Jae-Kyoung Shim

Tomato yellow leaf curl virus (TYLCV-IL): a seed-transmissible geminivirus in tomatoes

Tomato yellow leaf curl virus (TYLCV) is one of the most well-known tomato-infecting begomoviruses and transmitted by Bemisia tabaci. Seed transmission has previously been reported for some RNA viruses, but TYLCV has not previously been described as a seed-borne virus. In 2013 and 2014, without whitefly-mediated transmission, TYLCV was detected in young tomato plants germinated from fallen fruits produced from TYLCV-infected tomato plants in the previous cultivation season. In addition, TYLCV-Israel (TYLCV-IL) was also detected in seeds and their seedlings of TYLCV-infected tomato plants that were infected by both viruliferous whitefly-mediated transmission and agro-inoculation. The seed infectivity was 20–100%, respectively, and the average transmission rate to seedlings was also 84.62% and 80.77%, respectively. TYLCV-tolerant tomatoes also produced TYLCV-infected seeds, but the amount of viral genome was less than seen in TYLCV-susceptible tomato plants. When tomato plants germinated from TYLCV-infected seeds, non-viruliferous whiteflies and healthy tomato plants were placed in an insect cage together, TYLCV was detected from whiteflies as well as receiver tomato plants six weeks later. Taken together, TYLCV-IL can be transmitted via seeds, and tomato plants germinated from TYLCV-infected seeds can be an inoculum source of TYLCV. This is the first report about TYLCV seed transmission in tomato.

Upregulation of the immune protein gene hemolin in the epidermis during the wandering larval stage of the Indian meal moth, Plodia interpunctella.

Expression of hemolin, which generates an immune protein, was up-regulated in wandering fifth instar larval stage of Plodia interpunctella. The mRNA level peaked in the middle of the wandering stage. Major expression was in the epidermis, rather than in the fat body or gut. To test a possible ecdysteroid effect on hemolin induction we treated with RH-5992, an ecdysteroid agonist, and KK-42, which inhibits ecdysteroid biosynthesis in both feeding and wandering fifth instar larvae. When feeding larvae were treated with RH-5992 the hemolin mRNA level was increased. When wandering larvae were treated with KK-42 its level was reduced. In addition, when KK-42-treated larvae were subsequently treated with RH-5992 the hemolin mRNA level was recovered. These results strongly suggest that ecdysteroid up-regulates the expression of hemolin mRNA. Hormonal and bacterial effects on hemolin induction were further analyzed at the tissue level. Major induction of hemolin mRNA was detected following both RH-5992 treatment and bacterial injection in the epidermis of both feeding and wandering larvae. Minor induction of hemolin was detected in the fat body following a bacterial injection, but not RH-5992 treatment. We infer that in P. interpunctella larvae, the epidermis is the major tissue for hemolin induction in naïve insects and in insects manipulated with bacterial and hormonal treatments.

Tissue-specific variation of heat shock protein gene expression in relation to diapause in the bumblebee Bombus terrestris

Diapause‐associated expression patterns of heat shock protein genes (hsp) were determined in the queen of the bumblebee Bombus terrestris, a pollination insect. Partial cDNA sequences of three hsp genes –shsp, hsc70 and hsp90– were obtained, and the deduced amino acid sequences were found to be highly homologous with corresponding hsp of hymenopteran insects. Using northern hybridization, the transcript level of each gene was compared in six stages relating to diapause: pre‐mating, post‐mating, chilling for 1, 2 and 3 months, and post‐chilling. The transcript level was also compared in four tissues of adult queens: brain, thoracic muscle, gut and ovary. The transcript levels of the three hsp genes changed at various rates in relation to diapause, and each pattern was highly tissue‐specific. Overall patterns of hsc70 and hsp90 expression were similar in each tissue. The shsp level in the brain was downregulated after 1 month chilling, but its level in the ovary was upregulated during a long chilling period; levels in muscle and gut did not change in relation to diapause. The levels of both hsc70 and hsp90 in muscle were gradually upregulated in late diapause and postdiapause stages, but levels in the ovary were downregulated during the chilling period, while levels in the brain and gut did not change in relation to diapause. Our results show that the three hsp genes were differentially regulated in stage‐ and tissue‐specific manners throughout diapause, and suggest unique physiological roles for these genes in relation to diapause in each tissue of queen bumblebees.

Molecular identification and phylogenetic analysis of economically important acaroid mites (Acari: Astigmata: Acaroidea) in Korea

Molecular diagnosis is highly valuable for the species identification of microscopic mites. Here, we collected some economically important mites of the superfamily Acaroidea from various stored products in Korea. Those nucleotide sequences of ribosomal second internal transcribed spacer (ITS2) and the mitochondrial cytochrome oxidase subunit I (COI) regions were determined by PCR using universial primers. In nucleotide sequence comparison at GenBank database, seven species including Rhizoglyphus robini, R. echinopus, Sancassania sp. Acarus siro, Tyrophagus putrescentiae, T. similis in Acaridae and Suidasia medanensis in Suidasiidae were identified. Particularly, COI sequences of R. robini, R. echinopus, Sancassania sp. and T. similis were firstly determined. Our results suggest that the phylogenetic relationships inferred from the ITS2 region, rather than COI region, were similar to those derived based on their morphological classification. Our study provides molecular information for the identification and phylogenetic relationship of acaroid mites in Korea.

Development of species-specific primers for rapid diagnosis of Tetranychus urticae, T. kanzawai, T. phaselus and T. truncatus (Acari: Tetranychidae)

Species diagnosis is of the utmost importance to both pest management and plant quarantine services. Because of difficulties in the morphological diagnosis of spider mites, molecular techniques are of great value to rapidly and accurately diagnose closely related species. We examined four species of genus Tetranychus (the green and red forms of T. urticae, and T. kanzawai, T. phaselus and T. truncatus), which are found in Korea and are of significance to plant quarantine services. DNA samples isolated from a single egg, larva or adult weighed 64–188 ng. We designed species‐specific primers by performing sequence alignment for 107 sequences of the ribosomal internal transcribed spacer 2 (ITS2) region, which we obtained from GenBank, and sequences generated in this study. Specific nucleotides of each species were selected for designing primers specific for each species. Each species‐specific primer pair, when used to perform PCR analyses, detected only the species from which it originated. However, a T. urticae‐specific primer pair did not discriminate between the green and red forms of this species. These species‐specific primers can be applied in practice for the rapid and accurate diagnosis of spider mite species in plant quarantine and in agricultural fields.

Differential regulation of heat shock protein genes by temperature in relation to initial diapause in the egg of the katydid Paratlanticus ussuriensis

The lifecycle of Paratlanticus ussuriensis Uvarov can extend for longer than 1 year (plurennial) as a result of prolonged diapause. Eggs can overwinter by entering a facultative initial diapause in the early embryonic stage within the egg, followed by further overwintering by entering an obligatory final diapause at the fully developed embryonic stage. The initial diapause is temperature‐dependent and initiated by continuous incubation at 20 °C but not at 30 °C. To understand the mechanism of initial diapause at the molecular level, cDNAs of small heat shock protein genes (shsp), hsp90, as well as three hsp70 genes (hsp70a, hsp70b and hsp70c), are determined and the transcript levels of the five hsp genes are compared in eggs incubated at 20 or 30 °C for 60 days after oviposition using real‐time reverse transcriptase‐polymerase chain reaction analysis. At 30 °C, transcript levels of shsp, hsp70a, hsp70b and hsp90 increase in eggs with age. However, hsp70c levels uniquely peak only in 25‐day‐old eggs, corresponding to the time of initial diapause. At 20 °C, distinct levels of shsp, hsp70a and hsp90 appear in some stages and fluctuate throughout the 60‐day observation period, although both hsp70b and hsp70c are undetectable. These results suggest that each hsp may play a specific role in relation to initial diapause. In addition, up‐regulation of hsp70c may be associated with the mechanism of embryonic activation for averting initial diapause in warmer conditions.

Behavioral Attraction of Two Parasitoids, Venturia canescens and Bracon hebetor, to Silk Extracts of a Host Plodia interpunctella

Abstract Kairomonal activities of silk extracts of host Plodia interpunctella were determined by measuring the rates of behavioral responses of two parasitic wasps, Venturia canescens and Bracon hebetor. Silk of P. interpunctella larvae attracted both parasitic wasps but the cocoon silk of silkworm, Bombyx mori and the web silk of two-spotted spider mite, Tetranychus urticae did not. Silk components of P. interpunctella were extracted by using either hexane or methanol, and tested the rates of three serial responses of wasps; host location, antennal drumming and ovipositor probing behaviors. The patterns of each behavioral response were similar in two wasps. The rates of each response were increased at the higher concentrations of both extracts. Antennal drumming behavior was much more responsive to lower concentrations of both extracts than ovipositor probing behavior was. Furthermore, the rate of antennal drumming response was higher in hexane-extracts rather than methanol-extract in both wasps; V. canescens and B. hebetor for 20 and 17 times, respectively. However, ovipositor probing response was similar in two different extracts. Both extracts elicited 100% of antennal drumming response but ovipositor probing response was only 60 to 80% of all tested individuals. Our results were shown that silk extracts of host larvae elicited strong behavioral responses of two parasitic wasps and could be applied for practical application of parasitoids attraction in the biological control of agricultural pests.

Identification of glucose-regulated protein 78 (grp78) cDNA from Aphis gossypii and its expression during development, heat shock and nutritional ingestion

Glucose‐regulated protein 78 (GRP78) is a heat shock protein 70 (HSP70) specific to the endoplasmic reticulum (ER). It acts as a chaperone and a signalling regulator that responds to ER stresses in vertebrates. However, its role in invertebrates, including in insects, is uncertain. The present study determines the full grp78 cDNA sequence and expression patterns of grp78 in Aphis gossypii Glover, which is a major pest of numerous crop plants worldwide. The A. gossypii grp78 cDNA contains highly conserved motifs of the HSP70 family, as well as the C‐terminal KDEL motif that is characteristic of ER‐specific HSPs. The amino acid sequence also exhibits 97–98% identity with HSP70 of other aphid species. Quantitative real‐time reverse transcriptase‐polymerase chain reaction analysis shows that the grp78 level is higher in fourth‐instar nymphs than in the younger instar‐nymphs and adults. Its level is not significantly affected by exposure to 0–40 °C for 1 h. The grp78 level gradually increases after ingestion of sucrose solutions that range in concentration from 0% to 30% (w/v) using Parafilm feeding chambers. In addition, the grp78 levels vary among aphids that are fed on the leaves of 13 different plant species for 24 h. More specifically, grp78 levels are higher in aphids that fed on eggplant and pepper than any other host plants examined in the present study. Thus, the results demonstrate that grp78 expression is highly associated with the nutritional uptake of aphids and provide important insights for understanding the molecular aspects of relationships between insects and their host plants.

Molecular identification of four Panonychus species (Acari: Tetranychidae) in Korea, including new records of P. caglei and P. mori

Four species of the genus Panonychus are identified in Korea including two previously known species P. citri (McGregor) and P. ulmi (Koch) and two newly identified species P. mori Yokoyama and P. caglei Mellot. Morphological diagnostics were observed among the four species in the color of dorsal tubercles and in the shape of male aedeagus. The dorsal tubercles of P. citri, P. ulmi, P. mori and P. caglei are red, white, whitish red and light red, respectively. The aedeagi of male adults are diagnostic in sigmoid‐shaped at the distal end and its shape and length was different among four species. Molecular comparison of the internal transcribed spacer 2 (ITS2) sequence of nuclear DNA and cytochrome‐c oxidase subunit I (COI) nucleotide sequence of mitochondrial DNA among the four species showed divergences 8–12% and 9–12%, respectively. Molecular analysis of the ITS2 and COI sequences revealed their divergences were slightly different among four species. In addition, species‐specific primer sets were designed at the base on ITS2 sequences to precisely diagnose these four species at the molecular level.

Genetic variation of the hawthorn spider mite Amphitetranychus viennensis (Acari: Tetranychidae) in Korea: Genetic diversity of Amphitetranychus viennensis

Amphitetranychus viennensis (Zacher) is a serious economic pest of various fruit trees. Due to its restricted geographic distribution in Asia and Europe, it is an important quarantine species in the North American countries. Despite its significance in agricultural damage and quarantine, the genetic diversity of this species has not been studied in detail. In this study, we determined the nucleotide sequences of mitochondrial cytochrome c oxidase subunit I (COI) gene and internal transcribed spacer 2 (ITS2) region of the nuclear ribosomal DNA in A. viennensis collected from various regions of Korea. Variations in the COI and ITS2 sequences of the Korean samples ranged up to 6.92 and 4.02 %, respectively. A sequence comparison showed that the Korean samples were divided into two groups, one comprising the Geoje and Jinju samples, and the other comprising all samples except for the Geoje/Jinju samples. In most samples, except for the Geoje/Jinju samples, variations in both COI and ITS2 sequences were less than 1.04 and 0.99 %, respectively. However, the variations between two groups were 6.23–6.92 % and 3.02–4.02 %, respectively. In addition, the phylogenic analysis performed with sequences in the GenBank database showed that the samples from Geoje/Jinju, which are located in the southern region of Korea, were most similar to the Japanese samples, but the others were most similar to those of China and Taiwan. Thus, our study suggests that two genetically different groups of A. viennensis exist in Korea.