streak, maize stripe and maize mosaic diseases QJ in the tropics (Africa and islands ■ h in the Indian Ocean)

Dossier prepared by f C. JOURDAN-RUF, J.-L. MARCHAND, M. PETERSCHMITT ■ Impact of maize virus @®CA, BP 5035, 1 • 34032 Montpellier Cedex 1,iranee diseases and current research B. REYNAUD, J. DINTINGER I C tR i0 € A , station, de Ligne Pqljdis, ■ Vectors ■ 97410 Saint-Pierre, Réunion, Fw ce

The main research scientists that and epidemiology collaborated in these studies are: G. KONATE, O. TRAORE and S. TRAORE ■ Maize virus diagnosis and (Burkina Faso); #> f r; M. ESSEH-YOVO ¡Togo); C. THE (Cameroon): variability H. PHAM ¡Zimbabwe); P. MARKHAM (UK); C. BUDUCA, B. CLERGET and A. RODIER ■ Maize resistance [Réurtion, France) = } M i ' and breeding

Photos D. Debert: Bagging maize inflorescenles. p /Vaize fields in Burkina Fasfx Photo B. Reynaud: Peregrínuâmaidts. Impact of maize virus diseases and current research

Regions Madagascar, Réunion, and central in East Africa, Tanzania, Kenya, Africa. The disease is also sometimes Mauritius and Réunion, but MStpV found in West Africa, particularly has caused very little damage. The The three main tropical maize in Mali, Togo, Côte d'Ivoire virus was recently identified are: maize streak virus (MSV), and Senegal. In West Africa, in West Africa (Côte d'Ivoire, maize stripe virus (MStpV) and maize there was a widespread outbreak Togo, Nigeria, Burkina Faso and mosaic virus (MMV). Their impacts of MSV in 1983-1984, sometimes Cameroon). However, it seems to vary markedly between countries completely destroying all maize have had a greater impact in Latin (Figures 1 & 2). crops. This disease also occurs in America, particularly Venezuela and MSV was first detected in South India in wheat and millet, but it has the West Indies. It also occurs in the Africa at the beginning of the not been detected in the South Philippines and Australia. century, and similar virus diseases American tropics. have been described in MMV, first discovered in 1921 and in many wild Poaceae species. It MStpV is not as common. Peregrinus in Hawaii, is also transmitted by is transmitted by homopteran maidis Ashmead (Homoptera, P. maidis. It was later detected of the genus (Homoptera, Delphacidae), the vector, is in Cuba, Guadeloupe, Surinam, Cicadellidae). MSV infections have present in many African countries Venezuela, Guyana, Puerto Rico, been noted in many different (Côte d'Ivoire, Zaire, Nigeria, Burkina Faso, Côte d'Ivoire, Nigeria, countries, with varied incidence. It is Burkina Faso and Cameroon). Since Mozambique, Tanzania, Kenya, endemic to all East African countries, 1936, this insect has been reported Réunion, Mauritius and India.

Evolution of host plant/vector/virus complexes

P. maidis and the viruses it transmits evolved together in a few plant species. Maize is the favoured host of this vector/virus complex; the original host was teosinte or sorghum. The close host plant/vector relationship perpetuates the complex with the virus, despite the narrow range of host plants and susceptible species. The evolution of MSV, transmitted to maize by Naude, is different: the virus was revealed in maize crops, but also infects wild grasses through the vector, which is common in Africa and has a wide host range.

5 6 Agriculture et développement ■ Special issue - December 1995 and more regular maize yields could in resistance transfers to create maize Organizations be obtained with such varieties. varieties that will be more efficient under cropping conditions found in Maize breeding studies began in the involved Africa and throughout the Indian 1930s in East and South Africa with Ocean region. The developed the aim of obtaining MSV resistance. CIMMYT, Centro Internacional de diagnostic techniques can be applied Moreover, the 1972 MSV epidemic Mejoramiento de Maíz y Trigo, Mexico at a variety of sites, without special that occurred in West Africa promp­ CIRAD, Centre de coopération equipment, to identify viruses and ted certain research organizations internationale en recherche assess varietal susceptibility. (IITA and CIMMYT) to conduct agronomique pour le développement, similar studies. France CORAF, during the general assembly meeting of its maize network in CORAF, Conférence des responsables During the 1 970s, CIRAD (in Yaoundé (Cameroon) in 1987, deci­ de la recherche agronomique africains collaboration with African and European partners) focused research ded to launch an MSV research pro­ IRA, Institut de la recherche ject because of the urgency of the agronomique, Cameroon on this topic in order to deal with the serious maize virus epidemic situation. For resistance transfers, INCV, Institut national des cultures problems — the three mairi maize national CORAF correspondents vivrières, Togo viruses and their vectors are present chose a number of varieties from INERA, Institut national d'études rurales including local resistant varieties. among already (or soon to be) et agricoles, Burkina Faso widely grown maize varieties. IITA, International Institute of Tropical The research This multidisciplinary project brings Agriculture, Nigeria together virologists, entomologists John Innés Institute, Norwich, UK and who is involved and geneticist/breeders: Since the 1980s, CIRAD and partners - virus studies by John Innes Institute, The incidence of MMV is generally have been focusing on the following Norwich (UK) and CIRAD, Montpellier (France); low in all of these countries. topics: Nevertheless, serious MMV - insect vectors and their - interactions between viral isolates epidemics have been reported in epidemiology; and resistant genotypes in Réunion some regions of maize growing - maize virus diagnosis; (CIRAD) and Burkina Faso (INERA); countries such as USA (Hawaii, - breeding and investigating - insect vectors in Réunion (CIRAD), Florida), Brazil and Mexico. the genetic factors determining Burkina Faso (INERA) and Cameroon resistances, and transferring them (IRA); Control into susceptible maize genotypes. - maize virus dynamics in host Virus-resistant varieties of maize are These investigations revealed the plants in the tropical phytovirology laboratory (CIRAD); of considerable economic interest vectors and factors involved in due to the extent of damage caused triggering and spreading viral - resistance genetics in Réunion by maize viruses in the tropics epidemics in maize crops and led to (MSV, MStpV, MMV) (CIRAD) and in (Africa, South America) and the the development of specific maize Togo (MSV) (INCV); lack of efficient agronomical and virus diagnosis techniques. Resistant - creation of resistant varieties in chemical control techniques. Higher ecotypes in Réunion have been used Réunion, Togo and Cameroon. ■

Agriculture et développement ■ Special issue - December 1995 5 7 Symptoms

Maize streak disease

Maize streak is characterized by chlorotic spots along the leaf veins, forming discontinuous streaks of varying thickness. Dwarfism is very marked in young infected plants.

Dwarfed young plant.

MSV leaf symptoms.

Maize mosaic disease

Maize mosaic is characterized by regular continuous chlorotic lines running along the whole length of the leaf, parallel to the veins. The symptoms vary according to the thickness of these lines, their spacing and discontinuous appearance.

Limited growth of infected plants.

Continuous chlorotic stripes caused by MMV.

Maize stripe disease

Maize stripe occurs in two forms. The first, simply called stripe, is characterized by chlorotic stripes of various widths along the leaves, and the apex is typically curved. The second form, called chlorotic stripe, is characterized by the formation of chlorotic stripes, and the leaves eventually become totally chlorotic, with the reappearance of thick green discontinuous stripes. Young infected plants present severe dwarfism, and then dry up and die.

Chlorotic stripes on maize leaves due to MStpV. Photos P. Baudin and B. Reynaud Infected plants are dwarfed and dry up.

5 8 Agriculture et développement ■ Special issue - December 1 995 S t # * *

Virus reservoir plants Vectors and A 3-year (1988-1990) wide ranging survey of herbaceous plants was conducted in Burkina Faso, resulting in the detection of 41 Poaceae epidemiology species with MSV symptoms. ELISA tests revealed the presence of the virus in 25 species.

Analysis of the spatiotemporal distribution of virus reservoirs (host As early as 1973, Insect vectors plants) highlighted three main points: severe epidemics of several reservoirs were greater in maize crop In Réunion, Cicadulina mbila is the zones; the incidence of MSV in maize viruses were noted main MSV vector even though this maize fields was proportional to the insect occurs in relatively small in Réunion. In 1985, three number of reservoirs; and there were quantities on maize, the final host of very few infectious reservoirs in May, viruses were identified the disease. It is a very active and with an increase 3-4 weeks after the efficient Poaceae-specific vector. by serological techniques: onset of the rains to reach a peak C. storeyi is the next important MSV in August. In Burkina Faso, the MSV, MStpV and MMV. vector in Réunion. Insect vectors are required incidence of MSV is generally 6% or In Burkina Faso, five Cicadulina less in maize plants presenting virus for their transmission. species have been identified in symptoms; it increases to 30-40% MSV is transmitted all ecological zones: C. mbila, during severe epidemics. C. similis, C. arachidis, C. triangula In Sudano-Sahelian regions, the risk by at least eight different and, less commonly, C. hartmansi. of early infection (often severe) could Cicadulina species. The virus transmission potential be minimized by early sowing varies in the different leafhopper MStpV and MMV (mid-May to mid-June), during a species, with C. mbila being the most period when the inoculum source is are transmitted by Peregrinus efficient and C. arachidis the least. minimal. However, this recommen­ dation cannot always be met on maidis. Research carried P. maidis is a sedentary vector that is account of the cotton and rice out to date has specific to maize and sorghum which it colonizes in relatively cropping calendar. mainly focused high densities. Its maize virus on the Sudano-Sahelian transmission efficiency is intrinsi­ The importance cally low. region (particularly of climatic factors Burkina Faso) and Réunion. In Burkina Faso, 10 years of MSV host plants agroclimatic studies demonstrated and vector the effects of climate on MSV incidence, in order of importance: epidemiology in the relative humidity in February; relative humidity in January; and Sudano-Sahelian temperatures in May. region In January, at the beginning of the dry season, insect densities are high A detailed study, representative of and they are obliged to migrate to Sudano-Sahelian regions, was set up refuge areas (sites with permanent in Burkina Faso in 1984. water supplies, irrigated crops) H i g h I y MSV-sensiti ve maize because of the lack of host plants. cultivars (Safita II and Jaune flint de The relative air humidity decreases Saria) were used for all tests of virus through January and February, which transmission with the leafhopper can affect insect breeding and C. triangula. The serological ELISA survival and lead to high mortality. (enzyme-linked immunosorbent The number of MSV vectors assay) technique was used to detect available in the following rainy MSV in collected samples. season is determined by the size of

Agriculture et développement ■ Special issue - December 1995 5 9 optimal for C. mbila and preci­ pitation favours development of Poaceae hosts. With hot temper­ atures, there is a higher risk of epidemics of MSV than MStpV, with a reversal of the situation at the end of the dry season. The cooler weather is more suitable for P. maidis than C. mbila.

P. maidis can survive at more moderate temperatures, which means that epidemics can occur at high elevations. Since P. maidis generally infests maize, areas where this crop is grown intensively are Cicadulina mbila nymph and adult. seriously affected by MStpV. The MMV infection rate of maize is not very high in irrigated lowland zones. However, it should be noted that the symptom-based technique the residual insect population, which for evaluating viral infection levels in in turn determines the extent of MSV maize plants often underestimates development. May precipitation was MMV incidence; this is particularly found to be very important in the true when there are secondary MSV MSV epidemics of 1 983-84. In and MStpV infections, since these Zimbabwe, March-June precipitation two virus diseases mask MMV is also a factor that determines symptoms. Cicadulina spp. population densities from July to September, and thus MSV infection rates. Moreover, in Efficiency of virus Zimbabwe and Réunion, the rising temperatures during the southern hot transmission season promotes development of to the insect vector Peregrinus maidis adult. vector populations. Photos B. Reynaud The intrinsic transmission ability expresses the specificity of the Impact of three infectious agent/vector relationship. maize virus With persistent viruses, the cycle of virus transmission to the insect diseases in Réunion occurs in the following steps: - acquisition access period, when In Réunion, the population dynamics the insect acquires the virus after of maize virus insect vectors were feeding on an infected plant; studied by sowing maize on a - latent period in the insect, or time bimonthly basis from 1983 to 1985, and on a weekly basis from 1985 required for the insect to become to 1988. infective; - inoculation access period, when The virus symptom observations and the plant is contaminated through an vector trapping results for lowland infective insect puncture. areas in Réunion indicated that epidemics of the three virus diseases Moreover, there is a virus retention differ in terms of their locations, period in the insect, during which extents and seasonal patterns. At low the vector remains infective. elevations, MSV causes very high to The cycle of MSV transmission to the total maize losses. It is the most com­ vector is short, i.e. 24 h or less; those mon maize virus disease during the of MStpV and MMV are about hot season, when temperatures are 15 days.

6 0 Agriculture et développement ■ Special issue - December 1995 No MSV multiplication in the insect was noted since, after a short acquisi­ tion access period, there, was an overall reduction in transmission rates and viral concentrations. This insect vector is efficient because of its rapid uptake of the virus during acquisition and the low viral concentrations required during inoculation. The virus takes a long time to disappear in the insect, thus explaining its excellent long-standing infective potential.

MSV could be considered as a circulative non-propagative virus.

MStpV transmission by P. maidis

P. maidis populations have a relatively low MStpV transmission Virus acquisition can be followed by capacity, i.e. about 20% in Réunion, a latency period and the inoculation. Depending on the virus and insect vector, which is quite close to levels noted in various transmission barriers have been Transmission studies revealed that other regions. identified. In different insect organs, some MSV, MStpV and MMV have cell membranes can act as barriers or as characteristic circulative-type virus MStpV transmission by P. maidis is pathways linked to a specific virus patterns, with a latency period in the relatively inefficient. Transmission recognition mechanism. insect, maintenance of the infective barriers have been detected: the first potential during moulting, presence involves hindering penetration of the Figure 3. Diagram of the infection of the virus in the insect and inocula­ virus through the intestinal wall process in the insect vector tion of the plant with insect saliva during acquisition feeding, and (from REYNAUD, 1988). (Figure 3). Very little is known about secondly through the salivary gland the latency period in the insect — it walls. Some females can transmit the includes the transit time of the virus virus to their offspring through their through the insect's body and its ovaries. The virus can multiply in uptake in the salivary glands to a most of these insects from the egg specific concentration threshold. stage onwards. Percentages of infective insects in a For artificial infestations (in varietal population are determined to assess screening tests for resistance), vector transmission efficiencies. it is now possible to select for the Captures and tests of different alimentary and ovarian acquisition leafhopper populations showed that ability. Populations of vectors with a 50% (on average) of C. mbila insects high intrinsic transmission ability can in a population are MSV vectors, thus be obtained within a few while 20% of P. maidis insects are generations. MStpV and MMV vectors.

MSV transmission MMV transmission by C. mbila by P. maidis

In Réunion, C. mbila is an excellent In Réunion, P. maidis has a low MSV vector with a high intrinsic MMV intrinsic transmission capacity, transmission capacity (above 50%), i.e. about 20% infectious insects. which corresponds to levels noted in Two transmission barriers, similar populations studied in South Africa to those described for MStpV and Burkina Faso. transmission, were found. ■

Agriculture et développement ■ Special issue - December 1995 Ó 1 Maize virus diagnosis and maize streak virus variability

Symptomatic diagnosis of virus diseases is often imprecise Features of and should be confirmed with various serological tests or electronic microscopy. This is especially true for early monoclonal stages of infection when diagnoses could be faulty if not antibodies confirmed by reliable serological techniques. ELISA Antiserum prepared against a given (enzyme-linked immunosorbent assay) techniques were used virus will naturally contain a set of to obtain clear diagnoses of maize virus diseases present antibodies that recognize several capsid protein sites in the viral particle. In in maize selection plots in Réunion. MSV, MStpV and M M V contrast, a monoclonal antibody only symptoms can be characteristic of each of the three viruses recognizes one capsid protein site, thus enabling detection of slight variations in at relatively advanced stages of the disease, such proteins. Monoclonal antibodies but the appearances sometimes vary, especially are produced through fusion of lymphocyte cells and myeloma cells. for MMV. Secondary infections of several viruses These hybrid cells (hybridoma) have the can also occur. perpetual traits of myeloma cells along with the ability of lymphocyte cells to produce single antibodies. CIRAD, in collaboration with the Institut de Virus purification d'Ivoire, Mali and Burkina Faso. biologie moléculaire et cellulaire Conventional ELISA tests can only be (Strasbourg, France), have developed and diagnostic carried out in the laboratory. five monoclonal antibodies that Nevertheless, CIRAD (Montpellier, recognize MSV. techniques France) has developed a modified ELISA test for detections of all The three maize viruses (MSV, three viruses on nitrocellulose Initially, on the basis of biological MStpV and MMV) have been membranes, without requiring any tests, serological techniques purified. The extracts were injected laboratory equipment. This new (with monoclonal antibodies) and into rabbits to produce specific technique will facilitate studies on electronic microscopy, the isolates immunoglobulins. geographical distributions of these were all considered to be MSV viruses and help in determining ELISA tests were developed for maize strains. Later, nucleotide sequence specific requirements for breeding virus diagnosis. These quick simple analyses of the viral genomes resistant varieties (Figure 4). tests are suitable for routine virus revealed that some of these strains disease diagnosis. are actually distinct viruses.

These tests helped to pinpoint MSV MSV variability in eight African countries. The In 1986 at CIRAD, serological tests presence of MStpV and MMV has Virus isolates that induce streak leaf on streak viruses of grasses led to the rarely been proven in Africa, but they symptoms have been described in characterization of three different were recently detected in Côte several grasses, including maize. serotypes.

6 2 Agriculture et développement ■ Special issue - December 1 995 >%eS

1 2 3 4 Virus diagnosis kit

A kit was developed for serological field If, on the other hand, the user also diagnosis of viruses using nitrocellulose wants to conduct the analysis, membranes. It can now be purchased everything required for the testing and (from CIRAD) by anyone wanting to interpretation is supplied in the kits, serologically confirm a visual diagnosis. i.e. unrevealed membranes and This diagnosis kit is very easy to use: positive/negative controls. - the test kit contains everything Samples from Zimbabwe and Réunion required for the analysis; were tested in various different ways to A - it is very compact; check the validity of kit for field - the test can be stopped after plant analyses: extracts are deposited on the 1 2 3 4 - tests performed completely at the membrane, the membrane just has to be sampling site; dried. In this form, the test can be completed within the next 20 days; - plant extracts deposited at the - these dried untested membranes can sampling sites and membranes analysed be sent by mail in a simple envelope elsewhere; and tested later. - tests performed completely at CIRAD For instance, someone not wanting to (Montpellier, France) with leaves sent spend the time required to perform the from Réunion and Zimbabwe. serological tests can simply deposit plant extracts on the membranes and send The results showed that this virus them to a correspondent for analysis. diagnosis kit is highly reliable and valid. B

Figure 4. Immunoenzyme detection of MMV (A) and MSptV (B) using ELISA from four crude extracts of maize from Réunion, infected by MSV (1 ), M M V (2), Sugarcane, Digitaria sp. and Setaria countries, or in wild Poaceae isolates MStpV (3) and uninfected (4). Each plant sp. isolates could thus be distingui­ — some of which showed different extract was tested at dilutions of 1 /1 0, shed from various maize and virulence when tested on maize. The 1/100 and 1/400 (leaf weight/buffer Poaceae isolates belonging to the serological technique was found to volume). same serotype. be unsuitable in several locations, Photo M . Peterschmitt particularly Réunion, for distingui­ The gene sequences also confirmed shing isolates with different these distinctions with respect to pathogenicities. other isolates — even indicating that sugarcane and Digitaria sp. isolates Studies carried out in the Sudano-Sahelian region, where the are distinct streak viruses that have Sudano-Sahelian zone confirmed the maize serotype and two others have been called sugarcane streak virus presence of one dominant serotype been identified, molecular analyses (SSV) and Digitaria streak virus in maize crops. In more than will reveal the presence of streak (DSV), respectively. Similarly, a 300 maize isolates analysed, 99% viruses of grasses that differ Panicum sp. isolate was reclassified, showed a characteristic serological from MSV. i.e. after it was initially identified as maize profile (which was previously an MSV isolate, it is now considered determined by CIRAD). However, as a distinct virus called Panicum half of about 100 isolates tested were MSV resistance streak virus. Further viruses could found to contain both the maize thus be distinguished in the future serotype and another serotype that mechanisms with other isolates. Setaria sp. had been identified in wild Poaceae isolates have not yet been analysed species. This modifies previous in maize in detail, only serological tests have conclusions, indicating that major been carried out. In summary, the serotype in maize can occur in the Various mechanisms can be serological techniques used in association with other strains involved in a plant's virus resistance. Réunion did not enable serotype whose serological profiles and Such mechanisms were compared differentiation in MSV isolates, or in pathogenicities might be masked by in the resistant maize variety other maize isolates even though the maize serotype. As in other IRAT 297 and the susceptible they originated from 11 different locations, it is likely that in the variety INRA 508.

Agriculture et développement ■ Special issue - December 1 995 6 3 Location of MSV in the maize plant Maize resistance Virus colonization processes in the plant (MSV capsid antigens) have been defined using the ELISA technique. After a plant leaf is and breeding inoculated by C. mbila, the virus migrates through phloem canals to the stem. MSV colonizes the leaves through young dividing cells in which viral DNA replication can occur. Virus spread can only be The main aim of the research carried out in Réunion detected in tissues that form is to transfer the resistance of some local maize varieties after infection. to three maize viruses (MSV, MStpV and MMV) into other With this mode of infection, the younger the plant is at inoculation, susceptible varieties of high agronomic interest. the higher is the percentage of MSV-resistance transfers are also under w ay in Togo, and infected tissues and the greater the yield losses. Since the virus is able to very recently in Cameroon. This breeding work with varieties migrate from the inoculated leaf to that are already being grown by farmers, or promising ones, the stem in less than 2 h, no prepro­ interests many African CORAF-member countries, and also pagation of the virus seems necessa­ ry. Leafhopper virus transmission countries in Latin America and the West Indies. tests showed that the virus can only be acquired from leaves showing symptoms.

Resistance mechanisms Overall, more than 800 maize Analysis of virus distributions in varieties from tropical and temperate maize plants indicated identical Resistance sources regions have been screened in distributions in the susceptible found in Réunion Réunion under natural virus infection variety (INRA 508) and in a resistant conditions. None of them are variety (IRAT 297). On the sufficiently resistant for cultivation in other hand, a viral concentration In Réunion, resistance was discovered in Réunion, except for ecotypes from difference between these varieties local maize varieties, especially in Rodrigues which are genetically was detected by measuring virus coastal varieties. This adaptation was similar to those found in Réunion. nucleoproteins using the ELISA acquired through natural selection. In One long-term objective is to improve technique, i.e. these nucleoproteins 1 974, the Réunion variety "Revolution" yields and agronomic qualities in were 10- to 90-fold more concentra­ was found to be MSV resistant in Benin. local maize varieties. The main ted in INRA 508 than in IRAT 297. This was confirmed in several countries, problem is that these varieties have These results indicated that virus i.e. in Kenya in 1976, USA in 1983 low yield potentials and they are all resistance in cv IRAT 297 involves and Nigeria in 1982. early producers. They have a poor mechanisms that hinder viral About 100 ecotypes were collected in a improvement potential because of propagation rather than blocking 1 979-80 survey conducted in Réunion their very narrow genetic bases. migration of the virus. and Rodrigues. They were tested under Hence, resistance transfers would be natural infestation conditions of MSV, The lower viral propagation in IRAT quicker and more efficient, and MStpV and MMV, and 41 of them were 297 is thus responsible for reduced enable creation of a wide range of found to be resistant to maize viruses. severity of the initial infection; this is resistant varieties from already This resistance was later confirmed shown by less severe symptoms and improved plant material of high under artificial infestation conditions. a longer period before symptoms agronomic value. These ecotypes were polycrossed to appear as compared to the suscep­ produce the "Composite viroses tible variety (in comparative tests, résistant" (CVR), which was further Preliminary studies 5 days versus 3 days, respectively). improved through three recurrent Secondary infections are less severe The following four conditions breeding cycles, and then used as a in resistant plants since they are a had to be met to be able to transfer resistance donor in transfers. poor inoculum source. ■ resistance to these three viruses

6 4 Agriculture et développement ■ Special issue - December 1 995 ‘ 1 ^ ;^e through backcrossing: - pinpoint resistance in germplasm Rating of the Indian Ocean islands, and select the most resistant populations Virus symptoms are graded visually on basis of these individual plant scores. to polycross them in a "Composite a semi-quantitative 0-5 scale, while For instance, in tests carried out in virose résistant" (CVR); taking the seriousness of the symptoms Réunion, it is considered that plants with into account. - carry out mass rearing of insect scores of 2 or less, contrary to those Plant symptom scoring: vectors to be used for inoculations. scored 3-5, will not produce lower 0 - the plant shows no symptoms; C. mbila, present in Réunion, is yields because of the presence of the 1 - a few chlorotic spots are detected on considered to be the main MSV virus. The overall scores take this vector and was thus chosen as vector the plant during a detailed inspection; 2 - slight streaks are clearly visible on threshold into account, e.g. a variety for the varietal improvement the plant; with no plants scored 3 or higher is programme. P. maidis is the only known vector of MStpV and MMV, 3 - moderate streaks are visible; attributed an overall score of 0 and 4 - the plant presents serious streaking which is also present in Réunion; considered as a resistant variety. In with dwarfism; - develop a system to screen plants contrast, a variety with all plants scored 5 - the plant presents very serious virus for virus resistance, through artificial 5 is very susceptible and is allotted an infection with highly marked dwarfism. infestation techniques using C. mbila overall score of 500. The overall score is The symptoms do not change after (currently being perfected for calculated as follows: 35 days postinfection. Symptoms are P. maidis). A resistance scoring scale 100 X (3 X N3 + 4 N4 + 5 X N5)/ I N| thus scored and plants for resistance is established; transfers are chosen between this stage (0 to 5) - serological tests were developed to and flowering. Overall scores can be where N¡ = number of plants scored check, when necessary, whether the attributed to families or varieties on the ¡(1 ,2 , 3, 4, 5). transmitted virus is the right one. Preliminary studies demonstrated that resistance to MSV from Réunion ecotypes was high in many countries, and that the Réunion MSV Artificial infestation strain was especially virulent. Artificial infestations are essential for plant screening since the proportions of the three maize viruses can vary during the year: natural viral pressure is not MSV resistance homogeneous or constant enough for precise screening. These artificial infestations require highly productive mass rearing and active vectors to limit the number of insects research that have to be produced. The percentage of infective insects is increased by lengthening the virus acquisition period on diseased plants and by breeding insects C. mbila is easy to breed and to enhance their transmission ability. Insects used for transfers of MSV resistance are artificial infestation can be carried obtained from a selected C. mbila population in which all insects are able to transmit out without difficulty. Research on the virus. These are placed on MSV is therefore more advanced plants presenting very severe than on MMV and MStpV in symptoms and left for 3 days for Réunion. virus acquisition. Carbon dioxide This virus is also mainly found in anesthesized insects are set in the whorl of each plant. A plant's Africa, thus explaining the develop­ susceptibility decreases with ment of MSV resistance transfer age, which means that very programmes in Togo, and more early infestation is necessary. recently in Cameroon. Infestations are carried out 10 days after sowing for CVR improvement practical reasons (at this point the plants have grown to a sufficient Since preliminary tests revealed high size for depositing the vectors). internal variability in resistance, The infestations must be efficiently recurrent selection work was under­ conducted to certify that all plants taken to improve CVR, first under are infected. Experience has natural viral MSV pressure (cycle shown that complete infestation ______1, CVR-CI), then under artificial can be obtained by depositing infestation (cycles 2 and 3, CVR-C3). three viruliferous on Artificial field infestation with C. mbila. This markedly increased the level of each plant. Photo B. Reynaud MSV resistance in CVR.

Agriculture et développement ■ Special issue - December 1995 In 1986, variety CVR-C1 was registe­ red under the name IRAT 297 with Breeding techniques the Crop Science Society of America. Its MSV-resistance value was verified The three main maize breeding techniques are based on certain features in this plant's through comparisons of susceptible floral biology, i.e. its cross-pollination and selfing potential. varieties with IRAT 297 top-crosses. Pedigree selection. Pedigree selection involves selfing a certain number of plants, thus producing SI families. Families of interest are chosen according to various criteria and Genetic factors then selected plants selfed within these families. This produces S2 families, with subsequent selfing and choosing to produce S3 and S4 families, etc. The breeding determining MSV process is generally carried out up to S6-S8, thus producing homozygous lines with resistance low vigour, but which can be used to create hybrids through heterosis. Recurrent selection. Recurrent selection involves choosing some plants of a variety or Based on the improved CVR-C3 composite. Seeds of cobs from these plants can be sown in "ear to rows" for instance form, 500 S1 families were produced with each cob producing a seed line. Choices are then made according to one or by selfing, and tested through artifi­ several criteria and chosen families are crossed to produce a new variety. The variety cial MSV infestation. The very low is progressively improved through several choosing and crossing cycles. percentage of MSV symptom-free Backcrossing. Backcrossing is used to transfer a trait to an interesting variety that does plants, the variability in the symptom not already have it. The FI progeny of the receiving variety and the trait donor are scores in each family and a gauss screened for the trait. Plants with the trait are backcrossed with the receiving variety. distribution curve plotted for all After several backcrossing cycles, varieties are produced that somewhat resemble families indicated that factors the initial variety but contain the sought-after trait. determining MSV resistance are complex and likely controlled by a polygenic system. but this cannot yet be attributed to a maintained after the first backcros­ Nevertheless, a few S5 lines with separate (probably polygenic) system sing (Figures 5 & 6). It is useful to very high levels of resistance (even or to partial expression of a major begin screening at the F1 generation complete resistance in one line) were system. in order to discard as many recessive obtained after four more selfing genes as possible from the outset. cycles. Genetic analysis of this mate­ rial revealed a major system of very Resistance transfers The resistance transfer procedure is heritable complete resistance, with limited to a single initial cross with MSV resistance transfers are perfor­ positive incomplete dominance, the donor and then two backcrossing med through a series of conventional involving at least three different cycles. The final variety therefore backcrossings. genes. Moreover, the presence of contains the resistance trait and 90% partial resistance was indicated In Réunion, for each screening, of the genes of the receiving variety. during the breeding of some lines — 5 000 F1 plants are sown and Selfing is necessary to recover the infected 10 days later by depositing colour and grain-type of the three preinfected leafhoppers in the receiving variety — when they differ whorl of each plant. Resistant plants from those of CVR. This was done for are chosen before flowering through MSV resistance each screening in the Togo trials. In the elimination of susceptible plants. Réunion, selfing was carried out after research conducted The chosen plants (only 250-300) the second backcrossing cycle. by IITA in Africa are intercrossed. The F1 and F2 generations of each breeding cycle are thus screened. Current results In 1972, MSV resistance was detected by IITA in the cultivar Tropical Zea The same procedures are used in the Presently, five varieties from Yellow (TZY). This variety was then bred research programme carried out in the backcrossing programme in and resistance transferred to other Togo, but the artificial infestation Réunion, i.e. CN7, Tiémantié, higher-yielding varieties. The improved technique differs. Maize plants are Pool 16 SR, IRAT 171 and Suwan line IB 32 (obtained in 1979) was found sown in pots. Once the maize plant 8331, are considered to be modified to have high partial resistance. It was emerges, the pots are placed in cages after two backcrossing cycles and used as a resistance donor. A selection containing infected leafhoppers. The three selfing cycles. In Réunion, they programme was carried out but plants are then planted out in the have been found to possess excellent completely symptom-free plants were field. The chosen plants are selfed resistance. Most plants showed no rejected; many varieties have been and then crossed again. The resistan­ symptoms under severe artificial produced and distributed throughout ce screening is very efficient. infestation conditions with a highly Africa. Resistance is at least partially virulent virus isolate.

6 6 Agriculture et développement ■ Special issue - December 1 995 500 Concerning the maize breeding carried out in Togo, five varieties (Violet de Katiola, ZL 2BD, Pool 16 DR, early Blanc 2 and AB 21) are

400 - at the second backcrossing stage and are therefore converted. Different breeding procedures are being assessed in terms of MSV resistance O and preservation of grain texture and 300 husk coverage of maize cobs, traits that were sought in the selection process.

200 - MStpV and MMV

m Tiémantié resistance • IRAT 171 100 Comparison of the performance of Suwan 8331 the IRAT 297 composite with that of o CN7 the susceptible INRA 508 hybrid □ Pool 16 SR under natural conditions highlighted a high level of MStpV and MMV 0 i r ~ r ~ r T T T 1------1---- resistance in the local composite, as O riginal FI F2 F3 BC1F1 BC1F2 BC1F3 BC2F1 BC2F2 BC2S1 shown by complete resistance in variety Transfer steps more than 80% of the plants. The Figure 5. Efficiency of MSV resistance transfers: variations in the overall susceptibility genetic factors that determine scores during different transfer steps. resistance to these viruses seem to differ, despite the fact that they occur in the same geographical zones and are transmitted by the same insect Origin of converted varieties. vector. IRAT 297 is currently the only Origin Mali Burkina Benin CIMMYT CIMMYT Côte IITA- known source of MStpV resistance. Faso d'Ivoire Nigeria MMV resistance is now defined Variety Tiémantié IRAT 171 CN7 Suwan 8331 Tuxpeño CPJ 16 SR Pool and plant resistance to vector transmission of the virus has also been detected in IRAT 297 hybrid lines. Studies on this latter type of resistance o 4 500 —i are now under way. These two levels of resistance, i.e. to the virus and to- transmission, should be jointly taken into account for developing efficient genetic control techniques.

Prospects In the short term, it is essential to make between-country comparisons, under artificial MSV infestation, of resistance levels in varieties that were selected for MSV-resistance in Réunion (final forms obtained by Tiémantié IRAT 171 CN 7 Suwan 8331 Tuxpeño 16 SR Pool crossing S3 lines, with two backcros­ Varieties sing cycles) and Togo. Such tests are being carried out in Cameroon, O riginal form H Resistant form (F3 of the starting cross) Togo, Zimbabwe with CIMMYT, and Réunion. The same type of test will Figure 6. Transfer efficiency. Gram yields of converted varieties compared to those then be carried in more countries of the original varieties (kg/ha). under natural infestation conditions.

Agriculture et développement ■ Special issue - December 1 995 6 7 The results should highlight the DONSON J., ACCOTTO G.P., mosaïque par leurs vecteurs Cicadulina extent of similarity — in terms of BOULTON M.I., MULLINEAUX P.M., m bila (Naude, 1 924) et Peregrinus recovery of agronomic traits, grain DAVIES J.W., 1 987. The Nucleotide maidis (Asmead, 1890) (Homoptera). types, resistance other than to MSV Sequence of a Geminivirus from Digitaria Thèse de docteur en sciences, sanguinalis. Virology, 161:160-169. université des sciences et techniques du — between resistant and susceptible Languedoc, Montpellier, France. 173 p. forms of different varieties. ETIENNE J., RAT B„ 1973. Le stripe : une maladie importante du maïs à la Réunion. REYNAUD B., PETERSCHMITT M., In the longer term, transfers of MStpV L'Agronomie Tropicale, 28 : 11 -17. 1992. Study of the mode of transmission of and MMV resistance are planned, maize streak virus by Cicadulina mbila FULLER C., 1901. Mealie variegation. especially in open pollination (Naude). Annals Applied Biology, 121 : In 1st Report of the government varieties and hybrid varieties; the 85-94. entomologist. Natal, 1 899-1900, resulting plants could then be South Africa, p. 1 7-19. RODIER A., in press. Breeding distributed widely throughout the maize lines for complete and partial GRAHAM C.L., 1979. Inability of tropics. Research is currently under resistance to maize streak virus (MSV). certain vectors in North America to way on P. maidis transmission, resis­ Euphytica, in press. transmit maize streak. Environment tance transfers into elite maize lines, Entomology, 8: 228-230. SOTO P.E., BUDDENHAGEN I.W., and on the genetics of resistance to ASNANI V.L., 1982. Development of HAINZELIN E., MARCHAND J.-L., 1986. these two viruses. The preliminary streak-resistant maize populations through Registration of IRAT 297 maize results are promising. ■ improved challenge and selection germplasm. Crop Science, 26: methods. Annals Applied Biology, 100: 839-840. 539-546. KONATE G., TRAORE O., 1992. Les STOREY H.H., 1936. Virus Disease of hôtes réservoirs de la striure du maïs Bibliography East African plants. IV. A survey of (MSV) en zone soudano-sahélienne : the viruses attacking the graminae. identification spatio-temporelle. East African Agricultural Journal., AUTREY L.J.C., 1983. Studies of maize Phytoprotection, 73 (3) : 111 -11 7. mosaic virus and other maize mosaic 1: 333-337. virus disease in the islands of the Western KONATE G., TRAORE O., in press. Variabilité et adaptation d'hôte du STOREY H.H., HOWLAND AK„ 1967 a. Indian Ocean. In D.T. GORDON, Inheritance of resistance in maize to the J.K. KNOKE, L.R. NAULT, R.M. RITTER. virus de la striure du maïs en zone soudano-sahélienne. Phytoprotection, 75, virus of streak disease in East Africa. Proceedings International Maize Virus Annals Applied Biology, 59 (3): 429-436. Disease Colloquium and Workshop, in press. August 2-6 1983. Ohio State University, KULKARNI H.Y, 1973. Comparison and STOREY H.H., HOWLAND A.K., 1967 Ohio Agrie. Res. and Dev. Center, characterisation of maize stripe and maize b. Transfer of resistance to the streak virus Wooster, USA. p. 167-181. lines virus. Annals Applied Biology, into East African maize. East African 75 (2) : 205-216. Horticultural and Forestry Journal, BOCK K.R., 1980. Maize viruses. In Crop 33: 131-135. virology research project July 1973-July LE CONTE, 1974. La virose du maïs 1980. Final report. Kenya agricultural au Dahomey. L'Agronomie Tropicale, VON WECHMAR M.B., HUGHES F.L., research institute, Muguga. p.1-6. XXIX (4) : 831-832. 1982. A rapid technique for typing streak virus isolates using a panel of BRIDDON R.W., LUNNESS P., PETERSCHMITT M., 1988. Identification differential hosts. In Proceedings of 9th CHAMBERLIN L.C.L., PINNER M.S., sérologique et dynamique du maize streak South Africa Maize Symposium, BRUNDISCH H., MARKHAM P.G., 1992. virus dans le maïs et dans le vecteur Petermaritzburg, 1990. p 115-119. The nucleotide sequence of an Cicadulina mbila. Thèse de docteur en infectious insect-transmissible clone of the sciences, Université de Paris-Sud, Orsay, WEBB M.D., 1987. Species recognition geminivirus Panicum streak virus. Journal France. 179 p. in Cicadulina leafhoppers (: of General Virology, 73: 1041-1047. Cicadellidae) vectors of pathogens of PETERSCHMITT M., REYNAUD B., Graminea. Bulletin of Entomology DAMSTGEET V.D., 1983. Maize streak SOMMERMEYER G., BAUDIN P., 1 991. Research, 77: 683-712. virus: I. Host range and vulnerability of Characterization of maize streak maize germplasm. Plant disease, 67: virus isolates using monoclonal 734-737. and polyclonal antibodies and by Abstract... Resumen... DELPUECH I., BONFILS J., LECLANT F„ transmission to a few hosts. Plant Disease, 1986. Contribution à l'étude des virus du 75 (1): 27-32. Résumé... maïs transmis par homoptères PETERSCHMITT M., QUIOT J.-B., auchenorrynques à l'île de la Réunion. REYNAUD B., BAUDIN P., 1 992. J.-L. MARCHAND, M. PETERSCHMITT, B. REYNAUD, Agronomie, 6 (6) : 549-554. Detection of maize streak virus antigens J. DINTINGER— Maize streak, maize stripe and maize mosaic virus diseases in the tropics (Africa DINTINGER J., RODIER A., REYNAUD B., over time in different parts of maize plants and islands in the Indian Ocean). CLERGET B., MARCHAND J .-L., of a sensitive and so-called tolerant in press. Breeding maize for MSV cultivar by Elisa. Annals Applied Biology, Impact of maize virus disease and current research; 121:641-653. resistance in Reunion Island. In 11th South vectors and their epidemiology; maize virus diagnosis and Africa Maize Breeding Symposium, REYNAUD B., 1988. Transmission des variability; maize resistance and breeding. The three virus Cerada, 15-1 7 March 1994. In press. virus de la striure, du stripe et de la diseases have been known since the beginning of the

0 8 Agriculture et développement ■ Special issue - December 1995 century and are found in the tropics. They were studied in es transmitido por el m aize streak virus (MSV), el stripe es tropicales ont été plus précisément étudiées en Afrique et à Africa and Reunion. Research in virology, epidemiology transmitido por el maize stripe virus (MStpV) y el mosaico la Réunion. Des recherches en virologie, en épidémiologie and genetics is carried out by CIRAD in collaboration with por el maize mosaic virus (MMV). Los tres virus son et en génétique sont conduites par le CIRAD en several African institutions (CORAF, INCV, INERA and IRA), transmitidos obligatoriamente por insectos, del género collaboration avec plusieurs organismes africains (CORAF, international institutions (CIMMYT and IITA) and the John Cicadulina para el MSV (se han identificado numerosas INCV, INERA, IRA), internationaux (CIMMYT, IITA) et le Innes Institute in Great Britain. Maize streak virus (MSV), especies) y por Peregrinus maidis para el MStpV y el maize stripe virus (MStpV) and maize mosaic virus (MMV) MMV. Dichos insectos se han identificado en el continente John Innes Institute de Grande-Bretagne La striure est are always spread by insects. MSV is spread by members africano. Las epidemias de estriado provocan grandes causée par le maize streak virus (MSV), le stripe est of the genus Cicadulina (numerous species have been daños (en la Reunión y, algunos años, en Africa), provoqué par le maize stripe virus (MStpV) et la mosaïque identified). MStpV and MMV are spread by Peregrinus mientras que el impacto de las virosis provocado por el par le maize mosaic virus (M M V ). Ces virus sont maidis. These insects have been identified in Africa. MstpV y el MMV todavía se conoce mal. La eficacia de las obligatoirement transmis par des insectes, du genre Streak epidemics cause serious crop damage (in Reunion tronsmiciones virales al insecto vector es elevada para el Cicadulina pour le MSV (de nombreuses espèces sont and in Africa in certain years) but the impact of strike and MSV y C. mbila (50% de los insectos infecciosos) y más identifiées) et par Peregrinus maidis pour le MStpV et le mosaic is still not well known. The effectiveness of virus baja para el MStpV y el MMV y P. maidis (20% de los transmission to the insect vector was high for MSV and insectos infecciosos). Los síntomas de los 1res virus están MMV. Ces insectes ont été identifiés sur le continent C. mbila (50% of insects were infectious) and lower for descritos, y la epidemiologia está estudiada en Burkina y africain. Les épidémies de striure provoquent des dégâts MStpV and MMV and P. m aidis (20% of insects were en la Reunión. Los virus son identificados mediante importants (à la Réunion et certaines années en Afrique) ; infectious). The viruses were identified by enzyme-linked pruebas inmunoenzimáticas ELISA (Enzyme-linked l'impact des viroses causées par le MStpV et le MMV est immunosorbent assay (ELISA). The serological method was immunosorbent assay). El método serológico es más encore mal connu. L'efficacité des transmissions virales par the most reliable. A diagnosis kit was developed to test seguro que sólo la descripción de los sintomas. Se ha l'insecte vecteur est élevée pour le MSV et C. mbila (50 % the presence of the three viruses without special elaborado un kit de diagnóstico para probar la presencia des insectes infectieux), et plus faible pour le MStpV et le equipment. With the new techniques (monoclonal isolates, de estos tres virus sin necesitar un equipo de laboratorio. range of hosts and genome sequencing) used, MSV Otras técnicas utilizadas (gírmenes aislados monodonales, MM V par P. m aidis (20 % d'insectes infectieux). Les serotypes were identified. Detection of the virus in plants numerosos huéspedes, sequenciación del genoma), se han symptômes des trois virus sont décrits, et l'épidémiologie using ELISA made it possible to understand the mechanism distinguido serotipos de MSV. La investigación de virus en est étudiée au Burkina et à la Réunion. Les virus sont of resistance to MSV. Varietal resistance appeared to be la planta mediante la técnica ELISA ha permitido identifiés à l'aide des tests immunoenzymatiques ELISA caused by resistance to propagation of the virus. The comprender el mecanismo de resistencia al MSV. La lEnzyme-linked Immunosorbent assay). La méthode varietal selection work carried out in Reunion and in Togo resistencia varietal al virus se manifiesta aparentemente sérologique est la seule méthode fiable. Un kit de was aimed at transferring resistance (found in local por una resistencia a la multiplicación viral. Las diagnostic a été mis au point pour tester la présence de ces varieties) to varieties of clear agronomic interest. A investigaciones en selección varietal, tuvieron como "Composite viroses resistant" was form ed from local objetivo realizar transferencias de las resistencias, trois virus, sans nécessiter un équipement de laboratoire. varieties and then improved. The work on MSV is more presentes en variedades locales, a variedades de buen D'autres techniques (isolais monodonaux, gamme advanced than that on MStpV and MMV. Genetic analysis interés agronómico, en la Reunión y en Togo. A partir de d'hôtes, séquençage du génome) ont permis de distinguer of resistance to MSV revealed partial polygenic resistance variedades locales resistentes, se constituyó, y mejoró, un des souches différentes de MSV. La recherche du virus dans and total resistance by virtue of two or three genes. "Composite viroses résistant". Los trabajos relativos al la plante par la technique ELISA a permis de comprendre Research has been developped on the three viruses MSV están más avanzados que los relativos al MStpV y al le mécanisme de résistance de la plante au MSV, qui se resistance in Reunion, five varieties have been completed. MMV. El análisis genético de la resistencia al MSV Transfers have been carried out for the MSV resistance in demuestra que existiria un sistema mayor de resistencia traduirait par une résistance à la multiplication virale. Togo, five varieties have been formed. Trials are in total oligogenica (3 factores geneticos) y tal vez una Les sélections variétales, ont eu pour objectif de transférer progress in several countries (Cameroon, Togo, Zimbabwe resistencia parcial poligenica, la qual implica genes des résistances dans des variétés d'un bon intérêt and Reunion) to compare the levels of resistance achieved minores. Se están realizando transferencias en variedades agronomique, à la Réunion et au Togo. A partir de variétés and the agronomic conformity of the resistant varieties. africanas y efectuando investigaciones en la Reunión y en locales, un « Composite viroses résistant » a été constitué, Togo. Se han efectuado las investigaciones en los tres Key words: maize, streak, stripe, mosaic, virus, insect, puis amélioré. Les travaux sur le MSV sont plus avancés Cicadulina mbila, Peregrinus maidis, varieties, resistance, virus en la Reunión, cinco variedades se han terminado. que ceux sur le MStpV et le MMV. L'analyse génétique de ELISA, tropics, Africa, Reunion. Se han realizado transferencias para la resistencia al MSV en Togo, y se hon constituido cinco variedades. También la résistance au MSV montre qu’il existerait un système J.-L. MARCHAND, M. PETERSCHMITT, B. REYNAUD, se están haciendo pruebas en diferentes países (Camerún, majeur de résistance totale oligogénique (3 facteurs J. DINTINGER — Las virosis del estriada, del stripe Togo, Zimbabwe, la Reunión). génétiques) et éventuellement une résistance partielle y del mosaico en el maíz en región tropical (Africa Palabras clave: maíz, estriado, stripe, mosaico, virus, polygénique impliquant des gènes mineurs. Des transferts y las islas del Océano Indico). insecto, Cicadulina mbila, Peregrinus maidis, variedad, sont réalisés sur des variétés africaines ou introduites à la El impacto de las virosis y las investigaciones actuales; resistencia, técnica ELISA, región tropical, Africa, Reunión. Réunion et au Togo. A la Réunion, les travaux de sélection los vectores y su epidemiología; el diagnóstico de las sont conduits sur les trois virus, cinq variétés sont virosis y la variabilidad del virus MSI/; la resistencia del J.-L. MARCHAND, M. PETERSCHMITT, B. REYNAUD, converties. Au Togo, des transferts sont réalisés pour la m aíz y la selección varietal. Estos tres virosis, conocidas J. DINTINGER — Les viroses de la striure, du stripe résistance au MSV et cinq variétés sont converties. Un essai desde comienzos del siglo y difundidas en todas las et de la mosaïque sur le maïs en région tropicale regiones tropicales, han sido estudiadas más precisamente (Afrique et îles de l'océan Indien). multilocal est en place dans différents pays (Cameroun, en Africa y la isla de la Reunión. El CIRAD ha llevado a L'impact des viroses et les recherches actuelles ; les Togo, Zimbabwe, la Réunion). cabo investigaciones de virologia, epidemiología y vecteurs et leur épidémiologie ; le diagnostic des viroses et Mots-clés : maïs, striure, stripe, mosaïque, virus, insecte, genética, en colaboración con varios organismos africanos la variabilité du virus de la striure; la résistance du maïs et Cicadulina mbila, Peregrinus maidis, variété, résistance, (CORAF, INCV, INERA, IRA), internacionales (CIMMYT, la sélection variétale. Ces trois viroses, connues depuis le UTA) y el John Innes Institute de Gran Bretaña. El estriado début du siècle et répandues dans toutes les régions technique ELISA, région tropicale, Afrique, la Réunion.

Agriculture et développement ■ Special issue - December 1 995