Flagella-Mediated Adhesion and Extracellular DNA Release Contribute to Biofilm Formation and Stress Tolerance of Campylobacter jejuni

ABSTRACTSubminimal inhibitory concentrations of antibiotics have been shown to induce bacterial biofilm formation. Few studies have investigated antibiotic-induced biofilm formation inStaphylococcus aureus, an important human pathogen. Our goal was to measureS. aureusbiofilm formation in the presence of low levels of β-lactam antibiotics. Fifteen phylogenetically diverse methicillin-resistantStaphylococcus aureus(MRSA) and methicillin-sensitiveS. aureus(MSSA) strains were employed. Methicillin, ampicillin, amoxicillin, and cloxacillin were added to cultures at concentrations ranging from 0× to 1× MIC. Biofilm formation was measured in 96-well microtiter plates using a crystal violet binding assay. Autoaggregation was measured using a visual test tube settling assay. Extracellular DNA was quantitated using agarose gel electrophoresis. All four antibiotics induced biofilm formation in some strains. The amount of biofilm induction was as high as 10-fold and was inversely proportional to the amount of biofilm produced by the strain in the absence of antibiotics. MRSA strains of lineages USA300, USA400, and USA500 exhibited the highest levels of methicillin-induced biofilm induction. Biofilm formation induced by low-level methicillin was inhibited by DNase. Low-level methicillin also induced DNase-sensitive autoaggregation and extracellular DNA release. The biofilm induction phenotype was absent in a strain deficient in autolysin (atl). Our findings demonstrate that subminimal inhibitory concentrations of β-lactam antibiotics significantly induce autolysin-dependent extracellular DNA release and biofilm formation in some strains ofS. aureus.IMPORTANCEThe widespread use of antibiotics as growth promoters in agriculture may expose bacteria to low levels of the drugs. The aim of this study was to investigate the effects of low levels of antibiotics on bacterial autoaggregation and biofilm formation, two processes that have been shown to foster genetic exchange and antibiotic resistance. We found that low levels of β-lactam antibiotics, a class commonly used in both clinical and agricultural settings, caused significant autoaggregation and biofilm formation by the important human pathogenStaphylococcus aureus. Both processes were dependent on cell lysis and release of DNA into the environment. The effect was most pronounced among multidrug-resistant strains known as methicillin-resistantS. aureus(MRSA). These results may shed light on the recalcitrance of some bacterial infections to antibiotic treatment in clinical settings and the evolution of antibiotic-resistant bacteria in agricultural settings.

Download Full-text

Environmental Stress-Induced Bacterial Lysis and Extracellular DNA Release Contribute toCampylobacter jejuniBiofilm Formation

Applied and Environmental Microbiology ◽

10.1128/aem.02068-17 ◽

2017 ◽

Vol 84 (5) ◽

Cited By ~ 9

Author(s):

Jinsong Feng ◽

Lina Ma ◽

Jiatong Nie ◽

Michael E. Konkel ◽

Xiaonan Lu

Keyword(s):

Environmental Stress ◽

Campylobacter Jejuni ◽

Biofilm Formation ◽

Extracellular Dna ◽

Bacterial Cells ◽

Starvation Stress ◽

Content Type ◽

Bacterial Lysis ◽

Major Form ◽

Spatial Locations

ABSTRACTCampylobacter jejuniis a microaerophilic bacterium and is believed to persist in a biofilm to antagonize environmental stress. This study investigated the influence of environmental conditions on the formation ofC. jejunibiofilm. We report an extracellular DNA (eDNA)-mediated mechanism of biofilm formation in response to aerobic and starvation stress. The eDNA was determined to represent a major form of constitutional material ofC. jejunibiofilms and to be closely associated with bacterial lysis. Deletion mutation of the stress response genesspoTandrecAenhanced the aerobic influence by stimulating lysis and increasing eDNA release. Flagella were also involved in biofilm formation but mainly contributed to attachment rather than induction of lysis. The addition of genomic DNA from eitherCampylobacterorSalmonellaresulted in a concentration-dependent stimulation effect on biofilm formation, but the effect was not due to forming a precoating DNA layer. Enzymatic degradation of DNA by DNase I disruptedC. jejunibiofilm. In a dual-species biofilm, eDNA allocatedCampylobacterandSalmonellaat distinct spatial locations that protectCampylobacterfrom oxygen stress. Our findings demonstrated an essential role and multiple functions of eDNA in biofilm formation ofC. jejuni, including facilitating initial attachment, establishing and maintaining biofilm, and allocating bacterial cells.IMPORTANCECampylobacter jejuniis a major cause of foodborne illness worldwide. In the natural environment, the growth ofC. jejuniis greatly inhibited by various forms of environmental stress, such as aerobic stress and starvation stress. Biofilm formation can facilitate the distribution ofC. jejuniby enabling the survival of this fragile microorganism under unfavorable conditions. However, the mechanism ofC. jejunibiofilm formation in response to environmental stress has been investigated only partially. The significance of our research is in identifying extracellular DNA released by bacterial lysis as a major form of constitution material that mediates the formation ofC. jejunibiofilm in response to environmental stress, which enhances our understanding of the formation mechanism ofC. jejunibiofilm. This knowledge can aid the development of intervention strategies to limit the distribution ofC. jejuni.

Download Full-text

Important Contribution of the Novel LocuscomEBto Extracellular DNA-Dependent Staphylococcus lugdunensis Biofilm Formation

Infection and Immunity ◽

10.1128/iai.00775-15 ◽

2015 ◽

Vol 83 (12) ◽

pp. 4682-4692 ◽

Cited By ~ 12

Author(s):

Nithya Babu Rajendran ◽

Julian Eikmeier ◽

Karsten Becker ◽

Muzaffar Hussain ◽

Georg Peters ◽

...

Keyword(s):

Biofilm Formation ◽

Bacterial Species ◽

Laser Scanning Microscopy ◽

Extracellular Dna ◽

Staphylococcus Lugdunensis ◽

Content Type ◽

Competence Gene ◽

Independent Mechanism ◽

Dna Release ◽

The Impact

The coagulase-negative speciesStaphylococcus lugdunensisis an emerging cause of serious and potentially life-threatening infections, such as infective endocarditis. The pathogenesis of these infections is characterized by the ability ofS. lugdunensisto form biofilms on either biotic or abiotic surfaces. To elucidate the genetic basis of biofilm formation inS. lugdunensis, we performed transposon (Tn917) mutagenesis. One mutant had a significantly reduced biofilm-forming capacity and carried a Tn917insertion within the competence genecomEB. Site-directed mutagenesis and subsequent complementation with a functional copy ofcomEBverified the importance ofcomEBin biofilm formation. In several bacterial species, natural competence stimulates DNA release via lysis-dependent or -independent mechanisms. Extracellular DNA (eDNA) has been demonstrated to be an important structural component of many bacterial biofilms. Therefore, we quantified the eDNA in the biofilms and found diminished eDNA amounts in thecomEBmutant biofilm. High-resolution images and three-dimensional data obtained via confocal laser scanning microscopy (CSLM) visualized the impact of thecomEBmutation on biofilm integrity. ThecomEBmutant did not show reduced expression of autolysin genes, decreased autolytic activities, or increased cell viability, suggesting a cell lysis-independent mechanism of DNA release. Furthermore, reduced amounts of eDNA in thecomEBmutant biofilms did not result from elevated levels or activity of theS. lugdunensisthermonuclease NucI. In conclusion, we defined here, for the first time, a role for the competence genecomEBin staphylococcal biofilm formation. Our findings indicate thatcomEBstimulates biofilm formation via a lysis-independent mechanism of DNA release.

Download Full-text

Effect of nicotine on Staphylococcus aureus biofilm formation and virulence factors

Scientific Reports ◽

10.1038/s41598-019-56627-0 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Le Shi ◽

Yang Wu ◽

Chen Yang ◽

Yue Ma ◽

Qing-zhao Zhang ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Virulence Factors ◽

Biofilm Formation ◽

Alveolar Epithelial Cells ◽

Extracellular Dna ◽

Bacterial Invasion ◽

Initial Attachment ◽

Accessory Gene ◽

Nicotine Treatment ◽

Dna Release

AbstractStaphylococcus aureus is a common pathogen in chronic rhinosinusitis (CRS) patients, the pathogenesis of which involves the ability to form biofilms and produce various virulence factors. Tobacco smoke, another risk factor of CRS, facilitates S. aureus biofilm formation; however, the mechanisms involved are unclear. Here, we studied the effect of nicotine on S. aureus biofilm formation and the expression of virulence-related genes. S. aureus strains isolated from CRS patients and a USA300 strain were treated with nicotine or were untreated (control). Nicotine-treated S. aureus strains showed dose-dependent increases in biofilm formation, lower virulence, enhanced initial attachment, increased extracellular DNA release, and a higher autolysis rate, involving dysregulation of the accessory gene regulator (Agr) quorum-sensing system. Consequently, the expression of autolysis-related genes lytN and atlA, and the percentage of dead cells in biofilms was increased. However, the expression of virulence-related genes, including hla, hlb, pvl, nuc, ssp, spa, sigB, coa, and crtN was downregulated and there was reduced bacterial invasion of A549 human alveolar epithelial cells. The results of this study indicate that nicotine treatment enhances S. aureus biofilm formation by promoting initial attachment and extracellular DNA release but inhibits the virulence of this bacterium.

Download Full-text

Extracellular DNA release confers heterogeneity in Candida albicans biofilm formation

BMC Microbiology ◽

10.1186/s12866-014-0303-6 ◽

2014 ◽

Vol 14 (1) ◽

Cited By ~ 28

Author(s):

Ranjith Rajendran ◽

Leighann Sherry ◽

David F Lappin ◽

Chris J Nile ◽

Karen Smith ◽

...

Keyword(s):

Candida Albicans ◽

Biofilm Formation ◽

Extracellular Dna ◽

Dna Release

Download Full-text

Faculty Opinions recommendation of Low levels of β-lactam antibiotics induce extracellular DNA release and biofilm formation in Staphylococcus aureus.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.717952792.793458322 ◽

2012 ◽

Author(s):

Marvin Whiteley ◽

Holly Huse

Keyword(s):

Staphylococcus Aureus ◽

Biofilm Formation ◽

Extracellular Dna ◽

Dna Release ◽

Low Levels

Download Full-text

PspC domain-containing protein (PCP) determines Streptococcus mutans biofilm formation through bacterial extracellular DNA release and platelet adhesion in experimental endocarditis

PLoS Pathogens ◽

10.1371/journal.ppat.1009289 ◽

2021 ◽

Vol 17 (2) ◽

pp. e1009289

Author(s):

Chiau-Jing Jung ◽

Chih-Chieh Hsu ◽

Jeng-Wei Chen ◽

Hung-Wei Cheng ◽

Chang-Tsu Yuan ◽

...

Keyword(s):

Streptococcus Mutans ◽

Biofilm Formation ◽

Heart Valves ◽

Platelet Adhesion ◽

Bacterial Biofilm ◽

Extracellular Dna ◽

Dependent Manner ◽

Dna Release

Bacterial extracellular DNA (eDNA) and activated platelets have been found to contribute to biofilm formation by Streptococcus mutans on injured heart valves to induce infective endocarditis (IE), yet the bacterial component directly responsible for biofilm formation or platelet adhesion remains unclear. Using in vivo survival assays coupled with microarray analysis, the present study identified a LiaR-regulated PspC domain-containing protein (PCP) in S. mutans that mediates bacterial biofilm formation in vivo. Reverse transcriptase- and chromatin immunoprecipitation-polymerase chain reaction assays confirmed the regulation of pcp by LiaR, while PCP is well-preserved among streptococcal pathogens. Deficiency of pcp reduced in vitro and in vivo biofilm formation and released the eDNA inside bacteria floe along with reduced bacterial platelet adhesion capacity in a fibrinogen-dependent manner. Therefore, LiaR-regulated PCP alone could determine release of bacterial eDNA and binding to platelets, thus contributing to biofilm formation in S. mutans-induced IE.

Download Full-text