Determination of the bacterial diversity of a natural freshwater wetland impacted by acid mine drainage.
Thesis (MSc)--Stellenbosch University, 2015.
ENGLISH ABSTRACT: Freshwater resources in semi-arid countries, such as South Africa, are under constant threat from pollution. One of the major pollutants is acid mine drainage, which not only lowers the pH of the water, but also increases sulphate and metal concentrations. Primary producers, such as bacteria and algae, are the first organisms to respond to stressors such as reduced pH and elevated sulphate and metal concentrations. A natural freshwater wetland, the Zaalklapspruit wetland in Mpumalanga, impacted by acid mine drainage and industrial effluent was studied to determine the change in algal and bacterial populations. Five study sites were identified including a reference site and four sites displaying various degrees of degradation. Physical and chemical parameters were measured at each site. Algae were identified microscopically and chlorophyll-a concentrations were measured. The algal species present at the five study sites were species previously associated with the conditions present at the various sites. Gyrosigma rautenbachiae proved to be an ideal bioindicator for industrial pollution. The diatom species Synedra ulna, Nitzschia spp. and Cymbella spp. were found at the acidic sites. The filamentous green algae Microspora quadrata and Klebsormidium acidophilum were abundant at the sites the most impacted by AMD. Metal tolerant K. rivulare were also identified in this study. The cyanobacteria Oscillatoria tenius and Glaucospira sp. were associated with enriched conditions. The bacterial populations were sampled from both the water column and sediment and subjected to next generation sequencing for identification. The phyla that were highly represented throughout all the samples were the alpha-, beta- and gamma-Proteobacteria, Bacteriodetes and unclassified species. The Bacteriodetes phylum was observed at significantly higher numbers at sites 1, 2, 3 & 5 in the March 2013 water samples and sites 1 & 4 in the March 2013 sediment samples. Firmicutes had significantly higher numbers at sites 2 (January 2013), 3 (March 2013) & 4 (January 2013) in the water samples. Both water and sediment samples of sites 2 (March 2013) & 4 (January 2013) had significantly higher numbers of Actinobacteria. The Chloroflexi phylum had significantly higher numbers in the site 4 & 5 (January 2013) water samples and site 5 (January 2013) of the sediment samples. Acidobacteria were only detected in significantly higher numbers in the January 2013 sediment samples of sites 1 & 5. This study was the first to assess the total bacterial diversity in a natural, acid mine drainage impacted wetland in South Africa and also the first to identify sequences from the genus Marinobacterium. The wetland ecosystem health was also determined using a rapid bioassessment tool and a proposed bacterial bioindicator. The bioassessment tool scored the reference site as mostly natural, two sites as severely modified and the last two as modified. The proposed bacterial bioindicator was simplistic in use and reflected the stability of the populations at the five sites accordingly. Lastly, the bacterial bioindicator was incorporated into the established bioassessment tool and was found to correspond with the latter’s results.
AFRIKAANSE OPSOMMING: Varswater bronne in semi-droë lande soos Suid-Afrika is konstant onder druk van besoedeling. Een van die groot besoedeling bronne is suur myn-water, wat beide die pH van die water verlaag en die sulfaat en metal konsentrasies verhoog. Primêre produseerders soos bakterie en alge is die eerste organismes geaffekteer deur stresfaktore soos die bogenoemde. ‘n Natuurlike varswater vlei, die Zaalklapspruit vlei in Mpumalanga, besoedeld deur suur myn-waater en industriële uitvloei was bestudeer om die veranderinge in die alge en bakteriese populasies waar te neem. Vyf studie areas was geïdentifiseer, wat ‘n verwysings area en vier degradeerde areas insluit. Fisiese en chemiese parameters was gemeet by elke area. Alge was geïdentifiseer deur mikroskopie en chlorofil-a konsentrasies was gemeet. Die alge spesies teenwoordig by die vyf studie areas was voorheen gekoppel aan kondisies gemeet by elke area. Gyrosigma rautenbachiae was n ideale bioindikator vir industriële uitvloei. Die diatom spesies Sunedra ulna, Nitzchia spp. en Cymbella spp. was geïdentifiseer by studie areas met ‘n lae pH. Die filamentige, groen alge Microspora quadrata en Klebsormidium acidophilum was ook oorvloedig by areas geaffekteer deur die suur mynwater. Metaal-tolerante K. rivulare was ook gevind in hierdie studie. Die cyanobakterie Oscillatoria tenius en Glaucospira sp. was geassosieer met verrykde kondisies. Die bakteriese populasies was gemonster van beide die water kollom en die sediment en geanaliseer deur middle van volgende generasie volgordebepaling vir identifikasie. Die phyla wat hoogs verteenwoordig was in al die monsters was die alpha-, beta- en gamma- Proteobakterie, Bacteriodetes en ongeklassifiseerde spesies. Die Bacteriodetes phylum was teëgekom teen beduidende hoër getalle by areas 1, 2, 3 & 5 in die Maart 2013 water monsters en areas 1 & 4 in die Maart 2013 sediment monsters. Firmicutes het beduidende hoër getalle gehad by areas 2 (Januarie 2013), 3 (Maart 2013) & 4 (Januarie 2013) in die water monsters. Beide die water en sediment monsters van area 2 (Maart 2013) & 4 (Januarie 2013) het beduidende hoër getalle Actinobacteria gehad. Die Chloroflexi phylum het beduidende hoër getalle in die area 4 & 5 (Januarie 2013) water monsters en area 5 (Januarie 2013) sediment monster gehad. Acidobacteria was slegs verteenwoordig deur beduidende hoër getalle in die Januarie 2013 sediment monsters van areas 1 & 5. Hierdie studie was die eerste van sy soort om die totale bakteriese populasie in ‘n natuurlike, suur myn-water geïmpakteerde vleiland in Suid-Afrika te bestudeer asook die eerste studie om ‘n organisme van die genus Marinobacterium te identifiseer in ‘n vlei. Die vlei se ekosisteem gesondheid was bepaal deur middle van ‘n spoedige bioassesseeerings hulpmiddel en ‘n voorgestelde bioindikator. Die bioassesserings hulpmiddel het die verwysings area geklassifiseer as meestal natuurlik, twee studie areas as gemodifiseer en twee as ernstig gemodifiseer. Die voorgestelde bakteriese bioindikator was eenvoudig om te gebruik en het die stabiliteit van die populasies verteenwoordig by die verskillende studie areas. Die bakteriese bioindikator was geïnkorporeer in die bioassesseerings hulpmiddel en dit was gevind dat die resultate ooreen stem.