Seed-Associated Fungal Diversity and the Molecular Identification of Fusarium with Potential Threat to Ginseng (Panax ginseng) in China

The utility of traditional methods for detecting seed-borne fungi is limited by the fact some fungi are unculturable or difficult to isolate. The seed-borne pathogens affecting Panax ginseng cultivation have not been fully characterized. Seed-borne fungi can be identified based on the high-throughput sequencing of internal transcribed spacer (ITS) amplicons. A hierarchical clustering tree diagram analysis based on operational taxonomic units revealed a relationship between the seed-borne fungi and the region from which the seeds were collected. This study analyzed the fungal diversity on 30 ginseng seed samples from the main ginseng-producing areas of China. The 50 most abundant genera were identified including those responsible for ginseng diseases, Fusarium, Alternaria, Nectria, Coniothyrium, Verticillium, Phoma, and Rhizoctonia. Fusarium species, which are the primary causes of root rot, were detected in all seed samples. The results of a phylogenetic analysis indicated that the seed-borne fungal species originating from the same region were closely related. Fungi on ginseng seeds from eight different regions were divided into eight clades, suggesting they were correlated with the local storage medium. A total of 518 Fusarium isolates were obtained and 10 species identified, all of which can be detrimental to ginseng production. Pathogenicity tests proved that seed-borne Fusarium species can infect ginseng seedlings and 2-year-old ginseng root, with potentially adverse effects on ginseng yield and quality.

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Incidence and Etiology of Postharvest Fungal Diseases Associated with Bulb Rot in Garlic (Alllium sativum) in Spain

Foods ◽

10.3390/foods10051063 ◽

2021 ◽

Vol 10 (5) ◽

pp. 1063

Author(s):

Laura Gálvez ◽

Daniel Palmero

Keyword(s):

Negative Impact ◽

Fusarium Species ◽

Fungal Species ◽

Postharvest Disease ◽

Dry Rot ◽

Fusarium Dry Rot ◽

Causative Agents ◽

Postharvest Handling ◽

Pathogenicity Tests ◽

Bulb Rot

In recent years, different postharvest alterations have been detected in garlic. In many cases, the symptoms are not well defined, or the etiology is unknown, which further complicates the selection of bulbs during postharvest handling. To characterize the different symptoms of bulb rot caused by fungi, garlic bulb samples were collected from six Spanish provinces in two consecutive years. Eight different fungal species were identified. The most prevalent postharvest disease was Fusarium dry rot (56.1%), which was associated with six Fusarium species. Fusarium proliferatum was detected in more than 85% of symptomatic cloves, followed by F. oxysporum and F. solani. Pathogenicity tests did not show a significant correlation between virulence and mycotoxin production (fumonisins, beauvericin, and moniliformin) or the mycelial growth rate. Penicillium allii was detected in 12.2% of the samples; it was greatly influenced by the harvest season and garlic cultivar, and three different morphotypes were identified. Stemphylium vesicarium and Embellisia allii were pathogenic to wounded cloves. Some of the isolated fungal species produce highly toxic mycotoxins, which may have a negative impact on human health. This work is the first to determine the quantitative importance, pathogenicity, and virulence of the causative agents of postharvest garlic rot in Spain.

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Different Amplicon Targets for Sequencing-Based Studies of Fungal Diversity

Applied and Environmental Microbiology ◽

10.1128/aem.00905-17 ◽

2017 ◽

Vol 83 (17) ◽

Cited By ~ 40

Author(s):

Francesca De Filippis ◽

Manolo Laiola ◽

Giuseppe Blaiotta ◽

Danilo Ercolini

Keyword(s):

Microbial Ecology ◽

High Throughput ◽

Biological Samples ◽

Fungal Diversity ◽

High Throughput Sequencing ◽

Amplicon Sequencing ◽

Fungal Species ◽

Fungal Communities ◽

Its2 Region ◽

Mock Community

ABSTRACT Target-gene amplicon sequencing is the most exploited high-throughput sequencing application in microbial ecology. The targets are taxonomically relevant genes, with 16S rRNA being the gold standard for bacteria. As for fungi, the most commonly used target is the internal transcribed spacer (ITS). However, the uneven ITS length among species may promote preferential amplification and sequencing and incorrect estimation of their abundance. Therefore, the use of different targets is desirable. We evaluated the use of three different target amplicons for the characterization of fungal diversity. After an in silico primer evaluation, we compared three amplicons (the ITS1-ITS2 region [ITS1-2], 18S ribosomal small subunit RNA, and the D1/D2 domain of the 26S ribosomal large subunit RNA), using biological samples and a mock community of common fungal species. All three targets allowed for accurate identification of the species present. Nevertheless, high heterogeneity in ITS1-2 length was found, and this caused an overestimation of the abundance of species with a shorter ITS, while both 18S and 26S amplicons allowed for more reliable quantification. We demonstrated that ITS1-2 amplicon sequencing, although widely used, may lead to an incorrect evaluation of fungal communities, and efforts should be made to promote the use of different targets in sequencing-based microbial ecology studies. IMPORTANCE Amplicon-sequencing approaches for fungi may rely on different targets affecting the diversity and abundance of the fungal species. An increasing number of studies will address fungal diversity by high-throughput amplicon sequencing. The description of the communities must be accurate and reliable in order to draw useful insights and to address both ecological and biological questions. By analyzing a mock community and several biological samples, we demonstrate that using different amplicon targets may change the results of fungal microbiota analysis, and we highlight how a careful choice of the target is fundamental for a thorough description of the fungal communities.

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Best practices in metabarcoding of fungi: from experimental design to results

10.22541/au.163430390.04226544/v1 ◽

2021 ◽

Author(s):

Leho Tedersoo ◽

Mohammad Bahram ◽

Lucie Zinger ◽

Henrik Nilsson ◽

Peter Kennedy ◽

...

Keyword(s):

Experimental Design ◽

Best Practices ◽

Fungal Diversity ◽

High Throughput Sequencing ◽

Its Region ◽

Fungal Communities ◽

Sequence Variants ◽

Transcriptomic Data ◽

Operational Taxonomic Units ◽

Computational Analyses

The development of high-throughput sequencing (HTS) technologies has greatly improved our capacity to identify fungi and unveil their ecological roles across a variety of ecosystems. Here we provide an overview about current best practices in metabarcoding analysis of fungal communities, from experimental design through molecular and computational analyses. By re-analysing published datasets, we find that operational taxonomic units (OTUs) outperform amplified sequence variants (ASVs) in recovering fungal diversity, which is particularly evident for long markers. Additionally, analysis of the full-length ITS region allows more accurate taxonomic placement of fungi and other eukaryotes compared with the ITS2 subregion. We conclude that metabarcoding analyses of fungi are especially promising for co-analyses with the functional metagenomic or transcriptomic data, integrating fungi in the entire microbiome, recovery of novel fungal lineages and ancient organisms as well as barcoding of old specimens including type material.

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Characterization of Fusarium and Neocosmospora Species Associated With Crown Rot and Stem Canker of Pistachio Rootstocks in California

Plant Disease ◽

10.1094/pdis-11-18-2012-re ◽

2019 ◽

Vol 103 (8) ◽

pp. 1931-1939 ◽

Cited By ~ 4

Author(s):

Maria Crespo ◽

Daniel P. Lawrence ◽

Mohamed T. Nouri ◽

David A. Doll ◽

Florent P. Trouillas

Keyword(s):

Rna Polymerase Ii ◽

Fusarium Species ◽

Phylogenetic Analyses ◽

Elongation Factor ◽

Stem Canker ◽

The United States ◽

Fungal Species ◽

Crown Rot ◽

Pathogenicity Tests ◽

Pistachio Trees

California produces 99.1% of pistachios grown in the United States, and diseases affecting pistachio rootstocks represent a constant challenge to the industry. Field surveys of fungi associated with pistachio rootstocks with symptoms of crown rot and stem canker in three central California counties followed by phylogenetic analyses of translation elongation factor 1-α and second largest subunit of RNA polymerase II gene fragments identified three Fusarium species (Fusarium equiseti, Fusarium oxysporum, and Fusarium proliferatum) and two Neocosmospora species (Neocosmospora falciformis and Neocosmospora solani). F. oxysporum and N. falciformis were the fungal species most frequently recovered from symptomatic pistachio trees. Inoculations of detached twigs of cultivar Kerman pistachio Pioneer Gold I and clonal University of California, Berkeley I (UCBI) rootstocks showed that all five species could colonize pistachio wood and cause vascular discolorations. Pathogenicity tests in potted pistachio trees completed Koch’s postulates and confirmed that F. oxysporum, F. proliferatum, N. falciformis, and N. solani were capable of producing rot and discoloration in stems of clonal UCBI rootstocks, the most widely planted pistachio rootstock in California. To our knowledge, this study is the first to present insights into the biodiversity and biology of Fusarium and Neocosmospora species associated with pistachio trees in California.

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Field Fungal Diversity in Freshly Harvested Japonica Rice

International Journal of Biochemistry Research & Review ◽

10.9734/ijbcrr/2019/v26i130086 ◽

2019 ◽

pp. 1-14 ◽

Cited By ~ 1

Author(s):

Xiang Dong Sun ◽

Hong Shan ◽

Jing Lan ◽

Li Li Li ◽

Hai Tao Guan ◽

...

Keyword(s):

Fungal Diversity ◽

High Throughput Sequencing ◽

Fungal Species ◽

Climatic Conditions ◽

Rice Production ◽

Gibberella Zeae ◽

Japonica Rice ◽

Heilongjiang Province ◽

Significant Difference ◽

The Difference

Rice is a major food crop in China and Japonica rice production in Heilongjiang Province ranks No.1 in total annual rice production in the country. Rice is prone to invasion by fungi and mycotoxins produced by the fungi are proven to be serious threats to human health. The objective of the present study was to investigate fungal diversity of freshly harvested rice in the four main cultivation regions of Heilongjiang Province in order to find the difference of dominant fungi among the four regions. Through high throughput sequencing we detected Ascomycota accounts for absolute dominant phylum; Dothideomycetes, Sordariomycetes, Tremellomycetes, Microbotryomycetes, and Eurotiomycetes were dominant classes; Capnodiales, Hypocreales, and Pleosporales were the main orders; Cladosporiaceae, Pleosporaceae, Nectriaceae, Clavicipitaceae, Tremellaceae, Phaeosphaeriaceae, Trimorphomycetaceae, Sporidiobolaceae, Bionectriaceae, and Trichocomaceae were major family; Cladosporium, Epicoccum, Fusarium, and Alternaria were the most abundant phylotypes at genus level; Epicoccum nigrum, Gibberella zeae, and Fusarium proliferatum were the dominant fungal species. Great fungal diversity was observed in the rice samples harvested in the four major Japonica rice-growing regions in Heilongjiang province. However, no significant difference in diversity was observed among the four regions, likely due to the relatively close geographical proximity leading to very similar climatic conditions. Since some of the fungal species produce mycotoxins, it is necessary to take precautions to ensure the rice is stored under safe conditions to prevent the production of mycotoxins. This is the first report on investigation of field fungal diversity in freshly harvested Japonica rice in Heilongjiang Province in China.

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Endophytic Fungal Diversity on Different Aged Bark of Medicinal Taxus Plant Species

Journal of Biobased Materials and Bioenergy ◽

10.1166/jbmb.2019.1880 ◽

2019 ◽

Vol 13 (4) ◽

pp. 544-549

Author(s):

Xiaokuan Fan ◽

Surendra Sarsaiya ◽

Jing Yu ◽

Xiwu Pan ◽

Leilei Jin ◽

...

Keyword(s):

Sequence Analysis ◽

Endophytic Fungi ◽

Internal Transcribed Spacer ◽

Fungal Diversity ◽

Diversity Index ◽

Fungal Species ◽

Tree Age ◽

Distribution Data ◽

Operational Taxonomic Units ◽

Major Species

The present study investigated the endophytic biodiversity on different aged Taxus bark using highthroughput sequencing. Fungi from the five Taxus species, including Taxus yunnanensis, T. media, T. chinensis, and two unidentified Taxus species I and II, were identified by Internal Transcribed Spacer sequence analysis. The operational taxonomic units (OTUs) were 141, 114, 98, 67 and 36, respectively, while the genera numbers were 22, 14, 16, 9 and 10 from the species distribution data, respectively. The dominant fungal genera were Mycena (94.33%), Phageophora (43.97%) and Paraconiothyrium (34.02%), followed by Pestalotiopsis (1.58%) and Strelitziana (1.15%). Rankabundance curve and diversity index suggested that the major species from the bark of T. yunnanensis (YN) was endophytic fungi, whereas fungal species from the bark of unidentified Taxus species II (HDS 50) were quite rare. Furthermore, tree age notably influenced the bark endophytic fungal biodiversity.

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Toxicity, Pathogenicity, and Genetic Differentiation of Five Species of Fusarium from Sorghum and Millet

Phytopathology ◽

10.1094/phyto-95-0275 ◽

2005 ◽

Vol 95 (3) ◽

pp. 275-283 ◽

Cited By ~ 73

Author(s):

John F. Leslie ◽

Kurt A. Zeller ◽

Sandra C. Lamprecht ◽

John P. Rheeder ◽

Walter F. O. Marasas

Keyword(s):

Fusarium Species ◽

Toxin Production ◽

Fungal Species ◽

Banding Patterns ◽

Length Polymorphisms ◽

Amplified Fragment Length Polymorphisms ◽

Pathogenicity Tests ◽

Plant Pathogenicity ◽

Sorghum Seedling

Fusarium isolates recovered from sorghum and millet are commonly identified as F. moniliforme, but with the recognition of new species in this group, the strains given this name are being re-evaluated. We analyzed five strains each from five Fusarium species (F. andiyazi, F. nygamai, F. pseudonygamai, F. thapsinum, and F. verticillioides) often associated with sorghum and millet for their ability to produce fumonisin and moniliformin, their toxicity to ducklings, and their ability to cause disease on sorghum seedlings in vitro. These species can be distinguished with isozymes (fumarase, NADP-dependent isocitrate dehydrogenase, and malate dehydrogenase) and with banding patterns resulting from amplified fragment length polymorphisms. Two species, F. pseudonygamai and F. thapsinum, produced high levels of moniliformin, but little or no fumonisins, and were consistently highly toxigenic in the duckling tests. Two species, F. verticillioides and F. nygamai, produced high levels of fumonisins, but little or no moniliformin, and also were toxigenic in the duckling tests. F. andiyazi produced little or no toxin and was the least toxigenic in the duckling tests. In sorghum seedling pathogenicity tests, F. thapsinum was the most virulent followed by F. andiyazi, then F. verticillioides, and finally F. nygamai and F. pseudonygamai, which were similar to each other. Thus, these five species, which would once have all been called F. moniliforme, differ sufficiently in terms of plant pathogenicity and toxin production profile, that their previous misidentification could explain inconsistencies in the literature and differences observed by researchers who thought they were all working with the same fungal species.

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Preliminary analysis of amplicon high-throughput sequencing as a method for the assessment of fungal diversity in discolored wood

Holzforschung ◽

10.1515/hf-2017-0015 ◽

2017 ◽

Vol 71 (10) ◽

pp. 793-800 ◽

Cited By ~ 1

Author(s):

Xingxia Ma ◽

Mingliang Jiang ◽

Junliang Liu ◽

Hao Deng ◽

Shuangyong Wang

Keyword(s):

High Throughput ◽

Internal Transcribed Spacer ◽

Fungal Diversity ◽

Preliminary Analysis ◽

High Throughput Sequencing ◽

Fungal Species ◽

Quantitative Relation ◽

Core Microbiome ◽

Microbiome Analysis ◽

First Time

AbstractThe diversity of stain fungi is important if wood is inhabited with various fungi, and the discoloration mechanism will be better understood. MiSeq amplicon high-throughput sequencing (Illumina®) is able to detect species richness (the number of species within a community) and species evenness (the sizes of species populations within a community). This study detected fungal diversity in discolored Mongolian pine for the first time by the MiSeq approach, focusing on the nuclear ribosomal internal transcribed spacer-1 (ITS1). The results show that the discolored wood was inhabited by a combination of microorganisms, more than 90% of which belong toAscomycotafungi at the phylum level. The MiSeq method revealed not only all the inhabited fungal species but also their quantitative relation to each other. The dominant fungal species in sample A areHelotiales(34.1%) andHypocreales(20.7%). The dominant fungal species in sample B isNectriaceae(67.9%), whileHypocrea(34.7%) andSporothrix(27.6%) are the dominant fungal species in sample C. It was confirmed via core microbiome analysis that the following fungi were common stain fungi in the three discolored samples:Fusariumspp.,Aspergillusspp.,Sporothrixspp.,Penicilliumspp.,Trichodermaspp.,Alternariaspp. andCladophialophoraspp.

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Green Synthesis of Silver Nanoparticles by Allamanda cathartica L. Leaf Extract and Evaluation for Antimicrobial Activity

International Journal of Pharmaceutical Sciences and Nanotechnology ◽

10.37285/ijpsn.2013.6.4.9 ◽

2013 ◽

Vol 6 (4) ◽

pp. 2260-2268

Author(s):

M. Linga Rao ◽

Bhumi G ◽

Savithramma N

Keyword(s):

Antimicrobial Activity ◽

Silver Nanoparticles ◽

Leaf Extract ◽

Fusarium Species ◽

Inhibition Zone ◽

Fungal Species ◽

Green Technology ◽

Metallic Silver ◽

Synthesis Of Nanoparticles ◽

Aqueous Leaf Extract

Silver nanoparticles (SNPs) exhibit tremendous applications in medicine as antimicrobial agent. The use of different parts of plants for the synthesis of nanoparticles is considered as a green technology as it does not involve any harmful chemicals. In the present study, we report a rapid biosynthesis of silver nanoparticles from aqueous leaf extract of medicinal plant Allamanda cathartica. The active phytochemicals present in the plant were responsible for the quick reduction of silver ion to metallic silver nanoparticles. The reduced silver nanoparticles were characterized by using UV-Vis spectrophotometry, Scanning Electron Microscope (SEM), Energy Dispersive Analysis of X-ray (EDAX) and Atomic Force Microscopy (AFM). The spherical shaped silver nanoparticles were observed and it was found to 19-40 nm range of size. These phytosynthesized SNPs were tested for their antimicrobial activity and it analyzed by measuring the inhibitory zone. A. cathartica aqueous leaf extract of SNPs showed highest toxicity to Pseudomonas followed by Klebsiella, Bacillus and E. coli and lowest toxicity towards Proteus. In fungal species, highest inhibition zone was noted against Rhizopus followed by Curvularia, Aspergillus flavus and Aspergillus niger and minimum inhibition zone was observed against Fusarium species. These results suggest a promising potential of Indian plant-based green chemistry for production of SNPs for biomedical and nanotechnology applications.

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New Insights on the Zeugodacus cucurbitae (Coquillett) Bacteriome

Microorganisms ◽

10.3390/microorganisms9030659 ◽

2021 ◽

Vol 9 (3) ◽

pp. 659

Author(s):

Elias Asimakis ◽

Panagiota Stathopoulou ◽

Apostolis Sapounas ◽

Kanjana Khaeso ◽

Costas Batargias ◽

...

Keyword(s):

Relative Abundance ◽

Bacterial Communities ◽

High Throughput Sequencing ◽

Mutual Exclusion ◽

Bacterial Composition ◽

Operational Taxonomic Units ◽

Host Diet ◽

Dominant Bacteria ◽

High Degree

Various factors, including the insect host, diet, and surrounding ecosystem can shape the structure of the bacterial communities of insects. We have employed next generation, high-throughput sequencing of the 16S rRNA to characterize the bacteriome of wild Zeugodacus (Bactrocera) cucurbitae (Coquillett) flies from three regions of Bangladesh. The tested populations developed distinct bacterial communities with differences in bacterial composition, suggesting that geography has an impact on the fly bacteriome. The dominant bacteria belonged to the families Enterobacteriaceae, Dysgomonadaceae and Orbaceae, with the genera Dysgonomonas, Orbus and Citrobacter showing the highest relative abundance across populations. Network analysis indicated variable interactions between operational taxonomic units (OTUs), with cases of mutual exclusion and copresence. Certain bacterial genera with high relative abundance were also characterized by a high degree of interactions. Interestingly, genera with a low relative abundance like Shimwellia, Gilliamella, and Chishuiella were among those that showed abundant interactions, suggesting that they are also important components of the bacterial community. Such knowledge could help us identify ideal wild populations for domestication in the context of the sterile insect technique or similar biotechnological methods. Further characterization of this bacterial diversity with transcriptomic and metabolic approaches, could also reveal their specific role in Z. cucurbitae physiology.

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