BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, researchlibraries, and research funders in the common goal of maximizing access to critical research.

Boll Abscission Responses of Glyphosate-Resistant Cotton (Gossypium hirsutum) toGlyphosateAuthor(s): RYAN P. VIATOR, SCOTT A. SENSEMAN, JOE T. COTHRENSource: Weed Technology, 17(3):571-575. 2003.Published By: Weed Science Society of AmericaDOI: http://dx.doi.org/10.1614/WT02-166URL: http://www.bioone.org/doi/full/10.1614/WT02-166

BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the biological, ecological, andenvironmental sciences. BioOne provides a sustainable online platform for over 170 journals and books publishedby nonprofit societies, associations, museums, institutions, and presses.

Your use of this PDF, the BioOne Web site, and all posted and associated content indicates your acceptance ofBioOne’s Terms of Use, available at www.bioone.org/page/terms_of_use.

Usage of BioOne content is strictly limited to personal, educational, and non-commercial use. Commercial inquiriesor rights and permissions requests should be directed to the individual publisher as copyright holder.

571

Weed Technology. 2003. Volume 17:571–575

Boll Abscission Responses of Glyphosate-Resistant Cotton (Gossypium hirsutum)to Glyphosate1

RYAN P. VIATOR, SCOTT A. SENSEMAN, and JOE T. COTHREN2

Abstract: Response of glyphosate-resistant cotton to various rates of topically applied glyphosatewas investigated in growth chamber experiments to determine the relationship between glyphosaterate and boll abscission. Glyphosate at 0, 0.56, 1.12, or 2.24 kg ai/ha was applied to all exposedfoliage at the 12-leaf growth stage. Immediately after this treatment, 14C-glyphosate was applied tothe three uppermost fully expanded leaves at 0, 37, 74, or 148 kBq per leaf for the 0, 0.56, 1.12, or2.24 kg/ha treatment, respectively. After glyphosate application, glyphosate accumulated in repro-ductive tissue, and bolls were abscised. Abscission increased as the amount of glyphosate translocatedto fruiting sites increased.Nomenclature: Glyphosate; cotton, Gossypium hirsutum L., ‘Delta & Pine Land 5690RR’.Additional index words: Boll abscission; transgenic crops.Abbreviations: EPSPS, 5-enolpyruvylshikimate-3-phosphate synthase (EC 2.5.1.19); GR, glyphosateresistant; WAT, weeks after treatment.

INTRODUCTION

Glyphosate is a nonselective herbicide that binds to 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS)forming an EPSPS–shikimate-3-phosphate–glyphosatecomplex, which inhibits the biosynthesis of essential ar-omatic amino acids (Franz et al. 1997). Resistance toglyphosate has been transferred to cotton by addition ofthe glyphosate-insensitive CP4-EPSPS enzyme genecloned from Agrobacterium sp. strain CP4 (Barry et al.1992; Padgette et al. 1995). Use of glyphosate-resistant(GR) cotton allows improved utility of the nonselectiveherbicide glyphosate (Padgette et al. 1996).

The use of GR cotton is limited by the applicationwindow for topical treatments of glyphosate. Accordingto current label requirements, topical treatment with gly-phosate can only be applied before the fifth true-leafstage. Applications after this stage must be postdirectedwith limited exposure of the cotton plant (Anonymous1999). This application window limits the utility of top-ical applications of glyphosate, which may be necessaryto control in-row weeds (Matthews et al. 1998). Appli-cations of glyphosate inconsistent with label recommen-dations have resulted in abnormal flowers with partially

1 Received for publication September 17, 2002, and in revised form March4, 2003.

2 Research Assistant, Associate Professor, and Professor, Department of Soiland Crop Sciences, Texas A&M University, Box 2474, College Station, TX77843. Current address of first author: Crop Science Department, North Car-olina State University, Box 7620, Raleigh, NC 27695. Corresponding author’sE-mail: ryanviator@hotmail.com.

developed anthers (Barnea et al. 2000; Pline et al.2002a), boll abscission, and concomitant yield loss (Ka-laher et al. 1997). Moreover, yield losses have occurredwith the standard glyphosate program that consists of atopical application before the fifth true-leaf stage fol-lowed by a postdirected application at the seventh oreighth true-leaf stage (Brown and Bednarz 1998). Incontrast, other researchers have failed to show yield loss-es after labeled and nonlabeled glyphosate applications(Blackley et al. 1999; Jones and Snipes 1999; Matthewset al. 1998; Murdock 1999), but some reported reducedboll retention (Blackley et al. 1999; Jones and Snipes1999). Reduced boll retention can result in delayed ma-turity, which can cause harvest delays. Harvest delaysoften subject a crop to adverse weather, often reducingfiber quality and lint yield (Jones and Snipes 1999).

Environmental factors also are associated with bolldrop, abnormal bolls, and subsequent yield reductions(Kerby and Voth 1998). Preliminary studies with GRcultivars indicate that cool early-season temperatures orhigh nighttime temperatures in association with high hu-midity at midseason could cause boll abscission, abnor-mal bolls, and yield reductions (Kerby and Voth 1998).Previous greenhouse research on non-GR cotton indi-cated that glyphosate toxicity increased at 25 C com-pared with 35 C and that absorption and translocation ofglyphosate increased with 100% relative humidity com-pared with 40% (Wills 1978). Another report indicatesthat suboptimal temperatures have little effect on gly-phosate uptake and translocation (File et al. 1998).

VIATOR ET AL.: BOLL ABSCISSION RESPONSES TO GLYPHOSATE

572 Volume 17, Issue 3 (July–September) 2003

Wills (1978) reported that more glyphosate translo-cated after treatment to the mature lower stem than tothe mature lower leaves, immature upper stem, or upperleaves of non-GR cotton. Pline et al. (2001) found great-er absorption of 14C-glyphosate after application to lowerstems than when applied to upper leaves of GR cotton.This research also reported translocation of glyphosateto squares and bolls and that these fruiting structuresretained 0.2 to 3.7% of the applied 14C-glyphosate. Otherresearch, however, suggests that reproductive tissuesmay retain as much as 30% of absorbed glyphosate (Har-ris and Vencill 2000). Accumulation of glyphosate inreproductive tissue is expected because glyphosate fol-lows the same translocation patterns as photoassimilates(Franz et al. 1997). The objective of this research wasto determine the relationship between glyphosate rateand fruit abscission of GR cotton in a controlled envi-ronment.

MATERIALS AND METHODS

Glyphosate-resistant cotton cv. ‘Delta & Pine Land5690RR’ was grown under controlled conditions in agrowth chamber at temperatures of 26 and 24 C (dayand night, respectively) and with 90% relative humidity.Seeds were planted in a commercial potting medium3 in19-L pots. Light intensity was maintained at 1,000 mmol/m2 s photosynthetically active radiation, using a combi-nation of fluorescent and incandescent lights to providea 14-h day length.

Glyphosate was applied postemergence over-the-top at0, 0.56, 1.12, or 2.24 kg ai/ha to all exposed cotton fo-liage at the 12-leaf stage using a CO2-pressurized back-pack sprayer calibrated at a carrier volume of 187 L/ha.Previous research indicated that this stage of cotton de-velopment was susceptible for fruit abscission inducedby one application of glyphosate (Matthews et al. 1998).Furthermore, cotton plants at this growth stage containdeveloping fruit structures at different physiological ma-turity, from flower buds to bolls. Thus, it was possibleto determine glyphosate effects on fruiting structures atdifferent physiological stages with a single glyphosateapplication. Cotton was treated at this stage in an effortto abscise bolls; the treatment was different from thoselisted on the label. Furthermore, at this stage, there aremore fruiting structures present than during earliergrowth stages. Immediately after the glyphosate appli-

3 Metro-Mix 300, Scotts-Sierra Horticulture Products Co., 14111 Scotts-lawn, Marysville, OH 43041.

cation, 14C-glyphosate4 (7.40 MBq/ml) was applied witha microsyringe to the three uppermost, fully expandedleaves as five droplets (1 ml) at 0, 37, 74, or 148 kBqper leaf for the 0, 0.56, 1.12, or 2.24 kg/ha treatments,respectively. This method was previously used to inves-tigate glyphosate toxicity to non-GR cotton (Wills 1978)and shikimic acid accumulation in GR cotton (Pline etal. 2002b). Each plant was then returned to the growthchamber and placed in a screen box to ensure that thebolls that abscised from each plant remained associatedwith that plant.

Plant mapping5 was conducted to determine boll dis-tribution 8 wk after treatment (WAT), at which time allabscised bolls were collected individually with node andposition recorded (Heitholt 1997). This mapping waspossible because bolls did not form a complete abscis-sion zone, and there was limited air movement in thechamber; thus bolls, even though partially abscised anddried, clung to the plant. Experiment duration was in-creased past physiological cutout to 8 WAT to specifi-cally identify the biological significance of glyphosateapplication.

Leaves, stems, and roots were collected separately. Allmaterials were oven-dried for 1 wk at 50 C in a radiantheat oven6 and ground with a mechanical grinder7 usinga 20-mesh screen. A 0.1- to 0.2-g subsample from eachboll was oxidized using a biological oxidizer8 to captureabsorbed 14CO2. The amount of 14CO2 from the oxidizedmaterial was quantified using liquid scintillation spec-troscopy.9 The amount of evolved 14CO2 was then usedto calculate the amount of glyphosate from which it wasderived. The remaining plant matter, including plantroots, was collected, dried, oxidized, and analyzed to de-termine total 14CO2 recovery. Metabolism of glyphosatewas not examined as a result of negligible metabolismby plants (Duke 1988). Thus, in this study, all 14CO2 wasassumed to be from 14C-glyphosate.

To induce boll abscission, applications at rates equiv-alent to field application rates had to be made; thus, both14C-glyphosate and glyphosate were applied. Further-more, the total of both 14C-glyphosate and glyphosate

4 Amersham Life Science Inc., 2636 South Clearbrook Drive, ArlingtonHeights, IL 60005-4692.

5 Plant Map Analysis Program, Version 3.7, Texas A&M Research and Ex-tension Center, Route 2, Box 589, Corpus Christi, TX 78406-9704.

6 Lab-Line radiant heat oven, Lab-Line Instruments, Inc., 15 BloomingdaleAvenue, Melrose Park, IL 60160.

7 Mechanical grinder, Thomas Company, 704 Preston Street, Philadelphia,PA.

8 Harvey biological oxidizer, R. J. Harvey Instrument Corporation, 123 Pat-terson Street, Hillsdale, NJ 07642.

9 Beckman liquid scintillation counter, Beckman-Coulter, Inc., 4300 NorthHarbor, Fullerton, CA 92834-3100.

WEED TECHNOLOGY

Volume 17, Issue 3 (July–September) 2003 573

Figure 1. Glyphosate concentration in abscised bolls in response to increasingrate of glyphosate application. Data are the average of two trials with fourreplicates each.

Figure 2. Boll abscission in response to increasing rate of glyphosate appli-cation. Data are the average of two trials with four replicates each.

was used for rate, translocation, and accumulation cal-culations. The total amount of 14C-glyphosate quantifiedby liquid scintillation spectroscopy was divided by thetotal amount of 14C-glyphosate applied to determinetranslocation. Percent translocation was then multipliedby the concentration of glyphosate applied to determinethe amount of glyphosate translocated to fruiting struc-tures. Location of abscissions was analyzed by the fol-lowing zones: nodal positions 6 to 11, 12 to 16, and 17to 26. Nodal positions were grouped into these specificzones to analyze the lower, middle, and upper fruitingprofiles, as in previous fruit removal research (Jones etal. 1996). All horizontal fruiting positions on a particularnode were combined for this grouping.

Treatments were arranged in a randomized completeblock with four replications, with the experiment con-ducted twice in the same growth chamber over time.Data were analyzed regressing glyphosate rate vs. gly-phosate concentration, glyphosate rate vs. boll abscis-sion, glyphosate rate vs. boll retention, glyphosate ratevs. fruit retention, and glyphosate concentration vs. bollabscission using SAS (version 8). Linear, quadratic, cu-bic, log-logistic, segmented line, and nonlinear modelswere compared for the best fit. When possible, data fromglyphosate treatments were averaged across trials. Ho-mogeneity of error variances was tested as described byK. A. Gomez and A. A. Gomez (1984) to determinewhether trials could be combined.

RESULTS AND DISCUSSION

As rates of glyphosate applied topically to cotton in-creased, the amount of glyphosate translocated to bolls

increased (Figure 1). Furthermore, the number of bollsabscised per plant increased with increasing glyphosaterate (Figure 2). Glyphosate concentrations in the ab-scised fruit were 0 mg/g, 15 6 7 mg/g, 35 6 10 mg/g,or 126 6 37 mg/g when glyphosate was applied at 0,0.56, 1.12, or 2.24 kg/ha, respectively. Bolls retained onthe plant contained less glyphosate than those abscisedfor all glyphosate rates. Concentrations of glyphosate inretained bolls were 0 mg/g, 10 6 6 mg/g, 10 6 7 mg/g,and 7 6 3 mg/g for rates 0, 0.56, 1.12, and 2.24 kg/ha,respectively.

Concentrations of glyphosate in abscised bolls repre-sented 0.20, 0.25, and 0.43% of the total glyphosate ap-plied topically to the plant for rates 0.56, 1.12, and 2.24kg/ha, respectively. Pline et al. (2001) reported a 2.5%accumulation of applied 14C-glyphosate in bolls at cut-out. Differences in glyphosate accumulation betweenthese two studies are probably because the entire plantwas treated with glyphosate in the current study similarto that reported by Wills (1978), whereas Pline et al.(2001) treated only a 5-cm2 plant section. Furthermore,the duration of the current experiment was past physio-logical cutout to specifically identify the biological sig-nificance of glyphosate accumulation, which was abscis-sion. The additional 7 wk of experiment duration in thisstudy may explain the 10-fold difference in accumula-tion when compared with reports by Pline et al. (2001).

Glyphosate rates in this study did not affect the ver-tical location of the first reproductive node on the plant(data not shown). The vertical location of the first repro-ductive node was used as the criterion for early selectionof GR lines (Nida et al. 1996). However, glyphosate de-creased the first-position bolls and the first- and second-

VIATOR ET AL.: BOLL ABSCISSION RESPONSES TO GLYPHOSATE

574 Volume 17, Issue 3 (July–September) 2003

Figure 3. Boll retention of the first-position bolls and the first- and second-position bolls combined in response to increasing rate of glyphosate appli-cation. Data are the average of two trials with four replicates each. Figure 4. Fruit retention for the lower (nodes 6 to 11), middle (nodes 12 to

16), and upper (nodes 17 to 26) fruiting profiles in response to increasing rateof glyphosate application. Data are the average of two trials with four repli-cates each.

Figure 5. Boll abscission in response to increasing glyphosate concentrationsin bolls abscised per plant. Data are the average of two trials with four rep-licates each.

position bolls combined, with the 1.12 and 2.24 kg/hatreatments reducing boll numbers by 45% comparedwith the nontreated control (Figure 3). The reduction ofthe first- and second-position bolls is important becausethese positions contribute 84 to 96% of cotton yield (Jen-kins et al. 1990). Furthermore, the loss of bolls at thesepositions is not fully compensated for by bolls set furtherout on the same fruiting branch (Constable 1991). How-ever, cotton can compensate for both early- and midsea-son fruit loss by setting bolls on higher sympodialbranches, assuming a sufficiently long growing season.In situations where the growing season is shortened orenvironmental conditions are inadequate, boll abscissioncan lead to yield loss (Jones and Snipes 1999).

In addition to the effects on boll number, glyphosatealso reduced the number of fruiting structures (bolls,flowers, and squares) on the lower, middle, and upperparts of the plant (Figure 4). Previous research has notreported flower and square loss and boll loss only on thelower portion of the plant (Blackley et al. 1999; Jonesand Snipes 1999). However, these studies had earlier ap-plication timings, which could have affected the absorp-tion levels, translocation patterns, and location of ab-scission on the plant. Variations in environmental con-ditions, such as relative humidity, have affected the tox-icity of glyphosate in non-GR cotton (Wills 1978). Theamount of glyphosate at each nodal position did not dif-fer because of the late 12-leaf stage timing, suggestingthat glyphosate was distributed throughout the plant,rather than being concentrated in certain fruiting zoneson the plant. This distribution could further explain whyabscission occurred throughout the fruiting profile. In ad-dition, 14C-glyphosate was found in plant roots, leaves,

and stems, with 90% recovery for these experiments(data not shown).

This research demonstrates that after glyphosate ap-plication, glyphosate accumulates in reproductive tissueand bolls abscise in GR cotton. Moreover, abscission in-creased as the amount of glyphosate translocated to fruit-ing sites increased (Figure 5). Although abscission in-creased with increasing concentrations of glyphosate, en-vironmental factors, such as cool early-season tempera-tures and high midseason night humidity, may alsocontribute to the abscission problem. The unrelated en-vironmental factors that appear to compound fruit ab-scission caused by glyphosate applications warrant fu-

WEED TECHNOLOGY

Volume 17, Issue 3 (July–September) 2003 575

ture research. This research may help explain inconsis-tent responses to glyphosate on GR cotton under fieldconditions.

ACKNOWLEDGMENTS

We thank Cotton Incorporated and Monsanto Com-pany for partially funding this project. We also thank Dr.Wendy Pline for editing assistance and Dr. CavellBrownie for statistical consultation.

LITERATURE CITED

Anonymous. 1999. Roundup Ultra supplemental label 2137X3-20. St. Louis,MO: Monsanto.

Barnea, N., Y. Alon, N. Sibony, and B. Rubin. 2000. The effect of late seasonglyphosate applications on cotton plants and weeds. Proc. Beltwide Cot-ton Conf. 1:663–665.

Barry, G., G. Kishore, S. Padgette, et al. 1992. Inhibitors of amino acid bio-synthesis: strategies for imparting glyphosate resistance to crop plants.In B. K. Singh, H. Flores, and J. C. Shannon, eds. Biosynthesis andMolecular Regulation of Amino Acids in Plants. Rockville, MD: Amer-ican Society of Plant Physiology. Pp. 139–145.

Blackley, R. H., D. B. Reynolds, C. D. Rowland Jr., and S. L. File. 1999.Roundup Ready cotton tolerance to topical applications of Roundup Ul-tra. Proc. South. Weed Sci. Soc. 52:252–253.

Brown, S. M. and C. W. Bednarz. 1998. Tolerance of Roundup Ready cottonto mid- and late-post application of Roundup. Proc. Beltwide CottonConf. 1:849–850.

Constable, G. A. 1991. Mapping the production and survival of fruit on field-grown cotton. Agron. J. 83:374–378.

Duke, S. O. 1988. Glyphosate. In P. C. Kearney and D. D. Kaufman, eds.Herbicides: Chemistry, Degradation and Mode of Action. New York:Marcel Dekker. Pp. 1–70.

File, S. L., D. B. Reynolds, C. E. Snipes, and B. E. Serviss. 1998. RoundupReady cotton tolerance to topical and post-directed applications ofRoundup Ultra. Proc. Beltwide Cotton Conf. 1:850.

Franz, J. E., M. K. Mao, and J. A. Sikorski. 1997. Glyphosate: A UniqueGlobal Herbicide. Washington, DC: American Chemical Society. Pp.521–605.

Gomez, K. A. and A. A. Gomez. 1984. Analysis of data from a series ofexperiments. In K. A. Gomez and A. A. Gomez, eds. Statistical Proce-

dures for Agricultural Research. 2nd ed. New York: J. Wiley. Pp. 316–356.

Harris, H. M. and W. K. Vencill. 2000. Uptake and translocation of 14C-glyphosate in glyphosate-resistant cotton. Weed Sci. Soc. Am. Abstr. 40:65–66.

Heitholt, J. J. 1997. Floral bud removal from specific fruiting positions incotton: yield and fiber quality. Crop Sci. 37:826–832.

Jenkins, J. N., J. C. McCarty, and W. L. Parrot. 1990. Effectiveness of fruitingsites in cotton: yield. Crop Sci. 30:365–369.

Jones, M. A. and C. E. Snipes. 1999. Tolerance of transgenic cotton to topicalapplications of glyphosate. Cotton Sci. 3:19–26.

Jones, M. A., R. Wells, and D. S. Guthrie. 1996. Cotton response to seasonalpatterns of fruit removal II—growth and dry matter accumulation. CropSci. 36:639–645.

Kalaher, C. J., H. D. Coble, and A. C. York. 1997. Morphological effects ofRoundup application timings on Roundup Ready cotton. Proc. BeltwideCotton Conf. 1:780.

Kerby, T. and R. Voth. 1998. Roundup Ready introduction experiences in1997. Proc. Beltwide Cotton Conf. 1:26–29.

Matthews, S. G., G. N. Rhodes, T. C. Mueller, and R. M. Hayes. 1998. Effectsof Roundup Ultra on Roundup Ready cotton. Proc. Beltwide CottonConf. 1:850.

Murdock, E. C. 1999. Tolerance of Roundup Ready cotton to multiple post-emergence applications of glyphosate. Proc. South. Weed Sci. Soc. 52:5.

Nida, D. L., K. H. Kolacz, R. E. Buehler, et al. 1996. Glyphosate-resistantcotton: genetic characterization and protein expression. J. Agric. FoodChem. 44:1960–1966.

Padgette, S. R., K. H. Kolacz, X. Delannay, et al. 1995. Development, iden-tification, and characterization of a glyphosate-tolerant soybean line.Crop Sci. 35:1451–1461.

Padgette, S. R., D. B. Re, G. F. Barry, D. E. Eichholtz, X. Delannay, R. L.Fuchs, G. M. Gishgore, and R. T. Fraley. 1996. New weed control op-portunities. In S. O. Duke, ed. Herbicide Resistant Crops: Agricultural,Environmental, Economic, Regulatory, and Technical Aspects. NewYork: Lewis Publishers. Pp. 53–84.

Pline, W. A., A. J. Price, J. W. Wilcut, K. L. Edmisten, and R. Wells. 2001.Absorption and translocation of glyphosate in glyphosate-resistant cottonas influenced by application method and growth stage. Weed Sci. 49:460–467.

Pline, W. A., R. P. Viator, J. W. Wilcut, K. L. Edmisten, J. Thomas, and R.Wells. 2002a. Reproductive abnormalities in glyphosate-resistant cottoncaused by lower CP4-EPSPS levels in the male reproductive tissue. WeedSci. 50:438–447.

Pline, W. A., J. W. Wilcut, S. O. Duke, K. L. Edmisten, and Randy Wells.2002b. Tolerance and accumulation of shikimic acid in response to gly-phosate applications in glyphosate-resistant and nonglyphosate-resistantcotton (Gossypium hirsutum L.). J. Agric. Food Chem. 50:506–512.

Wills, G. D. 1978. Factors affecting toxicity and translocation of glyphosatein cotton (Gossypium hirsutum). Weed Sci. 26:509–513.