Comparative analysis of proteome changes induced by the two spotted spider mite Tetranychus urticae and methyl jasmonate in citrus leaves

Citrus plants are currently facing biotic and abiotic stresses. Therefore, the characterization of molecular traits involved in the response mechanisms to stress could facilitate selection of resistant varieties. Although large cDNA microarray profiling has been generated in citrus tissues, the available protein expression data are scarce. In this study, to identify differentially expressed proteins in Citrus clementina leaves after infestation by the two-spotted spider mite Tetranychus urticae, a proteome comparison was undertaken using two-dimensional gel electrophoresis. The citrus leaf proteome profile was also compared with that of leaves treated over 0-72 h with methyl jasmonate, a compound playing a key role in the defense mechanisms of plants to insect/arthropod attack. Significant variations were observed for 110 protein spots after spider mite infestation and 67 protein spots after MeJA treatments. Of these, 50 proteins were successfully identified by liquid chromatography-mass spectrometry-tandem mass spectrometry. The majority constituted photosynthesis- and metabolism-related proteins. Five were oxidative stress associated enzymes, including phospholipid glutathione peroxidase, a salt stressed associated protein, ascorbate peroxidase and Mn-superoxide dismutase. Seven were defense-related proteins, such as the pathogenesis-related acidic chitinase, the protease inhibitor miraculin-like protein, and a lectin-like protein. This is the first report of differentially regulated proteins after T. urticae attack and exogenous MeJA application in citrus leaves.     

Prey-related odor preference of the predatory mites Typhlodromalus manihoti and Typhlodromalus aripo (Acari: Phytoseiidae)

Typhlodromalus manihoti and Typhlodromalus aripo are exotic predators of the cassava green mite Mononychellus tanajoa in Africa. In an earlier paper, we showed that the two predators were attracted to odors from M. tanajoa-infested cassava leaves. In addition to the key prey species, M. tanajoa, two alternative prey mite species, Oligonychus ossypii and Tetranychus urticae also occur in the cassava agroecosystem. Here, we used a Y-tube olfactometer to determine the attraction of the predators to odors from O. gossypii- or T. urticae-infested cassava leaves and their prey-related odor preference. T. aripo but not T. manihoti was slightly attracted to odors from O. gossypii-infested leaves. Both predator species showed a stronger response to odors from cassava leaves infested by M. tanajoa over odors from cassava leaves infested by O. gossypii. Neither predator species was attracted to odors from T. urticae-infested leaves and the predators preferred the odors from M. tanajoa-infested leaves over those from T. urticae-infested leaves. When O. gossypii was present together with M. tanajoa on the same leaves or on different sets of leaves offered together as an odor source the two predators were attracted. In contrast, after mixing non-attractive odors from T. urticae-infested leaves with attractive odors from M. tanajoa-infested leaves, neither T. aripo nor T. manihoti was attracted. Ecological advantages and disadvantages of the predators’ behavior and possible implications for biological control of M. tanajoa are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

The influence of host plant species on parasitism of pollen beetles (Meligethes spp.) by Phradis morionellus

We investigated the response of the specialist insect predator Oligota kashmirica benefica (Coleoptera: Staphylinidae) to volatiles from lima bean leaves infested with the spider mite Tetranychus urticae (Acari: Tetranychidae), both in a Y-tube olfactometer and in a field in Kyoto, Japan. Adult male and female predators were significantly more attracted to T. urticae-infested leaves than to clean air. Adult male and female predators were not more attracted to uninfested leaves, artificially damaged leaves, or the spider mites and their visible products when compared to clean air. In a field trap experiment, 12 adult predators were caught in three traps containing T. urticae-infested lima bean plants over 13 days, whereas no adult predators were trapped in three traps containing uninfested lima bean plants during the same period. These results showed that O. kashmirica benefica adults responded to herbivore-induced plant volatiles from T. urticae-infested lima bean leaves under both laboratory and field conditions.     

A key volatile infochemical that elicits a strong olfactory response of the predatory mite <Emphasis Type="Italic">Neoseiulus californicus</Emphasis>, an important natural enemy of the two-spotted spider mite <Emphasis Type="Italic">Tetranychus urticae</Emphasis>

Herbivore-induced plant volatiles (HIPVs) emitted from lima bean leaves infested with the two-spotted spider mites Tetranychus urticae strongly attract the predatory mites Neoseiulus californicus. Among these HIPVs, methyl salicylate and linalool can attract the predators. Three green-leaf volatiles (GLVs) of (Z)-3-hexen-1-ol, (Z)-3-hexenyl acetate and (E)-2-hexenal, found in the odor blends from T. urticae-infested leaves and physically damaged leaves, can also attract the predators. To search for a strong predator attractant, the olfactory responses of N. californicus to each synthetic compound or their combinations were investigated in a Y-tube olfactometer. When presented a choice between a mixture of the five compounds (i.e. the two HIPVs and the three GLVs) and T. urticae-infested leaves, N. californicus did not discriminate between these odor sources. The same trend was observed when either a mixture of the two HIPVs or methyl salicylate vs. T. urticae-infested leaves were compared. In contrast, the predators preferred T. urticae-infested leaves to linalool, each of the three GLVs, or a mixture of the three GLVs. These results indicated that methyl salicylate is a strong predator attractant, and its potential attractiveness almost equaled that of the blend of HIPVs from T. urticae-infested leaves.     

The effect of host plants on Tetranychus evansi, Tetranychus urticae (Acari: Tetranychidae) and on their fungal pathogen Neozygites floridana (Entomophthorales: Neozygitaceae)

In a series of tritrophic-level interaction experiments, the effect of selected host plants of the spider mites, Tetranychus evansi and Tetranychus urticae, on Neozygites floridana was studied by evaluating the attachment of capilliconidia, presence of hyphal bodies in the infected mites, mortality from fungal infection, mummification and sporulation from fungus-killed mite cadavers. Host plants tested for T. evansi were tomato, cherry tomato, eggplant, nightshade, and pepper while host plants tested for T. urticae were strawberry, jack bean, cotton and Gerbera. Oviposition rate of the mites on each plant was determined to infer host plant suitability while host-switching determined antibiosis effect on fungal activity. T. evansi had a high oviposition on eggplant, tomato and nightshade but not on cherry tomato and pepper. T. urticae on jack bean resulted in a higher oviposition than on strawberry, cotton and Gerbera. Attachment of capilliconidia to the T. evansi body, presence of hyphal bodies in infected T. evansi and mortality from fungal infection were significantly higher on pepper, nightshade and tomato. The highest level of T. evansi mummification was observed on tomato. T. evansi cadavers from tomato and eggplant produced more primary conidia than those from cherry tomato, nightshade and pepper. Switching N. floridana infected T. evansi from one of five Solanaceous host plants to tomato had no prominent effect on N. floridana performance. For T. urticae, strawberry and jack bean provided the best N. floridana performance when considering all measured parameters. Strawberry also had the highest primary conidia production. This study shows that performance of N. floridana can vary with host plants and may be an important factor for the development of N. floridana epizootics.     

Induced Response of Tomato Plants to Injury by Green and Red Strains of Tetranychus Urticae

We studied the induced response of tomato plants to the green strain and the red strain of the spider mite Tetranychus urticae. We focused on the olfactory response of the predatory mite Phytoseiulus persimilis to volatiles from T. urticae-infested tomato leaves in a Y-tube olfactometer. Tomato leaves attracted the predatory mites when slightly infested with the red strain, or moderately or heavily infested with the green strain. In contrast, neither leaves that were slightly infested with green-strain mites, nor leaves that were moderately or heavily infested with the red strain attracted the predators. We discuss the specific defensive responses of tomato plants to each of the two strains.     

Functional expression and structural characterization of ORF cDNA encoding chitinase of the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae)

Chitin is an important component of the exoskeleton and peritrophic matrix in insects. Its bio-degradation is initiated by the endo-splitting chitinase. We cloned an ORF cDNA encoding chitinase from the last instar larva of the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae), into E. coli to confirm its functionality. Its amino acid sequence was compared with previously described lepidopteran chitinases. S. exigua chitinase expression enhanced cell growth approx. 1.5 fold in transformed E. coli than in the wild strain in a 1% colloidal chitin-containing medium with insufficient regular nutrients. Compared with the wild strain, the two intracellular chitin degradation derivatives, glucosamine and N-acetylglucosamine, increased approx. 5.8 and 1.5 fold, respectively, and extracellular chitinase activity in the transformed strain was about 1.6 fold higher. The ORF of S. exigua chitinase-encoding cDNA including stop codon was composed of 1674 bp nucleotides and the calculated molecular weight of the deduced 557 amino acid residues was about 62.6 kDa. The ORF consisted of an N-terminal leading signal peptide (AA 1-20), a catalytic domain (AA 21-392), a linker region (AA 393-493), and a C-terminal chitin-binding domain (AA 494-557) showing a typical molting fluid chitinase structure. Phylogenetic analysis determined that all 5 noctuid chitinases were grouped together, while two bombycid enzymes and one tortricid enzyme mapped together in one lineage. In the noctuid group, the sub-lineages reflected their taxonomic relationships at the Genus level.     

Cloning and functional expression of the mitochondrial alternative oxidase gene (aox1) of Aspergillus niger in Lactococcus lactis and its induction by oxidizing conditions

Lactococcus lactis is a widely used food bacterium mainly known for its fermentation metabolism. An important, and for long time overlooked, trait of this species is its ability to perform respiratory metabolism in the presence of heme and under aerobic conditions. There is no evidence however for the presence of an alternative respiration pathway and AOX activity. In this study, a cDNA fragment encoding the mitochondrial alternative oxidase, the enzyme responsible for alternative respiration, from a citric acid producing Aspergillus niger strain was cloned and expressed in L. lactis as a host strain. Expression of aox1 conferred on this organism cyanide-resistant and salicylhydroxamate-sensitive growth. Bioreactor cultures under fully aerobic conditions of the transformed L. lactis showed that the alternative respiratory pathway operates and improves significantly the microorganism's response to oxidizing stress conditions as it enhances biomass production, suppresses lactate formation, and leads to accumulation of large amounts of nisin.     

Effects of prey mite species on life history of the phytoseiid predators Typhlodromalus manihoti and Typhlodromalus aripo

The effects of prey mite suitability on several demographic characteristics of phytoseiid predators and the relationship of these effects to the potential of phytoseiid predators to control herbivorous mite populations are well documented. Evidence has also accumulated in the last 20 years demonstrating that phytoseiid predators utilize herbivorous prey mite-induced plant volatiles as olfactory cues in locating their herbivorous mite prey, but less well established is the predictability of reproductive success from the ability of the predators to utilize olfactory cues to locate their prey, and how these processes are related to the success of the predators as biological control agents of the herbivorous mite. In this study, we determined in laboratory no choice experiments, the development, survivorship and fecundity of the two neotropical phytoseiid predators Typhlodromalus manihoti Moraes and T. aripo DeLeon when feeding on three herbivorous mites, including the key prey species Mononychellus tanajoa (Bondar), and the two alternative prey species Oligonychus gossypii (Zacher) and Tetranychus urticae (Koch). Intrinsic rate of increase (r_m) of T. aripo was 2.1 fold higher on M. tanajoa as prey compared with T. urticae as prey, while it was almost nil on O. gossypii. For T. manihoti, r_m was 2.3 fold higher on M. tanajoa as prey compared with O. gossypii as prey, while reproduction was nil on T. urticae. An independent experiment on odor-related prey preference of the two predator species showed that T. manihoti and T. aripo preferred odors from M. tanajoa-infested leaves to odors from O. gossypii-infested leaves. Moreover, both predator species preferred odors from M. tanajoa-infested leaves over those from T. urticae-infested leaves. As reported here, life history of the two predatory mites matches odor-related prey preference if the key prey species is compared to the two inferior prey species. The implications of our findings for the persistence of T. manihoti and T. aripo and biological control of M. tanajoa in the cassava agroecosystem in Africa are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Differential induction of chitinase in Piper colubrinum in response to inoculation with Phytophthora capsici,the cause of foot rot in black pepper

Plant chitinases have been of particular interest since they are known to be induced upon pathogen invasion. Inoculation of Piper colubrinum leaves with the foot rot fungus, Phytophthora capsici leads to increase in chitinase activity. A marked increase in chitinase activity in the inoculated leaves was observed, with the maximum activity after 60 h of inoculation and gradually decreased thereafter. Older leaves showed more chitinase activity than young leaves. The level of chitinase in black pepper (Piper nigrum L.) upon inoculation was found to be substantially high when compared to P. colubrinum. RT–PCR using chitinase specific primers revealed differential accumulation of mRNA in P. colubrinum leaves inoculated with P. capsici. However, hyphal extension assays revealed no obvious differences in the ability of the protein extracts to inhibit growth of P. capsici in vitro.     

不同猎物饲喂对南方小花蝽捕食量和喜好性的影响

为探讨南方小花蝽对不同猎物的捕食喜好性,室内用西花蓟马、蚕豆蚜、二斑叶螨、混合饲料(同时饲喂3种猎物)分别饲喂南方小花蝽驯化两代,研究了4种饲喂处理的南方小花蝽初孵若虫、5龄若虫和雌成虫对西花蓟马、蚕豆蚜和二斑叶螨的捕食量和喜好性。结果显示不同猎物饲喂处理驯化的南方小花蝽1龄若虫对同一种猎物的捕食量和喜好性均不存在显著差异。南方小花蝽5龄若虫和雌成虫对某种猎物的捕食量因前期取食的猎物种类不同而有显著差异。南方小花蝽5龄若虫和雌成虫均表现出对西花蓟马2龄若虫的正喜好性。蚕豆蚜饲喂处理的5龄若虫和雌成虫对蚕豆蚜表现出正喜好性,除二斑叶螨饲喂处理外其余3种处理的南方小花蝽5龄若虫和雌成虫均表现出对二斑叶螨的负喜好性。以上结果表明4种饲喂驯化处理的南方小花蝽1龄若虫的喜好性不受前期取食猎物的影响,但5龄若虫和雌成虫对前期取食过的猎物的喜好性增强,存在一定的学习行为。     

盐胁迫下荒漠共生植物红砂与珍珠的根茎叶中离子吸收与分配特征

西北荒漠地区C3小灌木红砂(Reaumuria soongorica)和C4半灌木珍珠猪毛菜(Salsola passerina)在特定环境下混生在一起,分布面积广阔。以采自腾格里沙漠边缘荒漠地带的天然野生珍珠猪毛菜和红砂群落的幼苗为材料,经0、100、200、300、400mmol/L NaCl盐溶液共同胁迫10 d,检测它们的含水量、主要矿质离子在根茎叶的含量与分布,揭示二者耐盐的共生协同的离子平衡适应机制。试验结果发现,珍珠猪毛菜叶片具有"吸钾排钠的"的耐盐特征,红砂叶片具备"吸钠排钾"的特征,吸收利用无机矿质离子具备互补效应。二者耐盐Cl、Ca和Si离子吸收与累积能力存在很大差异:随着盐胁迫程度加剧,红砂的根茎叶中Cl离子含量持续增加,并且为珍珠猪毛菜的2—5倍;珍珠猪毛菜根中Ca离子含量为红砂的2—3倍,但含量变化不显著;红砂根中Si离子含量迅速降低后稳定,并且是珍珠猪毛菜根的3—5倍,其他器官变化差异较小。因此,红砂与珍珠猪毛菜的共培养盐胁迫下根中吸收的离子侧重不同,红砂以Na、Cl、Si为主,珍珠猪毛菜以K、Ca为主。随着盐胁迫的程度加强,离子选择吸收系数S k,Na的变化趋势降低,表明二者叶部对Na的选择性减小,K的选择性吸收积累增大,增强了它们的抗盐性,最终使叶片所受盐害减小。总之红砂与珍珠猪毛菜共生的耐盐离子稳态机制显著不同,离子吸收与分布具有互补互利的效应。     

UV tolerance in the two-spotted spider mite, Tetranychus urticae

The two-spotted spider mite, Tetranychus urticae was exposed to UV-C (250 nm), UV-B (300 nm), and UV-A (350 nm). In non-diapausing females, the median effective doses for 50% mortality plus escape incidence (ED₅₀) were 21 (UV-C) and 104 kJ m⁻² (UV-B); those for 50% oviposition rate in continuous darkness-treated mites were 6.2 (UV-C) and 41 kJ m⁻² (UV-B). No significant effects of UV-A on mortality and oviposition rate were observed. The ED₅₀ values for UV-B were similar to the natural UV-B observed for 2-5 days in summer when T. urticae inhabits the undersides of leaves. Therefore, T. urticae possibly uses leaves as a filter to avoid the deleterious effects of UV-B. In diapausing females, low mortality was observed even at high doses of UV radiation, but more than half escaped even at low doses. The orange body color of diapausing females results from accumulation of carotenoids, a scavenger for UV-induced reactive oxygen species; this may explain the low mortality of diapausing females. Diapausing females may overcome the deleterious effects of UV-B during winter in the absence of leaves by emigrating to UV-free environments and by accumulating carotenoids.     

Effects of Light on the Tritrophic Interaction Between Kidney Bean Plants, Two-Spotted Spider Mites and Predatory Mites, Amblyseius Womersleyi (Acari: Phytoseiidae)

By analyzing the volatiles from Tetranychus urticae-infested kidney bean plants (Phaseolus vulgaris) at different times for two days, we found that they were mainly produced in the light. Tetranychus urticae showed a higher oviposition rate and spent more time feeding during the day (in the light) than at night (in the dark). Infested leaves placed in the light attracted the predatory mite Amblyseius womersleyi, whereas those that were placed in the dark for at least 2h in daytime did not. This indicates that presence or absence of light affects the production of herbivore-induced plant volatiles. Amblyseius womersleyi dispersed more frequently and consumed more T. urticae eggs during the day (in the light) than at night (in the dark), whereas their oviposition rate did not differ between day and night. Presence or absence of herbivore-induced plant volatiles in the surroundings did not affect dispersal, predation or oviposition rates of A. womersleyi. These results show that A. womersleyis behavior coincides with the production pattern of herbivore-induced plant volatiles.