Lophodermium Piceae Research Articles

Evaluation of foliar fungal endophyte incidence in field-grown transgenicBtwhite spruce trees

A total of 770 transgenic Bt white spruce needles were collected and plated on potato dextrose agar to determine their foliar endophyte diversity. The ribosomal internal transcribed spacer regions for 310 foliar endophytes were amplified by polymerase chain reaction (PCR) and digested using CfoI and MspI, which created 21 restriction groups. Isolates from each restriction group were sequenced and compared with reference sequences in GenBank. Eighteen sequence groups were obtained, of which five were identified at the species level. The most common endophytic fungi identified by PCR-RFLP was Lophodermium piceae (incidence of 74.5%). The second and third most common ones were Hypoxylon fragiforme (3.63%) and Lophodermium nitens (3.18%). A statistical analysis performed on the most common endophyte groups showed no statistical difference in endophyte frequency or distribution between the control white spruce needles (nontransgenic) and saplings with constructs containing the reporter gene GUS or the Bt Cry1A(b) gene and kanamycin.

Biodiversity of foliar fungal endophytes in white spruce (Picea glauca) from southern Québec

The objective of this study was to investigate the foliar endophyte biodiversity of white spruce ( Picea glauca (Moench) Voss to establish a baseline for future comparative studies examining the impact of forestry practices. It identifies for the first time endophytic fungi living inside the needles of white spruce from 280 needles collected in seven natural stands in southern Québec. The endophyte colonization rate on surface sterilized needles was 53.2%. We performed a PCR-RFLP and sequence analysis on the ITS region of the nuclear ribosomal DNA to achieve molecular identification. Isolate ITS sequences were compared with data from GenBank presenting the best similarity and were analyzed by a maximum of parsimony and Bayesian inference. Relationships between morphological groups, digestion groups, and sequence groups were investigated. In all, 23 morphotypes were found to belong to 14 sequence groups and we demonstrated that morphological groups are poor indicators for estimating species diversity. This study is the first to establish species richness values for foliar endophytes. Among the 141 isolates in this study, 75.15% have a high sequence similarity with Lophodermium piceae , 10.95% with an unknown species of Mycosphaerella , and 5.5% with two species of the genus Hypoxylon . Distribution, incidence and biological significance of all the endophytes found in this study are discussed.

Diversity of endophytic fungi of single Norway spruce needles and their role as pioneer decomposers.

The diversity of endophytic fungi within single symptomless Norway spruce needles is described and their possible role as pioneer decomposers after needle detachment is investigated. The majority (90%) of all 182 isolates from green intact needles were identified as Lophodermium piceae. Up to 34 isolates were obtained from single needles. Generally, all isolates within single needles had distinct randomly amplified microsatellite (RAMS) patterns. Single trees may thus contain a higher number of L. piceae individuals than the number of their needles. To investigate the ability of needle endophytes to act as pioneer decomposers, surface-sterilized needles were incubated on sterile sand inoculated with autoclaved or live spruce forest humus layer. The dry weight loss of 13-17% found in needles after a 20-week incubation did not significantly differ between the sterilized and live treatments. Hence, fungi surviving the surface sterilization of needles can act as pioneer decomposers. A considerable portion of the needles remained green during the incubation. Brown and black needles, in which the weight loss had presumably taken place, were invaded throughout by single haplotypes different from L. piceae. Instead, Tiarasporella parca, a less common needle endophyte, occurred among these invaders of brown needles. Needle endophytes of Norway spruce seem thus to have different abilities to decompose host tissues after needle cast. L. piceae is obviously not an important pioneer decomposer of Norway spruce needles. The diversity of fungal individuals drops sharply when needles start to decompose. Thus, in single needles the decomposing mycota is considerably less diverse than the endophytic mycota.

Co‐occurrence of the ascomycete Lophodermium piceae and the rust fungus Chrysomyxa abietis in Norway spruce needles

The frequency of needles and the proportion of needle segments infected by Lophodermium piceae were compared in symptomless and Chrysomyxa abietis‐infected, 1‐year‐old needles of Picea abies. In late spring, symptomless needles from both rust‐infected and healthy saplings were sampled. In addition, rust‐infected, totally chlorotic needles and needles with chlorosis along about half their length from the diseased trees were examined. In all three stands, the proportion of segments infected by L. piceae was larger in the rust‐infected half of the needle than in the symptomless half; but the difference was statistically significant in only one of the stands. The proportion of L. piceae‐infected segments among the nonrust‐infected needles was the same as that found for the uninfected half of rust‐infected needles (after correction for size differences). No differences in the proportion of L. piceae‐infected segments were found between the totally chlorotic, rust‐infected needles and the green needles of diseased or healthy trees.

Seasonal patterns of colonization of Norway spruce needles by Lophodermium piceae

Summary Colonization of living Norway spruce needles by Lophodermium piceae was studied in southern Sweden. One‐month to 3‐year‐old needles were assessed for frequency of infection. Needles appearing in the growing season of 1996 were monitored and sampled at 1 to 4 week intervals over a period of 14 months. Infection frequency of the current‐year needles increased from around 20% in July to approximately 40% in August 1996. This level had then doubled to approximately 80% after another growing season in August 1997. Multiple infections were rare in the needles during the first growing season, but after the second growing season at least 70% of the needles had multiple infections. The average number of infections in 9‐, 21‐ and 33‐month‐old needles were 1.3, 6.3 and 8.4, respectively. The size of the individual mycelia was estimated to be small in all needles, none of the individuals could be isolated from more than two adjacent needle sections, about 1.5 mm long. In conclusion, the number of L. piceae infections per needle increased during ageing of needles but mycelial growth is probably postponed until needle senescence and death.

Effects of Irrigation, Fertilization and Drought on the Occurrence of Lophodermium piceae in Picea abies Needles

Effects of irrigation, drought and fertilization on the frequency of the fungal endophyte Lophodermium piceae in green needles was assessed in a 30-year-old experimental stand of Picea abies in southern Sweden. Frequencies of needles with L. piceae were lower in irrigation and ammonium sulphate fertilization treatments than in the control. Drought treatment frequencies were similar to the control. Needles were susceptible to colonization for at least 3 years; colonization increased with needle age. The results indicate that the increased availability of water to the root system as well as ammonium sulphate fertilization indirectly delays colonization of needles by L. piceae.

Effect of open‐air fumigation with sulphur dioxide and ozone on phyllosphere and endophytic fungi of conifer needles

ABSTRACTThe phyllosphere microbial populations inhabiting the needles of three conifer species, Scots pine (Pinus sylvestris L.), Sitka spruce (Picea sitchensis L.) and Norway spruce (Picea abies (L.) Karst.), exposed to SO2 and O3, in an open‐air fumigation experiment were analysed over a 3 year period using serial dilution after washing, direct plating and a fluorescein diacetate (FDA) enzyme assay. Total fungal populations ranged from 102 to 105 colonyforming units (CPU) g−1 fresh weight of needles. The dominant fungi isolated from needles varied with tree species and isolation technique; Aureobasidium pullulans (de Bary) Arnaud was most common on Scots pine and Norway spruce and white yeasts on Sitka spruce using the dilution plating method. However, direct plating of needle segments onto culture media indicated that Sclerophoma pythiophila (Corda) Hohnel was dominant on Scots pine and A. pullulans on Sitka and Norway spruce. Green needles of Sitka spruce were found to be endophytically colonized by Rhizosphaera kalkhoffii Bubak, but seldom by Lophodermium piceae (Fuckel) Hohn during extensive sampling in 1990. Statistical analyses revealed significant differences (P<0.05) between plots in the 3 year mean of the total fungal populations or the fungal biomass (FDA assay) on all three tree species. Differences between plots were also observed for a number of dominant component species. Data were also analysed for treatment effects. A significant effect of SO2 treatment was observed on the total fungal populations on Sitka spruce (P<0.05) which were reduced markedly by the low‐SO2 treatment, while the O3 treatment caused a significant increase in total fungal numbers on Scots pine (P<0.05). The FDA activity on needles of both Scots pine and Sitka spruce was noticeably higher in the 03‐only treatment plot, but the overall O3 effect was not significant. Treatment effects were also detected on the occurrence of component species. The serial dilution method revealed an SO2 effect (P<0.05) of a reduction in the occurrence of pink yeasts on Sitka spruce and an O3 effect (P<0.05) of an increase in the occurrence of S. pythiophila on Sitka spruce (P<0.01) but a decrease of Epicoccum nigrum Link and Cladosporium spp. on Scots pine. The direct‐plating method revealed an SO2 effect of an increase in S. pythiophila on Norway spruce (P<0.05). Ozone treatment caused a significant increase in the isolation of a black strain of A. pullulans on Norway spruce (P<0.05). Endophytic colonization of Sitka spruce needles by R. kalkhoffii was found to be increased on two occasions by O3 exposure.

Lophodermium piceae and Rhizosphaera kalkhoffii in fallen needles of Norway spruce (Picea abies)

AbstractFalling needles collected from under individual 70‐year‐old Norway spruce (Picea abies) trees for about four years were checked for fungal fruitbodies. The most common fungi were Lophodermium piceae and Rhizosphaera kalkhoffii. Maximum frequencies of fruitbodies occurred in late autumn for L. piceae (ca. 80% of the needles), in summer for R. kalkhoffii (ca. 60%) and in winter for the third most common fungus, Tiarosporella parca (ca. 10%). The frequencies of needles with fruiting fungi varied greatly within and between years. This is the first report of T. parca from Sweden.

Morphological studies of Lophodermium piceae (Fuckel) v. Höhnel on Norway spruce needles

AbstractMorphological structures of the teleomorphous and anamorphous stages of the ascomycete Lophodermium piceae have been studied in detail. This fungal species is widely distributed and frequent on Norway spruce needles. The examination of many samples indicates that all belong to one species, namely L. piceae.

Life cycle of Lophodermium piceae in Norway spruce needles

AbstractThe spruce needle inhabiting ascomycete Lophodermium piceae can live symptomless for several years, before it completes its life cycle within less than one year as detailed studies have shown.

Infektionsversuche mit Rhizosphaera kalkhoffii und Lophodermium piceae an unterschiedlich gestreßten Jungfichten (Picea abies [L.] Karst.)

Young spruce trees [Picea abies (L.) Karst.] artificially inoculated with Rhizosphaera kalkhoffii B. and Lophodermium piceae (F.) v. H. were exposed to fumigations with O3 and SO2 and drought. These treatments showed no influence on the infection-rates of the fungi. R. kalkhoffii proved to be a saprophyte living for a long period as an epiphyte on the needle surfaces.

Necrotic spots induced by simulated acid rain on needles of Abies balsamea saplings

Simulated acid rain (SAR) of pH 2.6, 3.6 and 5.6 (control) was applied to Abies balsamea (L.) Mill. saplings at 2-week intervals for 3 years. Ambient precipitation was excluded from the trees during this period. Light to dark brown necrotic spots along the midribs of needles, 0.1 to 0.5mm in diameter and sometimes having a white center, were associated with the most acidic (pH 2.6) SAR treatment. These spots appeared over a significantly (p = 0.05) larger area of the foliage in the pH 2.6 treatment than in controls (pH 5.6). Needles that received SAR of pH 3.6 had intermediate numbers of spots. The spots persisted for at least 12/3 years after cessation of SAR application. Needle retention, needle lengths, and the occurrence of epicormic shoots and the needle rust caused by Uredinopsis longimucronata Faull were not significantly (p = 0.05) affected by the SAR treatments. Needle retention tended to be lessened by the most acidic (pH 2.6) SAR treatment. The occurrence of Lophodermium piceae (Fuckel) Höhnel needle cast was significantly lower in the middle tree sections of the pH 5.6 and 3.6 treatments than in the pH 2.6, drought and ambient treatments. Damage by naturally occurring insects was negligible, and was not related to the SAR treatments.

Zur Taxonomie und Nomenklatur von Lophodermium piceae (Fuckel) v. Höhnel

AbstractThe systematic position and nomenclature of Lophodermium piceae are reviewed chronologically on basis of references cited since 1799. A critical analysis of the actual name of this fungus species of the ascomycete family of Rhytismataceae is given.

Structure spatio-temporelle de la mycoflore decomposant les aiguilles d' Abies alba: Place de Marasmius androsaceus

The spatio-temporal distribution of seven fungal species colonizing Abies alba Mill, needles during senescence [ Cytospora friesii Sacc., Rhizosphaera sp., Lophodermium piceae (Fuckel) Höhn.) and in the L ( Thysanophora penicillioides (Roum.) Kendrick, Nectria purtonii (Grev.) Berk., Ceuthospora pinastri (Fr.) Höhn.] and F1 litter layers [ Marasmius androsaceus (L.: Fr.) Fr.] has been described. Ecological niches of the seven fungi were separated in the spatio-temporal plane. Marasmius could colonize fir needles whatever the previous successions had been. At the end of Marasmius development, 16 elementary successions could be recognized. Only 4 complete successions were necessary to explain the occurrence of the 7 observed species ( Ceuthospora-Marasmius; Rhizosphaera-Thysanophora Marasmius; Cytospora-Nectria-Marasmius and Lophodermium Marasmius). Other successions could be derived from the complete ones by deletion of one or more species. In naturally fallen litter 24% of the needles showed no evidence of colonization by any of the 7 species studied. Microbial interactions did not explain the quantitative limitation of the colonization by Marasmius (27–74% of the needles in these experiments).

Identification of endophytic hyphae of Lophodermium piceae in tissues of green, symptomless Norway spruce needles by immunoelectron microscopy

Antiserum specific for Lophodermium piceae hyphae was obtained by absorbing a rabbit L. piceae antiserum with hyphal material of different fungal isolates. The specificity of this absorbed antiserum was tested with hyphae of endophytic fungi isolated from green, asymptomatic needles of Norway spruce (Picea abies Karst.) using the on-section immunogold labeling technique. With this specific, absorbed antiserum, a homogenous labeling was obtained with cultured hyphae of L. piceae, whereas all other spruce needle endophytes examined remained practically unlabeled. Insignificant cross-reactivity was also observed with needle tissues. The absorbed antiserum was then applied for the immunoelectron microscopical identification of L. piceae hyphae in situ in infected tissues of green, asymptomatic Norway spruce needles. The function of papillalike structures observed in infected needle mesophyll as well as the role of the hyphal sheath surrounding endophytic L. piceae hyphae are discussed in connection with the interaction between L. piceae and asymptomatic Norway spruce needles.

Endophytische Pilze in Nadeln von gesunden und geschädigten Fichten (Picea abies [L.] Karsten)

Abstract Endophytic fungi in needles of healthy‐looking and diseased Norway spruce (Picea abies [L.] Karsten). Endophytic fungi were isolated from surface‐sterilized, 5 to 6 year old needles of healthy‐looking and diseased trees. Two fungal species could be isolated frequently: Lophodermium piceae and Tiarosporella parca. At any site neither L. piceae nor T. parca could be isolated more frequently from needles of diseased trees. The possible types of relationship between these two fungi and Norway spruce are discussed.

Structure spatio-temporelle de la mycoflore des premiers stades de decomposition des aiguilles d' Abies alba

Mycofloras associated with the decomposition of coniferous needles have been frequently described. Their temporal structure (succession) is well-known, but the study of their spatial structure has been neglected. Spatial diversity of mycofloras can be important for the understanding of litter dynamics if it leads to a diversity of decomposition processes. In this study we show that three fungi [ Cytospora friesii Sacc., Rhizosphaera spp and Lophodermium piceae (Fuckl.) Höhn.] colonized silver fir needles ( Abies alba Mill.) during senescence with mutual exclusion. These species occupied different needles or segments of needles. After needle fall, Thysanophora penicillioides (Roum.) Kendrick developed on green needles that had not been colonized before. This fungus could also succeed on Rhizosphaera spp. Colonization by Cytospora prevented subsequent development of Thysanophora but allowed that of Nectria purtonii (Grev.) Berk. Persistent activity of Lophodermium in litter prevented colonization by Thysanophora. Ceuthospora pinastri (Fr.) Höhn. occurred only in litter, on green, uncolonized needles. These results lead to a description of the progressive differentiation of the spatio-temporal structure of this mycoflora and of the several elementary successions possible in this litter.

Internal hyphae in young, symptomless needles of Picea abies: electron microscopic and cultural investigation

Results from cultural investigations were correlated with scanning electron microscope observations and give an introduction to the interaction of endophytic hyphae of Lophodermium piceae (Fckl.) Höhn. and other fungi with living tissues of green, symptomless needles of Norway spruce (Picea abies Karst.). The frequency of endophytic needle fungi was determined by the culture of surface-sterilized needles harvested from two trees. Lophodermium piceae was found to be the most important colonizer of symptomless needles; the infection rates ranged from about 3 to 7%. The presence of fungal hyphae in living needle tissues of symptomless, current year and older Norway spruce needles was demonstrated by scanning electron microscopy. The observed mycelia were both intercellular and intracellular. The hyphal diameters varied significantly and striking swellings of unknown function were observed along internal hyphae. The observed needle colonization is discussed in connection with the spreading of fungal endophytes in the host tissues during the endophytic phase of L. piceae in Norway spruce needles.

Occurrence and pathogenicity of Lophodermium piceae appearing as an endophyte in needles of Picea abies

Lophodermium piceae (probably conspecific with L. nanakii ) was found to be the predominating endophyte in the laminae of I- to 4-year old needles of Picea abies in southern Sweden. The fungus did not appear to be pathogenic and was more common in young, healthy stands than in stressed diseased ones.

Microscopie de la mycoflore des aiguilles de sapin (Abies alba). III. Marasmius androsaceus

Marasmius androsaceus (L.: Fr.) Fr. spreads in the litter by means of its rhizomorphs. It colonizes needles in autumn, soon after litter fall. Penetration of mycelium into needles occurs through stomata. Rhizomorphs and fruit bodies appear 1 year later, in summer and autumn, emerging from the needles by rupturing the epi- and hypo-dermis. In the first stage of colonization hyphae invade needle tissues in extracellular spaces and along vessels. During a second stage, parenchyma (cellulosic cell walls and residual cytoplasm) is bleached and destroyed. Lignified cell walls are degraded, in particular those of the epi- and hypo-dermis. Marasmius androsaceus activity is restricted to L and F1 litter, reaching a maximum after those of Thysanophora penicillioides (Roum.) Kendrick and Lophodermium piceae (Fuckl.) Höhn. Obvious differences are observed among needles colonized by these three different fungi.