Abstract:
Seed-borne fungi of sorghum and pearl millet in Burkina Faso were surveyed. A total of 188 seed samples from various locations, collected in 1989 (42) and 2002 (146), were tested, using the blotter, dry inspection and washing methods. Infection experiments were carried out with the major fungi recorded on each crop by the blotter test. Six essential oils of plants were investigated for their inhibitory activity against eight pathogenic fungi. Thirty four and 27 fungal species were found in seed samples of sorghum and pearl millet, respectively. Phoma sp. and Fusarium moniliforme infected 95 to 100% of the seed samples of both sorghum and pearl millet. Sphacelotheca sorghi and Tolyposporium ehrenbergii were encountered in respectively, 75 and 33% of seed samples of sorghum. T. penicillariae, Sclerospora graminicola and Claviceps fusiformis were present in 88, 41 and 32% of seed samples of pearl millet, respectively. Seeds inoculated with Acremonium strictum, Curvularia oryzae, F. equiseti, F. moniliforme and F. subglutinans and sown in sterilized soil, showed considerable mortality of the seedlings. Three essential oils inhibited in vitro the mycelial growth of all the fungi used by 85 to 100% and reduced significantly sorghum and pearl millet seed infection rates of Phoma sp., Fusarium sp., Curvularia sp., Colletotrichum graminicola and Exserohilum sp. Presence of many pathogenic fungi in considerable number of seed samples indicates the need of field surveys for these and other pathogens. Development of plant extracts for the control of seed-borne pathogens and public awareness on seed-borne diseases management measures for maintaining quality seed should be increased.
INTRODUCTION
Sorghum (Sorghum bicolor (L.) Moench) and pearl millet (Pennisetum glaucum (L.) R. Br.) are the two major cereal of Burkina Faso. Annually grown on 2,641,089 hectares, representing 88% of the cereal areas, these two crops produced 2,100,046 tons of grain, corresponding to 79% of the total national cereal production (DSA/MAHRH, 2005). Sorghum and pearl millet are of vital importance because 70 and 95% of their respective production are consumed by populations in the main production regions (FAO and ICRISAT, 1997). Despite the socio-economic importance of these crops, their productivity has remained very low due to various constraints. The average yields recorded during the last ten years are 830 kg ha-1 for sorghum and 670 kg ha-1 for pearl millet (DSA/MAHRH, 2005; FAO, 2006). These yields are by far low from the world yields of sorghum and pearl millet which are 1332 and 816 kg ha-1, respectively (FAO, 2006). Amongst the biotic constraints to sorghum and pearl millet production in Burkina Faso, seed-borne fungal diseases are of great importance (Neya and Kabore, 1987; Sereme, 1995; Mathur and Manandhar, 2003). In Burkina Faso, sorghum and pearl millet seeds used by farmers are mainly from their own-saved seed lots; the only criterion considered by the farmers when selecting seeds is the physical appearance of the grain. Earlier investigations on sorghum seeds produced in different research stations in Burkina Faso (Kamboinse, Farako-ba, Kouare, Di, Niangoloko) and two localities (Nakamtinga, Boni), revealed that these seeds were infected by important fungal pathogens such as Colletotrichum graminicola (Ces.) Wilson, the causal agent of anthracnose, Gloeocercospora sorghi (zonate leaf spots), Phoma sp. and Fusarium sp. (seedlings blights) (Kabore et Couture, 1983; SOMIMA, 1994), but very little information is available on the occurrence of fungi on farmers`own saved seeds from various producing places. Therefore, a country-wide survey is necessary to identify the kind and amount of mycoflora associated with sorghum and pearl millet seeds in Burkina Faso and to assess their distribution in different ecological zones and their significance on seed and plant health. Breeding for genetic resistance and use of chemicals have been the most effective strategies recommended to control sorghum and pearl millet diseases (DPVC, 1995). But, improved varieties and chemicals are not always available or are often costly for the small-scale farmers; and the use of natural products as alternatives for synthetic pesticides is still limited. Furthermore, a repetitive use of synthetic pesticides can induce genetic resistance in target organisms, as resistance of Ustillago nuda (wheat smut) to carboxine, previously reported by Leroux (1986, 1987). In this context, development of simple and eco-friendly seed-borne diseases management methods is necessary to improve the quality of seed in general and farmers saved-seed in particular. In a recent study, Somda et al. (2007) demonstrated the efficacy of aqueous extracts of Cymbopogon citratus (DC) Stapf. for controlling sorghum seed infections by C. graminicola and P. sorghina. Many studies carried out in Burkina Faso underlined insecticidal properties of some plant species including Cymbopogon, Hyptis and Ocimum (Koumaglo et al., 1998; Djibo, 2000; Nebie, 2006). But information related to the role of such local plants in controlling crop pathogen agents is still limited.
The objectives of the study are to inventory fungi in farmers saved-seed samples of sorghum and pearl millet collected from various localities of Burkina Faso, to evaluate the effect of seed-borne inoculum of the main fungi recorded on seedlings emergence, seedlings mortality and diseases transmission and to investigate extracts from plants widely distributed in Swahilian and sudanian agro ecological zones, for their potential in improving seed quality and planting value of sorghum and pearl millet.
MATERIALS AND METHODS
Seed samples: A total of 86 seed samples of sorghum and 102 seed samples of pearl millet, weighted 500 to 1000 g each, were collected in 43 locations situated in the north-sudanian, south-sudanian and Swahilian zones of Burkina Faso (Fig. 1). Among the samples, 29 and 13 seed samples
of sorghum and pearl millet, respectively, were collected and tested in 1989 and data on eight major fungi recorded are analysed in the present study. Fifty seven and eighty nine seed samples of sorghum and pearl millet, respectively, were collected in 2002 and analysed in 2002 and 2003. The seed samples were packed in cotton or plastic bags and stored in a freezer at the Laboratory of Phytopathology of Kamboinse research station in Burkina Faso, or, in a storage room at 5°C at the Danish Seed Health Centre for Developing Countries (DSHC) in Denmark, until use. Seventy six and 112 seed samples were tested, respectively at DSHC and at Kamboinse research station.
Essential oils: Aerial parts of Cymbopogon schoenanthus (L.) Spreng., Hyptis spicigera Lam., Lantana camara L. and Ocimum americanum L., collected at Kamboinse (in the Centre of Burkina Faso) and C. nardus Chiov. and C. giganteus Chiov., collected, respectively at Porto-Novo (in the north of Benin) and Koupela (in the East of Burkina Faso), were air-dried for four days before oil isolation. The essential oils were extracted from the plant materials by the hydro distillation method using a Clevenger-type apparatus (Lamaty et al., 1987). The oils were separated from the water by decantation, dried over anhydrous sulphate and stored in darkness at 4°C.
Seed health testing methods: Three conventional seed health testing methods, reported by Mathur and Kongsdal (2003), were used to detect fungi associated with seeds:
Dry inspection method was used in this study to detect sclerotia of ergot in the seed samples. Fifteen and 90 g of seeds from each sample of pearl millet and sorghum, respectively, were visually inspected using a magnifying lamp. The sclerotia were selected from seeds, weighed and the result was expressed in percentage.
The standard blotter method was used to detect a wide range of fungi which are able to arise easily from seeds in presence of humidity. Four hundred untreated pure seeds from each sample were plated on moisten blotters in plastic Petri dishes at the rate of 25 seeds per dish and incubated for 7 days at 20-25°C under alternating cycles of 12 h near Ultraviolet Light (NUV) and 12 h darkness. Individual seeds were examined for the presence or absence of fungi under a stereomicroscope and identification was confirmed by examining mycelium and/or conidia under a compound microscope. The fungal species present on each seed were recorded and the percentage incidence of each fungus per sample was computed.
The washing test was used to detect teliospores of S. cruenta, S. reiliana, S. sorghi and T. ehrenbergii (causal agents of sorghum smuts), T. penicillariae (pearl millet smut) and oospores of Peronosclerospora sorghi and Sclerospora graminicola (causal agents of sorghum and pearl millet downy mildews), present on the surface of seeds. Sorghum seeds (5 g) and pearl millet seeds (10 g) were shaken in 25 mL of distilled water. The suspension was centrifuged at 3000 rpm for 5 min. The sediment was suspended in 1 mL of distilled water and examined under compound microscope. Teliospores of smuts and oospores of downy mildews were counted using a haemocytometer (type Fuchs-Rosenthal) and the total number of spores was calculated using the following formula (Mathur and Kongsdal, 2003):
with : 1 mm 3 =(1/1000) mL, so the formula can be written as follow:
Testing effect of major fungi on seedling emergence and mortality and evaluation of seed-to-seedling transmission: The effect of 17 and 15 fungi isolated, respectively from incubated seeds of sorghum and pearl millet was tested on seedling emergence and seedling mortality. One hundred and fifty seeds of sorghum and pearl millet previously disinfected in 70% ethanol for 5 min were inoculated by rolling on sporulating cultures of the different fungi, before sowing in pots (25 seeds per pot) containing sterilized soil, following the procedure used by Mathur et al. (1973). The pots were kept in a growing room for 10 days under 12 h fluorescent light/12 h darkness at 25-29°C. Untreated seeds, seeds disinfected with ethanol and seeds treated with a chemical called calthio (20% of Lindane, 25% of Thirame) were used as controls. For each crop, a randomized complete block design with three replicates of 50 seeds was used. Ten days after sowing, seedlings emergence and seedlings mortality were evaluated and percentage of emerged seedlings and percentage of dead seedling were calculated. To evaluate seed-to-seedling transmission of the fungi, ten seedlings from each treatment were cut at the level of the coleoptiles, disinfected in 70% ethanol for 2 min and plated on moistened blotter papers in plastic box for 5 days. Infected plants by the target fungus were counted using a stereomicroscope and the results expressed in percentage. The severity of infection (the capacity of the fungus to propagate inside the seedling) was estimated on the incubated seedlings by assigning score based on the presence or absence of the fungus in plant parts: score 1 is attributed to healthy plants; score 2 to slightly infected plants (fungus present on plant stem); score 3 to highly infected plants (fungus present on plant stem and/or leaves). An index of severity was calculated following the formula used by Williams and Singh (1981) and the result was expressed in percentage:
| Where: | ||
| xi | = | Note attributed to each plant from the class i |
| ni | = | Number of plants from the class i |
| E(xi) | = | Range of the scale of notation (3) |
| N | = | Total number of observed seedlings (10) |
| S | = | Severity of infection or capacity of the
fungus to invade the plant (%). Percentages of severity of infection
were transformed into Arcsine values before performing the statistical
analysis |
Effect of essential oils on the radial growth in vitro of eight seed-borne fungi: Different PDA growth media containing 0.2% of essential oils (v/v) were prepared for growing the tested fungi. The oils were filtered with Millipore filters (Ø = 0.45 μm) for sterilization and mixed to autoclaved PDA and distributed in Petri dishes. Non treated PDA and PDA mixed with calthio at the rate of 0.25 g for 100 mL PDA, were considered as controls.
The pathogenic fungi selected by the previous tests were grown on PDA. For each fungus, a fungal disc measuring 4 mm diameter from a 5-day-old culture, was placed in a Petri dish containing the tested growth medium. Five replicates were used for each kind of growth medium. The Petri dishes were incubated under laboratory conditions (25-30°C) and two diameters perpendicular to each other of the colony were measured 4 and 6 days after incubation. The results were expressed in percent of mycelial inhibition calculated following the formula used by Singh et al. (2003):
Where, dc and dt are average diameters of the mycelial colonies in the control and treated media, respectively.
Effect of essential oils on sorghum and pearl millet seed contamination by five fungi: The essential oils that showed potent growth inhibitory activity in vitro were tested for efficacy against Colletotrichum graminicola, Curvularia sp., Exserohilum sp., Fusarium sp. and Phoma sp., in naturally infected seeds of sorghum and pearl millet. Four doses: 5, 7.5, 10 and 15 μL of essential oil per gram of seeds were used. Before applying the oils, the seeds were mixed with 200 μL of distilled water in order to soften their teguments and allow essential oils absorption. The seeds were maintained hermetically closed in flasks for about 20 h (overnight) and evaluated for health using the standard blotter method described above. Untreated seeds, seeds mixed with distilled water and seeds treated with calthio at the rate of 2.5 g kg-1 of seeds, were used as controls. One hundred and fifty seeds were used for each treatment.
Data analyses: The main seed-borne fungi of sorghum and pearl millet, the effect of fungi on seedling emergence and seedling mortality, the main pathogenic and seed-transmitted fungi, the effect of essential oils on fungal growth and on seed infection were determined by Analyses of Variance (ANOVA) using a completely randomised design. The significance (p<0.05) of differences between treatments were determined, using the Duncan`s Multiple Range (DMR) test of Statistical Analysis System, version 8.
RESULTS
Sorghum seed mycoflora: A total of thirty four fungal species belonging to seventeen genera (Alternaria Nees, Bipolaris Shoem. (syn. Drechslera fide Ellis), Botryodiplodia (Sacc.) Sacc., Cercospora Fres., Colletotrichum Corda, Curvularia Boedijn, Drechslera Ito, Exserohilum Leonard and Suggs., Fusarium Link, Gloeocercospora Bain and Edgerton ex Deighton, Macrophomina Petrak, Myrothecium Tode, Nigrospora Zimm., Peronosclerospora (Ito) Shirai and K. Hara, Phoma Sacc., Sphacelotheca de Bary, Tolyposporium Woronin ex Schroter) were detected in sorghum seed samples (Table 1). All the seed samples tested in 1989 and 2002/2003 were infected by Phoma sp and F. moniliforme, respectively. F. moniliforme, C. lunata, C. graminicola, F. pallidoroseum and Drechslera sp. were found in 59 to 93% of the samples in 1989, while Phoma sp. and Curvularia lunata were detected in 96.5 and 93% of the samples, respectively, in 2002/2003. Sphacelotheca sorghi and Colletotrichum graminicola, causing important diseases on sorghum, were encountered in, respectively 75.4 and 24.6% of the samples tested. Fifteen fungi occurred in few number of samples (1.7-5.3% of the samples).
All the seed samples tested using dry inspection were free of sclerotia of Claviceps sorghi, the causal agent of sorghum ergot.
| Table 1: | Fungal species
detected in 86 seed samples of sorghum from Burkina Faso in 1989
and 2002/2003, by dry inspection, washing of seeds and blotter
tests methods |
| a: No. in the parentheses represent the number of seed samples tested, b: No. of spores per gram of seed | |
In the three ecological zones, Phoma sp., F. moniliforme, C. lunata and S. sorghi encountered in 72 to 100% of the samples were the widest spread fungi (Table 3). All of them, excepted S. sorghi, occurred more frequently in the north and south-sudanian zones than in the Swahilian zone. Among the major fungal species recorded in sorghum seeds five were not encountered in the Swahilian zone. In addition, seed samples from the Swahilian zone had the lowest rates of infection compared to those from the north and south-sudanian zones.
Pearl millet seed mycoflora: Twenty seven fungal species were associated with seed samples of pearl millet (Table 2). In 1989, F. moniliforme, Phoma sp. and Drechslera sp. were associated to all the samples, at high levels of infection. F. equiseti and F. pallidoroseum were also encountered frequently. Among the fungi recorded in 2002/2003, Phoma sp., F. moniliforme, T. penicillariae and C. lunata were encountered, respectively in 100, 95.4, 88.2 and 82.9% of the samples (Table 2). Other fungi with high frequency of dispersal included C. pallescens, E. rostratum, S. graminicola and C. fusiformis (Table 2).
T. penicillariae, F. moniliforme and A. strictum occurred at high levels in seeds from Swahilian zone. High infection levels of Phoma sp. ranging from 33.70-34.85% infection were recorded on seed samples from the three agro ecological zones (Table 4). The widest spread fungi in the three zones were Phoma sp., F. moniliforme, T. penicillariae and C. lunata with 66 to 100% of seed samples infected.
During examination of incubated seeds of sorghum and pearl millet, a number of saprophytic fungi, not reported in the tables, were present in the seed samples. These included mainly Aspergillus niger Tiegh., A. flavus Link, Cladosporium sp., Penicillium sp. and Rhizopus sp.
| Table 2: | Fungal species
detected in 102 seed samples of pearl millet from Burkina Faso
in 1989 and 2002/2003, by dry inspection, washing of seeds and
blotter tests methods |
| a: No. in the parentheses represent the number of seed samples tested, b: Percentage of sclerotia by weight of seed, c: No. of spores per gram of seed | |
| Table 3: | Major seed-borne fungi recorded in 86 seed samples of sorghum grown in three agro ecological zones in Burkina Faso in 1989 and 2002 |
Psp. : Phoma
sp.; Fm: Fusarium moniliforme; Cl: Curvularia lunata;
Cg: Colletotrichum graminicola; Cp: Curvularia pallescens;
Fs: Fusarium solani; Gloe: Gloeocercospora sorghi;
Er: Exserohilum rostratum; Fpalli: Fusarium pallidoroseum;
Fe: Fusarium equiseti; Ce: Curvularia eragrostidis;
Aalt: Alternaria alternata; Along: Alternaria longissima;
Fsub: Fusarium subglutinans; Ssorg: Sphacelotheca sorghi,
Means followed by the same letter within the same column are not
significantly different at level 5%, ( ): No. of infected seed
samples (%), [ ]: No. of tested seed samples |
|
Effect of seed-borne inoculum on seedling emergence, seedling mortality and disease transmission:
On pearl millet, only F. solani caused a negative influence on seedlings emergence compared to the chemical control (Table 6). High numbers of dead plants were observed with F. solani (19.3%), A. strictum (18.7%)
| Table 4: | Major seed-borne
fungi recorded in 101seed samples of pearl millet grown in three
agro ecological zones in Burkina Faso in 1989 and 2002 |
| Psp.
: Phoma sp.; Fm: Fusarium moniliforme; Cl: Curvularia
lunata; Bm: Bipolaris maydis; Cp: Curvularia pallescens;
Fs: Fusarium solani; As: Acremonium strictum; Er:
Exserohilum rostratum; Fpalli: Fusarium pallidoroseum;
Fe: Fusarium equiseti; Ce: Curvularia eragrostidis;
Fsub: Fusarium subglutinans; Sgram: Sclerospora graminicola;
Cfusi: Claviceps fusiformis; Tpenic: Tolyposporium penicillariae,
*: Means followed by the same letter within the same column are
not significantly different at level 5%, ( ): No. of infected
seed samples (%), [ ]: No. of tested seed samples |
|
| Table 5: | Seedlings emergence, seedlings mortality, disease transmission and severity of infection recorded after sowing inoculated seed of sorghum by different fungal species |
Means followed
by the same letter within the same column are not significantly
different at level 5%, No. in the parentheses represent the percentages
of severity of infection turned into Arcsines values on which
statistical analysis was performed |
|
and F. moniliforme (18%). Results on disease transmission revealed that F. solani was the most seed-transmitted fungus (100% transmission rate). Other fungi with high levels of seed-transmittance included F. moniliforme, A. strictum, C. eragrostidis and C. lunata. The highest index of severity (50%) was recorded with F. solani while the lowest was obtained with P. sorghina (4.4%). A. strictum (44.4%) and F. moniliforme (43.3%) infected severely the seedlings.
Inhibitory effect of essential oils on the mycelial growth of eight major seed-borne fungi: Essential oils of C. schoenanthus, C. nardus and C. giganteus reduced the mycelial growth of all the fungi at the rates of 86-100% and 85-100%, respectively, at 4 and 6 days after incubation (DAI) (Table 7). The percent mycelial inhibition obtained with these oils on F. solani, C. lunata, E. rostratum and P. sorghina were similar to those obtained with the chemical control. In addition these essential oils reduced better the mycelial growth of F. moniliforme, C. graminicola, B. sorokiniana and A. strictum than the chemical control. Essential oil of C. nardus completely inhibited the mycelial growth of all the fungi tested both at 4 and 6 DAI. Percent of growth inhibition due to L. camara, O. americanum and H. spicigera were relatively low (less than 50%). The growth of P. sorghina, F. solani, A. strictum and B. sorokiniana was favoured by essential of L. camara (negative values of percent of mycelial inhibition).
| Table 6: | Seedlings emergence, seedlings mortality, disease transmission and severity of infection recorded after sowing inoculated seed of pearl millet by different fungal species |
| Means followed by the same letter within the same column are not significantly different at level 5%, No. in the parentheses represent the percentages of severity of infection turned into Arcsines values on which statistical analysis was performed | |
| Table 7: | Inhibitory effect
of different essential oils on the radial growth in vitro
of eight seed-borne fungi at 4 and 6 Days after Incubation (DAI) |
CC = Chemical
Control; Cg = Cymbopogon giganteus; Cn = Cymbopogon
nardus; Cs = Cymbopogon schoenanthus; Hs = Hyptis
spicigera; Oa = Ocimum americanum; Lc = Lantana
camara; C = Control (untreated seeds), Means followed by the
same letter within the same column are not significantly different
at level 5% |
|
Efficacy of essential oils in controlling seed-borne fungi of sorghum and pearl millet: A seed health testing was performed after the treatment of the seeds by essential oils of C. nardus, C. giganteus and C. schoenanthus and percent of infected seeds by Phoma sp., Fusarium sp., Curvularia sp., Colletotrichum graminicola and Exserohilum sp. were recorded. The results presented in Table 8 showed that all the essential oils reduced significantly sorghum and pearl millet seed contamination by the selected fungi. The lowest rates of infected seeds
| Table 8: | Effect of treatment
by essential oils of C. giganteus, C. schoenanthus
and C. nardus at different doses on the percent
of infected seeds by five fungi involved in sorghum and pearl
millet seedlings infections in Burkina Faso |
C = Untreated
seeds; WC = Water Control (seeds mixed with distilled water);
CC = Chemical control (seeds treated with chemical); D1 = 5 μL
of essential oil per g of seeds; D2 = 7.5 μL of essential
oil per g of seeds; D3 = 10 μL of essential oil per g of
seeds; D4 = 15 μL of essential oil per g of seeds |
|
were generally recorded on seeds treated with 10 μL and/or 15 μL of essential per g of seeds. In most of the time, these doses were most efficient or so efficient as the chemical control. Oil of C. giganteus used at 15 μL g-1 of seeds eliminated completely pearl millet seed contamination by all the fungi tested. However, the dose 15 μL of essential oil g-1 strongly reduced seed germination in Petri dishes.
DISCUSSION
Good seed is recognized as an important input in any agricultural production system. One of the important aspects of good seed, besides high germination and purity, is the absence of seed-borne pathogens. At least 27 fungal species were encountered in high frequencies and infection percentages in different samples of sorghum and pearl millet collected from farmers own saved-seed in Burkina Faso (Table 1, 2). Phoma sp., F. moniliforme, C. lunata, S. sorghi, T. penicillariae, E. rostratum, S. graminicola and C. graminicola were the main fungi occurring frequently in sorghum and/or pearl millet seeds. Out of the fungi, S. sorghi, S. reilana, T. ehrenbergii, T. penicillariae, S. graminicola, important pathogens whose inoculum is present on the surface of seeds, were recorded for the first time in farmers saved-seeds of sorghum and pearl millet from Burkina Faso. About ten fungal species belonging to the genera Fusarium, Phoma, Curvularia and Exserohilum heavily infected both sorghum and pearl millet seeds. In previous surveys, Mathur et al. (1975) and Tarp et al. (1987) also found high frequencies of occurrence of Phoma sp., F. moniliforme and C. lunata in sorghum seeds from India and Mozambique. Beyond seed rots and seedling blights induced by these fungi, their presence at high levels in seeds is alarming since the genera Fusarium and Phoma are known to produce mycotoxins (CAST, 1989; Mathur and Manandhar, 2003).
Anthracnose and pearl millet downy mildew were frequently reported in the literature to be the most widespread diseases on sorghum and pearl millet in many parts of the world (ICRISAT, 1980; Singh, 1995) and in Burkina Faso (Kabore and Couture, 1983; Sereme, 1995). The present survey indicated that sorghum covered smut and pearl millet smut caused by S. sorghi and T. penicillariae, respectively, were widely scattered since their seed-borne inoculum were encountered more frequently in the samples than those of anthracnose (C. graminicola) and downy mildew (S. graminicola). Even if the role of secondary inoculum in anthracnose and downy mildew spread in field is of great importance (Warren, 1986; Singh and Williams, 1980), field surveys will be needed to complete the seed health surveys and to follow the evolution of the important diseases caused by these fungi. Other studies will also be necessary to establish the relationship between the amount of seed-borne inoculum of important pathogens such as S. sorghi, C. graminicola and S. graminicola in sorghum and pearl millet and the severity of the diseases its can produce in fields.
Occurring of all the major fungi in both sorghum and pearl millet seeds from the north-sudanian zone could be related to the large number of seed samples coming from this agro ecological zone. In addition, the sites of collection of the samples situated in the north-sudanian zone were more various than in the two other zones. In an other hand, the climate, drier in the Swahilian zone than the north and south-sudanian zones could justify the low representation of fungal species in the Swahilian zone. That was illustrated by, for example, the occurrence of C. graminicola, G. sorghi, F. solani and F. pallidoroseum in sorghum seeds from the north and south-sudanian zones and their absence in the seeds from the Swahilian zone.
According to the infection experiments carried out with the main fungi recorded by the blotter test, none of these fungi, except F. solani on pearl millet, had significant effect on sorghum and pearl millet seedlings emergence. These results were not in accordance with those previously reported by Mathur et al. (1973). These researchers observed reduction in germination rate of pearl millet by inoculating seeds with C. lunata, E. rostratum, B. spicifera and F. equiseti. Testing for germination of naturally infected seeds and seeds infected artificially at different dates could provide more information on the effect of these fungi on seedling establishment. Many fungi including F. moniliforme, F. equiseti, F. subglutinans, C. oryzae, A. strictum, F. solani, C. pallescens, E. rostratum and C. lunata, were frequently transmitted to sorghum and pearl millet seedlings through infected seeds. Among the seed-transmitted fungi, F. moniliforme, F. equiseti, F. subglutinans, C. oryzae, A. strictum and F. solani, caused high rates of dead seedlings. These results indicated that these seed-transmitted fungi could be the main fungal pathogens involved in sorghum and pearl millet seedlings diseases in Burkina Faso. In spite Phoma sp. being identified as the most frequent fungus recorded in the seeds, it did not appear to be so pathogenic (Table 6, 7). However, considering the high infection levels of Phoma sp. encountered in the seeds, further studies are necessary to elucidate the exact role of this fungus in seed. In contrast to Phoma sorghina, fungi as F. equiseti, F. solani, C. oryzae and A. strictum, occurring in low frequencies in seeds, were found to be very pathogenic to seedlings. Their presence in seeds, even in low levels, could be prejudicial to the seed lot. The main fungal pathogens involved in sorghum and pearl millet seedling diseases in Burkina Faso included the genera Acremonium, Curvularia, Fusarium and Exserohilum. Occurrence of these pathogens in many samples suggested that the pathogens were widely distributed in Burkina Faso.
Among the essential oils tested against the fungi, those extracted from C. giganteus, C. nardus and C. schoenanthus exhibited anti fungal activity by inhibiting in vitro, the mycelial growth of the tested fungi. In previous studies, Dawson-Andoh et al. (2000) observed an inhibitory effect of essential oil of C. nardus on Alternaria alternata and Aureobastum pullulans, two pathogenic fungal on wood. The Japan International Research Center for Agricultural Science (JIRCAS) (2005) also reported that oil of C. nardus completely inhibited the growth of food stocks fungi such as Aspergillus candidus, A. flavus, A. versicolor, Eurotium amstelodami, E. chevalieri, Penicillium adametzii, P. citrinum, P. griseofulvum and P. islandicum. By reducing in vivo, the seed contamination by the major fungi, the essential oils of C. giganteus, C. nardus and C. schoenanthus, confirmed their anti fungal properties and seemed to be the most potent extracts usable in seed-borne fungi management.
CONCLUSIONS
The presence of so many pathogenic fungi at high levels in farmers saved-seed from various geographical areas indicates a clear need for field surveys for these and other pathogens. There also is a clear need to increase public awareness on aspects related to seed health and to develop suitable management practices for improving the quality of the seeds. Testing seed health of major crops should be introduced in the national seed quality control system.
Plant species such as C. giganteus, C. nardus and C. schoenanthus, potent in controlling cereal crop seed-borne fungi, are available in all over tropical zone. The cost of intensive use of their essential oils should be relatively low and affordable to poor communities in many sorghum and pearl millet producing areas. Nevertheless, efficacy of these essential oils should be evaluated under field conditions. Moreover, the active constituents should be determined and quantified and the mammal toxicity tested.
ACKNOWLEDGMENTS
The authors are grateful to the Danish Seed Health Centre for Developing Countries (DSHC) for providing laboratory equipment, technical assistance and funds for research. The authors are very thankful to the International Foundation for Science for providing financial support and complementary equipment as a research grant given to Mrs Zoma-Zida P. Elisabeth. They wish to acknowledge Dr. Jan Torp (DSHC, Denmark) and Dr. Hamidou Traore (INERA,Burkina Faso) for going through the manuscript and giving us their valuable comments and suggestions and Dr. Nebie Roger (IRSAT, Burkina Faso) for extracting essential oils.