Transcriptome Changes during Major Developmental Transitions Accompanied with Little Alteration of DNA Methylome in Two Pleurotus Species.

Pleurotus tuoliensis (Pt) and P. eryngii var. eryngii (Pe) are important edible mushrooms. The epigenetic and gene expression signatures characterizing major developmental transitions in these two mushrooms remain largely unknown. Here, we report global analyses of DNA methylation and gene expression in both mushrooms across three major developmental transitions, from mycelium to primordium and to fruit body, by whole-genome bisulfite sequencing (WGBS) and RNA-seq-based transcriptome profiling. Our results revealed that in both Pt and Pe the landscapes of methylome are largely stable irrespective of genomic features, e.g., in both protein-coding genes and transposable elements (TEs), across the developmental transitions. The repressive impact of DNA methylation on expression of a small subset of genes is likely due to TE-associated effects rather than their own developmental dynamics. Global expression of gene orthologs was also broadly conserved between Pt and Pe, but discernible interspecific differences exist especially at the fruit body formation stage, and which are primarily due to differences in trans-acting factors. The methylome and transcriptome repertories we established for the two mushroom species may facilitate further studies of the epigenetic and transcriptional regulatory mechanisms underpinning gene during development in Pleurotus and related genera.

Biodiversity of Bacteria Associated with Eight Pleurotus ostreatus (Fr.) P. Kumm. Strains from Poland, Japan and the USA.

Few publications report the occurrence of bacteria associated with fungal cells. The presence of bacteria associated with one strain of Pleurotus ostreatus (Fr.) P. Kumm. was described in the literature. We describe the biodiversity of bacteria associated with eight oyster mushroom strains from Japan, Poland, and the USA. The presence of microorganisms associated with all tested P. ostreatus strains was confirmed using fluorescent microscopy. Among 307 sequences, 233 of clones representing 34 genera and 74 sequences were identified as Bacteria. Most of the bacteria associated with the strain PUSAS were related to E. coli and two clones were related to Cupriavidus genus. The biodiversity of clones isolated from fungal strains originating from Japan and Poland ranged from 15 to 32 different bacterial clones. The most often the bacteria related to genus Curvibacter, Pseudomonas, Bacillus, Cupriavidus, Pelomonas, and Propionibacterium were associated with the strains of fungi mentioned above. Laccase-like (LMCO) genes were identified in whole bacterial DNA isolated from the associated bacteria but ß-glucosidase and ß-xylanase genes were not detected.Few publications report the occurrence of bacteria associated with fungal cells. The presence of bacteria associated with one strain of Pleurotus ostreatus (Fr.) P. Kumm. was described in the literature. We describe the biodiversity of bacteria associated with eight oyster mushroom strains from Japan, Poland, and the USA. The presence of microorganisms associated with all tested P. ostreatus strains was confirmed using fluorescent microscopy. Among 307 sequences, 233 of clones representing 34 genera and 74 sequences were identified as Bacteria. Most of the bacteria associated with the strain PUSAS were related to E. coli and two clones were related to Cupriavidus genus. The biodiversity of clones isolated from fungal strains originating from Japan and Poland ranged from 15 to 32 different bacterial clones. The most often the bacteria related to genus Curvibacter, Pseudomonas, Bacillus, Cupriavidus, Pelomonas, and Propionibacterium were associated with the strains of fungi mentioned above. Laccase-like (LMCO) genes were identified in whole bacterial DNA isolated from the associated bacteria but ß-glucosidase and ß-xylanase genes were not detected.

Mapping the Secretome and Its N-Linked Glycosylation of Pleurotus eryngii and Pleurotus ostreatus Grown on Hemp Stalks.

Our previous research showed that Pleurotus eryngii and Pleurotus ostreatus were effective fungi for pretreatment of industrial hemp stalks to improve enzymatic saccharification. The secretomes of these two fungi were analyzed to search for the effective enzyme cocktails degrading hemp lignin during the pretreatment process. In total, 169 and 155 proteins were identified in Pleurotus eryngii and Pleurotus ostreatus, respectively, and 50% of the proteins involved in lignocellulose degradation were CAZymes. Because most of the extracellular proteins secreted by fungi are glycosylated proteins, the N-linked glycosylation of enzymes could be mapped. In total, 27 and 24 N-glycosylated peptides were detected in Pleurotus eryngii and Pleurotus ostreatus secretomes, respectively. N-Glycosylated peptides of laccase, GH92, exoglucanase, phenol oxidase, α-galactosidase, carboxylic ester hydrolase, and pectin lyase were identified. Deglycosylation could decrease enzymatic saccharification of hemp stalks. The activities of laccase, α-galactosidase, and phenol oxidase and the thermal stability of laccase were reduced after deglycosylation.

Pleurotus opuntiae revisited - An insight to the phylogeny of dimitic Pleurotus species with emphasis on the P. djamor complex.

The name Pleurotus opuntiae is indiscriminately used for describing mushrooms with white to off-white to white-grey pilei with short or absent stipe and dimitic hyphal system, which grow on plants of the genera Opuntia, Yucca, Agave, Phytolacca etc. However, the outcome of the present study evidences that this name should be reserved for specimens deriving from the Mediterranean area only; an epitype originating from Italy on Opuntia ficus-indica is designated. Pertinent material was sequenced by using the internal transcribed spacer region (ITS) and found to be phylogenetically related to P. djamor from Kenya and Nigeria, while members of the P. djamor complex from other continents were clearly more distant. Results were further corroborated by examining the large subunit of nuclear ribosomal DNA (LSU) and the second subunit of RNA polymerase II (RPB2). The P. djamor complex shows high intraspecific polymorphism evidenced by sequence divergence and genetic distance values, presents a cosmopolitan distribution and also comprises material initially identified as P. flabellatus, P. opuntiae, P. ostreatoroseus, P. parsonsiae and P. salmoneostramineus. An ITS tree including representative specimens from all major Pleurotus species is provided for the first time and ambiguous taxa are discussed in the context of new findings.

Comparison of non-volatile and volatile flavor compounds in six Pleurotus mushrooms.

BACKGROUND: Non-volatile and volatile flavor compounds of six Pleurotus mushrooms including Pleurotus citrinopileatus, P. cornucopiae, P. djamor, P. floridanus, P. ostreatus and P. sapidus were studied. RESULTS: The content of total free amino acids ranged from 21.80 to 40.60 g kg-1 and the content of monosodium glutamate (MSG)-like amino acids ranged from 3.10 to 8.64 g kg-1 . The content of total 5'-nucleotides ranged from 4.16 to 8.80 g kg-1 while the content of flavor 5'-nucleotides ranged from 2.00 to 4.51 g kg-1 . Sixty-three volatile compounds were identified in six Pleurotus mushrooms, including 17 aldehydes, 10 ketones, 14 alcohols, 2 ethers, 5 acids, 5 hydrocarbons, 10 heterocyclic and aromatic compounds. 1-Octen-3-one and 1-octen-3-ol were the key odor compounds in P. citrinopileatus, P. djamor, P. ostreatus, P. floridanus and P. sapidus, while 1-octen-3-one, 1-octen-3-ol and 2-octenal were the key odor compounds in P. cornucopiae. CONCLUSION: Pleurotus citrinopileatus had highest content of total free amino acids (40.60 g kg-1 ), total 5'-nucleotides (8.80 g kg-1 ) and flavor 5'-nucleotides (4.51 g kg-1 ) than other Pleurotus mushrooms. Moreover, eight-carbon compounds were the most abundant compounds in six Pleurotus mushrooms. Our study should be helpful in promoting the cultivation and consumption of these Pleurotus mushrooms. © 2018 Society of Chemical Industry.

Classification of fungal glucuronoyl esterases (FGEs) and characterization of two new FGEs from Ceriporiopsis subvermispora and Pleurotus eryngii.

Fungal glucuronoyl esterases (FGEs) catalyze cleavage of the ester bond connecting a lignin alcohol to the xylan-bound 4-O-methyl-D-glucuronic acid of glucuronoxylans. Thus, FGEs are capable of degrading lignin-carbohydrate complexes and have potential for biotechnological applications toward woody biomass utilization. Therefore, identification and characterization of new FGEs are of critical importance. Firstly, in this study, we built a phylogenetic tree from almost 400 putative FGEs obtained on BLAST analysis and defined six main clades. In the phylogenetic tree, all the putative FGEs of ascomycetes cluster in clades I to IV, and most of the putative FGEs of basidiomycetes (B-FGEs) cluster in clades V to VI. Interestingly, several B-FGEs were found to cluster in clade II; most FGEs of clade II were found to have higher theoretical isoelectric points than those in the other five clades. To gain an insight into the putative FGEs in the clades that have not been characterized yet, we chose the FGEs of Ceriporiopsis subvermispora (CsGE) and Pleurotus eryngii (PeGE), which belong to clades V and II, respectively. The catalytic domains of both CsGE and PeGE were successfully expressed using Pichia pastoris, and then purified. Benzyl glucuronic acid was used as a substrate to confirm the activities of the CsGE and PeGE, and the hydrolyzed product, glucuronic acid, was quantified spectrophotometrically. Both CsGE and PeGE clearly exhibited the esterase activity. Additionally, we demonstrated that PeGE exhibits high tolerance toward several denaturing agents, which may make it a potentially more applicable enzyme.

Genome-Wide Characterization and Expression Analyses of Pleurotus ostreatus MYB Transcription Factors during Developmental Stages and under Heat Stress Based on de novo Sequenced Genome.

Pleurotus ostreatus is a commercially grown mushroom species in China. However, studies on the mechanisms of the fruiting body development and stress response of P. ostreatus are still at a primary stage. In this study, we report the entire genome sequence of P. ostreatus CCMSSC03989. Then, we performed comprehensive genome-wide characterization and expression analysis of the MYB transcription factor family during a series of developmental stages and under the condition of heat stress. A 34.76 Mb genome was obtained through next-generation sequencing (NGS) and Bionano optical mapping approaches. The genome has a scaffold N50 of 1.1 Mb and contains 10.11% repeats, and 10,936 gene models were predicted. A total of 20 MYB genes (PoMYB) were identified across the genome, and the full-length open reading frames were isolated. The PoMYBs were classified into 1 repeat (1R), 2R, and 3R-MYB groups according to their MYB domain repeat numbers, and 3R-MYBs possessed relatively more introns than 1R and 2R-MYBs. Based on phylogenetic analysis, the PoMYBs were divided into four groups and showed close relationships with the MYB genes of plants and fungi. RNA-sequencing (RNA-Seq) and quantitative PCR (qPCR) analyses revealed that PoMYB expression showed stage-specific patterns in reproductive stages and could be induced by heat stress. The P. ostreatus draft genome will promote genome-wide analysis, and our study of PoMYBs will promote further functional analysis of MYB genes in mushrooms.

Comparative mitogenomics reveals large-scale gene rearrangements in the mitochondrial genome of two Pleurotus species.

In the present study, we assembled the mitogenomes of Pleurotus citrinopileatus and Pleurotus platypus. The circular mitogenome of the two Pleurotus species comprises a set of 14 conserved protein-encoding genes (PEGs), 2 RNA genes (small subunit ribosomal RNA and large subunit ribosomal RNA), and 24 tRNAs, with sizes of 60,694 and 73,807 bp, respectively. They contain 4 and 10 introns with 3 and 10 intronic open reading frames (ORFs), respectively. Thirteen position classes (Pcls) of introns were found in the cox1 gene of four Pleurotus species. The number and class of Pcl varied among different Pleurotus species, indicating that numerous events of loss and gain occurred during evolution of Pleurotus. Comparative mitogenomic and collinearity analyses reveal that large-scale gene rearrangements may have occurred during the evolution of Pleurotus citrinopileatus and Pleurotus platypus, including gene rearrangements and inversions, which may be related to the observed high amounts of repetitive DNA elements (5.62 and 5.45%, respectively). Phylogenetic analysis based on concatenated mitochondrial protein sequences reveals that concatenated mitochondrial genes are suitable as molecular markers for phylogenetic analysis. This serves as the first report on large-scale rearrangements in the mitochondria of the genus Pleurotus, thereby improving our understanding of the evolution of the Pleurotus genus and other macrofungi.

The evolution of genomic and epigenomic features in two Pleurotus fungi.

Pleurotus tuoliensis (Bailinggu, designated Pt) and P. eryngii var. eryngii (Xingbaogu, designated Pe) are highly valued edible mushrooms. We report de novo assemblies of high-quality genomes for both mushrooms based on PacBio RS II sequencing and annotation of all identified genes. A comparative genomics analysis between Pt and Pe with P. ostreatus as an outgroup taxon revealed extensive genomic divergence between the two mushroom genomes primarily due to the rapid gain of taxon-specific genes and disruption of synteny in either taxon. The re-appraised phylogenetic relationship between Pt and Pe at the genome-wide level validates earlier proposals to designate Pt as an independent species. Variation of the identified wood-decay-related gene content can largely explain the variable adaptation and host specificity of the two mushrooms. On the basis of the two assembled genome sequences, methylomes and the regulatory roles of DNA methylation in gene expression were characterized and compared. The genome, methylome and transcriptome data of these two important mushrooms will provide valuable information for advancing our understanding of the evolution of Pleurotus and related genera and for facilitating genome- and epigenome-based strategies for mushroom breeding.

Identification of pyrG Used as an Endogenous Reference Gene in Qualitative and Real-Time Quantitative PCR Detection of Pleurotus ostreatus.

As a well-known edible fungus rich in nutrients, Pleurotus ostreatus has been used as an alternative to expensive wild edible fungi. Specifically, the fact that using P. ostreatus instead of other expensive wild edible fungi has damaged the rights and interests of consumers. Among the existing methods for detection of food adulteration, the amplification of endogenous reference gene is the most accurate method. However, an ideal endogenous reference gene for P. ostreatus has yet to be developed. In this study, a DNA extraction method for P. ostreatus was optimized, and pyrG was selected as a species-specific gene through sequence alignment. This gene was subsequently subjected to qualitative and quantitative Polymerase Chain Reaction (PCR) assays with 3 different P. ostreatus varieties and 7 other species. A low detection limit of 5 pg/µL was obtained by TaqMan quantitative PCR, and no pyrG amplification product was observed in the 7 other species. No allelic variation was detected in P. ostreatus varieties. These experiments confirmed that pyrG was an ideal endogenous reference gene for the qualitative and real-time quantitative PCR detection of P. ostreatus. This method was also suitable for the examination of processed P. ostreatus samples and determination of adulteration in wild mushrooms. PRACTICAL APPLICATION: The pyrG gene was chosen as an ideal endogenous reference gene for the qualitative and real-time quantitative PCR detection of P. ostreatus, and the detection limit was 5 pg/µL for the quantification. This method is used not only for raw materials but also for processed P. ostreatus products and other processed mushroom foods.

Intra- and inter-isolate variation of ribosomal and protein-coding genes in Pleurotus: implications for molecular identification and phylogeny on fungal groups.

BACKGROUND: The internal transcribed spacer (ITS), RNA polymerase II second largest subunit (RPB2), and elongation factor 1-alpha (EF1α) are often used in fungal taxonomy and phylogenetic analysis. As we know, an ideal molecular marker used in molecular identification and phylogenetic studies is homogeneous within species, and interspecific variation exceeds intraspecific variation. However, during our process of performing ITS, RPB2, and EF1α sequencing on the Pleurotus spp., we found that intra-isolate sequence polymorphism might be present in these genes because direct sequencing of PCR products failed in some isolates. Therefore, we detected intra- and inter-isolate variation of the three genes in Pleurotus by polymerase chain reaction amplification and cloning in this study. RESULTS: Results showed that intra-isolate variation of ITS was not uncommon but the polymorphic level in each isolate was relatively low in Pleurotus; intra-isolate variations of EF1α and RPB2 sequences were present in an unexpectedly high amount. The polymorphism level differed significantly between ITS, RPB2, and EF1α in the same individual, and the intra-isolate heterogeneity level of each gene varied between isolates within the same species. Intra-isolate and intraspecific variation of ITS in the tested isolates was less than interspecific variation, and intra-isolate and intraspecific variation of RPB2 was probably equal with interspecific divergence. Meanwhile, intra-isolate and intraspecific variation of EF1α could exceed interspecific divergence. These findings suggested that RPB2 and EF1α are not desirable barcoding candidates for Pleurotus. We also discussed the reason why rDNA and protein-coding genes showed variants within a single isolate in Pleurotus, but must be addressed in further research. CONCLUSIONS: Our study demonstrated that intra-isolate variation of ribosomal and protein-coding genes are likely widespread in fungi. This has implications for studies on fungal evolution, taxonomy, phylogenetics, and population genetics. More extensive sampling of these genes and other candidates will be required to ensure reliability as phylogenetic markers and DNA barcodes.

Bioactive compounds and antioxidant activity exhibit high intraspecific variability in Pleurotus ostreatus mushrooms and correlate well with cultivation performance parameters.

Experimental data related with oyster mushroom production and nutritional properties usually derive from the examination of only one strain, and hence their representativeness/usefulness is questionable. This work aims at assessing intraspecific variability in Pleurotus ostreatus by studying 16 strains, under the same conditions, in respect to essential cultivation and mushroom quality aspects, and by defining the impact of intrinsic/genetic factors on such parameters. Hence, mushroom yield, earliness, crop length, biological efficiency, productivity, and their content in selected macro and microconstituents (e.g. fatty acids, sterols, individual phenolic compounds, terpenic acids, glucans) as well as their antioxidant properties (i.e., antiradical activity, ferric reducing potential, inhibition of serum oxidation) were assayed. The effect of intrinsic/genetic factors was evident, especially as regards earliness, yield of each production flush and mushroom weight, whereas biological efficiency was not particularly influenced by the cultivated strain. Moreover, phenolics, ergosterol and antiradical activity demonstrated significant variability among strains in contrast to what was observed for fatty acids, ß-glucans and ferric reducing potential. The observed heterogeneity reveals the limitations of using a low number of strains for evaluating mushroom production and/or their content in bioactive compounds, and as evidenced, it is valuable for breeding and commercial purposes.

Identification of two mutations that cause defects in the ligninolytic system through an efficient forward genetics in the white-rot agaricomycete Pleurotus ostreatus.

White-rot fungi play an important role in the global carbon cycle because they are the species that almost exclusively biodegrade wood lignin in nature. Lignin peroxidases (LiPs), manganese peroxidases (MnPs) and versatile peroxidases (VPs) are considered key players in the ligninolytic system. Apart from LiPs, MnPs and VPs, however, only few other factors involved in the ligninolytic system have been investigated using molecular genetics, implying the existence of unidentified elements. By combining classical genetic techniques with next-generation sequencing technology, they successfully showed an efficient forward genetics approach to identify mutations causing defects in the ligninolytic system of the white-rot fungus Pleurotus ostreatus. In this study, they identified two genes - chd1 and wtr1 - mutations in which cause an almost complete loss of Mn2+ -dependent peroxidase activity. The chd1 gene encodes a putative chromatin modifier, and wtr1 encodes an agaricomycete-specific protein with a putative DNA-binding domain. The chd1-1 mutation and targeted disruption of wtr1 hamper the ability of P. ostreatus to biodegrade wood lignin. Examination of the effects of the aforementioned mutation and disruption on the expression of certain MnP/VP genes suggests that a complex mechanism underlies the ligninolytic system in P. ostreatus.

Is Widely Cultivated "Pleurotus sajor-caju", Especially in Asia, Indeed an Independent Species?

In modern mycology, the name "Pleurotus sajor-caju" has 2 meanings: a strict taxonomical meaning refers to its nomenclatural synonym Lentinus sajor-caju, and a more widely distributed biotechnological meaning refers to a strain determined as "Pleurotus sajor-caju" but that belongs to true Pleurotus. The main purpose of this article is to highlight the taxonomic status and to give the correct name for this problematic member of Pleurotus. Both microscopic analysis and BLAST search results (internal transcribed spacer homology varies between 97% and 100%) provide the position of the fungus in question into the range of infraspecific radiation of P. pulmonarius. It was shown that the fungus represents a tropical ecotype of P. pulmonarius, and a new variety (P. pulmonarius var. stechangii) was described for its taxonomic representation. This variety was named in honor of Professor S.-T. Chang, a great mycologist who introduced this taxon to the mushroom industry. A formal change in the anamorphic name of the fungus in question is provided, too, and a new name is presented: Antromycopsis stechangii.

Aflatoxin B1 and M1 Degradation by Lac2 from Pleurotus pulmonarius and Redox Mediators.

Laccases (LCs) are multicopper oxidases that find application as versatile biocatalysts for the green bioremediation of environmental pollutants and xenobiotics. In this study we elucidate the degrading activity of Lac2 pure enzyme form Pleurotus pulmonarius towards aflatoxin B1 (AFB1) and M1 (AFM1). LC enzyme was purified using three chromatographic steps and identified as Lac2 through zymogram and LC-MS/MS. The degradation assays were performed in vitro at 25 °C for 72 h in buffer solution. AFB1 degradation by Lac2 direct oxidation was 23%. Toxin degradation was also investigated in the presence of three redox mediators, (2,2'-azino-bis-[3-ethylbenzothiazoline-6-sulfonic acid]) (ABTS) and two naturally-occurring phenols, acetosyringone (AS) and syringaldehyde (SA). The direct effect of the enzyme and the mediated action of Lac2 with redox mediators univocally proved the correlation between Lac2 activity and aflatoxins degradation. The degradation of AFB1 was enhanced by the addition of all mediators at 10 mM, with AS being the most effective (90% of degradation). AFM1 was completely degraded by Lac2 with all mediators at 10 mM. The novelty of this study relies on the identification of a pure enzyme as capable of degrading AFB1 and, for the first time, AFM1, and on the evidence that the mechanism of an effective degradation occurs via the mediation of natural phenolic compounds. These results opened new perspective for Lac2 application in the food and feed supply chains as a biotransforming agent of AFB1 and AFM1.

Taxonomic Identity, Geographic Distribution, and Commercial Exploitation of the Culinary-Medicinal Mushroom Pleurotus nebrodensis (Basidiomycetes).

An updated overview of the outcome of studies conducted on the culinary-medicinal mushroom Pleurotus nebrodensis is presented by placing emphasis on the clarification of the taxonomic identity of P. nebrodensis and other related taxa possessing entirely white to cream basidiomes, which grow in association with different plants of the family Apiaceae. Cultivation techniques, quality of the product sold and sales price, as well as nutritional and medicinal aspects are discussed. Taking also into consideration the high economic importance of P. nebrodensis, it is essential to proceed with the verification of the commercial strains currently available in the international market under the name of "P. nebrodensis" since it is very probable that many (or most) of them do not represent the real P. nebrodensis. TO confirm this hypothesis, an in silico analysis was conducted on a large of number of ITS1-5.8S-ITS2 rRNA sequences deposited in the National Center for Biotechnology Information database under the name P. nebrodensis. Results demonstrated that all "P nebrodensis" material examined from China (plus several sequences of no reported origin) corresponded to P. eryngii subsp. tuoliensis, with only 2 exceptions, which were grouped within P. eryngii sensu stricto. The real P. nebrodensis biological material from Italy and Greece is certified and is available upon request by the authors at the University of Palermo and the Agricultural University of Athens.

Comparative evaluation of five Pleurotus species for their growth behaviour and yield performance using wheat straw as a substrate.

Pleurotus spp. is one of the most important edible mushrooms cultivated in India. The present study was an attempt to compare five Pleurotus species in context of actual time required for each growth stage viz., spawn run period, number of days required for initiation of pin heads of sporophores, average weight of fruiting bodies in all the flushes and total yield. The spawn run period in all the five species were recorded between 18 days-21 days, similarly for initiation of pinheads 5 days -7 days were required after spawn run period. A total of 24 days to 27 days, 34 days to 37 days and 47 days to 53 days were required for harvesting the I, II and III flushes respectively. An average number of 41 to 70 sporophores per bag containing 1 kg of dry substrates were obtained from all the Pleurotus species. Maximum 14 g weight of single sporophore was recorded from P. florida, similarly, an average maximum diameter of 5.3 cm of sporophores of P. florida was observed whereas the diameter of sporophores in rest of the species ranged from 3.0 cm to 3.2 cm. The number of sporophores were obtained from P. sajor-caju (n-70) and all the species showed significant difference with respect to the number of sporophores in a bunch at probability level of P = 0.05. Maximum weight of single bunch was recorded (58 g) in P. florida and total yield of 740 gkg(-1) of dry matter was recorded in P. florida.

Morphological and molecular characterization of yellow oyster mushroom, Pleurotus citrinopileatus, hybrids obtained by interspecies mating.

Pleurotus citrinopileatus (yellow oyster mushroom) has an attractive shape and yellow colour but the fragile texture complicates packaging, and its strong aroma is unappealing to consumers. This study aimed to improve the characteristics and yield of P. citrinopileatus by interspecies mating between monokaryotic cultures of P. citrinopileatus and P. pulmonarius. Ten monokaryon cultures of the parental lines were crossed in all combinations to obtain hybrids. Eleven compatible mating pairs were obtained and cultivated to observe their sporophore morphology and yield. The selected hybrid, i.e. P1xC9, was beige in colour while hybrid P3xC8 was yellow in colour. Their sporophores had less offensive aroma, improved texture and higher yield. The DNA sequences of these hybrids were found to be in the same clade as the P. citrinopileatus parent with a bootstrap value of 99%. High bootstrap values indicate high genetic homology between hybrids and the P. citrinopileatus parent. The biological efficiencies of these hybrids P1xC9 (70.97%) and P3xC8 (52.14%) were also higher than the P. citrinopileatus parent (35.63%). Interspecies hybrids obtained by this mating technique can lead to better strains of mushrooms for genetic improvement of the Pleurotus species.

Notes on a New Productive Strain of King Oyster Mushroom, Pleurotus eryngii (Higher Basidiomycetes), a Prized Italian Culinary-Medicinal Mushroom.

In this paper, the authors provide data on a culinary-medicinal, host-specific variety of P. eryngii species-complex that is known in Italy as "cardoncello". A species description, the techniques of isolation of a new strain (C-142-c), and the preparation of the substratum are illustrated. Data on the productivity of substratum inoculated with C-142-c strain and the nutritional value of cultivated "cardoncello" mushrooms are also provided.

Genetic diversity of Kenyan native oyster mushroom (Pleurotus).

Members of the genus Pleurotus, also commonly known as oyster mushroom, are well known for their socioeconomic and biotechnological potentials. Despite being one of the most important edible fungi, the scarce information about the genetic diversity of the species in natural populations has limited their sustainable utilization. A total of 71 isolates of Pleurotus species were collected from three natural populations: 25 isolates were obtained from Kakamega forest, 34 isolates from Arabuko Sokoke forest and 12 isolates from Mount Kenya forest. Amplified fragment length polymorphism (AFLP) was applied to thirteen isolates of locally grown Pleurotus species obtained from laboratory samples using five primer pair combinations. AFLP markers and internal transcribed spacer (ITS) sequences of the ribosomal DNA were used to estimate the genetic diversity and evaluate phylogenetic relationships, respectively, among and within populations. The five primer pair combinations generated 293 polymorphic loci across the 84 isolates. The mean genetic diversity among the populations was 0.25 with the population from Arabuko Sokoke having higher (0.27) diversity estimates compared to Mount Kenya population (0.24). Diversity between the isolates from the natural population (0.25) and commercial cultivars (0.24) did not differ significantly. However, diversity was greater within (89%; P > 0.001) populations than among populations. Homology search analysis against the GenBank database using 16 rDNA ITS sequences randomly selected from the two clades of AFLP dendrogram revealed three mushroom species: P. djamor, P. floridanus and P. sapidus; the three mushrooms form part of the diversity of Pleurotus species in Kenya. The broad diversity within the Kenyan Pleurotus species suggests the possibility of obtaining native strains suitable for commercial cultivation.