African Crop Science Conference Proceedings, Vol. 7. pp. 1279-1283Printed in Uganda. All rights reservedISSN 1023-070X/2005 $ 4.00© 2005, African Crop Science Society

Occurrence and prevalence of cowpea virus diseases in uganda

M. ORAWU, R. MELIS, W. DE MILLIANO, M. LAING & E. ADIPALA1

African Centre for Crop Improvement, University of KwaZulu Natal, Scottsville 3209, PMB, South Africa 1Regional Universities Forum for Capacity Building in Agriculture, Makerere University, P.O. Box 7062, Kampala, Uganda

Abstract Field samples were obtained from 5-6 week old cowpea plants from 32 locations in eastern Uganda in 2004. Doubleantibody sandwich enzyme-linked immunosorbet assay (DAS-ELISA) was used to test the 108 virus symptomatic leaf samples.The virus incidences ranged from 46.6 to 96.2%, and severities ranged from 13.8 to 31.8%. The common viruses identified werecowpea aphid-borne mosaic virus, cowpea mild mottle virus and cowpea severe mosaic virus. Of the samples tested, 21.3% wereinfected by a single virus and 7.4% with two viruses. Cowpea severe mosaic virus was the most common (13% incidence) followedby cowpea mild mottle virus (6.5% incidence) and the least was cowpea aphid-borne mosaic virus (1.9% incidence). Cowpea severemosaic virus occurred in mixed infections with cowpea mild mottle virus and cowpea aphid-borne mosaic virus, but mixed infectionof cowpea mild mottle virus and cowpea aphid-borne mosaic virus was rare.

Key words: Infection, occurrence, Vigna unguiculata, virus

Résumé Des échantillons étaient obtenus 5-6 semaines après plantation du niébé de 32 endroits à l’est de l’Ouganda en 2004. Uneenzyme doublement anticorps associée aux essais immunosorbet (DAS-ELISA) étaient utilisée pour tester le 108 ème échantillond’une feuille avec les symptôme d’un virus. L’incidence du virus rangée de 46.6 a 96.2%, et la sévérité rangée entre 13.8 a 31.8%. Levirus commun était la mosaïque aphide du niébé, le virus du doux du niébé et la mosaïque sévère du niébé. Parmi les échantillonstestés 21.3 % étaient infectés par un seul virus et 7.4% avec deux virus. La mosaïque du niébé était le plus commun des virus(incidence 13%) suivi par motte douce du niébé (incidence 6.5%) et la faible incidence était celle de la mosaïque aphide du niébé(1.9%). Le virus sévère de la mosaïque du niébé apparu dans une infection mixte avec la motte douce et l’aphide du virus de lamosaïque. Mais une infection mixte de la motte douce du niébé et la mosaïque aphide du niébé était rare.

Mots clés: Infection, apparition, Vigna unguiculata, virus

Introduction

Cowpea (Vigna unguiculata L. Walp) is a major source ofprotein and of considerable importance for human nutritionin the semiarid and tropical regions of Africa (Gowda et al.,2000). In West Africa, cowpea is second in importance aftergroundnuts, with Nigeria accounting for over 70% of thetotal world production (Sing et al., 2000). In Uganda,cowpea is the third most important legume food crop afterthe common beans (Phaseolus vulgaris L.) and groundnuts(Arachis hypogea L.) (Sabiti et al., 1994). As a legume crop,its tolerance to moisture stress and marginal soils makes itsuitable for cultivation by most farmers in Uganda. Theleaves and seeds are consumed as an important supplementto staple cereal diet. In addition, cowpea is a majorcomponent of the cropping system on farmers’ holdingswhere it is grown in association with cereals such as maize,sorghum and millet, although it is sometimes grown as amonocrop. In eastern Uganda, where nearly 90% of thecountry’s crop is produced (Adipala et al., 1997), cowpeaproduction is on the increase due to high demand fromboth the local and external markets, and many farmers inthe region now grow cowpea for cash markets (Sabiti et al.,1994).

However, viral diseases are among the mostagriculturally important and biologically intriguing groupsof plant pathogens, and cause serious economic losses by

reducing yields and quality of the crop (Byoung-Cheorlet al., 2005). Cowpea yields, especially among subsistencefarmers, are generally low due to several factors, but viraldiseases remain a major constraint to production on alarge scale (Thottapilly and Rossel, 1992). Yield losses ofcowpea up to 87% due to infection by viruses have beenreported in Nigeria (Shoyinka et al., 1997), and thecomplete loss of cowpea crop in northern Nigeria wasattributed to cowpea aphid-borne mosaic virus disease(Raheja and Leleji, 1974).

Worldwide, up to 20 viruses have been reported tooccur on cowpea, but only eight viruses are known toinfect cowpea in Africa (Thottappilly and Rossel, 1992).The economically important viruses in Africa includecowpea aphid-borne mosaic virus potyvirus, cowpea mildmottle virus carlavirus, cowpea mosaic virus comovirus,cowpea chlorotic mottle virus carmovirus, cowpeagolden mosaic virus germinivirus, southern bean mosaicvirus sobemovirus, cucumber mosaic virus cucumovirusand tobacco mosaic virus tobamovirus (Thottappilly andRossel, 1992; Alegbejo and Kashina, 2001).

In Uganda, studies by Edema et al. (1997) showedcowpea aphid-borne mosaic virus as the most commonvirus, yet little attention has been given to it. Informationon the occurrence of other viruses in the cowpea growingregions in Uganda is not well known. With changes inthe cropping systems and population dynamics of virus

M. ORAWU et al.1280

vectors may influence virus occurrence (Wisler et al.,1998), and hence cowpea viruses have increasinglybecome very significant in Uganda. In order to provideinformation that would be essential for determiningpriorities in breeding for resistance to viruses, attemptswere made to verify and identify the occurrence of theeconomically important viruses on cowpea in easternUganda.

Materials and methods

Field survey and sampling. Field surveys were conductedon farmers’ fields during the growing season in Octoberand November, in the eastern Uganda. In 2004, surveyswere restricted to Soroti, Kumi, Pallisa and Tororo, themajor cowpea producing districts, where 32 locations weresurveyed. Cowpea fields in each district were surveyedand sampled at approximately 5 km apart. About five to sixweeks old cowpea trifoliate leaves were sampled in thefour districts and a diagonal pattern of sampling was used.The total number of plants within a quadrant (1 x 1 m) ofeach field was counted approximately 30 plants and virus-like symptomatic plants recorded. Thus, 120 plants wererecorded in each field of four quadrants. The virusincidence was calculated as percentages of the diseasedplants over the total number of plants assessed as theaverage of the cowpea fields per district. Disease severitywas also assessed visually during the study as theproportion of the leaves with virus symptoms onpercentage score of 0, 5, 20, 25, 30, 40, 45, 50, and 60.

Serological tests. One hundred eight virus symptomaticleaf samples were collected from 32 locations in fourdistricts namely Soroti, Kumi, Pallisa and Tororo. Fullyexpanded trifoliate leaves from each quadrant weresampled in a field of four quadrants and the samples wereplaced separately in small plastic polythene bag. Thesamples were then taken to Makerere University Biotechlaboratory and frozen at -200C before being subjected toDAS-ELISA. Double antibody sandwich enzyme-linked

immunosorbet assay was used for serological testing ofplant samples and was carried out using kits provided byDSMZ in Germany. The kits included specifically toparticular virus antiserum namely cowpea aphid-bornemosaic virus (CABMV), cowpea chlorotic mosaic virus(CCMV), cowpea mild mottle virus (CMMV), cowpeamosaic virus (CMV) and cowpea severe mosaic virus(CSMV). Three leaf-discs were taken from top, middle andlower parts of the leaf preferably with virus symptoms.The kits contained positive and negative controls of dried-ground leaf samples. The positive and negative reactionswere visually assessed, the degree of yellow colorationdetermining those regarded as positive with the ELISAtest. The absorbencies were recorded at 405nm (A405) after90 minutes substrate incubation period using a microplatereader. Readings indicating twice the values of the negativecontrols were considered positive.

Results

Virus disease occurrence. Figure 1 shows the meanincidence and severity of virus symptoms that wereobserved in the surveyed districts of Soroti, Kumi, Pallisaand Tororo though there were considerable differences inoccurrence and prevalence among the districts. Thehighest disease incidence was 96.2% with severity of31.8% in Pallisa and lowest disease incidence levels of46.6% with severity of 28.8% being recorded in Kumi (Fig.1). However, Tororo district that recorded high diseaseincidence levels of 85.8% had the lowest severity levelsof 13.8% compared to Soroti, Kumi and Pallisa districts.

Virus detection. One hundred eight leaf samples collectedfrom four cowpea growing districts were subjected toDAS-ELISA tests using antisera to CABMV, CCMV,CMMV, CMV and CSMV. Thirty five samples of the total108 collected were virus infected. Three viruses namelyCMMV, CABMV and CSMV were detected in the samples.Cowpea severe mosaic virus (CSMV) was detected in thesamples in the four districts surveyed. A total of 24 (22.2%)

0

20

40

60

80

100

120

Soroti Kumi Pallisa Tororo

Dise

ase

inci

denc

e (%

) and

seve

rity

(%)

Disease incidence (%)Disease severity (%)

Figure 1. Percentage incidence and severity of cowpea plants with viruslike symptoms in the major cowpea growing districts in eastern Uganda.

1281Occurrence and prevalence of cowpea virus diseases

symptomatic samples reacted positive with CSMVantibodies thus making CSMV the most prevalent virus inthe districts (Table 1). This was followed by CMMV witha total of 7 (6.5%) symptomatic samples occurring only inthree districts, with the exception of Tororo district.Cowpea aphid-borne mosaic virus was detected in 4 (3.7%)diseased plant samples obtained from Pallisa and Tororodistricts, and it was the least frequent virus among theviruses detected.

Single and multiple virus infections. A proportion of21.3% of symptomatic samples were infected by a singlevirus, whereas 7.4% were infected with two viruses. Themost common with single infection was that of cowpeasevere mosaic virus (13%), followed by cowpea mild mottlevirus (6.5%) and the least was cowpea aphid-borne mosaicvirus (1.9%). However, cowpea severe mosaic virusoccurred in mixed infections with other viruses namelycowpea severe mosaic virus and cowpea mild mottle virus(5.6%), and cowpea severe mosaic virus and cowpeaaphid-borne mosaic virus (1.9%) (Fig. 2).

Discussion

Cowpea viruses have become increasingly important inthe cowpea growing regions in Uganda and subsequentlyinflicted heavy losses to yields. The results reported hereinindicate substantial incidence and severity of virusesduring the survey conducted in the cowpea growing

districts in eastern Uganda. There were considerablevariations of incidences and severities among the districtssurveyed. For instance, virus incidences ranged between46.6% and 96.2%, and severities ranged between 13.8%and 31.8%. The incidences were high in Pallisa and Tororodistricts compared to Kumi and Soroti districts. Thesevariations in occurrence under different agro-ecologicalenvironments can be due to the fact that the low virusincidences in Kumi and Soroti may suggest presence oflow virus vectors and consequently low virus inoculumsource to cause high incidence in cowpea grown fields.Elsewhere, studies have shown that with changes in thefarming systems, cultivar types and dynamic populationof virus vectors may affect virus occurrence (Wilser et al.,1998). Edema et al. (1997) and Shoyinka et al. (1997)reported the effect of weather conditions within seasonsand cropping systems on distribution of viruses in thedifferent environments. In other studies, however,manipulation of plant populations especially at relativelyhigh plant densities also showed to affect the dynamicsof virus vectors that transmit viruses to plant crops(Ogenga-Latigo et al., 1992a,b). Similarly, according toMukankusi et al. (1999) on groundnut rosette disease andOrawu et al. (2001) on cowpea viral diseases, also reportedlow virus incidences under high plant populations,suggesting that unfavourable microclimate disfavoursvirus vectors to colonise plants.

Three cowpea viruses namely cowpea aphid-bornemosaic virus, cowpea severe mosaic virus and cowpea

Table 1. Number, percentage incidence and virus types when serologically tested in the symptomatic samples collected in four districts of Uganda.

District Samples tested Virus serological detection by DAS-ELISA

CABMV CCMV CMMV CMV CSMV

Soroti 32 - - 1 (3.1) - 11 (34.4)Kumi 32 - - 4 (12.5) - 1 (3.1)Pallisa 22 2 (9.1)* - 2 (9.1) - 7 (31.8)Tororo 22 2 (9.1) - - - 5 (22.7)

Total 108 4 0 7 0 24

* Figures in parentheses are percentage incidence. - Indicate no virus was detected in the samples in any of the districts.

0

2

4

6

8

10

12

14

CSMV CMMV CABMV CSMV + CMMV CSMV + CABMV

Perc

enta

ge p

lant

s inf

ecte

d

Figure 2. Occurrence of single and multiple virus infections in symptomatic cowpea plants.

M. ORAWU et al.1282

mild mottle virus were detected in the survey ofsymptomatic cowpea plants, suggesting their existencein the major cowpea growing areas in eastern Uganda.Cowpea severe mosaic virus was the most common viruswhile cowpea mild mottle virus was the second mostprevalent virus detected in all surveyed cowpea growingdistricts. Our results suggest that cowpea aphid-bornemosaic virus was the third and least frequent virusdetected in the samples from the four surveyed districts.Although an earlier report by Edema et al. (1997) indicatedcowpea aphid-borne mosaic virus as the most commonvirus in Uganda, but on contrary our results showedcowpea severe mosaic virus as the common virus basedon four surveyed districts in eastern Uganda. From thesurveyed districts, antibodies of cowpea chlorotic mosaicvirus and cowpea mosaic virus were included in theserological tests of cowpea samples, but no samplesreacted with antisera of these viruses, suggesting thatthey may not be present in Uganda.

Multiple-virus infections are common among samplesfrom field grown cowpeas and are known to modify andcomplicate symptoms, thus precluding in field diagnosisbased on symptoms (Shoyinka et al., 1997). In the study,however, mixtures of viruses were observed in only twoof the 108 cowpea samples. The shared viruses in thesamples were cowpea severe mosaic virus and cowpeamild mottle virus, and cowpea severe mosaic virus andcowpea aphid-borne mosaic virus. These results comparewell with other studies on virus occurrence in mixedinfections (Shoyinka et al., 1997). The higher incidence ofcowpea severe mosaic virus (13%) in the samplescompared to cowpea mild mottle virus (6.5%) and cowpeaaphid-borne mosaic virus (1.9%), could suggest its relativeabundance of beetle vectors to survive under adverseenvironmental conditions over the white fly and aphidvectors in the surveyed districts. However, there was noassociation between cowpea mild mottle virus and cowpeaaphid-borne mosaic virus in cowpea samples, probablythe aphid vectors and white fly vectors are not linkedtogether to cause synergism effect in cowpea plants.

Generally, the study confirmed three economicallyimportant cowpea viruses and their relative complexes inthe cowpea fields. This situation demands that urgentattention should be given to strengthening themanagement of cowpea viruses in Uganda. The onlyintervention to enhance production could be through theuse of resistant varieties, but this is often compoundedby pathogenic variability and occurrence of virus mixtures.However, exploitation of cowpea genotypes with multipleresistance genes would offer a better option in the breedingwork to control several viruses so as to circumvent theescalating problems of viruses and ultimately yield losses.

Acknowledgements

This study was funded by the Rockefeller Foundationthrough the African Centre for Crop Improvement in SouthAfrica. We gratefully appreciate the assistance extendedby director of SAARI and the Makerere University,Department of Crop Science for the facilities provided for

the work. We also thank the support of the farmers whoallowed us free access to their fields to conduct thesurveys.

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