Distribution and diversity of geminiviruses in trinidad and tobago

ABSTRACT Seven crop and eight weed species from 12 agricultural locations in Trinidad and Tobago were assayed for the presence of whitefly-transmitted geminiviruses (WTGs) by using dot blot hybridization and polymerase chain reaction (PCR) amplification of the N-terminal coat protein sequence with degenerate primers. The amplified fragments were cloned and analyzed by restriction enzyme digestion to determine fragment length polymorphism among the cloned fragments. Representative clones were then sequenced and subjected to phylogenetic analysis to determine the sequence similarity to known WTGs. WTGs were found in every location sampled and in 10 of the 15 species investigated: Lycopersicon esculentum(tomato), Capsicum annuum (pepper), Capsicum frutescens (sweet pepper), Abelmoschus esculentus (okra), Phaseolus vulgaris (beans), Alternanthera tenella, Desmodium frutescens, Euphorbia heterophylla, Malva alceifolia, and Sida acuta. The geminiviruses infecting these plants were closely related to potato yellow mosaic virus from Venezuela (PYMV-VE) and tomato leaf curl virus from Panama (ToLCV-PA). However, in pepper, sweet pepper, okra, Alternanthera tenella, Euphorbia heterophylla, Des-modium frutescens, and in one sample of tomato, a PYMV-VE-related virus was found in mixed infections with a virus related to pepper huasteco virus. Full-length infectious DNA-A and DNA-B of a tomato-infecting geminivirus from Trinidad and Tobago were cloned and sequenced. DNA-A appears to be a recombinant derived from PYMV-VE or ToLCV-PA, and Sida golden mosaic from Honduras. The implications of these findings in the control of WTGs are discussed.     

Cacao resistance to Phytophthora: Effect of pathogen species, inoculation depths and pod maturity

Two species of Phytophthora (P. palmivora and P. capsici) and inoculations at two depths (3 mm and 9 mm) were tested each on 10 clones of Theobroma cacao to determine their effects on pod resistance. Ripe and unripe pods were also assessed to determine the influence of physiological status of the pod on the expression of resistance. The two pathogens tested (P. palmivora and P. capsici) differed significantly in their reactions on pods, with P. palmivora being more aggressive than P. capsici. However, the lack of interaction between clones and pathogen species and the similarity in the ranking of clones based on lesion size suggested that selection for resistant clones can be based on one of the two pathogens, preferably the more aggressive one. Pod reactions differed between inoculation depths (3 mm and 9 mm), and between pod maturity stages (ripe and unripe pods) with relatively larger lesions being recorded at 9 mm depth and on unripe pods as compared to those observed at 3 mm depth and on unripe pods, respectively. The magnitude of increase in lesion sizes, however, varied with genotypes, indicating that inoculation depth and pod maturity stage should be standardized in screening cacao germplasm for resistance to Phytophthora.     

Quantitative resistance to Cercospora leaf spot disease caused by Pseudocercospora cruenta in cowpea

Six populations (Parent 1, Parent 2, F1, F2, BC1 and BC2) generated from each of four crosses involving four resistant and two susceptible varieties of cowpea (Vigna unguiculata L. Walp) were evaluated for resistance to Cercospora leaf spot (CLS) disease caused by Pseudocercospora cruenta under induced epiphytotic conditions, in four separate field experiments. Climatic conditions determined the onset of CLS disease in the susceptible cultivar and varied in the four experiments from 35 to 48 days after planting (DAP). Genetic analysis revealed that the mode of inheritance of resistance to P. cruenta can be oligogenic or polygenic depending upon the cross. This is the first report of polygenic inheritance of CLS resistance. Number of nodes infected fitted a simple additive dominance model with predominance of additive effects based on generation mean analysis. Oligogenic resistance was observed for the other three crosses, with the most plausible models being: a single gene model with incomplete dominance in CB27 × IT87D-939-1; a single gene model with complete dominance in CB27 × VRB-10; and a triger model in Los Banos Bush Sitao × IT86D-792, based on segregation analysis of symptomatic : non-symptomatic plants. The role of minor genes was also indicated in the above crosses. Suggested approaches to breeding for resistance to CLS are discussed.     

Detection of begomoviruses in clarified plant extracts: a comparison of standard, direct-binding, and immunocapture polymerase chain reaction techniques

ABSTRACT Three polymerase chain reaction (PCR) techniques-standard PCR (Std-PCR), direct-binding PCR (DB-PCR), and immunocapture PCR (IC-PCR)-using degenerate primers were optimized and evaluated for the detection of begomoviruses. Tomato leaf samples were ground in three different extraction buffers and subjected to Std-PCR. The effect of the buffers on the detection limits of amplification of the virus (detection of the initial and end points of dilution) was determined. With the optimal extraction buffer determined by the first experiment, the antibody concentration and incubation conditions for IC-PCR were evaluated to determine the requirements for maximum capture of antigens during the capture phase of the technique. The incubation conditions of DB-PCR were also investigated to determine the most favorable conditions for adsorption of the viral template. The reproducibility of all assays was evaluated. With the results of the optimization experiments, the applicability of the three techniques to different plant species was assessed. Extracts of plant species belonging to three families were subjected to the optimized Std-, DB-, and IC-PCR protocols. Std- and IC-PCR both achieved reproducible detection of begomoviruses, but the detection limits and amplified band intensity for all plant species tested were superior for the latter. DB-PCR was an unreliable method of detection, because of poor reproducibility and low intensity of amplified bands. These results indicate that the optimized IC-PCR detection system using degenerate primers is the most effective for the detection of begomoviruses in clarified plant extracts.     

Identification of resistance to Cercospora leaf spot of cowpea

Twelve selected cowpea cultivars were screened for resistance to Cercospora leaf spot (CLS) disease caused by Pseudocercospora cruenta and Cercospora apii s. lat. under artificial epiphytotic conditions in a replicated field trial, with the objective of developing a quantitative measure of disease resistance. CLS incidence, leaf spotting score, lesion density, lesion size, proportion of nodes infected, diseased leaf area, conidia number mg⁻¹ and fascicle density were assessed in 12 cowpea genotypes at crop maturity. Proportion of nodes infected and leaf spotting score were best able to quantitatively differentiate between the levels of resistance, and allow the exploitation of quantitative resistance to the disease. Both lesion density and lesion size were important in determining the final leaf spotting score but the former was epidemiologically more important than the latter, indicated by its correlation to most of the CLS symptom measures. There was differential resistance to the P. cruenta and C. apii s. lat. among the cowpea varieties screened. Among the cowpea lines screened, resistance to P. cruenta was more common than resistance to C. apii s. lat. Nevertheless, P. cruenta was considered the more aggressive and epidemiologically more important than C. apii s. lat. on the varieties tested evidenced by the strong correlation of P. cruenta incidence with acropetal spread of CLS, intensity of leaf spotting, conidia number mg⁻¹ and fascicle density. The highly susceptible varieties namely VRB7, Los Banos Bush Sitao no.1 and CB27 were susceptible to both Cercospora pathogens. The cowpea variety VRB-10 was completely resistant to both pathogens and is a useful source of resistance in CLS breeding programmes.     

Relationships Between Black Pod and Witches'-Broom Diseases in Theobroma cacao

ABSTRACT Field observations were conducted from 1998 to 2001 at the International Cocoa Genebank, Trinidad, to evaluate 57 cacao clones for resistance to black pod (BP) and witches'-broom (WB) diseases (caused by Phytophthora sp. and Crinipellis perniciosa, respectively). Each month ripe pods were harvested and the number of healthy and diseased was recorded. The number of brooms on vegetative shoots was recorded three times a year on selected branches. Twenty-three clones showed less than 10% of infection for both BP and WB on pods. Among those, eight clones showed an absence of brooms on the observed branches: IMC 6, MAN 15/60 [BRA], PA 67 [PER], PA 195 [PER], PA 218 [PER], PA 296 [PER], PA 303 [PER], and POUND 32/A [POU]. Broad-sense heritability was estimated at 0.38 and 0.57 for WB disease on pods and shoots, respectively, and at 0.51 for BP disease. Genetic correlation between WB disease on pods and on shoots was low and estimated at 0.39, whereas the correlation between WB and BP diseases on pods was 0.48. To choose putative parents for breeding schemes, it is suggested that clones are first assessed for their level of resistance to WB on shoots, and the most promising individuals are screened for BP with a detached pods test. Further studies are needed to confirm whether the level of resistance to WB on pods can be predicted using an early test on seedlings.     

A study of genotype-isolate interaction in cacao (Theobroma cacao L.): resistance of cacao genotypes to isolates of Phytophthora palmivora

The resistance of cacao (Theobroma cacao. L)leaves to 10 isolates of Phytophthora palmivora(Butler) Butler was investigated in 18 genotypes of cacao in three experiments. Tissue-paper-mount and punch-inoculation methods were used to distinguish between resistance at the penetration and post-penetration levels based on lesion frequency and size, respectively. There was a 5–6 fold difference in aggressiveness among isolates collected from different locations in the islands of Trinidad and Tobago for both types of resistance. However, host genotype ×isolate interaction effects were not significant for resistance at both the penetration and post-penetration levels. A strong positive relationship (r = 0.8) was observed in the abilities of isolates to effectively breach the two types of resistance. This suggests that the same isolate can be used in screening for both types of resistance. No significant relationship was found between resistance at the penetration and post-penetration levels suggesting that the resistance mechanisms were different. Subsequent experiments using a wider range of cacao genotypes from the International Germplasm Collection, Trinidad, confirmed the initial results. These experiments indicate that resistance found using one isolate would be equally valid using other isolates of Phytophthora palmivora. The implications of the findings are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic analysis of pod quality characteristics in vegetable cowpea (Vigna unguiculata L. Walp.)

A preliminary study of consumer preferences for pod characteristics in vegetable cowpea showed a general preference for greener, longer, fleshier pods that are less seedy. Methods of quantitatively assessing these traits were developed and used in understanding the inheritance of pod quality traits. Combining ability analysis, involving a 6 × 6 diallel mating design, showed that GCA effects were predominant for all pod characteristics. The additive effects were especially high for pod seediness. The dominance effect was, nevertheless, significant for all the characters. The dominance effects were unidirectional in the case of pod wall thickness and were in the direction of the thin walled parent. Dominance was ambidirectional for the other characteristics investigated. The study showed that it is relatively easy to improve these characters by carefully selecting the parents in hybridisation programmes.     

Assessing the Genetic Diversity in the International Cocoa Genebank,Trinidad (ICG,T) using Isozyme Electrophoresis and RAPD

Random amplified polymorphic DNA (RAPD) and isozyme electrophoresis (IE) techniques were used to estimate the level of genetic diversity in a sample of cacao germplasm existing at the International Cocoa Genebank, Trinidad. Twenty-six cocoa populations represented by 459 cocoa genotypes were analysed using IE and 22 populations represented by 353 cocoa genotypes were analysed using RAPD. Despite few differences in the classification of the populations, both techniques revealed three major groups: the indigenous trees, the cultivated Trinitario and the cultivated trees from Ecuador. Two-thirds of the partitioned diversity were found within populations and one-third between the populations, with both techniques.     

Inheritance and combining ability studies of pod physical and biochemical quality traits in vegetable pigeonpea (<Emphasis Type="Italic">Cajanus cajan</Emphasis> L. Millsp)

Pigeonpea (Cajanus cajan L. Millsp) is a popular vegetable crop in the Caribbean, grown for its immature green peas. Although breeding programmes, in the past, have made considerable strides in improving yield in a relatively photoperiod insensitive genetic background, the quality of peas continue to affect consumer acceptance. The mode of inheritance of a number of quality traits (physical and biochemical) known to affect consumer acceptance of pigeon peas was investigated in this study using a 6 × 6 half-diallel mating design and a M × N (5 × 6) mating design analysed using Griffing’s Method II Model I approach and the North Carolina design-II approach, respectively. The results showed that pod length, pod width, seeds per pod, shelling percentage and phenolic content were under the control of additive genetic effects with the non-additive effects generally being either not significant or much smaller compared to the additive genetic effects. Hundred seed weight was controlled by both additive and non-additive effects, while pod biochemical characteristics viz. sugar content, starch content and protein content, were governed by a preponderance of non-additive genetic effects. The strong genetic correlation between pod length, pod width and seed number indicates that these characteristics could be simultaneously improved in breeding programmes. The best general and specific combiners for the various characteristics were identified, and a method of improvement of quality in pigeonpea is discussed.