Asymmetry in the reproductive interference between two closely related species of spider mites, Panonychus citri and Panonychus osmanthi (Prostigmata: Tetranychidae).

An incomplete species recognition system may cause reproductive interference. Interspecific mating between Panonychus citri and Panonychus osmanthi produced apparently normal F1 hybrid females. However, the F1 females laid few eggs, and the produced eggs did not hatch (suggesting hybrid breakdown). Several common host plants of P. citri and P. osmanthi have been determined, including Japanese pear, peach, and citrus. However, P. osmanthi has been found only on its particular host plants in the genus Osmanthus. To assess the potential for reproductive interference between P. citri and P. osmanthi, we evaluated the competitiveness (1 ♀ × 2 ♂♂) and preference (2 ♀♀ × 1 ♂) of males in mating and guarding behavior and assessed the effect of second mating on offspring paternity using Japanese pear leaf disks. We found that P. citri males were superior competitors. Moreover, the species recognition ability of P. citri males was lower than that of P. osmanthi males, and frequent occurrence of reproductive interference should therefore be expected. In a mixed population (2 ♀♀ × 2 ♂♂), 41% of females copulated with heterospecific males, although the mating superiority of P. citri was not proven. The ratio of effective second mating with conspecific males after heterospecific mating was higher in the crosses to P. citri females (77.8%) than P. osmanthi females (14.3%). Therefore, P. citri females had more chance than P. osmanthi females to recover from the fitness cost imposed by heterospecific mating through subsequent conspecific mating. To date, P. osmanthi has not been detected on host plants other than Osmanthus species. Reproductive interference by P. citri males may therefore narrow the range of host plants available for P. osmanthi.

The flashy red color of the red velvet mite Balaustium murorum (Prostigmata: Erythraeidae) is caused by high abundance of the keto-carotenoids, astaxanthin and 3-hydroxyechinenone.

The red velvet mite, Balaustium murorum (Hermann), is a pollenophagous free-living mite with a flashy red body. This mite occurs in early spring and lives on sunny surfaces of human-made structures, such as concrete. Hence, it is inevitably exposed to a harsh environment due to solar ultraviolet-B (UV-B) radiation and radiant heat, which cause oxidative stress via the production of reactive oxygen species. The spider mite Panonychus citri that resides on upper leaf surfaces accumulates synthesized keto-carotenoids to protect against oxidative stress. Therefore, we evaluated carotenoid composition in the red pigment of B. murorum. To identify major carotenoids, we performed a high-performance liquid chromatography analysis of intact and de-esterified pigments of B. murorum females. The flashy red pigments of B. murorum consisted of the highly abundant keto-carotenoids astaxanthin and 3-hydroxyechinenone (60 and 38% of major carotenoids, respectively), and a small amount of ß-carotene (2%). Although P. citri is an astaxanthin-rich species, the astaxanthin concentration (per protein) in B. murorum is 127-fold that in P. citri. Due to their high antioxidant activities, those keto-carotenoids probably contribute to the survival of B. murorum in the harsh environment caused by solar UV-B radiation and radiant heat in inorganic habitats.

Combination of target site mutation and associated CYPs confers high-level resistance to pyridaben in Tetranychus urticae.

Pyridaben is a mitochondrial electron transport complex I inhibitor. The H110R mutation in the PSST subunit has been reported as a major factor in pyridaben resistance in the two-spotted spider mite, Tetranychus urticae. However, backcross experiments revealed that the mutant PSST alone conferred only moderate resistance. In contrast, inhibition of cytochrome P450 (CYP) markedly reduces resistance levels in a number of highly resistant strains. It was reported previously that maternal factors contributed to the inheritance of pyridaben resistance in the egg stage, but the underlying mechanisms have yet to be elucidated. Here, we studied the combined effects of the PSST H110R mutation and candidate CYPs, as metabolic resistance factors, on pyridaben resistance in T. urticae. We found that the maternal effects of inheritance of resistance in the egg stage were associated with CYP activity. Analysis of differential gene expression by RNA-seq identified CYP392A3 as a candidate causal factor for the high resistance level. Congenic strains, where the alleles of both PSST and CYP392A3 were derived from a resistant strain (RR_i; i = 1 or 2) and a susceptible strain (SS_i) in a common susceptible genetic background, were constructed by marker-assisted backcrossing. RR_i showed upregulation of CYP392A3 and high resistance levels (LC50 > 10,000 mg L-1), while SS_i had LC50 < 10 mg L-1. To disentangle the individual effects of PSST and CYP392A3 alleles, we also attempted to uncouple these genes in RR_i. We conclude that given the variation in LC50 values and expression levels of CYP392A3 in the congenic and uncoupled strains, it is likely that the high pyridaben resistance levels are due to a synergistic or cumulative effect of the combination of mutant PSST and associated CYPs, including CYP392A3, but other yet to be discovered factors cannot be excluded.

Disruption of a horizontally transferred phytoene desaturase abolishes carotenoid accumulation and diapause in Tetranychus urticae.

Carotenoids underlie many of the vibrant yellow, orange, and red colors in animals, and are involved in processes ranging from vision to protection from stresses. Most animals acquire carotenoids from their diets because de novo synthesis of carotenoids is primarily limited to plants and some bacteria and fungi. Recently, sequencing projects in aphids and adelgids, spider mites, and gall midges identified genes with homology to fungal sequences encoding de novo carotenoid biosynthetic proteins like phytoene desaturase. The finding of horizontal gene transfers of carotenoid biosynthetic genes to three arthropod lineages was unprecedented; however, the relevance of the transfers for the arthropods that acquired them has remained largely speculative, which is especially true for spider mites that feed on plant cell contents, a known source of carotenoids. Pigmentation in spider mites results solely from carotenoids. Using a combination of genetic approaches, we show that mutations in a single horizontally transferred phytoene desaturase result in complete albinism in the two-spotted spider mite, Tetranychus urticae, as well as in the citrus red mite, Panonychus citri Further, we show that phytoene desaturase activity is essential for photoperiodic induction of diapause in an overwintering strain of T. urticae, consistent with a role for this enzyme in provisioning provitamin A carotenoids required for light perception. Carotenoid biosynthetic genes of fungal origin have therefore enabled some mites to forgo dietary carotenoids, with endogenous synthesis underlying their intense pigmentation and ability to enter diapause, a key to the global distribution of major spider mite pests of agriculture.

Molecular monitoring of Neoseiulus californicus released from sheltered slow-release sachets for spider mite control in a Japanese pear greenhouse.

A novel system for spider mite control was developed with a slow-release sachet containing Neoseiulus californicus (McGregor) (Acari: Phytoseiidae) protected by a waterproof shelter. Monitoring the efficacy of the predator release system for spider mite control at a Japanese pear greenhouse requires discrimination of N. californicus from other indigenous phytoseiid mite species inhabiting the study site and subsequent identification of the released N. californicus. The report of our earlier study described a PCR-based method for discrimination of N. californicus species. For the present study, we first examined phytoseiid mite species composition in the greenhouse. Subsequently, we developed microsatellite markers to identify the released N. californicus. Finally, we installed the predator release system in the greenhouse and conducted a population survey of phytoseiid and spider mites. Results demonstrated that approximately 1 month is necessary for distribution of the released N. californicus on the leaves.

Convergent evolution of cytochrome P450s underlies independent origins of keto-carotenoid pigmentation in animals.

Keto-carotenoids contribute to many important traits in animals, including vision and coloration. In a great number of animal species, keto-carotenoids are endogenously produced from carotenoids by carotenoid ketolases. Despite the ubiquity and functional importance of keto-carotenoids in animals, the underlying genetic architectures of their production have remained enigmatic. The body and eye colorations of spider mites (Arthropoda: Chelicerata) are determined by ß-carotene and keto-carotenoid derivatives. Here, we focus on a carotenoid pigment mutant of the spider mite Tetranychus kanzawai that, as shown by chromatography, lost the ability to produce keto-carotenoids. We employed bulked segregant analysis and linked the causal locus to a single narrow genomic interval. The causal mutation was fine-mapped to a minimal candidate region that held only one complete gene, the cytochrome P450 monooxygenase CYP384A1, of the CYP3 clan. Using a number of genomic approaches, we revealed that an inactivating deletion in the fourth exon of CYP384A1 caused the aberrant pigmentation. Phylogenetic analysis indicated that CYP384A1 is orthologous across mite species of the ancient Trombidiformes order where carotenoids typify eye and body coloration, suggesting a deeply conserved function of CYP384A1 as a carotenoid ketolase. Previously, CYP2J19, a cytochrome P450 of the CYP2 clan, has been identified as a carotenoid ketolase in birds and turtles. Our study shows that selection for endogenous production of keto-carotenoids led to convergent evolution, whereby cytochrome P450s were independently co-opted in vertebrate and invertebrate animal lineages.

Combination of restriction endonuclease digestion with the ΔΔCt method in real-time PCR to monitor etoxazole resistance allele frequency in the two-spotted spider mite.

Monitoring resistance allele frequency at the early stage of resistance development is important for the successful acaricide resistance management. Etoxazole is a mite growth inhibitor to which resistance is conferred by an amino acid substitution in the chitin synthase 1 (CHS1; I1017F) in T. urticae. If the susceptible allele can be specifically digested by restriction endonuclease, the ΔΔCt method using real-time PCR for genomic DNA (RED-ΔΔCt method) may be available for monitoring the resistance allele frequency. We tested whether the etoxazole resistance allele frequency in a pooled sample was accurately measured by the RED-ΔΔCt method and validated whether the resistance variant frequency was correlated with etoxazole resistance phenotype in a bioassay. Finally, we performed a pilot test using field populations. Strong linearity of the measures by the RED-ΔΔCt method with practical resistance allele frequencies; resistance allele frequency in the range between 0.5% to at least 0.75% was strictly represented. The strong linear relationship between hatchability of haploid male eggs after the etoxazole treatments (phenotype) and resistance allele frequencies in their mothers provided direct evidence that I1017F is a primary resistance factor to etoxazole in the strains used for experiments. The pilot test revealed a significant correlation between egg hatchability (including both diploid female eggs and haploid male eggs) and estimators in field populations. Consequently, we concluded that the RED-ΔΔCt method is a powerful tool for monitoring a resistance allele in a pooled sample.

Photo-enzymatic repair of UVB-induced DNA damage in the two-spotted spider mite Tetranychus urticae.

Ambient ultraviolet-B (UVB) radiation induces lethal effects in the two-spotted spider mite Tetranychus urticae, whereas photoreactivation by irradiation with ultraviolet-A and visible light (VIS) plays an important role to increase survival of mites irradiated by UVB. The physiological mechanisms and ecological significance of photoreactivation in terrestrial arthropods have not been shown clearly. We verified the biological impact and accumulation of DNA lesions by UVB irradiation and the repair of them by photoreactivation in T. urticae larvae. Survival of UVB-irradiated larvae decreased with increasing UVB dose, but recovered remarkably with VIS exposure after UVB irradiation (photoreactivation). The DNA lesions, cyclobutane pyrimidine dimers (CPDs) and 6-4 pyrimidine-pyrimidine photoproducts (6-4PPs) linearly increased with the UVB dose. The CPDs were repaired after exposure to VIS, whereas the frequency of 6-4PPs was unaffected by VIS; CPD photolyase genes, but not (6-4) photolyase genes, have been found in the T. urticae genome. Therefore, DNA damage and CPD photo enzymatic repair (PER) is significant for survival in this mite under ambient UVB radiation. Unexpectedly, gene expression of CPD photolyase was unaffected by irradiation with UVB and VIS. Instead, expression of xeroderma pigmentosum A (XPA) was increased by irradiation. XPA is a core factor in nucleotide excision repair (NER), which is a repair system unrelated to photo energy. The relationship between gene expression and enzymatic repair remains unclear. To elucidate the PER process in T. urticae, further study will be necessary on the gene expression patterns and molecular functions of CPD photolyase in PER and of XPA in NER.

Effects of combination between web density and size of spider mite on predation by a generalist and a specialist phytoseiid mite.

Complicated three-dimensional webs of silk threads produced by Tetranychus spider mites provide protection from predation by many generalist phytoseiid mite species. Accessibility to prey may be the most significant criterion for successful predation in complicated webs. To assess the protective effects of combination between web size and density, we compared predation on eggs of the two-spotted spider mite, Tetranychus urticae, by a generalist (Euseius sojaensis) and a specialist (Neoseiulus womersleyi) phytoseiid mite in the laboratory under manipulated web size and density. Web sizes negatively affected to the predation. Significant interactions were found between phytoseiid mite species and prey distribution; egg consumption by E. sojaensis was more in uniform distribution, but that by N. womersleyi, in contrast, was larger in contagious distribution. However, the egg consumption by E. sojaensis and N. womersleyi were both mitigated in larger webs. This area size depending mitigation was more effective to the predation by E. sojaensis. Although the mechanism of web size depending mitigation is unknown, web sizes might affect prey searching efficiency of this phytoseiid mite. Consequently, combination between web density and size are likely to affect not only a generalist E. sojaensis but also a specialist N. wormersleyi.

Factors affecting photoreactivation in UVB-irradiated herbivorous spider mite (Tetranychus urticae).

Ambient ultraviolet-B (UVB) radiation causes lethal damage to spider mites, and the extent of photochemical effects is determined by cumulative irradiance in the two-spotted spider mite, Tetranychus urticae. However, the LD50 values obtained using a UVB lamp were much lower than those elicited by solar UVB radiation. As solar radiation includes intense visible light and UVA, we assumed that a photoreactivation mechanism would play a role to survive under solar radiation. We assessed the capacity for photoreactivation in T. urticae eggs and larvae, and found that the efficacy of photoreactivation was determined by the cumulative irradiance of visible light (VIS) after exposure to UVB radiation. The wavelength range effective for photoreactivation went from UVA to green. Next, we found that an increased time lag between UVB and VIS radiation reduced the photoreactivation efficacy in eggs. In contrast, a time lag ≤4 h did not affect that in larvae. We discussed the possibility that the timing of photoreactivation occurs related with phase-specific UVB vulnerability and outbreak symptoms due to UVB-induced DNA damage. Our results suggest that T. urticae depends on a photoreactivation mechanism, and that the photoreactivation efficiency probably caused the divergence in UVB impact between UV lamp and solar radiation.

Combined nighttime ultraviolet B irradiation and phytoseiid mite application provide optimal control of the spider mite Tetranychus urticae on greenhouse strawberry plants.

BACKGROUND: Tetranychus urticae is a hard-to-control pest of greenhouse strawberry production. Nighttime ultraviolet B (UV-B) radiation using light reflection sheets (LRS) has been applied as a physical method to control T. urticae through direct ovicidal effects (the UV method). However, because strawberry leaves grow more densely, UV-B radiation fails to reach the lower leaf surfaces inhabited by spider mites; therefore, a complementary method is required. We propose the supplemental application of phytoseiid mites in greenhouse strawberry production. We evaluated the control effects of UV-B irradiation, phytoseiid mite application and their combined use. The effects of UV-B irradiation on the degree of overlap relative to the independent distributions (ω) between predators and prey were also analyzed. RESULTS: The UV method alone maintained low T. urticae density levels from November to February; however, mite populations increased from March onward. Phytoseiid mite application in January and February without UV-B irradiation resulted in a temporary increase in spider mites in March and/or April. By contrast, combined application of the UV method and phytoseiid mites had a greater control effect during the strawberry growing season. The ω values were higher for the UV method compared with no UV-B irradiation, suggesting that UV-B irradiation increased phytoseiid mite foraging rates. CONCLUSION: The release of phytoseiid mites compensated for the shortcomings of the UV method, and UV-B irradiation promoted predation by phytoseiid mites by increasing the behavioral numerical response. Consequently, combined application of UV-B irradiation and phytoseiid mites is optimal for T. urticae control in greenhouse strawberry production. © 2023 Society of Chemical Industry.

PCR-RFLP analysis for identification of Tetranychus spider mite species (Acari: Tetranychidae).

A polymerase chain reaction (PCR)-restriction fragment length polymorphism (PCR-restriction fragment-length polymorphism)-based method for species identification was applied to 14 Tetranychus spider mite species, which were dominant species intercepted at Japanese import plant quarantine. We sequenced the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (rDNA), which included the partial ends of the 18S and 28S ribosomal RNA genes, 5.8S ribosomal RNA gene, and two internal transcribed spacers (ITS1 and ITS2) for 15 populations of the 14 species. We analyzed the recognition sites of four restriction endonucleases, which had been proposed for discrimination of Japanese Tetranychus species, and constructed a scheme for Tetranychus species identification by PCR-restriction fragment-length polymorphism. We then applied the scheme to 245 individuals from 199 populations, most of them were from foreign countries. As a result, all 14 species were correctly identified using PCR-restriction fragment-length polymorphism. This demonstrates the usefulness of the PCR-restriction fragment-length polymorphism method for the worldwide identification of Tetranychus species.

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.

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.

Vulnerability and behavioral response to ultraviolet radiation in the components of a foliar mite prey-predator system.

Ambient ultraviolet-B (UVB) radiation impacts plant-dwelling arthropods including herbivorous and predatory mites. However, the effects of UVB on prey-predator systems, such as that between the herbivorous spider mite and predatory phytoseiid mite, are poorly understood. A comparative study was conducted to determine the vulnerability and behavioral responses of these mites to ultraviolet (UV) radiation. First, we analyzed dose-response (cumulative irradiance-mortality) curves for the eggs of phytoseiid mites (Neoseiulus californicus, Neoseiulus womersleyi, and Phytoseiulus persimilis) and the spider mite (Tetranychus urticae) to UVB radiation from a UV lamp. This indicated that the phytoseiid mites were more vulnerable than the spider mite, although P. persimilis was slightly more tolerant than the other two phytoseiid mites. Second, we compared the avoidance behavior of adult female N. californicus and two spider mite species (T. urticae, a lower leaf surface user; Panonychus citri, an upper leaf surface user) in response to solar UV and visible light. N. californicus actively avoided both types of radiation, whereas P. citri showed only minimal avoidance behavior. T. urticae actively avoided UV as well as N. californicus but exhibited a slow response to visible light as well as P. citri. Such variation in vulnerability and avoidance behavior accounts for differences in the species adaptations to solar UVB radiation. This may be the primary factor determining habitat use among these mites on host plant leaves, subsequently affecting accessibility by predators and also intraguild competition.

Mechanisms underlying the impact and interaction of temperature and UV-B on the hatching of spider mite and phytoseiid mite eggs.

BACKGROUND: A spider mite control method using night-time ultraviolet (UV)-B irradiation was recently developed for strawberry greenhouses (UV method). The control effect of this UV method is negatively affected by increasing temperature. Tetranychus urticae eggs are more resistant to a single dose of UV-B irradiation than Neoseiulus californicus eggs. By contrast, N. californicus can better survive nightly UV-B irradiation with the UV method compared with T. urticae. To elucidate the mechanism underlying these phenomena, we explored the hypotheses that higher temperature promotes photoenzymatic repair (PER) and that mortality is determined by UV-B susceptibility in the embryonic stage exposed to UV-B. RESULTS: PER efficacy was not promoted by increasing temperature. The lowest hatchability (around zero) of T. urticae eggs after a single dose of UV-B irradiation (0.288 and 0.432 kJ m-2 ) without photoreactivation was seen in the morphogenesis stages between "cleavage ended" and "eye points became colored". Based on these results, we developed a linear function of daily UV-B irradiance and deviation of cumulative irradiance during vulnerable embryonic developmental phases from 50% lethal dose (LD50 ) after a single dose of UV-B irradiation. The difference between T. urticae and N. californicus and changes in UV-B vulnerability due to temperature could be explained by this simple relationship. CONCLUSION: Slower development in T. urticae than N. californicus in nature and developmental delay under low temperatures increase the ovicidal effects of the UV method. This shows the advantage of the simultaneous use of the UV method and biological control, contributing to the development of integrated pest management. © 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.

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.

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.

Co-occurrence of subunit B and C mutations in respiratory complex II confers high resistance levels to pyflubumide and cyenopyrafen in the two-spotted spider mite Tetranychus urticae (Acari: Tetranychidae).

BACKGROUND: Pyflubumide and cyenopyrafen are respiratory complex II (complex II) inhibitors. Previous quantitative trait locus analyses suggested associations of I260V and S56L in complex II subunit B (B-I260V) and subunit C (C-S56L) with pyflubumide and cyenopyrafen resistance, respectively, in Tetranychus urticae. However, although resistant strains had been selected separately by these acaricides, all strains were homozygous for both B-I260V and C-S56L. Hence, the effects of each mutation on resistance development remain unclear. RESULTS: We established strains homozygous for B-I260V with C-S56 (B-I260V_I260V/C-S56_S56) and for C-S56L with B-I260 (B-I260_I260/C-S56L_S56L). High resistance levels (LC50 > 1000 mg L-1 ) to pyflubumide and cyenopyrafen was not conferred by B-I260V or C-S56L alone. Next, we prepared intermixed strains by crossing B-I260V_I260V/C-S56_S56 and B-I260_I260/C-S56L_S56L. Selection of the intermixed strains by either acaricide caused very high resistance levels (LC50 ≥ 10 000 mg L-1 ) to both acaricides and fixed both mutations. Allele-selected recoupling of the mutations without acaricide selection also conferred very high resistance levels to both acaricides in the intermixed strains. Unlike these, B-I260V or C-S56L alone conferred very high and high resistance levels to cyflumetofen, respectively. CONCLUSION: We conclude that the effect of individual mutations characteristically varies among complex II inhibitors. Moreover, very high resistance levels to pyflubumide and cyenopyrafen is conferred by the co-occurrence of B-I260V and C-S56L mutations, which alone have limited effects on resistance level.