Analysis Of ITS Sequences Research Articles

Fungi associated with the potato taste defect in coffee beans from Rwanda

BackgroundPotato taste defect (PTD) of coffee is characterized by a raw potato like smell that leads to a lower quality taste in the brewed coffee, and harms the commercial value of some East African coffees. Although several causes for PTD have been proposed, none of them have been confirmed. Recently, high throughput sequencing techniques and bioinformatic analysis have shown great potential for identifying putative causal agents of plant diseases. Toward the goal of determining the cause of PTD, we examined raw coffee beans from Rwanda exhibiting varying PTD scores using an Illumina-based sequence analysis of the fungal rRNA ITS region.ResultsSix fungal amplicon sequence variants (ASVs) with high relative abundances correlated with coffee taste scores. Four of these ASVs exhibited negative correlations – Aspergillus versicolor, Penicillium cinnamopurpureum, Talaromyces radicus, and Thermomyces lanuginosus – indicating that they might be causing PTD. Two of these fungi exhibited positive correlations – Kazachstania humilis and Clavispora lusitaniae – indicating that they might be inhibiting organisms that cause PTD.ConclusionsThis study addressed PTD causality from a new angle by examining fungi with high throughput sequencing. To our knowledge, this is the first study characterizing fungi associated with PTD, providing candidates for both causality and biocontrol.

Pluteus variabilicolor and Volvopluteus earlei, new records for Pakistan

Pluteus variabilicolor and Volvopluteus earlei are reported as new records for the Pakistani funga, with the former representing a new record for South Asia and the latter, a second report from Asia. Morphological descriptions are accompanied by colored photographs of basidiomata and microscopic structures. Identity of both species is confirmed by ITS sequence analyses. Comparison with previous descriptions and data on their distributions are briefly discussed.

Isolation, Mutagenesis, and Organic Acid Secretion of a Highly Efficient Phosphate-Solubilizing Fungus.

The highly effective phosphate-solubilizing microorganisms are significant for making full use of the potential phosphorus resources in the soil and alleviating the shortage of phosphorus resources. In this study, a phosphate-solubilizing fungus was isolated from wheat and cotton rhizosphere soils in the lower reaches of the Yellow River in China and was identified as Penicillium oxalicum by morphological and ITS sequencing analysis. In order to obtain a fungus with more efficient phosphorus solubilization ability, we tested three positive mutant strains (P1, P2, and P3) and three negative mutant strains (N1, N2, and N3) through low-energy nitrogen ion implantation mutagenesis. Compared with the parental strain, the phosphate-solubilizing capacity of P1, P2, and P3 was enhanced by 56.88%, 42.26%, and 32.15%, respectively, and that of N1, N2, and N3 was weakened by 47.53%, 35.27%, and 30.86%, respectively. Compared with the parental strain, the total amount of organic acids secreted significantly increased in the three positive mutant strains and decreased in the negative mutant strains; the pH of culture medium was significantly lower in the positive mutant strains and higher in the negative mutant strains. The capacity of phosphate-solubilizing fungus to secrete organic acids and reduce the growth-medium pH was closely related to its phosphate-solubilizing ability. The changes in the amount of organic acids secreted by mutants can alter their acidification and phosphate-solubilizing capacity. In conclusion, this study offers a theoretical basis and strain materials for the exploration and application of phosphate-solubilizing fungi.

Molecular and metabolic traits of some Egyptian species of Cassia L. and Senna Mill (Fabaceae-Caesalpinioideae)

The genus Cassia and Senna have been classified under subfamily Caesalpinioideae of family Fabaceae (Leguminosae) of order Fabales. There is a scarce taxonomical studies of the genus Cassia and Senna inhabiting Egyptian environments, thus, the main objective of the current was to revise and authenticate the phylogenetic relationship between studied taxa of the species of the genera Cassia and Senna in Egypt using the recent tools of ITS barcoding, RAPD analysis and metabolic profiling, in comparing to the traditional taxonomical features. From the cluster analysis of the traditional 27 morphological characters, the studied taxa were categorized into two major clades with an average taxonomic distance of 4.3. The clade I include Cassia fistula, C. renigera, C. javanica L subsp. nodosa and C. roughiia that belongs to series Obolospermae, and C. grandis that belongs to series Grandes. The clade (II) includes Senna surattensis and S. alata at taxonomic level 3.6. The taxonomical description of the studied taxa was confirmed from the molecular analysis of ITS sequences and RAPD analysis. The ITS sequences of the tested plants species C. fistula L, C. grandis MD4, C. javanica subsp. nodosa MD7, C. roxburghii MD5, C. renigera MD5 were deposited at genbank with accession numbers MW367973, MZ960447, MW386305, MW326753 and MW32685, respectively. While, the ITS sequences of the S. surrattensis and S. alata were deposited into genbank accession # MD14 MW367670 and MD20 MW412635, respectively. Thus, from the molecular analysis, two clades were clearly separated into Clade I of Cassia and Clade II of Senna. The cluster I represented by C. fistula, C. renigera, C. roxburghii, and C. javanica sub nodosa, and the cluster II represented by S. alata and S. surattensis. From the PCA of RAPD, a clearly discrimination between the two Taxa was observed revealing the characteristic grouping of Cassia and Senna. The species Senna alata and Senna surattensis were grouped together, but the species of C. renigera, C. javanica, C. roxburghii and C. grandis was grouped on a distinct group. The separation of Cassia and Senna species into two clusters verify the segregation of the genus Cassia L. senso lato into two distinct genera namely Senna P. and Cassia L. The morphological, molecular traits of the studied plants were authenticated from the metabolic profiling by GC-MS analysis. Among the 23 identified metabolites, four compounds namely hexadecanoic acid, methyl ester, 9-Octadecenoic acid (Z)-ethyl ester and Vitamin E were detected with fluctuated concentrations, among C. fistula, C. grandis, C. javanica subsp. nodosa and C. roxburghii. Conclusively, the traditional morphological features, molecular barcoding using ITS sequences, RAPD analysis and metabolic traits by GC-MS analysis, authenticates the taxonomical diversity of the genus Cassia and Senna.

Crassiparies yunnanensis sp. nov. (Neohendersoniaceae, Pleosporales) from dead twigs of Coffea arabica in China

Arabica coffee (Coffea arabica) accounting for about 60% of the world’s coffee is popular all over the world as a refreshing beverage and food additive. During surveys of fungi occurring on coffee plants, we collected a species of Neohendersoniaceae on dead twigs of an Arabica coffee plant in Pu’er, Yunnan Province, China. Phylogenetic analyses of combined SSU, LSU, ITS, rpb2, and tef gene sequences confirmed that our fungus is a distinct new species within Crassiparies. Morphological characteristics also support this taxon to be a novel species in Crassiparies. Full description, photo plate, and a phylogenetic tree to show the placement of the new species in Crassiparies are provided.

Identification of a Cordyceps fumosorosea Fungus Isolate and Its Pathogenicity against Asian Citrus Psyllid, Diaphorina citri (Hemiptera: Liviidae).

Simple SummarySome entomopathogenic fungi are highly pathogenic to Diaphorina citri, which is the mainly transmitting vector of CLas. In our study, we isolated an entomopathogenic fungus strain from an adult cadaver of D. citri. The isolate was identified as Cordyceps fumosorosea, based on morphology and ITS sequence analysis, and named C. fumosorosea SCAU-CFDC01. We further evaluated its pathogenicity against D. citri nymphs and adults under laboratory and greenhouse conditions. The laboratory results showed that C. fumosorosea SCAU-CFDC01 was most pathogenic to young nymphs, followed by old nymphs and adults. Furthermore, the results on greenhouse experiments revealed that C. fumosorosea SCAU-CFDC01 not only had high pathogenicity against nymphs, but also had serious negative effects on adults from nymphs treated, including emergence rate of adults and female longevity. Our results showed C. fumosorosea SCAU-CFDC01 was highly pathogenic to D. citri, and these findings will facilitate mycoparasite development for biological control of D. citri.Diaphorina citri is the mainly transmitting vector of the citrus huanglongbing pathogen, which causes severe losses in in the citrus industry. In this study, we isolated a new entomopathogenic fungus, identified as member of Cordyceps fumosorosea based on morphology and ITS sequence analysis. We named C. fumosorosea SCAU-CFDC01 and evaluated its pathogenicity against D. citri nymphs and adults by immersion under laboratory and greenhouse conditions. Results showed that SCAU-CFDC01 was most pathogenic to young nymphs, followed by old nymphs and adults. The LC50 values of the fungus on nymphs and adults showed a declining trend over a 2–7-day period after inoculation. The LT50 (lethal time for a certain concentration to cause 50% mortality) values also presented a decreasing trend along with increasing conidia concentrations. For the results on greenhouse experiments, when 3rd and 5th instar nymphs were inoculated with 1 × 105 conidia mL−1, the survival rate of nymphs were lower, and the emergence rate of adults and female longevity was significantly reduced compared with the control. However, there were no significant effects on sex ratio of adults and male longevity. Our results showed SCAU-CFDC01 was highly pathogenic to D. citri, and may promote mycoparasite development for biological control of D. citri.

Fungal Richness of Cytospora Species Associated with Willow Canker Disease in China.

Species of Cytospora are considered important plant pathogens of a wide range of plant hosts, especially Salicaceae plants. Salix (Salicaceae, Malpighiales) has been widely cultivated in China because of its strong ecological adaptability, fast growth, and easy reproduction. In this study, a total of eight species of Cytospora were discovered on Salix in China, including C. ailanthicola, C. alba, C. chrysosperma, C. gigaspora, C. nivea, C. paracinnamomea, C. rostrata, and C. sophoriopsis. Among them, C. alba and C. paracinnamomea were identified as novel species based on morphology and phylogenetic analyses of ITS, act, rpb2, tef1-α, and tub2 gene sequences and were confirmed as pathogens of willow canker disease by pathogenicity tests. The mycelial growth rates of strains from these two novel species (C. alba and C. paracinnamomea) had optimum temperatures of 21 to 22 °C and an optimum pH value of 5 to 6. The effectiveness of six carbon sources on the mycelial growth showed that fructose and maltose had the highest influence. Cytospora species richness was significantly positively correlated with dry and wet areas. This study represents a significant evaluation of Cytospora associated with willow canker disease in China and provides a theoretical basis for predicting the potential risk of willow canker disease.

Community structure of yeast in fermented soy sauce and screening of functional yeast with potential to enhance the soy sauce flavor

Yeast plays an important role in the formation of desirable aroma during soy sauce fermentation. In this study, the structure and diversity of yeast communities in seven different soy sauce residues were investigated by ITS sequencing analysis, and then the aroma characteristics of selected yeast species were examined by a combination of gas chromatography–mass spectrometry (GC–MS), headspace solid-phase microextraction (SPME) and liquid-liquid extraction (LLE). A total of 18 yeast genera were identified in seven soy sauce residues. Among them, Candid and Zygosaccharomyces were detected in all samples, followed by Millerozyma, Wickerhamiella, Meyerozyma, Trichosporon and Wickerhamomyces, which were found in more than two-thirds of the samples. Subsequently, eight representative species, isolated from soy sauce residues, were subjected to environmental stress tolerance tests and aroma production tests. Among them, three isolated species were regarded as potential aroma-enhancing microbes in soy sauce. Wickerhamiella versatilis could increase the contents of ethyl ester compounds and alcohols, thereby improving the fruity and alcoholic aroma of soy sauce. Candida sorbosivorans enhanced sweet and caramel-like aroma of soy sauce by producing 4-hydroxy-2,5-dimethyl-3(2H)-furanone (HDMF) and 3-hydroxy-2-methyl-4h-pyran-4-one (maltol). Starmerella etchellsii could enhance the contents of 2,6-dimethylpyrazine, methyl pyrazine and benzeneacetaldehyde. This study is of great significance for the development and application of flavor functional yeasts in soy sauce fermentation.

Anthracnose in Samanea tubulosa caused by Colletotrichum karstii in Brazil

AbstractIn April 2019, Samanea tubulosa seedlings were observed with typical anthracnose symptoms in a forest nursery in São João Evangelista, Minas Gerais, Brazil. Based on morphological characteristics and phylogenetic analyses of ITS (5.8 S ribosomal RNA and the flanking internal transcribed spacer regions), act (actin), cal (calmodulin) and gapdh (glyceraldehyde‐3‐phosphate dehydrogenase) sequences, the fungus was identified as Colletotrichum karstii. Inoculation of the fungus in healthy S. tubulosa seedlings showed symptoms similar to those observed under natural conditions. To our knowledge, this is the first report of C. karstii causing anthracnose in S. tubulosa worldwide.

Host and Tissue Affiliations of Culturable Endophytic Fungi Associated with Xerophytic Plants in the Desert Region of Northwest China

Endophytic fungi isolated from plants under drought stress have been evidenced to confer hosts adaptive benefits to withstand drought. In this study, endophytic fungi associated with five typical desert shrubs in different tissue from extremely arid desert in Northwest China were investigated based on ITS sequence analysis. A total of 158 endophytic fungal stains were isolated from 1088 tissue segments of 12 samples, and 28 taxa represented by 25 species and 15 genera were identified as Ascomycetes. Alternaria sp. was the dominant genus with generic abundance ranging from 20% to 65%. The colonization rate of root was significantly lower, but the root-endophytic fungi (19 species) conversely presented a higher diversity than stem and leaf (11 and 7 species, respectively). Endophytic fungi had pronounced relative host and tissue preferences, while tissue explained more endophytic fungal variation than plant species. Additionally, soil pH, organic carbon, and phosphatase elicited significant responses from fungal species, which significantly affected the species richness of Fusarium redolens, Alternaria chlamydospore, Didymella glomerata, and Xylariales sp. This research provides a basis for the further understanding of the ecological distribution of endophytic fungi associated with xerophytic plants and their potential application for vegetative restoration and agricultural cultivation in drylands.

Endophytic fungi from Dongxiang wild rice (Oryza rufipogon Griff.) show diverse catalytic potential for converting glycyrrhizin.

Endophytic fungi inhabiting niche environments are novel biocatalyst resources that need to be exploited urgently. In this study, 63 endophytic fungi isolated from Dongxiang wild rice (Oryza rufipogon Griff.) were tested to assess their potentials to transform glycyrrhizin (GL) into glycyrrhetinic acid monoglucuronide (GAMG) or glycyrrhetinic acid (GA), of which 12 strains were shown to have β-d-glucuronidase activity. Based on morphological characteristics and rDNA ITS sequence analysis, the strains S59, L138, L55 and R57 with high GL molar conversion rates (55%, 45%, 65% and 89%) were further identified as Microsphaeropsis arundinis S59, Penicillium rubens L138, Aspergillus flavus L55 and Eupenicillium javanicum R57, respectively. These four strains with four different types of GL conversion processes were identified, i.e., (1) GL → GAMG in M. arundinis S59, (2) GL → GAMG and GA in A. flavus L55, (3) GL → GA in P. rubens L138, and (4) GL → GAMG → GA in E. javanicum R57, in which the bioconversion type (4) is reported for the first time. The study not only provided abundant and diverse β-d-glucuronidase resources that can be used for GL bioconversion, especially for GAMG biosynthesis from endophytic fungi, but also expanded our knowledge of potential roles of endophytes as new biocatalysts in biotransformation.

Biocontrol of Colletotrichum brevisporum in soybean using a new genistein-producing Paecilomyces strain

Colletotrichum brevisporum is a hazardous fungal pathogen that can cause anthracnose disease in several crops. In this study, Paecilomyces maximus NJC01 was isolated from C. brevisporum-infected soybean plants, and identified by ITS and TUB2 sequencing and morphological analysis. Interestingly, NJC01 inhibited the mycelial growth of plant fungal pathogens C. brevisporum, Sclerotinia sclerotiorum, Ceratocystis fimbriata and Valsa pyri. NJC01 showed various antifungal mechanisms against C. brevisporum, including competition for space and nutrients and the production of secondary metabolite genistein, which was detected at 91.8 µg/mL in the culture medium. NJC01 could colonize wounds in soybean pods, maintained its population in the inoculation site, and did not cause necrotic lesions. Combination of 1 × 108 cells/mL NJC01 and heat treatment reduced C. brevisporum lesions by 45%. The first biocontrol strategy for the management of C. brevisporum is reported in this study. This study provides new insights on the use of Paecilomyces species for the biocontrol of anthracnose disease.

First Report of Diaporthe cercidis Causing Leaf Blotch of Acer pictum subsp. mono in China.

Acer pictum subsp. mono (Maxim.) H. Ohashi is a common deciduous tree species that is widely distributed in Northeast and Northern China, including all provinces of the Yangtze River Basin (Liu et al. 2014). A foliar disease, with an incidence of ~90% (19/21 trees), occurred on A. pictum subsp. mono in a community park, Nanjing, Jiangsu, China in July 2019. On average, ~80% of the leaves per individual tree were infected by this disease. The symptoms initially appeared as brown, necrotic lesions at leaf tips, and half the leaf would become dark brown with time, and finally almost all of leaves were infected. Small pieces of leaf tissue (3 to 5 mm2) cut from the lesion margins were surface-sterilized in 75% ethanol for 30 s and 1% NaClO for 90 s, rinsed with sterile H2O three times, and placed on potato dextrose agar (PDA) at 25°C in the dark. The same fungus was isolated from 92% of the samples. The pure cultures were obtained by single-spore isolation. Three representative isolates (WJF1, WJF3 and WJF4) were obtained, and WJF1 was deposited in China's Forestry Culture Collection Center (CFCC 54806), and WJF3 and WJF4 were deposited at the Nanjing Forestry University (NFU 083 and NFU 084). The culture on PDA was white, with white vigorous aerial mycelia at the edge. Black pycnidia developed on the alfalfa stems at 25°C under a 14/10 h light/dark cycle for 20 days. Conidiophores were hyaline, branched, septate, straight, 16.4-34.7 × 1.5-3.0 μm (n = 30). Conidiogenous cells were 9.0-24.6 × 1.3-2.3 μm (n = 30). Alpha conidia were 7.0 ± 0.6 × 2.2 ± 0.2 μm (n = 30), fusiform, hyaline, smooth and multi-guttulate. Beta conidia were 25.5 ± 4.3 × 1.3 ± 0.1 μm (n = 30), hyaline, smooth and hamate. Morphological characters of all three isolates matched those of Diaporthe spp. (Gomes et al. 2013). DNA of three isolates were extracted and the internal transcribed spacer region (ITS), partial sequences of elongation factor 1-alpha (EF1-α), calmodulin (CAL), beta-tubulin (β-tub) and histone H3 (HIS) genes were amplified with primers ITS1/ITS4 (White et al. 1990), EF1-728F/EF1-986R and CAL228F/CAL737R (Carbone et al. 1999), βt2a/βt2b and CYLH3F/H3-1b (Glass and Donaldson, 1995, Crous et al. 2004), respectively. The sequences of WJF1, WJF3 and WJF4 were deposited in GenBank (WJF1: Accession Nos. MW301339 for ITS, MW363932 to MW363935 for EF1-α, β-tub, HIS, and CAL; WJF3: MW453062 and MW561566 to MW561569; WJF4: MW453063 and MW561570 to MW561573). BLAST results showed that the ITS, EF1-α, β-tub, HIS, and CAL sequences of WJF1 were similar with sequences of Phomopsis liquidambari C.Q. Chang, Z.D. Jiang & P.K. Chi JQ676191 (identity = 540/540; 100%), D. huangshanensis H. Zhou & C.L. Hou MN224671 (identity = 291/292; 99%), D. pescicola Dissan., J.Y. Yan, Xing H. Li & K.D. Hyde MK691230 (identity = 438/438; 100%), D. spinosa Y.S. Guo & G.P. Wang MK726170 (identity = 437/438; 99%), D. cercidis C.M. Tian & Qin Yang MK691114 (identity = 452/452; 100%), respectively. BLAST results of WJF3 and WJF4 are list in Table 1. A maximum likelihood and Bayesian posterior probability analyses using IQtree v. 1.6.8 and Mr. Bayes v. 3.2.6 with the concatenated sequences placed WJF1, WJF3 and WJF4 in the clade of D. cercidis. Based on the five-locus phylogeny and morphology, WJF1, WJF3 and WJF4 were identified as D. cercidis. The pathogenicity of three isolates were tested on potted 3-yr-old seedlings of A. pictum subsp. mono, grown in a greenhouse. Healthy leaves were wounded with a sterile needle and then inoculated with 10 μL of conidial suspensions (106 conidia/mL). Control leaves were treated with sterilized H2O. A total of twelve seedlings were used for the tests, 3 seedlings per treatment, and five leaves were inoculated per seedling. Each plant was covered with a plastic bag after inoculation and sterilized H2O was sprayed into the bag twice/day to maintain humidity and kept in a greenhouse at the day/night temperatures at 25 ± 2°C/16 ± 2°C. In 5 days, all the inoculated leaves had lesions similar to those observed in the field. D. cercidis was reisolated from the lesions of the inoculated leaves and was confirmed based on morphological characteristics and ITS sequence analysis. No symptoms were observed on the control leaves, and no fungus was isolated from them. D. cercidis was previously reported on twigs of Cercis chinensis (Yang et al. 2018) and causing pear shoot canker (Guo et al. 2020). This is the first report of D. cercidis causing leaf blotch on A. pictum subsp. mono. Identification of the pathogen is imperative for diagnosing and controlling this potentially high risk disease on A. pictum subsp. mono and also for the future studies.

Two new species of Fusicolla (Hypocreales) from China

Two new Fusicolla species are described and illustrated based on strains isolated from soil and rotten twig. Morphological features and DNA sequence analyses of combined ITS and LSU regions support their placements in Fusicolla and recognition of new species. Fusicolla gigas sp. nov. is characterized by C-shaped, 0−4(−5)-septate microconidia, falcate, (1–)4–9-septate macroconidia up to 80 µm long, and pinkish orange colonies on PDA. Fusicolla guangxiensis sp. nov. is distinguished by long-fusiform, falcate, (0–)1(–3)-septate macroconidia with acute ends, and orange colonies on PDA. Their macroconidia produced on OA are obvious larger than those on PDA. Distinctions between the new species and their close relatives are compared in detail.

A new endophytic fungus CJAN1179 isolated from the Cholistan desert promotes lateral root growth in Arabidopsis and produces IAA through tryptophan-dependent pathway

Fungi, important for growth of plants in arid lands, are expected to be involved in novel biochemical activities during fungal–plant interactions. We isolated 150 fungi associated with rhizosphere and root endosphere of two perennial grasses, Cymbopogon jwarancusa and Panicum antidotale, from Cholistan desert. The isolates were screened for their impact on plant growth and development using Arabidopsis thaliana (Col-0) as a model system. A root-endophytic fungus CJAN1179 from C. jwarancusa showed the highest plant growth-promoting effects. The most remarkable was enhanced number of lateral roots (3.1-fold). CJAN1179 produced indole-3-acetic acid (IAA) particularly in the presence of tryptophan. ITS sequence and phylogenetic analysis characterisation suggested the fungus to be a new species within Sordariomycetidae. CJAN1179 appears to promote plant growth by secreting IAA using tryptophan as a precursor. This fungus can be further explored for its suitability to promote growth of commercially important crops, particularly in arid regions.

Bioinformatic characterization of a triacylglycerol lipase produced by Aspergillus flavus isolated from the decaying seed of Cucumeropsis mannii

Lipases are enzymes of industrial importance responsible for the hydrolysis of ester bonds of triglycerides. A lipolytic fungus was isolated and subsequently identified based on the ITS sequence analysis as putative Aspergillus flavus with accession number LC424503. The gene coding for extracellular triacylglycerol lipase was isolated from Aspergillus flavus species, sequenced, and characterised using bioinformatics tools. An open reading frame of 420 amino acid sequence was obtained and designated as Aspergillus flavus lipase (AFL) sequence. Alignment of the amino acid sequence with other lipases revealed the presence GHSLG sequence which is the lipase consensus sequence Gly-X1-Ser-X2-Gly indicating that it a classical lipase. A catalytic active site lid domain composed of TYITDTIIDLS amino acids sequence was also revealed. This lid protects the active site, control the catalytic activity and substrate selectivity in lipases. The 3-Dimensional structural model shared 34.08% sequence identity with a lipase from Yarrowia lipolytica covering 272 amino acid residues of the template model. A search of the lipase engineering database using AFL sequence revealed that it belongs to the class GX-lipase, superfamily abH23 and homologous family abH23.02, molecular weight and isoelectric point values of 46.95 KDa and 5.7, respectively. N-glycosylation sites were predicted at residues 164, 236 and 333, with potentials of 0.7250, 0.7037 and 0.7048, respectively. O-glycosylation sites were predicted at residues 355, 358, 360 and 366. A signal sequence of 37 amino acids was revealed at the N-terminal of the polypeptide. This is a short peptide sequence that marks a protein for transport across the cell membrane and indicates that AFL is an extracellular lipase. The findings on the structural and molecular properties of Aspergillus flavus lipase in this work will be crucial in future studies aiming at engineering the enzyme for biotechnology applications. Communicated by Ramaswamy H. Sarma

Curvularia coatesiae XK8, a Potential Bioadsorbent Material for Adsorbing Cd(II) and Sb(III) Compound Pollution: Characteristics and Effects.

Soil heavy metal pollution is a common problem in mining areas. The soil of the Xikuangshan located in Lengshuijiang, Hunan Province, China contains various excessive heavy metals, especially antimony and cadmium. Previous studies have shown that heavy metal-tolerant microorganisms screened from mining areas have the potential to adsorb heavy metals. In this study, we screened out a cadmium and antimony tolerant fungus named XK8 from the slags collected from the Xikuangshan. Then, we explored the single and binary biosorption characteristics of Cd(II) and Sb(III) on it. In our results, the fungus XK8 was identified as Curvularia coatesiae XK8 by ITS sequencing analysis. Under the optimal conditions, in binary biosorption of the XK8, the main effect of the initial cadmium concentration on the cadmium removal rate of XK8 is negative, while the main effect of the initial antimony concentration, biosorption time, and initial pH on the cadmium removal rate of XK8 is positive. The initial pH has the greatest impact on the biosorption of cadmium on XK8, followed by the biosorption time; moreover, the effects of both are stronger than the coexisting ions. SAS analysis shows that under the optimal conditions, the theoretical maximum cadmium removal rate of XK8 is 100%, and the actual removal rate is 67.57%. Compared to the single biosorption with binary biosorption, the maximum biosorption capacity of XK8 for cadmium in the composite biosorption system increased to 23.6 mg g–1. It shows that under the background of high antimony, Sb(III) has a promoting effect on the biosorption of Cd(II) on XK8. In summary, a cadmium and antimony tolerant fungus with strong cadmium biosorption ability under the background of high antimony was screened out. It provides a potential microbial material for the bioremediation of heavy metal pollution.

Neohygrocybe pseudoingrata, a new grassland species from Slovakia and the Czech Republic.

Neohygrocybe pseudoingrata, a new waxcap species known from Slovakia and the Czech Republic, is characterised by its pale greyish coloured and often robust basidiomata (or sporocarps), nitrous smell, context without colour changes, hollow, contorted and compressed stipe and smooth or slightly fibrillose pileus surface. Based on morphology and DNA analysis of ITS and LSU sequences of the collected specimens, N. pseudoingrata belongs to Neohygrocybe sect. Neohygrocybe together with N. ovina, N. nitrata and N. ingrata. Collections of N. pseudoingrata form a well-supported clade in phylogenetic trees. Citation: Fuljer F, Zajac M, Boertmann D, Szabóová D, Kautmanová I (2022). Neohygrocybe pseudoingrata, a new grassland species from Slovakia and the Czech Republic. Fungal Systematics and Evolution 9: 11-17. doi: 10.3114/fuse.2022.09.02.

First report of Sclerotinia sclerotiorum on watercress (Nasturtium officinale) in aquaponic system in Hungary.

Watercress (Nasturtium officinale) is an aquatic dicotyledonous vegetable belonging to Brassicaceae (Aiton 1812). Watercress was grown in an aquaponic system on fired clay ball medium at the Aquaponic Research Station of the University of Debrecen, in the city of Debrecen (Hungary). During January 2020, 3-month-old plants showed symptoms in aquaponic cultivation. A visual survey showed 30% of plants with symptoms. Leaves and stems withered and showed white cotton-like mycelium. Mycelia from infected plants were placed on potato dextrose agar (PDA) and incubated at 25°C for seven days. Single hyphal tips were transferred to produce a pure culture. All ten fungal isolates showed similar morphological characteristics on PDA. Colonies consisted of white mycelia after three days and globoid to irregular and black 2.5 to 7 (average, 3) mm (n = 100 from ten plates) sclerotia formed ten days later, which are the typical morphological features of Sclerotinia sclerotiorum (Mordue et al. 1976). Molecular identification was performed with one of the ten isolates (Scl_B). Mycelia were grown in 250 ml of potato dextrose broth in a rotary shaker at 175 rpm at 24°C for six days. DNA was extracted from mycelium using a Nucleospin plant II (Macherey-Nagel, Germany) according to the manufacturer's protocol. PCR amplification (Kim et al. 2014) was performed with primers ITS1/ITS4 for the internal transcribed spacer region (White et al. 1990) on a Primus 96 thermal cycler (MWG Biotech, Germany). Specific polymerase chain reaction was performed with primers SSasprF/SSasprR (Abd-Elmagid et al. 2013). PCR products were sequenced by Microsynth Austria GmbH. NCBI BLAST analysis of the 440-bp ITS sequence (Genbank MW012403.1) showed 100% identity with the sequence of S. sclerotiorum (MT177267.1, etc.). The 170-bp specific gene sequence (Genbank MW959042.1) had a 100% similarity to hypothetical proteins (Genbank MK028159.1), with a 99.4% similarity to a portion of the S. sclerotiorum aspartyl protease gene (AF271387.1). Pathogenicity tests were carried out by inoculating surface-disinfested, 30-day-old watercress plants in plastic pots (15x15x12 cm). In three repeated experiments 90 watercress plants were measured. 15 plants (one plant per pot) were planted into the five-times autoclaved substrate (Biorgmix: pH 6.1±0.5%, N:1.5%, P2O5:0.7%, K2O:0.5%, organic matter content:50%) and inoculated by ten wheat kernels that were colonized by S. sclerotiorum (Scl_B) (Garibaldi et al. 2019). 15 plants were planted into the substrate with ten non-inoculated kernels as a control. Plants were kept in an MLR-352 climatic test chamber (PHCbi, Japan) at 21 ± 1°C for 12 hr light:dark cycle. On the first day of the experiment complex nutrient solution (Tek-Land: N:5%, P2O5:5%, K2O:5%, B:0.01%, Cu:0,01%, Mn:0.02%, Mo:0.002%, Zn:0.016%) was used, then autoclaved water daily. Eight days later white mycelium appeared on every inoculated plant and five days later dark sclerotia formed on the stems. Based on the morphological characteristics the re-isolated pathogen was S. sclerotiorum. Similar results were detected in three repeated experiments with white mold fungus being reisolated from all 45 infected watercress plants. The 45 non-inoculated plants did not show any symptoms and any diseases. This pathogen has already been reported on watercress in the field (Farr et al. 1989; Boland and Hall 1994; Garibaldi et al. 2019). This is the first reported case of white mold on watercress in aquaponic system in Hungary.

Screening and Identification of Indigenous Entomopathogenic Fungal Isolates from Agricultural Farmland Soils in Nile Delta, Egypt.

The compound negative impact of insect pests attacking agricultural ecosystems includes (i) direct yield losses from damaged crops, (ii) the economic cost of the attempt to prevent these losses and (iii) the negative short- and long-term hazard effects of chemical pesticides on human and environmental health. Entomopathogenic fungi (EMPF) are a group of microorganisms that represent the natural enemies of a number of crop pests, presenting an opportunity to harness their evolutionary fine-tuned relationship with their insect hosts as biocontrol agents in integrated pest management programs. The aim of this study was to establish an indigenous EMPF collection via the Galleria mellonella (greater wax moth) entrapment method from the soils of Nile Delta, Egypt. Obtained insect associated fungal isolates were bio-assayed for pathogenicity against the serious pest Spodoptera litura and Tenebrio molitor, and the seven outperforming isolates were selected for molecular identification and thermotolerance assay. Based on ITS sequence analysis and phylogeny, selected isolates were identified as Beauveria bassiana (four isolates), Metarhizium anisopliae (two isolates) and one isolate of Cordyceps javanica. The obtained results demonstrated (i) the efficacy of using insect baiting coupled with molecular identification and pathogenicity screening to isolate EMPF to control insect pests, and (ii) the availability of indigenous virulent EMPF in Nile Delta’s soil, which can be exploited for the development of sustainable crop protection strategies.