Application of plant growth promoting substances and arbuscular mycorrhizal fungi for phytostabilisation of mine tailings

Rossouw, Marthinus Jacob (2016-03)

Thesis (MSc)--Stellenbosch University, 2016.

Thesis

ENGLISH ABSTRACT: This study focused on investigating methods of phytostabilisation of mine tailings operated by Palabora Copper in South Africa. Capping material and mine tailing at various sites of the mine were collected and used in pot trials to investigate the effect of a number of plant growth promoting substances (PGPS) on several of grass species currently used in effort to stabilise the areas in question. Lumichrome, strigolactones (GR24), flavonoids (CropbioLife™), smoke-water (karrikins) and arbuscular mycorrhizal fungi (Mycoroot™) were used as PGPS to investigate growth-promoting effects on i) Anthephora pubescens, Cenchrus ciliaris, Chloris gayana, Cynodon dactylon, and Panicum maximum which are species currently used by Palabora Copper for rehabilitation of mine tailings, and ii) Additional grass species theoretically suited to surviving the environment. Treatments were applied on 2-week old transplanted grass seedlings in pot trials containing mine capping material as the substrate, to infer treatment responses. Trypan-Blue staining procedures were used to ascertain which grass species formed symbiotic relationship with arbuscular mycorrhizal fungi (AMF), which would potentially aid in their survival in deleterious areas. Germination rates were measured to determine the fastest germinating species of the selected grasses with Eragrostis teff and Melinis repens germinating the quickest in the mine capping material. Capping material and mine tailing samples were collected at sites under revegetation by Palabora Copper. This included samples of the rhizosphere of locally abundant plants at two sites: a recently (two years) capped mine tailing, and a rock dump site (capped 10-12 years previously). Five rhizosphere samples were collected from individuals of Cenchrus ciliaris, Enneapogon cenchroides, and Tephorisia polystachya (a locally abundant forb species) at site 1 and Cenchrus ciliaris, Stipagrostis hirtigluma, Tephrosia polystachya, and Pennisetum setaceum at site 2. At both sites the soil of open areas devoid of plants was also sampled. Metagenomic DNA was extracted from the collected samples, often following enrichment techniques. Dilution series spread plates to determine culturable bacteria present in the tailing samples were also utilised. Polymerase Chain Reactions were implemented to produce amplicons of conserved regions within AMF and bacteria present in the mine tailing site. The predominant genera of bacteria detected in the collected tailing samples belonged to Bacillus. However, due to the use of enrichment techniques it was not possible to comment on the relative abundance of different bacteria in the environment where the samples were collected. Due to the small-scale ex situ nature of the experiments the results gained from the PGPS treatment trials and microbial DNA isolation are not necessarily representative of the ecological environment present in situ. However, PGPS treatment of the selected grasses did not elicit any clear beneficial responses in the measured growth parameters, making application thereof of limited benefit for phytostabilisation purposes. Trypan staining revealed most of the grass species are capable of forming symbiotic relationships with mycorrhizal fungi, with trials indicating that AMF might benefit plants present in the mine tailings.

AFRIKAANSE OPSOMMING: Hierdie studie het gefokus op die ondersoek metodes van phytostabilisasie van mynuitskot wat bedryf word deur Palabora Copper in Suid-Afrika. Bedekkingsmateriaal en mynuitskot op verskillende terreine van die myn is versamel en gebruik in pot proewe om die rol van verbindings wat plantgroei bevorder (PGBV) op sewe gras spesies wat tans gebruik word in die stabilisering van die spesifieke gebiede te ondersoek. Lumichrome, strigolaktone (GR24), flavonoïede (CropbioLife™), rook-water (karrikins) en arbuskulêre mikorisaie swamme (Mycoroot™) is die PGBV wat gebruik was plantgroei bevorder te ondersoek in i) Anthephora pubescens, Cenchrus ciliaris, Chloris gayana, Cynodon dactylon, en Panicum maximum, wat tans gebruik word deur Palabora Copper rehabilitasie pogings, en ii) Addisionele grasspesies wat teoreties bepaal is as mees geskik om te oorleef in die omgewing was gebruik in die pot proewe. Behandelings (PGBV) is toegepas op 2 week oue oorgeplante gras saailinge in pot proewe met myn bedekkingsmateriaal, om die reaksie op die behandelinge af te lei. Trypan-Blue bevlekkking prosedures was geïmplementeer om vas te stel watter van die grasspesies simbiotiese verhoudings met arbuskulêre mikorisaie swamme (AMS) vorm en potensieel kan help in hul oorlewing in skadelike gebiede. Groeikoerse vir die grasse wat gebruik is gemeet om te bepaal watter van die gekose grasspesies groei die vinnigste. Beide Eragrostis teff en Melinis repens het die vinnigste ontkiem in die myn bedekkingsmateriaal. Bedekkingsmateriaal en mynuitskot monsters is versamel by terreine wat gebruik word deur Palabora Copper. Dit sluit monsters in van die risosfeer van geselekteerde plante op twee plekke: 'n onlangs (twee jaar) bedekte mynuitskot, en 'n rots stortingsterrein wat voorheen bedek is (10-12 jaar gelede). Vyf rhizosfeer monsters van die volgende is ingesamel: Cenchrus ciliaris, Enneapogon cenchroides, Tephrosia polystachya, (plaaslike volop forb spesies) by terrein 1 en Cenchrus ciliaris, Stipagrostis hirtigluma, Tephrosia polystachya, en Pennisetum setaceum monsters by terrein 2. Die monsters van oop gebiede sonder plante is ook gemonster in beide terreine. Metagenomiese DNS is onttrek uit die versamelde monsters, hoewel verrykingstegnieke toegepas. Verdunnings reeks verspreiding plate is gebruik om kweekbare bakterieë wat teenwoordig is in die uitskot monsters te identifiseer. Polimerase ketting reaksies was geïmplementeer om amplikons te produseer van konserveerde streke in genomiese DNS van AMS en bakterieë wat teenwoordig in die mynskot. Die oorheersende genera van bakterieë bespeur in die versamelde uitskot monsters behoort aan Bacillus¸ maar as gevolg van die verryking tegnieke wat gebruik is kan dit egter nie beslis verklaar word dat hierdie bakterie die volopste in die omgewing is vanwaar die monsters versamel is nie. As gevolg van die klein skaalse ex situ aard van die eksperimente, is die ooreenstemmende resultate wat verkry is uit hulle behandeling proewe en mikrobiese DNS isolasie nie heeltemal verteenwoordigend van die aard van die ekologiese omgewing teenwoordig in situ. Nietemin, PGBV behandeling op die gekose grasse het geen duidelike voordeel ontlok in gemeet groei parameters gebruik word vir doeleindes phytostabilisation. Trypan bevlekkkings prosedures het aangedui dat meeste van die grasspesies in staat is om simbiotiese verhoudings met AMS te vorm, terwyl proewe aangedui het dat AMS plante kan baat vat teenwoordig is in die mynuitskot.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/98721
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