Genome wide gene-expression analysis of facultative reproductive diapause in the two-spotted spider mite Tetranychus urticae

Background

Diapause or developmental arrest, is one of the major adaptations that allows mites and insects to survive unfavorable conditions. Diapause evokes a number of physiological, morphological and molecular modifications. In general, diapause is characterized by a suppression of the metabolism, change in behavior, increased stress tolerance and often by the synthesis of cryoprotectants. At the molecular level, diapause is less studied but characterized by a complex and regulated change in gene-expression. The spider mite Tetranychus urticae is a serious polyphagous pest that exhibits a reproductive facultative diapause, which allows it to survive winter conditions. Diapausing mites turn deeply orange in color, stop feeding and do not lay eggs.

Results

We investigated essential physiological processes in diapausing mites by studying genome-wide expression changes, using a custom built microarray. Analysis of this dataset showed that a remarkable number, 11% of the total number of predicted T. urticae genes, were differentially expressed. Gene Ontology analysis revealed that many metabolic pathways were affected in diapausing females. Genes related to digestion and detoxification, cryoprotection, carotenoid synthesis and the organization of the cytoskeleton were profoundly influenced by the state of diapause. Furthermore, we identified and analyzed an unique class of putative antifreeze proteins that were highly upregulated in diapausing females. We also further confirmed the involvement of horizontally transferred carotenoid synthesis genes in diapause and different color morphs of T. urticae.

Conclusions

This study offers the first in-depth analysis of genome-wide gene-expression patterns related to diapause in a member of the Chelicerata, and further adds to our understanding of the overall strategies of diapause in arthropods.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-14-815) contains supplementary material, which is available to authorized users.     

Comparative toxic and sublethal effects of fluvalinate on two-spotted spider mite and European red mite

Four fluvalinate formulations differed in their residual toxicity to female two-spotted spider mite (TSM), Tetranychus urticae adults; the emulsifiable concentrate (EC) was the most toxic. In contrast, there was little difference in toxicity between the formulations with the European red mite (ERM) Panonychus ulmi with the exception of the EC formulation which was the least toxic. Fluvalinate 2F caused minimal (<10%) TSM and ERM egg mortality. Fluvalinate 2F was more toxic and caused greater larval dispersal for the TSM compared to the ERM at the field concentration and below. The toxicity of fluvalinate 2F to TSM and ERM protonymphs, deutonymphs and adults was low, approximately <20% at field concentration. Dispersal was the main response to fluvalinate and this was positively correlated with increasing concentration. The combined mortality and dispersal LC₅₀ was five times lower for ERM protonymphs and adults, but 11 times higher for ERM deutonymphs compared to equivalent TSM life stages. Fluvalinate 2F reduced TSM development from the protonymph and deutonymph stages to a greater extent compared to the ERM. The mortality response to fluvalinate 2F was unaffected by host type (peach or apple) for the TSM whereas ERM mortality was higher on apple compared to peach. TSM dispersal was higher from apple compared to peach whereas ERM dispersal was similar on both host types. Oviposition by both mite species was lower on apple than peach leaves. A 1 h exposure to fluvalinate 2F reduced ERM oviposition for 12 days.     

Effects of fenvalerate and azinphosmethyl on two-spotted spider mite and phytoseiid mites

The pyrethroid fenvalerate showed significantly faster activity against adult two-spotted spider mite Tetranychus urticae Koch c.f. azinphosmethyl using broad bean leaf discs sprayed in a Potter tower. LC₅₀s for fenvalerate were similar at 24 and 48 hr (0.056 and 0.051g AI/1) while LC₅₀s for azinphosmethyl were significantly different at 24 and 48 hr (0.72 and 0.38g AI/1, respectively). Mortality was partitioned to run-off and direct mortality. Fenvalerate showed an increasing contribution to mortality by run-off with increasing concentration. Increasing concentrations of azinphosmethyl had no effect on the proportion of T. urticae running off the discs. Fenvalerate inhibited egg production c.f. azinphosmethyl (60% and 20% inhibition respectively c.f. control after 24hr). The effect was not permanent. Carbaryl showed no acaricidal or inhibitory effects at 1g AI/1. T. urticae detected fenvalerate residues as reflected by choice of oviposition sites on untreated halves of leaf discs c.f. treated halves. Azinphosmethyl had no effect on oviposition preference. Phytoseiid mites were highly sensitive to fenvalerate residues. Predators moved off the test arena into sticky barriers after feeding on fenvalerate-treated eggs or walking on glass slides treated at 0.00005g AI/1.

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Résumé Les insecticides pyrethroïdes ont été utilisés pour lutter contre les pullulations d'araignées rouges. Cette note examine les réponses de Tetranychus urticae Koch et des prédateurs phytoseiidés résistants aux organo-phosphorés, Amblyseius fallacis Garman et Typhlodromus occidentalis Nesbitt, au fenvalerate (pyrethroïde) et à l'azinphosmethyl (organophosphate). Quelques essais avec du carbaryl sont indiqués.Une femelle adulte de T. urticae est placée sur une rondelle de feuille de Phaseolus vulgaris L. pulvérisée dans une tour de Potter.Les résultats sur la mortalité en fonction de la dose obtenus montrent une activité plus rapide du fenvalerate que de l'azinphosmethyl. Les DL₅₀ du fenvalerate (0,056 et 0,051g AI/I) sont les mêmes à 24 et 48 h, tandis que l'azinphosmethyl montre une activité plus lente (DL₅₀ de 0,72 et 0,38g AI/I à 24 et 48 h). La mortalité se partage entre la sortie de la rondelle et la mortalité in situ.Le fenvalerate provoque une plus forte répulsion que l'azinphosmethyl. Contrairement à l'azinphosmethyl le fenvalerate inhibe la production d'oeufs 60% et 20% d'inhibition à DL50 au bout de 24 h par rapport au témoin. Cette inhibition n'est pas permanente. Le carbaryl n'a pas d'effets inhibiteur ou acaricide à 1g AI/l.Les femelles adultes de T. urticae décèlent les résidus de fenvalerate sur les rondelles et pondent leurs oeufs sur les moitiés non traitées ou traitées à l'azinphosmethyl.Les Phytoseiides sont très sensibles aux résidus de fenvalerate. Après consommation d'oeufs traités, A. fallacis est incapable d'éviter des bandes gluantes. T. occidentalis décèle des traitements à 0,00005g AI/l en quittant les lames traitées par les bandes gluantes.

     

Effects of fluvalinate on two-spotted spider mite dispersal, fecundity and feeding

Residual deposits of fluvalinate caused two-spotted spider mite,Tetranychus urticae Koch, dispersal by running off (run-off) and spinning down (spin-down) from treated leaves. Run-off occured at concentrations from 0.0005 g a.i. 1⁻¹ and reached a peak at 0.01 g a.i. 1⁻¹. Spin-down was more predominant at concentrations above 0.01 g a.i. 1⁻¹. Spin-down and run-off proportions varied with fluvalinate formulation. Oviposition was temporarily suppressed on fluvalinate-treated leaves, presumably because of an irritant effect which reduced feeding. An emulsifiable concentrate formulation caused the largest reductions in oviposition. Oviposition returned to the level exhibited by untreated mites two days after removal from treated surfaces. Oviposition and feeding were positively correlated and both declined reciprocally with increasing fluvalinate concentration.     

Alternative phenotypes of male mating behaviour in the two-spotted spider mite

Severe intraspecific competition for mates selects for aggressive individuals but may also lead to the evolution of alternative phenotypes that do not act aggressively, yet manage to acquire matings. The two-spotted spider mite, Tetranychus urticae, shows male mate-guarding behaviour and male–male combat for available females. This may provide opportunity for weaker males to avoid fighting by adopting alternative mating behaviour such as sneaker or satellite tactics as observed in other animals. We investigated male precopulatory behaviour in the two-spotted spider mite by means of video-techniques and found three types of male mating behaviour: territorial, sneaker and opportunistic. Territorial and sneaker males associate with female teleiochrysales and spend much time guarding them. Territorial males are easily disturbed by rival males and engage themselves in fights with them. However, sneaker males are not at all disturbed by rival males, never engage in fights and, strikingly, never face attack by territorial males. Opportunistic males wander around in search of females that are in the teleiochrysalis stage but very close to or at emergence. To quickly classify any given mate-guarding male as territorial or sneaker we developed a method based on the instantaneous response of males to disturbance by a live male mounted on top of a brush. We tested this method against the response of the same males to natural disturbance by two or three other males. Because this method proved to be successful, we used it to collect territorial and sneaker males, and subjected them to morphological analysis to assess whether the various behavioural phenotypes are associated with different morphological characters. However, we found no statistical differences between territorial and sneaker males, concerning the length of the first legs, the stylets, the pedipalps or the body.     

Identification of Wolbachia-responsive microRNAs in the two-spotted spider mite,Tetranychus urticae

Background

The two-spotted spider mite, Tetranychus urticae, is infected with Wolbachia, which have the ability to manipulate host reproduction and fitness. MicroRNAs (miRNAs) are small non-coding RNAs that are involved in many biological processes such as development, reproduction and host-pathogen interactions. Although miRNA was observed to involve in Wolbachia-host interactions in the other insect systems, its roles have not been fully deciphered in the two-spotted spider mite.

Results

Small RNA libraries of infected and uninfected T. urticae for both sexes (in total four libraries) were constructed. By integrating the mRNA data originated from the same samples, the target genes of the differentially expressed miRNAs were predicted. Then, GO and pathway analyses were performed for the target genes. Comparison of libraries showed that Wolbachia infection significantly regulated 91 miRNAs in females and 20 miRNAs in males, with an overall suppression of miRNAs in Wolbachia-infected libraries. A comparison of the miRNA and mRNA data predicted that the differentially expressed miRNAs negatively regulated 90 mRNAs in females and 9 mRNAs in males. An analysis of target genes showed that Wolbachia-responsive miRNAs regulated genes with function in sphingolipid metabolism, lysosome function, apoptosis and lipid transporting in both sexes, as well as reproduction in females.

Conclusion

Comparisons of the miRNA and mRNA data can help to identify miRNAs and miRNA target genes involving in Wolbachia-host interactions. The molecular targets identified in this study should be useful in further functional studies.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-1122) contains supplementary material, which is available to authorized users.     

Vertical dispersal of the two-spotted spider mite Tetranychus urticae, and the predatory mite Phytoseiulus persimilis on dwarf hops

1 The pattern of dispersion within plants of the two-spotted spider mite, Tetranychus urticae, and its predator, the phytoseiid Phytoseiulus persimilis, was studied on the dwarf hop variety First Gold from May to September in 1997 and 1998. 2 Spider mite populations developed on the lower leaves initially but, by late July, as the numbers of mites increased, most were found towards the top of plants. From early August, the numbers of spider mites decreased most rapidly on the upper parts of plants. 3 Where P. persimilis was released, the predator maintained the numbers of T. urticae below those found on non-release plots throughout the season. 4 By early August, the predator’s pattern of dispersion was similar to that of the pest. 5 Predators spread to non-release plots by 20 June in 1997 and 24 July in 1998 and eventually became more numerous than on the plots where they had been released.     

A description of the life cycle of Entomophthora sp. in the two-spotted spider mite

Entomophthora sp. killed its host, Tetranychus urticae, in 3.38 days at 25°C and in 11.02 days at 15°C. Development of hyphal bodies within the mite was studied with a light microscope, and a scanning electron microscope was used to examine spore development and structure. A comparison of this fungus with other mite-infecting fungi revealed that it is more closely related to E. floridana than to E. tetranychi, and should be called Entomophthora sp. near floridana until a more detailed study of E. floridana can be made.     

Effect of age at first mating on primary sex-ratio of the two-spotted spider mite

This paper determines the influence of initial age at mating on the primary sex-ratio of the two-spotted spider miteTetranychus urticae Koch. The specific question asked is whether females that mate at ages beyond ecdysis, or are inseminated at ecdysis by virgin males of set ages, have daughter productions different from control females mated at ecdysis by randomly aged males.Age of initial female mating did not effect survival. Females mated at ecdysis produced more daughters than those mated at age 5 or 10 days. However, daughter production was equal for all female treatments in the post-mating period. All treatments stopped producing daughters at approximately the same age, regardless of when mated. Son production was lowest for females mated at ecdysis. However, son production by females mated when 5 or 10 days old was lower than that of females mated at ecdysis.Male age at mating did not effect female survival. Females exposed to virgin males either 1 or 7 days old produced more daughters than those exposed to males either 11 or 20 days of age. However, a substantial proportion of females in these latter cohorts were not mated. If only mated females are considered, daughter production is the same between all cohorts.Delayed age at first mating influenced sex-ratio only via the females, specifically by changes in daughter production (as opposed to changes in survivorship). Daughters that would have been produced between ecdysis and mating were never recovered by females that mated at ages beyond ecdysis. Thus, these daughters were lost from a sex-ratio viewpoint. Initial age of male mating did not change sex-ratio.     

Post-embryonic development of the two-spotted spider mite Tetranychus urticae with special reference to the differentiation of the female reproductive system

Summary

The spider mite, Tetranychus urticae passes through four active stages in its life cycle: larva, protonymph, deutonymph, adult. The duration at 23°C and 40–60% relative humidity is 10–12 days. Each of the movable stages is followed by a resting phase (= chrysalis) where moulting processes are prepared. The general internal anatomy of the developmental stages does not differ from that of the adults. The gonadial rudiment of the female larva is an undifferentiated cell mass situated between the nervous system and the anus. In the nymphochrysalis, development of oocytes starts as indicated by synaptonemal complexes occuring in cells which are interconnected via cytoplasmic bridges. Cells exhibiting extranuclear material appear during the protonymphal stage. The presumptive uterus and vagina are formed during the deutochrysalis stage and consist of an undifferentiated tube, the distal portion of which is lined by a fine cuticle. The ovary of the deutonymph contains different cell types. Oogonia and growing oocytes are found in the cranial germ region. Oocytes, nurse cells and cells with large lobed nuclei can be observed in the caudal previtellogenic region. Oocytes protrude through the ovarian surface and invade ovarian pouches covered only by the basement membrane whereby each is connected to a tri-nucleate nurse cell via a cytoplasmic bridge. Oocytes increase in size but do not form yolk droplets. The uterus and vagina are differentiated during the late deutonymphal stage and copulation may take place as indicated by the presence of sperm in the lumen of the seminal receptacle.

The results are compared to previously published information on the female reproductive system of T. urticae and discussed with reference to co-operation of ovarian cell types and their origin.     

Variable fitness and reproductive effects of Wolbachia infection in populations of the two-spotted spider mite Tetranychus urticae Koch in China

Maternally inherited Wolbachia bacteria are widely distributed among insects, and their presence usually causes modifications of the host. To understand the evolutionary history of diverse host-Wolbachia associations, we investigated the symbiosis between Wolbachia and the two-spotted spider mite Tetranychus urticae Koch in China. The cytoplasmic incompatibility (CI) level, fecundity, female ratio, host longevity and host development time were examined. Our results indicate that Wolbachia bacteria had variable effects on the reproduction and fitness of Chinese populations of T. urticae. Variability of CI expression within T. urticae ranged from no CI to a strong level of CI in spite of the low variability of the wsp gene. Relative to uninfected mites, infected females in one of the three populations showed enhanced fecundity associated with the infection of Wolbachia. This is the first report of a Wolbachia infection promoting the fecundity of infected females in T. urticae. Furthermore, we found both positive and negative effects of Wolbachia infection on longevity and the development time. The differences in ecological characters may be attributed to both Wolbachia and host genotype.     

Chemical investigation of aggregation behaviour in the two-spotted spider mite Tetranychus urticae

The silk produced by the group-living mite Tetranychus urticae provides group protection and is used as an informative material during habitat settlement, egg laying, mating, and dispersal events. In this context, cues contained in the silk and other materials produced by mites [eggs, black faeces (BF) and white faeces + silk (WFS)] were investigated. Chemical compounds were extracted by hexane or methanol, and choice tests were used to determine the individual attractiveness of each extract. For both solvents, individuals did not respond to the extract from eggs and WFS. BF extracts were attractive for both solvents. After separating the BF methanol extract into four different chemical components using thin layer chromatography, no component was determined to be responsible for mite attraction. This work supports the evidence that the faeces of T. urticae do contain substances that promote behavioural changes. Not particular chemical compounds but combinations of them seem to induce the mites’ preference. Moreover, the response of mites to chemicals seemed to be context dependent as mites belonging to populations with different densities differed in their attraction to BF extracts.     

东亚小花蝽对西方花蓟马和二斑叶螨的捕食选择性

多食性天敌对猎物的捕食选择性及猎物密度对天敌捕食的干扰作用直接影响到天敌对不同猎物的控制作用.在实验室条件下研究了东亚小花蝽对西方花蓟马和/或二斑叶螨不同虫态的捕食选择性,以及两猎物中一种猎物的密度变化对小花蝽取食另一种猎物的影响.结果如下:东亚小花蝽5龄若虫和成虫对西方花蓟马2龄若虫的捕食选择性均强于对其成虫,对二斑叶螨的选择性为雌成螨>若螨>幼螨.实验中西方花蓟马2龄若虫是东亚小花蝽最喜好的虫态.二斑叶螨雌成螨密度固定为60头/19.63cm2,西方花蓟马若虫密度从10增加到60时显著地减少了东亚小花蝽对二斑叶螨的取食.反之,固定西方花蓟马同样密度,增加二斑叶螨密度却没有显著改变小花蝽对西方花蓟马的取食.此结果进一步表明,西方花蓟马是东亚小花蝽更喜好的猎物.     

Demographic examination of sex ratio in the two-spotted spider mite, Tetranychus urticae

Primary sex ratio of arrhenotokous tetranychid mites depends on the age-specific schedule of ratios of progeny produced by individual females. Since primary sex ratio is conditional upon female age and since the proportion of females of a given age in a population is influenced by the rate of increase, both the agespecific ratio of males to females and the overall ratio of males to females depends on the rate of increase. We given an expression for this dependence and compare it to experimental results for the two-spotted spiter mite. These results imply that there is no standard sex ratio of spider mites because of interpopulation variation in rates of increase.

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Résumé Le taux sexuel des acariens tétranychidés arrhénotoques est défini comme le rapport de la somme des nombres de mâles sur la somme des nombres de femelles pour chaque âge de la distribution en âges. Comme le taux sexuel primaire dépend de l'âge de la femelle et comme la proportion de femelles d'un âge donné dans la population est influencée par le taux d'accroissement, alors le rapport mâle/femelle spécifique d'un âge déterminé et le rapport global mâle/femelle dépendent du taux d'accroissement. Nous avons cherché à exprimer cette dépendance et à la comparer aux résultats expérimentaux pour Tetranychus urticae. Ces résultats signifient qu'il n'y a pas de taux sexuel caractéristique de Tétranyques par suite des variations du taux d'accroissement suivant les populations.

     

Genetics of diapause supression in the two-spotted spider mite,Tetranychus urticae Koch

Starting from the Sambucus strain of the two-spotted spider miteTetranychus urticae Koch, the strains ND₁ and ND₂ were selected for lack of diapause under a short-day regimen (LD 1014, 18°C). The genetic basis of the suppression of diapause in ND₁ and ND₂ was analysed by means of mendelian crosses and backcrosses, using the photoperiodic cycle LD 1014 as diagnostic for the distinction of diapausing and non-diapausing phenotypes. Suppression of diapause is inherited as a recessive trait. From backcross analysis it was concluded that the suppression of diapause in ND₁ and ND₂ is under monogenic control. The major genes for suppression of diapause in ND₁ and ND₂ are alleles at a locus designated asd. A mutation for albinism of the locusa-p is pleiotropic for suppression of diapause. It was shown from crosses between ND₁ and the albino strain, thatd anda-p are distinct loci.