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Title: The factors affecting bacterial colonisation on microplastics and the impact of tertiary treatment of wastewater on the attached bacteria and microplastics
Authors: Rajcoomar, Saieshna 
Keywords: Microplastics;Wastewater;6S rRNA analysis;Biofilms;Tertiary treatment
Issue Date: May-2023
Microplastics (MPs) in aquatic environments have become an environmental concern globally.
In addition to the direct impact of these plastics on aquatic organisms, their surfaces could
serve as a unique habitat for various microbial communities through the formation of biofilms.
Various factors could play a role in microbial attachment and biofilm formation in wastewater.
This study aimed to assess potential factors that lead to biofilm formation on different types of
MPs in wastewater and determine the impact of UV and chlorine treatment on these biofilms.
In a laboratory scale experiment, MPs (low density polyethylene (LDPE), high density
polyethylene (HDPE), and polypropylene (PP) were exposed to untreated wastewater under
various conditions of temperature (20°C, 25°C and 35°C), light and dark conditions, as well as
aerobic and anaerobic conditions for a period of five weeks. The formation of biofilms on MPs
was quantified using optical density (OD660) measurements. The highest biofilm formation was
observed in week 3, with an OD of 1.77. Thereafter, a decline in OD was observed, reaching
an OD of 1.1 by week 5. This change in biofilm concentration over the week corresponded to
changes in nutrient (nitrite, nitrate and ammonia) concentration in the media. A positive
correlation was observed between the changes in biofilm concentration and nitrite (r = 0.824)
and ammonia (r = 0.1) levels in the media. Meanwhile, a negative correlation observed for
nitrate concentration (r=-0.673). Factors such as dark conditions, 25 C, and aerobic conditions
presented the highest median biofilm formation with an OD value of 1.6, 1.7 and 1.6,
respectively. It was also observed that polyethylene had higher biofilm concentrations
compared to the polypropylene. Furthermore, rough MPs had higher biofilm formation than
smooth MPs, with median ODs of 1.7 and 1.6 respectively. The microbial communities in the
biofilms and wastewater medium were characterised by 16S rRNA amplicon sequencing. The
results revealed that the alpha diversity (richness, evenness, and diversity) was lower in
wastewater compared to the biofilms. It was observed that PP supported the most diverse
bacterial community ( H’= 2.51138 and Simpson index= 11.096), while HDPE supported the
least diverse bacterial community (H’= 0.88779 and Simpson index= 1.5324). Beta diversity
using the Jaccard distance index revealed that the most similar communities were observed
among biofilms from the three types of MPs while the most dissimilar communities were
observed between the biofilm and wastewater medium communities. The most dominant phyla
in both the biofilms and wastewater medium during the five weeks were Proteobacteria,
Bacteroidetes and Planctomycetes. The bacterial communities, however, varied for each type
of plastic and the wastewater medium. It was observed that Methylotenera, Hydrogenophaga, and Rhodanobacter was the most abundant genera in biofilms whereas C39(45.25%) and
Luteimonas(18.96%) were the abundant genera in the wastewater medium. Methylotenera
mobilis was the most common species among the three types of MPs. In addition, pathogenic
species such as Mycobacterium arupense and Methylobacterium adhaesivum were detected in
abundance on LDPE and PP. To assess the impact of UV treatment and chlorination on the
attached biofilms, the microplastics with attached biofilm were exposed to UV-C and Chlorine
(5 mg/L) treatment for 60 minutes. The biofilms were inactivated (100%) after 30 mins of UV
treatment, whereas 10 min was sufficient to achieve 100% inactivation of biofilm by chlorine
treatment. In conclusion, the research presented in this study has made substantial contributions
to our understanding of the role that environmental factors play in the formation of biofilm on
MP surfaces.
Submitted in fulfillment of the degree of Master of Applied Science: Biotechnology, Durban University of Technology, Durban, South Africa, 2022.
Appears in Collections:Theses and dissertations (Applied Sciences)

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