Four-locus phylogeny of Fusarium avenaceum and related species and their species-specific identification based on partial phosphate permease gene sequences

Stakheev, Alexander A.; Khairulina, D. R.; Завриев, С. К.

doi:10.1016/j.ijfoodmicro.2016.02.012

Cited by 29 publications

(25 citation statements)

References 81 publications

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“…This result agrees with the data in an earlier study of the polymorphism of other genes involved in mycotoxin biosynthesis, e.g. TRI1 [ 7 ], or the gene of enniatin synthetase ( Esyn1 ), a key enzyme of enniatin biosynthesis in F. avenaceum and closely related species [ 28 , 59 ].…”

Section: Discussionsupporting

confidence: 92%

“…Prior to DNA isolation, monospore cultures of fungi were grown on potato sucrose agar (PSA) at room temperature for 10 days, until abundant mycelium was obtained. DNA was isolated from monospore fungal cultures by a method based on the use of cetyltrimethylammonium bromide as a detergent, with allowance for the modifications described earlier [ 28 ]. The concentration and purity of DNA samples were evaluated using a NanoVue spectrophotometer (GE HealthCare, USA).…”

Section: Methodsmentioning

confidence: 99%

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Phylogenetic Analysis and Molecular Typing of Trichothecene-Producing Fusarium Fungi from Russian Collections

et al. 2018

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We performed a three-locus phylogenetic analysis of Fusarium strains presumably capable of trichothecene production, which were deposited in the Russian national collections. The intra- and interspecific polymorphism of partial sequences of the translation elongation factor 1 alpha (TEF1α) gene and two genes from the trichothecene cluster TRI5 and TRI14 was studied. A study of 60 strains of different origins using DNA markers confirmed, and in the case for several strains, clarified their taxonomic characteristics. As a result, a strain of F. commune (F-900) was identified in Russia for the first time. Furthermore, the strain F-846 proved to be phylogenetically distinct from any of the known Fusarium species. F. equiseti strains from Northwest Russia were found to belong to the North European group (I), whereas a strain from the North Caucasus – to the South European one (II). Partial TRI14 sequences from 9 out of 12 species were determined for the first time. Their comparative analysis demonstrated a relatively high level of intraspecific variability in F. graminearum and F. sporotrichioides, but no correlation between the sequence polymorphism and the geographic origin of the strains or their chemotype was found. Specific chemotypes of trichothecene B producers were characterized using two primer sets. The chemotyping results were verified by HPLC.

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Section: Discussionsupporting

confidence: 92%

Section: Methodsmentioning

confidence: 99%

Phylogenetic Analysis and Molecular Typing of Trichothecene-Producing Fusarium Fungi from Russian Collections

et al. 2018

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“…Каждый из образцов ДНК был протестирован в трех повторностях. Состав ПЦР-буфера и методика разделения продуктов амплификации с помощью гель-электрофореза описаны в более ранних публикациях (32,33).…”

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РАЗРАБОТКА НОВЫХ СИСТЕМ ПЦР-ИДЕНТИФИКАЦИИ НЕКАРАНТИННЫХ ПАТОГЕНОВ КАРТОФЕЛЯ (Solanum tuberosum L.), РАСПРОСТРАНЕННЫХ НА ТЕРРИТОРИИ РОССИИ

Stakheev¹

2020

S-h. biol.

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Российская Федерация-один из крупнейших производителей картофеля в мире: за 2018 год посевные площади этой культуры на территории страны составили 310,7 тыс. га, а валовые сборы в промышленном секторе-7157 тыс. т. Однако для современного картофелеводства характерно прогрессивное распространение вирусных болезней и бактериозов, приводящих к снижению урожая и катастрофическому ухудшению стандартного качества клубней. В настоящее время в картофелеводстве широко применяются классические методы диагностики, в частности растения-индикаторы, серологические и цитологические подходы. Они относительно надежны, но не всегда достаточно чувствительны, требуют высокой квалификации персонала и существенных затрат времени. Современной альтернативой этим методам служат диагностические системы на основе полимеразной цепной реакции (ПЦР), в частности ее модификации в формате реального времени (количественная ПЦР). В представленном исследовании для шести некарантинных патогенов (некротических штаммов Y вируса картофеля PVY N-NTN и PVY N:O , вируса погремковости табака, патогенных бактерий Dickeya solani, D. dianthicola, Pectobacterium atrosepticum) впервые в России были разработаны системы диагностики и идентификации в формате количественной ПЦР. Праймеры и флуоресцентно меченные зонды подбирали на основе последовательностей нуклеотидов, представленных в международной базе данных NCBI GenBank, с учетом размера ампликона не более 500 п.н. Специфичность предложенных систем была показана в тестах с генетическим материалом патогенов, таксономически близких или занимающих сходные экологические ниши: ординарный штамм PVY, вирус метельчатости верхушки картофеля, бактерии Pectobacterium carotovorum, D. zeae. Качество работы предложенных тест-систем также оценивали с использованием растительного материала, предположительно зараженного анализируемыми патогенами. Выделение нуклеиновых кислот осуществляли с использованием комплектов реактивов Проба-НК (для РНК) и Проба-ГС (для ДНК) (ООО «АгроДиагностика», Россия). Для проведения реакции обратной транскрипции использовали набор RevertAid Premium First Strand cDNA Synthesis Kit («Thermo Scientific», США). ПЦР проводили в амплификаторе Терцик («ДНК-технология», Россия), количественную ПЦР-в детектирующем амплификаторе ДТ-96 («ДНК-технология», Россия). Для оценки возможного ингибирования в реакционную смесь добавляли внутренний контрольный образец (ВКО), представляющий собой плазмиду со специфической вставкой (размер 560 п.н.). Положительные контрольные образцы (ПКО) клонировали с использованием набора Quick-TA kit («Евроген», Россия). Концентрацию плазмидной ДНК определяли с помощью спектрофотометра NanoVue («GE HealthCare», США). Секвенирование молекул ДНК проводили на автоматическом секвенаторе ABI PRISM 3730 («Applied Biosystems», США). Аналитическую чувствительность оценивали с использованием количественной ПЦР, при которой в качестве матрицы выступали последовательные 10-кратные разведения плазмидных ДНК (ПКО в четырех независимых повторностях) в диапазоне от 10 7 до 10 0 копий на реакцию. Бы...

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“…These are genes for 1 translation elongation factor (tef1) [30], β-tubulin (β-tub) [31], phosphate permease (РНО) [32,33], sterol 14-demethylase (CYP51C) [19], aminoadipate reductase (lys2) [20,21], intergenic spacers (IGS) of rRNA genes [22], and a number of genes involved in cascade reactions of chemical synthesis. Due to high inter-and intraspecific polymorphism they can be successfully used in phylogenetic studies of the genus.…”

mentioning

confidence: 99%

“…At the same time PHO gene is polymorphic enough to clearly distinguish F. culmorum from the closely related F. cerealis [41]. We used PHO as a marker for intra-and interspecific polymorphism study in F. avenaceum, F. tricinctum, F. acuminatum, F. torulosum, producing toxins of enniatin group and moniliformin [32]. It has been shown that for this group of species PHO has a higher variability than the «classical» markers such as tef1, β-tub, or enniatin synthetase (Esyn1) genes.…”

mentioning

confidence: 99%

MOLECULAR GENETIC APPROACHES FOR INVESTIGATION OF TAXONOMY AND SPECIFIC IDENTIFICATION OF TOXIN-PRODUCING Fusarium SPECIES: ACHIEVEMENTS AND PROBLEMS (review)

Stakheev¹,

Samokhvalova²,

Ryazantsev³

et al. 2016

S-h. biol.

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A b s t r a c tAscomycetous fungi of the genus Fusarium, which was separated as a single taxonomic group in 1809 by Heinrich Friedrich Link, are distributed worldwide and have different ecological niches. They have been found in soil and plants as endophytes, saprotrophs and parasites. Infections of agricultural plants, caused by Fusarium fungi, lead to annual damage, estimated in hundred million dollars. In addition to decrease harvest quality and quantity, Fusarium fungi are able to produce toxic metabolites (fusariotoxins). The consumption of fusariotoxin-contaminated winter grain led to deaths of tens of thousands of people in Volga and Ural regions in 1930s-1940s. These factors make necessary investigation of morphology, biochemistry and genetics of those organisms. Nowadays there is no universal taxonomic system of the genus Fusarium, so it is not possible to identify an isolate to species level in many cases. The high variability of morphological structures, and, on the other side, their similarity for closely related species, are major problems for researchers which use classical methods for identification and systematization of this group of fungi. Today, the molecular methods, based on the use of specific DNA sequence, are playing an increasingly important role in Fusarium systematics. The application of this approach has led to establishment of a number of new species in the genus. T. Yli-Mattila et al. (2009, 2011 used multilocus phylogenetic analysis to describe the novel species F. sibiricum and F. ussurianum which were found in Siberia and Far East and proved to be morphologically similar to F. graminearum and F. langsethiae, respectively. The authors of this paper identified the species F. torulosum (morphologically similar to F. avenaceum) in Russia for the first time based on the use of DNA markers. It is no less important that the analysis of interand intraspecific polymorphism makes possible the development of highly-specific assays for molecular detection of Fusarium fungi, including ones which infect plants and produce mycotoxins. The use of these assays (for example, ones described by M. Nicolaisen et al., 2009 and A.A. Stakheev et al., 2011) allows not only to detect the contamination of plants by mycotoxin producers, but also to estimate the quantity of their content in a probe. At the same time, a lot of questions are still unanswered: particularly, simultaneous application of different species concepts (morphological, biological and phylogenetic) makes it difficult to develop universal taxonomic system of the genus; also the most informative and reliable marker, which would reflect the evolutionary history of the genus, has not been found. The main achievements and problems of Fusarium phylogenetics, and perspectives of the application of molecular approach for studies of this group of organisms are discussed in this review.

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Four-locus phylogeny of Fusarium avenaceum and related species and their species-specific identification based on partial phosphate permease gene sequences

Cited by 29 publications

References 81 publications

Phylogenetic Analysis and Molecular Typing of Trichothecene-Producing Fusarium Fungi from Russian Collections

Phylogenetic Analysis and Molecular Typing of Trichothecene-Producing Fusarium Fungi from Russian Collections

РАЗРАБОТКА НОВЫХ СИСТЕМ ПЦР-ИДЕНТИФИКАЦИИ НЕКАРАНТИННЫХ ПАТОГЕНОВ КАРТОФЕЛЯ (Solanum tuberosum L.), РАСПРОСТРАНЕННЫХ НА ТЕРРИТОРИИ РОССИИ

MOLECULAR GENETIC APPROACHES FOR INVESTIGATION OF TAXONOMY AND SPECIFIC IDENTIFICATION OF TOXIN-PRODUCING Fusarium SPECIES: ACHIEVEMENTS AND PROBLEMS (review)

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