scholarly journals Molecular Characterization of Fungi Associated with Dump Site Soil

2021 ◽  
pp. 19-30
Author(s):  
N. G. Ogbuji ◽  
A. E. Ataga ◽  
P. M. Tari-Ukuta ◽  
C. J. Olisedeme
Keyword(s):  
Aspergillus Fumigatus ◽  
Trichoderma Harzianum ◽  
Its Region ◽  
Molecular Techniques ◽  
Nucleotide Sequence Analysis ◽  
Genetic Constitution ◽  
Species Identity ◽  
Dump Site ◽  
Aspergillus Flavipes ◽  
Port Harcourt

Aims: A study was conducted to identify fungal species isolated from dumpsite soil in University of Port Harcourt using molecular techniques. Methodology: Molecular methods for determining the species of a fungus based on the amplification and sequencing of the internal subscribed spacer (ITS) region of the fungal rRNA operon using molecular markers was applied. Soil sample was collected from a dumpsite in the University of Port Harcourt, Rivers State, Nigeria. Isolation of fungi associated with the dumpsite soil was carried out using spread plate method. Fungal genomic DNA was extracted using Quick-DNA Fungal/Bacterial Miniprep kit. The ITS1-2 gene of the isolates was amplified by Polymerase Chain Reaction (PCR) using the primer pair; ITS4 and ITS5. Results: The sequences of the amplified ITS region were blasted against known sequences on the National Centre for Biotechnology Information (NCBI) database. Nucleotide sequence analysis revealed the species identity of the fungal isolates to be: Aspergillus fumigatus, Trichoderma harzianum, Aspergillus felis, Aspergillus templicola, Aspergillus flavipes, Aspergillus fumigatus and Cunninghamella binariae. Phylogenetic analysis was carried out to ascertain the relationship between the isolates and other closely-related isolates on GenBank. Isolates 2 (Trichoderma harzianum) and 7 (Cunninghamella binariae), 3 (Aspergillus felis) and 6 (Aspergillus fumigatus), and 4 (Aspergillus templicola) and 5 (Aspergillus flavipes) were found to be more closely related to each other. Conclusion: The molecular techniques employed successfully identified the organisms to the species level as these techniques are based on the genetic constitution of organisms. The result obtained from this study will complement the information on the fungal organisms associated with dumpsite soil.

scholarly journals Isolation and identification of Aspergilli causing Banana fruit rot

2021 ◽  
Vol 4 (1) ◽  
pp. 8-18
Author(s):  
Fatima Ali ◽  
◽  
Naureen Akhtar ◽  
Shazia Shafique ◽  
Sobiya Shafique ◽  
...  
Keyword(s):  
Genetic Analysis ◽  
Aspergillus Fumigatus ◽  
Fungal Pathogens ◽  
Its Region ◽  
Fruit Rot ◽  
Nucleotide Sequence Analysis ◽  
Banana Fruit ◽  
Isolation And Identification ◽  
Pure Cultures ◽  
Different Strains

Banana is a commercially and nutritionally important food crop worldwide and is affected by several fungal diseases. The most important post-harvest disease is fruit rotting. Fruit rot is responsible for significant losses in banana. The present study was therefore, designed to isolate and explore the mycoflora associated with banana fruit rot. For this, infected samples were grown on different media to obtain pure cultures of isolated fungal pathogens. Identifications were made initially on morphological basis and then confirmed by genetic analysis. A comprehensive study of micro and macroscopic features revealed four Aspergillus species with two of <i> Aspergillus fumigatus</i>, one of <i> Aspergillus flavus</i> and one of <i> Aspergillus niger</i>. Genetic analysis by Nucleotide sequence analysis of ITS region of rDNA was performed. The sequence alignment of two different isolates of <i> Aspergillus fumigatus</i> showed \(99\%\) homology to different strains deposited in Genbank i.e., \(004(\text{KU}321562.1)\), \(\text{SK}1(\text{KM}207771.1)\), and \(98\%\) homology to \(\text{AHBR}16(\text{KF}305755.1), \ \ \text{SF}8(\text{KX}011021.1).\)

scholarly journals Molecular characterisation of fungi associated with sewage-impacted soil

2021 ◽  
Vol 27 (2) ◽  
pp. 107-113
Author(s):  
N. G. Iyanyi ◽  
A. E. Ataga ◽  
E. A Obichi ◽  
S. C. Agbasoga
Keyword(s):  
Pcr Amplification ◽  
Its Region ◽  
Fungal Species ◽  
Molecular Techniques ◽  
Septic Tank ◽  
Nucleotide Sequence Analysis ◽  
Molecular Method ◽  
Base Pairs ◽  
Universal Primer ◽  
Fungal Isolates

The decay of faecal matter from a septic system causes the arousal of fungi in the surrounding soil. These fungi can cause diseases if there is sewage spillage containing untreated or improperly treated wastewaters. Molecular techniques of identification of fungi have shown to be more dependable than traditional methods of identifying fungal species. This study was carried out to identify the fungal species associated with soil obtained from sewage-impacted soil near a septic tank using both traditional cultural techniques and molecular method. Fungi associated with the soil samples were isolated using serial dilution and Potato Dextrose Agar (PDA) method. Deoxyribonucleic Acid (DNA) was extracted from the pure cultures of fungal isolates using Quick DNA Fungal/Bacterial Miniprep kit. Polymerase Chain Reaction (PCR) amplification of internal transcribed spacer (ITS) region of the fungal isolates was carried out using universal primer pair; ITS4 and ITS5. The PCR products were sequenced and the sequences were blasted against National Centre for Biotechnology Information database. The result of the nucleotide sequence analysis revealed the identity of the isolates as Trichoderma harzanium with 580 base pairs and Aspergillus welwitschiae with 560 base pairs. Sequences of the isolates were aligned and compared with sequences on GenBank and a phylogenetic tree was constructed. The cultural method only aided in suggesting the suspected genera of the isolates while the molecular method was able to identify the isolates to the species level. This study will promote the knowledge of the fungal species associated with sewage-impacted soil and also aid researchers in proffering ways to enhance the prevention/control of diseases associated with sewage spill. Keyword: Septic tank, fungi, soil, phylogeny, sequencing

scholarly journals Antifungal activity of several isolates of Trichoderma against Cladosporium and Botrytis

2018 ◽  
Vol 8 (1) ◽  
pp. 88-91 ◽  
Author(s):  
M. Skaptsov ◽  
S. Smirnov ◽  
M. Kutsev ◽  
O. Uvarova ◽  
T. Sinitsyna ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Trichoderma Harzianum ◽  
Its Region ◽  
Botanical Garden ◽  
Antagonistic Activity ◽  
Antagonistic Effect ◽  
Soil Samples ◽  
Morphological Observation ◽  
The South

<p><em>Trichoderma</em> isolates (SSBGT07, SSBGT08, SSBGT09, SSBGT10) were isolated from the soil samples of the South-Siberian Botanical Garden and identified using morphological observation and ITS region analysis as <em>Trichoderma harzianum</em>, <em>T. asperellum, T. ghanense</em>, and <em>T. longibranchiatum</em>. Antagonistic activity against <em>Cladosporium </em>sp. and<em> Botrytis </em>sp. was evaluated <em>in vitro</em>. All isolates showed antagonistic effect by competition against <em>Cladosporium </em>sp. <em>T. asperellum </em>and <em>T. longibranchiatum</em> showed antagonism against <em>Botrytis </em>sp. All isolates showed hyper sporulation on the sclerotia of <em>Botrytis</em> sp. (except the <em>T. ghanense</em>) and colonies of the <em>Cladosporium</em> sp. Our study provides new isolates that affect the <em>Cladosporium </em>sp. and<em> Botrytis </em>sp.</p>

scholarly journals Biosorption potential and molecular characterization of metal resistant autochthonous microbes from tannery solid waste

2021 ◽  
Author(s):  
Hajira Younas ◽  
Aisha Nazir ◽  
Zakia Latif ◽  
Janice E Thies ◽  
Muhammad Shafiq ◽  
...  
Keyword(s):  
Solid Waste ◽  
Its Region ◽  
Metal Resistance ◽  
Molecular Techniques ◽  
Bacterial Strains ◽  
Fungal Strains ◽  
Resistant Strains ◽  
Tannery Solid Waste ◽  
Clean Metal

This study encompasses isolation and screening of heavy metal-resistant fungal and bacterial strains from tannery solid waste (TSW). Twelve fungal strains and twenty-five bacterial strains were isolated from TSW. The growth of fungal strains was observed against different heavy metals ranging from 10 mg L -1 to 1050 mg L -1 and the growth of bacteria was observed in metal concentrations ranging from 10 mg L -1 to 1200 mg L -1 . Five multi-metal resistant fungal isolates belonging to the genus Trichoderma and ten bacterial isolates belonging to the genus Bacillus showed good metal resistance and biosorption potential. They were identified through molecular techniques, fungi based on ITS region ribotyping, and bacteria based on 16S rRNA ribotyping. The fungal strains were characterized as T. hamatum (TSWF-06), T. harzianum (TSWF-11), T. lixii (TSWF-02) and T. pseudokoningii (TSWF-03, TSWF-10). The bacterial strains were characterized as Bacillus xiamenensis (TSW-02), B. velezensis (TSW-05), B. piscis (TSW-06), B. safensis (TSW-10), B. subtilis (TSW-14, TSW-15, TSW-17) B. licheniformis (TSW-19), B. cereus (TSW-20) and B. thuringiensis (TSW-22). The fungal strains namely, T. pseudokoningii (TSWF-03) and T. harzianum proved to be two multi-metal resistant strains with good biosorption efficiency. Unlike fungi, bacterial strains showed metal specific resistance. The strains Bacillus xiamenensis , B. subtilis (TSW-14) and B. subtilis (TSW-15) showed good biosorption efficiency against Cr, B. safensis against Cu, B. piscis and B. subtilis (TSW-17) against Pb and B. licheniformis and B. thuringiensis against Zn. The autochthonous fungal and bacterial strains can therefore be employed to clean metal contaminated environments.

scholarly journals B chromosome in Plantago lagopus Linnaeus, 1753 shows preferential transmission and accumulation through unusual processes

2017 ◽  
Vol 11 (2) ◽  
pp. 375-392 ◽  
Author(s):  
Manoj K. Dhar ◽  
Gurmeet Kour ◽  
Sanjana Kaul
Keyword(s):  
B Chromosome ◽  
Unreduced Gametes ◽  
Molecular Techniques ◽  
Root Tip ◽  
Genetic Constitution ◽  
Preferential Transmission ◽  
Female Sex ◽  
Process Formation ◽  
Plantago Lagopus ◽  
Mother Cells

Plantagolagopus is a diploid (2n = 2x =12) weed belonging to family Plantaginaceae. We reported a novel B chromosome in this species composed of 5S and 45S ribosomal DNA and other repetitive elements. In the present work, presence of B chromosome(s) was confirmed through FISH on root tip and pollen mother cells. Several experiments were done to determine the transmission of B chromosome through male and female sex tracks. Progenies derived from the reciprocal crosses between plants with (1B) and without (0B) B chromosomes were studied. The frequency of B chromosome bearing plants was significantly higher than expected, in the progeny of 1B female × 0B male. Thus, the B chromosome seems to have preferential transmission through the female sex track, which may be due to meiotic drive. One of the most intriguing aspects of the present study was the recovery of plants having more chromosomes than the standard complement of 12 chromosomes. Such plants were isolated from the progenies of B chromosome carrying plants. The origin of these plants can be explained on the basis of a two step process; formation of unreduced gametes in 1B plants and fusion of unreduced gametes with the normal gametes or other unreduced gametes. Several molecular techniques were used which unequivocally confirmed similar genetic constitution of 1B (parent) and plants with higher number of chromosomes.

scholarly journals Gyrodactylus magadiensis n. sp. (Monogenea, Gyrodactylidae) parasitising the gills of Alcolapia grahami (Perciformes, Cichlidae), a fish inhabiting the extreme environment of Lake Magadi, Kenya

Parasite ◽  
2019 ◽  
Vol 26 ◽  
pp. 76
Author(s):  
Quinton Marco Dos Santos ◽  
John Ndegwa Maina ◽  
Annemariè Avenant-Oldewage
Keyword(s):  
New Species ◽  
Cichlid Fish ◽  
Soda Lake ◽  
Its Region ◽  
Extreme Environment ◽  
Molecular Techniques ◽  
Diurnal Fluctuation ◽  
Lake Magadi ◽  
A New Species ◽  
Alcolapia Grahami

A new species of Gyrodactylus von Nordmann, 1832 is described from the gills of Alcolapia grahami, a tilapian fish endemic to Lake Magadi. This alkaline soda lake in the Rift Valley in Kenya is an extreme environment with pH as high as 11, temperatures up to 42 °C, and diurnal fluctuation between hyperoxia and virtual anoxia. Nevertheless, gyrodactylid monogeneans able to survive these hostile conditions were detected from the gills the Magadi tilapia. The worms were studied using light microscopy, isolated sclerites observed using scanning electron microscopy, and molecular techniques used to genetically characterize the specimens. The gyrodactylid was described as Gyrodactylus magadiensis n. sp. and could be distinguished from other Gyrodactylus species infecting African cichlid fish based on the comparatively long and narrow hamuli, a ventral bar with small rounded anterolateral processes and a tongue-shaped posterior membrane, and marginal hooks with slender sickles which are angled forward, a trapezoid to square toe, rounded heel, a long bridge prior to reaching marginal sickle shaft, and a long lateral edge of the toe. The species is also distinct from all other Gyrodactylus taxa based on the ITS region of rDNA (ITS1–5.8s–ITS2), strongly supporting the designation of a new species. These findings represent the second record of Gyrodactylus from Kenya, with the description of G. magadiensis bringing the total number of Gyrodactylus species described from African cichlids to 18.

scholarly journals Morphological and molecular identification of ectomycorrhizal fungi associated with Persian oak (Quercus brantii Lindl.) tree

2020 ◽  
Vol 66 (No. 6) ◽  
pp. 244-251
Author(s):  
Behnaz Yousefshahi ◽  
Masoud Bazgir ◽  
Samad Jamali ◽  
Fatemeh Valizadeh Kakhki
Keyword(s):  
Its Region ◽  
Molecular Techniques ◽  
Molecular Methods ◽  
Morphological Identification ◽  
Tree Roots ◽  
Microscopic Features ◽  
Zagros Zone ◽  
Oak Tree ◽  
Ecm Fungi ◽  
Quercus Brantii

Identification of ectomycorrhizal (ECM) fungi in different ecosystems has major significance. In this research, to identity ECM fungi, we used two methods including the morphological method and the molecular method that is more precise. Basidiocarp collection of fungi associated with oak tree (Quercus brantii Lindl.) roots was carried out in the spring season 2016 and was identified by morphological and molecular methods. We also checked macroscopic and microscopic features and measured each structure using BioloMICS Measures software. To verify the morphological identification, the internal transcribed spacer (ITS) region was amplified by PCR using the primer pair ITS1/ITS4, and the sequences were analyzed. According to the morphological observations, the identified species were Amanita crocea, Boletus comptus, Tricholoma giganteum, and Inocybe rimosa. Besides, based on molecular techniques by comparing sequences, we identified five species out of the eight ones as A. crocea and other species as T. giganteum, I. rimosa and B. comptus. Both morphological and molecular methods are necessary for identifying ECM fungi associated with tree roots in the Zagros zone in the west of Iran.

Detection and Fate of Bacillus anthracis (Sterne) Vegetative Cells and Spores Added to Bulk Tank Milk†

2003 ◽  
Vol 66 (12) ◽  
pp. 2349-2354 ◽  
Author(s):  
MICHAEL L. PERDUE ◽  
JEFF KARNS ◽  
JIM HIGGINS ◽  
JO ANN VAN KESSEL
Keyword(s):  
Bacillus Anthracis ◽  
Protective Antigen ◽  
Lethal Factor ◽  
Raw Milk ◽  
Molecular Data ◽  
Molecular Techniques ◽  
Nucleotide Sequence Analysis ◽  
Vegetative Cells ◽  
Bulk Tank Milk ◽  
Bulk Tank

A preparation of Bacillus anthracis (Sterne strain) spores was used to evaluate commercially available reagents and portable equipment for detecting anthrax contamination by using real-time PCR and was used to assess the fate of spores added directly to bulk tank milk. The Ruggedized Advanced Pathogen Identification Device (RAPID) was employed to detect spores in raw milk down to a concentration of 2,500 spores per ml. Commercially available primers and probes developed to detect either the protective antigen gene or the lethal factor gene both provided easily read positive signals with the RAPID following extraction from milk with a commercially available DNA extraction kit. Nucleotide sequence analysis of the vrrA gene with the use of DNA extracted from spiked milk provided molecular data that readily identified the spores as B. anthracis with a 100% BLAST match to the Sterne and Ames strains and easily distinguished them from B. cereus. Physical-fate and thermal-stability studies demonstrated that spores and vegetative cells have a strong affinity for the cream fraction of whole milk. A single treatment at standard pasteurization temperatures, while 100% lethal to vegetative cells, had no effect on spore viability even 14 days after the treatment. Twenty-four hours after the first treatment, a second treatment at 72°C for 15 s reduced the viability of the population by ca. 99% but still did not kill all of the spores. From these studies, we conclude that standard pasteurization techniques for milk would have little effect on the viability of B. anthracis spores and that raw or pasteurized milk poses no obstacles to the rapid detection of the spores by molecular techniques.

scholarly journals Trichothecium roseum Causes Fruit Rot of Tomato, Orange, and Apple in Pakistan

Plant Disease ◽  
2014 ◽  
Vol 98 (9) ◽  
pp. 1271-1271 ◽  
Author(s):  
M. I. Hamid ◽  
M. Hussain ◽  
M. U. Ghazanfar ◽  
M. Raza ◽  
X. Z. Liu
Keyword(s):  
Sequence Analysis ◽  
Its Region ◽  
Molecular Techniques ◽  
Fruit Rot ◽  
Control Treatment ◽  
Distilled Water ◽  
Trichothecium Roseum ◽  
Fruit Samples ◽  
The World ◽  
Stereo Microscope

During a field survey of greenhouses and fresh markets in 2013, fruits of tomato, oranges, and apples exhibited rot symptoms with white mycelial growth and salmon-color sporulation in the vicinity of Sargodha city (32°5′1″ N, 72°40′16″ E), Pakistan. Diseased fruit samples were collected in plastic bags and taken to laboratory on ice for further diagnosis. Diseased fruits were observed under a stereo microscope and single spores were removed using an inoculating needle. Isolation from single spores showed pink to white colonies on potato dextrose agar (PDA) containing hyaline, 2-celled, ellipsoid to pyriform conidia (17 to 24 × 7 to 11 μm) with slanting and truncate basal mark and produced in clusters. Conidiophores were branched (105 to 254 × 2 to 4 μm) and hyphae were hyaline (3 to 5 μm in diameter). These characteristics of the fungus were similar to Trichothecium roseum (Pers.) as reported by Inácio et al. (1). Genomic DNA was extracted by using CTAB buffer from a single pure colony of one isolate of the fungus and PCR analysis was performed for ITS region and part of the 5′ end of the beta tubulin (TUB) gene (2,3). Single fragments of 550 bp and 1.5 kb length from ITS and TUB gene were amplified and sequenced (GenBank Accession Nos. KF975702 and KJ607590, respectively). Sequence analysis showed 99% similarity with T. roseum isolates from different regions of the world. Phylogenetic analysis (MEGA version 5.2 with WAG model) showed the close relatedness to the isolates of T. roseum from Pakistan and isolates from other parts of the world that revealed the low genetic variability of ITS region. TUB gene sequence analysis indicated 100% homology with isolates of T. roseum and to the other species in Hypocreales. Pathogenicity tests were performed on tomato cvs. Nova Mech and Rio Grande, orange cv. Kinnow, and on apple cv. Golden Delicious by inoculating five fruits from each cultivar. Spore suspensions (105 conidia/ml of sterilized distilled water) were inoculated into all wounded fruits (9 wounds/fruit) of each cultivar and incubated at 25°C for the development of symptoms. Five wounded fruits of each cultivar were inoculated with sterilized distilled water as a control treatment. The fruits were kept in plastic boxes and incubated in humid chambers for 5 days. The symptoms on apples were observed as brown rot with pinkish spores on rotted tissue. The cross section of apple fruits also showed the brown rotted tissues internally. The fungus developed mycelium and spores on the surface and caused severe rotting inside the tomato and citrus fruits. T. roseum was re-isolated by picking a single spore from rotted tissues of fruits under a stereo microscope, and culturing on PDA. The re-isolated fungus was confirmed morphologically and by molecular techniques. Tomato and apple has been reported as a host for T. roseum (1,4,5) but oranges have not. To our knowledge, this is the first record of T. roseum infecting tomato, oranges, and apples in Pakistan. References: (1) C. A. Inácio et al. Plant Dis. 95:1318. 2011. (2) K. O'Donnell, and E. Cigelnik. Mol. Phylogenet. Evol. 7:103, 1997. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990. (4) Y. H. Yun et al. Afr. J. Microbiol. Res. 7:1128, 2013. (5) M. Žabka et al. Mycopathologia. 162:65, 2006.

A molecular phylogeny of the subtribe Pterostylidinae (Orchidaceae): resolving the taxonomic confusion

2010 ◽  
Vol 23 (4) ◽  
pp. 248 ◽  
Author(s):  
Jasmine K. Janes ◽  
Dorothy A. Steane ◽  
René E. Vaillancourt ◽  
Marco F. Duretto
Keyword(s):  
Bayesian Methods ◽  
Phylogenetic Trees ◽  
Taxonomic Status ◽  
Its Region ◽  
Population Level ◽  
Molecular Techniques ◽  
The Past ◽  
Single Well ◽  
Single Genus ◽  
Taxonomic Confusion

In the past decade, two major classification schemes of the subtribe Pterostylidinae have resulted in taxonomic confusion because a single well known genus was split into a large number of new genera and subgenera, many of which are difficult to discriminate accurately. These classifications have not been accepted widely among systematists because of poor phylogenetic support for several genera. Analyses of the internal transcribed spacer (ITS) region of nuclear rDNA in a large number of species and samples facilitate further clarification of the relationships within the Pterostylidinae. The phylogenetic trees were reconstructed using parsimony and Bayesian methods. These phylogenetic trees indicate that subtribe Pterostylidinae is monophyletic, and support the concept of a single genus, Pterostylis R.Br. sensu lato within the Pterostylidinae. Two clades representing subgenera correlate with the morphology of the lateral sepals, whereas several of the previously erected genera consistently have poor support. The proposed subgenera are divided further into 10 sections. Several closely related taxa with identical ITS sequences require further scrutiny by population-level molecular techniques to determine their taxonomic status.

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