Polymer-enhanced peroxidase activity of ceria nanozyme for highly sensitive detection of alkaline phosphatase.

Nanoceria have demonstrated a wide array of catalytic activity similar to natural enzymes, holding considerable significance in the colorimetric detection of alkaline phosphatase (ALP), which is a biomarker of various biological disorders. However, the issues of physiological stability and formation of protein corona, which are strongly related to their surface chemistry, limit their practical application. In this work, CeO2 nanoparticles characterized by enhanced dimensional uniformity and specific surface area were synthesized, followed by encapsulation with various polymers to further increase catalytic activity and physiological stability. Notably, the CeO2 nanoparticles encapsulated within each polymer exhibited improved catalytic characteristics, with PAA-capped CeO2 exhibiting the highest performance. We further demonstrated that the PAA-CeO2 obtained with enhanced catalytic activity was attributed to an increase in surface negative charge. PAA-CeO2 enabled the quantitative assessment of AA activity within a wide concentration range of 10 to 60 µM, with a detection limit of 0.111 µM. Similarly, it allowed for the evaluation of alkaline phosphatase activity throughout a broad range of 10 to 80 U/L, with a detection limit of 0.12 U/L. These detection limits provided adequate sensitivity for the practical detection of ALP in human serum.

Antimicrobial Peptide-Inspired Design of Amino-Modified Lignin with Improved Antimicrobial Activities.

A major challenge to make use of lignin as an antimicrobial material is the weak antimicrobial activity of industrial lignin. Inspired by the antimicrobial mechanism of actions of antimicrobial peptides, alkyldiamines were employed as lysine mimics for lignin modifications. Accordingly, aminoalkyl-modified lignins with different degrees of substitution of amino groups and different hydrophobicity were synthesized. The chemical structure, properties, and antimicrobial activities of the as-prepared aminoalkyl lignins were thoroughly characterized with state-of-the-art technologies. The results indicated that aminobutyl lignin showed enhanced antimicrobial activity against S. aureus and E. coli and performed even better than copper ions. The antimicrobial mechanism of action of the as-prepared aminobutyl lignin was similar to that of polylysine, which damaged the cell membrane, leading to the leakage of intracellular molecules and death of the cell. This study provides a feasible approach to afford modified lignin with enhanced antimicrobial performance, which would facilitate the high-value valorization of lignin as biological materials.

Fundicoccus culcitae sp. nov., a novel potential bacteriocin producing bacterium isolated from a spoiled eye mask.

A Gram-positive, facultatively anaerobic, oval beaded-shape, oxidase-negative, and non-motile bacterium designated DM20194951T was isolated from a spoiled eye mask obtained from Guangdong, China. Based on the 16S rRNA gene sequence, phylogenetic analysis indicated that strain DM20194951T showed the highest sequence similarity (95.8%) to Fundicoccus ignavus WS4937T. Meanwhile, strain DM20194951T could be distinguished from the type strains in the genus Fundicoccus by distinct phenotypic and genotypic traits. Strain DM20194951T grew variably with 1-2% (w/v) NaCl and tolerated pH 6.0-10.0. Growth was observed from 28 to 37 °C. The diagnostic diamino acids in the cell-wall peptidoglycan consisted of aspartic and glutamic acids as well as alanine. The predominant fatty acids were C18:1 ω9c, C16:0, and C16:1 ω9c. In the polar lipid profile, two glycolipids, three phospholipids, one phosphatidylglycerol, and one diphosphatidylglycerol were found. No respiratory quinones were detected. The DM20194951T genome is 3.2 Mb in size and contains a G + C content of 38.1%. A gene cluster for lactococcin 972 family bacteriocin production was found in the DM20194951T genome. Based on morphological, genotypic, and phylogenetic data, strain DM20194951T should be considered to represent a novel species in the genus Fundicoccus, for which the name Fundicoccus culcitae sp. nov. is proposed with the type strain DM20194951T (= KCTC 43472T = GDMCC 1.3614T).

Visible light-regulated cationic polymer coupled with photodynamic inactivation as an effective tool for pathogen and biofilm elimination.

BACKGROUND: Pathogenic microorganism pollution has been a challenging public safety issue, attracting considerable scientific interest. A more problematic aspect of this phenomenon is that planktonic bacteria exacerbate biofilm formation. There is an overwhelming demand for developing ultra-efficient, anti-drug resistance, and biocompatibility alternatives to eliminate stubborn pathogenic strains and biofilms. RESULTS: The present work aims to construct a visible light-induced anti-pathogen agents to ablate biofilms using the complementary merits of ROS and cationic polymers. The photosensitizer chlorin e6-loaded polyethyleneimine-based micelle (Ce6-TPP-PEI) was constructed by an amphiphilic dendritic polymer (TPP-PEI) and physically loaded with photosensitizer chlorin e6. Cationic polymers can promote the interaction between photosensitizer and Gram-negative bacteria, resulting in enhanced targeting of PS and lethality of photodynamic therapy, and remain active for a longer duration to prevent bacterial re-growth when the light is turned off. As expected, an eminent antibacterial effect was observed on the Gram-negative Escherichia coli, which is usually insensitive to photosensitizers. Surprisingly, the cationic polymer and photodynamic combination also exert significant inhibitory and ablative effects on fungi and biofilms. Subsequently, cell hemolysis assessments suggested its good biocompatibility. CONCLUSIONS: Given the above results, the platform developed in this work is an efficient and safe tool for public healthcare and environmental remediation.

Outer membrane protein of OmpF contributes to swimming motility, biofilm formation, osmotic response as well as the transcription of maltose metabolic genes in Citrobacter werkmanii.

Bacterial outer membrane proteins (Omps) are essential for environmental sensing, stress responses, and substance transport. Our previous study discovered that OmpA contributes to planktonic growth, biocide resistance, biofilm formation, and swimming motility in Citrobacter werkmanii, whereas the molecular functions of OmpF in this strain are largely unknown. Thus, in this study, the ompF gene was firstly knocked out from the genome of C. werkmanii using a homologous recombination method, and its phenotypical alternations of ∆ompF were then thoroughly characterized using biochemical and molecular approaches with the parental wild type (WT) and complementary (∆ompF-com) strains. The results demonstrated that the swimming ability of ∆ompF on semi-solid plates was reduced compared to WT due to the down-regulation of flgC, flgH, fliK, and fliF. Meanwhile, ompF deletion reduces biofilm formation on both glass and polystyrene surfaces due to decreased cell aggregation. Furthermore, ompF inactivation induced different osmotic stress (carbon sources and metal ions) responses in its biofilms when compared to WT and ∆ompF-com. Finally, a total of 6 maltose metabolic genes of lamB, malE, malK, malG, malM, and malF were all up-regulated in ∆ompF. The gene knockout and HPLC results revealed that the MalEFGK2 cluster was primarily responsible for maltose transport in C. werkmanii. Furthermore, we discovered for the first time that the upstream promoter of OmpF and its transcription can be combined with and negatively regulated by MalT. Overall, OmpF plays a role in a variety of biochemical processes and molecular functions in C. werkmanii, and it may even act as a targeted site to inhibit biofilm formation.

Synthesis, molecular docking, and evaluation of antibacterial activity of 1,2,4-triazole-norfloxacin hybrids.

A series of 1,2,4-triazole-norfloxacin hybrids was designed, synthesized, and evaluated for in vitro antibacterial activity against common pathogens. All the newly synthesized compounds were characterized by Fourier-transform infrared spectrophotometry, proton and carbon nuclear magnetic resonance, and electrospray ionization-mass spectrometry. Representative compounds from each step of the synthesis were further characterized by X-ray crystallography. Many of the compounds synthesized exhibited antibacterial activity superior to that of norfloxacin toward both, gram-positive and gram-negative bacteria. The toxicity of the 1,2,4-triazole-norfloxacin hybrids toward bacterial cells was 32-512 times higher than that toward mouse fibroblast cells. Moreover, hemolysis was not observed at concentrations of 64 µg/mL, suggesting good biocompatibility. Molecular docking showed a least binding energy of -9.4 to -9.7 kcal/mol, and all compounds were predicted to show remarkable affinity for the bacterial topoisomerase IV.

Roles of ompA of Citrobacter werkmanii in bacterial growth, biocide resistance, biofilm formation and swimming motility.

The genus Citrobacter is commonly found in environmental and industrial settings, some members of which have been used for bioremediation of heavy metals owing to the absorption ability of their biofilms. Although our previous studies have found that the outer membrane protein A (OmpA) contributes to the process of Citrobacter werkmanii biofilm formation, the underlying mechanisms remain elusive. Therefore, we deleted ompA from the genome of C. werkmanii and investigated its phenotypes in comparison to the wild type strain (WT) and the complementary strain using biochemical and molecular techniques including RNA-Seq. Our results demonstrated that the deletion of ompA led to an increase in biofilm formation on both polystyrene and glass surfaces due to upregulation of some biofilm formation related genes. Meanwhile, swimming ability, which is mediated by activation of flagellar assembly genes, was increased on semi-solid plates in the ∆ompA strain when compared with WT. Additionally, inactivation of ompA also caused increased 1,2-benzisothiazolin-3-one (BIT) resistance, differential responses to Ca2+ stress, curli protein expression and cellulose production. Finally, ∆ompA caused differential expression of a total of 1470 genes when compared with WT, of which 146 were upregulated and 1324 were downregulated. These genes were classified into different Gene Ontology (GO) and KEGG pathways. In summary, ompA in C. werkmanii contributes to a variety of biological functions and may act as a target site to modulate biofilm formation. KEY POINTS: • ompA is a negative regulator for biofilm formation by C. werkmanii. • ompA inhibits swimming motility of C. werkmanii. • ompA deletion causes different expression profiles in C. werkmanii.

The Connection between Czc and Cad Systems Involved in Cadmium Resistance in Pseudomonas putida.

Heavy metal pollution is widespread and persistent, and causes serious harm to the environment. Pseudomonas putida, a representative environmental microorganism, has strong resistance to heavy metals due to its multiple efflux systems. Although the functions of many efflux systems have been well-studied, the relationship between them remains unclear. Here, the relationship between the Czc and Cad systems that are predominantly responsible for cadmium efflux in P. putida KT2440 is identified. The results demonstrated that CzcR3, the response regulator of two-component system CzcRS3 in the Czc system, activates the expression of efflux pump genes czcCBA1 and czcCBA2 by directly binding to their promoters, thereby helping the strain resist cadmium stress. CzcR3 can also bind to its own promoter, but it has only a weak regulatory effect. The high-level expression of czcRS3 needs to be induced by Cd2+, and this relies on the regulation of CadR, a key regulator in the Cad system, which showed affinity to czcRS3 promoter. Our study indicates that the Cad system is involved in the regulation of the Czc system, and this relationship is important for maintaining the considerable resistance to cadmium in P. putida.

Antibacterial Nanorods Made of Carbon Quantum Dots-ZnO Under Visible Light Irradiation.

Due to secondary pollution from bactericidal substances, the importance of eliminating microbial contamination has become a controversial topic. Three antibacterial nanorod materials of carbon quantum dots-zinc oxide (1/3CQDs-ZnO, CQDs-ZnO, and 2CQDs-ZnO), in which ZnO nanorods are surrounded by carbon quantum dots (CQDs), were successful prepared via in-situ sol-gel chemistry. Antibacterial nanorods of CQDs-ZnO had strong antibacterial activity under visible light irradiation, and a concentration of 0.1 mg/L was able to kill more than 96% of bacteria. The photocatalytic antibacterial mechanism was also studied. CQDs-ZnO produced more than three times the free radicals than pure ZnO under visible light irradiation. These free radicals destroyed the bacterial matrix of the cell wall and released cell proteins and nucleic acids. Moreover, CQDs-ZnO showed low cytotoxicity and can be used in medical applications.

Biological functions of nirS in Pseudomonas aeruginosa ATCC 9027 under aerobic conditions.

Through our previous study, we found an up-regulation in the expression of nitrite reductase (nirS) in the isothiazolone-resistant strain of Pseudomonas aeruginosa. However, the definitive molecular role of nirS in ascribing the resistance remained elusive. In the present study, the nirS gene was deleted from the chromosome of P. aeruginosa ATCC 9027 and the resulting phenotypic changes of ΔnirS were studied alongside the wild-type (WT) strain under aerobic conditions. The results demonstrated a decline in the formations of biofilms but not planktonic growth by ΔnirS as compared to WT, especially in the presence of benzisothiazolinone (BIT). Meanwhile, the deletion of nirS impaired swimming motility of P. aeruginosa under the stress of BIT. To assess the influence of nirS on the transcriptome of P. aeruginosa, RNA-seq experiments comparing the ΔnirS with WT were also performed. A total of 694 genes were found to be differentially expressed in ΔnirS, of which 192 were up-regulated, while 502 were down-regulated. In addition, these differently expressed genes were noted to significantly enrich the carbon metabolism along with glyoxylate and dicarboxylate metabolisms. Meanwhile, results from RT-PCR suggested the contribution of mexEF-oprN to the development of BIT resistance by ΔnirS. Further, c-di-GMP was less in ΔnirS than in WT, as revealed by HPLC. Taken together, our results confirm that nirS of P. aeruginosa ATCC 9027 plays a role in BIT resistance along with biofilm formation and further affects several metabolic patterns under aerobic conditions.

Functional roles of norCB in Pseudomonas aeruginosa ATCC 9027 under aerobic conditions.

Nitric oxide (NO) reductase (NorCB) of Pseudomonas aeruginosa is an essential enzyme that metabolizes NO and alleviates anaerobic NO toxicity during denitrification processes under anaerobic conditions. However, the molecular functions of norCB in the presence of oxygen are poorly understood. This study utilized norCB knockout from P. aeruginosa ATCC 9027 to analyze the resulting phenotypic changes of ΔnorCB in comparison to the wild-type parental strain (WT) and the complementary strain (ΔnorCB-com). The results demonstrated an increase in planktonic growth and biofilm formation by ΔnorCB compared to WT and ΔnorCB-com in the presence of isothiazolones under aerobic conditions. Deletion of norCB led to increased swimming ability and decreased pyocyanin production. Inactivation of norCB also led to an increase of cellular H2 O2 concentration due to decreased activity of its catalases. In addition, the deletion of norCB also influenced the relative expressions of several other genes, including norD, nirS, hmgA, and hpd. These findings provide preliminary evidence that norCB in P. aeruginosa plays an essential role in bacterial life process under aerobic conditions and improves the application of denitrification in the next step.

Diallyl disulfide from garlic oil inhibits Pseudomonas aeruginosa virulence factors by inactivating key quorum sensing genes.

Garlic oil can disrupt the quorum sensing (QS) pathways of the opportunistic pathogen Pseudomonas aeruginosa; however, the underlying mechanisms for this effect are unclear. Diallyl disulfide (DADS) is one of the most abundant sulfur-containing compounds in garlic oil. This study investigated the effects of DADS on the growth, virulence factor production (elastase, pyocyanin, biofilm, and swarming motility), and essential gene expression of P. aeruginosa PAO1, particularly as they apply to QS and virulence. DADS at 1.28 mg/mL did not affect P. aeruginosa PAO1 growth, although it decreased elastase and pyocyanin production, biofilm formation, and swarming motility. Each of these phenomena is regulated by the three QS systems of P. aeruginosa PAO1 (las, rhl, and pqs). Real-time q-PCR revealed that DADS down-regulated the transcription levels of several important QS genes (lasI, lasR, rhlI, rhlR, pqsA, and pqsR) in the three systems. Furthermore, the transcription levels of QS-regulated virulence genes were also down-regulated. The lasB gene, encoding LasB elastase, is co-regulated by the las, rhl, and pqs systems, and thus the down-regulation of genes across the three systems further down-regulated lasB. Additionally, phzM (encoding pyocyanin), pslB (responsible for the production of a biofilm matrix polysaccharide), and chiC (encoding chitinase) were positively activated by LasR, and a decrease in lasR transcription further down-regulated the transcription of phzM, pslB, and chiC. Hence, DADS inhibits P. aeruginosa PAO1 virulence factors by inactivating the transcription of key genes across three different QS systems.

Role of Ttca of Citrobacter Werkmanii in Bacterial Growth, Biocides Resistance, Biofilm Formation and Swimming Motility.

To screen, identify and study the genes involved in isothiazolone resistance and biofilm formation in Citrobacter werkmanii strain BF-6. A Tn5 transposon library of approximately 900 mutants of C. werkmanii strain BF-6 was generated and screened to isolate 1,2-benzisothiazolin-3-one (BIT) resistant strains. In addition, the tRNA 2-thiocytidine (32) synthetase gene (ttcA) was deleted through homologous recombination and the resulting phenotypic changes of the ΔttcA mutant were studied. A total of 3 genes were successfully identified, among which ΔttcA mutant exhibited a reduction in growth rate and swimming motility. On the other hand, an increase in biofilms formation in ΔttcA were observed but not with a significant resistance enhancement to BIT. This work, for the first time, highlights the role of ttcA gene of C. werkmanii strain BF-6 in BIT resistance and biofilm formation.

Purification and molecular weight distribution of a key exopolysaccharide component of Bacillus megaterium TF10.

Extracellular polymeric substances (EPS) are organic metabolic compounds excreted by microorganisms. They largely impact microbial aggregate structures and functions. Extracellular polysaccharides (EP) in EPS are responsible for the formation of microbial aggregates. In this work, we successfully separated and characterized EP from EPS of the bacterium Bacillus megaterium TF10. Extraction of EP from EPS was optimized using Sevag's reagent. Chemical characteristics, functional groups, and molecular weight (MW) distribution of EP were compared with the harvested EPS and soluble microbial products (SMP). We found that the polymers of lower MW and free proteins were successfully removed by Sevag's reagent. The higher MW components of EPS were predominantly polysaccharides, while the polymers of lower MW tended to secrete to the supernatant and were described as SMP. A part of the proteins in the EP was polysaccharide-bonded. Our results can be further used in elucidating the complex flocculation mechanisms in which EP play a major role.

Complete genome sequence of Citrobacter werkmanii strain BF-6 isolated from industrial putrefaction.

BACKGROUND: In our previous study, Citrobacter werkmanii BF-6 was isolated from an industrial spoilage sample and demonstrated an excellent ability to form biofilms, which could be affected by various environmental factors. However, the genome sequence of this organism has not been reported so far. RESULTS: We report the complete genome sequence of C. werkmanii BF-6 together with the description of the genome features and its annotation. The size of the complete chromosome is 4,929,789 bp with an average coverage of 137×. The chromosome exhibits an average G + C content of 52.0%, and encodes 4570 protein coding genes, 84 tRNA genes, 25 rRNA operons, 3 microsatellite sequences and 34 minisatellite sequences. A previously unknown circular plasmid designated as pCW001 was also found with a length of 212,549 bp and a G + C content of 48.2%. 73.5%, 75.6% and 92.6% of the protein coding genes could be assigned to GO Ontology, KEGG Pathway, and COG (Clusters of Orthologous Groups) categories respectively. C. werkmanii BF-6 and C. werkmanii NRBC 105721 exhibited the closest evolutionary relationships based on 16S ribosomal RNA and core-pan genome assay. Furthermore, C. werkmanii BF-6 exhibits typical bacterial biofilm formation and development. In the RT-PCR experiments, we found that a great number of biofilm related genes, such as bsmA, bssR, bssS, hmsP, tabA, csgA, csgB, csgC, csgD, csgE, and csgG, were involved in C. werkmanii BF-6 biofilm formation. CONCLUSIONS: This is the first complete genome of C. werkmanii. Our work highlights the potential genetic mechanisms involved in biofilm formation and paves a way for further application of C. werkmanii in biofilms research.

Comparison of transcriptomes of wild-type and isothiazolone-resistant Pseudomonas aeruginosa by using RNA-seq.

Isothiazolone biocides (such as Kathon) are widely used in a variety of industrial and domestic applications. However, the mechanisms through which bacteria develop resistance to these biocides are not completely clear. A better understanding of these mechanisms can contribute to optimal use of these biocides. In this study, transcription profiles of a Kathon-resistant strain of Pseudomonas aeruginosa (Pa-R) and the wild-type strain were determined using RNA sequencing (RNA-Seq) with the Illumina HiSeq 2000 platform. RNA-Seq generated 18,657,896 sequence reads aligned to 7093 genes. In all, 1550 differently expressed genes (DEGs, log2 ratio ≥1, false discovery rate (FDR) ≤0.001) were identified, of which 482 were up-regulated and 1068 were down-regulated. Most Kathon-induced genes were involved in metabolic and cellular processes. DEGs significantly enriched nitrogen metabolism and oxidative phosphorylation pathways. In addition, Pa-R showed cross-resistance to triclosan and ciprofloxacin and showed repressed pyocyanin production. These results may improve our understanding of the resistance mechanisms of P. aeruginosa against isothiazolones, and provide insight into the development of more efficient isothiazolones.

The dynamics and mechanism of the antimicrobial activity of tea tree oil against bacteria and fungi.

Tea tree oil (TTO) is a yellow liquid extracted from Melaleuca alternifolia. Although the antimicrobial activity of TTO has been known for a long time, its specific antimicrobial effects and mechanism underlying these remain poorly characterized. The present study investigated the chemical composition of TTO and the dynamics and mechanism of its antimicrobial activities in two bacterial and two fungal strains. Gas chromatography-mass spectrometry analysis identified alkenes and alcohols as the main constituents of TTO. Terpinen-4-ol was the most abundant individual component, accounting for approximately 23 % of the TTO. Poisoned food technique assessment showed that the minimum inhibitory concentrations of TTO for bacterial strains (Escherichia coli and Staphylococcus aureus) and fungal strains (Candida albicans and Aspergillus niger) were 1.08 and 2.17 mg/mL, respectively. Antimicrobial dynamic curves showed that with increasing concentrations of TTO, the rate of cell killing and the duration of growth lag phase increased correspondingly. These data indicated that TTO produced concentration and time-dependent antimicrobial effects. The minimum bactericidal and fungicidal concentrations of TTO were 2.17, 4.34, and 4.34 against E. coli, S. aureus, and C. albicans, respectively. However, A. niger conidia were not completely eradicated, even after 3 days in the presence of 17.34 mg/mL TTO. Transmission electron microscopy images indicated that TTO penetrated the cell wall and cytoplasmic membrane of all the tested bacterial and fungal strains. TTO may also penetrate fungal organelle membrane. These findings indicated that TTO maybe exerts its antimicrobial effects by compromising the cell membrane, resulting in loss of the cytoplasm and organelle damage, which ultimate leads to cell death.

The Three Bacterial Lines of Defense against Antimicrobial Agents.

Antimicrobial agents target a range of extra- and/or intracellular loci from cytoplasmic wall to membrane, intracellular enzymes and genetic materials. Meanwhile, many resistance mechanisms employed by bacteria to counter antimicrobial agents have been found and reported in the past decades. Based on their spatially distinct sites of action and distribution of location, antimicrobial resistance mechanisms of bacteria were categorized into three groups, coined the three lines of bacterial defense in this review. The first line of defense is biofilms, which can be formed by most bacteria to overcome the action of antimicrobial agents. In addition, some other bacteria employ the second line of defense, the cell wall, cell membrane, and encased efflux pumps. When antimicrobial agents permeate the first two lines of defense and finally reach the cytoplasm, many bacteria will make use of the third line of defense, including alterations of intracellular materials and gene regulation to protect themselves from harm by bactericides. The presented three lines of defense theory will help us to understand the bacterial resistance mechanisms against antimicrobial agents and design efficient strategies to overcome these resistances.

Antifungal effect and mechanism of garlic oil on Penicillium funiculosum.

Garlic oil is a kind of fungicide, but little is known about its antifungal effects and mechanism. In this study, the chemical constituents, antifungal activity, and effects of garlic oil were studied with Penicillium funiculosum as a model strain. Results showed that the minimum fungicidal concentrations (MFCs, v/v) were 0.125 and 0.0313 % in agar medium and broth medium, respectively, suggesting that the garlic oil had a strong antifungal activity. The main ingredients of garlic oil were identified as sulfides, mainly including disulfides (36 %), trisulfides (32 %) and monosulfides (29 %) by gas chromatograph-mass spectrometer (GC/MS), which were estimated as the dominant antifungal factors. The observation results by transmission electron microscope (TEM) and scanning electron microscope (SEM) indicated that garlic oil could firstly penetrate into hyphae cells and even their organelles, and then destroy the cellular structure, finally leading to the leakage of both cytoplasm and macromolecules. Further proteomic analysis displayed garlic oil was able to induce a stimulated or weakened expression of some key proteins for physiological metabolism. Therefore, our study proved that garlic oil can work multiple sites of the hyphae of P. funiculosum to cause their death. The high antifungal effects of garlic oil makes it a broad application prospect in antifungal industries.

Efficacy of metal ions and isothiazolones in inhibiting Enterobacter cloacae BF-17 biofilm formation.

Enterobacter cloacae is a nosocomial pathogen. The E. cloacae strain BF-17, with a high capacity for biofilm formation, was screened and identified from industrially contaminated samples, carried out in our laboratory. To develop an efficient strategy to deal with biofilms, we investigated the effects of metal ions, including Na⁺, K⁺, Ca⁺, Mg⁺, Cu⁺, and Mn⁺, and 3 isothiazolones, on elimination of E. cloacae BF-17 biofilm formation by using a 0.1% crystal violet staining method. The results revealed that higher concentrations of Na⁺ or K⁺ significantly inhibited E. cloacae BF-17 biofilm development. Meanwhile, Ca²âº and Mn²âº stimulated biofilm formation at low concentration but exhibited a negative effect at high concentration. Moreover, biofilm formation decreased with increasing concentration of Mg²âº and Cu²âº. The isothiazolones Kathon (14%), 1,2-benzisothiazolin-3-one (11%), and 2-methyl-4-isothiazolin-3-one (10%) stimulated initial biofilm formation but not planktonic growth at low concentrations and displayed inhibitory effects on both biofilm formation and planktonic growth at higher concentrations. Unfortunately, the 3 isothiazolones exerted negligible effects on preformed or fully mature biofilms. Our findings suggest that Na⁺, K⁺, Mg²âº, and isothiazolones could be used to prevent and eliminate E. cloacae BF-17 biofilms.