浙江省五种实蝇记述

报道了中国浙江省分布的5种实蝇，即：瓜实蝇Bactrocera（Zeugodaeus）cucurbitae（Coquillett）、南亚果实蝇胁。Bactrocera（Z.）tau（Walker）、宽带果实蝇Bactrocera（Z.）scutellata（Hendel）、日本拟羽角实蝇Paragastrozona japonica（Miyake）、秀长痣实蝇Acidiostigma postsignata（Chen）。对这5种实蝇特征进行了详细描述并附有各种前翅照片。所有实蝇标本保存在浙江大学农业与生物技术学院应用昆虫研究所。     

桔小实蝇等五种实蝇的SSR鉴定

桔小实蝇Bactrocera (Bactrocera) dorsalis (Hendel)是一种为害多种水果、瓜类、茄类蔬菜的检疫性害虫,食性杂、寄主范围广泛,直接威胁水果和蔬菜的安全生产.采用改进的CTAB法提取单头实蝇的基因组DNA,从20对引物中筛选出2对重复性好、多态性高的引物,运用微卫星标记(SSR)技术,初步构建了桔小实蝇和重庆市常见南亚果实蝇B.(Zeugodacus) tau (Walker)、宽带果实蝇B.a(Z.) scutellata (Hendel)、瓜实蝇 B.(Z.) cucurbitae (Coquillett)、黑颜果实蝇B.(Z.) diaphora (Hendel) 5种实蝇的指纹图谱.通过这些图谱可容易区分重庆市常见4种实蝇和桔小实蝇,实现了采用2对引物1次扩增的简单快速有效方法将桔小实蝇与其他4种实蝇区分开.这为重庆市桔小实蝇非疫区的建设提供了快速、准确的鉴定方法.     

云南边境检疫性实蝇风险分析研究

近年来,云南边境口岸检验检疫机关在进境水果和蔬菜中检疫截获实蝇害虫17种,大部分属国家禁止传入的危险性害虫。从截获的批次和数量来看,优势种类有桔小实蝇（Bactrocera dorsalis Hendel)、瓜实蝇（Bactrocera cucurbitae （Coquillett)和南瓜实蝇（Bactrocera tau (Walker)等。针对缅甸、越南、老挝等国家的实蝇种类、云南边境情况、水果和蔬菜类型,结合实蝇的生物学习性,分析实蝇在云南的适生区域和非适生区,并提出检疫措施。     

台州果蔬混栽橘园用性诱剂监测实蝇种群动态及效果评价

在台州黄岩果蔬混栽园内,通过诱蝇醚（Methyl eugenol,Me）和诱蝇酮（Cuelure,Cue）性诱剂监测鉴定出4种果蔬有害实蝇：柑橘小实蝇（Bactrocera dorsalis Hendel）、南瓜实蝇（Bactrocera tau Walker）、具条实蝇（Bactrocera scutellata Hendel）、瓜实蝇（Bactrocera cucurbitae Coquillett）。柑橘挂果期内4种实蝇的群落动态监测结果表明：6-8月期间,4种实蝇种群数量较低;9-11月期间,橘小实蝇种群数量分别在9月中旬历经小高峰、10月下旬达到大高峰;南瓜实蝇种群数量则在10月下旬达到最大值;具条实蝇种群数量则在9月下旬达到最高峰;瓜实蝇种群数量相对较少,在9月下旬和10月下旬存在2个小高峰。另外,比较评价了Cue和ZG1引诱剂在低温月份（11-12月份）对南瓜实蝇的引诱能力,结果表明：在低温月份,ZG1引诱剂引诱能力明显强于Cue（南瓜实蝇引诱剂）。由此推测,Cue在台州地区低温月份用于监测种群的准确性较差。     

云南边境检疫性实蝇风险分析研究

近年来,云南边境口岸检验检疫机关在进境水果和蔬菜中检疫截获实蝇害虫17种,大部分属国家禁止传入的危险性害虫.从截获的批次和数量来看,优势种类有桔小实蝇(Bactrocera dorsalis Hendel)、瓜实蝇(Bactrocera cucurbitae(Coquillett)和南瓜实蝇(Bactrocera tau(Walker)等.针对缅甸、越南、老挝等国家的实蝇种类、云南边境情况、水果和蔬菜类型,结合实蝇的生物学习性,分析实蝇在云南的适生区域和非适生区,并提出检疫措施.     

Phylogenetic Relationships of Sorghum and Related Species Inferred from Sequence Analysis of the nrDNA ITS Region

Analysis of phylogenetic and evolution in six species of Sorghum was based on internal transcribed spacer （ITS） sequences in nuclear ribosomal DNA. Results showed that the length of the ITS regions among the six species ranged from 588 to 589 bp and the contents of G＋C in ITS （ITS1 ＋5.8S＋ITS2） regions ranged from 60.27 to 61.05%; the length of ITS1 ranged from 207 to 208 bp and the contents of G ＋ C in the ITS 1 regions ranged from 53.91 to 61.54%. The length of the 5.8S rDNA and ITS2 regions in the six species was 164 and 217 bp respectively, and the contents of G ＋ C ranged from 56.10 to 58.54% in the 5.8S rDNA region and 66.36 to 67,28% in the ITS2 region. Among regions of ITS, ITS1, ITS2, and 5.8S, the best sequence for genetic relationship analysis in the six species was the ITS region. On the basis of the Jaccard coefficient and phylogentic tree, S. sp. was more related to S, propinguum than to other species. This was consistent with the fact that S. sp. is derived from S. propinguum. From the phylogenetic tree based on ITS1, S. halepense, silk sorghum and S. sudanense, are identical in the ITS 1 sequence, whereas the phylogenetic tree based one shows that S. sudanense has a closer genetic association with S. almum rather than with S. halepense and silk sorghum.     

Au Elements and Their Evolution in Some AIIopolyploid Genomes of Aegilops

To study the sequences of short interspersed nuclear elements （SINEs） evolution in some allopolyploid genomes of Aegilops, 108 Au element fragments （a novel kind of plant SINE） were amplified and sequenced in 10 species of Aegilops, which were clustered into three different groups （A, B and C） based on their related geuome types. The sequences of these Au element fragments were heterogouous in di-, tetra-, and hexa-ploids, and the deudrograms of Au element obtained from phylogenetic analysis were very complex in each group and could be clustered into 15, 15 and 22 families, respectively. In this study, three rules about Au elements evolution have been drawn from the results： i. Most families were composed of Au element members with different host species in three groups; ii. Family 1-6 in Group A, Family 1-6 in Group B, Family 1-4 and Family 6-13 in Group C contained only one, apparently highly degenerate Au dement member （a single representative elemeut）; iii. Elements generally fell into clades that were species-specific with respect to their host species. The potential mechanisms of Au element evolution in Aegilops were discussed.     

Molecular Taxonomy of Conogethes punctiferalis and Conogethes pinicolalis（Lepidoptera： Crambidae） Based on Mitochondrial DNA Sequences

Conogethes punctiferalis（Guenée）（Lepidoptera： Crambidae） was originally considered as one species with fruit-feeding type（FFT） and pinaceae-feeding type（PFT）, but it has subsequently been divided into two different species of Conogethes punctiferalis and Conogethes pinicolalis. The relationship between the two species was investigated by phylogenetic reconstruction using maximum-likelihood（ML） parameter estimations. The phylogenetic tree and network were constructed based upon sequence data from concatenation of three genes of mitochondrial cytochrome c oxidase subunits I, II and cytochrome b which were derived from 118 samples of C. punctiferalis and 24 samples of C. pinicolalis. The phylogenetic tree and network showed that conspecific sequences were clustering together despite intraspecific variability. Here we report the results of a combined analysis of mitochondrial DNA sequences from three genes and morphological data representing powerful evidence that C. pinicolalisand C. punctiferalis are significantly different.     

Comparative Analysis of Population Genetic Structure in Bemisia tabaci （Gennadius） Biotypes B and Q Based on ISSR Marker

Bemisia tabaci （Gennadius） biotypes B and Q are two invasive biotypes in the species complex. The comparison of the population genetic structure of the two biotypes is of significance to show their invasive mechanism and to their control. The intersimple sequence repeats （ISSR） marker was used to analyze the 16 B-biotype populations and 4 Q-biotype populations worldwide with a Trialeurodes vaporariorum population in Shanxi Province, China, and a B. tabaci non-B/Qbiotype population in Zhejiang Province, China, was used as control populations. The analysis of genetic diversity showed that the diversity indexes of biotype Q including Nei＇s gene diversity index, Shannon informative index, and the percentage of polymorphic loci were higher than those of biotype B. The high genetic diversity of biotype Q might provide the genetic basis for the excellent ecological adaptation. Cluster analysis suggested that the ISSR could not be used in the phylogenetic analysis though it could easily distinguish the biotypes of B. tabaci. The difference of the population genetic structure between the biotype B and the biotype Q exists based on the ISSR marker. Meanwhile, the results suggested that the molecular marker has its limitation in the phylogenetic analysis among the biotypes of B. tabaci.     

Sequence Analysis of mtDNA COIGene and Molecular Phylogeny of Different Geographical Populations of Bemisia tabaci （Gennadius）

Bemisia tabaci(Gennadius) is a serious pest in many cropping systems worldwide and occurs in different biotypes. The mtDNA COIgene of the 12 Bemisia tabaci (Gennadius) populations from different regions and countries were analyzed. Based on mtDNA COI sequences, their biotypes were characterized and phylogenetic relationships among these populations were established with the method of UPGMA. The results indicated the genetic similarity between those populations from Beijing, Zhengzhou, Zaozhuang, Nanjing, Shanghai, Haikou, and the B-biotype populations from California, Texas, Arizona reached 99.8-100%, which meant the nation-wide infested populations of B.tabaci in China in recent years were B-biotypes. Another population collected from Kunming of Yunnan Province showed very high similarity with Q-biotype B.tabaci from Spain and Morocco, which meant the Kunming population was Q-biotype. This is the first report on the invasion of Q-biotype into China.     

Sequence Comparison of MHC Class Ⅱβ（Exon 2） and Phylogenetic Relationship Between Poultry and Mammalian

A fragment spanning over exon 2 and intron 2 of major histocompatibility complex B-LB Ⅱ genes was amplified using PCR,cloned and sequenced in 13 individuals from eight Chinese indigenous chicken breeds and one introduced breed. Another 41 sequences of MHC class Ⅱβ from ten vertebrate species were cited from the NCBI GenBank. Thirteen new B-LB Ⅱ alleles were found in the chicken breeds sampled. Alignment of the exon 2 sequences revealed 91.1-97.8% similarity to each other within the chickens sampled, and the chickens shared 84.1-87.0% homology to Phasianus colchicus, 78.5-81.5% similarity to Coturnixjaponica. The sequences in poultry showed 62.6-68.1% identity to HLA-DRB1, 50-61.5% similarity to DQB (HLA-, SLA- and H2-BB), 53.7-60% to HLA-DPB and 53.3-57.8% similarity to HLA-DOB. The frequency of nonsynonymous substitutions of nucleotide was higher than that of synonymous substitutions, and the frequencies of nonsynonymous and synonymous substitutions in poultry B-LB Ⅱ genes were lower than those observed in mammalian DRB1 and DQB1 genes. The deduced amino acid sequences of MHC class Ⅱ β1 domain exhibited extreme difference in conversed region and variable region patterns among the various species, but the two conserved cysteines forming disulfide-bond were shown consistent in poultry with that in mammalian species; and the carbohydrate attachment site was found more conserved in chicken, Homo sapiens, Bos taurus, Ovis aries and Capra hircus than in Sus scrofa and rodent animals. Compared with exon 2 of DQB1 genes of Homo sapiens, ruminant species and Sus scrofa, the differentia that the deletion of six nucleotides at position195 to 200 of exon 2 of DQB1 genes, and insertion of three nucleotides at position 247 to 249 of the exon 2 existed in rodent species were found, which led to the absence of three AA residues at position 65, 66,and 67 within β1 domain of DQB1 chain, and the insertion of one AA residue at position 85. The difference of the deletion of six nucleotides at position 72 to 77 of exon 2 of DPBI genes was observed with Homo sapiens DQB1, which caused absence of three AA residues at position 24, 25, and 26 of β1 domain of DPB1 chain. The phylogenetic tree revealed that the B-LB Ⅱ sequences from poultry are not orthologous to the class Ⅱ MHC β-chain genes of mammalian species. The tree indicated that genetic evolutionary relationship of chickens with Phasianus colchicus was much closer than with Coturnix japonica, and the DQB and DPB clusters are more tightly related to each other than to the remaining clusters.     

Analysis of Simple Sequence Repeats in Genomes of Rhizobia

Simple sequence repeats （SSRs） or microsatellites, as genetic markers, are ubiquitous in genomes of various organisms. The analysis of SSR in rhizobia genome provides useful information for a variety of applications in population genetics of rhizobia. We analyzed the occurrences, relative abundance, and relative density of SSRs, the most common in Bradyrhizobium japonicum, Mesorhizobium loti, and Sinorhizobium meliloti genomes se- quenced in the microorganisms tandem repeats database, and SSRs in the three species genomes were compared with each other. The result showed that there were 1 410, 859, and 638 SSRs in B. japonicum, M. loti, and S. meliloti genomes, respectively. In the genomes of B. japonicum, M. loti, and S. meliloti, tetranucleotide, pentanucleotide, and hexanucleotide repeats were more abundant and indicated higher mutation rates in these species. The least abundance was mononucleotide repeat. The SSRs type and distribution were similar among these species.     

应用SS-PCR技术设计种特异性引物快速鉴定瓜实蝇

[目的]为了促进果蔬进出口贸易和快速通关,防止瓜实蝇[Bactrocera cucurbitae(Coquillett)]进一步传入扩散,迫切需要开展适合实蝇快速鉴定技术的研究。[方法]应用SS-PCR技术,基于mtDNA COⅠ序列,设计了1对能够准确鉴定瓜实蝇的种特异性引物GF85和GR531。选用瓜实蝇作为阳性对照,南瓜实蝇[B.tau(Walker)]、具条实蝇[B.scutellata(Hendel)]等其他19种待测实蝇作为阴性对照,提取供试虫样基因组DNA,并进行PCR扩增和电泳检测。[结果]仅虫样瓜实蝇能在约447 bp位置扩增出一条清晰且单一的条带,其余的虫样未出现条带。[结论]将该试验建立的SS-PCR鉴定方法应用于实际进出境果蔬截获和疫情监测的虫样的鉴定,并得到验证,表明该方法具特异性、稳定性强,可在实际的实蝇鉴定工作中应用。     

Stem Borer Species Composition on Maize and Two Non-Cereal Hosts in the Forest Zone of Kisangani,DRC

Lepidopteran stem borers are the most damaging pests of maize in Sub-Saharan Africa. Despite the growing importance of maize in the forest zone of Democratic Republic of Congo, no data is available regarding stem borer pest species present and their relative importance. It is thus important to gather information likely to guide future research in this area. This study was undertaken to catalogue stem borer pest species identity and assess their relative infestation levels on maize. Surveys were carried out in wild and cultivated habitats in Kisangani. Five species were collected on maize, i.e., Sesamia calamistis Hampson （1910）, Eldana saccharina Walker （1865）, Busseola fusca Fuller （1901）, Chilo sp. Strand （1913）, and Mussidia nigrivenella Ragonot （1888）. In the wild habitats, Poenoma serrata Hampson, B. fusca and S. calamistis were collected on Pennisetum purpureum whereas Chilo sp. was collected on Panicum maximum. Our results suggest that P. maximum might affect the population dynamics of Chilo sp. whereas P. purpureum is expected not to influence the population dynamics of other stem borers owing to its scarcity in the interior of the forest.     

Immunocytochemical Identification and Localization of Diffuse Neuroendocrine System （DNES） Cells in Gastrointestinal Tract of Channel Catfish （Ictalurus punctatus）

To detect distribution and relative frequency of diffuse neuroendocrine system （DNES） cells in the gastrointestinal tract of channel catfish （lctalurus punctatus）, the intestinal tract of channel catfish was divided into seven portions from proximal to distal： the enlarged area after oesophagus, cardia, fundus, pylorus, and anterior, middle, and posterior intestine. Immunohistochemical method using the strept avidin-biotin-complex （SABC） was employed. All antisera between seven portions of the channel catfish were compared statistically using statistical package for the social science （SPSS）. Five types of DNES ceils were determined： neuropeptide Y-immunoreactive （NPY-IR） cells were demonstrated in both anterior and middle intestine; serotonin （5-HT） immunoreactive cells were detected throughout the whole gastrointestinal tract; vasoactive intestinal peptide （VIP） positive cells were at the highest frequency in pylorus; glucagon-immunoreactive （GLU-IR） cells were moderate in number in the fundus and anterior, middle intestine, and no immunoreactivity was determined in the other portions; somatostatin （SOM） positive cells were more abundant in the anterior and middle intestine. The regional distribution and relative frequency of immunoreactive cells in the channel catfish, Ictalurus punctatus, are essentially similar to those of other fish. However, some characteristics are observed in this species, which further proved that the diversity of the physiological function of DNES cells was based on their morphology.     

Cloning and Sequence Analysis of Prophenoloxidase from Haemocytes of the Red Swamp Crayfish, Procambarus clarkii

The full-length cDNA sequence of prophenoloxidase was obtained through RACE technology. The complete cDNA sequence is 3 721-bp long, containing an open reading frame （ORF） of 1 881 bp, a 154-bp 5′-untranslated region, and a 1 686- bp 3′-untranslated region with three potential functional poly（A） signals （AATAAA）. The molecular mass of the deduced amino acid sequence （627 aa） was 72.3 kDa with an estimatedpI of 5.88. It contained putative copper-binding sites （copper A： 131, 135, 167 and copper B： 301,305, 341）, and a tentative complement-like motif （GCGWPDHL）. Eight potential N-linked glycosylation sites were predicted to be present in P. clarkii prophenoloxidase. Similar to those in other arthropod prophenoloxidases reported so far, no signal peptide was detected in the crayfish prophenoloxidase. The phylogenetic trees confirmed that P. clarkii prophenoloxidase was most closely related to that of freshwater crayfish P. leniusculus and more closely related to other crustacean prophenoloxidases from shrimp, prawn, and lobster than to the insect prophenoloxidases. Besides, two putative introns were found in this sequence of genomic DNA.     

The Mining of Citrus EST-SNP and Its Application in Cultivar Discrimination

Single nucleotide polymorphisms （SNPs） are the most abundant sequence variations found in plant genomes and are widely used as molecular genetic markers in cultivar identification and genetic diversity studies. The objective of this study was to identify SNP markers useful for discrimination of citrus cultivars, since large numbers of expressed sequence tags （ESTs） of sweet orange are available from the National Center for Biotechnology Information （NCBI）. We now have the opportunity to discover SNP markers suitable for determining the haplotypes with which to distinguish very closely related cultivars and to assess genetic diversity within or between related species of citrus. SNPs and small insertions/deletions （Indels） from ESTs of sweet orange and satsuma were identified by the in silico SNP discovery strategy. 55 296 EST sequences of sweet orange and 2 575 of satsuma retrieved from the NCBI repository were mined for potential SNPs. Cleaved amplified polymorphic sequences （CAPS） and sequencing approaches were used to validate putative SNPs in a sample of 30 citrus accessions. A total of 3 348 putative SNPs were identified based on the abundance of sequences and haplotype cosegregation. Of these 3 348 SNPs, the transitions, transversions and Indels ratios were 47.9, 36.1 and 16.0%, respectively. The SNPs occurred on average at a frequency of 1 per 164 bp in the coding region of citrus. 14 SNPs were randomly selected and genotyped according to 30 citrus accessions including 23 accessions of sweet orange; 11 SNPs displayed polymorphism with an average polymorphism information content （PIC） of 0.20 among 30 citrus accessions. The genetic diversity present in sweet orange was low, so the 14 SNP markers failed to discriminate different cultivars of sweet orange, but they did succeed in distinguishing accessions of inter-species of citrus. In this study, SNPs were mined from EST sequences of sweet orange and satsuma, which displayed potential capability as molecular markers to discriminate inter-species accessions of citrus. It is anticipated that these putative SNPs could be applied in citrus genetics research and breeding.     

福建省实蝇种类监测和寄主初步调查

实蝇监测结果表明,福建省的实蝇种类有4种,包括桔小实蝇[Bactrocera(Bactrocera)dorsalis]、瓜实蝇[B.(zeugoda-cus)cucurbitae]、南瓜实蝇[B.(zeugodacus)tau]、具条实蝇(B.scutellata)。调查发现,福建省桔小实蝇寄主有35种,瓜实蝇寄主有26种,南瓜实蝇寄主有18种,有16种是这3种实蝇的共同寄主。     

The Midgut Bacterial Flora of Laboratory-Reared Hard Ticks,Haemaphysalis longicornis,Hyalomma asiaticum,and Rhipicephalus haemaphysaloides

Ixodid ticks play an important role in the transmission of a variety of zoonoses of viral, bacterial and protozoan origin, and they also harbor a wealth of microorganisms. To gain more detailed insights into the potential interactions between bacterial flora and tick-borne pathogens, we investigated the midgut bacterial flora of laboratory-reared Haemaphysalis longicornis, Hyalomma asiaticum and Rhipicephalus haemaphysaloides. Based on morphological, biochemical, and 16 S rDNA sequencing results, we identified 15 species belonging to 12 genera in the midgut of the three ticks. The bacterial communities were similar to those found in other studies of hematophagous arthropods. Kocuria sp. was the most frequently isolated species and its 16 S rDNA gene sequence was very similar to Kocuria koreensis P31 T. To our knowledge, this is the first report of the bacterial flora of tick midguts and the results show that there were many different bacterial species in each tick species. Among the most common genera, there may have been a novel species in the genus Kocuria. The results might be the first step for looking for different aspects of the pathogen and tick interaction.     

The mRNA Expression Profiles of Five Heat Shock Protein Genes from Frankliniella occidentalis at Different Stages and Their Responses to Temperatures and Insecticides

The westem flower thrips, Frankliniella occidental＆ （Pergande） is a highly invasive pest that is able to exploit many crops across a wide range of environmental conditions. Five full-length cDNAs of heat shock protein （HSP） genes （Fo-HSP90, Fo-HSP70, Fo-HSP60, Fo-HSP40 and Fo-HSP28.9） were cloned from F. occidentalis, and their expression profiles were investigated under conditions of thermal stress and insecticide exposure, and at different stages during development, using real-time quantitative PCR. All five gene sequences showed high similarity to homologs in other species, indicating the conserved fimction of this gene family. HSP60 represents an informative phylogenetic marker at the ordinal taxonomic level within Insecta, but HSP90, which has two homologous copies in Hymenoptera, was not informative. The expression of Fo-HSPs under thermal stress suggests that Fo-HSP90, Fo-HSP70, and Fo-HSP28.9 are inducible by both cold and heat stress, Fo-HSP40 is only heat-inducible, and Fo-HSP60 is thermally insensitive. There were two patterns of cold induction of Fo-HSPs： one is from 0 to 4℃ and the other is around -8℃. All five Fo-HSPs genes were induced by exposure to sublethal concentrations of the insecticide avermectin. The expression of the five Fo-HSPs during different developmental stages suggests that they all play a role in development of F. occidentalis.