XYLAN & ARABINOXYLAN (100 Assays per Kit) or (1000 Microplate Assays per Kit) or (1300 Auto-Analyser Assays per Kit) © Megazyme International Ireland 2012 INTRODUCTION:In nature, D-xylose occurs mainly in the polysaccharide form as xylan, arabinoxylan, glucuronoarabinoxylan, xyloglucan and xylogalacturonan. Mixed linkage D-xylans are also found in certain seaweed species and a similar polysaccharide is thought to make up the backbone of psyllium gum. Free D-xylose is found in guava, pears, blackberries, loganberries, raspberries, aloe vera gel, kelp, echinacea, boswellia, broccoli, spinach, eggplant, peas, green beans, okra, cabbage and corn. In humans, D-xylose is used in an absorption test to help diagnose problems that prevent the small intestine from absorbing nutrients, vitamins and minerals in food. D-Xylose is normally easily absorbed by the intestine. When problems with absorption occur, D-xylose is not absorbed and blood and urine levels are low. A D-xylose test can help to determine the cause of a child's failure to gain weight, especially when the child seems to be eating enough food. If in a polysaccharide, the ratio of D-xylose to other sugars etc. is known, then the amount of the polysaccharide can be quantitated from this knowledge plus the determined concentration of D-xylose in an acid hydrolysate. Xylans are a major portion of the polysaccharides that could potentially be hydrolysed to fermentable sugar for biofuel production.
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Microsoft word - 9 galley.docJ. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 Testing an attracticide hollow fibre formulation for control
of Codling Moth, Cydia pomonella
ALAN L. KNIGHT
YAKIMA AGRICULTURAL RESEARCH LABORATORY, AGRICULTURAL RESEARCH
SERVICE, USDA. 5230 KONNOWAC PASS RD., WAPATO, WA 98951
Laboratory and field tests were conducted to evaluate the use of an experimentalsprayable formulation of chopped hollow fibres loaded with codlemone and mixed with1.0% esfenvalerate and an adhesive to control codling moth, Cydia pomonella (L.)(Lepidoptera: Tortricidae). Moths were not repelled by the addition of the insecticide tothe adhesive and were rapidly killed following brief contact. A significantly greaterproportion of male moths flew upwind and contacted individual fibres for a longerperiod of time when fibres had been aged > 7 d versus fibres 0 – 7 days-old in flighttunnel tests. Field tests using sentinel fibres placed in 10.0 mg drops of adhesive onplastic disks stapled to the tree found that fibres were not touched until they had aged >8 d. Conversely, moth mortality following a 3-s exposure to field-collected fibresdeposited on the top of leaves was low in bioassays with fibres aged > 8 d. Thedeposition and adhesion of fibres within the apple canopy appear to be two majorfactors influencing the success of this approach. Fibres were found adhering to foliage,fruit, and bark within the orchard; however, visual recovery of fibres following each ofthe three applications was < 5.0%. Both the substrate and the positioning of the fibre onthe substrate influenced fibre retention. The highest proportion of fibres was foundinitially on the upper surface of leaves and this position also had the highest level offibre retention. Fibres on the underside of leaves or partially hanging off of a substratewere dislodged within two weeks.
Key words: sex pheromone, codling moth, attracticide, apple
A variety of approaches have been developed that utilize the sex pheromone of codling moth, Cydia pomonella L., for its effective management in deciduous tree fruit and nutcrops, including the application of hand applied dispensers (Charmillot and Pasquier1992), sprayable microencapsulated materials (Charmillot and Pasquier 2001), widely-spaced aerosol emitters (Shorey and Gebers 1996), and paste droplets formulated withinsecticides (Charmillot et al. 2000). Chopped hollow fibres loaded with codlemone havebeen used for codling moth both in hand-applied formulations (Cardé et al. 1977) and in asprayable formulation in which the fibres were mixed with an adhesive (Moffitt andWestigard 1984). Fibres provided high levels of disruption throughout the season in thesestudies.
Hollow fibres have been widely used in aerial applications in cotton for management of pink bollworm, Pectinophora gossyptella (Saunders) (Baker et al. 1990). Cotton growerscombined the use of the hollow fibres and synthetic pyrethroids to develop an attracticideformulation (Beasley and Henneberry 1984). Studies with pink bollworm showed that thisattracticide approach had minimal effect on natural enemies (Butler and Las 1983) and hadsignificant lethal and sublethal effects, which reduced the pest population (Floyd and J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 Crowder 1981). This attracticide approach has not been tested with codling moth. Hereinare presented preliminary studies that evaluated the potential of this approach and the useof chopped fibres for communication disruption of codling moth.
MATERIALS AND METHODS
Laboratory test protocol. The response of male codling moths to an experimental
formulation of black hollow Celcon fibres (200 µm i.d. x 15 mm) loaded with 15%codlemone diluted in hexane (Scentry Inc., Buckeye, AZ) was observed in a flight tunnel.
Fibres were formulated to release 0.1 µg/h codlemone at 20 oC (Weatherston et al. 1985).
The tunnel was 1.65 m long, 0.56 m wide and 0.56 m high and constructed from 6 mmthick acrylic sheeting. A blower was used to push air within the room (maintained at 22 –24 oC and 50 – 60% RH) into a plenum, through a charcoal filter, and through a series ofscreens before passing into the working section of the tunnel. An identical blower wasused on the opposite end to pull air through the tunnel. Power to the blowers was providedby two 12-volt battery chargers attached to 115-volt AC variable resistors. By carefullyadjusting the speed of each blower, laminar airflow was created which passed through thetunnel at the rate of 0.13 m/sec (measured by movement of smoke). Exhaust was expelledto the outside of the building. Red lights installed above the working section of the tunnelprovided enough light (4.3 lux) to make observations.
Insects were obtained as mature larvae inside corrugated cardboard strips from a laboratory colony reared on a soybean diet at the Yakima USDA Insectary (Toba andHowell 1991). Virgin male moths were collected daily and conditioned in constant lightfor 24 – 48 h at 21 oC and 60% RH. Prior to testing, moths were placed in completedarkness for 30 min.
Technical esfenvalerate (Dupont Agricultural Products, Wilmington, DE) was diluted in acetone and mixed with adhesive at a 1.0% wt/wt concentration. A single hollow fibrewas placed on a 10.0 mg droplet of a polybutene adhesive (Biotac 100, ScentryBiologicals, Billings, MT) in the center of an 18.0 mm diameter plastic disk. The plasticdisk with the fibre was placed on a small metal platform suspended 30 cm from the top ofthe flight tunnel at the air inlet end. Moths were released from a 30 cm high platformplaced near the air outlet end of the tunnel. The number and duration of individual visitsto the fibre were recorded for 7 min with an infrared motion detector coupled to acomputer.
Attractiveness and toxicity of laboratory-aged fibers. Two types of tests were
conducted in the flight tunnel to assess the attractiveness of individual fibres for malecodling moths and the toxicity and possible repellency of adding an insecticide to theadhesive. In the first test, hollow fibres were aged for 7 d at 24 oC prior to testing. Tenreplicates of 10 moths were flown to a fibre placed either in adhesive or in adhesive withinsecticide. Treatments were run alternately for each replicate, n = 10. The attractivenessand toxicity of fibres placed on adhesive treated with 1.0% esfenvalerate and aged in agreenhouse were evaluated in the second test. Disks were collected after 0, 1, 4, 7, 14, 21,and 28 d and kept frozen at –10 oC. Five fibres from each age class were tested in the flighttunnel in a random order and each fibre was tested twice, n = 10 replicates. Six maleswere released simultaneously for each fibre and allowed to fly for 7 min. Moths werecollected individually in vials at the end of each flight test and mortality was scored after24 h at 24 oC.
Field test protocol. Field studies of fibres were conducted in 1992 and 2003. Three
applications of fibres were made to a 0.3 ha (214 trees) 5-year-old ‘Golden Delicious'block trained on a M-16 rootstock with central leader architecture on 5 May, 1 June, and28 July 1992. The mean (SE) height of trees was 2.1 (0.1) m. A standard spinning cone J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 applicator used for ground application of the fibre in field crops (Moffitt and Short 1982)was supplied by Scentry personnel and attached to a tractor. The tractor and sprayer werecalibrated to deliver 100.0 g of fibres (15.0 g a.i.) in 6.0 L adhesive per hectare. Thedeposition and retention of fibres were evaluated following an application on 9 July 2003in a 4.0-ha orchard of mixed apple cultivars. The orchard was treated with 250.0 g of a10.0% a.i. fibre mixed with 4.7 L adhesive per hectare using a specialized tractor-pulledoverhead applicator (Blue Line Manufacturing, Wenatchee, WA). The same adhesive andfibre were used in both years.
Attractiveness and toxicity of field-aged fibres. The attractiveness and toxicity of
field-aged fibres were evaluated throughout the 1992 season. On each of the three spraydates one fibre was placed on a 10.0 mg adhesive drop in the center of each of 120 plasticdisks that were stapled to the wooden posts of a wire deer fence situated > 50 m from theapple orchard. On each subsequent sampling date 10 disks without any moth scales wereplaced in the upper third of the apple orchard's canopy in a horizontal position. Thesesentinel fibres were left in the orchard for 5 – 7 d and then reexamined with a microscopefor the presence of moth scales. In addition, on each sampling date 10 fibres deposited bythe spray application on the upper surface of leaves were collected from the orchard andreturned to the laboratory. Five 1 – 2 d-old chilled laboratory-reared moths were touchedto each fibre using a suction hose for 3 s. Moth mortality was scored after 24 h at 24 oC.
Deposition and retention of fibres. Several studies were conducted during the season
to assess the deposition and retention of fibres in the apple orchard. A trial was conductedon 12 September to estimate the number of fibres applied per hectare. Blank white celconfibres (200 µm i.d. x 15 mm) were mixed with the adhesive and applied at the standard rate(100.0 g in 6.0 liters adhesive). Five dark blue tarps (2.92 m x 2.92 m) were placed in arow on a grassy strip. The sprayer was started 50 m away from the first tarp and oncefibres had begun to be released from the spinning cone the tractor was driven forward at aspeed of 4.0 km per h. The number of fibres deposited on each sheet was counted and usedto estimate the number of fibres applied to the entire orchard (area equivalent to 426 tarps).
Deposition of fibres within the canopy of the orchard was estimated following each sprayapplication by visually examining 60 trees for fibres. Individual trees were inspected for 3to 5 minutes from the ground. The retention of marked fibres within the canopy of theapple orchard was evaluated following the June application in 1992. Fifty-seven fibreswere located on leaves and their location was marked with flagging. The retention of thesefibres was checked after 2 and 7 wk.
One hundred and twenty-two fibres were located and marked with flagging one day after the application in 2003. The position of each fibre was recorded with respect tosubstrate and whether the fibre was in full contact with the substrate or if a portion of thefibre was detached from the substrate (overhanging). Their retention in the canopy wassubsequently evaluated on 14 and 21 July and 22 August.
Statistical analyses. An unpaired t-test and analysis of variance on transformed data
(square root [x+0.01]) were used to compare the attractiveness and toxicity of fibres placedin adhesive either with or without insecticide and to fibres aged from 0 – 28 d to cohorts ofmoths, respectively (Analytical Software 2000). Means in significant ANOVA's wereseparated with Fisher's LSD test, P < 0.05.
Attractiveness and toxicity of laboratory-aged fibers. No difference was found in
the number of moth visits to fibres placed in either clean (mean ± SE = 13.4 ± 1.7) orinsecticide-impregnated (17.4 ± 2.3) adhesive during the 7-minute bioassay in the flighttunnel (t = 1.40, df = 18, P = 0.18). Similarly, no difference was found in the duration of amoth visit between fibres placed in clean (1.9 ± 0.5) or insecticide-impregnated adhesive J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 (1.8 ± 0.4) (t = -0.09, df = 18, P = 0.93). Subsequent tests showed that the age of the fibrewas a significant factor affecting moth contact and moth mortality (Table 1). Fibres placedin insecticide-impregnated adhesive and aged for < 7 d had significantly fewer mothcontacts and reduced visitation time. No difference in either factor was found for fibresaged 14 – 28 d. A significantly greater proportion of moths per cohort were killed whenflown to fibres aged 14 - 28 d versus < 1 d-old fibres. The highest moth mortality occurredwith fibres aged 14 d (Table 1). The lack of a significant difference in moth mortalityfollowing exposure to fibres aged 4 – 7 d versus 21 – 28 d may have been due to a declinein the toxicity of the insecticide in the older drops.
Influence of age on the attractiveness and toxicity of an individual hollow fibre loadedwith 15% codlemone and placed on a 10.0 mg drop of adhesive treated with 1.0%esfenvalerate for male codling moths flown in a flight tunnel.
Mean (SE) time (s) proportion of dead Age of fibre (d)a per source contact b Statistical analysis F = 6.82; df = 6, 63; F = 3.93; df = 6, 46; F = 9.82; df = 6, 46; P < 0.0001 P < 0.01 P < 0.001 a Fibres were aged in a greenhouse maintained between 20 – 24 oC for up to 28 days.
b Ten cohorts of six moths were flown in the flight tunnel for 7 minutes for each fibre ageclass. The mean number of moth contacts and time per source visit per cohort weremeasured with an infrared motion detector hooked to a computer.
c Following each tests moths which contacted adhesive were collected and placedindividually in vials. Mortality was scored after 24 h at 24 oC.
Attractiveness and toxicity of field-aged fibres. No moth scales were found on the
sentinel fibres placed in the apple orchard during the first eight days after any of the threeapplications (Table 2). The proportion of fibres aged from 8 – 51 d visited by mothsranged from 0.43 – 0.85 during the season. Mortality of moths in the 3-s touch bioassaywas > 85% for fibres collected on the day of the spray application. In general, fibres wereinitially sticky and associated with several milligrams of adhesive. Moth mortalitydropped sharply with field aging of the fibres, however, 65 – 80% mortality occurred with8 d and 5 d old fibres after the second and first applications, respectively. Moth mortalitywas much lower with 7 d-old fibres following the third application (Table 2). Mothmortality with field-collected fibres collected 2 – 7 wk after the application ranged from0.0 – 30.0%.
Deposition and retention of fibres. The mean (SE) number of fibres counted per tarp
was 32.9 (15.3). Extrapolating the deposition of fibres on the tarps to the area of the entireorchard (equivalent to 426 tarps) estimated 14,015 fibres were applied. Following the 5May spray application a mean (SE) of 0.9 (0.3) fibres were sampled per tree. This firstapplication was made a few days past full bloom and the growth of green foliage waslimited. The mean density of fibres following the 1 June application when trees hadabundant foliage increased to 2.5 (0.4) fibres per tree. However, fibre density following the J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 third application on 28 July was somewhat lower, 1.5 (0.4) fibres per tree. The highestdensity of fibres found on a single tree during the season was 17. Extrapolating the meandensity of fibres sampled per tree (1.5 – 2.5) multiplied by the number of trees in the block(214) suggests that only 2.3 – 3.8% of the estimated number of fibres sprayed in theorchard (14,015) were deposited on the trees.
Proportion of sentinel hollow fibres placed in 10 mg adhesive on a plastic disk at varioustimes following a spray application that contained moth scales and moth mortalityfollowing a 3-s touch to field-aged fibres deposited on the upper surface of apple leaves.
Post-spray interval Proportion of fibres % moth mortality in (d) fibre was in field a Positive visitation of codling moth to sentinel fibres was based on the microscopicdetection of moth scales in the adhesive surrounding each sentinel fibre. Fibres andadhesive were placed in the center of plastic disks that were stapled horizontally in theupper third of the tree canopy and left in the field for 5 – 7 d.
b Moth mortality was assessed 24 h following a 3 s touch exposure to a field-collected fibreon the upper surface of a leaf. Five moths were tested per fibre and ten fibres werecollected on each date.
Retention of hollow fibres loaded with codlemone and mixed with an adhesive in thecanopy of an apple orchard following a spray application on 9 July 2003.
% fibres lost after Position of fibre Top of leaf, overhanging Bottom of leaf, overhanging J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 Retention of fibres on apple trees was short-lived. Following the 1 June application in 1992, < 50% of marked fibres on leaves were retained on trees after 2 wk andapproximately 10% were retained after 7 wk. The 2003 study showed that the retention offibres is variable based on differences in their location and alignment on various substrates(Table 3). Following the 9 July application 58% of fibres were located on the top of leaves.
Deposition of fibres on the bottom of leaves and on fruit was similar with about 20% each.
Fibres deposited on the trunk and branches of trees accounted for < 5% of the total. Alarge proportion of fibres deposited initially on leaves were found overhanging the edge ofthe leaf. This was more common for fibres deposited on the underside of leaves with 64%of fibres overhanging. Retention of fibres was highest on the top of leaves with fruit beingthe second best. Fibres overhanging on leaves and all fibres deposited on the underside ofleaves were lost within 43 d. In comparison, 60% and 80% of the fibres deposited on fruitand bark were loss within 6 wk, respectively. Only a quarter of the fibres deposited on thetop of leaves and not overhanging were lost.
The experimental formulation of hollow fibres loaded with codlemone and mixed with an insecticide in this study was ineffective as an attracticide due to several factorsincluding the emission rate of the fibre and the toxicity of the adhesive. The initialemission rate of codlemone from individual fibres was apparently too high to allow mothcontact. Fibres had to be aged for > 7 d before male codling moths would contact fibresunder both flight tunnel and field conditions. Moth mortality was high following briefcontact with newly applied fibres but dropped rapidly with time. Modifications are neededto improve the performance of this attracticide approach.
The emission characteristics of sex pheromones from hollow fibres are well studied (Ashare et al. 1982). Fibres typically have an initial high release and then have a lower andfairly constant rate over an extended period of time. Previous studies with hollow fibresloaded with codlemone have shown that fibres can be long lived. Cardé et al. (1977)reported complete shutdown of lure-baited traps for 10 wk. Moffitt and Westigard (1984)reapplied fibres every 4 – 5 wk during the season. The emission rate of hollow fibres canbe adjusted by modifying either the internal diameter of the fibre or by changing the lengthof the fibre (Ashare et al. 1982). Modifications of these factors could likely improve theuse of fibres as an attracticide for codling moth.
Proper choice of an adhesive is critical in developing an effective attracticide. The viscosity of the adhesive affects both the application and the adhesion of the fibres. Thepolybutene adhesive Biotac has been widely used with hollow fibres (Beasley andHenneberry 1984, Moffitt and Westigard 1984) and is available in several formulationsthat differ in their viscosity and are appropriate for the range of temperatures experiencedfrom early spring to late summer. Yet, fibres were generally associated with limitedamounts of adhesive, < 1.0 mg; and were often poorly attached to the plant. In contrast,the initial laboratory studies placed fibres on large 10.0 mg drops of adhesive. This limitedamount of adhesive associated with fibres under field conditions formed a dry film thatwas not effective in transferring a toxic dose of insecticide to codling moth adults. Incomparison, the large drop of Biotac was toxic for several weeks in laboratory bioassays.
The use of non-drying grease or a different type of adhesive instead of Biotac might extendthe toxicity of the insecticide under field conditions.
Future improvements of the attracticide method for codling moth could include the use of a more concentrated insecticide dose. A 1.0% sticker formulation with permethrin andfenvalerate did not cause mortality of P. gossypiella while a 10.0% concentration waseffective (Haynes and Baker 1986). Yet, formulations with only 0.1% concentrations ofcyfluthrin killed 100% of codling moths when formulated in a castor oil-based paste (Lösel J. ENTOMOL. SOC. BRIT. COLUMBIA 100, DECEMBER 2003 et al. 2000). One attract and kill paste formulation currently registered for control ofcodling moth contains 6.0% permethrin (Charmillot et al. 2000). These paste formulationsremain effective against codling moth for at least 6 wk (Charmillot et al. 2000, Lösel et al2000).
The impacts of an attracticide approach can include both lethal and sublethal effects such as the interference with mate location by males (Haynes and Baker 1986). Whilesublethal effects were not examined in our field studies, previous flight tunnel tests withcodling moth found significant effects on male flight behaviours with concentrations ofesfenvalerate as low as 0.04% (unpublished data). Further studies that can characterize thesublethal effects of the range of attracticide formulations for codling moth would beuseful.
Depositing more fibres in the canopy would improve the effectiveness of this formulation both as an attracticide and for mating disruption of codling moth. Theapplication methods used to apply fibres have included specialized and expensive groundand air equipment (Moffitt and Short 1982). Results reported here suggest that thisapproach is ineffective in placing a significant number of hollow fibres in the tree canopy.
Fibres deposited in the apple tree canopy were primarily deposited in the middle of theupper leaf surface. This fibre position also appeared to be the most stable over time withnearly 75% of fibres retained after 6 wk. Unfortunately, the adhesion of fibres to bark orthe underside of leaves was low and short-lived. Increasing the number of fibres sprayedper hectare is one approach that could be used to increase the density of deposited fibres.
Ground applications in orchards with larger trees or denser canopies or perhaps the use ofaerial applications might improve the deposition rates of hollow fibres and needs to befurther examined.
I would like to thank John Turner (U.S.D.A., A.R.S., Yakima, WA) for his help in conducting the field studies and Tom Weissling (University of Nebraska, Lincoln, NE) forhis help in conducting the statistical analysis. Rick Hilton, Oregon State University,Eugene Miliczky, USDA, ARS, Wapato, WA, and Peter Shearer, Rutgers University,Princeton, NJ provided helpful reviews. This project received partial funding from theWashington Tree Fruit Research Commission, Wenatchee, WA.
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Excretion & internal environment of the body. In vertebrates the two functions of excretion and osmoregulation are • Excretion is the elimination to waste products from performed by kidneys and their associated structures in urinary system. • Waste products are unwanted and toxic by-products • The organs which form, store and void the urine