB development

Transcriptome and gene expression analysis of three developmental stages of the coffee berry borer, Hypothenemus hampei

Coffee production is a global industry valued at approximately 173 billion US dollars. One of the main challenges facing coffee production is the management of the coffee berry borer (CBB), Hypothenemus hampei, which is considered the primary arthropod pest of coffee worldwide. Current control strategies are inefficient for CBB management. Although biotechnological alternatives, including RNA interference (RNAi), have been proposed in recent years to control insect pests, characterizing the genetics of the target pest is essential for the successful application of these emerging technologies. In this study, we employed RNA-seq to obtain the transcriptome of three developmental stages of the CBB (larva, female and male) to increase our understanding of the CBB life cycle in relation to molecular features. The CBB transcriptome was sequenced using Illumina Hiseq and assembled de novo. Differential gene expression analysis was performed across the developmental stages. The final assembly produced 29,434 unigenes, of which 4,664 transcripts were differentially expressed. Genes linked to crucial physiological functions, such as digestion and detoxification, were determined to be tightly regulated between the reproductive and nonreproductive stages of CBB. The data obtained in this study help to elucidate the critical roles that several genes play as regulatory elements in CBB development

Transcriptome and gene expression analysis of three developmental stages of the coffee berry borer, Hypothenemus hampei

Coffee production is a global industry valued at approximately 173 billion US dollars. One of the main challenges facing coffee production is the management of the coffee berry borer (CBB), Hypothenemus hampei, which is considered the primary arthropod pest of coffee worldwide. Current control strategies are inefficient for CBB management. Although biotechnological alternatives, including RNA interference (RNAi), have been proposed in recent years to control insect pests, characterizing the genetics of the target pest is essential for the successful application of these emerging technologies. In this study, we employed RNA-seq to obtain the transcriptome of three developmental stages of the CBB (larva, female and male) to increase our understanding of the CBB life cycle in relation to molecular features. The CBB transcriptome was sequenced using Illumina Hiseq and assembled de novo. Differential gene expression analysis was performed across the developmental stages. The final assembly produced 29,434 unigenes, of which 4,664 transcripts were differentially expressed. Genes linked to crucial physiological functions, such as digestion and detoxification, were determined to be tightly regulated between the reproductive and nonreproductive stages of CBB. The data obtained in this study help to elucidate the critical roles that several genes play as regulatory elements in CBB development

As enzimas presentes no trato digestivo dos insetos : um alvo susceptível de inibição

Tese (doutorado)—Universidade de Brasília, Instituto de Ciências Biológicas, Departamento de Biologia Celular, 2009.Sementes do feijão (Phaseolus coccineus) foram analisadas pela presença e atividade do inibidor de amilase (-AI). Por meio do uso de anticorpos policlonais gerados contra o inibidor -AI-1 do feijão comum (P. vulgaris), foi possível detectar polipeptídios típicos deste inibidor (Mr. 14 18 kDa), além do polipeptídeo de 32 kDa. Os resultados encontrados nos testes de inibição permitem sugerir que o inibidor presente nas sementes de P. coccineus 35590 seria o melhor candidato para a transformação genética do café visando à geração de plantas com resistência a larvas da broca-do-café. A digestão proteolítica in vitro do inibidor puro causou uma perda rápida da sua atividade inibitória, afetando seriamente sua capacidade natural de interagir com as -amilases. Também, neste trabalho foram estudadas as propriedades das -amilases presentes no trato digestivo de larvas de Tecia solanivora, um inseto-praga que gera um dano importante à batata (Solanum tuberosum). O inseto tem no mínimo três isoformas principais de -amilases digestivas, as quais apresentaram pontos isoelétricos de 5.30, 5.70 e 5.98. A atividade da -amilase digestiva tem um pH ótimo entre 7.0 e 10.0, com um pico de atividade em pH 9.0. A enzima foi inibida por inibidores de natureza protéica presentes nas sementes de P. coccineus (70% de inibição) e de P. vulgaris cv Radical (87% de inibição) em pH 6.0. Os resultados mostram também que o inibidor de - amilase de amaranto pode ser o melhor candidato para gerar batatas modificadas geneticamente resistentes a este inseto-praga. Futuras pesquisas precisam ser desenvolvidas para esclarecer se estes inibidores de -amilase são resistentes às proteases presentes no trato digestivo da lagarta de batata, T. solanivora. Em um ensaio subseqüente, foi desenvolvido e testado um novo e sensível método espectrofotométrico para a detecção da atividada leucine aminopeptidase (-aminoacyl-peptide hydrolase, EC 3.4.11.1) proveniente do trato digestivo de larvas de Telchin licus (Lepidoptera: Castniidae), fazendo uso do substrato L-leucyl-2- naphthylamide. _______________________________________________________________________________ ABSTRACTSeeds of scarlet runner bean (Phaseolus coccineus L.) were analyzed for -amylase inhibitor (-AI) activity. By using polyclonal antibodies raised against pure a−AI-1 from common bean (Phaseolus vulgaris L.) it was possible to detect the typical -AI polypeptides (Mr 14 -18 kDa) as well as a large polypeptide of Mr 32,000 Da. Differential inhibition curves lead us to suggest that the gene encoding one of the inhibitors in P. coccineus (in accession G35590) would be a good candidate for genetic engineering of coffee resistance towards the coffee berry borer Hypothenemus hampei Ferrari (Coleoptera: Scolytidae). An in vitro proteolytic digestion treatment of pure a−AI−1, resulted in a rapid loss of the inhibitory activity, seriously affecting its natural capacity to interact with mammalian -amylases. In addition, the properties of the digestive -amylase from T. solanivora larvae, an important insect pest of potato (Solanum tuberosum), were studied. This insect has three major digestive -amylases with isoelectric points 5.30; 5.70 and 5.98 respectively, which were separated using native and isoelectric focusing gels. The -amylase activity has an optimum pH between 7.0 and 10.0 with a peak at pH 9.0. The enzyme was inhibited by proteinaceous inhibitors from P. coccineus (70% inhibition) and P. vulgaris cv. Radical (87% inhibition) at pH 6.0. The results show that the -amylase inhibitor from Amaranthus may be a better candidate to make genetically modified potatoes resistant to this insect. However, it is necessary to know if these -amylase inhibitors are resistant to the gut proteases from T. solanivora. On the other hand, a simple and sensitive spectrophotometric assay to determine the activity of leucine aminopeptidase (-aminoacyl-peptide hydrolase, EC 3.4.11.1) from insect intestinal tracts of Telchin licus (Lepidoptera: Castniidae), using L-leucyl-2- naphthylamide as substrate, was developed

Diabrotica undecimpunctata virus 2, a Novel Small RNA Virus Discovered from Southern Corn Rootworm, Diabrotica undecimpunctata howardi Barber (Coleoptera: Chrysomelidae)

The genome of Diabrotica undecimpunctata virus 2 (DuV2), a positive-sense, single-stranded RNA virus identified from the southern corn root- worm transcriptome, comprises 5,313 nucleotides, including a short poly(A) tail. The two open reading frames encode a nonstructural polyprotein (p156) and a putative capsid protein (p25)

Genome Sequence of a Small RNA Virus of the Southern Corn Rootworm, Diabrotica undecimpunctata howardi Barber (Coleoptera: Chrysomelidae)

The genome sequence of a novel small RNA virus, tentatively named Diabrotica undecimpunctata virus 1 (DuV1), was discovered from the transcriptome of the southern corn rootworm, Diabrotica undecimpunctata howardi Barber. DuV1 has a positive-sense, single-stranded RNA genome that encodes a single polyprotein of 3,401 amino acids with limited similarity to other viruses