福州实蝇监测初报

自2005年5月29日至2006年5月28日在福州金山福建农林大学校园内进行了实蝇监测。采用甲基丁香酚(M e)、诱蝇酮(Cue)和地中海实蝇诱芯(T),诱集到橘小实蝇[Bactrocera(Bactrocera)dorsalis]、瓜实蝇[B.(Zeugodacus)cucurbi-tae]、南瓜实蝇[B.(Zeugodacus)tau]和具条实蝇(B.scutellata)4种实蝇,但未诱到地中海实蝇(Ceratitis capitata)。其中,橘小实蝇诱集量最大,8月中下旬达到高峰。本研究为掌握福州地区实蝇发生动态提供了基础资料。     

Comparative evaluation of spinosad and phloxine B as toxicants in protein baits for suppression of three fruit fly (Diptera: Tephritidae) species

Spinosad and phloxine B are two more environmentally friendly alternative toxicants to malathion for use in bait sprays for tephritid fruit fly suppression or eradication programs. Laboratory tests were conducted to assess the relative toxicity of these two toxicants for melon fly, Bactrocera cucurbitae Coquillett; oriental fruit fly, Bactrocera dorsalis Hendel; and Mediterranean fruit fly, Ceratitis capitata (Wiedemann) females. Field tests also were conducted with all three species to compare these toxicants outdoors under higher light and temperature conditions. In laboratory tests, spinosad was effective at much lower concentrations with LC50 values at 5 h of 9.16, 9.03, and 4.30 compared with 250.0, 562.1, and 658.9 for phloxine B (27, 62, and 153 times higher) for these three species, respectively. At 16 ppm spinosad, LT50 values were lower for all three species (significantly lower for C. capitata and B. dorsalis) than 630 ppm phloxine B LT50 values. At 6.3 ppm spinosad, the LT50 value for C. capitata (3.94) was still significantly less than the 630 ppm phloxine B LT50 value (6.33). For all species, the 100 ppm spinosad concentrations gave LT50 values of < 2 h. In comparison among species, C. capitata was significantly more sensitive to spinosad than were B. cucurbitae or B. dorsalis, whereas B. cucurbitae was significantly more sensitive to phloxine B than were C. capitata or B. dorsalis. LC50 values were reduced for both toxicants in outdoor tests, with greater reductions for phloxine B than for spinosad for B. dorsalis and B. cucurbitae. Fly behavior, though, is likely to keep flies from being exposed to maximum possible outdoor light intensities. Comparable levels of population suppression for any of the three species tested here will require a much higher concentration of phloxine B than spinosad in the bait.     

The Hobo Transposable Element Excises and Has Related Elements in Tephritid Species

Function of the Drosophila melanogaster hobo transposon in tephritid species was tested in transient embryonic excision assays. Wild-type and mutant strains of Anastrepha suspensa, Bactrocera dorsalis, B. cucurbitae, Ceratitis capitata, and Toxotrypana curvicauda all supported hobo excision or deletion both in the presence and absence of co-injected hobo transposase, indicating a permissive state for hobo mobility and the existence of endogenous systems capable of mobilizing hobo. In several strains hobo helper reduced excision. Excision depended on hobo sequences in the indicator plasmid, though almost all excisions were imprecise and the mobilizing systems appear mechanistically different from hobo. hobo-related sequences were identified in all species except T. curvicauda. Parsimony analysis yielded a subgroup including the B. cucurbitae and C. capitata sequences along with hobo and Hermes, and a separate, more divergent subgroup including the A. suspensa and B. dorsalis sequences. All of the sequences exist as multiple genomic elements, and a deleted form of the B. cucurbitae element exists in B. dorsalis. The hobo-related sequences are probably members of the hAT transposon family with some evolving from distant ancestor elements, while others may have originated from more recent horizontal transfers.     

The complete nucleotide sequence of the mitochondrial genome of the oriental fruit fly, Bactrocera dorsalis (Diptera: Tephritidae)

The complete mitochondrial genome of the oriental fruit fly Bactrocera dorsalis s.s. has been sequenced, and is here described and compared with the homologous sequences of Bactrocera oleae and Ceratitis capitata. The genome is a circular molecule of 15,915 bp, and encodes the set of 37 genes generally found in animal mitochondrial genomes. The structure and organization of the molecule is typical and similar to the two closely related species B. oleae and C. capitata, although it presents an interesting case of putative intra-molecular recombination. The relevance of the growing comparative dataset of tephritid complete mitochondrial genomes is discussed in relation to the possibility to develop robust assays for species discrimination in quarantine and agricultural monitoring practices, as well as basic phylogeography/population genetic studies.     

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。     

Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae) revealed by COI and 16S DNA sequences

The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies.     

Microsatellite markers for the pest fruit fly,Bactrocera papayae (Diptera: Tephritidae) and other Bactrocera species

The dorsalis complex contains some of the most economically important fruit fly pests of the Asia–Pacific regions, including Bactrocera dorsalis, Bactrocera papayae and Bactrocera carambolae. These species are morphologically indistinct and genetically very similar. We describe the development of 12 microsatellite markers isolated from a representative of the dorsalis complex, B. papayae. We show the potential utility of the B. papayae microsatellites and a set of microsatellites isolated from Bactrocera tryoni as population and species markers for the dorsalis complex.     

A comparative assessment of the response of three fruit fly species (Diptera: Tephritidae) to a spinosad-based bait: effect of ammonium acetate, female age, and protein hunger

Ammonia-releasing substances are known to play an important role in fruit fly (Diptera: Tephritidae) attraction to food sources, and this information has been exploited for the development of effective synthetic food-based lures and insecticidal baits. In field studies conducted in Hawaii, we examined the behavioural response of wild female oriental fruit fly (Bactrocera dorsalis (Hendel)), melon fly (B. cucurbitae (Coquillett)), and Mediterranean fruit fly (Ceratitis capitata (Wiedemann)) to spinosad-based GF-120 NF Naturalyte Fruit Fly Bait(?) formulated to contain either 0, 1 or 2% ammonium acetate. Use of visually-attractive yellow bait stations for bait application in the field allowed for proper comparisons among bait formulations. Field cage tests were also conducted to investigate, using a comparative behavioural approach, the effects of female age and protein starvation on the subsequent response of F1 generation B. cucurbitae and B. dorsalis to the same three bait formulations that were evaluated in the field. Our field results indicate a significant positive effect of the presence, regardless of amount, of AA in GF-120 for B. dorsalis and B. cucurbitae. For C. capitata, there was a significant positive linear relationship between the relative amounts of AA in bait and female response. GF-120 with no AA was significantly more attractive to female C. capitata, but not to female B. dorsalis or B. cucurbitae, than the control treatment. Our field cage results indicate that the effects of varying amounts of AA present in GF-120 can be modulated by the physiological stage of the female flies and that the response of female B. cucurbitae to GF-120 was consistently greater than that of B. dorsalis over the various ages and levels of protein starvation regimes evaluated. Results are discussed in light of their applications for effective fruit fly suppression.     

Host status of Vaccinium reticulatum (Ericaceae) to invasive tephritid fruit flies in Hawaii

Ohelo (Vaccicinium reticulatum Small) (Ericaceae) is a native Hawaiian plant that has commercial potential in Hawaii as a nursery crop to be transplanted for berry production or for sale as a potted ornamental. No-choice infestation studies were conducted to determine whether ohelo fruit are hosts for four invasive tephritid fruit fly species. Ohelo berries were exposed to gravid female flies ofBactrocera dorsalis Hendel (oriental fruit fly), Ceratitis capitata (Wiedemann) (Mediterranean fruit fly), Bactrocera cucurbitae Coquillet (melon fly),or Bactrocera latifrons (Hendel) in screen cages outdoors for 24 h and then held on sand in the laboratory for 2 wk for pupal development and adult emergence. Only B. dorsalis successfully attacked and developed in ohelo berries. In total, 1570 berries produced 10 puparia, all of which emerged as adults, for a fruit infestation rate of 0.0064% and an average of 0.0053 puparia per gram of fruit. By comparison, papaya fruit used as controls produced an average of 1.44 B. dorsalis puparia per g of fruit. Ohelo berry is a marginal host for B. dorsalis and apparently a nonhost for C. capitata, B. cucurbitae, and B. latifrons. Commercial plantings of ohelo will rarely be attacked by fruit flies in Hawaii.     

地中海实蝇幼虫分子鉴定

通过对从秘鲁进口的葡萄中截获实蝇类幼虫进行ITS区和线粒体COⅠ,COⅡ,COⅢ、ND5基因序列的扩增和测序,并与GenBank中对应的序列进行比对,结果表明,截获样品ITS区序列和地中海实蝇Ceratitis capitata(Wiedemann)同源性为95·16%(其中ITS1为99·52%,ITS2为86·2%),线粒体COⅠ,COⅡ,COⅢ,ND5基因序列和地中海实蝇C.capitata同源性为100%,99·9%,99·5%,99·8%;基于COⅠ序列构建的系统发育树中,幼虫样品和地中海实蝇最为接近。根据序列分析和系统发育关系分析的结果,将截获的实蝇类幼虫鉴定为地中海实蝇C.capitata。     

Toxicity of spinosad in protein bait to three economically important tephritid fruit fly species (Diptera: Tephritidae) and their parasitoids (Hymenoptera: Braconidae)

The feeding toxicity of the natural insecticide spinosad in Provesta protein bait was evaluated for three economically important fruit fly species, the Mediterranean fruit fly, Ceratitis capitata (Wiedemann); the melon fly, Bactrocera cucurbitae Coquillett; and the oriental fruit fly, Bactrocera dorsalis Hendel. Both females and males were evaluated. Spinosad was remarkably similar in toxicity to all three fruit fly species. Male C. capitata (24 h LC50 values and 95% fiducial limits = 2.8 [2.60-3.0] mg/liter spinosad) were significantly, although only slightly more susceptible to spinosadthan females (4.2 [3.8-4.6] mg/liter). Male (5.5 [4.7-6.6] mg/liter) andfemale (4.3 [3.7-4.9] mg/liter) B. cucurbitae were equally susceptible to spinosad. Female (3.3 [3.1-3.6] mg/liter) and male (3.1 [2.9-3.3] mg/liter) B. dorsalis also were equally susceptible to spinosad. Provesta bait containing spinosad also was evaluated against two parasitoids of tephritid fruit flies, Fopius arisanus (Sonan) and Pysttalia fletcheri (Silvestri). These parasitoids did not feed on the bait, so a contact toxicity test was conducted. Significant amounts of mortality were found only after exposure of parasitoids to spinosad-coated glass vials with concentrations > or =500 mg/liter spinosad. Parasitoids were less susceptible than fruit flies to such a degree that use of spinosad in bait spray should be compatible with these parasitoid species. Because the fruit flies tested in this study were so susceptible to spinosad, this product seems to be promising as a bait spray additive and a replacement for malathion for control of these species.     

Comparison of in vitro heat and cold tolerances of the new invasive species Bactrocera invadens (Diptera: Tephritidae) with three known tephritids

Bactrocera invadens Drew, Tsuruta & White (Diptera: Tephritidae) is spreading throughout central Africa attacking a variety of fruit; quarantines are placed on fruit from this region that are considered hosts. The only phytosanitary treatment that is commercially available is an ionizing irradiation treatment for all Tephritidae at 150 Gy. The development of other treatments, such as heat, cold, or fumigation, usually requires testing tens of thousands of insects at a dose that provides efficacy and may take several years. It may be possible to shorten the time required to develop treatments by comparing tolerance of a new quarantine pest to tolerances of pests with similar behaviors and modes of infestation for which treatment schedules are available. Cold and heat tolerance ofB. invadens was compared with tolerance of Anastrepha ludens (Loew), Bactrocera dorsalis (Hendel), and Ceratitis capitata (Wiedemann) in vitro. Third-instar B. invadens was no more cold tolerant than the other species when treated in diet at 0.94 +/- 0.65 degrees C and no more heat tolerant than C. capitata when immersed in vials in water at 44.7 +/- 0.1 degrees C. The data at 0.94 +/- 0.65 degrees C was used to include B. invadens in a USDA cold treatment schedule for citrus fruit from Africa so that trade would not be interrupted while protecting U.S. agriculture from this invasive pest.     

Trap capture of three economically important fruit fly species (Diptera: Tephritidae): evaluation of a solid formulation containing multiple male lures in a Hawaiian coffee field

Invasive fruit flies (Diptera: Tephritidae) pose a global threat to agriculture through direct damage to food crops and the accompanying trade restrictions that often result. Early detection is vital to controlling fruit flies, because it increases the probability of limiting the growth and spread of the invasive population and thus may greatly reduce the monetary costs required for eradication or suppression. Male-specific lures are an important component of fruit fly detection, and three such lures are used widely: trimedlure (TML), cue lure (CL), and methyl eugenol (ME), attractive to Mediterranean fruit fly, Ceratitis capitata (Wiedemann); melon fly, Bactrocera cucurbitae (Coquillett); and oriental fruit fly, Bactrocera dorsalis (Hendel), respectively. In California, Florida, and Texas, the two Bactrocera lures are applied to separate species-specific traps as liquids (with a small amount of the insecticide naled added), whereas TML is delivered as a solid plug in another set of traps. Thus, the detection protocol involves considerable handling time as well as potential contact with a pesticide. The purpose of this study was to compare trap capture between liquid male lures and "trilure" wafers that contain TML, ME, raspberry ketone (RK, the hydroxy equivalent of CL), and the toxicant DDVP embedded within a solid matrix. Field studies were conducted in a Hawaiian coffee (Coffea arabica L.) field where the three aforementioned species co-occur, showed that the wafer captured at least as many flies as the liquid baits for all three species. This same result was obtained in comparisons using both fresh and aged (6-wk) baits. Moreover, the wafers performed as well as the single-lure traps in an ancillary experiment in which TML plugs were substituted for liquid TML. Additional experiments demonstrated explicitly that the presence of ME and RK had no effect on captures of C. capitata males and similarly that the presence of TML had no effect on the capture of B. cucurbitae or B. dorsalis males.     

Novel bait stations for attract-and-kill of pestiferous fruit flies

A novel, visually-attractive bait station was developed in Hawaii for application of insecticidal baits against oriental fruit fly, Bactrocera dorsalis (Hendel), melon fly, Bactrocera cucurbitae (Coquillett), and Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (all Diptera: Tephritidae). The bait station developed represents a supernormal visual stimulus of papaya foliage and takes advantage of the flies' strong response to the high light-reflecting properties of yellow color and of their need for shelter, while fully protecting the bait against rainfall. Field studies revealed that the behavioral response of female fruit flies, in particular C. capitata and B. cucurbitae , to yellow-painted bait stations sprayed with GF-120 NF Naturalyte Fruit Fly Bait was significantly enhanced compared with similarly sprayed bait stations that mimicked the green color of fully grown papaya leaves. Field studies conducted with B. cucurbitae indicated that the period of bait attractiveness can be extended for at least 1 week after bait application due to the rain-fastness properties of the bait stations and the use of a visually-attractive color. Our studies provide the behavioral basis for the development of improved attract-and-kill bait stations for fruit flies in Hawaii. These devices also provide a standardized way of evaluating bait spray formulations, thus allowing for proper comparisons over time, across species, and among geographical areas.     

基于mtDNA Cytb 的六种果实蝇的分子鉴定

本研究首次对果实蝇属的桔小实蝇Bactrocera dorsalis、瓜实蝇B. cucurbitae、南瓜实蝇B. tau、番石榴实蝇B. correcta、具条实蝇B. scutellata、黑漆实蝇B. scutellaris等6种实蝇mtDNA Cytb基因进行了测序。对这6种实蝇72个个体mtDNA Cytb基因中段420 bp的碱基序列进行分析,得到38种单倍型,发现了116个变异位点,其中30个位点较为稳定。对这6种实蝇与其各自鉴别位点的对应关系研究表明,mtDNA Cytb基因可以作为这6种实蝇种类鉴别的分子标记。     

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.     

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。     

Autophagy is Required for the Degeneration of the Ovarian Follicular Epithelium in Higher Diptera

Autophagy is a major pathway for the degradation of long-lived proteins and cytoplasmic organelles and an essential part of programmed cell death, as well. Our findings indicate that programmed cell death of the ovarian follicle cells in the higher Diptera species Bactrocera oleae and Ceratitis capitata manifests features of autophagic cell death. The follicle cells during the developmental stage 14 contain autophagic vacuoles and they do not exhibit caspase activity in any area of the egg chamber. Their nuclei are characterized by condensed chromatin, accompanied with high—but not low—molecular weight DNA fragmentation events exclusively detected in distinct cells of the anterior pole. The above results are likely associated with the abundant phagocytosis observed at the entry of the lateral oviducts, where numerous cell bodies are massively engulfed by epithelial cells. The similarity of the cell death process among B. oleae, C. capitata and Drosophila melanogaster species strongly suggests that autophagy-mediated cell death is conserved in higher Diptera species.

Addendum to:

Programmed Cell Death of Follicular Epithelium during the Late Developmental Stages of Oogenesis in the Fruit Flies Bactrocera oleae and Ceratitis capitata (Diptera, Tephritidae) is Mediated by Autophagy

I.P. Nezis, D.J. Stravopodis, L.H. Margaritis and I.S. Papassideri

Dev Growth Differ 2006; 48:189-98     

MONOPHYLY OF THE GENUS CLOSTERIUM AND THE ORDER DESMIDIALES (CHAROPHYCEAE, CHLOROPHYTA) INFERRED FROM NUCLEAR SMALL SUBUNIT rDNA DATA

We newly sequenced the nuclear-encoded small subunit (SSU) rDNA coding region for 21 taxa of the genus Closterium. The new sequences were integrated into an alignment with 13 known sequences of conjugating green algae representing six traditional families (i.e. Zygnemataceae, Mesotaeniaceae, Gonatozygaceae, Peniaceae, Closteriaceae, and Desmidiaceae) and five known charophycean sequences as outgroups. Both maximum likelihood and maximum parsimony analyses supported with high bootstrap values one large clade containing all placoderm desmids (Desmidiales). All the Closterium taxa formed one clade with 100% bootstrap support, indicating their monophyly, but not paraphyly, as suggested earlier. As to the taxa within the genus Closterium , we found two clades of morphologically closely related taxa in both maximum likelihood and maximum parsimony trees. They corresponded to the C. calosporum species complex and the C. moniliferum-ehrenbergii species complex. It is of particular interest that the homothallic entity of C. moniliferum v. moniliferum was distinguished from and ancestral to all other entities of the C. moniliferum-ehrenbergii species complex. Superimposing all 50 charophycean sequences on the higher order SSU rRNA structure model of Closterium , we investigated degrees of nucleotide conservation at a given position in the nucleotide sequence. A characteristic "signature" structure to the genus Closterium was found as an additional helix at the tip of V1 region. In addition, eight base deletions at the tip of helix 10 were found to be characteristic of the C. calosporum species complex, C. gracile , C. incurvum , C. pleurodermatum , and C. pusillum v. maius. These taxa formed one clade with an 82% bootstrap value in maximum parsimony analysis.