PMAS-ARID AGRICULTURE UNIVERSITY RAWALPINDI DEPARTMENT OF PLANT PATHOLOGY Synopsis for Degree of Ph.

D Plant Pathology

Title: Spatio-temporal and disease predictive modeling of bacterial seed and boll rot of cotton based on climate change pattern and predictive disease management strategies through induced systemic resistance under changing environmental conditions.

Name of the student

Muhammad Ehetisham-ul-Haq

Abstract Elevated CO2 atmospheric level continuously changing environmental patterns. Shifts in climatic events effecting plant pathosystems. Cotton is an important commercial crop of global importance known as white gold. Bacterial seed and boll rot is a newly emerging threat to cotton economy of Pakistan. Systemic acquired resistance (SAR) is novel in plant disease management. Present study will be aimed to assess spatio-temporal disease distribution of bacterial seed and boll rot of cotton based on climatic change events. Disease predictive model based on three year data will be developed under changing environmental pattern. Assessing the impact of elevated temperature on induced systemic resistance in lab and in field conditions, a simulation model will be devised.

Need for the Project

Cotton is an important commercial crop of global importance. It is known as the king of fiber and is called white gold playing a key role in many socio-economic aspects of the world. It is grown in more than 60 countries of the world. Total world cotton production in 2011 was 123.3 million bales, more than 6 percent from the preceding year. Globally in 201112, harvested area was recorded 35.7 million hectares which was 7 percent above the previous year. Global cotton yield in 2011-12 was 752 kg ha-1 about unchanged from 2010-11 (Johnson et al., 2012). In Pakistan 2012, the crop was sown under the area of 3350 thousands hectares and now total production of 14.34 million bales is being expected (SUPARCO, 2012). Cotton accounts for 8.2 % of the value-added in the agriculture sector and about 2 percent to GDP, adds over $2.8 billion to the national economy which is the 60% of total foreign exchange earned through the export of raw cotton and cotton products (Economic Survey, 2012). Population explosion is a continuous alarming threat to the environment. Producing more to meet human needs, industries have been tremendously increased since last decades (Gleick, 2003). Despite coming up with human needs, waste products are posing a serious menace to biotic and abiotic environment directly or indirectly. Emitting gasses from factories and automobiles are the major fact of global warming. A considerable increase in CO2 level (more than 400 ppm) has been assessed since last couple of decades which was 150-300 ppm measuring from ice-core since 650,000 years ago (Chakraborty et al., 2008). Havier in weight than other gasses, CO2 is continuously depositing biosphere absorbing the sunlight and making the globe warmer (Mikkelsen et al., 2008). Shift in temperature increase from 0.6 -0.7 oC globally as compared to last century posing a serious change in shorter and warmer winters in many regions of the world (Benvenuti, 2009). Host, Pathogen and environment primarily factorizing in pathogen establishment and disease progression. Host susceptibility, pathogen virulence and optimum environmental conditions (vary from pathogen to pathogen) are the key factors for disease induction. Predisposing factors play significant role on plant-pathogen interactions. Climate change may alter stage and rate of pathogen development, modify host resistance and may induce change in physiology of plant pathosystem. Elevated temperature events may induce plant stress resulting into wilting, leaf burn, leaf folding, abscission and changes in physiological responses including RNA metabolism and protein synthesis may tend to favor host susceptibility (Clark et al., 1982). Specific time

period and host plant age require to interact pathogen on host for disease progression (Agrios, 2005). Change in global temperature may prolong this time period to interact plant and pathogen increases compatibility chances. Globally Continuous temperature elevation pattern directly effecting crops yield. Schlenker and Lobell (2010) found a decrease in production of 8% for cassava with much more severe impacts for maize (-22%), sorghum (-17%) and millets (-17%). Systemic acquired resistance (SAR) develops in plants at a distance from the local lesion phenomenon firstly pioneered by Kuc. Plants given an initial inoculum results in local necrotic lesions become resistance latterly to secondary inoculum. The resistance ingressed may delay or reduce symptoms expression. Pathogen used as a secondary inoculum may be the same as for the primary inoculum or entirely different one since SAR is often effective against wide range of plant pathogens. The period over SAR develops varies according to particular plant and primary inoculum and may be as short as a few hours to two to three weeks (Strange, 2006). Cotton is attacked by a number of diseases inducing severe reduction in yield by influencing germination, killing the plants, reducing plant productivity and affecting the quality of lint. Root-rot, fungal wilt, bacterial wilt, anthracnose, cotton leaf curl and bacterial rust are the major diseases of cotton. Bacterial seed and boll rot was firstly observed in South Carolina, USA in 1999 (Jones et al., 2000). The disease costs 10 to 15 percent yield loss in USA (Hudson, 2000). In 2006, it was observed in Xinjiang Province of China (Ren et al., 2008) where yield losses are up to 20 percent in many fields. Bacterial seed and boll rot is caused by a bacterium Pantoea agglomerans(Medrano & Bell, 2007; Ren et al., 2008) and is transmitted by southern green stink bug (Nezara viridula L) (Medrano et al., 2007; Medrano et al., 2009a; Medrano et al., 2009b). Early detection of disease is very complicated, boll seems to be healthy because no symptoms appear on the outer surface of boll. Disease can only be observed when bolls are cross sectioned or opened. Immature diseased locule is attributed as hard lock having brown and dense lint. At harvesting time, the infected bolls either drop earlier or not picked efficiently (Hudson, 2000). In Pakistan, bacterial seed and boll rot is a new emerging and devastating disease of cotton. It was observed in different areas of the country including Faisalabad and Multan regions where cotton is extensively grown.Disease hit severely cotton fields of the country last

year and caused huge damage. In future, if no attention is given, the disease will become epidemic and will crush the cotton fields resulting poor yield and bad quality lint fiber. It will be a great threat for cotton growers, textile industry and ultimately on Pakistans economy. Objectives To assess the spatio-temporal distribution of bacterial seed and boll rot of cotton based on climatic changes. To develop a disease predictive model based on three year data under changing environmental pattern. To develop a predictive model assessing the impact of elevated temperature on induced systemic resistance in lab and in field conditions.

REVIEW OF LITERATURE
Bacterial boll rot of cotton was firstly reported in July, 1999 from Hampton County, South Carolina, USA. (Jones et al., 2000). Bacteria were isolated from diseased locules of cotton (Hollis, 2001; Hudson, 2000). It was recorded in Xinjiang, China in 2006 by Ren et al. (2008). Edmisten (1999), found the exterior walls of carpels of unopened diseased bolls are non-symptomatic and hence the early detection is complicated in affected fields. Cross-sections of diseased fruit reveal necrotic seed and discolored fiber tissue. The disease symptoms can only be observed when infected immature bolls are cross sectioned or opened, discolored lint and diseased locule attributed as hard lock are the prominent symptoms (Hudson, 2000). Anatomical and developmental differences between embryos dissected from typical cotton seed and bolls exhibiting hollow seed was described by (Mauney et al., 2004). Ren et al. (2008), isolated bacteria on Kings media B and characterized forty eight strains with yellow pigmentation. He also studied the physico-chemical properties of the bacterium and described it as the gram negative, facultatively anaerobic, unable to produce indole from tryptophan, able to reduce nitrate to nitrite and produce acid from glucose, cellobiose, lactose, melibiose, and from melonate. Medrano and Bell (2007), took flame sterilized glass rod, rubbed it on discolored seed and streaked on tryptic soy agar (TSA) medium. Secondly, seeds were soaked in sterile water and extracct was spread on TSA. Plates were incubated at 25C until colonies appeared. Individual colonies were purified. Bacterium was identified on the basis of phenotype testing,

fatty acid profiling and 16s ribosomal DNA sequence analysis and categorized as Pantoea agglomerans. In the greenhouse study, they used a syringe and needle to inject a suspension of P. agglomerans rifampicin (Rif)-resistant mutant (Sc 1-R) into healthy cotton bolls, after two weeks of exposure to the bacterium, inoculated bolls were harvested and infection symptoms were similar to those observed in diseased field-grown South Carolina. Marois and Wright (2004) grew four cotton varieties (Suregrow 501 BR, Delta Pine, Land (DPL) 458 B/RR and DPL 555 B/RR) to study the relationship between boll rot of cotton and disease severity. Disease severity ranged from 40 to 95% after 8 weeks of sowing. A strong positive correlation (r = 0.935) was observed between disease severity and environmental factors. Khan et al. (1999), recorded disease severity of bacterial blight of cotton with six cotton varieties (CIM-1100, CIM-109, CIM-435, NIAB-78,NIAB-Krishma and SLS-1) in five different districts (Rajanpur, D.G.Khan, Muzaffarghar, Multan and Jhang) of Punjab province. Disease severity data was correlated with environmental factor (maximum and minimum air temperature, rainfall, relative humidity). All environment variable except maximum temperature responded significantly to disease severity. Maximum disease severity was noticed at 100-147 mm rainfall, 26-27 0C maximum temperature and 67-77 % relative humidity. Qamar et al. (2003)who studied the correlation of disease severity of potato virus X, Y and PLRV and aphid population with environmental conditions on five potato varieties/lines i.e. Adora, Desiree, Lesieta, Mondial and Rodeo on weekly basis. They found the overall correlation of maximum (24-42C) and minimum (15-22C) air temperature with disease severity. Maximum temperature, solar radiation, dew point and wind speed were significantly correlated with aphid population which was significantly correlated with PLRV. Batool et al. (2011) studied disease severity and aphid populationregarding to environmental conditions . Significant relationship was observed between all environmental vriables and PLRV disease severity except wind velocity. Air temperature (maximum and minimum), relative humidity, pan evaporation and wind speed siginificantly influenced on disease severity. Except pan evaporation,all environmental parameters significantly effected aphid popualtion. Ali (2011), found a significant (P<0.05) but negative correlation between maximum temperature and CLCuD. The relationship of rainfall with CLCuD was found non significant. Wind velocity was found non significant but negatively correlated with CLCuVD. Whitefly had significant and positive correlation with CLCuVD. The relative humidity contributed significantly disease and the wind velocity did not contribute as effectively as compared to

other variables. in the buildup of whitefly population. Relative humidity did play a prominent role in the spread of the. Hamid et al. (2012) correlated bacterial blight disease severity of 25 varieties with environmental conditions (maximum and minimum air temperature, rainfall, relative humidity, wind speed, pan evaporation and radiation). Except wind speed and radiation, significant impact of enviromental factor (maximum and minimum air temperature, rainfall, relative humidity and pan evaporation) on disease severity was noticed.

MATERIALS AND METHODS

Collection of three years data of bacterial seed and boll rot cotton, red cotton bug and environmental conditions from different districts. Data of bacterial seed and boll rot disease, red cotton bug population density and environmental conditions will be collected from different districts of cotton growing areas of Punjab (Pakistan). The influence of each environmental variable (Maximum and minimum air temperature, rainfall, relative humidity wind velocity and CO2) on seed and boll rot and red cotton bug population density will be determined by correlation and regression analysis. Regression analysis Regression analysis uses quantitative independent variables to explain variation in quantitative dependent variables. Linear regression models can be separated into two broad categories: simple regression models and multiple regression models. In simple linear regression models, the dependent variable (Y) is a direct function of an independent (or explanatory) variable (X). The following is the general equation for simple linear regressions: Y = 0 + 1X Where 0 is the intercept and 1 is the slope. For multiple linear regression models, multiple independent variables are used to predict the response of the dependent variable. (Graham, 2003) Area under disease progress curve Area under disease progressive curve (AUDPC) was calculated by the trapezoidal integration of the disease incidence over time for each variety/advance line, considering the whole period evaluated (Shaner & Finney, 1980), as follows.

Where n is the number of assessment; X, disease incidence (%); and (ti+1-ti), duration between two consecutive assessments. The disease assessments (bacterial seed and boll rot incidence) over specific periods of time interval (bi-weekly) will be recorded during the experiment and will be interpreted according to the above mentioned formula.

Simple growth model and Linked differential equation (LDE) Growth models have been used almost exclusively as statistical means to describe the observed patterns and then use the estimated model parameters for comparing epidemics. To assess the epidemic disease pattern of bacterial seed and boll rot of cotton, Gompert growth curve model will be used to assess the disease spatio-temporal patterns.

REFERENCES
Agrios, G. 2005. Plant Pathology. 5th eds, New York: Academic Press. Ali, S. 2011. Prediction of Cotton Leaf Curl Virus Disease and its vector based on epidemiological factors and its management, University of Agriculture, Faisalabad. Batool, A., Khan, M.A., Farooq, J., Mughal, S.M., Iftikhar, Y. 2011. ELISA-based screening of potato germplasm against potato leaf roll virus. Journal of Agricultural Research, 49(1), 57-63. Benvenuti, S. 2009. Potenziale impatto dei cambiamenti climatici nellevoluzione floristica di fitocenosi spontanee in agroecosistemi mediterranei. Rivista Italiana di Agronomia, S1, 45-67. Chakraborty, S., Luck, J., Hollaway, G., Freeman, A., Norton, R., Garrett, K.A., Percy, K., Hopkins, A., Davis, C., Karnosky, D.F. 2008. Impacts of Global Change on Diseases of Agricultural Crops and Forest Trees. Perspectives in Agriculture, Veterinary Science, Nutrition and Natural Resources, 3(054), doi:10.1079/PAVSNNR20083054. Clark, R., Cristiansen, M., Lewis, C. 1982. Breeding plants for less favorable environments. Breeding plants for less favorable environments. Edmisten, K. 1999. Supernumerary carpel syndrome and cotton seed rot. Carolina Cotton Notes Available at: http://www.cotton.ncsu.edu/ccn/1999/ccn-99-8a.htm (accessed: November 2012). Gleick, P.H. 2003. Global freshwater resources: soft-path solutions for the 21st century. Science, 302(5650), 1524-1528. Graham, M.H. 2003. Confronting multicollinearity in ecological multiple regression. Ecology, 84(11), 2809-2815. Hamid, M.I., Khan, M.A., iqbal, Z., Ghazanfar, M.U., Iftikhar, Y., Akhtar, N. 2012. correlation of environmental condition with bacterial blight disease of cotton. Pakistan Journal of Phytopathology, 24(1), 39-43.

Hollis, P. 2001. Seed rot has lowered South Carolina cotton yields. Southeast Farm Press. Available at: http://southeastfarmpress.com/mag/farming_seed_rot_lowered/ (accessed: November 2012). Hudson, J. 2000. Seed rot hits South Carolina cotton. Southeast Farm Press. Available at: http://southeastfarmpress.com/mag/farming_seed_rot_hits/ 2012). Johnson, J., Kiawu, J., MacDonald, S., Meyer, L., Skelly, C. 2012. The world and United States cotton outlook. Association: United States Department of Agricultural> Agricultural Outlook Forum 2012. Jones, M., Mueller, J., Kluepfel, D., Sullivan, M., Walker, J., Roof, M., Stewart, J., Linvill, D. 2000. Preliminary investigations on cotton seed rot in South Carolina. Clemson University Station Bulletin, 675, 21. Khan, M.A., Khan, H.A., Ilyas, M.B., Rashid, A. 1999. Correlation of environmental conditions with bacterial blight disease on six commercially grown cotton cultivars in five districts of the punjab. Pakistan Journal of Agricultural Sciences, 36, 1-2. Marois, J., Wright, D. 2004. Etiology, epidemiology, and control of Fusarium hardlock of cotton in the southeast: the possibilities. Proceedings Beltwide Cotton Conference, San antonio, TX. pp. 5-9. Mauney, J.R., Stewart, J.M., Jones, M.A. 2004. Onset and progression of the Hollow Seed (seed rot) malady of South Carolina. in: 20022004 CD-ROM version of Proceedings Beltwide Cotton Conference. San Antonio, TX, USA. Medrano, E., Bell, A. 2007. Role of Pantoea agglomerans in opportunistic bacterial seed and boll rot of cotton (Gossypium hirsutum) grown in the field. Journal of Applied Microbiology, 102(1), 134-143. Medrano, E., Esquivel, J., Bell, A. 2007. Transmission of cotton seed and boll rotting bacteria by the southern green stink bug (Nezara viridula L.). Journal of Applied Microbiology, 103(2), 436-444. (accessed: November

Medrano, E.G., Esquivel, J., Bell, A., Greene, J., Roberts, P., Bacheler, J., Marois, J., Wright, D., Nichols, R., Lopez, J. 2009a. Potential for Nezara viridula (Hemiptera: Pentatomidae) to transmit bacterial and fungal pathogens into cotton bolls. Current Microbiology, 59(4), 405-412. Medrano, E.G., Esquivel, J.F., Nichols, R.L., Bell, A.A. 2009b. Temporal analysis of cotton boll symptoms resulting from southern green stink bug feeding and transmission of a bacterial pathogen. Journal of Economic Entomology, 102(1), 36-42. Mikkelsen, T.N., Beier, C., Jonasson, S., Holmstrup, M., Schmidt, I.K., Ambus, P., Pilegaard, K., Michelsen, A., Albert, K., Andresen, L.C. 2008. Experimental design of multifactor climate change experiments with elevated CO2, warming and drought: the CLIMAITE project. Functional Ecology, 22(1), 185-195. Qamar, N., Khan, M.A., Rashid, A. 2003. Relationship of environmental conditions with potato virus Y (PVY) disease development on six varieties/advanced lines of potato. International journal of Agriculture and Biology, 5, 172-174. Ren, Y., Liu, Y., Ding, S., Li, G., Zhang, H. 2008. First Report of Boll Rot of Cotton Caused by Pantoea agglomerans in China. Plant Disease, 92(9), 1364-1364. Schlenker, W., Lobell, D.B. 2010. Robust negative impacts of climate change on African agriculture. Environmental Research Letters, 5(1), 014010. Shaner, G., Finney, R.E. 1980. New sources of slow leaf rusting resistance in wheat. Phytopathology, 70(12), 1183-1186. Strange, R.N. 2006. Introduction to plant pathology. John Wiley & Sons. SUPARCO. 2012. Crop Situation. Pakistan Satellite Based Crop Monitoring System Bulletin, 2(1).