CAFTA (ALC)
Introduction
coconut cultivation (Cocos nucifera L) is of economic, social and ecological to the Dominican Republic, which produces the currency, jobs and income, food and other products for local and international market. Besides coconut palms grace the beaches of the main tourist areas, providing its tropical beauty to the landscape, and therefore it which encourages tourists visit the country.
In Dominican Republic, the cultivated area is 22.491 ha. 359, 856 TAS. With an average annual production of 2,227, 872.8, units in the east and west coastal areas (Ministry of Agriculture, 2006). The production of fresh coconuts and products are intended for national and international markets, the latter generating U.S. $ 4, 553,143.00 Dollars (CEI-RD, 2005) by way of exports.
In 2006 the Dominican Republic was visited by 2, 612.256 tourists from different parts of the world, generating tourism input $ 3.750 and more than 300 thousand jobs (Martínez, 2006).
The coconut lethal yellowing disease (ALC) is considered the most devastating affects this crop, which has killed hundreds of thousands of palms in Key West, Florida, USA, from the Caribbean 1936.En has caused the disappearance of this crop, destroying millions of coconut palms in the last 40 years (OIRSA, 2002). CLA is caused by a phytoplasma, transmitted by the insect vector crudus Myndus Van Duzee, of the order Homoptera: Family Cixiidae. (Howard, 1983).
The ALC has been reported in the Dominican Republic several times since 1962, affecting domestic plantations in locations the province of Puerto Plata, Santiago and Dajabón (Hichez, 1971). Recently it was reported an outbreak in the town of Andrés, Boca Chica, Santo Domingo Province. (Martinez et al, 2007).
The detection of this disease is carried by the technique of reaction polymerase chain (PCR), which is a molecular technique that amplifies nucleic acid bands.
Because these pathogens can not grow invitro, recommended diagnostic technique using Chain Reaction (PCR), which is a molecular technique that amplifies nucleic acid bands. The PCR is recommended for detection of phytoplasmas and genetic characterization of these. Phytoplasma known races allow management to devise strategies to prevent the spread of disease and damage in commercial plantations.
I.
PROBLEM 1.1. Description and formulation of the problem.
The coconut lethal yellowing disease (ALC) has been reported in the Dominican Republic since 1962, affecting domestic plantations in the villages of the provinces of Puerto Plata, Santiago and Dajabón. In each occurrence of the disease have been implemented measures such as slashing and burning of diseased plants to prevent the rapid dispersion the same (Hichez, 1971).
The most recent occurrence of CLA was reported in 2006 in the east coast in the town of Andrés, Boca Chica, with a larger number of symptomatic plants, which reported outbreaks in previous years in the North, (Martinez, personal communication 2006).
This disease represents a potential threat to the production of coconuts in the Dominican Republic, as this has been reported as the most devastating crop of coconut palms; Initial symptoms are characterized by the premature fall of the fruits of all sizes, the lower leaves, intermediate and higher turn yellow, the heads and knives become necrotic, in the final stage all the leaves fall, leaving only the stem erect (Córdova, 1995). 1.2 OBJECTIVES
objective 1.2.1:
Diagnosis and characterization of strains of phytoplasmas coconut lethal yellowing.
1.2.2 Specific objectives
a) determine the presence of phytoplasma.
b) Genetic differentiation of phytoplasma strains in the country.
c) Establish the relationship of the phytoplasmas found in the Dominican Republic with other países.México, Honduras, Belize, Guatemala, USA (Florida), Jamaica, Cuba, Cayman and Haiti.
1.3 - Justification
For the Dominican Republic coconut cultivation is of great importance because it is used in agriculture and tourism, generating foreign exchange for the country as part of the export of the product and concept as tourism, attracted by the coconut.
The production of fresh coconuts, grated, dried, and products for the cosmetics industry, food and medicine is intended for local and international markets, both the United States, European Union, Canada, West Indies Children, Virgin Islands, Brazil, Puerto Rico, Haiti and others. Foreign exchange earnings in 2005, this concept was worth $ 2, 282, 987.80, USD. In 2004 and $ 4, 553,143.00, USD (CEI-RD, 2005). It is important to note that Nationwide thousands of families depend on this crop for their livelihood.
coconut palms beaches located in major tourist areas, provide the tropical beauty of the landscape, which encourages tourists visit. In 2006 the Dominican Republic was visited by 2,612,256 tourists from different parts of the world, generating tourism $ 3.750 input. More than 300 thousand. Jobs (Martínez, 2006).
business opportunities for The Dominican Republic is developing exports, as the nations of North America, Europe, South America and the Caribbean require greater amount of fresh coconut, processed and a wide variety of products. Today has resumed the use of coconut oil for culinary purposes, such as virgin coconut oil. In the United States and Europe, ½ liter sells for $ 16.00 and U.S. $ 15.75, respectively. This represents an opportunity to diversify production and increase foreign exchange earnings from this crop (CEI-RD 2006). 1:4
Diagnosis and Characterization
The diagnosis and understanding of the genetic diversity of the pathogen in the country help design management strategies to prevent the spread of disease and damage to areas of commercial production. So the knowledge of the races will help design management strategies to prevent the spread and damage to commercial plantations.
This research would provide the basis for the continuity of production and the benefits it provides to more than 1500 coconut farmers, processors, exporters and other members of the Dominican Republic in the production chain as well as foreign exchange earnings and employment sources both sectors and the preservation of the natural landscape that give the coconut palms to the country's beaches. Creating the basis for both sectors are more competitive and sustainable, especially in times of opening new markets.
1.4 Scope and limitations
1.4.1 Scope The study will include the towns of the North (Puerto Plata, Santiago, Dajabón) and Southeast (Andrés, Boca Chica) where the disease has been reported and in the production regions Northeast Commercial (Maria Trinidad Sanchez, Samana) and Eastern (Hato Mayor, El Seibo, La Altagracia, La Romana) in the country. Which has not been reported. and to ensure that these areas are free of the disease. 1.4.2 Limitations
To determine the sequence of base pairs of possible phytoplasma in the country, will be sent to international centers engaged in such activity, as in the Dominican Republic there are no possibilities for these techniques.
11. THEORETICAL FRAMEWORK 2.1 General
Cocos nucifera L.
(Belonging to the family Arecaceae), commonly known as coconut palm. It is probably native to the Pacific islands, and today is cultivated almost throughout the tropics. It
the most cultivated and palm in the world, and currently is the main species producing vegetable fat. It is a plant that provides a wider range of products in the world, being a primary source of food, drink and shelter. The distribution of the coconut palm extends over most of the islands and the tropical coasts and in some places outside the tropics.
The main export is raw copra desiccated coconut followed.
diversity and potential of coconut contributes significantly to the economic sector in producing countries. 2.2 Background
coconut lethal yellowing (ALC) is considered as one of the most important diseases that affect the production of coconut, coconut trees has destroyed up to 100% in some countries (Harries 1978). It is caused by a phytoplasma, This devastating disease is found in Mexico, Honduras, Belize, Guatemala, USA (Florida), Jamaica, Cuba, Grand Cayman, Haiti and the Dominican Republic (Beakbane et al 1972).
Many of these countries have had disastrous experiences as in the case of Jamaica, where 100% of their plantations were destroyed in a period of four years, estimated losses in five million palms in a period of 20 years (Romney ( 1983). In Honduras has destroyed 90% of the coconut (Plavsic-Banjac et al., 1972).
is an organism that has characteristics similar to bacteria, have cytoplasm, ribosomes, DNA and is restricted to phloem cells. It is transmitted by insects is one of the most important Myndus crudus, Van Duzz (Homoptera: Cixidae) (Howard et al, 1983, Howard, 1995).
pathogen dispersal apparently occurs through two mechanisms, (McCoy et al 1983). One is through a central localized infection, ie, the disease appears in one or both palms and then randomly extends adjacent palms. The second mechanism of dispersion of the FTA is at large distances as discontinuous, followed by localized dispersal. The distance covered by these jumps can be tens of kilometers and are favored by high winds such as hurricanes. Therefore, the pathogen can spread very quickly as has happened in Mexico and Honduras, (Cordova, 1995).
M. crudus feed on diseased plants and then moved to healthy plants, thus spreading the disease. It is believed that the ornamental grass used to beautify the surroundings of the tourist areas is used for development (Piña, 1995). Have been reported in different strains of phytoplasma causing lethal yellowing of coconut in the Caribbean and Africa (Harrison et al.1994).
Symptoms of LAC in the Caribbean and Africa show similar, but differences were observed in the epidemiology and susceptibility in the materials, so it is assumed that different phytoplasmas cause disease. The coconut trees and palms likely die quickly, in a period between 4 and 6 months after the onset of symptoms (Eden-Green, 1995)
. ALC
symptoms observed in Puerto Plata, have been confirmed by the Scientific Research Centre of Yucatan (CICY) of Mexico, in collaboration with the University of Florida, United States, in samples submitted in 1997. These samples were analyzed by the technique of Chain Reaction Polymerase-PCR, they were positive to LAC. This technique has been. Used to detect the phytoplasma in the embryo of the fruit. (Cordova, et al, 2003).
This disease was reported for more than 40 years until 2006, being detected and confirmed a new outbreak in the south coast in the town of Boca Chica, (Martinez et al, 2007)
111. MATERIALS AND METHODS. 3.1 Location
The study will cover the North (Puerto Plata, Dajabón) and South (Andres Boca Chica) where the disease has been reported commercial plantations in the Northeast (Maria Trinidad Sanchez, Samana) and Eastern (Hato Mayor, El Seibo, La Altagracia, La Romana) of the Dominican Republic, where no disease has been reported. In order to confirm that the area remains free of ALC.
3.2 Type of research.
is a descriptive research to detect and characterize the genetic variability of phytoplasma LAC coconut palms in areas affected by CLA.
3.3. Selection of plants sampled.
In every place where the disease has been reported to be selected 5 plants showing symptoms typical of CLA and 5 asymptomatic plants. In each plant, take samples of leaves, stems and inflorescences to determine the presence or phytoplasma. In areas free of the disease, samples were taken from 10 plants.
receiver will use Global Positioning System (GPS) to mark the areas of advocacy, the points located were used to construct a map of disease distribution
3.4.
Sampling Samples were taken from the young leaves and inflorescence stem. For the sample was chosen with symptoms characteristic of the early stage of FTA. For the inflorescence, cut is closed before the open inflorescence stems and samples were taken by a hole 10 to 12 cm. deep, at a height of 1.5 meters de1.0 soil using an electric drill.
The collected samples will be placed on damp paper and deposited in a plastic bag previously identified. They are then placed in a cooler to be transported to the laboratory.
3.3 Analysis Laboratory
The samples will be taken to the laboratory of Virology of the Centre for Agricultural Technology (CENTA) of the Dominican Institute of Agriculture and Forestry Research (IDIAF). These are analyzed through Chain Reaction (PCR). The research
Three types of analysis:
a) nested PCR for detection of phytoplasmas.
b) Restriction fragments length polymerase (RFLPs) for the differentiation of genetic diversity.
c) Sequencing of the selected samples.
The DNA samples were amplified using universal primers P1-P7 for the presence of phytoplasma in the samples, then perform a nested PCR with specific primers and LY50F LysR. 3.4
phytoplasma detection
The PCR product will be fragmented to electrophoresis in agarose gel 5% to analyze the patterns of bands obtained. For detection of phytoplasma. The PCR product obtained will be sequenced to relate the similarity with other phytoplasmas in the regions area. Molecular characterization of races, we will make comparisons with different isolates of the pathogen to determine if there are different strains of phytoplasma in the Dominican Republic.
3.6 - Schedule of activities
Activity
EFMAMJJASONDEFMAM
Buy reactive
Sample Collection xxx xxx
Laboratory analysis results Analysis Presentation thesis
XX
Publication
3.5-budget for this research. ARTICLE
ITEM SPECIFICATIONS UNIT UNIT PRICE AMOUNT VALUE Value U.S. $ RD $
1 0.5 ul eppendorf tubes cover / 500 1 2000 2000 60.60 1.5
2 ul eppendorf tubes cover / 500 1 2500 2500 3 75.75
Punta micropipettes 1 -10 ul Cover / 1000 2700 1400 4 42.42
Case Punta ul micropipettes 200-1000 / 1000 2 1200 2400 5 72.72
Disposable Gloves Medium / Long Box / 100 3195585 17.72 6
Vials/50 started 24 bp 2 ul 2 7 7821.02 3910.51 237.00
Taq polymerase 250 units / ul 1 2 vials
280.00 4620 9240 8 10 mM dNTPs dATP, dCTP, dGTP and dTTP Kit 0.5 mL 1 160.00 5280 5280 9 PCR mix
September 1 4780 4780.00 144.80 liters liquid nitrogen
10 4,170,680 20.60
11 Grade Agarose PCR. Intermediate solution. Molecular weight resealable plastic bag < 1000 bp. Sobre /100 gr 1 7755 7755 235.00
12 6x10/10x 14 "boxes / 100 3,300,900 27.27
Binoculars 13 3500 3500 1 106.00
14 48841.02 1480.03 Sub-Total Operating Expenses
15 16 Fuel 12 viajes/35 gl / Km Travel 12 1770 2124 0 643.00
17 Per diem 2 technicians / Driver 36 300 1080 0 327.27
18 Salary / Labor Day 1 inflorescence sampling
8200 48.48 1600 19 6400.00 Thesis Preparation and printing 193.94
19 Sub-Total Expenditure 40040.00 1019.39
Op 20 Sub-Total Op Expenses / Reagents
88881.02 2499.42 8888.102 21 Contingencies 10%
249.90 Grand Total 97769.12 2749.36 22
Note: This is
The estimated budget for this inquiry, but have yet
the Agricultural Technology Centre IDIAF, laboratory equipment muestras.en analyzing the yucatan mexico is where conditions until the time for fulfillment.
Bibliography:
1 - Almeyda, Isidro, molecular 1998.Diagnóstico Coconut lethal yellowing,
Regional Research Center Southeast, Yucatan, Mexico.
2 - Beakbane, AB, Slater, CHW and Posnett, AF1972. Mycoplasmas in the phloem
of coconut, Cocos nucifera L, with lethal yellowing disease.
Journal of Horticulture Science.
3 - Export and Investment Center of the Dominican Republic (CEI-RD), 2005
4 - Cordova, I.1995.Estudiola intraplant distribution and dispersal of lethal yellowing in cocoteeros
using the chain Reccion polymerase.
5 - Eden-Green, SJ 1995. Brief history of yellowing research. In: C. Oropeza.,
WF Howard and GR. Absburner lethal yellowing research and practical aspects. pp. 17-33.
6- Harrison, N.A., Richardson, P.A., Kramer, J.B. and Tsai, J.H. 1994. Detection of
the Mycoplasmas-like organism associated with lethal yellowing disease of palm
in Florida by polymerase chain reaction Plant Pathology.
7- Harries, H.C.1978. Lethal yellowing disease of coconut in global perspective.
Philippines journal of coconut science. Florida, Estados Unidos
8- Howard, F.W., Norris, R.C. and Thomas, D.L. (1983). Evidence of transmission of palm lethal yellowing agents by a planthopper, Myndus crudus (Homptera: Cixiidae). Tropical Agriculture, Trinidad.
9- Howard, F.W. (1995). lethal vector yellowing study. Methods of experimental
transmission. In: C. Oropeza., WF Howard., And GR. Absburmer: lethal yellowing research
and Practical aspects.
10 - Hichez, E.1971. Lethal yellow in the Dominican Republic. Bulletin No.3
Plant Health Department, Secretary of State for Agriculture.
11 - Martinez, Desiré, 2006.Informe National Association of Hotels and
Restaurantes.Listin Journal, 19 marzo2006.
12-Ministry of Agriculture, 2006. Diagnosis of the Agricultural Sector, Assistant Secretary for Planning, Department of Agricultural Economics, Santo Domingo, Dominican Republic.
11 - Plavsic-Benjac, B., Hunt, P. and Maramorosch, K. (1972). Mycoplasma-like Bodies
associated with lethal yellowing disease of coconut palm. Phytopathology, 62: 298-2 - 12 - Romney, DH (1983). Brief review of coconut lethal yellowing. Indian coconut journal.
(This is a research work done by the Ing.Andrea
Felix)
. NOTICE
0 comments:
Post a Comment