Sorghum Physiological Responses to Sugarcane Aphid Feeding
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Sorghum Physiological Responses to Sugarcane Aphid Feeding Juan D. Raya Vaca 1 , Adryenna Perez 1 , Rupesh Kariyat 1 , Sajjan Grover 2 , and Joe Louis 2,3 1 Department of Biology, University of Texas Rio Grande Valley 2 Department of Entomology and 3 Department of Biochemistry, University of Nebraska-Lincoln • Sugarcane aphids (Melanaphis sacchari) cause severe yield losses in sorghum (Sorghum bicolor) every year • Sugarcane aphids (SCA) use their slender stylets to penetrate plant tissue intercellularly to access the sieve elements for feeding. • Plants have evolved a wide range of defense mechanisms against insect infestations 1 , including callose deposition 2 • Callose deposits can serve as phloem blockers as a response to biotic stress, such as aphid and pathogen attack 2 • SC265 demonstrates resistance while SC1345 exhibits susceptibility towards SCA infestation (Fig. 2A) • SCA spent significantly less time in phloem phase while feeding on SC265 as compared to RTx430 (Fig. 2B) • SC265 had highest callose deposits per mm 2 at 24 h when compared to RTx430 and SC1345 (Fig. 2C) 1. Tetreault, H. M., et al. (2019). Global responses of resistant and susceptible sorghum (Sorghum bicolor) to sugarcane aphid (Melanaphis sacchari). Frontiers in plant science, 10, 145. 2. Amsbury, S., Kirk, P. and Benitez-Alfonso, Y. (2017). Emerging models on the regulation of intercellular transport by plasmodesmata-associated callose, Journal of Experimental Botany, 69, 105- 115. Additional reading on plant defenses • Plants and Insects: Sorghum plants were grown in the University of Nebraska-Lincoln greenhouse until reaching their third leaf stage 1 . SCA colony was reared in a 12:12hr light:dark cycle, at 26˚C • Aphid Bioassay: Ten adult wingless aphids were placed on each plant and total number of aphids were counted 7 days following initial infestation • Electrical Penetration Graph (EPG): The aphid stylet actions below the surface of the plant including probing, various feeding activities and the uptake of phloem sap studied using EPG technique (Fig. 1A). Data was analyzed using the EPG analysis Stylet+ software • Callose Quantification: Leaves were clip- caged with aphids (Fig. 1B) Harvested leaves were cleared in 100% Ethanol, then stained with 0.01% Aniline Blue [50 mM K 2 HPO 4 , pH 9.5] for 4 h. Slides were made with 50% glycerol and imaged with a flourescent microscope (Fig. 2D) BACKGROUND • Callose quantification at early time points (3, 6 and 12 h) • Expand studies on other β-glucan polymers used for plant’s defense mechanisms • Understand how other defense mechanisms are modulated with callose accumulation to limit phloem sap-feeding This research was supported by the National Science Foundation CAREER grant awarded to Dr. Joe Louis. RESEARCH OBJECTIVE To understand the physiological defense responses in sorghum against SCA feeding MATERIALS & METHODS RESULTS 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 0hr 24hr 48hr Mean Number of Callose Spots (mm 2 ) Time (h) RTx430 SC1345 SC265 C 0 1 2 3 4 5 6 Pathway Xylem Phloem Non-probing Time (h) RTx430 SC265 * * FUTURE DIRECTIONS REFERENCES ACKNOWLEDGEMENTS Fig. 2: (A) Mean number of aphids found on sorghum genotypes after 7 days of infestation (n=10). B) Comparative EPG waveform analysis for genotypes RTx430 (n=17) and SC265 (n=16) after a single, wingless adult aphid was wired and allowed to feed for 8 h. C) Mean number of callose spots identified under fluorescent microscope per mm 2 found on different sorghum genotypes (n=3) D) Arrows pointing to fluorescent Callose deposits in sorghum under 10X magnification. Asterisks above bars represent significant differences relative to RTx430. SIGNIFICANCE • Baseline study on novel sources of resistance in sorghum against SCA • Reduction of chemical usage in agriculture 0 20 40 60 80 100 120 140 160 180 RTx430 SC1345 SC265 Mean Number of Aphids Sorghum genotypes A B D 400um * ** A B Fig. 1: (A) The circuitry for a typical Electrical Penetration Graph (EPG) setup. (B) Illustration of aphid clip cages used for the callose quantification assay. EPG parameters
Transcript of Sorghum Physiological Responses to Sugarcane Aphid Feeding
Sorghum Physiological Responses to Sugarcane Aphid FeedingJuan D. Raya Vaca1, Adryenna Perez1, Rupesh Kariyat1, Sajjan Grover2, and Joe Louis2,3
1Department of Biology, University of Texas Rio Grande Valley2Department of Entomology and 3Department of Biochemistry, University of Nebraska-Lincoln
• Sugarcane aphids (Melanaphis sacchari) cause severe yield losses in sorghum (Sorghum bicolor) every year
• Sugarcane aphids (SCA) use their slender stylets to penetrate plant tissue intercellularlyto access the sieve elements for feeding.
• Plants have evolved a wide range of defense mechanisms against insect infestations1, including callose deposition2
• Callose deposits can serve as phloem blockers as a response to biotic stress, such as aphid and pathogen attack2
• SC265 demonstrates resistance while SC1345 exhibits susceptibility towards SCA infestation (Fig. 2A)
• SCA spent significantly less time in phloem phase while feeding on SC265 as compared to RTx430 (Fig. 2B)
• SC265 had highest callose deposits per mm2
at 24 h when compared to RTx430 and SC1345 (Fig. 2C)
1. Tetreault, H. M., et al. (2019). Global responses of resistant and susceptible sorghum (Sorghum bicolor) to sugarcane aphid (Melanaphis sacchari). Frontiers in plant science, 10, 145.
2. Amsbury, S., Kirk, P. and Benitez-Alfonso, Y. (2017). Emerging models on the regulation of intercellular transport by plasmodesmata-associated callose, Journal of Experimental Botany, 69, 105-115.
Additional reading on plant defenses
• Plants and Insects: Sorghum plants were grown in the University of Nebraska-Lincoln greenhouse until reaching their third leaf stage1. SCA colony was reared in a 12:12hr light:dark cycle, at 26˚C
• Aphid Bioassay: Ten adult wingless aphids were placed on each plant and total number of aphids were counted 7 days following initial infestation
• Electrical Penetration Graph (EPG): The aphid stylet actions below the surface of the plant including probing, various feeding activities and the uptake of phloem sap studied using EPG technique (Fig. 1A). Data was analyzed using the EPG analysis Stylet+ software
• Callose Quantification: Leaves were clip-caged with aphids (Fig. 1B) Harvested leaves were cleared in 100% Ethanol, then stained with 0.01% Aniline Blue [50 mM K2HPO4, pH 9.5] for 4 h. Slides were made with 50% glycerol and imaged with a flourescent microscope (Fig. 2D)
BACKGROUND
• Callose quantification at early time points (3, 6 and 12 h)
• Expand studies on other β-glucan polymers used for plant’s defense mechanisms
• Understand how other defense mechanisms are modulated with callose accumulation to limit phloem sap-feeding
This research was supported by the National Science Foundation CAREER grant awarded to Dr. Joe Louis.
RESEARCH OBJECTIVETo understand the physiological defense
responses in sorghum against SCA feeding
MATERIALS & METHODS
RESULTS
00.5
11.5
22.5
33.5
44.5
0hr 24hr 48hr
Mea
n N
umbe
r of C
allo
seSp
ots
(mm2
)
Time (h)
RTx430 SC1345 SC265C
0
1
2
3
4
5
6
Pathway Xylem Phloem Non-probing
Tim
e (h
)
RTx430 SC265
**
FUTURE DIRECTIONS
REFERENCES
ACKNOWLEDGEMENTS
Fig. 2: (A) Mean number of aphids found on sorghum genotypes after 7 days of infestation (n=10). B) Comparative EPG waveform analysis for genotypes RTx430 (n=17) and SC265 (n=16) after a single, wingless adult aphid was wired and allowed to feed for 8 h. C) Mean number of callosespots identified under fluorescent microscope per mm2 found on different sorghum genotypes (n=3) D) Arrows pointing to fluorescent Callose deposits in sorghum under 10X magnification. Asterisks above bars represent significant differences relative to RTx430.
SIGNIFICANCE• Baseline study on novel sources of resistance
in sorghum against SCA• Reduction of chemical usage in agriculture
020406080
100120140160180
RTx430 SC1345 SC265
Mea
n N
umbe
r of A
phid
s
Sorghum genotypes
A B
D
400um
*
**
A B
Fig. 1: (A) The circuitry for a typical Electrical Penetration Graph (EPG) setup. (B) Illustration of aphid clip cages used for the callose quantification assay.
EPG parameters
