Ganoderma boninense basidiospores in oil palm plantations: evaluation of their possible role in stem rots of Elaeis guineensis

Basidiospores are implicated in the distribution and genetic diversity of Ganoderma boninense, cause of basal stem rot (BSR) and upper stem rot (USR) of oil palm (Elaeis guineensis). Measurement of aerial basidiospores within plantations in Sumatra showed continuous and high production over 24 h (range c. 2–11 000 spores m^?3) with maximum release during early evening. Basidiospores applied to cut surfaces of fronds, peduncles and stems germinated in situ. Equivalent, extensive wounds are created during plantation harvesting and management and represent potential sites for formation of infective heterokaryons following mating of haploid basidiospore germlings. Use of spore‐sized micro‐beads showed that basidiospores could be pulled up to 10 cm into severed xylem vessels, where they are relatively protected from dehydration, UV irradiation and competing microflora. Diversity of isolates from five locations on two plantations was assessed by RAMS fingerprinting. Isolates from within individual palms with USR were identical and represent single infections, but different USR infections had unique band patterns and revealed separate infections. Some BSR‐affected trees contained more than one isolate, and thus had multiple infections. There was one example of adjacent BSR palms with the same isolate, indicating vegetative spread, but there were no identical genets from BSR infections and adjacent fallen palms. Isolate diversity was as great within a plantation as between plantations. It is evident that basidiospores play a major role in spread and genetic variability of G. boninense. Evidence for direct basidiospore infection via cut fronds, indirectly through roots via colonized debris and less frequently, infection by vegetative, clonal spread is considered.     

Disease progression of a lethal decline caused by the 16SrIV‐D phytoplasma in Florida palms

Recently, a new phytoplasma was discovered in Hillsborough County in the state of Florida, USA. This phytoplasma belongs to the 16SrIV taxonomic group and is classified as subgroup D. It is the causal agent of lethal bronzing disease (LBD) of palm. Since the discovery of LBD in 2006, the disease has spread throughout much of the state. In 2014 and 2015, stands of cabbage palm and queen palms that had been present at the University of Florida's Fort Lauderdale Research and Education Center in Davie, FL began showing symptoms of LBD. After confirming the presence of the LBD phytoplasma in initially infected palms by nested PCR and RFLP analysis, all palms were systematically sampled over the period of 1 year to monitor and quantify disease spread. A total of 30 cabbage palms were tested monthly by qPCR, with five testing positive on the first sample date. By the end of the study period, 16 cabbage palms had died from the infection. A total of 16 queen palms were surveyed, with three palms initially testing positive. By the end of the study, four queen palms had tested positive and died from the infection. To the authors’ knowledge, this study is the first to document and quantify spread of palm‐infecting phytoplasmas. This data provides important insights into the ecology of palm‐infecting phytoplasmas and highlights the impact that the movement of infective insects can pose to established stands of palms.     

Effects of inoculum potential, shading and soil temperature on root infection of oil palm seedlings by the basal stem rot pathogen Ganoderma boninense

Ganoderma boninense causes severe losses to oil palm in South East Asia. The disease typically manifests itself as basal stem rot, but there remains controversy over the route of infection and source of inoculum. Using isolates differing in aggressiveness, infection via roots was confirmed; it was also shown that large inoculum provided as Ganoderma -infested palm- or rubber-wood blocks (12 × 6 × 6 cm) is necessary for soil infection of seedlings after 6–8 months. Smaller blocks (3 × 3 × 3 cm) produced rapid (≤ 3 months) infection of roots and lower stem when physically attached to roots. Therefore fragmentation of infested palm wood from a felled, mature plantation before subsequent replanting may provide inoculum. Failure of G. boninense to grow through non-sterile soil or organic debris from frond bases, suggests it is a poor competitor and that roots must contact inoculum directly. Severe disease occurred after 8 months on inoculated seedlings under shade, but not on seedlings exposed to sun. Soil temperatures in sunlight frequently rose above 40°C and reached 45°C, whereas in shade they never exceeded 32°C. Ganoderma boninense is probably inhibited in exposed soil since optimal growth in vitro was 25–30°C, and there was no recovery from 45°C. Soil temperature may explain why symptoms often first appear in mature plantations when canopy formation creates shade. Infection is not peculiar to senescing palms but can occur throughout the growth cycle.     

Genetic structure of a population of Ganoderma boninense on oil palm

Ganoderma boninense (the causal agent of basal stem rot of oil palm in Papua New Guinea) has a tetrapolar mating system with multiple alleles. Investigations into the population structure of G. boninense , using interfertility between isolates as a marker, revealed that the population on oil palm was comprised predominantly of genetically distinct individuals, although a number of isolates were found to share single mating alleles. No direct hereditary relationship was found between isolates on neighbouring or spatially separated diseased palms, indicating that outcrossing had probably occurred over several generations in the founder population prior to colonization of oil palm. In this study, a total of 81 A and 83 B mating type alleles (factors) were detected with 18 allelic repeats at the A locus and 17 at the B locus. Alleles appeared to be randomly dispersed throughout the population in each study block, although there was a significantly ( P  < 0·01) higher proportion of allelic repeats between estates separated by several kilometres compared to within estates. This is the first study on the population structure of this species and confirms that sexual reproduction is a significant component of the epidemiology of this disease.     

Transmission of the coconut cadang-cadang viroid to six species of palm by inoculation with nucleic acid extracts

Seedlings of Areca catechu (betel nut palm), Corypha elata (buri palm), Adonidia merrillii (manila palm), Elaeis guineensis (oil palm), Chrysalidocarpus lutescens (palmera) and Oreodoxa regia (royal palm) were inoculated with nucleic acid extracts from coconut palms with cadang-cadang disease. Within 2 years of inoculation, analysis using a ³²P-labelled DNA probe complementary to the coconut cadang-cadang viroid (CCCV) showed that RNA sequences identical to CCCV were present in the inoculated seedlings. Electrophoresis in polyacrylamide gels showed that these palms also contained an RNA with mobility identical to CCCV. Four to five years after inoculation, the infected palms of four species were usually stunted compared with uninoculated palms, while betel nut and palmera were not stunted. Yellowing of leaflets was observed with defined spots or mottling of the older fronds in all except betel nut palms. All infected palms showed mild or severe yellow-leaf spotting. These results widen the known host range and. hence, the potential number of viroid reservoir species in the field.     

Identification and Spread of Fomitiporia punctata Associated with Wood Decay of Grapevine Showing Symptoms of Esca

ABSTRACT A full understanding of the pathology of esca, a chronic disease of grapevines, has been problematic, in part because the identity of the pathogen (or pathogens) has been difficult to determine. The wood decay symptoms of esca have been most often associated with Phellinus igniarius or Fomitiporia punctata. However, Koch's postulates have not been completely fulfilled because symptoms take many years to develop. The goal of this study was to determine the identity and mode of spread of basidiomycetes associated with wood decay in vines showing esca symptoms in Italian vineyards. Vineyards were intensively studied for the presence of basidiocarps, and mycelium was isolated from symptomatic vines. Fruiting bodies were identified by morphology, while mycelial isolates were identified by restriction fragment length polymorphism analysis of the internal transcribed spacer region of the nuclear ribosomal RNA gene cluster. Fomitiporia punctata fruiting bodies and mycelium were associated with approximately 50% of the vines showing esca symptoms in two vineyards; P. igniarius was not found in any samples. Fruiting bodies of F. punctata were found in five of six vineyards examined, but at low frequencies except in one vineyard. The diversity of somatic incompatibility types was very high; isolates from almost every vine had different somatic incompatibility types. With few exceptions, symptomatic and dead vines were not spatially aggregated within 12 vineyards. The combination of diverse somatic incompatibility types and lack of spatial aggregations are not consistent with the hypothesis that the disease is spread clonally through roots or by pruning tools. The correct identity of basidiomycetes associated with wood decay of vines with esca symptoms is important for understanding the epidemiology of this disease because F. punctata is found commonly on many woody hosts in Europe, which may represent a potential inoculum source for this disease.     

Basal stem rot of oil palm (Elaeis guineensis); mode of root infection and lower stem invasion by Ganoderma boninense

Reproducible infection of intact roots of oil palm ( Elaeis guineensis ) with Ganoderma boninense , the cause of basal stem rot, showed penetration followed by rapid longitudinal progression of hyphae and colonization of the lower stem (bole). Light and transmission electron microscopy showed invasion of the root cortex, with no evidence of selective progression through the vascular system or lacunae. In newly colonized tissue the fungus behaved as a hemibiotroph, with numerous, wide, intracellular hyphae occupying entire host cells that possessed intact cell walls and contained discernible cytoplasm and organelles. In the bole this phase coincided with a complete depletion of previously abundant starch grains in advance of invasion. Subsequently, in the roots and colonized stem base, widespread necrotrophic, enzymatic attack of all layers of the host cell walls occurred. Hyphae were intra- and intercellular and intramural and associated host cell wall degradation was often at a distance from hyphae, resulting in cavities within cell walls. A third developmental stage was the formation of an extensive, melanized, tough mycelium or pseudo-sclerotium which surrounded roots and comprised many very thick-walled cells encasing more typical thin-walled hyphae. Macroscopic observation of and isolation from the bole of randomly felled, commercial palms provided confirmatory evidence that multiple infections originated in the roots before spreading into the base of long-established palms.     

Vegetative incompatibility in Diaporthe ambigua

Vegetative compatibility of strains of Diaporthe ambigua has not previously been examined. Single ascospore and single ascus strains, originating from individual apple, pear and plum rootstocks, were paired on freshly prepared oatmeal agar to determine if vegetative incompatibility could be detected in D. ambigua . Barrage reactions were evident as black lines along the zone of mycelial contact between expanding colonies (vegetative incompatibility reaction). Strains from cankers within an area were of numerous vegetative compatibility groups (VCGs). Strains from adjacent rootstocks usually differed in VCG. D. ambigua has the ability to outcross, and does so, despite its apparent homothallic nature. DsRNA-containing strains of D. ambigua developed a broad, clear zone when paired with a dsRNA-containing strain from a different VCG.     

Evaluation of Malaysian oil palm progenies for susceptibility,resistance or tolerance to Fusarium oxysporum f. sp. elaeidis and defence‐related gene expression in roots

Vascular wilt of oil palm caused by Fusarium oxysporum f. sp. elaeidis (Foe) is a devastating disease in West and Central Africa. As the oil palm industry in southeast Asia is still expanding, so is the oil palm germplasm collection through the importation of seed and pollen from Africa, the centre of diversity for Elaeis guineensis. There is a risk of inadvertent spread of the disease on contaminated seed or pollen. Regular re‐evaluation of the reaction of currently grown palm genotypes towards Foe is clearly required for biosecurity. This study has demonstrated that four Malaysian oil palm progenies, three in current or recent commercial use, are highly susceptible to infection by at least one of two African isolates of Foe, representing different countries, aggressiveness and vegetative compatibility groups. Symptoms and reduction of palm growth generally reflected the extent and intensity of systemic colonization by Foe. Progeny PK 5463 expressed partial resistance to Foe isolate F3, but not to isolate 16F, displaying significantly milder symptoms and supporting less widespread vascular colonization. This relatively incompatible interaction was used to study expression of potential defence‐related genes during root infection when compared to a susceptible palm–isolate combination. The only significant response was an early up‐regulation of chitinase in resistant palms. The research revealed at least one progeny–isolate differential interaction, and the associated resistance expression suggests a component of tolerance, because colonization by Foe was systemic in both compatible and incompatible combinations.     

Mitochondrial DNA Variability in Fusarium Proliferatum (Gibberella Intermedia)

Restriction fragment length polymorphisms (RFLP) were used to assess genetic diversity of mitochondrial DNA (mtDNA) among 184 isolates of Fusarium proliferatum recovered from maize, asparagus, palms and reed. All strains were cross-fertile with standard mating type tester strains of Gibberella intermedia. Sixteen mitochondrial haplotypes were identified following digestion of DNAs with HaeIII, with seven, seven, five and six different haplotypes from maize, asparagus, palms and reed, respectively. Four haplotypes (I, III, IV and VII) were found on more than one host. Of these four, haplotype I was dominant on maize, representing 71% of the isolates. The banding patterns for haplotypes III and IV were >90% similar to the banding pattern of haplotype I. Haplotypes I, III and IV accounted for 87% of the isolates from maize, but were less common on the other hosts, accounting for 70%, 52% and 33% of the isolates from asparagus, palms and reed, respectively. Thirteen of the 16 haplotypes were recovered from only a single host plant species. When comparing the banding patterns and frequencies of these haplotypes, at least five were recovered at a higher frequency from one host relative to the others. Our results suggest that mtDNA RFLP analysis is a useful indicator of genetic divergence in Fusarium proliferatum.     

Molecular records of micro-evolution within the Algerian population of Fusarium oxysporum f. sp. albedinis during its spread to new oases

The genetic diversity of the date palm wilt pathogen Fusarium oxysporum f. sp. albedinis in Algeria was assessed using vegetative compatibility, restriction fragment length polymorphism (RFLP) of mitochondrial DNA (mtDNA), and random amplified polymorphic DNA (RAPD). Ninety-eight isolates were collected from the main infested regions, Touat, Gourara and Mzab, and 6 isolates from Morocco were added for comparison. All isolates were vegetatively compatible and belonged to VCG 0170. No variation was detected in the mtDNA of a subset of 73 isolates and the RAPD analysis indicated that they were genetically very closely related. However, some geographic substructuring was apparent, suggesting that local diversification of the pathogen might have occurred. These results provide evidence that the Algerian isolates of F. oxysporum f. sp. albedinis belong to a same clonal lineage and support the hypothesis that they were probably founded by a single virulent clone that originated from the Moroccan oases where the date palm wilt (Bayoud disease) was first detected. Based on similarity of RAPD patterns occurring in different oases, and on historical records of the Bayoud disease in Algeria, spread of the pathogen in the different regions is discussed.     

Detection of the mycoplasma-like organism associated with lethal yellowing disease of palms in Florida by polymerase chain reaction

DNA amplification by polymerase chain reaction (PCR) was used specifically to detect the mycoplasma-like organism (MLO) associated with lethal yellowing disease of palms in Florida. For PCR, a pair of oligonucleotide primers was synthesized according to partial sequences of a cloned 1·3 kbp fragment of lethal yellowing MLO-specific genomic DNA isolated from a diseased windmill palm ( Trachycarpus fortunei ). A DNA product of about 1 kbp was specifically amplified by PCR in reaction mixtures containing template DNA derived from either heart, inflorescence or leaf tissues of lethal yellowing-affected palms. PCR performed for 35 cycles with as little as 5 pg of DNA template, in some instances, was sufficient consistently to amplify the same lethal yellowing MLO DNA product from hearts of 11 species comprising 30 symptomatic palms. Similar reliable and reproducible detection of the lethal yellowing MLO in palm inflorescence spikelets was also achieved after 35 cycles of PCR. When template DNA for PCR was derived from tissues of the the most immature emerging leaf, a 40-cycle reaction was sufficient for consistent foliar detection of the pathogen in all coconut palms including palms with earliest visible symptoms of disease.     

Can increasing infestations by Rhynchophorus ferrugineus threaten endemic Chamaerops humilis in Sardinia (Italy)?

The red palm weevil, Rhynchophorus ferrugineus, is the most devastating pest of the Canary palm, Phoenix canariensis, in the Mediterranean Basin. Sporadic infestations have also been observed in other cultivated palm species and, above all, in the endemic European fan palm, Chamaerops humilis. Considering the high ecological importance of C. humilis in coastal ecosystems, the present paper gives an update on the status of R. ferrugineus infestations on the European fan palm in Sardinia (Italy). Since the introduction of R. ferrugineus into Sardinia, attacks of this pest on C. humilis have been limited but have increased in the last 2 years, including infestations on eight naturally growing palms in a Natura 2000 site. Male and female red palm weevils that emerged from C. humilis were similar in size to those of P. canariensis, indicating that the quality of these two feeding substrates is similar. The limited number of infested European fan palms observed so far suggests caution when predicting a rapid spread of this pest in cultivated and non‐cultivated C. humilis palms. Nonetheless, natural populations of C. humilis along western Mediterranean shores should be monitored in order to highlight the potential establishment of R. ferrugineus in these habitats.     

Fine structure of phloematic trypanosomatid–coconut tree interaction

Phytomonas wilt or Hartrot is a fatal disease of palm (Arecaceae) species including Cocos nucifera (coconut) and is caused by a phloematic trypanosomatid, a promastigote parasite that inhabits phloem sieve elements of disease palms. In the present work, we described the morphology of the interaction between a phloematic trypanosomatid (Phytomonas staheli) and C. nucifera. Two varieties and one ecotype of the adult coconut palm from northeast and southeast Brazil were analyzed, totaling 34,000 plants. Coconut palm losses due to Hartrot varied according to the variety or ecotype and geographic area. Occurrence of Hartrot was insignificant in Rio de Janeiro state (southeast), but in Bahia state (northeast) losses were substantial when appropriate cultural practices were not applied. Symptomatic and healthy palm tissues were analyzed by light and electron microscopy. Laboratory diagnoses revealed the twisted promastigote form of phloematic trypanosomatids in the extracts of shoot apex, leaves, stems and inflorescence in diseased plants, but not in the healthy ones. No parasites were found in the roots. Although the general anatomy of healthy and diseased palms was similar, callose deposition in the sieve plates was revealed by histochemistry and immunocytochemistry in the diseased tissue. Plugging by the P-protein and plastid alterations was also observed. Our observations strongly suggest that parasite traffic between sieve elements took place, although their cell bodies were larger than the sieve pores. Phloematic trypanosomatid proliferation in the sieve tube elements might interrupt the transport of phloem or/and consume plant nutrients. In addition, an association between the percentage of sieve elements colonized by pathogen in palm tissues and disease severity was established.     

Red palm weevil (Rhynchophorus ferrugineus), an invasive pest recently found in the Caribbean that threatens the region

Rhynchophorus ferrugineus, an important palm pest, was accidentally introduced into the Caribbean. A monitoring programme was established to determine the population level and distribution of infestations on Aruba and Curacao through the use of commercially available pheromone traps. Due to the small size of the islands and limited distribution of palms, eradication may be feasible using a combination of trapping, timely disposal of infested palms and curative and prophylactic chemical treatments. These studies on the pest in the Caribbean were used to help design a USDA plant health emergency response through the development of Animal Plant Health Inspection Service, Plant Protection and Quarantine New Pest Response Guidelines and provide an effective emergency response programme for other Caribbean Islands and the Americas.     

Field efficacy of imidacloprid and Steinernema carpocapsae in a chitosan formulation against the red palm weevil Rhynchophorus ferrugineus (Coleoptera: Curculionidae) in Phoenix canariensis

BACKGROUND: The invasive red palm weevil, Rhynchophorus ferrugineus (Olivier), has become the major pest of palms in the Mediterranean Basin. Chemical control against this species is difficult because of its cryptic habits and is mainly based on the repeated application of large quantities of synthetic insecticides. The aim of this work has been to evaluate in the field the efficacy of imidacloprid (Confidor^® 240 OD) and Steinernema carpocapsae Weiser with chitosan (Biorend R^® Palmeras) as soil and stipe treatments respectively, alone or in combination, against this pest. RESULTS: All treatments significantly reduced the mean number of immature stages of R. ferrugineus per palm. However, there were no significant differences among the different treatments considered. Efficacies ranged from 83.8 to 99.7% for the mean number of immature stages found in the palms and resulted in a significant increase in palm survival compared with the untreated control (75.0–90.0% versus 16.5% respectively). CONCLUSION: Both imidacloprid and S. carpocapsae in a chitosan formulation proved highly effective against R. ferrugineus in the field, and their efficacies did not significantly change when used in combination. Copyright © 2009 Society of Chemical Industry     

A set of novel RNAs transcribed from the chloroplast genome accumulates in date palm leaflets affected by brittle leaf disease

Brittle leaf disease or maladie des feuilles cassantes (MFC) is a lethal disorder of date palms that has assumed epidemic proportions in the oases of southern Tunisia. After a prolonged period during which palms are declining, the disease ends with the death of the palms. Whereas no pathogen could ever be associated with the disease, leaflets of affected palms have been previously shown to be deficient in manganese. Analysis of RNA preparations from leaflets of MFC-affected palms revealed the presence of a set of novel RNAs (MFC-RNAs) of sense and antisense polarities, which are homologous to various regions of the date palm chloroplast genome, such as the regions containing genes rrn5S-trnR(ACG) and trnM(CAU)-atpE. In the RNA preparations obtained from leaflets of affected palms, some of these RNAs are present as double-stranded species (MFC-dsRNAs), as witnessed by results from cellulose chromatography, end labeling, RNase digestion, and northern hybridization with strand specific probes. These MFC-RNAs represent a novel type of host-derived RNAs, and their presence in MFC-affected date palms is of diagnostic value.     

Comparison of the ribosomal RNA operon from Texas Phoenix decline and lethal yellowing phytoplasmas

A comparison was made of the two palm yellows phytoplasmas affecting palms to determine if the entire ribosomal RNA operon portion of the phytoplasma genome, or portions thereof, could account for the observed palm host differences. Polymerase chain reaction (PCR) was used to amplify a 5.0?kb DNA fragment consisting of the entire ribosomal RNA operon from a subgroup 16SrIV-D phytoplasma that causes Texas Phoenix palm decline (TPD) in cabbage (Sabal palmetto) palm in west central Florida and from a subgroup 16SrIV-A phytoplasma that causes lethal yellowing (LY) in coconut (Cocos nucifera) palm in Jamaica. Before the PCR reaction, we sequenced by 454 sequencing a draft genome of the coconut LY phytoplasma, strain LYFL, that infects C. nucifera in Florida, and obtained from this draft sequence both copies of the entire ribosomal operon. Sequence analysis of the ribosomal RNA operons from both the LY and TPD phytoplasmas revealed the gene composition and orientation for the operons to be 5′16S rRNA-tRNA^Ile-23S rRNA-5S rRNA3′ and a tRNA^Val3′ downstream of the 5S rRNA gene. Based on molecular comparisons using the sequences of the ribosomal RNA operon, the TPD (16SrIV-D) strain was 98?% similar to the LY (16SrIV-A) strains.     

Responses of Rhynchophorus ferrugineus adults to selected synthetic palm esters: electroantennographic studies and trap catches in an urban environment

BACKGROUND: The red palm weevil (RPW), Rhynchophorus ferrugineus Olivier, accidentally introduced into the European countries facing the Mediterranean basin, is becoming the most serious pest for ornamental palms in the urban environment. In the present work, tests were conducted on the electroantennogram (EAG) and behavioural responses of RPW adults to five selected synthetic palm esters, ethyl propionate, ethyl butyrate, ethyl isobutyrate, ethyl lactate and ethyl acetate, to assess the application of a mass trapping technique in an urban environment. RESULTS: In the laboratory, EAG bioassays on RPW adults with the synthetic palm esters showed: (1) positive dose‐dependent responses; (2) differences among the treatments; (3) a greater response to ethyl propionate; (4) a higher sensitivity of female antennae compared with male antennae. In the urban environment, RPW behavioural responses, evaluated over six biweekly observations on pheromone‐ and molasses‐ baited traps, showed that more adults were caught by traps supplemented with ethyl propionate and ethyl acetate than by traps supplemented with only ethyl propionate. Similar catches were recorded in the traps supplemented with ethyl propionate or ethyl acetate. CONCLUSION: This study supports the application of the mass trapping implemented with synthetic palm esters as a potential tool for the management of RPW populations to protect ornamental palms in urban environments. Copyright © 2010 Society of Chemical Industry     

Use of nitrate non-utilising (Nit) mutants to determine vegetative compatibility in Botryotinia fuckeliana (Botrytis cinerea)

Nitrate non-utilising (Nit) mutants of six field strains and three single-ascospore strains of Botryotinia fuckeliana were selected by plating mycelial plugs onto a nitrate-containing minimal medium amended with chlorate. Mutants were characterised by growth responses on minimal medium amended with various sole nitrogen sources. For each parental strain two mutants were produced: nit1, defective in nitrate reductase apoenzyme; and NitM, defective in the molybdenum cofactor pathway. Complementation on nitrate minimal medium was observed between nit1 and NitM mutants from the same parent in all instances. However, complementation was not observed between any such combinations when the mutants were derived from different parents. It is concluded that a vegetative incompatibility system operates in B. fuckeliana resulting in multiple vegetative compatibility groups. One of the single-ascospore strains was derived from a cross between two of the field strains, indicating that new compatibility groups can be generated during sexual reproduction. Mycelial interaction zones were formed between all parental strains when they were paired on NaCl-amended medium, indicating congruence between mycelial incompatibility and vegetative incompatibility.