Gene_locus Report for: spolt-ACHE1

BACKGROUND: The fall armyworm (FAW), Spodoptera frugiperda; J.E. Smith (Lepidoptera: Noctuidae), is now an economically important pest that causes huge losses to maize productivity in sub-Saharan Africa. Variations in sub-population genetics and the processes of rapid adaptation underpinning the invasion remain unclear. For this, the genetic identity and diversity of FAW populations in Uganda were revealed by sequencing 87 samples (collected across the country). Based on the partial mitochondrial cytochrome oxidase I (COI) gene polymorphisms, we further examined the mitochondrial haplotype configuration and compared the FAW in Uganda with sequences from other parts of the world. The molecular target for organophosphate and carbamate resistance, acetylcholinesterase, was also investigated. RESULTS: Analysis of the partial COI gene sequences showed the presence of both rice (predominant) and corn strain haplotypes, with a haplotype diversity of 0.382. Based on the COI marker, pairwise difference distribution analyses, and neutrality tests, showed that the FAW populations in Uganda and the rest of Africa are evolving neutrally, but those in America and Asia are undergoing expansion. Our findings support observations that invasive FAW populations throughout the rest of Africa and Asia share a common origin. Sequencing of the S. frugiperda ace-1 gene revealed four amino acid substitutions, two of which (A201S and F290V) were previously shown to confer organophosphate resistance in both S. frugiperda and several other insect species. The other two previously reported new variations in positions g-396 and g-768, are presumed to be related to the development of insecticide resistance. CONCLUSIONS: This research has increased our knowledge of the genetics of FAW in Uganda, which is critical for pest surveillance and the detection of resistance. However, due to the low gene polymorphism of COI, more evolutionary studies incorporating the Spodoptera frugiperda whole-genome sequence are required to precisely understand the FAW population dynamics, introduction paths, origin, and subsequent spread.

Control of the beet armyworm, Spodoptera exigua depends heavily on chemical insecticides. Chlorpyrifos, an acetylcholinesterase (AChE) inhibitor, has been used in beet armyworm control for many years in China. Here we describe high level resistance to chlorpyrifos in a S. exigua strain, FX19-R, which was developed from a field-collected Chinese strain (FX) by selection with chlorpyrifos in the laboratory. FX19-R showed 1001-fold resistance to chlorpyrifos compared with the laboratory reference strain WH-S. The esterase inhibitor triphenyl phosphate (TPP) provided significant but small synergism (only 3.5-fold) for chlorpyrifos and neither of the glutathione s-transferase depletor diethyl maleate and the cytochrome P450s inhibitor piperonyl butoxide provided any detectable synergism, indicating that AChE insensitivity may play the major role in the resistance in FX19-R. Consistent with this, an amino acid substitution, F443Y (F331Y in standard Torpedo californica numbering) in AChE1 was identified in the FX19-R strain and shown to be tightly linked to chlorpyrifos resistance. Precisely homologous substitutions have been associated with organophosphate resistance in other pest species. A novel amino acid substitution, G311S (or G198S in standard numbering), was also identified in the reference strain WH-S. Recombinantly expressed AChE1 proteins carrying the G311S and F443Y substitutions were about 4.2-fold and 210-fold less sensitive to inhibition by chlorpyrifos oxon than wild-type AChE1, respectively. These results enhance our understanding of the mechanisms of chlorpyrifos resistance and provide a basis for resistance management based on monitoring the F443Y and G311S substitutions.

Two acetylcholinesterase genes (SlAce1 and SlAce2) were cloned from Spodoptera litura, which is an important pest that causes widespread economic damage to vegetables and ornamental plants. We analyzed their expression patterns and compared their biological functions by using RNA interference. Our results showed that SlAce1 and SlAce2 cDNA contains 2085bp and 1917bp nucleotides and encoding proteins of 694 and 638 amino acid residues, respectively. Phylogenic analysis indicated that the lineage of SlAce genes and SlAce1 was completely different from SlAce2. Although both genes were expressed in all developmental stages and majorly in the brain. The expression levels of the both genes were suppressed by inserting their related dsRNA in the 6th instar larvae, which led to 47.3% (SlAce1) and 37.9% (SlAce2) mortality. Interestingly, the suppression of the SlAce2 transcripts also led to significant reductions in the fecundity, hatching, and offspring in the parental generation of S. litura. It is concluded that SlAce2 is responsible for the hydrolysis of acetylcholine and also plays role in female breeding, embryo progress, and the development of progeny. Considerable larval mortality was observed after both AChE genes (i.e. Ace1 and Ace2) were silenced in S. litura confirms its insecticidal effectiveness, which provided a molecular basis in biological pest control approach.

BACKGROUND: The fall armyworm (FAW), Spodoptera frugiperda; J.E. Smith (Lepidoptera: Noctuidae), is now an economically important pest that causes huge losses to maize productivity in sub-Saharan Africa. Variations in sub-population genetics and the processes of rapid adaptation underpinning the invasion remain unclear. For this, the genetic identity and diversity of FAW populations in Uganda were revealed by sequencing 87 samples (collected across the country). Based on the partial mitochondrial cytochrome oxidase I (COI) gene polymorphisms, we further examined the mitochondrial haplotype configuration and compared the FAW in Uganda with sequences from other parts of the world. The molecular target for organophosphate and carbamate resistance, acetylcholinesterase, was also investigated. RESULTS: Analysis of the partial COI gene sequences showed the presence of both rice (predominant) and corn strain haplotypes, with a haplotype diversity of 0.382. Based on the COI marker, pairwise difference distribution analyses, and neutrality tests, showed that the FAW populations in Uganda and the rest of Africa are evolving neutrally, but those in America and Asia are undergoing expansion. Our findings support observations that invasive FAW populations throughout the rest of Africa and Asia share a common origin. Sequencing of the S. frugiperda ace-1 gene revealed four amino acid substitutions, two of which (A201S and F290V) were previously shown to confer organophosphate resistance in both S. frugiperda and several other insect species. The other two previously reported new variations in positions g-396 and g-768, are presumed to be related to the development of insecticide resistance. CONCLUSIONS: This research has increased our knowledge of the genetics of FAW in Uganda, which is critical for pest surveillance and the detection of resistance. However, due to the low gene polymorphism of COI, more evolutionary studies incorporating the Spodoptera frugiperda whole-genome sequence are required to precisely understand the FAW population dynamics, introduction paths, origin, and subsequent spread.

Control of the beet armyworm, Spodoptera exigua depends heavily on chemical insecticides. Chlorpyrifos, an acetylcholinesterase (AChE) inhibitor, has been used in beet armyworm control for many years in China. Here we describe high level resistance to chlorpyrifos in a S. exigua strain, FX19-R, which was developed from a field-collected Chinese strain (FX) by selection with chlorpyrifos in the laboratory. FX19-R showed 1001-fold resistance to chlorpyrifos compared with the laboratory reference strain WH-S. The esterase inhibitor triphenyl phosphate (TPP) provided significant but small synergism (only 3.5-fold) for chlorpyrifos and neither of the glutathione s-transferase depletor diethyl maleate and the cytochrome P450s inhibitor piperonyl butoxide provided any detectable synergism, indicating that AChE insensitivity may play the major role in the resistance in FX19-R. Consistent with this, an amino acid substitution, F443Y (F331Y in standard Torpedo californica numbering) in AChE1 was identified in the FX19-R strain and shown to be tightly linked to chlorpyrifos resistance. Precisely homologous substitutions have been associated with organophosphate resistance in other pest species. A novel amino acid substitution, G311S (or G198S in standard numbering), was also identified in the reference strain WH-S. Recombinantly expressed AChE1 proteins carrying the G311S and F443Y substitutions were about 4.2-fold and 210-fold less sensitive to inhibition by chlorpyrifos oxon than wild-type AChE1, respectively. These results enhance our understanding of the mechanisms of chlorpyrifos resistance and provide a basis for resistance management based on monitoring the F443Y and G311S substitutions.

The fall armyworm (FAW), Spodoptera frugiperda, is a major pest native to the Americas that has recently invaded the Old World. Point mutations in the target-site proteins acetylcholinesterase-1 (ace-1), voltage-gated sodium channel (VGSC) and ryanodine receptor (RyR) have been identified in S. frugiperda as major resistance mechanisms to organophosphate, pyrethroid and diamide insecticides respectively. Mutations in the adenosine triphosphate-binding cassette transporter C2 gene (ABCC2) have also been identified to confer resistance to Cry1F protein. In this study, we applied a whole-genome sequencing (WGS) approach to identify point mutations in the target-site genes in 150 FAW individuals collected from China, Malawi, Uganda and Brazil. This approach revealed three amino acid substitutions (A201S, G227A and F290V) of S. frugiperda ace-1, which are known to be associated with organophosphate resistance. The Brazilian population had all three ace-1 point mutations and the 227A allele (mean frequency = 0.54) was the most common. Populations from China, Malawi and Uganda harbored two of the three ace-1 point mutations (A201S and F290V) with the 290V allele (0.47-0.58) as the dominant allele. Point mutations in VGSC (T929I, L932F and L1014F) and RyR (I4790M and G4946E) were not detected in any of the 150 individuals. A novel 12-bp insertion mutation in exon 15 of the ABCC2 gene was identified in some of the Brazilian individuals but absent in the invasive populations. Our results not only demonstrate robustness of the WGS-based genomic approach for detection of resistance mutations, but also provide insights for improvement of resistance management tactics in S. frugiperda.

Acetylcholinesterase (AChE) is a vital enzyme that hydrolyzes acetylcholine. Here, full-length complementary DNAs (cDNAs) of two acetylcholinesterase genes (SeAce1 and SeAce2) were obtained from Spodoptera exigua, a widespread phytophagous pest in agriculture. The complete SeAce1 cDNA comprised 5447 nucleotides including an open reading frame (ORF) encoding 694 amino acids, while SeAce2 cDNA encompassed a 1917-bp ORF which would likely yield 638 amino acids. Both SeAce1 and SeAce2 contained specific characteristics of functional AChE. A phylogenetic tree of all lepidopteran insect Aces showed S. exigua clustered with S. litura, Helicoverpa assulta, and H. armigera, all of which are Noctuidae. In S. exigua, SeAce1 gene expression levels (reverse transcription polymerase chain reaction [RT-PCR] and quantitative RT-PCR) were markedly increased compared with SeAce2 in all developmental phases and tissue types. Both genes were down regulated by inserting the corresponding dsRNAs in 5th instar larvae, which resulted in 56.7% (SeAce1) and 24.6% (SeAce2) death. Downregulation of both SeAce1 and SeAce2 significantly reduced fecundity and vitellogenin gene expression in S. exigua. These results revealed the biological functions of the two Ace genes (SeAce1 and SeAce2), providing novel insights into the development of strategies for controlling insect pests.

Fall armyworm, Spodoptera frugiperda (J.E. Smith), is an important pest that damages agriculture worldwide. Organophosphorus insecticides are used for its control; however, intensive and inadequate use of insecticides has selected for resistance. The ace-1 gene confers cross resistance to organophosphorus and carbamate insecticides in cholinergic synapses. SNPs associated with resistance to organophosphorus insecticide were searched using resequencing of the ace-1 gene in populations of S. frugiperda from eight municipalities at Zacatecas, Mexico: Calera, Fresnillo, General R. Murguia, Loreto, Ojocaliente, Tepechitlan, Tlaltenango, and Villanueva. The ace-1 gene was amplified by PCR and bidirectionally sequenced in third and fifth larval instars. Results indicated the presence of the g-301 mutation in the population at Loreto, and the g-565 mutation in populations at Calera, Ojocaliente, Tepechitlan, and Villanueva (50% of the populations) with 25% homozygous and 25% heterozygous. The g-380 mutation was not observed at any town. New variations in positions g-396, g-498, and g-768 also were found. Using association analysis with a dendrogram, four groups of populations were two homozygous resistant and two homozygous susceptible. The results suggested an extensive analysis of resistance of populations in the State of Zacatecas and in Mexico to establish better strategies for controlling the pest.

Two acetylcholinesterase genes (SlAce1 and SlAce2) were cloned from Spodoptera litura, which is an important pest that causes widespread economic damage to vegetables and ornamental plants. We analyzed their expression patterns and compared their biological functions by using RNA interference. Our results showed that SlAce1 and SlAce2 cDNA contains 2085bp and 1917bp nucleotides and encoding proteins of 694 and 638 amino acid residues, respectively. Phylogenic analysis indicated that the lineage of SlAce genes and SlAce1 was completely different from SlAce2. Although both genes were expressed in all developmental stages and majorly in the brain. The expression levels of the both genes were suppressed by inserting their related dsRNA in the 6th instar larvae, which led to 47.3% (SlAce1) and 37.9% (SlAce2) mortality. Interestingly, the suppression of the SlAce2 transcripts also led to significant reductions in the fecundity, hatching, and offspring in the parental generation of S. litura. It is concluded that SlAce2 is responsible for the hydrolysis of acetylcholine and also plays role in female breeding, embryo progress, and the development of progeny. Considerable larval mortality was observed after both AChE genes (i.e. Ace1 and Ace2) were silenced in S. litura confirms its insecticidal effectiveness, which provided a molecular basis in biological pest control approach.

The fall armyworm Spodoptera frugiperda is an economically important pest of small grain crops that occurs in all maize growing regions of the Americas. The intensive use of chemical pesticides for its control has led to the selection of resistant populations, however, to date, the molecular mechanisms underlying resistance have not been characterised. In this study the mechanisms involved in the resistance of two S. frugiperda strains collected in Brazil to chlorpyrifos (OP strain) or lambda-cyhalothrin (PYR strain) were investigated using molecular and genomic approaches. To examine the possible role of target-site insensitivity the genes encoding the organophosphate (acetylcholinesterase, AChE) and pyrethroid (voltage-gated sodium channel, VGSC) target-site proteins were PCR amplified. Sequencing of the S. frugiperda ace-1 gene identified several nucleotide changes in the OP strain when compared to a susceptible reference strain (SUS). These result in three amino acid substitutions, A201S, G227A and F290V, that have all been shown previously to confer organophosphate resistance in several other insect species. Sequencing of the gene encoding the VGSC in the PYR strain, identified mutations that result in three amino acid substitutions, T929I, L932F and L1014F, all of which have been shown previously to confer knockdown/super knockdown-type resistance in several arthropod species. To investigate the possible role of metabolic detoxification in the resistant phenotype of the OP and PYR stains all EST sequences available for S. frugiperda were used to design a gene-expression microarray. This was then used to compare gene expression in the resistant strains with the susceptible reference strain. Members of several gene families, previously implicated in metabolic resistance in other insects were found to be overexpressed in the resistant strains including glutathione S-transferases, cytochrome P450s and carboxylesterases. Taken together these results provide evidence that both target-site and metabolic mechanisms underlie the resistance of S. frugiperda to pyrethroids and organophosphates.