Browsing by Author "Zuo, Pengxiao"

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    Biofilm control using encapsulated phages and the impact of these phage interactions on antibiotic resistance dissemination
    (2022-08-05) Zuo, Pengxiao; Alvarez, Pedro J.J.
    Biofilms pose a major challenge to water security because they may harbor and protect pathogens, including antibiotic resistant bacteria. Conventional chemical disinfectants are not efficient at penetrating biofilms and produce undesirable disinfection by products, hence the need for a greener chemical free biocide. Bacteriophages (phages) are one such green alternative for microbial control, however, inefficient delivery and lack of precise targeting greatly lowers their efficacy and the consequences of phage treatment on antibiotic resistance dissemination are not well understood. This dissertation provides technology with potential to improve the precision of phage treatment and mechanistic understanding of how phages affect antibiotic resistant bacteria and resistance dissemination. To enhance phage-based biofouling mitigation in water storage system, crosslinked chitosan was used to form a pH-responsive phage encapsulation that can release phages at around pH 5, but otherwise remain stable for over one month in pH neutral tap water. This encapsulation allows for controlled phage release only as needed in response to the presence of a nascent biofilm because the biofilm inner layers tend to be acidic, as shown by confocal microscopy with pH-indicator dye SNARF-4F. A facile method was developed to coat encapsulated phages onto plastic and fiberglass surfaces and demonstrated that this coating mitigates biofilm formation in tap water amended with glucose and Pseudomonas aeruginosa. Furthermore, this coating allows phages to attack biofilms from a bottom-up approach, bypassing the limitation of poor biocide penetration into the biofilm matrix. Overall, this offers a potential green alternative for biofilm mitigation and highlights the possible applications and benefits encapsulation can bring to biocides. Phages are known to control bacteria populations; yet, much is unknown about their differential effects on antibiotic resistant bacteria (ARB) proliferation. Chapter 4 shows that beta-lactam resistance may fortuitously confer phage resistance as a critical factor for enhanced ARB proliferation. Following sub-lethal exposure to amoxicillin, Escherichia coli experienced lipopolysaccharides (LPS) modifications and became cross-resistant to various phages that adsorb to receptors on LPS as the first infection step. The relevance of this phenomenon was demonstrated using activated sludge microcosms where cross-resistant E. coli experienced significantly less decay after five days than the wildtype. Furthermore, cross-resistant E. coli (but not wildtype) proliferated in Luria broth-fed microcosms. A demonstratable fitness cost associated with amoxicillin resistance exists; however, due to acquired phage resistance, cross-resistant E. coli had greater fitness than the wildtype. Overall, this demonstrates that antibiotics can alter interactions between phages and bacteria, resulting in an overlooked competitive advantage for antibiotic resistance propagation. The common co-occurrence of antibiotics and phages in both natural and engineered environments underscore the need to understand their interactions and implications for bacterial control and antibiotic resistance propagation. Chapter 5 reports that aminoglycoside antibiotics (acting as bacterial protein synthesis inhibitors) impeded replication of various phages. This hindered the efficacy of phages against bacterial growth and biofilm formation, and diminished bacterial fitness cost that suppress antibiotic resistance emergence. Therefore, this highlights an overlooked antagonistic effect of aminoglycosides on phages, which may provide an advantage to affected bacteria in the common presence of phages due to less hindrance to bacterial growth and antibiotic resistance development. Overall, encapsulation technology can improve the application of phages as self-replicating biocides by enabling controlled phage release only as needed in response to nascent biofilm formation. Furthermore, coating methods for encapsulated phages can selectively protect just the surfaces vulnerable to biofouling rather than the entire system and release phages to attack from under the biofilm, avoiding challenge of penetrating the protective biofilm matrix. However, some ARB may have fortuitous cross resistance to phages, resulting in a significant competitive advantage for ARB proliferation under selective pressure from phages. Additionally, antibiotics like aminoglycosides (acting as bacterial protein synthesis inhibitors) impeded replication of various phages, which may attenuate phage suppression of bacterial growth, biofilm formation, antibiotic tolerance, and maintenance of antibiotic resistance genes.
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    Redistribution of intracellular and extracellular free & adsorbed antibiotic resistance genes through a wastewater treatment plant by an enhanced extracellular DNA extraction method with magnetic beads
    (Elsevier, 2019) Yuan, Qing-Bin; Huang, Ya-Meng; Wu, Wen-Bin; Zuo, Pengxiao; Hu, Nan; Zhou, Yong-Zhang; Alvarez, Pedro J.J.
    Due to the limitations of current extraction methods, extracellular DNA (eDNA) is rarely discerned from intracellular DNA (iDNA) despite having unique contributions to antibiotic resistance genes (ARGs) propagation. Furthermore, eDNA may be free (f-eDNA) or adsorbed to or suspended solids, including cells (a-eDNA), which affects ARG persistence and transmissivity. We developed a novel method using magnetic beads to separate iDNA, a-eDNA, and f-eDNA to assess how these physical states of ARGs change across a wastewater treatment plant. This method efficiently extracted eDNA (>85.3%) with higher recovery than current methods such as alcohol precipitation, CTAB-based extraction, and DNA extraction kits (<10%). Biological treatment and UV disinfection decreased the concentration of intracellular ARGs (iARGs) and adsorbed extracellular ARGs (a-eARGs), causing an increase of released free extracellular ARGs (f-eARGs). More ARGs were discharged through the wasted biosolids than in the effluent; iARGs and a-eARGs are prevalent in wasted biosolids ((73.9 ± 22.5) % and (23.4 ± 15.3) % of total ARGs respectively), while f-eARGs were prevalent in the effluent ((90.3 ± 16.5) %). Bacterial community analysis showed significant correlations between specific genera and ARGs (e.g., Aeromonas, Pseudomonas and Acinetobacter were strongly correlated with multidrug-resistance gene blaTEM). This treatment system decreased the discharge of iARGs to receiving environments, however, increased eARG concentrations were present in the effluent, which may contribute to the environmental resistome.