The diversity of root fungi associated with Erica species occurring in the Albany Centre of Endemism
- Authors: Bizabani, Christine
- Date: 2015
- Subjects: Ericaceae , Ericas , Roots (Botany) -- Diseases and pests , Mycorrhizal fungi , Polymerase chain reaction , Fungi -- Classification
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4160 , http://hdl.handle.net/10962/d1018575
- Description: South Africa has the highest species diversity of ericaceous plants belonging to the Erica genus. There are over 850 identified species in the Cape Floral Region. The Albany Centre of Endemism (ACOE) is located within this region and is a hotspot of diversity consisting of various plant genera. The success of Erica plants is ubiquitously attributed to mycorrhizal relationships they engage in with a diverse group of fungi. This symbiosis is known as the ericoid mycorrhizal (ERM) association. The overall aim of this study was to establish the diversity of root fungi associated with Erica plants using morphological, molecular and 454 pyrosequencing techniques. Six Erica species were identified using leaf and flower morphology according to taxonomic keys. The identified plants were Erica cerinthoides, Erica demissa, Erica chamissonis, Erica glumiflora, Erica caffra and Erica nemorosa. Roots from sampled plants were stained and examined microscopically to determine their mycorrhizal status. Ericoid mycorrhizal associations together with dark septate endophyte (DSE) structures and hyphae that did not form any specific structure were observed in all the roots. In addition arbuscular mycorrhizal (AM) structures in the form of vesicles were detected in E. glumiflora and E. cerinthoides. In order to identify the culturable fungi associated with the respective hosts, sterilised roots were placed on various culture media for cultivation. Thereafter isolated fungi were morphologically classified into 67 morphotypes. These were mostly sterile and darkly pigmented. Non-sporulating mycelia of variable colouration such as white, cream-yellowish, beige, green and brown were also observed. Further identification was carried out using molecular techniques. DNA was extracted separately from pure cultures and amplified using ITS1 and ITS4 primers in a polymerase chain reaction (PCR). Thereafter sequencing and Basic Local Alignment Search Tool (BLAST) were used to identify the isolates to generic level. The fungi were taxonomically classified into 54 operational taxonomic units and 94 percent were Ascomycetes and Helotiales was the dominant order. Unclassified Helotiales with affinities to fungi currently identified as Epacrid root fungus was common in all hosts. Other isolates that were identified included Oidiodendron, Meliniomyces, Phialocephala, Cadophora, Lachnum, Leohumicola Cryptosporiopsis, Chaetomium, Acremonium and Epicoccum species. Basidiomycetes were represented by two OTUs belonging to the genus Mycena. Four OTUs comprised fungi that had no significant alignments in the reference databases. Direct root DNA extraction together with 454 pyrosequencing was used to detect the diversity of culturable and unculturable fungi associated with the identified hosts. The ITS2 region was targeted for sequencing. Although Ascomycetes remained the dominant phyla, Basidiomycetes were also detected in all host plants. Glomeromycota was present in E. caffra and E. cerinthoides. Helotiales was dominant in all Erica plants with the exception of E. cerinthoides and E. chamissonis which were dominated by the order Chaetothyriales. The OTUs identified to genus level included Epacris pulchella root fungus, Oidiodendron cf. maius, Acremonium implicatum, Leohumicola, Lachnum, Capronia and Mycena species. Culture-based techniques and pyrosequencing detected similar fungal composition comprising Ascomycetes, while, pyrosequencing was able to detect Glomeromycetes and Basidiomycetes.
- Full Text:
- Date Issued: 2015
The diversity of root fungi associated with Erica species occurring in the Albany Centre of Endemism
- Authors: Bizabani, Christine
- Date: 2015
- Subjects: Ericaceae , Ericas , Roots (Botany) -- Diseases and pests , Mycorrhizal fungi , Polymerase chain reaction , Fungi -- Classification
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4160 , http://hdl.handle.net/10962/d1018575
- Description: South Africa has the highest species diversity of ericaceous plants belonging to the Erica genus. There are over 850 identified species in the Cape Floral Region. The Albany Centre of Endemism (ACOE) is located within this region and is a hotspot of diversity consisting of various plant genera. The success of Erica plants is ubiquitously attributed to mycorrhizal relationships they engage in with a diverse group of fungi. This symbiosis is known as the ericoid mycorrhizal (ERM) association. The overall aim of this study was to establish the diversity of root fungi associated with Erica plants using morphological, molecular and 454 pyrosequencing techniques. Six Erica species were identified using leaf and flower morphology according to taxonomic keys. The identified plants were Erica cerinthoides, Erica demissa, Erica chamissonis, Erica glumiflora, Erica caffra and Erica nemorosa. Roots from sampled plants were stained and examined microscopically to determine their mycorrhizal status. Ericoid mycorrhizal associations together with dark septate endophyte (DSE) structures and hyphae that did not form any specific structure were observed in all the roots. In addition arbuscular mycorrhizal (AM) structures in the form of vesicles were detected in E. glumiflora and E. cerinthoides. In order to identify the culturable fungi associated with the respective hosts, sterilised roots were placed on various culture media for cultivation. Thereafter isolated fungi were morphologically classified into 67 morphotypes. These were mostly sterile and darkly pigmented. Non-sporulating mycelia of variable colouration such as white, cream-yellowish, beige, green and brown were also observed. Further identification was carried out using molecular techniques. DNA was extracted separately from pure cultures and amplified using ITS1 and ITS4 primers in a polymerase chain reaction (PCR). Thereafter sequencing and Basic Local Alignment Search Tool (BLAST) were used to identify the isolates to generic level. The fungi were taxonomically classified into 54 operational taxonomic units and 94 percent were Ascomycetes and Helotiales was the dominant order. Unclassified Helotiales with affinities to fungi currently identified as Epacrid root fungus was common in all hosts. Other isolates that were identified included Oidiodendron, Meliniomyces, Phialocephala, Cadophora, Lachnum, Leohumicola Cryptosporiopsis, Chaetomium, Acremonium and Epicoccum species. Basidiomycetes were represented by two OTUs belonging to the genus Mycena. Four OTUs comprised fungi that had no significant alignments in the reference databases. Direct root DNA extraction together with 454 pyrosequencing was used to detect the diversity of culturable and unculturable fungi associated with the identified hosts. The ITS2 region was targeted for sequencing. Although Ascomycetes remained the dominant phyla, Basidiomycetes were also detected in all host plants. Glomeromycota was present in E. caffra and E. cerinthoides. Helotiales was dominant in all Erica plants with the exception of E. cerinthoides and E. chamissonis which were dominated by the order Chaetothyriales. The OTUs identified to genus level included Epacris pulchella root fungus, Oidiodendron cf. maius, Acremonium implicatum, Leohumicola, Lachnum, Capronia and Mycena species. Culture-based techniques and pyrosequencing detected similar fungal composition comprising Ascomycetes, while, pyrosequencing was able to detect Glomeromycetes and Basidiomycetes.
- Full Text:
- Date Issued: 2015
Ericoid mycorrhizal fungi and potential for inoculation of commercial berry species (Vaccinium corymbosium L.)
- Authors: Bizabani, Christine
- Date: 2011
- Subjects: Ericaceae , Mycorrhizas , Fynbos
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4136 , http://hdl.handle.net/10962/d1016127
- Description: Ericaceous plants are the richest growth form of the fynbos vegetation of South Africa. The fynbos is characterized by highly leached acidic soils, low mineral nutrients and climatically it is a winter rainfall and dry summer region. Ericoid mycorrhizal fungi associate with Erica species enhancing their ability to access essential nutrients for survival under unfavourable growth conditions. The aim of this study was to select local Ericaceae plant species and to isolate, identify and characterize the ericoid endophytes and assess these isolates as potential inocula for commercial berry species. Two ericaceous plants Erica cerinthoides L. and Erica demmissa Klotzsch ex Benth. were identified from the Mountain Drive area of Grahamstown, Eastern Cape. Root staining was used to confirm the mycorrhizal status of both plants. Hyphal coils typical of ericoid association were observed within the epidermal cells of the hair roots under a light microscope. The endophytes were successfully isolated in pure culture on 2% malt extract agar (MEA) and modified Fontana medium. Cultural morphology and microscopy were used for initial identification. Two slow growing isolates were selected. These isolates were further subjected to molecular identification; extracted DNA was amplified using ITS1 and ITS4 fungal primers. The rDNA gene internal transcriber spacer (ITS) was then sequenced and analyzed by comparison to sequences in the GenBank. On the basis of percentage sequence identity Lachnum Retz. species and Cadophora Lagerb. & Melin species were identified as the ericoid endophytes of E. cerinthoides and E. demmissa respectively. The optimum growth parameters of the fungal isolates were determined in 2% MEA incubated at varying temperatures and pH. It was established that both species had optimum growth at 27⁰C and pH 5. The Ericaceae species are sometimes found in metal contaminated sites were ericoid fungi have been proved to alleviate toxicity of their host. The fungal isolates were grown in increasing concentration of Cu²⁺ and Zn²⁺ in 2% MEA. The growth of Lachnum species decreased with increasing Zn²⁺ ions above 2.7 mM while Cadophora species showed a change in morphology and also decreased in growth with increased ion concentration. However there were no significant differences recorded in the growth of Cadophora and Lachnum species on increasing Cu²⁺ concentration. Lachnum and Cadophora isolates were formulated into a semi solid inoculum and inoculated onto micropropagated Vaccinni corymbosum L. plantlets of 5 different varieties. Colonization was low for all varieties, Elliott and Brightwell varieties recorded the highest colonization of 35% and 31% respectively. Lachnum species infected roots showed potential ericoid structures while the Cadophora inoculated plantlets had hyphal coils within the cortical cells typical of ericoid mycorrhizas. Inoculation significantly enhanced the shoot growth of Brightwell and Elliott varieties. The Chandler variety inoculated with Lachnum species showed improved shoot dry weight. The Bluecrop and Elliott varieties inoculated with Cadophora and Lachnum accumulated more root biomass. All inoculated Bluecrop plantlets had an improved canopy growth index. Brightwell plantlets inoculated with Lachnum species also had an enhanced canopy growth index. The growth responses were variable within varieties and between varieties. Treatments with the Cadophora and Lachnum have shown potential in the promotion of growth of the Blueberry species. The findings indicate the need to conduct trials under conditions which simulate the commercial growth conditions so as explore the optimum potential of the isolates.
- Full Text:
- Date Issued: 2011
- Authors: Bizabani, Christine
- Date: 2011
- Subjects: Ericaceae , Mycorrhizas , Fynbos
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4136 , http://hdl.handle.net/10962/d1016127
- Description: Ericaceous plants are the richest growth form of the fynbos vegetation of South Africa. The fynbos is characterized by highly leached acidic soils, low mineral nutrients and climatically it is a winter rainfall and dry summer region. Ericoid mycorrhizal fungi associate with Erica species enhancing their ability to access essential nutrients for survival under unfavourable growth conditions. The aim of this study was to select local Ericaceae plant species and to isolate, identify and characterize the ericoid endophytes and assess these isolates as potential inocula for commercial berry species. Two ericaceous plants Erica cerinthoides L. and Erica demmissa Klotzsch ex Benth. were identified from the Mountain Drive area of Grahamstown, Eastern Cape. Root staining was used to confirm the mycorrhizal status of both plants. Hyphal coils typical of ericoid association were observed within the epidermal cells of the hair roots under a light microscope. The endophytes were successfully isolated in pure culture on 2% malt extract agar (MEA) and modified Fontana medium. Cultural morphology and microscopy were used for initial identification. Two slow growing isolates were selected. These isolates were further subjected to molecular identification; extracted DNA was amplified using ITS1 and ITS4 fungal primers. The rDNA gene internal transcriber spacer (ITS) was then sequenced and analyzed by comparison to sequences in the GenBank. On the basis of percentage sequence identity Lachnum Retz. species and Cadophora Lagerb. & Melin species were identified as the ericoid endophytes of E. cerinthoides and E. demmissa respectively. The optimum growth parameters of the fungal isolates were determined in 2% MEA incubated at varying temperatures and pH. It was established that both species had optimum growth at 27⁰C and pH 5. The Ericaceae species are sometimes found in metal contaminated sites were ericoid fungi have been proved to alleviate toxicity of their host. The fungal isolates were grown in increasing concentration of Cu²⁺ and Zn²⁺ in 2% MEA. The growth of Lachnum species decreased with increasing Zn²⁺ ions above 2.7 mM while Cadophora species showed a change in morphology and also decreased in growth with increased ion concentration. However there were no significant differences recorded in the growth of Cadophora and Lachnum species on increasing Cu²⁺ concentration. Lachnum and Cadophora isolates were formulated into a semi solid inoculum and inoculated onto micropropagated Vaccinni corymbosum L. plantlets of 5 different varieties. Colonization was low for all varieties, Elliott and Brightwell varieties recorded the highest colonization of 35% and 31% respectively. Lachnum species infected roots showed potential ericoid structures while the Cadophora inoculated plantlets had hyphal coils within the cortical cells typical of ericoid mycorrhizas. Inoculation significantly enhanced the shoot growth of Brightwell and Elliott varieties. The Chandler variety inoculated with Lachnum species showed improved shoot dry weight. The Bluecrop and Elliott varieties inoculated with Cadophora and Lachnum accumulated more root biomass. All inoculated Bluecrop plantlets had an improved canopy growth index. Brightwell plantlets inoculated with Lachnum species also had an enhanced canopy growth index. The growth responses were variable within varieties and between varieties. Treatments with the Cadophora and Lachnum have shown potential in the promotion of growth of the Blueberry species. The findings indicate the need to conduct trials under conditions which simulate the commercial growth conditions so as explore the optimum potential of the isolates.
- Full Text:
- Date Issued: 2011
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