Oxidative stress induced by Etoposide anti-cancer chemotherapy drives the emergence of tumor-associated bacteria resistance to fluoroquinolones.

INTRODUCTION: Antibiotic-resistant bacterial infections, such as Pseudomonas aeruginosa and Staphylococcus aureus, are prevalent in lung cancer patients, resulting in poor clinical outcomes and high mortality. Etoposide (ETO) is an FDA-approved chemotherapy drug that kills cancer cells by damaging DNA through oxidative stress. However, it is unclear if ETO can cause unintentional side effects on tumor-associated microbial pathogens, such as inducing antibiotic resistance. OBJECTIVES: We aimed to show that prolonged ETO treatment could unintendedly confer fluoroquinolone antibiotic resistance to P. aeruginosa, and evaluate the effect of tumor-associated P. aeruginosa on tumor progression. METHODS: We employed experimental evolution assay to treat P. aeruginosa with prolonged ETO exposure, evaluated the ciprofloxacin resistance, and elucidated the gene mutations by DNA sequencing. We also established a lung tumor-P. aeruginosa bacterial model to study the role of ETO-evolved intra-tumoral bacteria in tumor progression using immunostaining and confocal microscopy. RESULTS: ETO could generate oxidative stress and lead to gene mutations in P. aeruginosa, especially the gyrase (gyrA) gene, resulting in acquired fluoroquinolone resistance. We further demonstrated using a microfluidic-based lung tumor-P. aeruginosa coculture model that bacteria can evolve ciprofloxacin (CIP) resistance in a tumor microenvironment. Moreover, ETO-induced CIP-resistant (EICR) mutants could form multicellular biofilms which protected tumor cells from ETO killing and enabled tumor progression. CONCLUSION: Overall, our preclinical proof-of-concept provides insights into how anti-cancer chemotherapy could inadvertently allow tumor-associated bacteria to acquire antibiotic resistance mutations and shed new light on the development of novel anti-cancer treatments based on anti-bacterial strategies.

Simultaneous Dissemination of Nanoplastics and Antibiotic Resistance by Nematode Couriers.

Nanoplastics (NPs) are increasingly recognized as a newly emerging pollutant in the environment. NPs can enable the colonization of microbial pathogens on their surfaces and adsorb toxic pollutants, such as heavy metals and residual antibiotics. Although the dissemination of plastic particles in water bodies and the atmosphere is widely studied, the dissemination of NPs and adsorbed pollutants on land, via biological means, is poorly understood. Since soil animals, such as the bacterivorous nematode Caenorhabditis elegans (C. elegans), are highly mobile, this raises the possibility that they play an active role in disseminating NPs and adsorbed pollutants. Here, we established that antibiotic-resistant bacteria could aggregate with antibiotic-adsorbed NPs to form antibiotic-adsorbed NP-antibiotic resistant bacteria (ANP-ARB) aggregates, using polymyxins (colistin) as a proof-of-concept. Colistin-resistant mcr-1 bearing Escherichia coli from a mixed population of resistant and sensitive bacteria selectively aggregate with colistin-ANPs. In the soil microcosm, C. elegans fed on ANP-ARB clusters, resulting in the rapid spread of ANP-ARB by the nematodes across the soil at a rate of 40-60 cm per day. Our work revealed insights into how NPs could still disseminate across the soil faster than previously thought by "hitching a ride" in soil animals and acting as agents of antibiotic-resistant pathogens and antibiotic contaminants. This poses direct risks to ecology, agricultural sustainability, and human health.

Biofilm matrix cloaks bacterial quorum sensing chemoattractants from predator detection.

Microbes often secrete high levels of quorum sensing (QS) autoinducers into the environment to coordinate gene expression and biofilm formation, but risk detection and subsequent predation by bacterivorous predators. With such prominent signaling molecules acting as chemoattractants that diffuse into the environment at alarmingly high concentrations, it is unclear if bacterial cells can mask their chemical trails from predator detection. Here, we describe a microbial-based anti-detection adaptation, termed as "biofilm cloak", where the biofilm prey produced biofilm matrix exopolysaccharides that "locked" and reduced the leaching of autoinducers into the milieu, thereby concealing their trails to the detection by the bacterivorous Caenorhabditis elegans nematode. The exopolysaccharides act as common good for the non-producers to hide their autoinducers from predator detection. Deficiency in chemosensory gene odr-10 in mutant animals abrogated their ability to detect autoinducers and migrate toward their prey in a directed manner, which led to lower population growth rate of animals. Hence, restriction of bacterial communication activities to the confinements of biofilms is a novel approach for predator evasion, which plays a fundamental role in shaping ecological dynamics of microbial communities and predator-prey interactions.

Biofilm matrix disrupts nematode motility and predatory behavior.

In nature, bacteria form biofilms by producing exopolymeric matrix that encases its entire community. While it is widely known that biofilm matrix can prevent bacterivore predation and contain virulence factors for killing predators, it is unclear if they can alter predator motility. Here, we report a novel "quagmire" phenotype, where Pseudomonas aeruginosa biofilms could retard the motility of bacterivorous nematode Caenorhabditis elegans via the production of a specific exopolysaccharide, Psl. Psl could reduce the roaming ability of C. elegans by impeding the slithering velocity of C. elegans. Furthermore, the presence of Psl in biofilms could entrap C. elegans within the matrix, with dire consequences to the nematode. After being trapped in biofilms, C. elegans could neither escape effectively from aversive stimuli (noxious blue light), nor leave easily to graze on susceptible biofilm areas. Hence, this reduced the ability of C. elegans to roam and predate on biofilms. Taken together, our work reveals a new function of motility interference by specific biofilm matrix components, and emphasizes its importance in predator-prey interactions.

Vanillin inhibits PqsR-mediated virulence in Pseudomonas aeruginosa.

Reduced efficacy of antibiotics in bacterial diseases is a global concern in clinical settings. Development of anti-virulence compounds which disarm bacterial virulence is an attractive therapeutic agent for complementary antibiotics usage. One potential target for anti-virulence compounds is quorum sensing (QS), the intercellular communication system in most pathogens, such as Pseudomonas aeruginosa. QS inhibitors (QSIs) can inhibit QS effectively, attenuate QS-mediated virulence, and improve host clearance of infections. While studies focused on developing homoserine-based las QSI, few targeted the quinolone-based pqs QS, which implicated host cytotoxicity and biofilm formation. It is imperative to develop novel anti-pqs-QS therapeutics for combinatorial antibiotic treatment of microbial diseases. We employed a gfp-based transcriptional pqs biosensor to screen a natural compounds library and identify vanillin (4-hydroxy-3-methoxybenzaldehyde), the primary phenolic aldehyde of vanilla bean. The vanillin inhibited pqs expression and its associated phenotypes, namely pyocyanin production and twitching motility in P. aeruginosa. Molecular docking results revealed that vanillin binds to the active site of PqsR, the PQS-binding response regulator. Combinatorial treatment of vanillin with antimicrobial peptide (colistin) inhibited biofilm growth in vitro and improved treatment in the in vivo C. elegans acute infection model. We demonstrated that vanillin could dampen pqs QS and associated virulence, thus providing novel therapeutic strategies against P. aeruginosa infections.