Fusarium oxysporum f.sp. elaeidis (fusarium wilt of oil palm)

An African isolate pathogenic to oil palm was identified as F. oxysporum var. redolens (Wollenw.) Gordon due to its cultural morphology. This variety is now widely regarded as a taxonomic synonym of F. oxysporum (Brayford, 1992).

Culture pigmentation is white, peach, salmon, vinaceous grey to purple to violet on media pH 6.5-7; mycelium is striate, felted to floccose. Microconidia, always present, are unicellular or bicellular, ellipsoidal, cylindrical, straight or curved (5-12 x 2.2-3.5 µm) and borne on lateral phialides or on phialides produced from short lateral conidiophores. Macroconidia are falcate, of the 'elegans' type in F. oxysporum but tending towards the 'Martiella' type in F. oxysporum var. redolens, generally 3-5 septate when mature, 27-60 x 3-5 µm and initially formed from simple lateral phialides, later forming slimy sporodochia. Chlamydospores, intercalary or terminal, on short lateral branches, solitary or in chains, hyaline, smooth to rough-walled. Stromatic pustules occasionally develop resembling Gibberella perithecia but no asci or ascospores have been reported (Booth, 1971).See also Brayford (1992).

The disease was first described in Zaire (Wardlaw, 1946) and has subsequently been diagnosed in several countries in central and western Africa: Côte d'Ivoire, Nigeria, Ghana, Cameroon and Congo (Wardlaw, 1948; Renard and Quillec, 1984; Oritsejafor, 1989). Localized outbreaks have also occurred in Brazil (Van de Lande, 1984) and Ecuador (Renard and de Franqueville, 1989). Early reports of the disease in Suriname (Anon., 1951) and Colombia (Sanchez Potes, 1966) remain unconfirmed.

RISK CRITERIA CATEGORY

ECONOMIC IMPORTANCE Low; potentially high
DISTRIBUTION Africa, South America
SEEDBORNE INCIDENCE Yes
SEED TRANSMITTED Yes
SEED TREATMENT Yes

OVERALL RISK Moderate


Notes on phytosanitary risk

Although contaminated seed and pollen have been exported in vast quantities for many years from western Africa to Asia without introducing the disease to this region, the recent outbreaks in South America appear to have originated from contaminated seed imported from western Africa. Therefore, importation of seed and pollen to any country outside western Africa does pose some phytosanitary risk, although with seeds this can be removed by vacuum infiltration with fungicides (Cooper, 2011; 2012). Brayford (1992) considered F.oxysporum f.sp. elaedis as one of the principal quarantine risks in the movement of germplasm from the centre of origin of the oil palm. Importation of oil palm seeds from its centre of diversity is often required for breeding programmes in South-East Asia but importation of oil palm seeds from West Africa is strictly controlled in Malaysia and Indonesia with quarantine procedures (Ritchie et al., 2000).

F. oxysporum f.sp. elaeidis is also pathogenic to the artificially inoculated South American oil palm, Elaeis oleifera (Renard et al., 1980). Isolates of F. oxysporum obtained from the root tissue of symptomless weed species (Amaranthus spinosus, Eupatorium odoratum, Mariscus alternifolius and Imperata cylindrica) from a Nigerian plantation were pathogenic to seedling oil palm (Oritsejafor, 1986). Under laboratory conditions, isolates of F. oxysporum pathogenic to oil palm can cause vascular wilt of date palm, while date palm isolates (F. oxysporum f.sp. albedinis) are pathogenic to oil palm (Paul, 1995).

The pathogen can attack oil palm at all ages from seedling to mature palm, and Prendergast (1957) suggested that in mature palms the disease can exist in two forms. In the chronic form, the older leaves become desiccated, the rachis breaks near or at some distance from the base and hangs down around the trunk. The disease progresses gradually, with younger leaves becoming successively affected whilst the erect young leaves in the crown are much reduced in size and may become chlorotic; the apex of the trunk may also reduce in diameter. Palms can exist in this condition for several years.

Alternatively, a palm may display the acute form of the disease, in which leaves dry out and die rapidly while retaining their original erect positions on the plant until broken off, usually several feet from the base, by wind action. The disease progresses rapidly and palms die within 2 or 3 months (Cooper, 2011).

Various intermediate stages between the acute and chronic forms may occur. De Franqueville and Renard (1990) suggested a third category of temporary wilt where palms develop leaf symptoms but later recover. On immature palms, leaves in the middle of the crown become yellow or brown; this first spreads to lower neighbouring leaves but eventually the palm will totally desiccate and die (de Franqueville and Diabate, 1996). At the nursery stage, infected palms show progressive shortening of younger leaves and desiccation and death of older leaves (Prendergast, 1957). These symptoms are thought to result from a combination of water stress (caused by xylem vessel blockage) and changes to plant gibberellin levels or activity (Mepsted et al., 1995a).

Internally this disease is characterized by discoloration and blockage of xylem vessels with tyloses and gums (Wardlaw, 1950; Prendergast, 1957; Cooper, 2011). Vascular discoloration (from healthy cream to infected dark brown) is always observed in palm stems, and in severely infected plants it can spread systemically to the petioles (Turner, 1981). However, even in highly diseased field palms most roots show no signs of infection (Wardlaw, 1950; Prendergast, 1957; Mepsted, 1992).

Although xylem discoloration of stem tissue is diagnostic for this disease, infection can be confirmed by surface sterilization of samples then plating onto Fusarium-selective medium (Papavizas, 1967); after 3-5 days mycelium of F. oxysporum emerging initially from xylem vessels ought to be visible. The pathogenicity of isolates should be checked by root inoculation of young seedlings (Flood et al., 1989; de Franqueville and Renard, 1990), symptoms take 3-6 months to develop. See also comments on DNA-based diagnosis.

Seed Contamination Tests

On seed, F. oxysporum is generally present in much lower quantities (about one-tenth) than F. solani, but can be detected by plating a dilution series onto a Fusarium selective medium (Flood et al., 1990).

- Each seed is agitated in 10 ml of sterile distilled water for 10 s, allowed to stand for 10 min and shaken again for 10 s before a dilution series is prepared.

- 1 ml of each dilution is plated onto a Papavizas selective medium (containing 1 g/l of pentachloronitrobenzene, 0.05 g/l chloramphenicol, 0.3 g/l penicillin and 0.13 g/l streptomycin sulphate).

- Plates are incubated at 28°C for 2-4 days, and colonies resembling F. oxysporum can be subcultured onto potato dextrose agar for identification.

Pathogenicity Test

Almost all batches of seed (unless vacuum treated with fungicide) may be contaminated with F. oxysporum, and the pathogenicity of these isolates can be confirmed by inoculation of seedlings (Flood et al., 1989).

External symptoms of this disease can be confused with Ganoderma (basal stem rot), Armillaria trunk rot or lightning strike.

The pathogen is generally regarded as soilborne and has been shown to penetrate roots through the loosely packed cells at the base of pneumathodes (Locke and Colhoun, 1977). In the field, spread is thought to occur through root contact with dead infected palm tissue (Prendergast, 1957; Renard and de Franqueville, 1989). However, due to the low incidence of root infection, Wardlaw (1950) suggested that colonization of the stem could follow infection of only a few roots, or that the pathogen could directly enter the stem through fissures at the base of the stem. Movement from tree to tree is supported by the statistical occurrence of infected palms in pairs or groups and the greater infection of palms with missing neighbours than those without (Dumortier et al., 1992; Rusli 2012).

Even in an area with a high incidence of wilt, the vast majority of soil isolates of F. oxysporum are non-pathogenic (Renard, 1967; JL Renard, CIRAD, Montpellier, France, personal communication, 1991; Mepsted, 1992, Mouyna, 1996). However, in a limited survey, all isolates of F. oxysporum from pollen, the surface of male inflorescences, the air and both the outside and inside of seeds were found to be pathogenic (Flood et al., 1990; Mepsted, 1992). The isolates from plant material all came from apparently uninfected palms. The epidemiological consequences of the aerial spread of the pathogen and the production of significant numbers of spores on the large male inflorescence are unknown, while contaminated seed and pollen may spread the pathogen to previously disease-free areas.

Environmental factors have been suggested to influence disease incidence; for example, higher levels of wilt were observed in areas of low rainfall (Prendergast, 1957; Aderungboye, 1981) and at the end of the rainy season (Waterson, 1953).

Seed Contamination and Transmission

Contamination by F. oxysporum of both the outside of seeds (Locke and Colhoun, 1973) and the kernel surface (Flood et al., 1990) have been reported. Levels of contamination vary considerably between seed consignments and between individual seeds. F. oxysporum was detected on the seed surface in five out of 10 commercial seed samples (10 seeds examined per sample) at levels up to 5000 c.f.u./seed and, with the same seeds, on the kernel surface in 30% of samples at levels up to 100 c.f.u./kernel (Flood et al., 1990). The pathogenicity of only two shell and two kernel isolates was checked, but all caused disease in inoculated seedlings.

Under glasshouse conditions in the UK two out of 60 artificially infested seeds (mean 50 c.f.u./shell and 7 c.f.u./kernel) developed wilt (Flood et al., 1994), yet many tons of seed have been exported from plantations affected by wilt in western Africa to Asia without transmitting the disease to that area. Recently, however, the disease has been observed in South America and fungal isolates from these outbreaks have identical restriction fragment length polymorphism (RFLP) patterns, and are vegetatively compatible with isolates from Côte d'Ivoire from where seeds were exported to these plantations (Flood et al., 1992; Mouyna et al., 1994). A worldwide collection of 76 F. oxysporum f.sp. elaeidis isolates and 21 F. oxysporum isolates from the soil of several palm groves was investigated using RFLPs (Mouyna et al., 1996). DNA fingerprint similarities produced 10 groups consisting of isolates with the same geographic origin. Isolates from Brazil and Ecuador had the same restriction pattern as some pathogenic isolates from the Côte d'Ivoire suggesting that they may have originated in Africa.

Transmission on contaminated palm seed is therefore possible, although the pathogen could also have been exported on contaminated seed of a cover crop (H. de Franqueville, IRHO/CIRAD, Dabou, Côte d'Ivoire, personal communication, 1995).

Pollen Contamination

F. oxysporum was isolated from 15 out of 30 randomly selected samples of commercial freeze-dried pollen at up to 40,000 c.f.u./g. The pathogenicity of only two isolates was checked, but both caused disease in inoculated seedlings (Flood et al., 1990). Preliminary experiments with the application of a range of fungicides in organic solvent carriers failed to produce complete decontamination combined with acceptable pollen viability (Mepsted, 1992).

Incidence

Contamination by F. oxysporum f.sp. elaeidis of both the outside of seeds of oil palm (Locke and Colhoun, 1973) and the kernel surface (Flood et al., 1990) have been reported. Levels of contamination vary considerably between seed consignments and between individual seeds. F. oxysporum f.sp. elaeidis was detected on the seed surface in five out of 10 commercial seed samples (10 seeds examined per sample) at levels up to 5000 c.f.u./seed and, with the same seeds, on the kernel surface in 30% of samples at levels up to 100 c.f.u./kernel (Flood et al., 1990). The pathogenicity of only two shell and two kernel isolates was checked, but all caused disease in inoculated seedlings.

Effect on Seed Quality

No effect on seed quality has been noted (Flood et al., 1994).

Pathogen Transmission

Under glasshouse conditions in the UK two out of 60 artificially infested seeds (mean 50 c.f.u./shell and 7 c.f.u./kernel) developed wilt (Flood et al., 1994), yet many tons of seed have been exported from plantations affected by wilt in western Africa to Asia without transmitting the disease to that area. Recently, however, indirect evidence for seed transmission was indicated by the occurrence of the disease in South America. Fungal isolates from these outbreaks had identical restriction fragment length polymorphism (RFLP) patterns and vegetative compatibility with isolates from Côte d'Ivoire from where seeds were exported to these plantations (Flood et al., 1992; Mouyna et al., 1994, 1996).

Seed Treatment

Normal seed treatments have no substantial effect on contamination levels (Flood et al., 1990). Vacuum infiltration ensures penetration of solutions via germ pores to the contaminated kernels and use of  prochloraz plus carbendazim completely eradicated all F. oxysporum f.sp. elaeidis and had no effect on seed germination or seedling vigour (Flood et al., 1994).

Seed Health Tests

Culture plate (Flood et al., 1990)

- Each seed is fragmented (e.g. using a vice) then agitated in 10 ml of sterile distilled water for 10 s, allowed to stand for 10 minutes and shaken again for 10 s before a dilution series is prepared.
- 1 ml of each dilution is plated onto a Fusarium selective medium (Papavizas, 1967)
- Plates are incubated at 28°C for 2-4 days, and colonies resembling F. oxysporum can be subcultured onto potato dextrose agar for identification.

Pathogenicity test

Seeds

Almost all batches of seed (unless vacuum treated as described in Seed Treatment) may be contaminated with F. oxysporum, and the pathogenicity of these isolates may be confirmed by inoculation of seedlings (Flood et al., 1989), but this takes several months.

Inoculate 100 ml of liquid medium in a 250 ml flask with three 5 ml agar plugs from the margin of the culture. Sucrose-salts solution or Czapek Dox with added casamino acids (Cooper and Wood, 1975) is suitable but any general purpose liquid medium will be satisfactory. This will produce inoculum for about 10 plants.

Agitate culture (100 r.p.m., or shake flask several times a day) for 5 days at 25-27°C.

Filter through sterile muslin and adjust spore concentration to approx. 10,000,000 spores per ml.

Inoculate seedlings (20 per isolate) or clones (10 per isolate) at the two to three leaf stage by exposing roots at the base of the stem and applying 10 ml of inoculum to each plant.

External symptoms of yellowing and necrosis of older leaves and stunting of young leaves will develop 3-6 months after inoculation.

At the end of the experiment, split the base of the seedling longitudinally and examine tissue for brown discoloration; re-isolate onto a Fusarium-selective medium.

Incidence - pollen

F. oxysporum was isolated from 15 out of 30 randomly selected samples of commercial freeze-dried pollen at up to 40,000 c.f.u./g (Flood et al., 1990).

Pathogen Transmission - pollen

As with seeds, pollen has been exported from western Africa to Asia for many years without introducing the disease to this area, although the pathogen may have been introduced by this method.

Pollen Treatment

F. oxysporum was isolated from 15 out of 30 randomly selected samples of commercial freeze-dried pollen at up to 40,000 c.f.u./g. The pathogenicity of only two isolates was checked, but both caused disease in inoculated seedlings (Flood et al., 1990). Preliminary experiments with the application of a range of fungicides in organic solvent carriers failed to produce complete decontamination combined with acceptable pollen viability (Mepsted, 1992).

Pollen Health Tests

This is the same as for seeds, with 0.03 g of pollen per sample.

Fusarium wilt is the most important disease of oil palm in western and central Africa (Turner, 1981). Losses of up to 50% have been recorded for palms under 10 years old in some plantations (Wardlaw, 1950; Waterson, 1953; Guldentops, 1962; Renard and Quillec, 1983). DuMortier et al (1992) recorded yield from palms with acute wilt in the year before detah as c. 54% and from palms with chronic wilt 30% that of healthy palms. However, in general, losses are currently low and have been estimated to range between 1 and 2% per annum (Bachy, 1970; de Franqueville and Renard, 1990) and in some areas of western Africa wilt has never been observed in groves or plantations (Waterson, 1953; Aderungboye, 1981). Prendergast (1957) suggested that no yield reduction could be expected until more than 20% of palms had died, due to the increased vigour of adjacent palms. However, Renard and de Franqueville (1989) observed a 6-16% yield reduction in 6-year-old palms when only 2.5-5.5% of plants showed external symptoms. They attributed most of the yield reduction to the 20-30% of palms that appeared to be healthy yet were infected without having any obvious external wilt symptoms. Planned introductions of palms lacking resistance/tolerance to new areas of Africa may be vulnerable.

Brown discoloration of xylem vessels in the trunk, and in the leaf bases of severely infected palms, is diagnostic of this disease. However, care must be taken to distinguish between brown xylem vessels and discoloration of the sclerenchyma tissue of vascular bundles, which can occur in old healthy palms or following attack by Ganoderma or Armillaria (Wardlaw, 1950). Trunk samples can be obtained by felling the palm or preferably with an increment borer, which avoids destructive sampling of entire palms (Mepsted et al., 1991). Samples can be examined microscopically for Fusarium hyphae or vascular occlusion with gels or more reliably plated onto agar; the pathogen can also be isolated from seeds and pollen (see Diagnostic Methods).

Currently, only the species F. oxysporum can be diagnosed at reasonable speed by DNA-based diagnostics using PCR with primers based on the translation elongation factor (TEF) gene (Geisser et al., 2004; Rusli 2012). Fusaria are common contaminants but detection of F. oxysporum is enough for a seed batch to be destroyed to avoid disease spread to new areas. There is a clear need for pathotype (f. sp. elaeidis)-specific primers (Cooper, 2012).

Due to the variable regulations around (de)registration of pesticides, your national list of registered pesticides or relevant authority should be consulted to determine which products are legally allowed for use in your country when considering chemical control. Pesticides should always be used in a lawful manner, consistent with the product's label.

There is a need for pathotype (f. sp. elaeidis)-specific primers (Cooper, 2012).