A mutation in chitin synthase I associated with etoxazole resistance in the citrus red mite Panonychus citri (Acari: Tetranychidae) and its uneven geographical distribution in Japan.

BACKGROUND: High-levels of etoxazole resistance have not yet been frequently reported in Panonychus citri. Although a highly resistant strain was discovered in 2014, etoxazole resistance has not become a significant problem in areas of citrus production in Japan. A target site mutation in chitin synthase 1 (CHS1), I1017F, is a major etoxazole-resistance factor in Tetranychus urticae. To investigate the mechanisms of etoxazole resistance and the dispersal of resistance genes, we analyzed target-site mutations in a highly resistant strain and their geographical distribution in Japan. RESULTS: High-level etoxazole resistance was completely recessive. The I1017F mutation was detected in CHS1 of the highly resistant strain, and its frequency was correlated with the hatchability of eggs treated with etoxazole. Sequencing and variant frequency analyses of local populations by quantitative polymerase chain reaction revealed that I1017F is restricted to the Ariake Sea area of Kyushu Island. Although a new nonsynonymous substitution, S1016L, accompanied by I1017F was found in CHS1 of the highly resistant strain, CRISPR/Cas9 engineering of flies showed that S1016L had no effect on the etoxazole resistance conferred by I1017F. CONCLUSION: I1017F is a major target site mutation that confers high-level etoxazole resistance on P. citri. Dispersion of I1017F possibly was suppressed as a result of the completely recessive inheritance of resistance together with low gene flow between local populations. © 2022 Society of Chemical Industry.

A feasibility trial of genomics-based diagnosis detecting insecticide resistance of the diamondback moth.

BACKGROUND: Insecticide resistance management has been key for crop protection for over 70 years and is increasingly important because the development of new active ingredients has decreased in recent years. By monitoring the development of resistance in a timely manner, we can effectively prolong insecticide efficacy. Genomic-based diagnosis can reliably predict resistance development if information on resistant mutations against major pesticides is available. Here, we developed a feasibility trial of genomics-based diagnosis of insecticide resistance in diamondback moth (Plutella xylostella) populations in Nagano Prefecture, Japan. Amplicon sequencing analyses using a next-generation sequencer (Illumina MiSeq) for major insecticides, including diamides, pyrethroids, Bacillus thuringiensis (Bt) toxin (Cry1Ac), organophosphates, and spinosyns, were conducted. RESULTS: Mutations related to the resistance of pyrethroids, organophosphates, and diamides (flubendiamide and chlorantraniliprole) prevailed, while those of a diamide (cyantraniliprole), Bt (Cry1Ac), and spinosyns were scanty, suggesting that they are still effective. The results of the genomics-based diagnosis were generally concordant with the results of bioassays. Resistance development tendencies were generally uniform across Nagano. CONCLUSION: An insecticide-resistance management campaign can be conducted in Nagano Prefecture with a quick genomic-based diagnosis in early spring while bioassay is the only option for monitoring resistances whose mutations are unavailable. Our study is the first step in the future management of insecticide resistance in all significant pests. © 2022 Society of Chemical Industry.

freqpcr: Estimation of population allele frequency using qPCR ΔΔCq measures from bulk samples.

PCR techniques, both quantitative (qPCR) and nonquantitative, have been used to estimate the frequency of a specific allele in a population. However, the labour required to sample numerous individuals and subsequently handle each sample renders the quantification of rare mutations (e.g., pesticide resistance gene mutations at the early stages of resistance development) challenging. Meanwhile, pooling DNA from multiple individuals as a "bulk sample" combined with qPCR may reduce handling costs. The qPCR output for a bulk sample, however, contains uncertainty owing to variations in DNA yields from each individual, in addition to measurement errors. In this study, we have developed a statistical model to estimate the frequency of the specific allele and its confidence interval when the sample allele frequencies are obtained in the form of ΔΔCq in the qPCR analyses on multiple bulk samples collected from a population. We assumed a gamma distribution as the individual DNA yield and developed an R package for parameter estimation, which was verified using real DNA samples from acaricide-resistant spider mites, as well as a numerical simulation. Our model resulted in unbiased point estimates of the allele frequency compared with simple averaging of the ΔΔCq values. The confidence intervals suggest that dividing the bulk samples into more parts will improve precision if the total number of individuals is equal; however, if the cost of PCR analysis is higher than that of sampling, increasing the total number and pooling them into a few bulk samples may also yield comparable precision.

Estimating the proportion of resistance alleles from bulk Sanger sequencing, circumventing the variability of individual DNA.

Specimens should be examined as much as possible to obtain a precise estimate of the proportion of resistance alleles in agricultural fields. Monitoring traps that use semiochemicals on sticky sheets are helpful in this regard. However, insects captured by such traps are ordinarily left in the field until collection. Owing to DNA degradation, the amount of DNA greatly varies among insects, causing serious problems in obtaining maximum likelihood estimates and confidence intervals of the proportion of the resistance alleles. We propose a statistical procedure that can circumvent this degradation issue. R scripts for the calculation are provided for readers. We also propose the utilization of a Sanger sequencer. We demonstrate these procedures using field samples of diamide-resistant strains of the diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae). The validity of the assumptions used in the statistical analysis is examined using the same data.

Optimal management strategy of insecticide resistance under various insect life histories: Heterogeneous timing of selection and interpatch dispersal.

Although theoretical studies have shown that the mixture strategy, which uses multiple toxins simultaneously, can effectively delay the evolution of insecticide resistance, whether it is the optimal management strategy under different insect life histories and insecticide types remains unknown. To test the robustness of this management strategy over different life histories, we developed a series of simulation models that cover almost all the diploid insect types and have the same basic structure describing pest population dynamics and resistance evolution with discrete time steps. For each of two insecticidal toxins, independent one-locus two-allele autosomal inheritance of resistance was assumed. The simulations demonstrated the optimality of the mixture strategy either when insecticide efficacy was incomplete or when some part of the population disperses between patches before mating. The rotation strategy, which uses one insecticide on one pest generation and a different one on the next, did not differ from sequential usage in the time to resistance, except when dominance was low. It was the optimal strategy when insecticide efficacy was high and premating selection and dispersal occur.

Is a larger refuge always better? Dispersal and dose in pesticide resistance evolution.

The evolution of resistance against pesticides is an important problem of modern agriculture. The high-dose/refuge strategy, which divides the landscape into treated and nontreated (refuge) patches, has proven effective at delaying resistance evolution. However, theoretical understanding is still incomplete, especially for combinations of limited dispersal and partially recessive resistance. We reformulate a two-patch model based on the Comins model and derive a simple quadratic approximation to analyze the effects of limited dispersal, refuge size, and dominance for high efficacy treatments on the rate of evolution. When a small but substantial number of heterozygotes can survive in the treated patch, a larger refuge always reduces the rate of resistance evolution. However, when dominance is small enough, the evolutionary dynamics in the refuge population, which is indirectly driven by migrants from the treated patch, mainly describes the resistance evolution in the landscape. In this case, for small refuges, increasing the refuge size will increase the rate of resistance evolution. Our analysis distils major driving forces from the model, and can provide a framework for understanding directional selection in source-sink environments.

Stellate hairs on leaves of a deciduous shrub Viburnum erosum var. punctatum (Adoxaceae) effectively protect Brevipalpus obovatus (Acari: Tenuipalpidae) eggs from the predator Phytoseius nipponicus (Acari: Phytoseiidae).

The eggs of the herbivorous false spider mite Brevipalpus obovatus Donnadieu have a longer incubation period than those of spider mites and are not protected by webs. Brevipalpus obovatus often lays its eggs in the gaps among the hairs on host leaves. We examined the effects of stellate hairs of Viburnum erosum var. punctatum (VEP) leaves on the survival of B. obovatus eggs. Adult B. obovatus and Phytoseius nipponicus Ehara, a generalist predator, were introduced to VEP leaf disks; each B. obovatus egg was inspected daily until hatching. More eggs (63 vs. 42 %) survived on the abaxial surfaces of VEP leaves, where the stellate hairs are more complicated, than on the adaxial surfaces. Predation hazard decreased rapidly with increasing egg age and a substantial portion of the eggs hatched. Phytoseius nipponicus preyed on eggs regardless of egg age when mixed-age eggs were provided. Manipulative experiments with bent stellate hairs showed that the normal hairs reduced the predation risk of B. obovatus eggs by P. nipponicus. Therefore, the predation hazard was considered to decrease since the stellate hairs hindered the search for B. obovatus eggs by the phytoseiid mite.

Tolerance to solar ultraviolet-B radiation in the citrus red mite, an upper surface user of host plant leaves.

Plant-dwelling mites are potentially exposed to solar ultraviolet-B (UVB) radiation that causes deleterious and often lethal effects, leading most mites to inhabit the lower (underside) leaf surfaces. However, in species of spider mite belonging to the Genus Panonychus, a substantial portion of individuals occur on upper leaf surfaces. We investigated whether the upper leaf surfaces of citrus trees are favorable for P. citri, and to what extent they are tolerant to UVB radiation. If eggs are not adequately protected from UVB damage, females may avoid ovipositing on the upper surfaces of sunny leaves. To test this, we conducted laboratory experiments using a UVB lamp, and semioutdoor manipulative experiments. As a result, P. citri eggs are tolerant to UVB. Field studies revealed that the ratio of eggs and adult females on upper leaf surfaces were larger for shaded than for sunny leaves. However, 64-89% of eggs hatched successfully even on sunny upper leaf surfaces. Nutritional evaluation revealed that whether on sunny or shaded leaves, in fecundity and juvenile development P. citri reaped the fitness benefits of upper leaf surfaces. Consequently, P. citri is tolerant to UVB damage, and inhabiting the upper surfaces of shaded leaves is advantageous to this mite.

Geotaxis and leaf-surface preferences mitigate negative effects of a predatory mite on an herbivorous mite.

Reproductive success and population growth of an herbivorous mite are limited by activities of phytoseiid predators. However, occurrences on upper versus lower leaf surfaces are sometimes mismatched between these prey and predators. The mismatch potentially mitigates predation risk for the prey species. We assessed factors that affect mite distributions on leaf surfaces, testing whether the presence of the phytoseiid mite Phytoseius nipponicus alters the leaf-surface distribution and reproductive success of the herbivorous false spider mite Brevipalpus obovatus. The host plant was Viburnum erosum var. punctatum (Adoxaceae). Leaves were set in natural (TRUE) and reversed (upside down; INVERTED) orientations using experimental devices. Both surfaces were accessible to mites. We detected lower and abaxial leaf-surface preferences in P. nipponicus. In contrast, upper and adaxial surfaces were preferred by B. obovatus. Thus, prey and predatory mites accumulated on different sides of leaves. Presence of the predator also indirectly decreased egg production in B. obovatus. Brevipalpus obovatus females actively avoided leaf surfaces with elevated predator numbers; these females shifted their distributions and changed oviposition sites to leaf surfaces with fewer predators. In consequence, B. obovatus eggs on the upper sides of leaves were less frequently preyed upon than were those on lower sides. We suggest that upper leaf-surface exploitation in this particular herbivorous mite species mitigates predation risk from phytoseiid mites, which prefer lower leaf surfaces.

Do plant mites commonly prefer the underside of leaves?

The adaxial (upper) and abaxial (lower) surfaces of a plant leaf provide heterogeneous habitats for small arthropods with different environmental conditions, such as light, humidity, and surface morphology. As for plant mites, some agricultural pest species and their natural enemies have been observed to favor the abaxial leaf surface, which is considered an adaptation to avoid rain or solar ultraviolet radiation. However, whether such a preference for the leaf underside is a common behavioral trait in mites on wild vegetation remains unknown. The authors conducted a 2-year survey on the foliar mite assemblage found on Viburnum erosum var. punctatum, a deciduous shrub on which several mite taxa occur throughout the seasons, and 14 sympatric tree or shrub species in secondary broadleaf-forest sites in Kyoto, west-central Japan. We compared adaxial-abaxial surface distributions of mites among mite taxa, seasons, and morphology of host leaves (presence/absence of hairs and domatia). On V. erosum var. punctatum, seven of 11 distinguished mite taxa were significantly distributed in favor of abaxial leaf surfaces and the trend was seasonally stable, except for Eriophyoidea. Mite assemblages on 15 plant species were significantly biased towards the abaxial leaf surfaces, regardless of surface morphology. Our data suggest that many mite taxa commonly prefer to stay on abaxial leaf surfaces in wild vegetation. Oribatida displayed a relatively neutral distribution, and in Tenuipalpidae, the ratio of eggs collected from the adaxial versus the abaxial side was significantly higher than the ratio of the motile individuals, implying that some mite taxa exploit adaxial leaf surfaces as habitat.

Reconfigurable multilevel transmitter using monolithically integrated quad Mach-Zehnder IQ modulator for optical 16-QAM and 8-PSK generation.

We propose and demonstrate a reconfigurable multilevel transmitter using a monolithically-integrated quad Mach-Zehnder in-phase/quadrature (QMZ-IQ) modulator with binary driving electronics. Different from previous parallel-integrated quadrature amplitude modulation (QAM) transmitter solutions, only one electrode is required to adjust the relative phase offset among embedded sub-Mach-Zehnder modulators in the proposed IQ superstructure. By feeding different RF driving electronics and operating the integrated modulator as different bias conditions, different advanced multilevel modulation formats, such as QAM and 8-ary phase-shift keying (8-PSK), could be synthesized. In this paper, a 40-Gb/s 16-QAM and a 30-Gb/s 8-PSK are generated using the proposed multilevel transmitter, respectively. Offline digital processing is employed for bit-error rates estimation and constellation reconstruction.