(Mazhar)isolation & control of banana bunchy top virus


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(Mazhar)isolation & control of banana bunchy top virus

  1. 2. Detection & Control Of Banana Bunchy Top Virus Mazhar Ullah Class # 08
  2. 3. Introduction <ul><li>Banana is the common name for herbaceous plants of the genus Musa and for the fruit they produce. </li></ul><ul><li>Bananas come in a variety of sizes and colors when ripe, including yellow, purple, and red. </li></ul>
  3. 4. Continued… <ul><li>Bananas are a staple starch for many tropical populations. </li></ul><ul><li>Depending upon cultivar and ripeness, the flesh can vary in taste from starchy to sweet, and texture from firm to mushy. </li></ul><ul><li>Both skin and inner part can be eaten raw or cooked. </li></ul><ul><li>Bananas' flavor is due, amongst other chemicals, to isoamyl acetate which is one of the main constituents of banana oil. </li></ul>
  4. 5. Continued… <ul><li>Banana bunchy top virus (BBTV) is one of the most serious diseases of banana. </li></ul><ul><li>Once established, it is extremely difficult to eradicate or manage. </li></ul><ul><li>BBTV is widespread in Southeast Asia, the Philippines, Taiwan, most of the South Pacific islands, and parts of Pakistan, India and Africa. (Saleem et al., 2005) </li></ul>
  5. 6. Continued… <ul><li>Banana bunchy top virus is spread by the banana aphid, which acquires the virus after at least four (but usually about 18) hours of feeding on an infected plant (Naseem et al., 2010). </li></ul><ul><li>The aphid can retain the virus through its adult life, for a period of 15–20 days. </li></ul><ul><li>During this time, the aphid can transmit the virus to a healthy banana plant by feeding on it, possibly for as little as 15 minutes but more typically for about two hours. </li></ul><ul><li>Disease symptoms usually appear about a month after infection . </li></ul>
  6. 7. Continued… <ul><li>The initial symptoms of banana bunchy top virus consist of dark green streaks in the veins of lower portions of the leaf midrib and the leaf stem. </li></ul><ul><li>Also, dark green, hook-like extensions of theleaf lamina veins can be seen in the narrow, light-green zone between the midrib and the lamina. </li></ul><ul><li>The short hooks point down along the midrib toward the petiole. They can best be seen by back-lighting the leaf against the sky. </li></ul>
  7. 8. Annual Losses In Pakistan <ul><li>In Pakistan Sindh is the main producer of banana </li></ul><ul><li>Annual loses of banana in Pakistan Because of BBTV: </li></ul><ul><li>Sindh 19 Million Rupees </li></ul><ul><li>KPK 8 Million Rupees </li></ul><ul><li>Punjab 13 Million Rupees </li></ul><ul><li>Balochistan 5 Million Rupees </li></ul>
  8. 9. Review Of Literature <ul><li>Mazhar & Waheed (1988) isolated & characterized BBTV from different banana species using ELISA </li></ul><ul><li>Humayoun et al (1999) confirmed that BBTV is an RNA virus. </li></ul><ul><li>Dmtzgen & Franeki (1988) reported that BBTV can be isolated & detected through ELISA. </li></ul>
  9. 10. Objectives <ul><li>To detect BBT virus. </li></ul><ul><li>To control BBT virus. </li></ul><ul><li>To characterize BBT virus. </li></ul>
  10. 11. Methodology <ul><li>The present research study will be conducted at Institute Of Biotechnology And Genetic Engineering (IBGE) Khyber Pakhtunkhwa Agricultural University Peshawar during 2011-12. </li></ul><ul><li>ELISA kit will be purchased from KPL companies Lahore. </li></ul>
  11. 12. Continued… <ul><li>This research will be carried out on the virus isolated banana X 2 specie. </li></ul><ul><li>Samples will be collected from different areas of the country. </li></ul>
  12. 13. Sample Preparation <ul><li>Leaf samples of about 1g will be taken from the infected banana tree. </li></ul><ul><li>The sample will be crushed in 5ml of extraction buffer using fine abrasive. </li></ul><ul><li>Then this viral extract will be filtered with nylon cloth. </li></ul>
  13. 14. Detection Of Viral Presence <ul><li>The presence of viral particles will be detected through ELISA test. </li></ul><ul><li>The ELISA plates will be coated with BBTV specific antibodies & will be incubated at 34C for 2-3 hours. </li></ul><ul><li>Then the plates will be washed. </li></ul><ul><li>Then the viral sample (antigen) will be introduced to the wells of ELISA plates. </li></ul>
  14. 15. Continued… <ul><li>The antigen added ELISA plates will be incubated at 4C overnight. </li></ul><ul><li>Washing will be done again. </li></ul><ul><li>Then the ELISA plates will be coated with specific antibodies linked with enzymes that will bind to the antigen. </li></ul><ul><li>Then Phenol phospho nitrate (PNP) will be added to break the antibody-antigen assembly. </li></ul>
  15. 16. Continued… <ul><li>It will give yellow colour which indicates the presence of virus. </li></ul><ul><li>The intensity of the yellow colour shows the intensity of viral infection. </li></ul>
  16. 17. Control Of BBTV By Pathogen Derived Resistance (PDR) <ul><li>DNA will be extracted from the isolated viral particles . </li></ul><ul><li>Core Protein (CP) gene will be isolated from the viral genome using proper restriction enzymes. </li></ul><ul><li>The CP gene will be amplified through PCR. </li></ul><ul><li>The gene will be loaded on a binary vector. </li></ul>
  17. 18. Continued… <ul><li>Then this gene will be introduced to banana by A.tumefaciens. </li></ul><ul><li>The gene will be incorporated to banana’s genome by A.tumefaciens through lateral DNA transfer. </li></ul><ul><li>This gene will confer banana plant resistance against BBTV. </li></ul>
  18. 19. Literature Cited <ul><li>Naseem, Z., M. Ihsan & M. Rameez. (2010). The study of BBTV symptoms in different species of banana. Journal of Virology 10(15):313-320. </li></ul><ul><li>Dmtzgen, R. G. & Franeki, R. I. B. (1988). Analysis of lettuce necrotic yellows virus structural proteins with monoclonal antibodies and concanavalin A. Virology 166, 486-494. </li></ul><ul><li>Saleem, A., Q.A.Arbab & H.U.Jan. (2005). Prevalence of BBTV in different areas of Nigeria. African journal of virology 9(6):107-110. </li></ul><ul><li>Humayoun, F., F.Ali & W. A. Khan. (2006).Isolation & genotyping of BBTV using ELISA. International Journal Of Food Microbiology 87(03):109-112. </li></ul>
  19. 20. Budget: <ul><li>Fixed Budget: </li></ul>200000 Grand total 1,00000 1,00000 PCR - - 3. 1,00000 - - 50,000 50,000 ELISA Kit Incubator 1. 2. Total Cost Year 2 Cost Year 1 S.No
  20. 21. Recurrent budget 9,10,000 - - - - Total 400,000 200,000 Miscellaneous 200,000 Miscellaneous 8.. 200,000 100,000 seminars 100,000 seminars 7. 100,000 50,000 Travelling 50,000 Travelling 6. 60,000 30,000 Honraria 30,000 Honraria 5. 1,00000 50,000 Researcher,s salary 50,000 Researcher,s salary 4. 20,000 10,000 Stationary 10,000 Stationary 3. 10,000 5,000 Glasswares 5,000 Glasswares 2. 20,000 10,000 Chemicals 10,000 Chemicals 1. Total amount Year 2 Amount Year 1 S.No
  21. 22. Total Budget <ul><li>Total Budget = Fixed Budget + Recurrent Budget </li></ul><ul><li>= 200,000 +9,10,000 </li></ul><ul><li>=11,10,000 </li></ul>
  22. 23. Thanks!!!