Real-Time PCR Method for HPV DNA Detection

Human papillomavirus (HPV) infection is an important etiologic factor in cervical carcinogenesis. Various HPV DNA detection methods have been evaluated for clinicopathological level. For the specimens with normal cytological finding, discrepancies among the detection methods were frequently found and adequate interpretation can be difficult. 6,322 clinical specimens were submitted and evaluated for real-time PCR and Hybrid Capture 2 (HC2). 573 positive or "Not Detected but Amplified" (NDBA) specimens by real-time PCR were additionally tested using genetic analyzer. For the reliability of real-time PCR, 325 retests were performed. Optimal cut-off cycle threshold ( ) value was evaluated also. 78.7% of submitted specimens showed normal or nonspecific cytological finding. The distributions of HPV types by real-time PCR were not different between positive and NDBA cases. For positive cases by fragment analysis, concordance rates with real-time PCR and HC2 were 94.2% and 84.2%. In NDBA cases, fragment analysis and real-time PCR showed identical results in 77.0% and HC2 revealed 27.6% of concordance with fragment analysis. Optimal cut-off value was different for HPV types. NDBA results in real-time PCR should be regarded as equivocal, not negative. The adjustment of cut-off value for HPV types will be helpful for the appropriate result interpretation

Thripinema nicklewoodi (Tylenchida: Allantonematidae), a potential biological control agent of Frankliniella occidentalis (Thysanoptera: Thripidae)

Frankliniella occidentalis Pergande (Thysanoptera: Thripidae) is the most important insect pest for greenhouse flower crops. Thripinema nicklewoodi Siddiqi (Tylenchida: Allantonematidae) is an entomoparasitic nematode attacking and sterilizing the thrips. Methods to propagate and study T. nicklewoodi were developed. I observed an excretion rate of 21.4 nematodes per day by parasitized female thrips. The sex ratio of the excreted nematodes was 6:1 (female:male). After exposing 50 healthy first instars to four parasitized female thrips in a rolled bean leaf, I obtained a 75.3% mean parasitization rate in the adult stage of the thrips. In contrast to previous reports, male thrips were found to be parasitized as readily as females. Parasitism reduced the longevity of both adult female and male thrips by 26% and 61%, respectively. T. nicklewoodi when presented with various thrips life stages achieved the highest attack rate in first and second instars and prepupa. Free-living nematodes were found to escape from hosts through the anus and penetrate new host thrips through the intersegmental membranes of the thorax and abdomen. While nematode parasitization affected tospovirus propagation, it did not reduce transmission of impatiens necrotic spot virus even though parasitism reduced feeding activity of adult female thrips by 81% on leaves, 38% on pollen, and 22% on honey. However, despite lowered total feeding, probing by parasitized thrips (in honey) was not reduced, and this may explain why lowered feeding does not result in lowered virus transmission. In a study of the population dynamics on caged impatiens in greenhouse, nematode transmission persisted for seven host generations and populations of normal (i.e., not parasitized) female thrips declined by 39–79% in the nematode treatment compared to the control. However, no significant reductions were found in numbers of larval thrips between nematode treatments and the control. A higher proportion of male thrips occurred in populations with nematodes in which adult female thrips declined significantly compared to the control population. T. nicklewoodi released seven times on caged impatiens in a greenhouse did not provide preventative control of thrips, though the population growth of second instar, adult female, and male thrips was suppressed by 44, 68, and 49%, respectively

Thripinema nicklewoodi (Tylenchida: Allantonematidae), a potential biological control agent of Frankliniella occidentalis (Thysanoptera: Thripidae).

EntomologyDoctor of Philosophy (Ph.D.

Puncturing apple fruits increases survival of Grapholita molesta (Lepidoptera: Tortricidae) in laboratory rearing.

Grapholita molesta (Busck) is a major pest in orchards of apple, peach, and plum. For better rearing in the laboratory, we compared the life history characteristics of G. molesta by providing larvae with either punctured or unpunctured apple fruits. The development time of immatures and the fecundity of adult females were similar between punctured and unpunctured apples. However, the overall survival rate of G. molesta (larvae to adult emergence) was 1.7 times higher on punctured apples than unpunctured ones, resulting in a higher intrinsic rate of population increase. Therefore, punctured apples would be a better food source for rearing of G. molesta

Development and fecundity performance of Grapholita molesta and Grapholita dimorpha (Lepidoptera: Tortricidae) on different immature fruits.

Both Grapholita molesta (Busck) and Grapholita dimorpha Komai (Lepidoptera: Tortricidae) are important pests of pome and stone fruits. Mature fruits of peach and plum have been tested as food sources for G. molesta, but no studies have examined the suitability of immature fruits, which are the fruit stage more likely to be available for the spring populations of both G. molesta and G. dimorpha. Thus, we evaluated immature fruits of peach, plum, and apple as food sources by assessing their effects on biological and behavioral attributes of these moths in the laboratory. Immature fruits were collected in May and June of 2016 and again in 2017. The first-instar larvae of G. molesta preferred either peach or plum, which showed exit rates of mature larvae of 81.0 and 100.0% for the two fruits, respectively. On peach, development time was shorter, and fecundity was lower than plum. However, G. dimorpha preferred plum and apple, showing the lowest fruit-boring rate and lowest mature larval exit rate on peach, from which only one female moth emerged but produced no eggs. In conclusion, it seems that at the immature fruit stage, plum and apple are better food sources for both G. molesta and G. dimorpha than is peach

Identification and evaluation of a new entomopathogenic fungal strain against Riptortus pedestris (Hemiptera: Alydidae) and its two egg parasitoids.

A strain (ARP14) of the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin was isolated from field-collected Riptortus pedestris (Fabricius) (Hemiptera: Alydidae). The lethal median concentration of the ARP14 strain was compared with that of a commercialized strain (GHA) of the same fungus against R. pedestris and its two egg parasitoids, Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae) and Gryon japonicum (Ashmead) (Hymenoptera: Platygastridae). Mortality and mycosis rates were evaluated after exposure to five concentrations of the fungus, i.e., 1×109, 1×108, 1×107, 1×106, and 1×105 conidia/mL, using a glass scintillation vial as an exposure arena in 25.0 ± 0.5°C and 93.7 ± 2.9% RH. The lethal median concentrations (LC50) for 2nd and 4th instar nymphs, and adults of R. pedestris were not significantly different between the two strains of B. bassiana. However, the mycosis rate of ARP14 was 1.3 and 1.8 times higher than that of the GHA strain in 4th instar nymphs and adult females of R. pedestris, respectively, at the 1×108 conidia/mL concentration. More interestingly, the mycosis rates at 1×108 conidia/mL concentration in the parasitoids G. japonicum and O. nezarae were much lower in the ARP14 strain (15.0 and 0%) than in the GHA strain (73.3 and 66.0%), respectively, suggesting that the B. bassiana strain ARP14 is less virulent to these parasitoids than the commercially available strain. Our results suggest that B. bassiana ARP14 may be a potential new biopesticide against R. pedestris with fewer negative effects on beneficial parasitoids than currently available options

Extract of Nicotiana tabacum as a potential control agent of Grapholita molesta (Lepidoptera: Tortricidae).

Oriental fruit moth, Grapholita molesta (Busck) (Lepidoptera: Tortricidae), is an important pest of stone and pome fruits. Growers usually depend on chemical insecticides to control this pest, but demand for more environmentally-friendly means of controlling pests is increasing. At least 91 plant extracts have been reported to be effective against other lepidopterans, but their acute toxicity against G. molesta has rarely been studied. Among these 91 materials, we assessed the residual toxicity of 32 extracts against first instar larvae (< 5 h old) of G. molesta in the laboratory. Nicotiana tabacum L., used at the concentration of 2 mg/ml, showed the highest corrected mortality (92.0%) with a lethal time (LT50) value of 12.9 h. The extract was followed in its efficacy by Allium sativum L. (88.0%), Zanthoxylum piperitum (L.) De Candolle (70.0%), and Sapindus mukorossi Gaertner (65.0%), when mortality was assessed at 20 h after exposure. Against adult fruit moths (< 5 d old), N. tabacum also showed the highest corrected mortality among tested extracts, being 85 and 100% in adult females and males, respectively, at 168 h after exposure. However, there was no synergistic effect of the combined application of any of the top four extracts in either laboratory or greenhouse assays. Oviposition by G. molesta on peach twigs was reduced 85-90% when N. tabacum was applied at 4 ml/ twig compared to control (methanol), demonstrating that N. tabacum may have potential for use as a botanical insecticide against G. molesta

Behavioral response of western flower thrips to visual and olfactory cues

Behavioral responses of Frankliniella occidentalis Pergande (Thysanoptera: Thripidae) to visual and olfactory cues were assessed in a cylindrical black box and a Y-tube olfactometer. Frankliniella occidentalis preferred circular shapes to other five geometrical patterns (rectangle, triangle, inverted triangle, diamond, and modified circle imitating flower petals) in multiple choice tests. In pair wise choice tests, the thrips preferred the yellow artificial flower shape to the geometrical patterns tested. Frankliniella occidentalis stayed on the artificial flower about four times longer than on the geometrical patterns. Higher numbers of thrips responded to a combination of p-anisaldehyde and artificial flower compared to the arm with only the olfactory or the visual cue. These results suggest that shape is an important cue for F. occidentalis.11 page(s

Circular yellow sticky trap with black background enhances attraction of Frankliniella occidentalis (Pergande) (Thysanoptera : Thripidae)

Yellow sticky flat traps were cut into six geometrical shapes (square, circle, semi-circle, diamond, triangle, and inverted triangle) to evaluate their attractiveness to Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae). The square was used as a standard to compare with other shapes in the laboratory. Circle was the most effective, attracting 2.7 and 1.5 times more thrips in choice and no-choice experiments, respectively. Four different background colors were also employed in the circular trap, and their efficacies were compared. In both choice and no-choice tests, the circular yellow sticky trap on a black background attracted a significantly higher number of thrips than on a yellow, blue, or green background. In addition, the higher ratio of a black background to yellow sticky foreground enhanced thrips' attraction in the choice test. Based on these results, the efficacy of circular yellow sticky traps (5 cm diameter) on a black background (12 cm wide×12 cm length) was compared to commercial yellow sticky traps (5 cm wide×8 cm length) in a strawberry greenhouse. The modified trap attracted 2.3–21 times more F. occidentalis than the commercial yellow sticky trap. Therefore, the modification of yellow sticky flat traps to a circular shape on a black background would be a good addition to the thrips management program.7 page(s

Annual pattern of occurrence of Riptortus pedestris (Hemiptera : Alydidae) and its egg parasitoids Ooencyrtus nezarae Ishii and Gryon japonicum (Ashmead) in Andong, Korea

Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) is one of the major pests of soybean (Glycine max [L.] Merr.) in Korea and Japan. The occurrence of two important egg parasitoids of R. pedestris, Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae) and Gryon japonicum (Ashmead) (Hymenoptera: Scelionidae), before and after soybean cultivation is poorly known. To record the seasonal occurrences of R. pedestris and parasitism during 2008–2010, aggregation pheromone traps baited with R. pedestris eggs were placed in a field in Andong, Korea where a series of crops, including barley (Hordeum vulgare L.), sesame (Sesamum indicum L.), and soybean, were cultivated during this study. R. pedestris was first recorded in the second week of April on barley and was most abundant in August on soybean. G. japonicum first appeared in May and was the dominant parasitoid until the first week of August on barley, sesame, and early soybean. From late August to September or October when soybean was cultivated, O. nezarae became the dominant parasitoid. The differential pattern of seasonal occurrence between the two parasitoids is discussed.6 page(s