In Silico Characterisation of the Aedes aegypti Gustatory Receptors.

Aedes aegypti, also known as the dengue mosquito or the yellow fewer mosquito, is the vector of dengue, chikungunya, Zika, Mayaro and yellow fever viruses. The A. aegypti genome contains an array of gustatory receptor (GR) proteins that are related to the recognition of taste. In this study, we performed in silico molecular characterization of all 72 A. aegypti GRs reported in the latest version of A. aegypti genome AaegL5. Phylogenetic analysis classified the receptors into three major clads. Multiple GRs were found to encode multiple transcripts. Physicochemical attributes such as the aliphatic index, hydropathicity index and isoelectric point indicated that A. aegypti gustatory receptors are highly stable and are tailored to perform under a variety of cellular environments. Analysis for subcellular localization indicated that all the GRs are located either in the extracellular matrix or the plasma membrane. Results also indicated that the GRs are distributed mainly on chromosomes 2 and 3, which house 22 and 49 GRs, respectively, whereas chromosome 1 houses only one GR. NCBI-CDD analysis showed the presence of a highly conserved 7tm_7 chemosensory receptor protein superfamily that includes gustatory and odorant receptors from insect species Anopheles gambiae and Drosophila melanogaster. Further, three significantly enriched ungapped motifs in the protein sequence of all 72 A. aegypti gustatory receptors were found. High-quality 3D models for the tertiary structures were predicted with significantly higher confidence, along with ligand-binding residues. Prediction of S-nitrosylation sites indicated the presence of target cysteines in all the GRs with close proximity to the ligand-bindings sites within the 3D structure of the receptors. In addition, two highly conserved motifs inside the GR proteins were discovered that house a tyrosine (Y) and a cysteine (C) residue which may serve as targets for NO-mediated tyrosine nitration and S-nitrosylation, respectively. This study will help devise strategies for functional genomic studies of these important receptor molecules in A. aegypti and other mosquito species through in vitro and in vivo studies.

Fitness costs in chlorfenapyr-resistant populations of the chive maggot, Bradysia odoriphaga.

The chive maggot, Bradysia odoriphaga (Yang and Zhang) is an economically important insect pest, affecting many key vegetables, including Chinese chive, especially in northern China. Chlorfenapyr, a halogenated pyrrole insecticide that interferes with mitochondrial oxidative phosphorylation is widely used against B. odoriphaga. In this study, we evaluated selection-induced resistance to chlorfenapyr and fitness costs in B. odoriphaga. The results showed that B. odoriphaga developed 43.32-fold resistance after continuous exposure to chlorfenapyr for over 10 consecutive generations. The life-history traits of chlorfenapyr-resistant and susceptible strains were compared using age-stage, two-sex life table approach. No significant effects were observed on the longevity and pre-adult period. However, reduction in the total pre-oviposition period (TPOP) and fecundity (eggs/female) were observed in the resistant strain. Moreover, the demographic parameters such as intrinsic rate of increase (r), net reproductive rate (R0) and finite rate of increase (λ) were also decreased significantly in the resistant strain compared to the susceptible strain. These results showed the potential of B. odoriphaga to develop resistance against chlorfenapyr under continuous selection pressure. Furthermore, there was a fitness cost linked with chlorfenapyr resistance in B. odoriphaga. We conclude that a better knowlegde on the trade-off at play between resistance degree and fitness cost could be crucial for developing further management of B. odoriphaga in China.

Acaricidal efficacy of Calotropis procera (Asclepiadaceae) and Taraxacum officinale (Asteraceae) against Rhipicephalus microplus from Mardan, Pakistan.

Medicinal plants are used by traditional folk healers, modern physicians, and veterinarians as an alternative to conventional drugs to treat a wide range of disorders including parasitic diseases. Some compounds from these plants have been shown to have acaricidal activity and repel arthropods. The cattle tick Rhipicephalus microplus is one of the most destructive pests to the livestock industry in tropical and subtropical parts of the world. The potential to develop herbal acaricides to control R. microplus infestations is critical in maintaining cattle herd productivity, reducing economic losses, and curtailing the overuse of synthetic chemical acaricides. Calotropis procera, the apple of Sodom, and Taraxacum officinale, the common dandelion, were evaluated for acaricidal activity against R. microplus larvae and adults in vitro. Both plant species tested are common indigenous species of Pakistan where R. microplus infestations are widespread across livestock species including cattle, sheep, and goats. Whole-plant extracts derived from C. procera and T. officinale significantly reduced the index of egg laying (P < 0.01) and increased the percent inhibition of oviposition of adult female ticks at a concentration of 40 mg/mL when assessed by the adult immersion test (AIT). Calotropis procera and T. officinale treatments at the same concentration also resulted in larval mortality of 96.0% ± 0.57 and 96.7% ± 0.88, respectively, as measured using the larval packet test (LPT). An increasing range of extract concentrations was tested to determine the LD50 and LD90 for C. procera, 3.21 and 21.15 mg/mL, respectively, and T. officinale, 4.04 and 18.92 mg/mL, respectively. These results indicate that further studies are warranted to determine the relative contribution of individual phytochemicals from whole-plant extracts on acaricidal activity. This information will guide the design of further acaricidal efficacy tests using livestock infested with R. microplus.

Salicylic acid-mitigates abiotic stress tolerance via altering defense mechanisms in Brassica napus (L.).

Under the changing climate due to global warming, various abiotic stresses including drought (D) and salinity (S) are expected to further trigger their devastating effects on the already vulnerable crop production systems. This experiment was designed to unravel and quantify the potential role of exogenous application of salicylic acid (SA) in mitigating both D and S stresses and their combination (D+S), with three replications using CRD (Completely Randomized Design). The obtained results of the current study demonstrated significant effects of all three types of stresses (D, S, and D+S) on various parameters in Brassica napus plants. Quantifying these parameters provides a more informative and precise understanding of the findings. Current results revealed that all three stress types (D, S, and D+S) resulted in a reduction in leaf area (13.65 to 21.87%), chlorophyll levels (30 to 50%), gaseous exchange rate (30 to 54%) and the concentration of mineral ions compared to non-stressed plants. However, application of SA helped in mitigating these stresses by ameliorating the negative effects of these stresses. Moreover, Malondialdehyde (MDA) contents, an indicator of lipid per-oxidation and oxidative stress, the levels of antioxidants, proline content, an osmolyte associated with stress tolerance, and sugar content in the leaves were elevated in response to all stress conditions. In addition, the ultra-structures within the leaves were negatively affected by the stresses, while an application of SA considerably minimized the deterioration of these structures thus providing protection to the brassica plants against the stresses. In a nutshell, the findings of this study suggest that SA application in S, D and S+ D stresses provides evasion to the plants by improving different physiological and growth indices. The application of Salicylic Acid (SA) mitigated the negative effects of the stresses on all the above parameters, reducing MDA contents (47%), antioxidants (11 to 20%), proline (28%), sugar contents (20.50%), and minimizing the deterioration of ultra-structures. The findings emphasize the potential mitigatory role of SA in mitigating D and S stresses and highlight the need for further research to understand the underlying mechanisms in detail and explore its practical application in farming practices.

Genomes of the cosmopolitan fruit pest Bactrocera dorsalis (Diptera: Tephritidae) reveal its global invasion history and thermal adaptation.

INTRODUCTION: The oriental fruit fly Bactrocera dorsalis is one of the most destructive agricultural pests worldwide, with highly debated species delimitation, origin, and global spread routes. OBJECTIVES: Our study intended to (i) resolve the taxonomic uncertainties between B. dorsalis and B. carambolae, (ii) reveal the population structure and global invasion routes of B. dorsalis across Asia, Africa, and Oceania, and (iii) identify genomic regions that are responsible for the thermal adaptation of B. dorsalis. METHODS: Based on a high-quality chromosome-level reference genome assembly, we explored the population relationship using a genome-scale single nucleotide polymorphism dataset generated from the resequencing data of 487 B. dorsalis genomes and 25 B. carambolae genomes. Genome-wide association studies and silencing using RNA interference were used to identify and verify the candidate genes associated with extreme thermal stress. RESULTS: We showed that B. dorsalis originates from the Southern India region with three independent invasion and spread routes worldwide: (i) from Northern India to Northern Southeast Asia, then to Southern Southeast Asia; (ii) from Northern India to Northern Southeast Asian, then to China and Hawaii; and (iii) from Southern India toward the African mainland, then to Madagascar, which is mainly facilitated by human activities including trade and immigration. Twenty-seven genes were identified by a genome-wide association study to be associated with 11 temperature bioclimatic variables. The Cyp6a9 gene may enhance the thermal adaptation of B. dorsalis and thus boost its invasion, which tended to be upregulated at a hardening temperature of 38 °C. Functional verification using RNA interference silencing against Cyp6a9, led to the specific decrease in Cyp6a9 expression, reducing the survival rate of dsRNA-feeding larvae exposed to extreme thermal stress of 45 °C after heat hardening treatments in B. dorsalis. CONCLUSION: This study provides insights into the evolutionary history and genetic basis of temperature adaptation in B. dorsalis.

Range expansion decreases the reproductive fitness of Gentiana officinalis (Gentianaceae).

Plants living at the edge of their range boundary tend to suffer an overall decline in their fitness, including growth and reproduction. However, the reproductive performance of plants in artificially expanded habitats is rarely investigated, although this type of study would provide a better understanding of range limitations and improved conservation of ex situ plants. In the current study, we transplanted a narrowly dispersed species of Gentiana officinalis H. Smith (Gentianaceae) from its natural area of distribution to two different elevations and natural elevation to comprehensively study its pollination biology, including flowering phenology and duration, floral display, reproductive allocation, pollinator activity, and seed production. The findings indicated that the starting point and endpoint of the flowering phenology of G. officinalis were earlier at the low elevation, but the peak flowering periods did not differ significantly between any of the experimental plots. When transplanted, the flowering duration, especially the female phase, was reduced; the floral display, including spray numbers, flower numbers, and flower size (length and width), decreased, especially at high elevations. Moreover, the pollen numbers and pollen-ovule ratio were decreased at both high and low elevations, although the ovule numbers showed no change, and aboveground reproductive allocation was decreased. Furthermore, pollinator richness and activity were significantly decreased, and the seed-set ratio decreased under both natural conditions and with supplemental pollination. Finally, more severe pollen limitation was found in transplanted individuals. These results indicated an overall decrease in reproductive fitness in plants living outside their original area of distribution when the geographical range of G. officinalis was expanded.

A novel gene, CaATHB-12, negatively regulates fruit carotenoid content under cold stress in Capsicum annuum.

BACKGROUND: Carotenoids, the secondary metabolites terpenoids, are the largest factors that form the fruit color. Similar to flavonoids, they are not only safe and natural colorants of fruits but also play a role as stress response biomolecules. METHODS: To study the contribution of the key genes in carotenoids biosynthesis, fruit-color formation, and in response to cold stress, we characterized the key regulatory factor CaATHB-12 from the HD-ZIP I sub-gene family members in pepper. RESULTS: Cold stress enhanced carotenoid accumulation as compared with the normal condition. CaATHB-12 silencing through virus-induced gene silencing changed the fruit color by regulating the carotenoid contents. CaATHB-12 silencing increased the antioxidant enzyme activities in the fruits of pepper, exposed to cold stress, whereas CaATHB-12 overexpression decreased the activities of antioxidant enzymes in the transgenic Arabidopsis lines, exposed to cold stress, suggesting that CaATHB-12 is involved in the regulation of cold stress in the pepper fruits. CONCLUSION: Our research will provide insights into the formation of fruit color in pepper and contribution of CaATHB-12 in response to cold stress. Further study should be focused on the interaction between CaATHB-12 and its target gene.

RNAi-Mediated Knockdown of Chitin Synthase 1 (CHS1) Gene Causes Mortality and Decreased Longevity and Fecundity in Aphis gossypii.

Chitin is a vital part of the insect exoskeleton and peritrophic membrane, synthesized by chitin synthase (CHS) enzymes. Chitin synthase 1 (CHS1) is a crucial enzyme in the final step of chitin biosynthetic pathway and consequently plays essential role towards insect growth and molting. RNA interference (RNAi) is an agent that could be used as an extremely target-specific and ecologically innocuous tactic to control different insect pests associated with economically important crops. The sole purpose of the current study is to use CHS1 as the key target gene against the cotton-melon aphid, Aphis gossypii, via oral feeding on artificial diets mixed with dsRNA-CHS1. Results revealed that the expression level of CHS1 gene significantly decreased after the oral delivery of dsRNA-CHS1. The knockdown of CHS1 gene caused up to 43%, 47%, and 59% mortality in third-instar nymph after feeding of dsCHS1 for 24, 48, and 72 h, respectively, as compared to the control. Consistent with this, significantly lower longevity (approximately 38%) and fecundity (approximately 48%) were also found in adult stage of cotton-melon aphids that were fed with dsCHS1 for 72 h at nymphal stage. The qRT-PCR analysis of gene expression demonstrated that the increased mortality rates and lowered longevity and fecundity of A. gossypii were attributed to the downregulation of CHS1 gene via oral-delivery-mediated RNAi. The results of current study confirm that CHS1 could be an appropriate candidate target gene for the RNAi-based control of cotton-melon aphids.

Insect pollinators diversity and abundance in Eruca sativa Mill. (Arugula) and Brassica rapa L. (Field mustard) crops.

Studies on the insect pollinators diversity and their relative abundance in Eruca sativa Mill. (Arugula) and Brassica rapa L. (field mustard) was carried out during spring season from February to April consecutively during all the three years of 2016-18. Insect pollinators observed belonged to four orders i.e. Hymenoptera, Diptera, Lepidoptera, and Coleoptera. A total of 20 major species of insect pollinators were recorded. The highest abundance of pollinator species belonged to Hymenoptera. The most prominent insect pollinator species were Apis mellifera followed by other three honey bee species of A. cerana, A. florea, and A. dorsata respectively. Some species of solitary bees were also recorded. From Diptera, four species of syrphid fly and one species from Muscidae family were also recorded. Insect pollinators recorded from order Lepidoptera were Pieris brassicae, Vanessa cardui, and Papilio demoleus. Lady bird beetle Coccinella septempunctata was recorded from Coleoptera order as occasional visitor. It was noticed that E. sativa attracted more insect pollinators than B. rapa which may be attributed to different amount and chemical properties of nectar, with number of pollen grains, and flower canopy of both crops. Further studies are needed to confirm the reasons for higher pollinator visitation to E. sativa than B. rapa through chemical analysis of nectar, amount of pollens, flower physiology and phenology of both crops.