Can bacteriophages in wastewater treatment plants do more?

Biological Processes for Wastewater Treatment and the Role of Bacteriophages

Biological processes are the foundation of municipal wastewater treatment, as they involve the decomposition of pollutants. These processes rely on the use of concentrated biomass from various microorganisms, known as activated sludge. Wastewater treatment plants serve as connections between environmental bacteria, the wastewater microbiome (including the human microbiome), and opportunistic pathogens. The main focus of treatment processes is the removal of sludge, suspended solids, phosphorus, and nitrogen compounds. However, the traditional method of using activated sludge is not effective in dealing with persistent industrial contaminants, pharmaceutical compounds, or antibiotic-resistant microorganisms [1, 2]. This article explores the potential solution of using bacteriophages in wastewater treatment to address these challenges.

Why Modification of Modern Wastewater Treatment Processes is Necessary

Modern wastewater treatment processes need to be modified on a micro scale due to the variable conditions that microbe-based processes for removing phosphorus and nitrogen compounds are exposed to. For example, the process of nitrification, which involves the conversion of ammonium nitrogen to nitrate by microorganisms, requires a constant level of nutrients in the wastewater entering the treatment plant. However, the nutrient composition can change depending on the season or time of day. Additionally, bacterial biomass grows slowly, and losses due to leaching or heavy rainfall can disrupt the transformation processes of nitrogen compounds. This leads to a decrease in pollutant removal efficiency and an accumulation of toxic chemical compounds that are harmful to the microorganisms themselves [2].

Furthermore, recent studies have shown that wastewater treatment plants are hotspots for the migration of antibiotic-resistant microorganisms into the environment and the transfer of antibiotic resistance genes between microbes [3]. The current treatment processes are not designed to effectively reduce the abundance of antibiotic resistance genes, and the methods used may only have an indirect impact. The high density of various microorganisms, surfactants, and disinfectants in treatment plants promotes the selection and horizontal transfer of genes. Additionally, microbes in treatment plants are exposed to stress from microplastics and heavy metals. As a result, new resistant pathogens are evolving and differ from the genotypes already present in the environment [3, 4].

The Potential Role of Bacteriophages in Wastewater Treatment

Bacteriophages offer potential solutions to some of the challenges mentioned above. Studies have shown that bacteriophages can be used to optimize wastewater treatment and methane digestion processes. Treatment plants are particularly interesting environments for studying bacteriophages due to their constant and large influx of microorganisms, including phages [5].

Bacteriophages are much smaller than bacterial cells, typically ranging from 20 nm to 200 nm in size. They have a simple structure consisting of a protein capsid containing the phage genome, which can be single- or double-stranded DNA or RNA. Some bacteriophages also have a lipid membrane surrounding the genetic material. Bacteriophages are the most abundant biological entities on Earth, outnumbering their bacterial hosts by up to 10 times. They can infect various types of bacteria, including gram-negative, gram-positive, and multidrug-resistant strains. The infection process begins with the phage adhering to the bacterial cell wall and injecting its genome into the host. Bacteriophages are essentially bacterial “eaters,” highlighting their high bactericidal capacity and specificity in action [5-7].

Most bacteriophages have high specificity against a particular type of bacteria and replicate at the site of infection. They take control of the bacterial machinery and resources for their own replication, inhibiting host growth and ultimately causing host death through lysis. Bacteriophages can achieve infection rates as high as 1023 infections per second [5].

Bacteriophage-bacteria interactions can have significant effects on the composition, function, and evolution of the microbiome. These properties make bacteriophages increasingly valuable in engineering, environmental, and medical sciences.

Applications of Bacteriophages in Wastewater Treatment

The use of bacteriophages can help modify the microbial community in wastewater treatment plants by eliminating undesirable strains. This modification can enhance aerobic processes in biological chambers and improve treatment efficiency. For example, one technological problem associated with activated sludge is swelling, which reduces sedimentation capacity and makes it difficult to separate sludge from treated liquid. Swelling leads to an increase in sediment volume and the accumulation of extracellular polysaccharide substances with high viscosity. These substances increase water retention in the sludge, hinder drainage, and affect floc stability. Filamentous bacteria, such as Gordonia spp., have been identified as the cause of these problems. Current chemical disinfection methods can produce toxic byproducts. The reduction of bacteria responsible for technological problems can be achieved by using single phage species or phage cocktails consisting of multiple species [7].

However, most bacteria in biological wastewater treatment systems are not suitable for culture under laboratory conditions, making it difficult to detect corresponding phages using classical methods. Advanced shotgun metagenomics methods combined with bioinformatics tools are needed to identify the phages [11, 12].

Wastewater treatment plants often contain large numbers of pathogenic microorganisms, such as E. coli, Salmonella spp., Staphylococcus aureus, and Campylobacter jejuni. Although these pathogens can be adsorbed in flocs and removed with excess sludge or ingested by protozoa, they can still pose a potential risk to the environment if the biological processes are disrupted. Bacteriophages can increase the safety of wastewater reuse and show promise in combating antibiotic resistance by reducing the number of organisms exposed to antibiotics and interrupting the transfer of antibiotic resistance genes [11, 12].

Opportunities and Obstacles

During phage infection, bacteria can develop defense mechanisms to resist the phages. They can change or lose their receptors, secrete substances that prevent phage adhesion, inhibit phage replication, or destroy phage-specific nucleic acids to prevent infection. Biofilms formed by microorganisms pose a particular challenge to phage infection. To overcome these defense mechanisms, phage cocktails consisting of multiple phage species can be used. It is important to improve strategies for predicting bacteriophage-host and bacteriophage-environment relationships. Molecular biology techniques can aid in tracking bacteria and bacteriophages, alleviating concerns about the use of phages in wastewater treatment biotechnology [5, 6].

Bacteriophages play an important ecological role in regulating the structure of microbial communities in wastewater treatment plants. Their predation can impact the efficiency of biological removal processes for nitrogen and phosphorus compounds by controlling swelling, foaming, or eliminating certain pathogens.

While most studies have focused on the use of bacteriophages for biological control in laboratory systems, further research is needed to explore the full-scale potential of phages in wastewater treatment plants. The variability of the activated sludge microbiome and the presence of microbial dark matter continue to challenge researchers [13]. Nonetheless, bacteriophages offer an alternative to traditional operational control methods and can become valuable tools for environmental and economic sustainability [5, 6].

Dr.-Ing. Edyta Łaskawiec – water and wastewater technologist, assistant professor at the Department of Environmental Biotechnology, Silesian University of Technology

References

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SDGs, Targets, and Indicators

SDGs Addressed or Connected to the Issues Highlighted in the Article:

1. SDG 6: Clean Water and Sanitation
2. SDG 9: Industry, Innovation, and Infrastructure
3. SDG 12: Responsible Consumption and Production
4. SDG 14: Life Below Water
5. SDG 15: Life on Land

Specific Targets Under Those SDGs Based on the Article’s Content:

• SDG 6.3: By 2030, improve water quality by reducing pollution, eliminating dumping and minimizing release of hazardous chemicals and materials, halving the proportion of untreated wastewater and substantially increasing recycling and safe reuse globally.
• SDG 9.4: By 2030, upgrade infrastructure and retrofit industries to make them sustainable, with increased resource-use efficiency and greater adoption of clean and environmentally sound technologies and industrial processes.
• SDG 12.4: By 2020, achieve the environmentally sound management of chemicals and all wastes throughout their life cycle, in accordance with agreed international frameworks, and significantly reduce their release to air, water and soil in order to minimize their adverse impacts on human health and the environment.
• SDG 14.1: By 2025, prevent and significantly reduce marine pollution of all kinds, in particular from land-based activities, including marine debris and nutrient pollution.
• SDG 15.1: By 2020, ensure the conservation, restoration and sustainable use of terrestrial and inland freshwater ecosystems and their services, in particular forests, wetlands, mountains and drylands, in line with obligations under international agreements.

Indicators Mentioned or Implied in the Article:

• Efficiency of pollutant removal in wastewater treatment plants
• Abundance of antibiotic-resistant microorganisms in wastewater treatment plants
• Transfer of antibiotic resistance genes between microbes in wastewater treatment plants
• Reduction of antibiotic resistance genes in wastewater treatment plants
• Presence of pathogenic microorganisms in wastewater treatment plants
• Effectiveness of bacteriophages in wastewater treatment and methane digestion processes
• Reduction of bacteria responsible for technological problems in activated sludge
• Detection of phages using advanced shotgun metagenomics methods combined with bioinformatics tools
• Impact of bacteriophages on community composition, function, and evolution of the microbiome

Table: SDGs, Targets, and Indicators

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