Coffee berry disease in Kenya. I. Colletotrichum spp. colonizing the bark of Coffea arabica

Several species ofColletotrichum occur in maturing bark ofCoffea arabica branches in Kenya. TheColletotrichum population inhabits the bark tissue external to the developing phellogens in the cortex. TheColletotrichum species are unable to invade green bark tissue, where the phellogen has not yet been differentiated, while colonization ceases on the phelloderm of the true bark. Only one of theColletotrichum species discussed in this paper,C. coffeanum, can infect green coffee berries. Shortly after the initiation of the first phellogen in the cortex the parasite is in a small area in the bark. It cannot be found in bark tissues where more phellogens have been formed and where the colour of the bark has changed from yellow-green to brown or black. From the bark ofC. liberica trees, grown in Kenya, andC. arabica cv. ’lsBourbon’, grown in greenhouses in the Netherlands one of the saprophytic components of theColletotrichum population could be isolated. It was impossible to induce ddie-back symptoms or mere infection by inoculation of green internodes even after wounding of live branches ofC. arabica with any of theColletotrichum components colonizing the bark. It is suggested that die-back systems of coffee in Kenya are primarily caused by unfavourable growing conditions.     

Genetic and morphological diversity of mono-spore isolates of glomerella cingulata associated with coffee berry disease

In earlier experiments the perfect state,Glomerella cingulata, had been found associated withColletotrichum coffeanum. As the linkage betweenG. cingulata and the coffee berry disease (CBD) pathogen had still been doubtful, further investigations were done. It was found that mono-spore cultures derived fromG. cingulata perithecia from two different CBD isolates were not compatible. This would suggest genetic differences between these two CBD pathogens.Preliminary tests with mono-ascospore isolates of oneG. cingulata source resulted in a wide range of segregation types. It has not yet been possible to obtain a conidial form fromG. cingulata, which has the morphological and pathogenic characteristics ofC. coffeanum. The results of the experiments suggest, however, that a CBD pathogen may arise through genetic segregation from aG. cingulata state.     

Cytogenomic characterization of Colletotrichum kahawae,the causal agent of coffee berry disease,reveals diversity in minichromosome profiles and genome size expansion

Colletotrichum kahawae is an emerging fungal pathogen, which has recently undergone a speciation process from a generalistic ‘C. gloeosporioides species complex' background by acquiring the unique capacity to infect green coffee berries, thus causing coffee berry disease. This is a severe and widespread disease in Africa and an imminent threat to Arabica coffee cultivation in Asia and America, if the pathogen enters those continents. Genetic diversity within C. kahawae is low but notorious differences in pathogen aggressiveness have been described. This work characterized two cytogenomic traits (genome size and minichromosome profiles) of a collection of C. kahawae isolates, representing the breadth of its genetic diversity and distinct aggressiveness classes, along with closely related taxa. The results obtained constitute the first flow cytometry‐based genome size estimation in the genus Colletotrichum and show a c. 8 Mb genome size expansion between C. kahawae (79·5 Mb on average) and its closest relatives (71·3 Mb), corroborating evidence indicating that C. kahawae (i.e. the coffee berry disease pathogens) should remain as a distinct species. Results have also shown the presence of two to five minichromosomes in C. kahawae, suggesting a positive relationship between the number of minichromosomes and the level of aggressiveness of the different isolates analysed, while no correlation could be established between aggressiveness and whole genome size. Overall, these results may be the basis for the identification of pathogenicity/aggressiveness‐related factors in such minichromosomes, and may provide clues to the characterization of specific markers for aggressiveness classes.     

Identification of <Emphasis Type="Italic">Colletotrichum</Emphasis> species associated with anthracnose disease of coffee in Vietnam

Twenty-three isolates of Colletotrichum gloeosporioides, five isolates of C. acutatum, two isolates of C. capsici and six isolates of C. boninense associated with anthracnose disease on coffee (Coffea spp.) in Vietnam were identified based on morphology and DNA analysis. Phylogenetic analysis of DNA sequences from the internal transcribed spacer region of nuclear rDNA and a portion of mitochondrial small subunit rRNA were concordant and allowed good separation of the taxa. We found several Colletotrichum isolates of unknown species and their taxonomic position remains unresolved. The majority of Vietnamese isolates belonged to C. gloeosporioides and they grouped together with the coffee berry disease (CBD) fungus, C. kahawae. However, C. kahawae could be distinguished from the Vietnamese C. gloeosporioides isolates based on ammonium tartrate utilization, growth rate and pathogenicity. C. gloeosporioides isolates were more pathogenic on detached green berries than isolates of the other species, i.e. C. acutatum, C capsici and C. boninense. Some of the C. gloeosporioides isolates produced slightly sunken lesions on green berries resembling CBD symptoms but it did not destroy the bean. We did not find any evidence of the presence of C. kahawae in Vietnam.     

Control of benomyl-resistant strains of Colletotrichum coffeanum, the cause of coffee berry disease

Sprays containing difolatan, dithianon, chlorothalonil and cuprous oxide were tested for the control of coffee berry disease (CBD) in 1979 and 1980 at an estate in Kenya with abundant benomyl-resistant strains of Colletotrichum coffeanum. In 1979 all fungicides gave good CBD control and increased yields of coffee. In 1980 only difolatan increased yield. Isolates of C. coffeanum capable of growing on agar with 8 μg/ml benomyl were detected throughout the 2 years of these trials.     

Evidence of cork barrier formation as a resistance mechanism to berry disease (Colletotrichum coffeanum) in arabica coffee

The histology of resistance to coffee berry disease (CBD) was studied in artificially inoculated berries and hypocotyls of 6-week old seedlings of a number of varieties ofCoffea arabica L. In resistant varieties a phellogen was quickly formed some cell layers below the site of infection and progress of the fungal invasion was effectively blocked by a complete barrier of suberized cells. Such barriers were absent or incompletely developed in CBD susceptible varieties. A highly significant correlation (r=0.87) was found between the frequencies of complete barrier formation in berries and in hypocotyls of young seedlings, while both were also highly correlated to observed mature plant resistance (r>-0.93). Resistance to CBD in arabica coffee may to an important extent be based on the formation of cork barriers. These cork barriers confine the pathogen to the small volume of tissue external to the barrier so that its growth is severely restricted. Such a resistance mechanism is likely to be stable (race-nonspecific). The almost identical response to infection observed in berries and hypocotyls provides further evidence that plants with resistance to CBD can be reliably preselected by the hypocotyl inoculation test.     

落叶松早期落叶病子囊孢子扩散形式及药剂防治试验的初步研究

落叶松早期落叶病子囊孢子在空中的扩散,在长春地区,以7月份为盛期。林外水平距1600米及垂直高55米处均可捕捉到不低于林内孢子的数量,其方向符合夏季主风方向。噴药防治的效果,无論成林或幼林,均以0.5度石灰硫黄合剂最好,尤其噴药3次的更佳。地面噴药抑止产生子囊壳,以3度石灰硫黄合剂及每株20克石灰比二硝基磷甲酚效果好。     

Role of rainfall in the development of coffee berry disease in Coffea arabica caused by Colletotrichum kahawae,in Cameroon

The development of coffee berry disease (CBD) epidemics (caused by Colletotrichum kahawae) in Cameroon was monitored over two successive years (2004 and 2005) on coffee trees protected from rainfall by transparent plastic sheets and on unprotected control trees. This work was done to assess how rain affected disease development when it did not fall directly onto the coffee trees and to determine the influence of primary inoculum on the severity of CBD. Weekly observations over the 2 years showed that there were 1·1% diseased berries on coffee trees completely protected from rainfall, compared with 45% diseased berries on unprotected coffee trees. Disease severity on unprotected trees during the 2 years of the experiment was estimated at 53% diseased berries, compared with 27% on trees only protected in the first year. These results confirmed rainfall as one of the key physical factors in the development of Arabica CBD. They also provided evidence of a subsequent effect of protecting coffee trees from rainfall in 2004 on the severity of CBD in 2005. This suggested some practices that might lead to very effective cultural control of CBD in regions where severe epidemics of the disease occur.     

Spatio-temporal development of pycnidia and perithecia and dissemination of spores of Mycosphaerella pinodes on pea (Pisum sativum)

The pattern of development of pycnidia and perithecia of Mycosphaerella pinodes was studied in the glasshouse on pea plants (cv. Solara) sprayed with a pycnospore suspension and in field plots inoculated with barley grains colonized by the fungus. The numbers of pycnidia and perithecia were estimated on each stipule and internode of infected plants, and were related to ratings of disease severity (0–5 scale). Pycnidia were produced on both green and senescent organs, whereas perithecia only appeared on senescent organs. The development and quantity of pycnidia were related to initial inoculum concentration and the physiological stage of the plants. The formation of fruiting bodies progressed from the bottoms to the tops of plants during crop development. Spore trapping showed that both pycnospore dispersal and ascospore discharge were initiated by rainfall or dew. Pycnospores were principally trapped in the first 20 cm above the soil surface while ascospores were also trapped above the crop canopy. Pycnospores and ascospores were dispersed throughout the growing season, suggesting that ascospores also play an important role in secondary infections.     

Use of a selective medium to study the dispersal of ascospores ofSclerotinia sclerotiorum

A selective medium (potato dextrose agar with pentachloronitrobenzene and streptomycin) was used to assess dispersal of ascospores ofSclerotinia sclerotiorum in the field. This medium inhibited development ofMucor spp. andRhizopus spp. without affecting the germinability ofS. sclerotiorum ascospores or the production of sclerotia. The number of viable ascospores started increasing in December, was maximal from January to February, and decreased sharply in March. Deposition of ascospores on plates was maximal (57%) between 10:00 and 13:00 hours. Seventy-seven to 90% of the ascospores were deposited within the first 100 m from the source of inoculum and the rest were transported farther.     

Aggressiveness profiling of the coffee pathogen Colletotrichum kahawae

Colletotrichum kahawae is a specialized plant pathogen of arabica coffee in Africa, able to infect green berries. The economic impact of this pathogen means there is an urgent need to better understand its pathogenic lifestyle, in particular its aggressiveness. In this study, several quantitative traits including disease severity, latent period and incubation period were measured to concomitantly assess the aggressiveness of 26 C. kahawae isolates. The results show that the area under disease progression curve is the most informative variable, particularly when joined together with the index disease intensity 10 days after inoculation and latency period, while the incubation period is not a reliable trait to distinguish aggressiveness levels in C. kahawae. This study also confirms the suitability of hypocotyls and detached green berries to perform C. kahawae aggressiveness assays, revealing that hypocotyls are a more reproducible testing material. Based on isolate profiles, three aggressiveness classes were established (high, moderate and low). A cytological analysis of representative isolates from each class showed that aggressiveness can be related to the development of post-penetration stages, rather than conidia germination and appressoria differentiation. This study provides, for the first time, the best metrics to evaluate C. kahawae aggressiveness, characterizing the profile of a broad range of isolates, and defining a set of parameters that can be used to classify new isolates. Furthermore, the collected information will contribute to the improvement of coffee breeding programmes, through the selection of tester isolates for prescreening of resistant coffee materials, and offers the opportunity to engage on future genotype–phenotype studies.     

Management and cross-infectivity potential of Colletotrichum acutatum causing anthracnose on bell pepper in Florida

Early anthracnose caused by Colletotrichum acutatum has become an increasingly serious disease on green, unripe bell pepper fruit in Florida. This contrasts with earlier reports of anthracnose occurring on bell pepper primarily as a ripe-rot disease of mature, colored pepper fruit caused by Colletotrichum gloeosporioides. Management of anthracnose on green bell pepper fruit using fungicides and a commercial inducer of systemic acquired resistance, acibenzolar-S-methyl (ASM), was evaluated during three seasons. In two of the three trials, all the fungicides tested including azoxystrobin, fludioxonil + cyprodinil, mancozeb, famoxadone + cymoxanil, copper hydroxide, and ASM significantly increased the number of marketable fruit compared with control plants. These trials identified fungicides that could contribute to a successful pest management program on pepper for controlling anthracnose caused by C. acutatum. The cross-infectivity potential of C. acutatum was investigated on tomato and strawberry by in vitro and field inoculation. Anthracnose lesions formed readily on wound-inoculated detached fruits of all hosts in in vitro assays. Under field conditions, after inoculation, anthracnose lesions occurred on pepper fruit but no lesions of anthracnose were found on either ripe or unripe tomato or strawberry fruit in adjacent plots.     

苹果炭疽叶枯病病原Glomerella cingulata及其侵染过程

为明确苹果炭疽叶枯病病原菌围小丛壳Glomerella cingulata的侵染致病特征,在分离获得该病原菌的基础上,采用形态学观察、ITS序列分析和致病性测定对其进行了鉴定,并利用光学和扫描电子显微镜对病原菌在嘎啦苹果叶片上的侵染过程进行了研究.结果表明,在陕西咸阳地区分离获得的9株病原菌均为围小丛壳G.cingulata.25 ℃下接种9 h后,分生孢子中间产生隔膜,双胞化,并萌发产生芽管和附着胞;24 h后分生孢子的2个细胞均可萌发并形成芽管及附着胞,部分芽管顶端可产生次级分生孢子;48 h后次级分生孢子萌发形成附着胞;72 h后,附着胞下形成的侵染钉可直接入侵寄主,在表皮细胞内形成初生菌丝和次生菌丝,此时叶片表面已出现褐色斑点.接种7 d后叶片病斑处出现分生孢子盘和子囊壳.表明陕西省近年出现的苹果炭疽叶枯病病原菌为围小丛壳G.cingulata,该病菌在嘎啦叶片上的一些特殊侵染行为可能是导致该病害易在短时间内暴发的重要原因.     

Effect of cultural practices on the development of arabica coffee berry disease,caused by <Emphasis Type="Italic">Colletotrichum kahawae</Emphasis>

In the high altitude regions of Africa, coffee berry disease (CBD), caused by Colletotrichum kahawae, is the main constraint for arabica coffee (Coffea arabica) production. However, certain agricultural practices can reduce losses caused by the disease and thereby promote optimum production. On small family farms in Cameroon, mixed cropping with fruit trees, intercropping with food crops and maintenance pruning of coffee trees are very widespread agricultural practices that can affect CBD epidemics. Consequently, an epidemiological study was conducted to assess how cultural practices affected the disease in an arabica coffee smallholding in Cameroon. The disease was monitored on a weekly basis over four successive years (2002–2005) on coffee trees in diverse cultural situations. Cultural practices likely to reduce losses due to CBD were identified. The infection rate was significantly lower on coffee trees grown intensively than on coffee trees grown in the traditional manner. Coffee trees located under the shade of fruit trees were significantly less attacked than those located in full sunlight. In addition, berries on the leafless parts of branches, near the main trunk of the coffee tree, were less infected than those on leafy sections. These results show that maintenance pruning, removal of mummified berries, and mixed cropping with shade plants are cultural practices which create environmental conditions that limit CBD development.     

Grey Mould Damage and Control in Hungary1

Grey mould is one of the most important fungal diseases of grapevine, and for this reason the successful control of Botrytis cinerea determines the result of grape growing. As far as damage is concerned, the most important infection occurs at the end of flowering and just before ripening. The conidia of the fungus infect the berries through wounds, which may be caused by hail or storms, or by chewing insects, or also, most importantly, by physiological berry splitting. This occurs when a long dry period in summer is followed by a rainy autumn at ripening time. The prevention of wounds is one of the most important aspects of control. For the first half of the year, we propose using the traditional contact fungicides against grey mould. The use of benzimidazole-type systemic fungicides is proposed only for the last two treatments, because they have a low persistence and because in this way we avoid the development of resistance to these systemic fungicides.     

Diversity of <Emphasis Type="Italic">Colletotrichum</Emphasis> species in coffee (<Emphasis Type="Italic">Coffea arabica</Emphasis>) plantations in Mexico

The aim of this study was to identify the Colletotrichum species associated with anthracnose symptoms in coffee (Coffea arabica L.) plantations in northern Puebla, Mexico. In 2013, five surveys were conducted in different production areas and at different altitudes. Symptomatic leaves, shoots, and ripe and unripe fruits of the coffee variety Red Caturra were collected. Isolates were obtained and the Colletotrichum species were identified morphologically and characterized by multilocus sequence analyses of the ACT, CAL, GAPDH, and TUB2 genes and the rDNA region. Additionally, pathogenicity tests were conducted using six isolates. We identified C. gigasporum, C. gloeosporioides, C. karstii (two isolates), C. siamense, and C. theobromicola. This is the first report of these five species infecting leaves of coffee. The symptoms caused by these species were characterized, but the species causing Coffee Berry Disease was not found. This is the first report of a complex of species affecting coffee plants in the same geographical area in Mexico, and suggests that other complexes of species may be important pathogens in coffee-producing areas elsewhere.     

Variation among Colletotrichum gloeosporioides isolates from infected coffee berries at different locations in Vietnam

Isolates of Colletotrichum gloeosporioides associated with anthracnose disease on coffee berries in Vietnam were characterized by morphological and molecular methods. Random amplified polymorphic DNA (RAPD) and microsatellite-primered PCR (MP-PCR) analyses were employed to investigate the genetic variation among 38 and 51 isolates of C. gloeosporioides , respectively. According to both methods, the isolates mainly grouped in accordance with geographical origins. Higher genetic variation ( H  = 0·312 and 0·335) in the northern population of C. gloeosporioides than in the southern population ( H  = 0·261 and 0·186), according to the RAPD and MP-PCR markers, respectively, was indicative of a difference between the northern and southern populations. Moderate gene differentiation ( G st = 0·1) between populations from the north and the south was found. However, there was no differentiation between locations within the northern or southern populations, indicating significant gene flow. A four-gamete test indicated a high level of recombination, particularly in the south. The geographic differences may be explained by different histories of coffee cultivation in different parts of Vietnam. The symptoms caused by the Vietnamese isolates on both hypocotyls and green berries were less severe than symptoms caused by the reference CBD (coffee berry disease; Colletotrichum kahawae ) isolates originating from Africa.     

Role of perithecia as an inoculum source for stem rot type of pepper root rot caused by Fusarium solani f. sp. piperis (teleomorph: Nectria haematococca f. sp. piperis)

Pepper (Piper nigrum L.) root rot caused by Fusarium solani f. sp. piperis (FSP; teleomorph: Nectria haematococca f. sp. piperis) includes two symptom types called root rot (RR) type and stem rot (SR) type. In this study, the temporal and spatial associations between perithecial formation of FSP and development of SR were investigated in naturally infested fields to verify the hypothesis that ascospores from the perithecia are the major inoculum source of the SR type on vines in the field. In surveys of all vines in two neighboring pepper fields every month from December 2005 to November 2006, I mapped the locations of all vines with perithecia and all vines the SR type. The frequency of vines with perithecia increased during April and May, the late rainy season. In June, the early dry season, the number of vines with SR type greatly increased. The vertical range of perithecial formation on the vines extended to 200 cm in height, but was restricted to 30 cm in the dry season in both fields. The join-count statistics showed a significant spatial association between vines with perithecia and vines with SR type in one field (P = 0.042), while no significant spatial association was recognized in another field. The results suggested that ascospores from perithecia of FSP on pepper vines are likely to be one of the main inoculum sources of the SR type of the disease on adjacent vines, but they may not be the exclusive source.     

Combining sanitation and disease modelling for control of grapevine powdery mildew

Chasmothecia of Erysiphe necator form in one season, survive winter and discharge ascospores that cause primary infections and trigger powdery mildew epidemics in the next season. A strategy for powdery mildew control was developed based on (i) the reduction in overwintering chasmothecia and on (ii) spring fungicide applications to control ascosporic infections timed based on estimate risk (two to five sprays per season). Several fungicides, the hyperparasite Ampelomyces quisqualis, and a mineral oil product were first tested as separate applications in a greenhouse and in vineyards. In the greenhouse, A. quisqualis suppressed chasmothecia formation by 41 %; fungicides and mineral oil suppressed chasmothecia formation by 63 % and ascospore viability by 71 %. In vineyards, application of boscalid + kresoxim-methyl or meptyldinocap once after harvest, as well as application of A. quisqualis pre- and post-harvest, delayed disease onset and epidemic development in the following season by 1 to 3 weeks and lowered disease severity (up to the pea-sized berry stage) by 56 to 63 %. Risk-based applications of sulphur and of synthetic fungicides provided the same control as the grower spray program but required fewer applications (average reduction of 47 %). Sanitation strategies were then tested by combining products and application times (late-season, and/or pre-bud break, and/or spring). Adequate disease control with a reduced number of sprays was achieved with the following combination: two applications of A. quisqualis (pre- and post-harvest), one application of mineral oil before bud break, and model-based applications of sulphur fungicides between bud break and fruit set.     

A Serious Disease of Annato Caused by Glomerella cingulata

Abstract

The occurrence of a disease of annato apparently restricted to the pods is reported, and is caused by a Colletotrichum state of Glomerella cingulata.