从东南亚进口水果截获桔小实蝇复合种五近缘种的鉴别(双翅目:实蝇科)

介绍了近年从进口东南亚水果中经常截获的、最具经济重要性的桔小实蝇复合种5个近缘种:杨桃实蝇B.carambolae、桔小实蝇B.dorsalis、芒果实蝇B.occipitalis、木瓜实蝇B.papayae及菲律宾实蝇B.philippinensis,分别记述了翅、胸、足、腹及雌虫产卵器主要鉴别特征,并列出桔小实蝇复合种5近缘种鉴别特征检索表。     

Rapid identification of Bactrocera latifrons (Dipt., Tephritidae) by real-time PCR using SYBR Green chemistry

Abstract:  The solanum fruit fly, Bactrocera latifrons (Hendel), is a major agricultural pest in Asia and Hawaii, and it is important to prevent its widespread invasion in plant quarantine. In this study we introduced a real-time polymerase chain reaction (PCR) essay, using SYBR Green I dye, to rapidly identify B. latifrons on an ABI PRISM 7700 sequence detection system. A latifron-specific PCR primer set was obtained based on mtDNA COI gene of B. latifrons . Nine Bactrocera fruit flies, B. latifrons , Bactrocera dorsalis , Bactrocera papayae , Bactrocera carambolae , Bactrocera philippinensis , Bactrocera occipitalis , Bactrocera correcta , Bactrocera cucurbitae and Bactrocera tau , were used to determine the specificity of primers lati1 and lati2. A series of genomic DNA dilutions of B. latifrons (0.01, 0.1, 1, 10, 20, 40 and 100 ng) were used to assess the sensitivity of the SYBR Green PCR. Template DNA concentration was one of the sources of variability in cycle threshold values (CT) and the optimum DNA concentration was between 1 and 20 ng. Genomic DNA isolated from larvae, pupae and adult specimens of B. latifrons were used to assess the specificity of the SYBR Green PCR. Melting curve analysis and agarose gel electrophoresis was employed to check the specificity of PCR products. Similar amplification plots were obtained using DNA from the three different stages of B. latifrons with primer set lati1/lati2. The melting temperature ( T _m) of PCR products was 77.5 ± 0.1°C, and the length of the amplified fragment 366 bp. Given the specificity and sensitivity of the assay, combined with high speed, low cost and the possibility of automating, SYBR Green PCR can be used as a rapid and specific technique for pest species identification in plant quarantine.     

Ecological significance of male attractant in the defence and mating strategies of the fruit fly, Bactrocera papayae

After being acclimatized to feeding on fruit flies, the Asian house gecko, Hemidactylus frenatus Duméril & Bibron (Gekkonidae:Squamata), consumed fewer methyl eugenol (ME) fed male fruit flies, Bactrocera papayae Drew & Hancock (Tephritidae: Diptera) than when offered ME-deprived males. After one-day exposure to only ME-fed males, the geckos avoided feeding on female flies when in the presence of ME-fed males. When mechanically disturbed, the ME-fed males spontaneously ejaculated a rectal secretion which contains phenyl propanoids that deter the predator.The ME-fed males also competed significantly better than normal (ME-deprived) males for virgin females. Male B. papayae converts ME to three other phenyl propanoids which act in concert as a sex pheromone to attract females during courtship and as an allomone to the gecko.     

Two new species and a new Australian record of Bactrocera Macquart (Diptera: Tephritidae: Dacinae) from northern Queensland, Torres Strait and Papua New Guinea

Abstract  Bactrocera (Bactrocera) speewahensis Fay & Hancock sp. n. is described from northern Queensland and B. (B.) torresiae Huxham & Hancock sp. n. is described from Boigu, Saibai and Dauan islands in Torres Strait and southern Papua New Guinea. Bactrocera (B.) nigrovittata Drew is newly recorded from Australia. All records are of male flies responding to chemical lures.     

Comparison of methyl eugenol metabolites, mitochondrial COI, and rDNA sequences of Bactrocera philippinensis (Diptera: Tephritidae) with those of three other major pest species within the dorsalis complex

Males of the oriental fruit fly, Bactrocera dorsalis (Hendel) and some of its sibling species have strong affinity for methyl eugenol (ME). Methyl eugenol ingested by male flies is biotransformed in the crop to two ME metabolites that eventually accumulate in the rectal gland, which is known to serve as a reservoir for B. dorsalis sex pheromones. When fed with ME, males of laboratory and wild B. philippinensis Drew and Hancock selectively accumulated two metabolites, 2-allyl-4,5-dimethoxyphenol and (E)-coniferyl alcohol, in the rectal gland, as was seen for B. dorsalis sensu stricto, B. invadens Drew, Tsuruta and White, and B. papayae Drew and Hancock. Phylogenetic analysis of COI and rDNA sequence data of these four taxa also revealed a close relationship among B. philippinensis, B. dorsalis s.s., B. invadens, and B. papayae (all four are members of the dorsalis species complex). This result corroborates pheromone analysis. The usefulness of pheromonal analysis as a chemotaxonomy tool to complement molecular and other analysis in differentiation of closely related sibling species within the Bactrocera dorsalis complex, for which use of morphological characters had been inadequate, is highlighted.     

桔小实蝇、木瓜实蝇和番石榴实蝇耐热性的热动力学分析

果实蝇属是重要的检疫性有害生物之一, 其中桔小实蝇Bactrocera dorsalis (Hendel)、 木瓜实蝇B. papayae Drew & Hancock和番石榴实蝇B. correcta (Bezzi)近几年在口岸检出率居高。本研究利用热水浸泡处理法比较这3种实蝇的卵、 1龄幼虫和3龄幼虫在44~47℃下的耐热性, 利用热动力学模型和活化能比较每种实蝇的耐热虫态以确定耐热种。结果表明： 在测定温度范围内, 3种实蝇卵、 1龄幼虫和3龄幼虫的死亡率都与处理时间呈正相关。热动力学系数取0.5时, 最适合热动力学模型预测3种实蝇在各实验温度LT_99.9968的致死时间。3种实蝇在44~47℃耐热性均为： 卵>1龄幼虫>3龄幼虫。番石榴实蝇在3种实蝇中最耐热。本研究结果为实蝇热处理指标的制定提供了实验依据。     

云南景洪果实蝇属三新种及一新纪录种记述(双翅目,实蝇科)(英文)

记述采自中国云南景洪的果实蝇属3新种和中国1新纪录种:滇黑寡鬃实蝇Bactrocera(Bactrocera)nigrifacia sp.nov.,哈迪氏果实蝇B.(B.)hardyisp.nov.,景洪果实蝇B.(B.)jinghongensis sp.nov.和黑颜面实蝇Bactrocera(Zeugodacus)vultus(Hardy,1973),3新种均符合DrewetHancock(1994)对桔小实蝇复合体的定义。模式标本保存于福建农林大学益虫研究所。滇黑寡鬃实蝇,新种Bactrocera(Bactrocera)nigrifacia sp.nov.(图1～4)新种与黑胫实蝇B.(Bactrocera)nigr of emoralis Whiteet Tsuruta相近,颜均为黑色,与后者的区别在于:2条平行的缝后侧黄色条终止于翅内鬃之前,第2腹背板橙棕色且前缘具黑色横带。正模♂,云南景洪,2010-09-02,Cue-lure引诱,张南南采。副模3♂♂,同正模。词源:新种种名根据颜面黑色而命名。哈迪氏果实蝇,新种Bactrocera(Bactrocera)hardyisp.nov.(图5～7)新种与印尼实蝇Bactrocera(Bactrocera)indonesiae Drewet Hancock相近,与后者的区别为:第3～5腹背板黑褐色,前足腿节黄褐色,且端部具黑褐色斑,中足和后足腿节为黄褐色,且端部具很短的黑褐色条纹。正模♂,云南景洪,2010-09-04,Cue-lure引诱,张南南采。副模3♂♂,同正模。词源:新种种名以双翅目分类专家DilbertE.Hardy的姓氏命名。景洪果实蝇,新种Bactrocera(Bactrocera)jinghongensis sp.nov.(图8～11)新种与基纳巴卢实蝇Bactrocera(Bactrocera)kinabalu Drewet Hancock相近,与后者的区别在于:缝后侧黄色条终止于翅内鬃之前,中足腿节端部1/2黑色。正模♂,云南景洪,2010-09-04,Cue-lure引诱,张南南采。副模4♂♂,同正模。词源:新种种名以模式产地命名。     

One and the same: integrative taxonomic evidence that Bactrocera invadens (Diptera: Tephritidae) is the same species as the Oriental fruit fly Bactrocera dorsalis

The invasive fruit fly Bactrocera invadens Drew, Tsuruta & White, and the Oriental fruit fly Bactrocera dorsalis (Hendel) are highly destructive horticultural pests of global significance. Bactrocera invadens originates from the Indian subcontinent and has recently invaded all of sub‐Saharan Africa, while B. dorsalis principally occurs from the Indian subcontinent towards southern China and South‐east Asia. High morphological and genetic similarity has cast doubt over whether B. invadens is a distinct species from B. dorsalis. Addressing this issue within an integrative taxonomic framework, we sampled from across the geographic distribution of both taxa and: (i) analysed morphological variation, including those characters considered diagnostic (scutum colour, length of aedeagus, width of postsutural lateral vittae, wing size, and wing shape); (ii) sequenced four loci (ITS1, ITS2, cox1 and nad4) for phylogenetic inference; and (iii) generated a cox1 haplotype network to examine population structure. Molecular analyses included the closely related species, Bactrocera kandiensis Drew & Hancock. Scutum colour varies from red‐brown to fully black for individuals from Africa and the Indian subcontinent. All individuals east of the Indian subcontinent are black except for a few red‐brown individuals from China. The postsutural lateral vittae width of B. invadens is narrower than B. dorsalis from eastern Asia, but the variation is clinal, with subcontinent B. dorsalis populations intermediate in size. Aedeagus length, wing shape and wing size cannot discriminate between the two taxa. Phylogenetic analyses failed to resolve B. invadens from B. dorsalis, but did resolve B. kandiensis. Bactrocera dorsalis and B. invadens shared cox1 haplotypes, yet the haplotype network pattern does not reflect current taxonomy or patterns in thoracic colour. Some individuals of B. dorsalis/B. invadens possessed haplotypes more closely related to B. kandiensis than to conspecifics, suggestive of mitochondrial introgression between these species. The combined evidence fails to support the delimitation of B. dorsalis and B. invadens as separate biological species. Consequently, existing biological data for B. dorsalis may be applied to the invasive population in Africa. Our recommendation, in line with other recent publications, is that B. invadens be synonymized with B. dorsalis.     

Sexual maturity and intraspecific mating success of two sibling species of the Bactrocera dorsalis complex

Bactrocera carambolae Drew & Hancock and B. papayae Drew & Hancock (Diptera: Tephritidae), two closely related sibling species in the B. dorsalis (Hendel) complex were shown to have different rates of sexual maturity. The response of B. carambolae males to methyl eugenol was observed to begin 10 days after emergence and increased with age. The attractancy peaked at 28 days after emergence and above. The males were also found to require a higher dosage of methyl eugenol (1 mg) for optimal response. These results are in contrast with those of B. papayae reported previously. The response to methyl eugenol was also observed to correspond with the age when first mating was performed.After eclosion, both sexes of B. carambolae mated approximately 2 weeks later and showed significantly lower matings/individual when compared with B. papayae in a 60-day observation period. Both sexes of B. papayae and males of B. carambolae were able to remate on the next day. However, B. carambolae females required a minimum refractory period of 8 days in order to be sexually receptive again.During the 60-day period, the mating propensity in B. papayae (100% success) was higher than B. carambolae (71–87% success) (P < 0.05). This observation was substantiated by a higher intraspecific mating success of B. papayae as compared to B. carambolae in a field cage (P < 0.001).     

Primers containing universal bases reduce multiple amoA gene specific DGGE band patterns when analysing the diversity of beta-ammonia oxidizers in the environment

The gene encoding the active site of the ammonia monooxygenase (amoA) has been exploited as molecular marker for studying ammonia-oxidizing bacteria (AOB) diversity in the environment. Primers amplifying functional genes are often degenerated and therefore produce multiple band patterns, when analysed with the Denaturing gradient gel electrophoresis (DGGE) approach. To improve the DGGE band patterns we have designed new primer sets which contain inosine residues and are specific for the amoA gene. Primers were evaluated analysing pure AOB cultures and two habitats (wastewater treatment plant, soda pools). We found that the application of inosine primers helped to reduce the apparent complexity of the DGGE band pattern. Comparison of sequences from environmental samples using either degenerated or inosine containing amoA primers retrieved both identical and additional sequences. Both primer sets seem to be limited in their ability to detect the presence of all AOB by DGGE analyses.     

Bridging the morphological and biological species concepts: studies on the Bactrocera dorsalis (Hendel) complex (Diptera: Tephritidae: Dacinae) in South-east Asia

Defining species accurately is a critical need in fundamental disciplines such as ecology and evolutionary biology and in applied arenas such as pest management. The validity of species designations depends on agreement of different methods of species diagnosis for unique biological species. The Bactrocera dorsalis complex of fruit flies provide an excellent opportunity for such a test of the congruence of different techniques (e.g. morphological, molecular, host-plant based, chemotaxonomy) used for species diagnosis. The complex contains a large number of closely-related species, is distributed over a wide geographical range in South-east Asia and considerable information has been compiled on some species. In the present study, the morphological and biological species boundaries were compared using new data from morphometric analyses of reproductive and body parts, together with a review of data on morphology, chemistry of male pheromones that are important in courtship and mating, molecular analyses, and endemic rainforest host plants. For the populations studied ( Bactrocera carambolae , Bactrocera dorsalis , Bactrocera occipitalis , Bactrocera papayae , Bactrocera philippinensis , Bactrocera kandiensis and Bactrocera invadens ) there appears to be significant congruence between the morphological and biological species boundaries.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 93 , 217–226.     

Population dynamics of three Bactrocera spp. fruit flies (Diptera: Tephritidae) and two introduced natural enemies,Fopius arisanus (Sonan) and Diachasmimorpha longicaudata (Ashmead) (Hymenoptera: Braconidae), after an invasion by Bactrocera dorsalis (Hendel) in Tahiti

Oriental fruit fly, Bactrocera dorsalis (Hendel), invaded French Polynesia in 1996. In 2002 a natural enemy, Fopius arisanus (Sonan), was released and established. By 2009 mean (±SD) F. arisanus parasitism for fruit flies infesting Psidium guajava (common guava), Inocarpus fagifer (Polynesian chestnut) and Terminalia catappa (tropical almond) fruits on Tahiti Island was 64.8 ± 2.0%. A second parasitoid, Diachasmimorpha longicaudata (Ashmead), was released and established in 2008. Although widespread, parasitism rates have not been higher than 10%. From 2003 (parasitoid establishment) to 2009 (present survey) numbers of B. dorsalis, Bactrocera tryoni (Froggatt), Queensland fruit fly, and Bactrocera kirki (Froggatt) emerging (per kg of fruit) declined. For example, for P. guajava there was a decline of 92.3%, 96.8%, and 99.6%, respectively. Analysis of co-infestation patterns (1998–2009) of B. dorsalis, B. tryoni, and B. kirki, suggest B. dorsalis is now the most abundant species in many common host fruits. Establishment of F. arisanus is the most successful example of classical biological control of fruit flies in the Pacific outside of Hawaii and can be introduced if B. dorsalis spreads to other French Polynesian islands, as was the recent case when B. dorsalis spread to the Marquesas Islands. These studies support F. arisanus as a prime biological control candidate for introduction into South America and Africa where Bactrocera carambolae Drew and Hancock and Bactrocera invadens Drew, Tsuruta, and White, respectively, have become established.     

Wing shape as a potential discriminator of morphologically similar pest taxa within the Bactrocera dorsalis species complex (Diptera: Tephritidae)

Four morphologically cryptic species of the Bactrocera dorsalis fruit fly complex (B. dorsalis s.s., B. papayae, B. carambolae and B. philippinensis) are serious agricultural pests. As they are difficult to diagnose using traditional taxonomic techniques, we examined the potential for geometric morphometric analysis of wing size and shape to discriminate between them. Fifteen wing landmarks generated size and shape data for 245 specimens for subsequent comparisons among three geographically distinct samples of each species. Intraspecific wing size was significantly different within samples of B. carambolae and B. dorsalis s.s. but not within samples of B. papayae or B. philippinensis. Although B. papayae had the smallest wings (average centroid size=6.002 mm±0.061 SE) and B. dorsalis s.s. the largest (6.349 mm±0.066 SE), interspecific wing size comparisons were generally non-informative and incapable of discriminating species. Contrary to the wing size data, canonical variate analysis based on wing shape data discriminated all species with a relatively high degree of accuracy; individuals were correctly reassigned to their respective species on average 93.27% of the time. A single sample group of B. carambolae from locality 'TN Malaysia' was the only sample to be considerably different from its conspecific groups with regards to both wing size and wing shape. This sample was subsequently deemed to have been originally misidentified and likely represents an undescribed species. We demonstrate that geometric morphometric techniques analysing wing shape represent a promising approach for discriminating between morphologically cryptic taxa of the B. dorsalis species complex.     

三种常见实蝇的形态学、生物学和生态学比较

对瓜实蝇Bactrocera (Tetradacus) minax (Enderlein)、桔小实蝇Bactrocera (Bactrocera) dorsalis (Hendel)和桔大实蝇Bactrocera (Zeugodacus) cucurbitae (Coquillett)的形态学、生物学、生态学等方面进行了比较和分析,包括三种实蝇在国内外的分布情况,对寄主选择的差异,各种虫态的形态特征,发育历期和生活史,并对它们的危害状况和防治方法分别作了介绍,可为3种实蝇的鉴定和防治提供参考.     

Molecular identification of Bactrocera passiflorae (Diptera: Tephritidae): Challenge and solution for DNA barcoding

Fruit flies cause significant damage to crop and fruit production worldwide. Therefore, it is essential to identify these organisms to species level; however, immature stages are often impossible to be identified morphologically; thus, the application of DNA barcoding has greatly assisted in species identification. Nuclear, mitochondrial pseudo-COI (NUMT) can be co-amplified with mitochondrial DNA when using generic primers and therefore impair the efficacy of DNA barcoding. This study detected two types of NUMTs from Bactrocera passiflorae, one of them is novel. Therefore, the new finding will assist future species identification by avoiding misidentification using ambiguous NUMT sequences. In addition, this study has developed primers to target the COI gene of B. passiflorae, not the NUMT copies. The newly designed primers have demonstrated its efficiency in amplifying the Mt-COI of B. passiflorae and can be used in routine diagnostics.     

番石榴果实蝇与桃实蝇的形态区别

介绍我国口岸截获频率较高的2种形态相似的实蝇---番石榴果实蝇Bactrocera (Bactrocera) correcta(Bezzi) 和桃实蝇Bactrocera zonata Saunders的鉴别特征。两者主要区别在于桃实蝇具1对卵圆形黑色颜斑,中胸背板红褐色。     

Piecing together an integrative taxonomic puzzle: microsatellite,wing shape and aedeagus length analyses of Bactrocera dorsalis s.l. (Diptera: Tephritidae) find no evidence of multiple lineages in a proposed contact zone along the Thai/Malay Peninsula

Bactrocera dorsalis (Hendel) and B. papayae Drew & Hancock represent a closely related sibling species pair for which the biological species limits are unclear; i.e. it is uncertain if they are truely two biological species, or one biological species which has been incorrectly split taxonomically. The geographical ranges of the two taxa are thought to abut or overlap on or around the Isthmus of Kra, a recognised biogeographic barrier located on the narrowest portion of the Thai Peninsula. We collected fresh material of B. dorsalis s.l. (i.e. B. dorsalis s.s.+ B. papayae) in a north–south transect down the Thai Peninsula, from areas regarded as being exclusively B. dorsalis s.s., across the Kra Isthmus, and into regions regarded as exclusively B. papayae. We carried out microsatellite analyses and took measurements of male genitalia and wing shape, both used previously to separate the taxa. No significant population structuring was found in the microsatellite analysis, consistent with one, predominantly panmictic population. Both morphological datasets showed consistent, clinal variation along the transect, without disjunction. No evidence supported historical vicariance driven by the Isthmus of Kra, and no dataset supported the current taxonomy of two species. Rather, within and across the area of range overlap or abutment between the two species, only continuous morphological and genetic variation was recorded. Recognition that morphological traits previously used to separate these taxa are continuous, and that there is no genetic evidence for population segregation in the region of suspected species overlap, is consistent with a growing body of literature that reports no evidence of biological differentiation between these taxa.     

Polymorphic microsatellite markers for population analysis of a tephritid pest species, Bactrocera tryoni

To obtain a set of microsatellite markers for the Queensland fruit fly Bactrocera tryoni , a genomic library was screened with a number of simple repeat oligonucleotide probes. Sequencing recovered 22 repeat loci. The microsatellite sequences were short, with repeat numbers ranging from five to 11. Of these, 16 polymerase chain reaction (PCR) primer sets yielded amplifiable products, which were tested on 53 flies from five widely separated sites. All loci showed polymorphism in the population sample, with the number of alleles ranging from two to 16. Several dinucleotide repeats showed alleles separated by single-base differences and multiple steps, suggesting a mutation process more complex than the stepwise mutation model.     

Tropical fruitflies (Diptera: Tephritidae) of the Krakatau Archipelago in 1990 and comments on faunistic changes since 1982

Three of the Krakatau islands, in Sunda Strait, some 40 km from both Java and Sumatra, are believed to have been totally devastated by the famous cataclysmic eruption of 1883. The fourth, Anak Krakatau, is an emergent volcanic island which rose from Krakatau's submerged 1883 caldera in 1930, suffered a self-devastating eruption in 1952/3 and several severely damaging ones since then. In 1990 the tephritid fauna was monitored on all the islands with Steiner traps. Nine species were found, five of which appeared to have been found by Yukawa in 1982 (Yukawa 1984) using similar methods and lures, when he noted a total of six species. As in 1982, the fauna in 1990 was characterized by the numerical dominance ofBactrocera albistrigata (de Meijere) although this appeared to be less marked than in 1982 andBactrocera papayae Drew & Hancock appeared to have increased in relative numbers. The distribution of three species over the islands of the archipelago had increased and four species not found in 1982 were present. Rakata, Krakatau's remnant and by far the largest and highest island, carried seven species (six in 1982), Sertung three (two in 1982), Panjang four (one in 1982) and Anak Krakatau four (one in 1982). Capture rates were lower than in 1982 but still higher than those obtained on the Javan mainland in 1982. There was a marked difference between the tephritid faunas of Anak Krakatau's two forelands, which were at different stages of biotic succession.     

应用种特异性PCR技术快速鉴定辣椒实蝇

【目的】辣椒实蝇 Bactrocera latifrons (Hendel)为我国重要的检疫性有害生物,其寄主范围广泛,危害严重。由于传统鉴定方法受到饲养周期、饲养条件、虫态等因素的限制,使得果蔬进出口贸易通关速度、疫情快速鉴定受到较大的影响,因此迫切需要开发关于实蝇的快速鉴定识别的技术。【方法】本研究基于mtDNA COI序列设计了一对能够准确鉴定辣椒实蝇的种特异性引物FL680和RL1057,选用辣椒实蝇作为阳性对照,选用番石榴实蝇B. correcta (Bezzi)、桔小实蝇 B. dorsalis (Hendel)和颜带实蝇 B. cilifer (Hendel)等20种实蝇作为阴性对照,进行PCR扩增并将PCR产物进行电泳检测。【结果】仅目标种辣椒实蝇能够扩增出清晰且单一的约378 bp的条带,其余实蝇种类均未出现条带。将本实验建立的种特异性PCR（SS-PCR）鉴定方法应用于实际检疫工作中并得到了验证,表明该方法具有强的种特异性。【结论】本文提出辣椒实蝇快速鉴定识别技术可应用于实蝇的疫情监测和口岸的检疫检测工作。