Studies On The Intraspecific Differentiations Of Oligonychus Ununguis (Acari: Tetranychidae)

The spruce spider mite, Oligonychus ununguis (Acari: Tetranychidae), is the main pest mite of Chinese fir, Cunninghamia lanceolata, and chestnut, Castanea mollissima. Chinese fir is the timber tree species in southern China and chestnut is the economic tree species in northern China. According to the previous records, spruce spider mite can feed on both coniferous trees such as species in Pinaceae, Cupressaceae, Taxodiaceae and deciduous trees such as species in Fagaceae. However, Sun Xugen and his colleagues studied and found that the population on the conifer Cunninghamia lanceolata failed to complete its life history when raised on the broadleaf trees Castanea mollissima and Quercus acutissima. And all individuals from broadleaf trees died at nymphal stage when reared on conifers such as Cunninghamia lanceolata, Pinus thunbergii and P. firma etc. In addition, partial reproductive isolation was found between the fir population and the chestnut population. Have the conifer population and broadleaf population of O.ununguis formed different biotypes, subspecies, or different species because of long-term host isolation and/or geographical isolation? To determine the differentiation degree between the conifer population and the broadleaf population of O.ununguis, the morphological characteristics of individuals from chestnut and fir were observed carefully and their sequence variation of the second internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA and part of the mitochondrial cytochrome oxidase I gene (COI) and zymogram of esterase isozyme were analyzed. The crossing experiments between the two populations were also improved. In addition, the genetic differentiations in molecular and biochemical level between the other populations of O.ununguis were compared and the affecting factors were analyzed. The main results are as follows:1. The morphological characteristics of individuals from the chestnut population of Shandong Province and the fir population of Zhejiang Province were observed with the help of optical microscope and scanning electron microscope (SEM). By means of optical microscope, the differences of leg chaetotaxy between the two populations were discovered: adult female mites, Tarsus I with 3 tactile setae and 1 sensory seta proximal to duplex setae from the chestnut population, but 4 tactile setae and 1 sensory seta or 5 tactile setae and 1 sensory seta from the fir population; adult male mites, tarsus II with 3 tactile setae and 1 sensory seta or only 3 tactile setae proximal to duplex setae from the chestnut population, but 4 tactile setae and 1 sensory seta or 3 tactile setae and 2 sensory setae from the fir population. With the help of scanning electron microscope, differences between the chestnut population and the fir population in the aedeagus and the sensillum of palpal tarsus of male adults, the epidermis of genital flap and its anterior area of female adults and the empodium I of both males and females were discovered. Male aedeagus of the two populations were all curved ventrally, the hook of individuals from the chestnut population was long and slender, tapering gradually from the curved position to the terminal, but that from the fir population was shorter and finger-like, tapering sharply. The epidermis of genital flap of adult femal mites from the two populations were all latitudinal and the anterior area epidermis of genital flap were all longitudinal, but the texture was thick in the chestnut population and thin in the fir population. The terminal sensillum of palpal tarsus of male adults from the chestnut population was conical, while that of the fir population was terete. Empodium I of the two populations were all claw-like, that of the chestnut population with sharp terminal and stocky proximal,while that of the fir population was slender with a clear hooked terminal.2. Esterase isozyme of different populations of O.ununguis was examined by non-denaturing vertical slab polyacrylamide gel electrophoresis. Results showed that the electrophoretogram of esterase isozyme of all broadleaf populations was identical. There were three bands in the zymogram of the fir population and two of the bands were identical with broadleaf populations. The spruce population of Jilin Province had two bands, one of which was identical with broadleaf populations. The spruce population and the fir population had no shared bands. The similarity coefficient was 0.667, 0.4 and 0 respectively between the broadleaf populations and the fir population, the broadleaf populations and the spruce population and between the fir population and the spruce population.3. The complete sequence of ITS2 and part sequence of COI of individuals from different populations were sequenced directly. The sequence divergence and genetic distance calculated based on COI sequence between populations were different. The sequence divergence was 8.9%-9.2%, 10.3%-10.8% and 8.9%-9.5% respectively between the fir population and the spruce population, the fir population and the broadleaf populations, and between the spruce population and the broadleaf populations, which was much higher than the sequence divergence within population (0-0.3%) and the divergence between different broadleaf populations (0-1.7%). The genetic distance was 0.096, 0.111-0.113 and 0.095-0.099 respectively between the fir population and the spruce population, the fir population and the broadleaf populations, and between the spruce population and the broadleaf populations, which was much higher than the distance between different broadleaf populations (0-0.015). The length of ITS2 was 469 bp, 513 bp and 495bp respectively in the broadleaf populations, the fir population and the spruce population. The sequence divergence and genetic distance calculated based on ITS2 sequence between the fir population and the spruce population, the fir population and the broadleaf populations, and between the spruce population and the broadleaf populations were all far greater than the sequence divergence and genetic distance between different broadleaf populations. In the phylogenetic trees based on the sequences of COI and ITS2, all the examined broadleaf populations were clustered in the same branch with 100% bootstrap value, while the individuals from the fir population and the spruce population were clustered in two different branches respectively. In the phylogenetic tree based on COI sequences, all deciduous populations were divided into three branches. Inter-branch genetic distance was significantly greater than that of intra-branch and there were no shared haplotypes between populations in the three branches. Further analysis showed that geographical isolation and pesticide stress were possibly the important factors affecting deciduous population differentiation of O.ununguis, while the divergence in COI had no correlation with host plants.4. All the females in the cross experiments between the chestnut population and the fir population produced eggs and the average hatching rate of eggs was higher than 98%, but the offspring were all males, which showed that completely reproductive incompatibility was existed between the two populations.