Wolbachia-induced 'hybrid breakdown' in the two-spotted spider mite Tetranychus urticae Koch

The most common post-zygotic isolation mechanism between populations of the phytophagous mite Tetranychus urticae is 'hybrid breakdown', i.e. when individuals from two different populations are crossed, F1 hybrid females are produced, but F2 recombinant male offspring suffer increased mortality. Two-spotted spider mites collected from two populations, one on rose and the other on cucumber plants, were infected with Wolbachia bacteria. These bacteria may induce cytoplasmic incompatibility in their hosts: uninfected (U) females become reproductively incompatible with infected (W) males. We report on the effect of Wolbachia infections in intra- and interstrain crosses on (i) F1 mortality and sex ratios (a test for cytoplasmic incompatibility), and (ii) the number of haploid offspring and mortality in clutches of F1 virgins (a test for hybrid breakdown). U x W crosses within the rose strain exhibited partial cvtoplasmic incompatibility. More interestingly, F2 males suffered increased mortality, a result identical to the hybrid breakdown phenomenon. The experiments were repeated using females from the cucumber strain. In interstrain U x W and U x U crosses, hybrid breakdown was much stronger in the former (80 versus 26%). This is the first report of a Wolbachia infection causing a hybrid breakdown phenotype. Our results show that Wolbhachia infections can contribute to reproductive incompatibility between populations of T. urticae.     

High Temperatures Eliminate Wolbachia, a Cytoplasmic Incompatibility Inducing Endosymbiont, From the Two-spotted Spider Mite

Wolbachia can induce cytoplasmic incompatibility (CI) in the arrhenotokous two-spotted spider mite between uninfected females and infected males. Cytoplasmic incompatibility is expressed through a male-biased sex ratio and a low hatchability, and can be suppressed by removing Wolbachia from spider mites reared on a diet with antibiotics. Here we investigated whether heat-treatment can elimate Wolbachia from infected mites. Using a PCR assay with a Wolbachia-specific primer pair (ftsZ), and by standard crosses, we were able to show that 71 per cent of the mites had lost the Wolbachia infection after rearing the infected population at 32 ± 0.5 °C for four generations. The infection could be completely removed when mites were reared at 32 ± 0.5 °C for six generations. Curing through high temperatures could be one of the reasons why mixed infected/uninfected populations occur in the field. An additional consequence of rearing mites at 32 ± 0.5 °C was the shortened development time. The effect of environmental temperature on the abundance of Wolbachia and possible behavioural consequences for the spider mite are discussed.     

Cardinium symbionts cause cytoplasmic incompatibility in spider mites

Intracellular symbiotic bacteria belonging to the Cytophaga-Flavobacterium-Bacteroides lineage have recently been described and are widely distributed in arthropod species. The newly discovered bacteria, named Cardinium sp, cause the expression of various reproductive alterations in their arthropod hosts, including cytoplasmic incompatibility (CI), induction of parthenogenesis and feminization of genetic males. We detected 16S ribosomal DNA sequences similar to those of Cardinium from seven populations of five spider mite species, suggesting a broad distribution of infection of Cardinium in spider mites. To clarify the effect of Cardinium on the reproductive traits of the infected spider mites, infected mites were crossed with uninfected mites for each population. In one of the populations, Eotetranychus suginamensis, CI was induced when infected males were crossed with uninfected females. The other six populations of four species showed no reproductive abnormalities in the F(1) generation, but the possibility of CI effects in the F(2) generation remains to be tested. One species of spider mite, Tetranychus pueraricola, harbored both Cardinium and Wolbachia, but these symbionts seemed to have no effect on the reproduction of the host, even when the host was infected independently with each symbiont.     

Genetic Differentiation in Tetranychus Urticae (Acari: Tetranychidae): polymorphism,host races or sibling species?

Based on allozyme electrophoresis at the Pgm locus and nuclear ribosomal DNA (ITS2) sequences, we studied the genetic variation of the two-spotted spider mite Tetranychus urticae Koch collected on rose bay, Nerium oleander L. (Apocynaceae), from several localities around the Mediterranean basin. In addition, we compared these results with those of Navajas et al. (1998) and Tsagkarakou (1997) who collected from several other host plants from the Mediterranean. In the western part of this area (Spain, France, Tunisia), we found the individuals collected from rose bay to be clearly genetically differentiated from other samples. No evidence of such host-associated differentiation was detected in the eastern Mediterranean (Italy and Greece). The genetic differentiation of mites collected on rose bay was investigated further by studying the reproductive incompatibilities between populations in Greece and in France and a laboratory strain reared on bean, Phaseolus vulgaris, in France. Reciprocal crosses performed between these strains revealed variable levels of incompatibility, spanning from partial to complete reproductive isolation. In all cases incompatibility was asymmetric. We designed a test based on double-mating to establish the fertilization status of females in fully incompatible crosses. These crosses showed that the females had been inseminated, which suggests that the barrier to reproduction is not of a prezygotic behavioral nature. The data raises the question of the relative role of ecological factors (host plant) and geographical distance, in the ongoing differentiation process potentially leading to speciation.     

Euseiusfinlandicus (Acari: Phytoseiidae) as a potential biocontrol agent against Tetranychus urticae (Acari: Tetranychidae): life history and feeding habits on three different types of food

Life history and reproductive parameters of the generalist predatory mite Euseius (Amblyseius) finlandicus (Oudemans) were studied in the laboratory at 25 +/- 1 degrees C, with a 16L:8D photoperiod and 60 +/- 15% RH, to investigate its response to different food sources: an eriophyid mite Aceria sp., tulip pollen Tulipa gesnerana L., and two-spotted spider mite Tetranychus urticae Koch. Total developmental time of the immature stages was the shortest on eriophyid mites, followed by pollen, and then spider mites. Fecundity was highest on pollen (43.69 eggs; 1.63 eggs/female/day), then eriophyid mites (39.73 eggs; 1.37 eggs/female/day) and lowest on spider mites (18.16 eggs; 0.80 eggs/female/day). Intrinsic rate of increase (Rm), net reproductive rate (Ro) and finite rate of increase (lambda) followed the same pattern [pollen (0.168, 27.96 and 1.183, respectively), eriophyid mites (0.153, 20.81 and 1.167), spider mites (0.110, 9.44 and 1.119)]. Mean generation time (days) was the shortest on pollen (19.90), followed by eriophyid mites (20.02), and then spider mites (20.59). Average spider mite larvae consumed by E. finlandicus during immature stages were 9.18 for males and 11.85 for females. Adult E. finlandicus females consumed an average of 166.38 spider mite protonymphs during adult stage compared to an average of 66.55 by males. The number of prey protonymphs consumed per day by females was highest in the oviposition period, lower in the pre-oviposition period and the lowest in the post-oviposition period. The eriophyid mite as a prey recorded the shortest developmental time, while pollen as food recorded the highest oviposition rate in E. finlandicus. The potential of this predator as a biocontrol agent against T. urticae is discussed.     

Allocating time between feeding, resting and moving by the two-spotted spider mite, Tetranychus urticae and its predator Phytoseiulus persimilis

This study characterizes the timing of feeding, moving and resting for the two-spotted spider mite, Tetranychus urticae Koch and a phytoseiid predator, Phytoseiulus persimilis Athias-Henriot. Feeding is the interaction between T. urticae and plants, and between P. persimilis and T. urticae. Movement plays a key role in locating new food resources. Both activities are closely related to survival and reproduction. We measured the time allocated to these behaviours at four ages of the spider mite (juveniles, adult females immediately after moult and adult females 1 and 3 days after moult) and two ages of the predatory mite (juveniles and adult females). We also examined the effect of previous spider mite-inflicted leaf damage on the spider mite behaviour. Juveniles of both the spider mite and the predatory mite moved around less than their adult counterparts. Newly emerged adult female spider mites spent most of their time moving, stopping only to feed. This represents the teneral phase, during which adult female spider mites are most likely to disperse. With the exception of this age group, spider mites moved more and fed less on previously damaged than on clean leaves. Because of this, the spider mite behaviour was initially more variable on damaged leaves. Phytoseiulus persimilis rested at all stages for a much larger percentage of the time and spent less time feeding than did T. urticae; the predators invariably rested in close proximity to the prey. Compared to adult predators, juveniles spent approximately four times as long handling a prey egg. The predator-prey interaction is dependent upon the local movement of both the predators and prey. These details of individual behaviours in a multispecies environment can provide an understanding of population dynamics.     

Altered host plant preference of Tetranychus urticae and prey preference of its predator Phytoseiulus persimilis (Acari: Tetranychidae, Phytoseiidae) on transgenic Cry3Bb-eggplants

The behavior of the two-spotted spider mite, Tetranychus urticae Koch and the predatory mite Phytoseiulus persimilis A.-H. was investigated in laboratory experiments with transgenic Bt-eggplants, Solanum melongena L., producing the Cry3Bb toxin and corresponding isogenic, non-transformed eggplants. In bitrophic experiments, dual-choice disc tests were conducted to reveal the effects of transgenic eggplants on host plant preference of T. urticae. Adult spider mite females were individually placed on leaf discs (2 cm diameter) and were observed during five days. Females occurred significantly more frequently on transgenic halves on which also significantly more T. urticae eggs were found. The effects of a Cry3Bb-eggplant fed prey on the feeding preference of P. persimilis were investigated in tritrophic experiments. Sixteen spider mite females, eight of which had been taken from transgenic and eight from isogenic eggplants, were offered to well-fed females of P. persimilis and numbers of respective spider mites consumed were registered 12 h later when the predators were offered new spider mites again. This procedure was repeated six times. The results revealed that predatory mites consumed significantly less Bt-fed spider mites than prey that had been raised on control eggplants. These results indicate that eggplants expressing the Cry3Bb toxin for resistance against the Colorado potato beetle are more preferred by spider mites but are less preferred by their predator P. persimilis. Possible consequences of these findings for biological control of spider mites on eggplants are discussed.     

Possible ecological consequences of heterospecific mating behavior in two tetranychid mites

Interspecific mating between the two-spotted spider mite,Tetranychus urticae Koch, and the Banks grass mite,Oligonychus pratensis (Banks), was documented using laboratory populations. The incidence of mating betweenT. urticae males andO. pratensis females was 26.0%, while that for the reciprocal mating was 18.8%. The incidence of mating was affected by both male and female species. Such matings may have several important ecological consequences. Interspecific matings resulted in all-male progenies. Thus, progeny sex ratios may be distorted by misdirected mating behavior. In addition, heterospecific mating resulted in lower fecundity than conspecific matings in the two-spotted spider mite, although not in the Banks grass mite. Aerial dispersal behavior of the two-spotted spider mite was also affected. Under crowded conditions and deteriorating resource quality, female mites exhibit an aerial dispersal posture that helps them to become airborne, and allows them to disperse long distances. Forty-two percent ofT. urticae females that mated with conspecific males exhibited this dispersal behavior, compared to only 3.6% for virgin females. The incidence of aerial dispersal behavior for females that mated with heterospecific males was intermediate (27.3%). The effects of these behavioral alterations on male and female fitness may depend on the population structure and resource distribution.     

Voluntary Falling in Spider Mites in Response to Different Ecological Conditions at Landing Points

We examined voluntary-falling behaviour by adult females of the two-spotted spider mite Tetranychus urticae Koch (Acari: Tetranychidae) and one of its major predators Neoseiulus californicus McGregor (Acari: Phytoseiidae). Experiments were conducted using a setup in which mites could only move onto one of two landing points by falling. Significantly more T. urticae females fell onto available food leaves compared to non-food or heavily infested leaves, whereas significantly fewer females fell onto leaves with the predatory mite N. californicus compared to leaves without the predator. This suggests that spider mites can actively choose on which patch to land on the basis of food quality and predation risk on the patch. Using the same experimental setup, starved N. californicus females never fell, suggesting that falling T. urticae females gain the potential advantage of predator avoidance.     

Wolbachia在山楂双叶螨中的感染及对寄主生殖的影响

【目的】共生菌Wolbachia在多种叶螨寄主中引起细胞质不亲和及适合度改变,影响寄主的生物学特性。山楂双叶螨Amphitetranychus viennensis是重要的果树害螨,常暴发成灾。本研究旨在明确Wolbachia在山楂双叶螨中的感染情况及对寄主生殖的影响。【方法】采集自然种群的山楂双叶螨,运用多位点序列分型技术（multilocus sequence typing, MLST）对其体内Wolbachia感染率及株系进行分析;通过杂交试验及生物学观察,分析感染Wolbachia对山楂双叶螨单雌产卵量、后代孵化率、性比及死亡率的影响。【结果】山楂双叶螨自然种群感染一种株系的Wolbachia （wVie）,该Wolbachia株系与小黑花椿象Orius strigicollis和丽蝇蛹集金小蜂Nasonia vitripennis中的Wolbachia株系亲缘关系较近,而与叶螨属Tetranychus叶螨感染的Wolbachia株系亲缘关系较远。Wolbachia与4种分化较小的线粒体单倍型相关联。Wolbachia感染雌虫与不感染雌虫产卵量没有显著差异（P>0.05）。不感染雌虫与感染雄虫交配,卵孵化率显著低于其他杂交组合 (P<0.05),但孵化率仍达近75%。各交配组合的后代性比及死亡率变化不明显（P>0.05）。【结论】Wolbachia在山楂双叶螨种群中的侵染历史较短,对山楂双叶螨的产卵力、后代性比、死亡率没有影响。Wolbachia在山楂双叶螨中诱导产生弱的CI表型。     

Predatory mirids of the green apple aphidAphis pomi,the two-spotted spider miteTetranychus urticae and the European red mitePanonychus ULMI in apple orchards in Québec

The incidence of predation of eight species of predacious mirids (Hemiptera: Miridae) present in an apple orchard of Québec on the green apple aphid, two-spotted spider mite and European red mite were investigated. The daily consumption rates varied from 1–2 green apple aphids forHyaliodes vitripennis Say andCampylomma verbasci Meyer to 7–9 aphids forDeraeocoris fasciolus Knight andLepidopsallus minisculus Knight.H. vitripennis consumed significantly more mites than the other mirid species with 26 and 18 mites per day for the two-spotted spider mite and the European red mite respectively. The combined use ofH. vitripennis andL. minisculus is suggested for the control of phytophagous mites. This paper is contribution No. 335/91.06.02R, Research Station, Agriculture Canada, Saint-Jean-sur-Richelieu, Québec, Canada.     

Wolbachia-Host Interactions: Host Mating Patterns Affect Wolbachia Density Dynamics

Wolbachia are maternally inherited intracellular bacteria that infect a wide range of arthropods and cause an array of effects on host reproduction, fitness and mating behavior. Although our understanding of the Wolbachia-associated effects on hosts is rapidly expanding, our knowledge of the host factors that mediate Wolbachia dynamics is rudimentary. Here, we explore the interactions between Wolbachia and its host, the two-spotted spider mite Tetranychus urticae Koch. Our results indicate that Wolbachia induces strong cytoplasmic incompatibility (CI), increases host fecundity, but has no effects on the longevity of females and the mating competitiveness of males in T. urticae. Most importantly, host mating pattern was found to affect Wolbachia density dynamics during host aging. Mating of an uninfected mite of either sex with an infected mite attenuates the Wolbachia density in the infected mite. According to the results of Wolbachia localization, this finding may be associated with the tropism of Wolbachia for the reproductive tissue in adult spider mites. Our findings describe a new interaction between Wolbachia and their hosts.     

Cardinium is associated with reproductive incompatibility in the predatory mite Metaseiulus occidentalis (Acari: Phytoseiidae)

Cardinium, a bacterium from the Bacteroidetes group, is associated with reproductive manipulations such as cytoplasmic incompatibility, parthenogenesis, and feminization in some arthropod species. We, and others, have shown that Cardinium, but not Wolbachia, is an endosymbiont in some populations of Metaseiulus occidentalis, a phytoseiid mite that is an important predator of spider mite pests of agricultural crops. However, the precise biological effects that Cardinium may have on M. occidentalis remain unclear. In this study we show, in two sets of crosses between different colonies of Cardinium-free (C-) M. occidentalis females and Cardinium-containing (C+) males, that fecundity was reduced in parental females, F1 progeny survival rates were reduced, and fewer female progeny were produced when compared to the reciprocal and control crosses. There were no differences in these attributes in the reciprocal and two control crosses. Cardinium was transmitted maternally but there was no observed paternal transmission. Finally, Cardinium did not cause asexual (thelytoky) reproduction in M. occidentalis. Thus, Cardinium is associated with nonreciprocal reproductive incompatibility in M. occidentalis and our results support the hypothesis that Cardinium is a reproductive parasite in this agriculturally important predator. Cardinium may therefore affect the evolution and ecology of M. occidentalis and biological control efforts using this mite.     

Wolbachia plays no role in the one-way reproductive incompatibility between the hybridizing field crickets Gryllus firmus and G. pennsylvanicus

Wolbachia are cytoplasmically inherited alpha-proteobacteria that can cause cytoplasmic incompatibility (CI) in insects. This incompatibility between sperm and egg is evident when uninfected females mate with infected males. Wolbachia-driven reproductive incompatibilities are of special interest because they may play a role in speciation. However, the presence of Wolbachia does not always imply incompatibility. The field crickets Gryllus firmus and G. pennsylvanicus exhibit a very clear unidirectional incompatibility and have been cited as a possible example of Wolbachia-induced CI. Here, we conduct curing experiments, intra- and interspecific crosses, cytological examination of Wolbachia in testes and Wolbachia quantifications through real-time PCR. All of our data strongly suggest that Wolbachia are not involved in the reproductive incompatibility between G. firmus and G. pennsylvanicus.     

Wolbachia distribution and cytoplasmic incompatibility based on a survey of 42 spider mite species (Acari: Tetranychidae) in Japan

Wolbachia are a group of maternally inherited bacteria that infect a wide range of arthropods. Wolbachia infections are known to result in the expression of various abnormal reproductive phenotypes, the best known being cytoplasmic incompatibility. The first systematic survey of 42 spider mite species in Japan revealed that seven species (16.7%) were infected with Wolbachia. Wolbachia in the spider mites were grouped into three subgroups in supergroup B by phylogenetic analyses of the wsp gene. Most spider mites did not show cytoplasmic incompatibility when infected males were crossed with uninfected females. However, all infected populations of Panonychus mori and Oligonychus gotohi (five and four populations, respectively) possessed modification-positive strains of Wolbachia, and the cytoplasmic incompatibility decreased egg hatchability and female ratio of the spider mites. Thus, some Wolbachia strains cause sex ratio distortion in their hosts.     

Pest species diversity enhances control of spider mites and whiteflies by a generalist phytoseiid predator

To test the hypothesis that pest species diversity enhances biological pest control with generalist predators, we studied the dynamics of three major pest species on greenhouse cucumber: Western flower thrips, Frankliniella occidentalis (Pergande), greenhouse whitefly, Trialeurodes vaporariorum (Westwood), and two-spotted spider mites, Tetranychus urticae Koch in combination with the predator species Amblyseius swirskii Athias-Henriot. When spider mites infested plants prior to predator release, predatory mites were not capable of controlling spider mite populations in the absence of other pest species. A laboratory experiment showed that predators were hindered by the webbing of spider mites. In a greenhouse experiment, spider mite leaf damage was lower in the presence of thrips and predators than in the presence of whiteflies and predators, but damage was lowest in the presence of thrips, whiteflies and predators. Whitefly control was also improved in the presence of thrips. The lower levels of spider mite leaf damage probably resulted from (1) a strong numerical response of the predator (up to 50 times higher densities) when a second and third pest species were present in addition to spider mites, and (2) from A. swirskii attacking mobile spider mite stages outside or near the edges of the spider mite webbing. Interactions of spider mites with thrips and whiteflies might also result in suppression of spider mites. However, when predators were released prior to spider mite infestations in the absence of other pest species, but with pollen as food for the predators, we found increased suppression of spider mites with increased numbers of predators released, confirming the role of predators in spider mite control. Thus, our study provides evidence that diversity of pest species can enhance biological control through increased predator densities.     

Does a vascular fungus of tomato induce a defence response or a change in host plant quality that also affects the oviposition of spider mites?

It has been suggested that previous infection by a vascular fungus causes induced resistance against two-spotted spider mites. To test the generality of this phenomenon, a series of experiments was carried out using two lines of tomato, differing only in resistance againstFusarium. In addition, tests were done in order to see whether the defense response against the fungus also affects the phytophagous mite directly. Inoculation of tomato plants with a vascular fungus (Fusarium oxysporum f.sp.lycopersici race 1) prior to infestation with spider mites caused a decrease in the rate of oviposition of two-spotted spider mites (Tetranychus urticae) on aFusarium-susceptible line, but only when plants were moderately to severely wilted. Spider mite oviposition did not change significantly of a previously inoculatedFusarium-resistant line.AsFusarium causes vascular occlusion and wilting of the plants, drought stress was experimentally induced to determine its influence on the reduction of oviposition. Drought caused a significant reduction in spider mite oviposition. We conclude that the effect of previousFusarium-inoculation on spider mite oviposition is primarily due to the fungus affecting the quality of the host plant (including the effect it may have on the composition of defensive compounds), rather than due to the stimulation of the defense system of the plant. SinceFusarium seals off the xylem vessels, thereby causing wilting of susceptible plants, the reduction in mite oviposition may well be due to drought stress in the leaves, rather than due to the production of phytoalexins.     

二斑叶螨两种群中Wolbachia诱导的胞质不亲和作用的影响因子比较研究

Wolbachia诱导胞质不亲和（cytoplasmic incompatibility, CI）是对寄主的生殖调控中最常见的一种方式,在不同种群中CI表达的差异较大。以二斑叶螨Tetranychus urticae辽宁兴城（LN）和江苏徐州（JS）两个地理种群为实验材料,经筛选获得100%感染Wolbachia和不感染Wolbachia的品系,通过杂交实验和实时定量PCR的方法研究寄主遗传背景、雄螨日龄、温度以及雄螨体内Wolbachia菌量等因子对我国二斑叶螨体内Wolbachia诱导CI能力的影响。结果表明：1,3,5和7日龄的雄螨诱导的CI的强度没有差异,表明雄螨日龄对我国二斑叶螨体内Wolbachia诱导CI的能力没有影响。二斑叶螨分别放在20℃的低温、 25℃的适温和30℃的高温条件下饲养时,Wolbachia诱导CI的能力也没有任何变化,表明温度对我国二斑叶螨体内Wolbachia诱导CI的能力也没有影响。江苏徐州种群所感染Wolbachia菌量显著高于辽宁兴城种群,并且这两个种群感染Wolbachia菌量都随着雄螨日龄的增大而显著增加,表明Wolbachia菌量对我国二斑叶螨体内Wolbachia诱导CI的能力没有影响;江苏徐州种群内Wolbachia不能诱导CI可能是Wolbachia株系与寄主的遗传背景共同作用的结果。研究结果为进一步了解Wolbachia的生殖调控机理提供了重要依据。     

DNA sequences and cross-breeding experiments in the hawthorn spider mite Amphitetranychus viennensis reveal high genetic differentiation between Japanese and French populations

Sequence variation of the complete second internal transcribed spacer (ITS2, 445 bp) of nuclear ribosomal DNA and part of the mitochondrial cytochrome oxidase I gene (COI, 350 bp) was examined in Amphitetranychus viennensis (Zacher) mites (Acari:Tetranychidae) from four French and four Japanese locations. Sequence analysis consistently revealed the separation of the samples in two major groups: French mites differed from Japanese by 3.8–4.1% of the nucleotide divergence in COI sequences. These two groups also displayed distinct ITS2 consensus sequences (2.1% nucleotide divergence). A few variations, not affecting the diagnostic sites around the consensus sequence, were revealed among cloned copies of the same individual. Reciprocal crosses and backcrosses between one French and two Japanese populations disclosed strong reproductive incompatibility. However, fertile hybrid females were obtained, indicating the conspecificity of the tested mites. Despite the presence of Wolbachia in the French strain, but not in the Japanese ones, our crosses did not display the unidirectional incompatibility typically produced by this microorganism, but rather a bidirectional – although asymmetrical – incompatibility pattern. The post-zygotic incompatibilities in A. viennensis cannot be explained by the presence of Wolbachia but to some extent by mite genome divergence resulting from limited gene exchange between allopatric populations. Experiments of Wolbachia elimination by antibiotic treatment and subsequent crosses with cured strains are still needed to fully understand the reproductive incompatibility patterns in this mite species.     

Cold storage of the predatory mite Neoseiulus californicus is improved by pre-storage feeding on the diapausing spider mite Tetranychus urticae

Low air temperature accompanied with high humidity is effective for long-term cold storage of the predatory mite Neoseiulus californicus (McGregor) (Acari: Phytoseiidae). To further improve this storage method, we investigated the effect of pre-storage nutrition on survival during storage and on post-storage quality in terms of survival, oviposition, and progeny viability. The predatory mite was fed from the egg to adult stage on the diapausing two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), non-diapausing spider mites, or Japanese pear pollen, Pyrus pyrifolia Nakai. Newly emerged N. californicus adult females and males were mated, and then both were stored at 7.5 °C and a vapor pressure deficit of 0.0 kPa for up to 75 days. Survival during storage and post-storage quality was significantly better with the diapausing spider mite diet than with the other diets. No effects on the survival or sex ratio of the progeny of the stored adults were observed, regardless of diet or storage duration. Providing diapausing spider mites as a pre-storage diet therefore significantly improves the long-term storage of N. californicus. We discuss the possibility that ingestion of the cryoprotectants, antioxidants, and energy reserves that are present in rich amounts in diapausing spider mites mitigates chilling injury.