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E M Scott - One of the best experts on this subject based on the ideXlab platform.
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in vitro susceptibility of trichophyton mentagrophytes Arthroconidia to clotrimazole and griseofulvin in human corneocyte suspensions
Mycoses, 2009Co-Authors: Salih H.m. Aljabre, G.s. Shankland, E M Scott, M D RichardsonAbstract:The use of corneocytes in suspension as a medium for the study of the effect of antifungal drugs on Trichophyton mentagrophytes Arthroconidia was investigated. In the presence of clotrimazole or griseofulvin, Arthroconidia germination in a suspension of corneocytes and in Sabouraud glucose liquid medium was significantly reduced compared with germination in drug-free media. Where antifungals were added to Arthroconidia after an activation period during which germination occurred, any further germination was inhibited. The data showed that the growth of T. mentagrophytes in the presence of corneocytes offered a simple, rapid, inexpensive and relevant model for the assessment of antifungal activity of compounds for the treatment of dermatophytosis.
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Early events in the invasion of the human nail plate by Trichophyton mentagrophytes.
The British journal of dermatology, 1995Co-Authors: A. Rashid, E M Scott, Malcolm D. RichardsonAbstract:A new in vitro model for the study of nail invasion by dermatophyte fungi was developed. The dermatophyte Trichophyton mentagrophytes, and fragments of finger-nails and toe-nails were used. Arthroconidia were inoculated on the ventral surface of the nails. After 6 h, adherence and germination of Arthroconidia was observed. By 16 h, small germ tubes with side branches were evident. At about 24 h, micro-colonies had become established. At 48 h, a mycelium had formed, and at about 72 h most of the nail fragment was covered with fungal growth. Nail penetration occurred from the ventral surface through the intercellular spaces, and with longer incubation all three layers were invaded by Arthroconidia growing through channels. Nail invasion occurred in the absence of added nutrients. Dermatophyte fungi appeared to invade the nail by a combination of mechanical and chemical factors. The model provides a substrate to study the pharmacokinetics and bioavailability of new antifungal agents in situ.
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Inhibitory effect of terbinafine on the invasion of nails by Trichophyton mentagrophytes.
Journal of the American Academy of Dermatology, 1995Co-Authors: A. Rashid, E M Scott, M D RichardsonAbstract:Terbinafine has a broad spectrum of action in vitro and primary fungicidal action against many pathogenic fungi. Its mode of action against dermatophyte fungi in nail keratin is little understood. Our purpose was to determine the bioavailability of terbinafine in a nail fragment model. The effect of terbinafine on adherence and germination of Arthroconidia of Trichophyton mentagrophytes on nail fragments was assessed by gross examination and light and electron microscopy. Preexposure of nail fragments to terbinafine concentrations (0.001 to 10 mg/L) inhibited fungal growth and acted as a barrier to dermatophyte invasion. Damaged Arthroconidia and distorted hyphae on the surface of nail fragments were observed. This in vitro model provides an alternative system for studying the activity of antifungal agents in nail and demonstrates the morphologic changes in dermatophyte fungi after exposure to terbinafine.
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Inhibitory effect of terbinafine on the invasion of nails by Trichophyton mentagrophytes
Journal of the American Academy of Dermatology, 1995Co-Authors: Azer Rashid, E M Scott, Malcolm RichardsonAbstract:Abstract Background: Terbinafine has a broad spectrum of action in vitro and primary fungicidal action against many pahtogenic fungi. Its mode of action against dermatophyte fungi in nail keratin is little understood. Objective: Our purpose was to determine the bioavailability of terbinafine in a nail fragment model. Methods: The effect of terbinafine on adherence and germination of Arthroconidia of Trichophyton mentagrophytes on nail fragments was assessed by gross examination and light and electron microscopy. Results: Preexposure of nail fragments to terbinafine concentrations (0.001 to 10 mg/L) inhibited fungal growth and acted as a barrier to dermatophyte invasion. Damaged Arthroconidia and distorted hyphae on the surface of nail fragments were observed. Conclusion: This in vitro model provides an alternative system for studying the activity of antifungal agents in nail and demonstrates the morphologic changes in dermatophyte fungi after exposure to terbinafine.
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Adherence of Arthroconidia and germlings of anthropophilic and zoophilic varieties of Trichophyton mentagrophytes to human corneocytes as an early event in the pathogenesis of dermatophytosis
Clinical and Experimental Dermatology, 1993Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, A. Rashid, E M Scott, G.s. ShanklandAbstract:The association (adherence) between human corneocytes and Arthroconidia of Trichophyton mentagrophytes strains 121 and 126, and T. interdigitale strain 4 was studied in vitro. Adherence of Arthroconidia to corneocytes from either the palm and sole occurred and increased with time up to 6 h, by which time germination of Arthroconidia had started. Significant differences were seen between the T. mentagrophytes strains and T. interdigitale in their adherence to corneocytes from the palm. When adherence values for plantar corneocytes were compared, significant differences were found between T. mentagrophytes and T. interdigitale. Not all corneocytes from either site had adherent Arthroconidia, although there was a time-dependent increase in the numbers of corneocytes with adherent fungal cells. By scanning and transmission electron microscopy it was seen that there was a loose association between Arthroconidia and corneocytes with no apparent damage to the corneocyte membrane. Adherence of germlings of T. interdigitale to corneocytes from the palm appeared to be mediated by germling outer cell wall fibrils. Hyphal branches and secondary germlings were observed to enhance the attachment of the parent hypha to adjacent corneocytes.
Malcolm Richardson - One of the best experts on this subject based on the ideXlab platform.
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Inhibitory effect of terbinafine on the invasion of nails by Trichophyton mentagrophytes
Journal of the American Academy of Dermatology, 1995Co-Authors: Azer Rashid, E M Scott, Malcolm RichardsonAbstract:Abstract Background: Terbinafine has a broad spectrum of action in vitro and primary fungicidal action against many pahtogenic fungi. Its mode of action against dermatophyte fungi in nail keratin is little understood. Objective: Our purpose was to determine the bioavailability of terbinafine in a nail fragment model. Methods: The effect of terbinafine on adherence and germination of Arthroconidia of Trichophyton mentagrophytes on nail fragments was assessed by gross examination and light and electron microscopy. Results: Preexposure of nail fragments to terbinafine concentrations (0.001 to 10 mg/L) inhibited fungal growth and acted as a barrier to dermatophyte invasion. Damaged Arthroconidia and distorted hyphae on the surface of nail fragments were observed. Conclusion: This in vitro model provides an alternative system for studying the activity of antifungal agents in nail and demonstrates the morphologic changes in dermatophyte fungi after exposure to terbinafine.
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Adherence of Arthroconidia and germlings of anthropophilic and zoophilic varieties of Trichophyton mentagrophytes to human corneocytes as an early event in the pathogenesis of dermatophytosis
Clinical and Experimental Dermatology, 1993Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, A. Rashid, E M Scott, G.s. ShanklandAbstract:The association (adherence) between human corneocytes and Arthroconidia of Trichophyton mentagrophytes strains 121 and 126, and T. interdigitale strain 4 was studied in vitro. Adherence of Arthroconidia to corneocytes from either the palm and sole occurred and increased with time up to 6 h, by which time germination of Arthroconidia had started. Significant differences were seen between the T. mentagrophytes strains and T. interdigitale in their adherence to corneocytes from the palm. When adherence values for plantar corneocytes were compared, significant differences were found between T. mentagrophytes and T. interdigitale. Not all corneocytes from either site had adherent Arthroconidia, although there was a time-dependent increase in the numbers of corneocytes with adherent fungal cells. By scanning and transmission electron microscopy it was seen that there was a loose association between Arthroconidia and corneocytes with no apparent damage to the corneocyte membrane. Adherence of germlings of T. interdigitale to corneocytes from the palm appeared to be mediated by germling outer cell wall fibrils. Hyphal branches and secondary germlings were observed to enhance the attachment of the parent hypha to adjacent corneocytes.
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Effects of terbinafine exposure on the ultrastructure of Trichophyton interdigitale
Medical Mycology, 1993Co-Authors: A. Rashid, E M Scott, Malcolm RichardsonAbstract:The morphological modifications of Arthroconidia and germ tubes of Trichophyton interdigitale exposed to terbinafine (6·25–100 mgl-1) incorporated into stratum corneum were studied by scanning and transmission electron microscopy after 16 h and 20 h. Changes in Arthroconidial morphology were apparent at 16 h. Pores and erosions were present in the cell wall with layers peeling off. The cell membrane was destroyed. Dilated vacuoles and small electron dense areas were evident in the Arthroconidial cytosol. Although germination was not arrested, inhibition of hyphal extension was seen. Germ tubes were susceptible to terbinafine with pores appearing along their length following exposure to the drug. Germ tubes were seen to have collapsed at higher drug concentrations. The study suggests that the outer and inner layers of the Arthroconidial cell wall are the initial targets of terbinafine action followed by alterations to the cytosol and intracellular organelles.
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Germination of Trichophyton mentagrophytes on human stratum corneum in vitro
Medical Mycology, 1992Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, E M Scott, G.s. ShanklandAbstract:The emergence of germ tubes from Arthroconidia of Trichophyton mentagrophytes on stripped sheets of stratum corneum from different body areas was measured. Arthroconidia increased in size and started germination by 4 h at 37°C. Germ tubes originated from a point on the arthroconidium surface mid-way between the points of attachment to adjacent conidia. With further incubation Arthroconidial germination increased and germ tubes extended across the stratum corneum. Histological staining of transverse sections of infected stratum corneum showed hyphae penetrating longitudinally and perpendicularly through the thickness of the stratum corneum. By 7 days' incubation hyphae started to form Arthroconidia thereby completing the vegetative growth cycle of the fungus. Scanning electron microscopy revealed penetration of corneocytes by germ tubes resulting in damage to the corneocyte surface.
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Germination of Trichophyton mentagrophytes on human stratum corneum in vitro.
Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology, 1992Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, E M Scott, G.s. ShanklandAbstract:The emergence of germ tubes from Arthroconidia of Trichophyton mentagrophytes on stripped sheets of stratum corneum from different body areas was measured. Arthroconidia increased in size and started germination by 4 h at 37 degrees C. Germ tubes originated from a point on the arthroconidium surface mid-way between the points of attachment to adjacent conidia. With further incubation Arthroconidial germination increased and germ tubes extended across the stratum corneum. Histological staining of transverse sections of infected stratum corneum showed hyphae penetrating longitudinally and perpendicularly through the thickness of the stratum corneum. By 7 days' incubation hyphae started to form Arthroconidia thereby completing the vegetative growth cycle of the fungus. Scanning electron microscopy revealed penetration of corneocytes by germ tubes resulting in damage to the corneocyte surface.
Bridget M Barker - One of the best experts on this subject based on the ideXlab platform.
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Proper Care and Feeding of Coccidioides: A Laboratorian's Guide to Cultivating the Dimorphic Stages of C. immitis and C. posadasii.
Current protocols in microbiology, 2020Co-Authors: Heather L Mead, Marley C. Caballero Van Dyke, Bridget M BarkerAbstract:Coccidioidomycosis ("Valley fever") is caused by Coccidioides immitis and C. posadasii. These fungi are thermally dimorphic, cycling between mycelia and Arthroconidia in the environment and converting into spherules and endospores within a host. Coccidioides can cause a broad spectrum of disease that can be difficult to treat. There has been a steady increase in disease, with an estimated 350,000 new infections per year in the United States. With the increase in disease and difficulty in treatment, there is an unmet need to increase research in basic biology and identify new treatments, diagnostics, and vaccine candidates. Here, we describe protocols required in any Coccidioides laboratory, such as growing, harvesting, and storing the different stages of this dimorphic fungal pathogen. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Growth and harvest of liquid mycelia cultures for extractions Alternate Protocol 1: Large-volume growth and harvest of liquid mycelia cultures Basic Protocol 2: Mycelial growth on solid medium Alternate Protocol 2: Maintaining mycelial growth on solid medium Basic Protocol 3: Harvesting and quantification of Arthroconidia Alternate Protocol 3: Long-term storage of Arthroconidia Basic Protocol 4: Parasitic spherule growth and harvest Alternate Protocol 4: Obtaining endospores from spherules Basic Protocol 5: Intranasal infection of murine models.
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characterizing in vitro spherule morphogenesis of multiple strains of both species of coccidioides
Medical Mycology, 2019Co-Authors: Marcus De Melo Teixeira, Bridget M Barker, John N Galgiani, Heather L MeadAbstract:The disease San Joaquin Valley Fever (coccidioidomycosis) is caused by the inhalation of Coccidioides Arthroconidia. In vivo, Arthroconidia transform into pathogenic structures termed spherules. Exposure to the host milieu triggers spherule development; however, the molecular mechanisms responsible for the morphological shift are not well characterized. This study compared the morphogenesis of five strains of both species of Coccidioides in two media types to improve the in vitro model of dimorphism that can be easily reproduced, and is amenable to tissue culture. We also sought to establish a modern record of the morphological switch among commonly used lab strains through a detailed account of growth under various conditions. Spherules from five strains were grown in standard (Converse) and experimental media (RPMI-sph). Strain behavior was quantified by median spherule size and spherule concentration, beginning 3 days after inoculation and followed for 10 days of growth. There were significant differences observed among Coccidioides immitis and C. posadasii strains, as well as differences between the in vitro systems.
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PEARLS Dust Devil: The Life and Times of the Fungus That Causes Valley Fever
2016Co-Authors: Eric R. G. Lewis, Bridget M Barker, Jolene R. Bowers, Coccidioides BiologyAbstract:Coccidioides immitis and C. posadasii are pathogenic, dimorphic, soil-dwelling Ascomycetes in the Onygenales order. On average, both Coccidioides species have 29 Mb haploid genomes, containing approximately 10,000 open reading frames (ORFs) on five chromosomes [1]. Cocci-dioides ’ most recent common ancestor underwent gene family expansions for proteases and keratinases, membrane biology genes, and toxin production, all likely utilized for survival in animal tissues and morphological changes; and a loss of genes associated with degradation of plant tissue, such as tannases, cellulases, and cutinases [1]. Coccidioides and other fungi in the family Onygenaceae are able to degrade keratin and may cause skin disease in humans and ani-mals. Both species of Coccidioides are distantly related to other dimorphic human pathogens, such as Histoplasma (Ajellomyces) capusulatum, in the new family Ajellomycetaceae [2]. Both Coccidioides species have similar biology, with a well-characterized asexual life cycle with distinct saprobic and parasitic stages, and only molecular evidence of a sexual cycle (Fig 1). In the saprobic phase, Coccidioides cycles between mycelial and Arthroconidial stages. Arthroconidia are abscised and become airborne by soil disturbance. Inhalation of arthroconi-dia by a potential host can lead to coccidioidomycosis, commonly known as (San Joaquin) Val
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effective disinfectants for coccidioides immitis and c posadasii
Applied Biosafety, 2015Co-Authors: Amy J Vogler, Roxanne Nottingham, Katy L Parise, Paul Keim, Bridget M BarkerAbstract:The lack of published data on effective disinfectants and contact times for use on the fungal pathogens Coccidioides immitis and C. posadasii prompted the authors to investigate the fungicidal activity of three commonly used laboratory disinfectants on Arthroconidia harvested from C. immitis strain 2009. They tested the ability of 10% bleach, 70% ethanol, and Vesphene® IIse to inactivate 107 Arthroconidia in an aqueous suspension within 1, 2, 5, 10, or 20 minutes of contact time. Both 10% bleach and 70% ethanol provided a 7-log10 reduction in Arthroconidia in less than 1 minute, with no growth observed at any of the tested time points. Vesphene® IIse was less effective, providing a 6-log10 reduction in Arthroconidia after 5 minutes, but was unable to completely inactivate all of the Arthroconidia, even after 20 minutes of contact time.
G.s. Shankland - One of the best experts on this subject based on the ideXlab platform.
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in vitro susceptibility of trichophyton mentagrophytes Arthroconidia to clotrimazole and griseofulvin in human corneocyte suspensions
Mycoses, 2009Co-Authors: Salih H.m. Aljabre, G.s. Shankland, E M Scott, M D RichardsonAbstract:The use of corneocytes in suspension as a medium for the study of the effect of antifungal drugs on Trichophyton mentagrophytes Arthroconidia was investigated. In the presence of clotrimazole or griseofulvin, Arthroconidia germination in a suspension of corneocytes and in Sabouraud glucose liquid medium was significantly reduced compared with germination in drug-free media. Where antifungals were added to Arthroconidia after an activation period during which germination occurred, any further germination was inhibited. The data showed that the growth of T. mentagrophytes in the presence of corneocytes offered a simple, rapid, inexpensive and relevant model for the assessment of antifungal activity of compounds for the treatment of dermatophytosis.
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Adherence of Arthroconidia and germlings of anthropophilic and zoophilic varieties of Trichophyton mentagrophytes to human corneocytes as an early event in the pathogenesis of dermatophytosis
Clinical and Experimental Dermatology, 1993Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, A. Rashid, E M Scott, G.s. ShanklandAbstract:The association (adherence) between human corneocytes and Arthroconidia of Trichophyton mentagrophytes strains 121 and 126, and T. interdigitale strain 4 was studied in vitro. Adherence of Arthroconidia to corneocytes from either the palm and sole occurred and increased with time up to 6 h, by which time germination of Arthroconidia had started. Significant differences were seen between the T. mentagrophytes strains and T. interdigitale in their adherence to corneocytes from the palm. When adherence values for plantar corneocytes were compared, significant differences were found between T. mentagrophytes and T. interdigitale. Not all corneocytes from either site had adherent Arthroconidia, although there was a time-dependent increase in the numbers of corneocytes with adherent fungal cells. By scanning and transmission electron microscopy it was seen that there was a loose association between Arthroconidia and corneocytes with no apparent damage to the corneocyte membrane. Adherence of germlings of T. interdigitale to corneocytes from the palm appeared to be mediated by germling outer cell wall fibrils. Hyphal branches and secondary germlings were observed to enhance the attachment of the parent hypha to adjacent corneocytes.
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Germination of Trichophyton mentagrophytes on human stratum corneum in vitro
Medical Mycology, 1992Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, E M Scott, G.s. ShanklandAbstract:The emergence of germ tubes from Arthroconidia of Trichophyton mentagrophytes on stripped sheets of stratum corneum from different body areas was measured. Arthroconidia increased in size and started germination by 4 h at 37°C. Germ tubes originated from a point on the arthroconidium surface mid-way between the points of attachment to adjacent conidia. With further incubation Arthroconidial germination increased and germ tubes extended across the stratum corneum. Histological staining of transverse sections of infected stratum corneum showed hyphae penetrating longitudinally and perpendicularly through the thickness of the stratum corneum. By 7 days' incubation hyphae started to form Arthroconidia thereby completing the vegetative growth cycle of the fungus. Scanning electron microscopy revealed penetration of corneocytes by germ tubes resulting in damage to the corneocyte surface.
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Germination of Trichophyton mentagrophytes on human stratum corneum in vitro.
Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology, 1992Co-Authors: Salih H.m. Aljabre, Malcolm Richardson, E M Scott, G.s. ShanklandAbstract:The emergence of germ tubes from Arthroconidia of Trichophyton mentagrophytes on stripped sheets of stratum corneum from different body areas was measured. Arthroconidia increased in size and started germination by 4 h at 37 degrees C. Germ tubes originated from a point on the arthroconidium surface mid-way between the points of attachment to adjacent conidia. With further incubation Arthroconidial germination increased and germ tubes extended across the stratum corneum. Histological staining of transverse sections of infected stratum corneum showed hyphae penetrating longitudinally and perpendicularly through the thickness of the stratum corneum. By 7 days' incubation hyphae started to form Arthroconidia thereby completing the vegetative growth cycle of the fungus. Scanning electron microscopy revealed penetration of corneocytes by germ tubes resulting in damage to the corneocyte surface.
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Dormancy of Trichophyton mentagrophytes Arthroconidia.
Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology, 1992Co-Authors: Salih H.m. Aljabre, E M Scott, Malcolm D. Richardson, G.s. ShanklandAbstract:Arthroconidia of Trichophyton mentagrophytes demonstrated an exogenous type of dormancy under environmental conditions unsuitable for germination, provided that these conditions were not lethal. Germination did not occur at 4 degrees C unless Arthroconidia were shifted up to 37 degrees C. At 45 degrees C germination was inhibited even when Arthroconidia were shifted down to 37 degrees C. Germination was dependent on humidified incubation conditions. The presence of human corneocytes enhanced the germination of Arthroconidia in the presence of moisture.
Barry Scott - One of the best experts on this subject based on the ideXlab platform.
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Formation of Arthroconidia during regeneration and selection of transformed Epichloë festucae protoplasts.
Fungal biology, 2014Co-Authors: Gemma Maree Cartwright, Aiko Tanaka, Carla Jane Eaton, Barry ScottAbstract:Transformation is an essential tool for modern fungal research and has played a fundamental role in gaining insight into gene function. Polyethylene glycol (PEG)-mediated transformation of protoplasts is the most commonly used method for genetic transformation of filamentous fungi. Selectable marker genes, that confer resistance to antibiotics, are generally incorporated with the DNA of interest, allowing transformed cells to grow through the antibiotic overlay. Colonies arising from transformed fungal cells are sub-cultured and further analysed. However, the morphological state of the fungal cells during the transformation procedure has been largely overlooked. We investigated the morphological appearance of transformed fungal cells prior to their emergence through the antibiotic overlay. Hyphae appeared to segment and bulge, reminiscent of Arthroconidia, an asexual spore typically produced by segmentation of pre-existing hyphae. Selective expression of eGFP under the control of a spore specific promoter, PcatA, in these cells confirmed their spore-like nature. Reducing the oxygen availability to surface-grown cultures partially recapitulated this morphological form. A GFP fusion to the cell wall integrity MAP kinase MpkA localised to the Arthroconidia nuclei suggesting the cell wall integrity signalling pathway modulates cell wall stress responses in Arthroconidia. This dramatic morphological change was also observed in transformed Magnaporthe oryzae cells suggesting it may be a more general phenomenon in filamentous fungi. Given the changes in cellular structure and spore-like appearance, these observations may have technical implications for deleting genes involved in these processes in Epichloë festucae and, more broadly, a range of fungal species.
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Formation of Arthroconidia during regeneration and selection of transformed Epichloë festucae protoplasts
Fungal Biology, 2014Co-Authors: Gemma Maree Cartwright, Aiko Tanaka, Carla Jane Eaton, Barry ScottAbstract:Transformation is an essential tool for modern fungal research and has played a fundamental role in gaining insight into gene function. Polyethylene glycol (PEG)-mediated transformation of protoplasts is the most commonly used method for genetic transformation of filamentous fungi. Selectable marker genes, that confer resistance to antibiotics, are generally incorporated with the DNA of interest, allowing transformed cells to grow through the antibiotic overlay. Colonies arising from transformed fungal cells are sub-cultured and further analysed. However, the morphological state of the fungal cells during the transformation procedure has been largely overlooked. We investigated the morphological appearance of transformed fungal cells prior to their emergence through the antibiotic overlay. Hyphae appeared to segment and bulge, reminiscent of Arthroconidia, an asexual spore typically produced by segmentation of pre-existing hyphae. Selective expression of eGFP under the control of a spore specific promoter, PcatA, in these cells confirmed their spore-like nature. Reducing the oxygen availability to surface-grown cultures partially recapitulated this morphological form. A GFP fusion to the cell wall integrity MAP kinase MpkA localised to the Arthroconidia nuclei suggesting the cell wall integrity signalling pathway modulates cell wall stress responses in Arthroconidia. This dramatic morphological change was also observed in transformed Magnaporthe oryzae cells suggesting it may be a more general phenomenon in filamentous fungi. Given the changes in cellular structure and spore-like appearance, these observations may have technical implications for deleting genes involved in these processes in Epichloe festucae and, more broadly, a range of fungal species.
