Effect of three species of bacteria on damping-off,root rot development,and ectomycorrhizal colonization of lodgepole pine and white spruce seedlings

Interactions between three species of bacteria (Burkholderia cepacia, Pseudomonas chlororaphis and Pseudomonas fluorescens), an ectomycorrhizal fungus (Paxillus involutus), and three root pathogenic fungi (Fusarium moniliforme, Fusarium oxysporum and Rhizoctonia solani) were studied. Burkholderia cepacia significantly reduced the in vitro mycelial growth of P. involutus, whereas, B. cepacia, Ps. chlororaphis, Ps. fluorescens and P. involutus reduced the mycelial growth of F. moniliforme, F. oxysporum and R. solani. Culture filtrates of B. cepacia, Ps. chlororaphis, Ps. fluorescens and P. involutus reduced conidial germination and increased the formation of chlamydospores of F. moniliforme and F. oxysporum. Burkholderia cepacia also reduced the formation of ectomycorrhizal short roots by P. involutus on lodgepole pine (Pinus contorta) and white spruce (Picea glauca) seedlings 2 months after inoculation. However, no significance difference in mycorrhizal short roots was observed after 4 months. The fungicide Anchor (a mixture of carboxine and thiram) significantly reduced root rot severity and increased the survival of lodgepole pine seedlings grown in a growth mix infested with F. moniliforme, F. oxysporum and R. solani. Control of the diseases of white spruce caused by these pathogens was not as successful. Treatment of seeds with either B. cepacia or P. involutus alone significantly increased the survival of seedlings grown in a mix that was inoculated with F. moniliforme and reduced the root rot severity caused by F. moniliforme and F. oxysporum, but not R. solani. Higher seedling survival and lower root rot severity were observed when conifer seeds were concomitantly inoculated with one of the bacterial species, P. involutus and Anchor.     

Virulence of Fusarium oxysporum and F. commune to Douglas‐fir (Pseudotsuga menziesii) seedlings

Fusarium species can cause damping‐off and root rot of young conifer seedlings, resulting in severe crop and economic losses in forest nurseries. Disease control within tree nurseries is difficult because of the inability to characterize and quantify Fusarium spp. populations with regard to disease potential because of high variability in isolate virulence. Fusarium isolates were collected from healthy and diseased seedlings of Douglas‐fir (Pseudotsuga menziesii) and western white pine (Pinus monticola) from a nursery in Idaho, USA. Molecular markers such as DNA sequences (mitochondrial small subunit and nuclear translation elongation factor 1‐alpha) and amplified fragment length polymorphism were used to identify isolates as either F. oxysporum or F. commune. In addition, diagnostic primers were developed to detect and distinguish F. commune from F. oxysporum. In vitro and greenhouse virulence tests were completed on Douglas‐fir germinants and seedlings. For Douglas‐fir germinants and seedlings, F. oxysporum isolates generally caused less severe symptoms, whereas most F. commune isolates caused mortality through damping‐off. This is the first report of direct evidence that F. commune can cause damping‐off disease on Douglas‐fir seedlings under greenhouse conditions.     

Association of seed‐borne Fusarium species on Pinus ponderosa with germination and seedling viability in Argentina

Pinus ponderosa seedlots from eight seed orchards in Western Patagonia, Argentina, were evaluated for the presence of Fusarium species, and isolates obtained were subjected to pathogenicity tests. The following species were isolated: Fusariumoxysporum, Fusarium equiseti, Fusarium verticillioides, Fusarium incarnatum, Fusarium acuminatum, Fusarium graminearum and Fusarium proliferatum. With the exception of F. proliferatum and F. graminearum all Fusarium species were recovered from P. ponderosa seedlings with symptoms of damping‐off. Five Fusarium species were re‐isolated from necrotic roots of seedlings showing chlorosis (F. oxysporum, F. equiseti, F. incarnatum, F. acuminatum and F. verticillioides). High damping‐off losses occurred in Fusarium‐contaminated seed of P. ponderosa and low disease incidence occurred on less contaminated seed. The role of seed‐borne inoculum in disease spread within forest nurseries is discussed.     

Fungi Isolated from Diseased Nursery Seedlings in Spain

Fungal endophytes in container-grown seedlings in forest nurseries located in the North Plateau of Spain have been studied. Influences of nursery, season (spring and autumn), host genus and plant part on those fungal assemblages have been also evaluated. Four nurseries were sampled in spring and three in autumn. Analysed material consisted of stems, roots and rhizosphere from five host species, Quercus ilex, Q. pyrenaica, Pinus pinea, P. nigra and P. sylvestris. A total of 581 isolates corresponding to 13 taxa were found in this study. Fusarium moniliforme, F. oxysporum and Trichoderma viride were the most frequently isolated fungi. Number of fungal species per observation differed significantly among the sampled parts. Multivariate Correspondence Analysis allowed the discrimination of samples in five groups based on nursery, season, host genus and plant part. Isolation frequencies of F. moniliforme and F. oxysporum were higher in the roots and in spring, whereas those of T. viride were higher in the rhizosphere and in autumn. A relationship was observed between presence of T. viride and absence of Fusarium spp. in forest nurseries. T. viride, could serve as a biological control agent against Fusarium spp. in forest nurseries.     

Seedborne Fusarium on Douglas-fir: Pathogenicity and seed stratification method to decrease Fusarium contamination

Twelve Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) seedlots from coastal British Columbia were assayed for seedborne Fusarium. All of the seedlots were contaminated with Fusarium. Percent of nonstratified seeds from individual seedlots harboring Fusarium ranged from 0.3% to 95.4%. Sixty-seven percent of the seedlots had Fusarium on less than 2% of the seeds. Post-stratification seedborne Fusarium levels were significantly less for running water imbibition compared to standing water imbibition. However, seedling growth at a container nursery was not consistently different for stratified seed imbibed initially in standing or running water. Fusarium disease symptoms were not observed in the nursery environment. The species of Fusarium isolated from seed were F. acuminatum, F. avenaceum, F. lateritium, F. moniliforme, F. oxysporum, F. poae and F. sambucinum. Twelve Fusarium isolates, comprising six species, were assessed for pathogenicity. Disease symptoms were observed after four weeks incubation and Fusarium isolates ranged in virulence from low to high. Fusarium oxysporum isolates were the most pathogenic.     

Fusarium spp. and Pinus strobus seedlings: root disease pathogens and taxa associated with seed

Eastern white pine (Pinus strobus L.) seeds weresown in soil infested with Fusarium proliferatum, rootnecrosis developed on seedling roots, and F. proliferatumwas reisolated from symptomatic roots; thus, demonstrating thatF. proliferatum is pathogenic toeastern white pine seedlings. Soils infested with F.acuminatum or F. sporotrichioides resulted infew diseased seedlings. Seedlings with root rot generally showed reductions inseedling height. All Fusarium species tested wererecoveredfrom rhizosphere soil samples. Three seedlots of Pinusstrobus were examined for Fusarium infestation.Fusarium species were recovered from most seeds in twoseedlots. Fusarium proliferatum and F.sporotrichioides were the predominant species isolated. Additionalspecies not previously reported from P. strobus included:F. acuminatum, F. chlamydosporum,F. equiseti, F. graminearum,F. moniliforme, F. poae, F.polyphialidicum, F. heterosporum, F.sambucinum, and F. semitectum.     

Effect of post-planting inoculation with Bradyrhizobium sp and mycorrhizal fungi on the growth of Brazilian rosewood,Dalbergia nigra Allem. ex Benth., in two tropical soils

Six months after planting, seedlings of the leguminous treeDalbergia nigra, known as Brazilian rosewood, wereinoculated with a selected Bradyrhizobium strainBHICB-DN 15,either alone or in combination with mycorrhizal fungi to determine their effecton the growth of D. nigra in Atlantic forest andeucalyptussoils. Height growth of D. nigra was similar in both soilsand six months after inoculation Bradyrhizobium did notaffect the D. nigra height growth, but it did improve drymass and especially in the nitrogen content of plants grown in eucalyptus soil.The success of the delayed inoculation with BHICB-DN 15 confirms its competitiveability vis-à-vis indigenous soil rhizobia. Co-inoculationof BHICB-DN 15 and mycorrhizal fungi did not increase plant dry weight,nitrogenand phosphorus content or mycorrhizal colonization. These results suggest thatthe BHICB-DN 15 strain had no synergistic relationship with the mycorrhizalfungi or that there was incompatibility between symbionts, in both soils.     

Effect of an ectomycorrhizal fungus,Laccaria laccata,on Fusarium damping-off in Pinus banksiana seedlings

Damping-off caused by Fusarium oxysporum Schlecht in Pinus banksiana Lamb, was reduced significantly when inoculated with an ectomycorrhizal fungus, Laccaria laccata (Scop. ex Fr.) Berk. and Br. In paired culture, growth of F. oxysporum was significantly reduced by L. laccata. The number of colony forming units of F. oxysporum was reduced significantly in the rhizosphere of P. banksiana seedlings when inoculated with L. laccata. Spore germination and germ tube length of F. oxysporum was inhibited strongly by culture filtrate of L. laccata and root exudate of P. banksiana inoculated with L. laccata. Mycorrhizal seedlings had significantly higher amount of total soluble phenols than nonmycorrhizal ones.     

Influence of Microbial Inoculations on Cedrus deodara in Relation to Survival,Growth Promotion and Nutrient Uptake of Seedlings and General Soil Microflora

Abstract

Selected microorganisms (Laccaria laccata, Trichoderma viride, Bacillus subtilis, Pseudomonas corrugata and Azotobacter chroococcum) were used as inoculants to study their influence on seed germination, seedling survival and subsequent growth in Cedrus deodara. The microbial inoculations positively affected these parameters over control, in the following order: Azotobacter chroococcum Pseudomonas corrugata Bacillus subtilis Trichoderma viride Laccaria laccata.The major cause of seedling mortality was found to be (1) wilting of emerged seedlings by Fusarium oxysporum, and (2) cutworm attack. Bacterial inoculations were found to be superior in suppressing the proliferation of Fusariumsp., thereby enhancing seedling survival. One of the bacterial inoculants, B. subtilis, resulted in 76% seed germination, compared to 54% in control. Though inoculation with L. laccataresulted in maximum mycorrhizal infection (up to 72%) compared to48%in control seedlings, it afforded least protection against Fusariumsp., as well as minimal growth promotion. On the other hand, inoculation with T. viridesignificantly controlled wilting of seedlings and improved plant growth. Microbial inoculations were found to influence the soil microflora, nutrient status of the rhizosphere soil and that of different parts of seedlings. Enrichment of carbon was recorded in the rhizosphere soil. Enhanced nitrogen, phosphorus and potassium content of various plant parts indicated a positive influence also on nutrient uptake. The present study has implications for large scale forestry programs.     

Pathogenicity of Fusarium verticillioides and Fusarium oxysporum on Pinus nigra seedlings in northwest Spain

Fusarium verticillioides may be responsible for causing significant damping-off damage similar to that incited by F. oxysporum on forest seedlings, resulting in considerable losses in nurseries in northwest of Spain. Traditionally, F. oxysporum has been considered the most important agent of this disease in Spanish forest nurseries. However, recent studies have showed that F. verticillioides also has been frequently isolated from diseased plants. This has increased the necessity for a more comprehensive knowledge of the behaviour and pathogenicity of both Fusarium spp. isolated from these sites. The effect of Fusarium spp. on seed germination and on seedling mortality was analysed by inoculating the fungus at seeding. The in vitro growth of the two species was studied and is discussed in relation to in vivo virulence. Both species caused a reduction in seed germination and an increase in seedling mortality. Mortality caused by F. verticillioides treatments occurred sooner than that for F. oxysporum and the growth rate of F. verticillioides was also greater.     

Seed source and vesicular-arbuscular mycorrhizal symbiont affects growth of Juglans nigra seedlings

Seedlings of Juglans nigra from three seed sources were grown in fumigated soil without vesicular-arbuscular mycorrhizal (VAM) fungi or inoculated with Gigaspora margarita, Glomus deserticola or Glomus etunicatum. Vesicular-arbuscular mycorrhizal development varied significantly between fungal symbionts within a black walnut source. Glomus deserticola and G. etunicatum produced the highest levels of root colonization in all sources. Significant differences in seedling shoot and root growth were attributed to root colonization by specific VAM fungi in each black walnut source. Glomus deserticola stimulated seedling leaf area and root weight 26 and 52%, respectively, in one seed source. Seedling leaf N, P and K concentrations were significantly improved by VAM in two seed sources. Juglans nigra seedlings respond favorably to VAM colonization. However, differences between seed sources suggest a strong host-symbiont interaction.     

Selection of efficient VA mycorrhizal fungi for Casuarina equisetifolia—second screening

An investigation was carried out to screen and select efficient vesicular arbuscular mycorrhizal (VAM) fungi for inoculating the forest tree species, Casuarina equisetifolia. The seedlings were inoculated with 10 different VAM fungi, obtained from various sources. Inoculated seedlings generally had greater plant height, stem girth, biomass and P content than uninoculated plants. They also had more mycorrhizal root colonization and spore numbers in root zone soil. C. equisetifolia seedlings responded best (in biomass) to inoculation with Glomus mosseae (Nicolson and Gerdemann) Gerdemann and Trappe, closely followed by Acaulospora laevis Gerdemann and Trappe and G. fasciculatum (Thaxter Sensu Gerdemann) Gerdemann and Trappe; all the three being statistically on par with each other.     

Aggressiveness of Fusarium oxysporum and F. verticillioides isolates on stone and scots pine under greenhouse conditions

Scots pine (Pinus sylvestris) and Stone pine (Pinus pinea) are two important species used in re-forestation that are subject to damage by damping-off fungi in forest nurseries. Twenty-two isolates of Fusarium oxysporum and F. verticillioides from diseased seedlings of eight different hosts were tested for aggressiveness on seeds and seedlings of both pine species, including the effects on seedling emergence and mortality. Scots pine was more susceptible to damping-off than Stone pine, as indicated both by reduced seedling emergence and elevated seedling mortality. The impact of F. oxysporum and F. verticillioides on seeds and seedlings did not differ significantly for either pine species. Our findings support previous studies that found that these are damping-off pathogens on the studied pines. Whereas most isolates proved to be pathogenic, some isolates of both Fusarium species showed to be non-pathogenic.     

Ectomycorrhizal inoculation of Douglas-fir transplanted container seedlings with commercially produced inoculum

Commercially produced vegetative inocula of Laccaria laccata and Hebeloma crustuliniforme successfully formed ectomycorrhizae with Douglas-fir transplanted container (plug+1) seedlings. After 4.5 months in containers, 83% and 90%, respectively, of short roots were mycorrhizal. L. laccata- or H. crustuliniforme-inoculated seedlings had significantly more mycorrhizal and total short roots than Pisolithus tinctorius-inoculated (4% mycorrhizal root tips) or uninoculated control seedlings. No significant differences were detected in seedling growth at the end of the container phase.After transplantation and growth in nursery beds for 17 months, mean new short root colonization of all seedlings was 80%. H. crustuliniforme persisted as a dominant mycorrhizal fungus on seedlings initially inoculated with this fungus. L. laccata-inoculated seedlings had 40% of their short roots colonized by L. laccata and another 40% by native fungi Rhizopogon and Thelephora spp. All mycorrhizae of control seedlings and those inoculated with P. tinctorius were formed by fungi native to the nusery beds. A significant fungal treatment effect was detected for shoot height only. Control seedlings were significantly taller than L. laccata-inoculated seedlings after transplanting.This article is part of senior author's thesis in partial fulfillment of the Ph.D. degree in the Department of Forest Science at Oregon State University, Corvallis.     

Fusarium root infection of container-grown Douglas-fir: effect on survival and growth of outplanted seedlings and persistence of the pathogen

Greenhouse-grown Douglas-fir (Pseudotsuga menziesii var.glauca [Beissn.] Franco) seedlings with roots infected with eitherFusarium oxysporum Schlect. orFusarium proliferatum (Matsushima) Nirenberg were outplanted on a forest site in northern Idaho, U.S.A. No residentFusarium populations were detected in the forest soil.Fusarium persisted the first four years on roots initiated during the greenhouse phase, but occurred sparingly or was absent on roots that grew after outplanting. Height growth was unaffected, and mortality was not often associated withFusarium. A seasonal pattern ofFusarium activity was observed. Low levels (10–40%) of initial root infection apparently have little adverse effect on outplanting performance of Douglas-fir seedlings.University of Idaho, Idaho Forest, Wildlife and Range Experiment Station Contribution No. 640.     

Effects of ectomycorrhizal fungi on damping‐off and induction of pathogenesis‐related proteins in Pinus tabulaeformis seedlings inoculated with Amanita vaginata

The formation of mycorrhiza on Pinus tabulaeformis (Pt) seedlings by ectomycorrhizal fungi (ECMF), the protective effect of the ECMF on damping‐off caused by Rhizoctonia solani and the expression of pathogenesis‐related proteins (PR‐proteins) induced by Amanita vaginata and/or R. solani in Pt seedlings were investigated. The results indicated that: (i) among the 10 ECMF tested, five species (Gomphidius viscidus, Suillus laricinus, Suillus tomentosus, A. vaginata and Basidiomycetes no. 058) formed mycorrhiza with Pt seedlings. There was a positive correlation between mycorrhizal colonization levels and protective effects (r = 0.9207). Among the five species, A. vaginata had the highest colonization level and the best protective effect against damping‐off; however, G. viscidus had no protective effect even though it had a high mycorrhization level. (ii) Chitinases, β‐1,3‐glucanases and PR‐proteins in the seedlings were not induced by A. vaginata colonization, but were induced when A. vaginata‐mycorrhizal seedlings were attacked by R. solani. It is possible that chitinases, β‐1,3‐glucanases and 108, 97, 78, 44, 37 and 14‐kDa PR‐proteins induced both by A. vaginata and R. solani protect Pt seedlings from R. solani infection.     

板栗接种食用菌根真菌的菌根化效果及对幼苗促生效应的研究

[目的]通过板栗幼苗人工接种食用菌根菌固体菌剂,分析其对板栗幼苗生长指标和根系菌根化的效果,以验证大红菇(菌株编号2014-10)、红绒盖牛肝菌(菌株编号2014-14)和美味牛肝菌(菌株编号50559)这3种食用菌根菌固体菌剂作为板栗接种体的有效性及其促生效应,为进一步探索"板栗食用菌根菌新型经济林栽培模式"提供基础。[方法]在板栗播种时分别接种上述3种食用菌根菌固体菌剂,并设空白作对照。对当年生幼苗苗高、地径、根系干质量、侧根数、侧根总长、幼苗侵染率、吸收根侵染率等指标进行调查和分析,并制作菌根石蜡切片进行微观结构观察。[结果]表明:(1)上述3种食用菌根菌在人工接种下均能侵染板栗幼苗根系,平均幼苗菌根化率分别达95.8%、87.0%和96.7%,平均吸收根菌根化率分别达57%、53%和50%,达较高的侵染强度;经微观观察,板栗幼苗根系均形成典型的菌根结构——菌丝套和哈蒂氏网。(2)与对照幼苗相比,3个菌种处理幼苗的平均苗高分别提高32.5%、24.0%和22.7%,平均地径分别提高19.4%、14.1%和5.0%,差异显著(P0.05);平均根系干质量分别提高18.9%、25.3%和14.1%,平均侧根数分别提高16.0%、14.1%和12.1%,平均侧根总长分别提高26.8%、28.9%和44.8%,差异极显著(P0.01)。[结论](1)上述供试食用菌根菌均与板栗根系建立了共生关系,幼苗菌根化效果良好,吸收根菌根化强度高,形成了菌根的典型结构,即菌丝套和哈蒂氏网,验证了上述菌种固体菌剂的有效性,认为其可以作为板栗食用菌根菌栽培的接种体而用于生产实践,为探索"板栗食用菌根菌新型经济林栽培模式"提供了基础。(2)上述供试菌种接种后,对板栗幼苗各项主要生长指标均有不同程度的促进作用,说明上述食用菌根菌对板栗幼苗具有显著的促生效应。     

Field survival of containerized red and jack pine seedlings inoculated with mycelial slurries of ectomycorrhizal fungi

Red pine and jack pine seedlings growing in styroblocks were inoculated 8 wk after sowing with mycelium/agar slurries of 3 mycorrhizal fungi (Laccaria bicolor, Scleroderma citrinum, and an unidentified basidiomycete), and one suspected mycorrhizal fungus (Cantharellula umbonata). Seedlings inoculated with L. bicolor developed mycorrhizae earlier and in greater numbers than the other inoculation treatments, with red pine out-performing jack pine in both respects. At 34 wk following sowing, seedlings were outplanted on a cleared xeric site in Baraga Co., in Michigan's Upper Peninsula. Seedlings inoculated with C. umbonata failed to form mycorrhizae and were not outplanted. Inoculation treatments did not affect shoot or root weight at outplanting. Red pine inoculated with L. bicolor averaged 21% and 19% greater survival compared with control seedlings after one and two years in the field, respectively. Other inoculation treatments failed to increase seedling survival for either tree species. Jack pine demonstrated higher overall survival than did red pine for both years in all corresponding inoculation treatments.     

Gliocladium virens in an alginate prill ineffective as a biological control of Fusarium root disease in container-grown Douglas-fir

An alginate prill formulation of Gliocladium virens (GL-21) was added as a top-dressings (54 g per m²) or incorporated into medium (1.2 kg per m³) used to grow Douglas-fir seedlings in styrofoam containers. Seedlings in the top-dress treatment were similar to control seedlings; infection and colonization by naturally-occurring Fusarium was unaffected by treatment. Incorporated G. virens reduced seedling growth and increased occurrence and colonization intensity of Fusarium. In a laboratory experiment, inoculating Douglas-fir seedlings with G. virens (10% w/w) prior to inoculation with Fusarium increased survival time when compared to concurrent inoculations of fungi.University of Idaho, Idaho Forest, Wildlife and Range Experiment Station Contribution No. 742.     

Root diseases in bareroot and container nurseries of the Pacific Northwest: epidemiology, management, and effects on outplanting performance

In forest and conservation nurseries in the Pacific Northwest USA, seedling production can be limited by root diseases caused by fungi in the genera Fusarium Link:Fr., Cylindrocarpon Wollenw., Phytophthora de Barry, and Pythium Pringsh. Fusarium, Cylindrocarpon, and Pythium are the most ubiquitous, whereas incidence of Phytophthora is mostly associated with coastal bareroot nurseries. All of these root pathogens are encouraged by water saturated soils or media. Seedlings infected with Fusarium, Phytophthora, or Pythium often appear chlorotic or necrotic with extensive root decay. Cylindrocarpon often causes serious root decay without shoot symptoms. The best approach to mitigate losses from these diseases is to use a holistic integrated pest management program. This program should combine chemical controls with cultural practices, particularly those that increase soil permeability and drainage and reduce potential sources of inoculum, especially by disinfesting seeds and containers reused for crops. In general, we found that seedlings meeting nursery specifications for outplanting on forest soil (proper height, root collar diameter, healthy shoot color, lack of disease symptoms) but having these disease organisms on their root systems perform as well as non-infected seedlings.