First report of flyspeck of mango caused by <Emphasis Type="Italic">Stomiopeltis</Emphasis> sp. in Japan

Flyspeck symptoms were found on mango (Mangifera indica L.) in Okinawa Prefecture, Japan in June 2014. Just before harvest, surfaces of fruits and green branches developed groups of tiny black dots within dark smudges. A fungus was isolated from the dots on both fruits and branches. The isolates were identified as Stomiopeltis sp. based on pathogenicity, morphology and molecular phylogenetic analyses with rDNA-ITS and LSU sequences. This is the first report of flyspeck (susuten-byo in Japanese) on mango caused by Stomiopeltis sp. in Japan.     

Construction of a recombinant strain of <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> producing both phenazine-1-carboxylic acid and cyclic lipopeptide for the biocontrol of take-all disease of wheat

Four Neofabraea species are responsible for bull’s eye rot, which is an important postharvest disease of apples and pears. The species diversity of its causal agents in Europe has not been thoroughly explored using molecular genetic methods. Eighty-one Neofabraea isolates were obtained mostly from apples with bull’s eye rot symptoms in the Czech Republic over a two year period. The isolates were identified using PCR fingerprinting and DNA sequencing of the ITS rDNA region, the mitochondrial SSU rDNA and the β-tubulin and EF1α genes. The most common species was N. alba (89 %), followed by N. perennans (5 %) and N. kienholzii (5 %). This is the third published record of N. kienholzii in Europe. The species identity of the isolate CPPF507, which was placed close to N. kienholzii, remains unclear. EF1α was shown to be a suitable marker for the identification of species of the genus Neofabraea and was comparable to the previously used β-tubulin gene. Furthermore, the aggressiveness of individual species was compared and species distribution across Europe was summarized. N. perennans and isolate CPPF507 proved to be the most aggressive, whereas the least aggressive was N. kienholzii. Two N. alba isolates isolated from symptomless apple fruits and leaves were pathogenic to apples in the infection tests.     

Secondary spread of Zygophiala wisconsinensis on the surface of apple fruit

Although the fungus Zygophiala wisconsinensis is a prominent component of the sooty blotch and flyspeck (SBFS) disease complex of apple, few studies have characterized in situ morphological adaptations of this or other SBFS species that accompany colonization of the epicuticular wax layer of fruit. Scanning electron microscope was used to monitor stages in the formation of sclerotium-like bodies on the wax surface, as well as ramifications from these structures to form secondary sclerotium-like bodies. At a mature stage in the formation of sclerotium-like bodies, the matrix of hyphae, conidiophores, and conidia between these bodies collapsed, resulting in formation of the independent clusters of typical colonies of species in the flyspeck mycelial type. Formation of conidiophores and secondary conidia in situ were also described for the first time for any flyspeck species. Observation of conidiophores on naturally and artificially inoculated apple peels with light microscope raises the possibility that formation of secondary conidia may be an important means of spatial dispersion of Z. wisconsinensis in apple orchards and the conidia might function in secondary spread.     

Phylogenetic Analysis of Sugarcane Rusts Based on Sequences of ITS, 5.8 S rDNA and D1/D2 Regions of LSU rDNA

Phylogenetic analysis of sugarcane rusts based on sequences of ITS and the 5.8 S rDNA revealed two highly divergent ITS groups among isolates of Puccinia sp. sensu Muta, 1987 and P. kuehnii specimens. Although there is sufficient divergence (exceeding normal intraspecific variation) between the ITS regions of the two groups to support separation into different species, unusually high homology of the ITS group I sequences with those of members of Cronartium and identical sequences of the D1/D2 regions of the LSU rDNA for all the isolates of “Puccinia sp.” and P. kuehnii that otherwise exhibited different ITS sequences, suggest that the two highly divergent sequences may have resulted from abnormal genetic events leading to non-orthologous, intraspeciflc polymorphisms. The other sugarcane rust, P. melanocephala and the grass rusts, P. miscanthi and P. rufipes, were separated from “Puccinia sp.” and P. kuehnii and from each other in D1/D2 region analyses, indicating that D1/D2 region sequences may more correctly reflect phylogenetic relationships in these rusts than do the ITS regions. Further studies to examine differences in patho-genicity or finer morphological features within P. kuehnii that may be correlated with the high divergence in ITS sequences and experiments to determine if these two sequence types represent intraspeciflc polymorphism are necessary. Received 11 October 2000/ Accepted in revised form 24 November 2000     

<Emphasis Type="Italic">Fibulorhizoctonia psychrophila</Emphasis> is the causal agent of lenticel spot on apple and pear fruit in the Netherlands

In a survey for postharvest diseases of apples and pears in the Netherlands, an unknown postharvest fruit rot was observed. The disease appeared to originate from infected lenticels. A fungus was consistently isolated from the decayed fruits. The fungal pathogen was isolated on potato dextrose agar, and at low temperatures development of a fast-growing whitish mycelium was observed. Growth of this fungus was observed between 1 and 20 °C with an optimum at 15 °C, while incubation of mycelium at 25 °C resulted in no growth. The isolates did not produce asexual or sexual spores. The isolates were characterized and identified by morphology and molecular phylogenetic analysis. Genomic DNA was isolated and amplified using ITS1-ITS4, EF1 and RPB2 primers, and BLAST searches in GenBank placed the fungus taxonomically in the genus Fibulorhizoctonia, with the highest matches to F. psychrophila. Pathogenicity of representative isolates from apple and pear fruit was confirmed under laboratory conditions. To the best of our knowledge this is the first report of F. psychrophila causing lenticel spot on apple and pear, and also the cause of a whitish mould on storage bins.     

Ribosomal DNA Sequence Comparisons of Colletotrichum Graminicola from Turfgrasses and other Hosts

The 5.8S ribosomal gene and the flanking internal transcribed spacers (ITS) 1 and 2 from Colletotrichum graminicola isolates causing anthracnose disease of Agrostis palustris and Poa species were sequenced. Although bootstrap support was not high, two major groups were observed with both UPGMA and parsimony algorithms, one containing isolates from A. palustris and another with isolates from Poa spp. The ITS sequences were also compared with those of isolates of C. graminicola and C. sublineolum from Sorghum spp., Zea mays and Rottboellia cochinchinesis as well as other Colletotrichum species. Except for one isolate from P. annua in Texas, the ITS1 and ITS2 sequences of turfgrass isolates always grouped separately from C. graminicola or C. sublineolum from non-turfgrass hosts with high bootstrap support. ITS sequences of the turfgrass isolates were more similar to those of other species of Colletotrichum, such as C. coccodes and C. dematium, than they were to C. graminicola isolates from other hosts. Turfgrass isolates have ITS sequences which are not identical to those of isolates from Zea mays and Sorghum species demonstrating diversity among fungi conventionally classified as C. graminicola.     

Characteristic of Monilinia spp. fungi causing brown rot of pome and stone fruits in Poland

Brown rot, caused by fungi belonging to the genus Monilinia, is one of the most important diseases of stone and pome trees in the world. During the summers of 2010 and 2011, a total of 670 Monilinia spp. isolates were obtained from infected fruits. They were collected from different commercial stone and pome fruit orchards, located in northern, southern and central Poland. All isolates were identified using multiplex PCR. Twenty isolates obtained from plum, peach and apple fruits were identified as M. polystroma and 5 isolates from plums as M. fructicola. The remaining isolates were identified as M. fructigena or M. laxa. The identification of the isolates was also confirmed on the basis of growth characteristics in culture according to the EPPO standard PM 7/18. A comparison of morphological features of four Monilinia spp. growing on two selective growth media, APDA-F500 and CHA, indicated significant differences between these species. In artificial inoculation of fruits, all the examined Monilinia spp. isolates were pathogenic. The species affiliation of M. polystroma and M. fructicola isolates collected from orchards in Poland was confirmed on the base of phylogenetic and sequence analysis of the internal transcribed spacer (ITS1/5.8S rDNA/ITS2) region of ribosomal DNA.     

Occurrence of soilborne Phytophthora species in declining broadleaf forests in Hungary

Invasive Phytophthora species are responsible for severe tree diseases in many forest ecosystems in Europe. In Hungary, the symptoms were first noted when P. alni infection led to a serious decline and mortality of alder stands in the late 1990s. Between 2001 and 2009, over 300 soilborne Phytophthora isolates were collected from declining broadleaf forests in Hungary, and 10 Phytophthora species identified based on morphological traits and the molecular characteristics of the internally transcribed spacer (ITS) regions of the nuclear ribosomal DNA. The most diverse species spectrum, found in diseased alder stands, included P. gonapodyides, P. gregata, P. inundata, P. lacustris, P. megasperma, P. plurivora, one informally designated taxon: P. taxon hungarica, and one unnamed species P. sp.1. P. cactorum and P. plurivora isolates were prevalent in the soil of a declining eastern black walnut forest, and three species, P. gonapodyides, P. multivora and P. plurivora were recovered from a declining oak stand. More than one ITS-based genotype was identified for four species, including six genotypes for P. gonapodyides, and two each for P. cactorum, P. plurivora and P. inundata. The high genetic diversity of the P. gonapodyides isolates may indicate that the species is indigenous to the region. In contrast, the frequently recovered, widely distributed P. lacustris with a single ITS genotype may represent a recent colonizer. The P. multivora isolates are, to date, the first reported from a European native forest.     

Susceptibility of cider apple cultivars to the sooty blotch and flyspeck complex in Spain

Cultivar may be an important factor associated with the occurrence and severity of sooty blotch and flyspeck (SBFS) fungi in apple. The aim of the study was to analyse the influence of 28 cultivar features on the occurrence of the SBFS complex in apple and to categorize the importance of each using a multivariate approach. Features were classified as agronomic (related to fruiting and growing behaviour and to susceptibility to other diseases; 11 features), morphological (describing fruit morphology; 11) and technological (related to the chemical composition of the fruit juice; 6). Susceptibility to SBFS varied widely among cultivars. The predictive value of some features was not the same for the signs of SB and FS, possibly because each sign is produced by a different species complex which may behave differentially against environmental variables, including the cultivar. Our results showed that agronomic features as a whole were more significant than morphological or technological ones for explaining the cultivar variability in damage. Those related to fruit development phenology were the more explanatory among the agronomic features: late-maturing cultivars suffered higher damage. The intensity of colour was the most explanatory variable among the features describing fruit morphology, showing that SBFS severity was greater on light-coloured apples. Russeting level or skin texture, amongst other variables, were significant but less predictive. The technological features were least explanatory: FS damage was favoured by lower sugar content and higher phenol content at harvest, whereas variability in SB signs was not explained by any juice parameter. The multivariate approach allowed us to select the most important variables in relation to SBFS damage and to discard those without influence, which may be a valuable contribution to understand SBFS development. This research highlights the need of a proper choice of cultivar to reduce disease severity.     

Characterization of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">melongenae</Emphasis> isolates from Turkey with ISSR markers and DNA sequence analyses

Fusarium oxysporum f. sp. melongenae (Fomg), causal agent of Fusarium wilt of eggplant, is a serious pathogen in open fields and greenhouses. Inter-simple sequence repeat (ISSR) banding profiles, sequence analyses of inter-transcribed-spacer (ITS), translation elongation factor 1-alpha (TEF-1α), and actin (actA) DNA regions were employed in this study to determine genetic diversity and population structure of Fomg isolates obtained from Turkey. For ISSR study, (ACTG)₅, (GACAC)₃, (GACA)₄, (GATA)₄, HVH(TG)₇ and (CA)₈RG primers were selected from a set of 16. Discriminative ability of the primers revealed with various indices including polymorphic information content (PIC), and mean PIC value was calculated as 0.26. The ISSR data revealed 31 loci belonging to 202 Fomg isolates and 14 of them were found to be polymorphic. The isolates on neighbor joining ISSR tree were grouped into two major clusters which separated Fomg and outgroup isolates. Population structure was investigated based on bayesian modeling and results indicated five subpopulations (K = 5, ?K = 205.42). Mean genetic and geographical distances among sampling locations revealed only a weak and insignificant correlation (r = 0.583, P = 0.06). Phylogenetic analyses were carried out with ITS, TEF-1α and actA DNA regions with a selected subset of 30 Fomg, along with one non-host and one outgroup isolates. Since ITS region were not able to provide a meaningful separation, TEF-1α and actA sequences of each organism were concatenated individually to build a dendrogram. The clustering tree successfully separated the Fomg, non-host and outgroup isolates in which all Fomg were located on the same branch, forming a monophyletic group in the dendrogram.     

Two new species,Phytophthora nagaii sp. nov. and P. fragariaefolia sp. nov., causing serious diseases on rose and strawberry plants,respectively, in Japan

A new disease of rose was noticed in Chiba Prefecture of Japan in 1968, and the pathogen was initially identified as Phytophthora megasperma based on morphological characteristics. Similar Phytophthora isolates have since been collected from rose plants in Chiba, Kanagawa, and Shizuoka Prefectures. In 2005, several Phytophthora isolates were recovered from crowns of strawberry plants in Hokkaido Prefecture. These were considered to be members of a new species. In this study, we re-examined all these isolates using morphological and physiological studies and a multilocus phylogenetic analysis. The rose and strawberry isolates were mostly similar morphologically and physiologically, with some exceptions. The rose isolates differed significantly from P. megasperma sensu stricto and other related Phytophthora species. The rose and strawberry isolates had external proliferation of sporangia, characteristic funnel-shaped oogonia, predominantly paragynous antheridia, and fast growth rates of 10.5 mm/24 h at an optimum temperature of 28 °C. In the multilocus phylogenetic tree constructed using sequences from the rDNA ITS regions, rDNA LSU, and the translation elongation factor 1-α, β-tubulin and coxI genes, they formed a distinct monophyletic group in clade 7 with strong bootstrap support. The rose and strawberry isolates separated into two distinct groups. The results indicate that the rose and strawberry isolates constitute two separate species, designated here as Phytophthora nagaii and P. fragariaefolia.     

Multiple Alternaria species groups are associated with leaf blotch and fruit spot diseases of apple in Australia

Leaf blotch and fruit spot of apple caused by Alternaria species occur in apple orchards in Australia. However, there is no information on the identity of the pathogens and whether one or more Alternaria species cause both diseases in Australia. Using DNA sequencing and morphological and cultural characteristics of 51 isolates obtained from apple leaves and fruit with symptoms in Australia, Alternaria species groups associated with leaf blotch and fruit spot of apples were identified. Sequences of Alternaria allergen a1 and endopolygalacturonase gene regions revealed that multiple Alternaria species groups are associated with both diseases. Phylogenetic analysis of concatenated sequences of the two genes resulted in four clades representing A. arborescens and A. arborescens‐like isolates in clade 1, A. tenuissima/A. mali isolates in clade 2, A. alternata/A. tenuissima intermediate isolates in clade 3 and A. longipes and A. longipes‐like isolates in clade 4. The clades formed using sequence information were supported by colony characteristics and sporulation patterns. The source of the isolates in each clade included both the leaf blotch variant and the fruit spot variant of the disease. Alternaria arborescens‐like isolates were the most prevalent (47%) and occurred in all six states of Australia, while A. alternata/A. tenuissima intermediate isolates (14%) and A. tenuissima/A. mali isolates (6%) occurred mostly in Queensland and New South Wales, respectively. Implications of multiple Alternaria species groups on apples in Australia are discussed.     

Isolation and characterization of gut bacteria of fruit fly, Bactrocera tau (Walker)

Gut bacteria of fruit fly, Bactrocera tau (Walker) (Diptera: Tephritidae), were isolated and the isolates attractive to B. tau adults were characterized using morphological, biochemical and 16S rRNA analyses to determine their taxonomic position. Based upon morphological, biochemical and 16S rRNA sequences (on the basis of closest match), five gut bacterial species of B. tau were characterized as Delftia acidovorans, Pseudomonas putida, Flavobacterium sp., Defluvibacter sp. and Ochrobactrum sp., of which four bacterial isolates, viz., Delftia acidovorans, Flavobacterium sp., Defluvibacter sp. and Ochrobactrum sp. are new records from guts of the fruit fly species.     

建兰胶孢炭疽菌ITS序列分析及其PCR快速检测

由胶孢炭疽菌Colletotrichum gloeosporioides引起的炭疽病是建兰的重要病害.为建立快速检测该病原菌的方法,以ITSl/ITS4为引物,对15个建兰胶孢炭疽菌的ITS进行PCR扩增及测序,将测定的序列与炭疽菌属其它种的ITS序列进行比对分析,设计特异性引物CFl/CR1,并通过常规和巢式PCR对建兰胶孢炭疽菌进行检测.结果显示,15个菌株中有13个菌株ITS序列与该菌的模式种序列相似性高达99％以上,而另外2个菌株相似性则为86％;供试菌株在系统发育树上聚为2个不同的分支;引物CFl/CR1通过常规PCR可从1 ng的建兰胶孢炭疽菌基因组DNA中扩增到目的条带,而利用引物ITSl/ITS4和CF1/CR1通过巢式PCR可从1 pg的基因组DNA中扩增到目的条带,即巢式PCR反应检测灵敏度较常规PCR至少高1 000倍.表明建立的巢式PCR法可从自然感病的建兰叶片组织中检测到胶孢炭疽菌.     

Studies of canker and dieback of oak tree in China,with two Cytospora species described

Oak (Quercus sp.) is an excellent tree species as a windbreak, for water conservation, and for fireproofing in forests in China. However, several trees of this genus were found to be suffering from various fungal diseases. In this study, we evaluated 15 fungal pathogens that can cause dieback and canker disease in oak in China, and discovered two Cytospora species. They were identified as Cytospora quercinum sp. nov. and C. vinacea, based on detailed morphological comparisons and phylogenetic analysis of ITS, LSU, act, rpb2, tef1-α, and tub2 loci. This study is the first record of C. quercinum and C. vinacea as causal agents of dieback in oak based on pathogenicity tests conducted on 2-year-old plants in a greenhouse. In addition, this study also revealed the influence of different conditions on the growth rate of mycelia. Mycelial growth of C. quercinum and C. vinacea occurred at optimum temperatures of 20.1 and 20.8 °C, and optimum pH of 5.4 and 5.3, respectively. For these two species, utilization of glucose and fructose was highly efficient, and sucrose was the least efficient. The habitat of Quercus mongolica indicated that more attention and management are needed to prevent the occurrence of Cytospora disease in the summer in north-eastern China, and in spring and autumn in eastern and northern China. This study contributes to the understanding of the species causing canker or dieback diseases in important economic forest trees, and provides useful information for effective disease management of oak trees in China.     

A new ‘forma specialis’ of Fusarium solani causing leaf yellowing of Phalaenopsis

The pathogenicity of 35 Fusarium solani isolates obtained from diseased leaves of greenhouse‐grown Phalaenopsis plants in Taiwan was tested on different orchids, including Phalaenopsis sp., Cymbidium spp., Oncidium sp., Dendrobium sp. and Cattleya sp., plus pea (Pisum sativum), chrysanthemum (Chrysanthemum indicum) and cucurbit [melon (Cucumis melo) and cucmber (C. sativus)] plants. Isolates of F. solani from Phalaenopsis spp. caused severe leaf yellowing on Phalaenopsis and mild symptoms on Cymbidium spp., but no visual symptoms on Oncidium sp., Dendrobium sp., Cattleya sp., pea, chrysanthemum or melon. Fusarium solani isolates collected from Phalaenopsis, pea and cucurbits were molecularly characterized by internal transcribed spacer (ITS), intergenic spacer (IGS) and β‐tubulin gene analyses. Phylogenetic trees constructed by distance and parsimony methods indicated that isolates from Phalaenopsis were grouped into one type based on ITS, IGS and β‐tubulin sequences with high bootstrap value (> 84%) support, compared to ‘formae speciales’ of F. solani from the other hosts. These analyses show that isolates of F. solani from Phalaenopsis are distinct from F. solani isolates from other hosts in Taiwan. Therefore, it is proposed that F. solani isolates that incite Phalaenopsis leaf yellowing be designated F. solani f. sp. phalaenopsis.     

Identification and pathogenicity of Cryphonectriaceae species associated with chestnut canker in China

Cryphonectriaceae species cause serious canker diseases on chestnut, oak and eucalypt trees. Recently, canker symptoms with typical orange fruiting bodies were observed on Chinese chestnut and oak trees in Hebei, Hubei, Shaanxi and Shandong Provinces in China. In the present study, isolates of these fungi were identified based on phylogenetic and morphological evidence, and their pathogenicities were tested on detached chestnut (Castanea mollissima) branches. DNA sequence comparisons of the internal transcribed spacer (ITS) regions and two regions of β-tubulin (TUB1/TUB2) indicate that these isolates represent five species in the Cryphonectriaceae, viz. Cryphonectria japonica, Cryphonectria parasitica, Aurantiosacculus castaneae sp. nov., Cryphonectria neoparasitica sp. nov. and Endothia chinensis sp. nov. The sexual morph of Aurantiosacculus is discovered for the first time and can be distinguished from the other genera in Cryphonectriaceae by dark brown ascospores and tubiform appendages at both ends. Cryphonectria neoparasitica sp. nov. is different from the other Cryphonectria species by its aseptate ascospores. Endothia chinensis sp. nov. is the sole species in Endothia infecting the host genus Castanea. Additionally, it is much smaller than E. gyrosa and narrower than E. singularis in ascospores. The inoculation results showed that these five Cryphonectriaceae fungi isolated from chestnut or oak are all pathogenic to tested chestnut branches. Cryphonectria parasitica appears to be the most aggressive fungus, followed by C. neoparasitica sp. nov., C. japonica, E. chinensis sp. nov. and A. castaneae sp. nov.     

Characterization of Monilinia species associated with brown rot in stone fruit in Brazil

Fungi of the Monilinia genus occur worldwide and affect a wide range of economically important stone fruits. Several Monilinia species are responsible for brown rot. Although this disease is common in Brazil, Monilinia sp. genetic variability in Brazilian orchards has generally been poorly characterized. The present study represents the first report on the genetic diversity of Monilinia sp. from Brazilian orchards. The genetic structure of the Brazilian population was also compared to isolates from other countries, together with some morphological characteristics and aggressiveness. Sixty‐one isolates belonging to the Monilinia genus were obtained from different orchards in Brazilian states. Ten Monilinia fructicola isolates from the United States and one isolate from a fruit imported into Brazil were also evaluated. Phylogenetic analysis of the ITS1‐5.8S‐ITS2 region (internal transcribed spacer) clustered most Brazilian and American isolates with M. fructicola authentic strains from Q‐Bank. Two isolates (one from an imported fruit) clustered as Monilinia laxa. The results revealed M. fructicola as the prevalent species associated with brown rot in Brazilian orchards. To evaluate the intraspecific diversity of M. fructicola and M. laxa, multigene sequence analysis was performed using ITS1‐5.8S‐ITS2 and TEF1 (elongation factor 1). Whilst TEF1 is the most phylogenetically informative gene for intraspecific studies of M. fructicola, RPB2 (RNA polymerase II gene) displayed low variation in intraspecific analysis, but was an informative locus for assigning isolates to M. fructicola or M. laxa species. The amova suggests that Brazilian isolates from the States of the main producing regions belong to a single genetic population, which is genetically distinct from the US (Californian) population of M. fructicola.     

Phylogeny, diversity, and species delimitation in some species of the Xiphinema americanum-group complex (Nematoda: Longidoridae), as inferred from nuclear and mitochondrial DNA sequences and morphology

During nematode surveys in southern Spain and Italy 14 populations of Xiphinema species tentatively identified as Xiphinema americanum-group were detected. Morphological and morphometrical studies identified three new species and six known Xiphinema americanum-group species, viz.: Xiphinema parabrevicolle n. sp., Xiphinema parapachydermum n. sp., Xiphinema paratenuicutis n. sp., Xiphinema duriense, Xiphinema incertum, Xiphinema opisthohysterum, Xiphinema pachtaicum, Xiphinema rivesi, and Xiphinema santos. The Xiphinema americanum-group is the most difficult Xiphinema species group for diagnosis since the morphology is very conservative and morphometric characters often overlap. This group includes vectors of several important plant pathogenic viruses that cause significant damage to a wide range of agricultural crops. Molecular characterisation of these species using D2-D3 expansion regions of 28S rRNA, 18S rRNA, ITS1-rRNA and the protein-coding mitochondrial gene, cytochrome oxidase c subunit 1 was carried out and maximum likelihood and Bayesian inference analysis were used to reconstruct phylogenetic relationships among these species and with other Xiphinema americanum-group species.     

Resistance assessment and biochemical responses of sugar beet lines against <Emphasis Type="Italic"> Pythium aphanidermatum,</Emphasis> causing root rot

Wood-staining fungi, moulds and decay fungi colonize freshly cut wood. Of these, only ophiostomatoid fungi are considered major agents of sapstain on logs and freshly sawn lumber because they can cause permanent staining and discoloration. Sapstain in pine pallets significantly reduces their market value and use in the food industry. The aim of this study was to identify the sapstaining fungi that colonize Scots pine wood used in pallet production. In addition, we evaluated the growth and stain intensity of six isolates of ophiostomatoid fungi on freshly cut Scots pine billets. Fungi were isolated from samples of Scots pine and identified based on morphology and DNA sequence comparisons for three gene regions (ITS, β-tubulin, TEF-1α). A total of 1259 isolates representing 31 fungal species were obtained from the pine samples in Poland during July and September. The isolates represented different ecological and taxonomical groups and belonged to the categories of staining fungi, decay fungi and surface moulds. The most frequently isolated fungi were ophiostomatoid fungi (14 species including an unknown Leptographium sp. and Ophiostoma sp.) and moulds (mainly Trichoderma and Mucor spp.). Of the ophiostomatoid species, Ophiostoma floccosum, O. piceae, O. piliferum and Endoconidiophora pinicola were the predominant species. The results of pine billet inoculation showed that among the six fungal species tested, E. pinicola exhibited the fastest growth in all three directions on the billets. Ophiostoma minus and O. piliferum displayed moderate growth rates in pine billets, while O. floccosum, O. piceae, and Leptographium lundbergii grew very slowly, especially in the tangential and radial directions. The information provided in this paper will help develop more effective control strategies for sapstain prevention in Scots pine.