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1.
Molecular mechanisms of monocrotophos resistance in the two-spotted spider mite (TSSM), Tetranychus urticae Koch, were investigated. A monocrotophos-resistant strain (AD) showed ca. 3568- and 47.6-fold resistance compared to a susceptible strain (UD) and a moderately resistant strain (PyriF), respectively. No significant differences in detoxification enzyme activities, except for the cytochrome P450 monooxygenase activity, were found among the three strains. The sensitivity of acetylcholinesterase (AChE) to monocrotophos, however, was 90.6- and 41.9-fold less in AD strain compared to the UD and PyriF strains, respectively, indicating that AChE insensitivity mechanism plays a major role in monocrotophos resistance. When AChE gene (Tuace) sequences were compared, three point mutations (G228S, A391T and F439W) were identified in Tuace from the AD strain that likely contribute to the AChE insensitivity as predicted by structure analysis. Frequencies of the three mutations in field populations were predicted by quantitative sequencing (QS). Correlation analysis between the mutation frequency and actual resistance levels (LC50) of nine field populations suggested that the G228S mutation plays a more crucial role in resistance (r2 = 0.712) compared to the F439W mutation (r2 = 0.419). When correlated together, however, the correlation coefficient was substantially enhanced (r2 = 0.865), indicating that both the F439W and G228S mutations may work synergistically. The A391T mutation was homogeneously present in all field populations examined, suggesting that it may confer a basal level of resistance.  相似文献   

2.
The toxicities of eight structurally different acaricidal compounds to six‐legged larvae (first motile stage) of three laboratory strains of the two‐spotted spider mite, Tetranychus urticae, and the European red mite, Panonychus ulmi, were evaluated following spray application. The larvae of five field‐derived strains of T urticae originating from France, Italy, Brazil, California and Florida were also tested for their susceptibilities to discriminating concentrations of several acaricides resulting in 95% mortality when applied to the organophosphate‐resistant laboratory reference strain WI. The spray bioassay used was robust and gave repeatable results with a wide range of acaricidal compounds, irrespective of their mode of action (ovo‐larvicides or primarily acting on motile life stages). Compounds tested were abamectin, azocyclotin, chlorpyrifos, clofentezine, deltamethrin, fenpyroximate, hexythiazox and pyridaben. Larvae of one of the laboratory strains of T urticae, AK, originally collected in Japan in 1996 and maintained without further selection pressure, exhibited 2000‐ and >4000‐fold resistance to the mitochondrial electron transport inhibitors pyridaben and fenpyroximate, respectively. Another strain of T urticae, AU, obtained from Australia and maintained in the laboratory under selection with hexythiazox and clofentezine since 1987 showed >770‐ and >1000‐fold resistance to clofentezine and hexythiazox, respectively. The same resistance pattern was observed against larvae of a laboratory strain of P ulmi, CE, also selected with hexythiazox. Larvae of one of the field‐derived strains of T urticae, BR, showed a lower susceptibility to a number of compounds, whilst the others were susceptible to all compounds except the organophosphates. © 2001 Society of Chemical Industry  相似文献   

3.
BACKGROUND: Heavy and constant use of organophosphorus (OP) larvicides selected Culex pipiens L. resistant populations through two main mechanisms of genetic resistance, the increased activity of detoxifying esterase and the production of alterate acetylcholinesterase‐1 (AChE1) by G119S mutation. The aim of this study was the assessment of the distribution of Cx. pipiens populations resistant to temephos and chlorpyrifos in the north‐eastern regions of Italy and the occurrence of the insensitive AChE in these populations. Data describe the situation in the last years before European legislation prohibited the use of OP larvicides in mosquito control, up until 2007. RESULTS: For the first time a high level of OP resistance in the samples from Ravenna (182‐fold, 80% A4/B4 or A5/B5 esterases and 38.3% Ester5), Emilia Romagna region, was detected; therefore, new data from the Veneto and Friuli Venezia Giulia regions were obtained and reinforced existing knowledge about resistance previously studied along the Adriatic coast. Nearby, in the Villa Verucchio locality, the highest (87.5%) AChE1R was found. CONCLUSION: Cx. pipiens resistance esterases A5/B5 and A4/B4 spread southward along the Adriatic coastal plain while OPs were being used in mosquito control, as confirmed by the first molecular screening of the AChE1 gene in these populations. Copyright © 2010 Society of Chemical Industry  相似文献   

4.
Insecticide resistant strains of the kanzawa spider mite, Tetranychus kanzawai, with insensitive AChE have spread widely throughout Japan. To clarify the molecular mechanism of this insensitivity, acetylcholinesterase (AChE) cDNA of the resistant strains of T. kanzawai was determined based on the AChE cDNA sequence of Tetranychus urticae and the sequences compared between the two spider mite species. The cDNA encoded 687 amino acids of AChE primary structure showing high homology to T. urticae. Amino acid homology indicated that the AChE is an Ace paralogous type of insect AChE. There were only three substitutions of amino acid residues between the AChEs of the two species. In the AChE of the resistant strain of T. kanzawai, one of the three amino acid substitutions was Phe439Trp, which lines the acyl pocket of the enzyme active site. Considering that the same substitution was found at the equivalent position of Ace paralogous AChE in the resistant strain of Culex tritaeniorhynchus, Phe439Trp substitution likely plays an important role in the insecticide insensitivity of the mite AChE.  相似文献   

5.
BACKGROUND: The tomato red spider mite, Tetranychus evansi (Baker and Pritchard), is a serious pest of solanaceous crops in many African countries. In this study an investigation has been conducted to establish whether mutation of the para‐type sodium channel underlies pyrethroid resistance in T. evansi strains collected in Southern Malawi. RESULTS: Two T. evansi strains from Malawi showed tolerance to the organophosphate chlorpyrifos and resistance (20–40‐fold) to the pyrethroid bifenthrin, but were susceptible to two contemporary acaricides (abamectin and fenpyroximate) in insecticide bioassays. Cloning of a 3.1 kb fragment (domains IIS5 to IVS5) of the T. evansi para gene from pyrethroid‐resistant and pyrethroid‐susceptible strains revealed a single non‐synonymous mutation in the resistant strains that results in an amino acid substitution (M918T) within the domain II region of the channel. Although novel to mites, this mutation confers high levels of resistance to pyrethroids in several insect species where it has always been associated with another mutation (L1014F). This is the first report of the M918T mutation in the absence of L1014F in any arthropod species. Diagnostic tools were developed that allow sensitive detection of this mutation in individual mites. CONCLUSION: This is the first study of pyrethroid resistance in T. evansi and provides contemporary information for resistance management of this pest in Southern Malawi. Copyright © 2011 Society of Chemical Industry  相似文献   

6.
The two-spotted spider mite Tetranychus urticae Koch is one of the most important pests of a wide range of crops worldwide. Its control is still largely based on the use of acaricides. However, due to its short life-cycle, high fecundity and arrhenotokous reproduction, it is able to develop resistance to these compounds very rapidly. Preliminary studies for milbemectin resistance in T. urticae showed that, under laboratory conditions, such resistance was unstable in the absence of a selection pressure. The aim of this study was to evaluate the possible fitness costs associated with milbemectin resistance in T. urticae. Comparison of biological traits between resistant and susceptible strains indicated the occurrence of fitness costs associated with milbemectin resistance. The resistant strain showed a longer juvenile development period for females, lower fecundity and a higher proportion of males among the progeny. The figures for net reproductive rate (R 0), intrinsic rate of natural increase (r m) and finite rate of increase (λ) were higher in the susceptible strain. The instability of milbemectin resistance can be exploited in resistance management programmes.  相似文献   

7.
BACKGROUND: A major problem of crop protection in Crete, Greece, is the control of Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae) with chemical insecticides owing to the rapid development of resistance. The aim of this study was to investigate the establishment of resistance and the underlying mechanisms to major insecticide classes with classical bioassays and known biochemical resistance markers. RESULTS: During a 2005–2007 survey, 53 Q biotype populations were collected. Application history records showed extensive use of neonicotinoids, organophosphates, carbamates and pyrethroids. High resistance levels were identified in the majority of populations (>80%) for imidacloprid (RF: 38–1958×) and α‐cypermethrin (RF: 30–600×). Low resistance levels (RF < 12) were observed for pirimiphos‐methyl. A strong correlation between resistance to imidacloprid and the number of applications with neonicotinoids was observed. Significant correlations were observed between COE and P450‐dependent monoxygenase activity with resistance to α‐cypermethrin and imidacloprid respectively. A propoxur‐based AChE diagnostic test indicated that iAChE was widespread in most populations. Resistance levels for α‐cypermethrin were increased when compared with a previous survey (2002–2003). Differentiation of LC50 values between localities was observed for imidacloprid only. CONCLUSION: Bemisia tabaci resistance evolved differently in each of the three insecticides studied. Imidacloprid resistance seems less established and less persistent than α‐cypermethrin resistance. The low resistance levels for pirimiphos‐methyl suggest absence of cross‐resistance with other organophosphates or carbamates used. Copyright © 2008 Society of Chemical Industry  相似文献   

8.
棉叶螨也称为棉红蜘蛛,属蛛形纲叶螨科,其种类繁多,分布范围广,世代周期短,是为害棉花的一类重要害螨。目前,用于防治棉叶螨的化学药剂主要是神经毒剂及呼吸抑制剂2大类,且棉叶螨对多数药剂产生了不同程度的抗性,以二斑叶螨Tetranychus urticae为首的植食性害螨已成为世界上抗药性最严重的节肢动物之一。美国路易斯安那州棉田二斑叶螨种群对阿维菌素产生了1 415倍抗性,而国内棉花上棉叶螨主要对有机磷类药剂产生了较强抗性,最高为467倍。棉叶螨产生抗药性的机制主要涉及靶标突变及解毒代谢增强,其中靶标突变主要涉及乙酰胆碱酯酶、电压门控钠离子通道和谷氨酸门控氯离子通道等;细胞色素P450单加氧酶、羧酸酯酶和谷胱甘肽S-转移酶等一种或多种解毒酶共同参与害螨对化学药剂的解毒代谢。该文主要从棉叶螨的种类及分布、用于防治棉叶螨的化学药剂、棉叶螨的抗药性现状、抗药性机制解析和抗药性治理策略5个方面进行阐述,提出因地制宜的抗药性治理策略,旨在为棉叶螨的田间防治提供指导。  相似文献   

9.
为了实现二斑叶螨Tetranychus urticae Koch对阿维菌素抗药性的快速检测,本研究建立了一种叶螨对阿维菌素抗药性的快速分子检测技术,即酶切扩增多态性序列(cleaved amplified polymorphic sequences,CAPS)标记技术,检测二斑叶螨谷氨酸门控氯离子通道(Glu Cl3)基因片段上G326E的突变频率,并采用琼脂浸叶法测定田间5个种群对阿维菌素的抗药性。结果表明,北京密云、顺义、昌平、浙江宁波和海南吉阳5个种群对阿维菌素的抗性倍数分别达1 155.63、1 317.10、844.19、314.95和1 799.69倍,均为极高抗性水平;CAPS检测发现我国二斑叶螨抗性基因存在G326E位点突变,但在供试的5个不同种群中突变频率变异很大,其中北京昌平种群G326E突变频率高达90.00%,密云和顺义种群均为26.67%,浙江宁波种群为5.00%,而海南吉阳种群不存在该位点的突变。表明该CAPS标记还需与其它抗性机制或检测技术结合来实现叶螨对阿维菌素抗性的早期检测和预警。  相似文献   

10.
A resistance management programme comparing rotations, mosaics and single use of insecticides for residual house-spraying against the insect vectors of malaria is being carried out in Southern Mexico. The area was chosen because of its prior history of insecticide use, relatively sedentary vector, and physical features of the area which limit inward migration of insects to the study area. A high level of resistance to DDT and low levels of organophosphorus (OP), carbamate and pyrethroid resistance were detected by WHO discriminating-dose assays in field populations of Anopheles albimanus in the pre-spray period in the region where this resistance management project is being undertaken. After the first year of spraying, resistance, as measured by a discriminating-dose assay, was still at a high level for DDT and had risen for all the other insecticides. Biochemical assays showed that DDT resistance was primarily caused by elevated levels of glutathione S-transferase (GST) activity leading to increased rates of metabolism of DDT to DDE. The numbers of individuals with elevated GST and DDT resistance were well correlated, suggesting that this is the only major DDT resistance mechanism in this population. The carbamate resistance in this population was conferred by an altered acetylcholinesterase (AChE) mechanism. The level of resistance in bioassays correlated well with the frequency of individuals homozygous for the altered AChE allele. This suggests that the level of resistance conferred by this mechanism in its heterozygous state is below the level of detection of the bioassay. The low levels of OP and pyrethroid resistance could be conferred by either the elevated esterase or monooxygenase enzymes. The esterases, however, are elevated only with p-nitrophenyl acetate (PNPA), and are unlikely to be causing broad-spectrum OP resistance. The altered AChE mechanism may also be contributing to the OP but not the pyrethroid resistance. There were significant differences in some resistance gene frequencies for insects obtained by different indoor and outdoor trapping methods. To determine whether the different sampling methods were effectively sampling the same interbreeding population, RAPD analysis of insects obtained by different collection methods in different villages was undertaken. There was no observed variability in the RAPD patterns for the different mosquito samples with a number of primers. ©1997 SCI  相似文献   

11.
The activity and sensitivity of acetylcholinesterase (AChE, EC 1.1.1.7) from an organophosphate-susceptible (OSS) and three resistant (OR-0, OR-1 and OR-2) strains of the greenbug (Schizaphis graminum) were biochemically compared. All three resistant strains displayed higher frequencies of individuals with respect to both increased activity of AChE toward the model substrate acetylthiocholine (ATC) and reduced sensitivity of AChE to inhibition by paraoxon as compared with the OSS strain. Kinetic study indicated that AChE from the OR-0, OR-1 and OR-2 strains had 3.3-, 2.7- and 2.3-fold, respectively, lower affinity, but 1.5-, 2.2- and 2.0-fold, respectively, higher catalytic activity toward ATC than AChE from the OSS strain. Significantly increased activity of AChE in the resistant strains was also confirmed by non-denaturing polyacrylamide gel electrophoresis, and appeared to be associated with the increase of general esterase activity. Inhibition kinetics revealed that AChE from the OR-0, OR-1 and OR-2 strains was 2.1-, 2.2- and 2.7-fold less sensitive to inhibition by paraoxon than that from the OSS strain. The study suggested that both qualitative and quantitative modifications of AChE had evolved in the resistant strains and were likely to significantly enhance the overall resistance level in greenbugs. © 1999 Society of Chemical Industry  相似文献   

12.
A biochemical approach was used to characterize acetylcholinesterase (AChE) insecticide insensitivity in several sweetpotato whitefly (Bemisia tabaci; SPW) populations. Discriminating doses of insecticide were established to differentiate between sensitive and insensitive SPW strains and to genotype individual whitefly. This technique was then used to examine the frequency of insensitive AChE alleles in several SPW populations and to isolate a line homozygous for insensitive AChE from a heterogenous B-type population. Inheritance of putative altered AChE genotypes was consistent with the proposed haplo-diploid status of B. tabaci. This biochemical diagnostic was also employed to determine the role of insensitive AChE in the observed resistance profiles of several laboratory populations subjected to different selection regimes. In keeping with previous studies on insecticide resistance in SPW, resistance does not appear to be uniquely associated with the B-type but rather with SPW populations found in crop systems. © 1998 SCI.  相似文献   

13.
Pyrethroid and organophosphate resistance-associated mutations have been recently reported in the whitefly Bemisia tabaci (Gennadius), a major pest of protected and outdoor crops worldwide. Here, we developed simple PCR–agarose gel visualization based assays for reliably monitoring the L925I and T929V pyrethroid resistance mutations in the B. tabaci para-type voltage gated sodium channel and the iAChE F331W organophosphate resistance mutation in the acetylcholinesterase enzyme ace1.PCR-RFLP assays were developed for detecting the L925I and the F331W resistance mutations. A highly specific PASA was developed for detecting the T929V mutation. The molecular diagnostic tools were used to monitor the frequency of the resistance mutations in a large number of field caught Q biotype B. tabaci from Crete (Greece), where both organophosphates and pyrethroids are extensively used. The F331W mutation was fixed in all field individuals examined. The pyrethroid resistance mutations were detected in high frequencies: 0.38 and 0.54 for L925I and T929V, respectively. The simple diagnostics are accurate and robust, to be used alongside classical bioassays to prevent ineffective insecticide applications, and for early identification of the spreading of resistant Q biotype populations into new regions around the globe.  相似文献   

14.
Strains of sheep louse Bovicola ovis (Schrank) with various levels of resistance to pyrethroid and one strain with high degree of resistance to organophosphate (OP) insecticides were used to investigate the biochemical mechanisms of insecticide resistance, i.e., enhanced levels of general esterases, specific acetylcholinesterases (AChE), glutathione S-transferase (GST), and mixed function oxidases. Native gel electrophoresis combined with quantitative enzyme assays showed analogous expression profiles of several esterase isozymes in all the strains tested. The determination of the sensitivity of each esterase isozyme to five inhibitors (acetylthiocholine iodide, butyrylthiocholine iodide, paraoxon eserine sulfate, and pCMB) led to the identification of nine esterases in the B. ovis strain. Gel electrophoresis results are supported by enzyme assay studies where, except for the OP resistant strain, no differences in esterase activities were detected in all the pyrethroid resistant and susceptible strains assayed. Statistical analyses demonstrated that some strains have elevated GST activities compared to the susceptible reference strain.  相似文献   

15.
BACKGROUND: Laodelphax striatellus (Fallén) is a major pest of cultivated rice and is commonly controlled in China with the organophosphate insecticides. To develop a better resistance management strategy, a chlorpyrifos‐resistant strain of L. striatellus was selected in the laboratory, and its cross‐resistance to other insecticides and possible mechanisms of the chlorpyrifos resistance were investigated. RESULTS: After 25 generations of selection with chlorpyrifos, the selected strain of L. striatellus developed 188‐fold resistance to chlorpyrifos in comparison with the susceptible strain, and showed 14‐ and 1.6‐fold cross‐resistance to dichlorvos and thiamethoxam respectively. There was no apparent cross‐resistance to abamectin. Chlorpyrifos was synergised by the inhibitor triphenyl phosphate; the carboxylesterase synergistic ratio was 3.8 for the selected strain, but only 0.92 for the susceptible strain. The carboxylesterase activity of the selected strain was approximately 4 times that of the susceptible strain, whereas there was no significant change in the activities of alkaline phosphatase, acid phosphatase, glutathione S‐transferase and cytochrome P450 monooxygenase between the strains. The Michaelis constant of acetylcholinesterase, maximum velocity of acetylcholinesterase and median inhibitory concentration of chlorpyrifos‐oxon on acetylcholinesterase were 1.7, 2.5 and 5 times higher respectively in the selected strain. CONCLUSION: The high cross‐resistance to the organophosphate dichlorvos in the chlorpyrifos‐resistant strain suggests that other non‐organophosphate insecticides would be necessary to counter resistance, should it arise in the field. Enhanced activities of carboxylesterase and the acetylcholinesterase insensitivity appear to be important mechanisms for chlorpyrifos resistance in L. striatellus. Copyright © 2010 Society of Chemical Industry  相似文献   

16.
Molecular mechanisms of carbofuran resistance in the brown planthopper, Nilaparvata lugens Stål, were investigated. A carbofuran-resistant strain (CAS) showed approximately 45.5- and 15.1-fold resistance compared with a susceptible strain (SUS) and a non-selected field strain (FM), respectively. Activities of the esterase and mixed-function oxidase were approximately 2.8- and 1.6-fold higher, respectively, in the CAS strain than in the SUS strain, suggesting that these enzymes play a minor role in carbofuran resistance. Interestingly, the insensitivity of acetylcholinesterase (AChE) to carbofuran was approximately 5.5- and 3.7-fold higher in the CAS strain compared to the SUS and FM strains, respectively, indicating that AChE insensitivity is associated with carbofuran resistance. Western blot analysis identified two kinds of AChEs, of which the type-1 AChE (encoded from Nlace1, which is paralogous to the Drosophila AChE gene) was determined to be the major catalytic AChE in N. lugens. The open reading frame of Nlace1 is composed of 1989 bp (approximately 74 kD) and revealed 52.5% and 24.3% amino acid sequence identities to those of Nephotettix cincticeps and Drosophila melanogaster, respectively. Screening of point mutations identified four amino acid substitutions (G119A, F/Y330S, F331H and H332L) in the CAS strain that likely contribute to AChE insensitivity. The frequencies of these mutations were well correlated with resistance levels, confirming that they are associated with reduced sensitivity to carbofuran in N. lugens. These point mutations can be useful as genetic markers for monitoring resistance levels in field populations of N. lugens.  相似文献   

17.
Nine different strains of the two-spotted spider miteTetranychus urticae Koch (Acari: Tetranychidae) were collected on cotton from Adana, Antalya, Izmir, Manisa and Urfa in Turkey. Their responses to bifenthrin were investigated using conventional bioassay and biochemical assays. LC50 and LC90 values of bifenthrin were determined for all strains by using a residual bioassay with a petri dish-spray tower. Resistance ratios were determined by comparing the samples with a standard susceptible strain, GSS. The resistance ratios of the strains ranged from <1 to 669-fold (at LC50). Of the investigated field strains, only three (two from Adana and one from Urfa) were resistant to bifenthrin. There was a correlation between esterase enzyme activity and bifenthrin resistance according to polyacrylamide gel electrophoresis and microtiter plate assays in the three resistant strains. http://www.phytoparasitica.org posting May 17, 2005.  相似文献   

18.
In this paper we analysed the basis of insecticide resistance in 59 Italian strains of the peach potato aphid Myzus persicae using both molecular and biochemical assays. Our data as a whole clearly indicate that most M. persicae strains (76.3%) have high or extremely high production of an esterase enzyme which sequester and detoxify insecticides with esteric group. Kdr genotypes conferring resistance towards pyrethoids are present in 57.7% of the analysed populations. Moreover, 26.5% of the kdr positive strains possess also the M918T mutation conferring super-kdr phenotype. Strains with modified AChE (MACE) are not so numerous (27.1%), although they can be found almost everywhere in Italy. Considering all the strains analysed, both MACE and kdr phenotypes are associated with high levels of esterase activity. In Central–Southern regions, kdr and MACE resistance mechanisms resulted in linkage disequilibrium. Bioassays performed in order to evaluate the efficacy of a pyrethroid insecticide against a strain possessing a F979S mutation within its para-type sodium channel gene suggests that this amino acid substitution could affect the sodium channel responsivity to pyrethroids.  相似文献   

19.
A strain (R) of Aphis gossypii from Southern France was found to be resistant to several insecticides, particularly to pirimicarb, as compared to a susceptible strain (S). Resistance levels were determined by biological tests, and the highest resistance factor (1350) was for pirimicarb. Resistance was mainly restricted to anticholinesterase inhibitors. Use of synergists, DEF and PB, suggested that resistance mechanisms based on detoxification were involved to a minor extent, since a good correlation was observed between I50 values and ki values of AChE and in-vivo bioassay data. The two strains differed in esterase activity, with a 27·7-fold increase in the R strain. Resolution of esterases by polyacrylamide gel electrophoresis showed different patterns in the S and R strains, and two isozymes were less sensitive to pirimicarb in the S strain; however, no in-vitro degradation of [14C]pirimicarb was observed. These data suggest that the main mechanism of resistance was through a decrease in the sensitivity of the target, AChE, to the insecticides. © 1997 SCI.  相似文献   

20.
Han J  Kim SI  Choi BR  Lee SG  Ahn YJ 《Pest management science》2011,67(12):1583-1588
BACKGROUND: This study was aimed at assessing the fumigant toxicity of 14 essential oil constituents from lemon eucalyptus, Eucalyptus citriodora Hook, and another ten known compounds to females of acaricide‐susceptible, chlorfenapyr‐resistant, fenpropathrin‐resistant, pyridaben‐resistant and abamectin‐resistant strains of Tetranychus urticae Koch. RESULTS: Menthol (LC50, 12.9 µg cm?3) was the most toxic compound, followed by citronellyl acetate (16.8 µg cm?3), against the susceptible females. High toxicity was also produced by β‐citronellol, citral, geranyl acetate and eugenol (LC50, 21.7–24.6 µg cm?3). The fumigant toxicity of these compounds was almost identical against females from either of the susceptible and resistant strains, indicating that the compounds and acaricides do not share a common mode of action or elicit cross‐resistance. CONCLUSION: Global efforts to reduce the level of highly toxic synthetic acaricides in the agricultural environment justify further studies on materials derived from lemon eucalyptus oil, particularly menthol and citronellyl acetate, as potential acaricides for the control of acaricide‐resistant T. urticae as fumigants with contact action. Copyright © 2011 Society of Chemical Industry  相似文献   

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