Psychrotolerant mucoralean fungi present in pristine mountain fynbos soil and vineyard soil from the Stellenbosch region

Samson, Heidi E. (Heidi Estrelita) (2001-03)

Thesis (MSc)--University of Stellenbosch, 2001.

Thesis

ENGLISH ABSTRACT: Mucoralean fungi are mostly saprotrophs that are frequently encountered in soil habitats. Using an isolation temperature of circa 25°C, other workers obtained these fungi from a wide diversity of geographical areas in southern Africa. However, it is known that psychrotolerant mucoralean fungi, able to grow at 25°C as well as at 5°C, occur in pristine Alti Mountain Grassland. Nothing is known about the diversity of these psychrotolerant soil fungi in other vegetation types of South Africa. Consequently, in this study, the psychrotolerant fungal taxa and numbers in soil from a vineyard and from pristine Mountain Fynbos were determined using an incubation temperature of 4°C and a complex isolation medium. The latter contained agar, malt extract, peptone, yeast extract, penicillin and streptomycin sulphate. Soil samples were analysed in late summer, autumn and mid-winter. It was found that, for the samples taken in late summer and autumn, the diversity of mucoralean species in the soil differed between fynbos and vineyard. In winter however, no significant difference was detected between the Shannon's diversity indices of mucoralean species in the soil samples taken from the two habitats. It was found that in both soil types, the percentage mucoralean fungi on the plates increased from summer to winter. In addition, the numbers of detectable Morlierella subgenus Morlierella on the plates were higher in winter than in late summer. The diversity of mucoralean species obtained during winter in fynbos and vineyard soil was significantly less than the diversity of these species in Alti Mountain Grassland soil. To determine if the Morlierella subgenus Morlierella isolates from the fynbos and vineyard soil, and those obtained from Alti Mountain Grassland, differ in the ability to grow at low temperatures, the radial growth rate on malt extract agar at 4°C and BOC was determined for each isolate. The results indicate that not only did seasonal changes occur in the taxa within Morlierella subgenus Morlierella, but that the isolates dominating the soil in different seasons also differed in the ability to grow at low temperatures. The percentage of isolates that had reached greater colony diameters after B days of incubation at 4°C, was higher for the isolates obtained in the cold wet month of July than for those obtained in the warmer dryer month of February. Similar results were obtained with the radial growth experiments conducted at BOC. The Morlierella subgenus Morlierella isolates obtained in winter from fynbos and vineyard soil showed less variation in low temperature growth rate than the isolates of this taxon obtained in winter from Alti Mountain Grassland soil during a previous study. This variation corresponds to the greater number (20) of Morlierella subgenus Morlierella species found in the grassland soil. Altogether only seven species of this subgenus was detected during the present study in the fynbos and vineyard soil samples. It was speculated that this difference in diversity between the fynbos and vineyard isolates, and the grassland isolates obtained in a previous study, might have been as a result of differences in the habitat or the enumeration methods used. The phylogenetic relationship between different psychrotolerant isolates of Morlierella subgenus Morlierella originating from the soil of the fynbos, vineyard and Alti Mountain Grassland, was subsequently determine through comparison of ITS regions, within ribosomal RNA repeats. Consequently, 45 psychrotolerant Morlierella subgenus Morlierella isolates originating from the three soil habitats was compared on the basis ITS 1 nucleotide sequence composition and radial growth rate at 4°C. Phylogenetic analyses showed that the isolates could be grouped into two clusters correlating with the ability to grow at low temperatures. Each cluster was further subdivided into two subgroups. It was found that except for one subgroup and the reference strain occurring in another subgroup, all the subgroups contain isolates originating from a single soil habitat. Therefore, the ITS 1 sequence of these fungi seems to indicate the original habitat and ability to grow at low temperatures. This correlation of the ITS sequence with the ecological habitat of a fungus has also been observed by other workers for other fungal groups.

AFRIKAANSE OPSOMMING: Mucoraliese fungi is meestal saprotrofe wat dikwels in grondhabitatte aangetref word. Deur gebruik te maak van 'n isolasietemperatuur van circa 25°C, het ander werkers dié fungi van 'n wye verskeidenheid geografiese gebiede in suidelike Afrika verkry. Dit is egter bekend dat die psigrotolerante mucoraliese fungi, wat in staat is om by 2SoC en ook by SaC te groei, in ongeskonde Alti Berg-Grasland voorkom. Niks is egter bekend oor die diversiteit van dié psigrotolerante grondfungi in ander veldtipes van suidelike Afrika nie. Die psigrotolerante fungustaksa en -getalle in grond van 'n wingerd en van ongeskonde Berg Fynbos is gevolglik in dié studie bepaal deur gebruik te maak van 'n inkubasietemperatuur van 4"C en 'n komplekse isolasiemedium. Laasgenoemde het agar, moutekstrak, peptoon, gisekstrak, penisillien en streptomisiensulfaat bevat. Grondmonsters is in die laatsomer, herfs en midwinter geanaliseer. Daar is 'n verskil gevind tussen die diversiteit van die mucoraliese spesies in die grond van fynbos en dié van wingerd in die monsters wat in die laatsomer en midwinter geneem is. In die winter is daar egter geen beduidende verskil gevind tussen die Shannon diversiteitsindekse van mucoraliese spesies in die grondmonsters wat uit die twee habitatte getrek is nie. In albei grondtipes is daar gevind dat die persentasie mucoraliese fungi op die plate toegeneem het van somer tot winter. Daarby was die aantal waarneembare Morlierella subgenus Morlierella op die plate meer in die winter as in die laatsomer. Die diversiteit van mucoraliese spesies wat in die winter uit fynbos- en wingerdgrond verkry is, was beduidend minder as die diversiteit van dié spesies in Alti Berg-Grasland grond. Om te bepaal of die Morlierella subgenus Morlierella isolate van die fynbos- en wingerdgrond en dié van Alti Berg-Grasland van mekaar verskil ten opsigte van hul vermoë om by lae temperature te groei, is die radiale groeitempo op moutekstrak by 4"C en aoc vir elke isolaat bepaal. Die resultate dui aan dat daar nie alleen seisoenale veranderinge in die taksa binne Morlierella subgenus Morlierella voorkom nie, maar dat die isolate wat tydens verskillende seisoene uit die grond verkry is, ook ten opsigte van hul groeivermoë by lae temperature van mekaar verskil. Die persentasie isolate wat groter kolonie diameters bereik het ná B dae inkubasie by 4°C, was hoër vir die isolate van die koue, nat Juliemaand as vir dié wat in die warmer en droër Februariemaand verkry is. Soortgelyke resultate is verkry met radiale groei-eksperimente wat by BOC gedoen is. Die MortierelIa subgenus MortierelIa isolate wat in die winter uit fynbos- en wingerdgrond verkry is, het In kleiner variasie in hul groeitempo by lae temperature getoon as die isolate in dié takson wat tydens 'n vorige studie in die winter uit Alti Berg-Grasland grond verkry is. Dié variasie stem ooreen met die groter aantal (20) MortierelIa subgenus MortierelIa spesies wat in die graslandgrond gevind is. Slegs sewe spesies van dié subgenus is gedurende die huidige studie in die fynbos- en wingerdgrondmonsters waargeneem. Daar is gespekuleer dat dié verskil in diversiteit tussen die fynbos- en wingerdisolate en die graslandisolate van die vorige studie die gevolg mag wees van verskille tussen die habitat of die enumerasiemetodes wat gebruik is. Die filogenetiese verwantskap tussen verskillende psigrotolerante isolate van MortierelIa subgenus MortierelIa uit die grond van die fynbos, wingerd en Alti Berg-Grasland, is vervolgens bepaal deur 'n vergelyking van interne getranskribeerde spasieerder (ITS) areas, binne ribosomale RNS herhalings. Daar is gevolglik 45 psigrotolerante MortierelIa subgenus MortierelIa isolate uit die drie grondhabitatte met mekaar vergelyk op grond van die basis ITS 1 nukleotied opeenvolgingsamestelling en radiale groeitempo by 4°C. Filogenetiese analises het die isolate in twee groepe verdeel op grond van hul vermoë om by lae temperature te groei. Elke groep is verder in twee subgroepe verdeel. Daar is gevind dat behalwe vir een subgroep en die verwysingstam wat in 'n ander subgroep voorgekom het, elkeen van die subgroepe bestaan het uit isolate wat van 'n enkele grond habitat verkry is. Dit wil dus voorkom of die ITS 1 opeenvolging van dié fungi 'n aanduiding gee van die oorspronklike habitat en die vermoë om by lae temperature te groei. Dié korrelasie tussen die ITS opeenvolging en die ekologiese habitat van 'n fungus is ook deur ander werkers vir ander fungusgroepe waargeneem.

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