scholarly journals Biofilm Development on Urinary Catheters Promotes the Appearance of Viable but Nonculturable Bacteria

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Sandra A. Wilks ◽  
Verena V. Koerfer ◽  
Jacqui A. Prieto ◽  
Mandy Fader ◽  
C. William Keevil
Keyword(s):  
Clinical Trials ◽  
Silver Alloy ◽  
Viable But Nonculturable ◽  
Content Type ◽  
Vbnc Bacteria ◽  
Staining Techniques ◽  
Tract Infections ◽  
Biofilm Development ◽  
Microscopy Techniques ◽  
The Impact

ABSTRACT Catheter-associated urinary tract infections have serious consequences, for both patients and health care resources. Much work has been carried out to develop an antimicrobial catheter. Although such developments have shown promise under laboratory conditions, none have demonstrated a clear advantage in clinical trials. Using a range of microbiological and advanced microscopy techniques, a detailed laboratory study comparing biofilm development on silicone, hydrogel latex, and silver alloy-coated hydrogel latex catheters was carried out. Biofilm development by Escherichia coli, Pseudomonas aeruginosa, and Proteus mirabilis on three commercially available catheters was tracked over time. Samples were examined with episcopic differential interference contrast (EDIC) microscopy, culture analysis, and staining techniques to quantify viable but nonculturable (VBNC) bacteria. Both qualitative and quantitative assessments found biofilms to develop rapidly on all three materials. EDIC microscopy revealed the rough surface topography of the materials. Differences between culture counts and quantification of total and dead cells demonstrated the presence of VBNC populations, where bacteria retain viability but are not metabolically active. The use of nonculture-based techniques showed the development of widespread VBNC populations. These VBNC populations were more evident on silver alloy-coated hydrogel latex catheters, indicating a bacteriostatic effect at best. The laboratory tests reported here, which detect VBNC bacteria, allow more rigorous assessment of antimicrobial catheters, explaining why there is often minimal benefit to patients. IMPORTANCE Several antimicrobial urinary catheter materials have been developed, but, although laboratory studies may show a benefit, none have significantly improved clinical outcomes. The use of poorly designed laboratory testing and lack of consideration of the impact of VBNC populations may be responsible. While the presence of VBNC populations is becoming more widely reported, there remains a lack of understanding of the clinical impact or influence of exposure to antimicrobial products. This is the first study to investigate the impact of antimicrobial surface materials and the appearance of VBNC populations. This demonstrates how improved testing is needed before clinical trials are initiated.

scholarly journals Biofilm development on urinary catheters promotes the appearance of viable but non-culturable (VBNC) bacteria

2020 ◽  
Author(s):  
Sandra A. Wilks ◽  
Verena V. Koerfer ◽  
Jacqui A. Prieto ◽  
Mandy Fader ◽  
C. William Keevil
Keyword(s):  
Clinical Trials ◽  
Bacteriostatic Effect ◽  
Silver Alloy ◽  
Silicone Hydrogel ◽  
Vbnc Bacteria ◽  
Staining Techniques ◽  
Tract Infections ◽  
Biofilm Development ◽  
Microscopy Techniques ◽  
The Impact

ABSTRACTCatheter-associated urinary tract infections have serious consequences, both for patients and in impacting on healthcare resources. Much work has been carried out to develop an antimicrobial catheter. Although such developments have shown promise under laboratory conditions, none have demonstrated a clear advantage in clinical trials.Using a range of microbiological and advanced microscopy techniques, a detailed laboratory study comparing biofilm development on silicone, hydrogel latex and silver alloy coated hydrogel latex catheters was carried out. Biofilm development by Escherichia coli, Pseudomonas aeruginosa and Proteus mirabilis on three commercially available catheters was tracked over time. Samples were examined with episcopic differential interference contrast (EDIC) microscopy, culture analysis and staining techniques to quantify viable but non-culturable (VBNC) bacteria.Both qualitative and quantitative assessment found biofilms to develop rapidly on all three materials. EDIC microscopy revealed the rough surface topography of the materials. Differences between culture counts and quantification of total and dead cells demonstrated the presence of VBNC populations, where bacteria retain viability but are not metabolically active.The use of non-culture based techniques showed the development of widespread VBNC populations. These VBNC populations were more evident on silver alloy coated hydrogel latex catheters, indicating a bacteriostatic effect at best. The laboratory tests reported here, that detect VBNC bacteria, allow more rigorous assessment of antimicrobial catheters offering an explanation for why there is often minimal benefit to patients.IMPORTANCESeveral antimicrobial urinary catheter materials have been developed but, although laboratory studies may show a benefit, none have significantly improved clinical outcomes. The use of poorly designed laboratory testing and lack of consideration to the impact of VBNC populations may be responsible. While the presence of VBNC populations is becoming more widely reported, there remains a lack of understanding of the clinical impact or influence of exposure to antimicrobial products. This is the first study to investigate the impact of antimicrobial surface materials and the appearance of VBNC populations. This demonstrates how improved testing is needed prior to clinical trials uptake.

scholarly journals Light-Mediated Decreases in Cyclic di-GMP Levels Inhibit Structure Formation in Pseudomonas aeruginosa Biofilms

2020 ◽  
Vol 202 (14) ◽  
Author(s):  
Lisa Juliane Kahl ◽  
Alexa Price-Whelan ◽  
Lars E. P. Dietrich
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Infections ◽  
Prolonged Exposure ◽  
Light Exposure ◽  
Research Attention ◽  
Content Type ◽  
Dependent Inhibition ◽  
Matrix Production ◽  
Biofilm Development ◽  
The Impact

ABSTRACT Light is known to trigger regulatory responses in diverse organisms, including slime molds, animals, plants, and phototrophic bacteria. However, light-dependent processes in nonphototrophic bacteria, and those of pathogens in particular, have received comparatively little research attention. In this study, we examined the impact of light on multicellular development in Pseudomonas aeruginosa, a leading cause of biofilm-based bacterial infections. We grew P. aeruginosa strain PA14 in a colony morphology assay and found that growth under prolonged exposure to low-intensity blue light inhibited biofilm matrix production and thereby the formation of vertical biofilm structures (i.e., “wrinkles”). Light-dependent inhibition of biofilm wrinkling was correlated with low levels of cyclic di-GMP (c-di-GMP), consistent with the role of this signal in stimulating matrix production. A screen of enzymes with the potential to catalyze c-di-GMP synthesis or degradation identified c-di-GMP phosphodiesterases that contribute to light-dependent inhibition of biofilm wrinkling. One of these, RmcA, was previously characterized by our group for its role in mediating the effect of redox-active P. aeruginosa metabolites called phenazines on biofilm wrinkle formation. Our results suggest that an RmcA sensory domain that is predicted to bind a flavin cofactor is involved in light-dependent inhibition of wrinkling. Together, these findings indicate that P. aeruginosa integrates information about light exposure and redox state in its regulation of biofilm development. IMPORTANCE Light exposure tunes circadian rhythms, which modulate the immune response and affect susceptibility to infection in plants and animals. Though molecular responses to light are defined for model plant and animal hosts, analogous pathways that function in bacterial pathogens are understudied. We examined the response to light exposure in biofilms (matrix-encased multicellular assemblages) of the nonphotosynthetic bacterium Pseudomonas aeruginosa. We found that light at intensities that are not harmful to human cells inhibited biofilm maturation via effects on cellular signals. Because biofilm formation is a critical factor in many types of P. aeruginosa infections, including burn wound infections that may be exposed to light, these effects could be relevant for pathogenicity.

scholarly journals Bacteriophage-Mediated Control of a Two-Species Biofilm Formed by Microorganisms Causing Catheter-Associated Urinary Tract Infections in anIn VitroUrinary Catheter Model

2014 ◽  
Vol 59 (2) ◽  
pp. 1127-1137 ◽  
Author(s):  
Susan M. Lehman ◽  
Rodney M. Donlan
Keyword(s):  
Mixed Species ◽  
Biofilm Inhibition ◽  
Biofilm Growth ◽  
Content Type ◽  
Patient Morbidity ◽  
Phage Cocktail ◽  
Artificial Urine ◽  
Tract Infections ◽  
Biofilm Development

ABSTRACTMicroorganisms from a patient or their environment may colonize indwelling urinary catheters, forming biofilm communities on catheter surfaces and increasing patient morbidity and mortality. This study investigated the effect of pretreating hydrogel-coated silicone catheters with mixtures ofPseudomonas aeruginosaandProteus mirabilisbacteriophages on the development of single- and two-species biofilms in a multiday continuous-flowin vitromodel using artificial urine. Novel phages were purified from sewage, characterized, and screened for their abilities to reduce biofilm development by clinical isolates of their respective hosts. Our screening data showed that artificial urine medium (AUM) is a valid substitute for human urine for the purpose of evaluating uropathogen biofilm control by these bacteriophages. Defined phage cocktails targetingP. aeruginosaandP. mirabiliswere designed based on the biofilm inhibition screens. Hydrogel-coated catheters were pretreated with one or both cocktails and challenged with approximately 1 × 103CFU/ml of the corresponding pathogen(s). The biofilm growth on the catheter surfaces in AUM was monitored over 72 to 96 h. Phage pretreatment reducedP. aeruginosabiofilm counts by 4 log10CFU/cm2(P≤ 0.01) andP. mirabilisbiofilm counts by >2 log10CFU/cm2(P≤ 0.01) over 48 h. The presence ofP. mirabiliswas always associated with an increase in lumen pH from 7.5 to 9.5 and with eventual blockage of the reactor lines. The results of this study suggest that pretreatment of a hydrogel urinary catheter with a phage cocktail can significantly reduce mixed-species biofilm formation by clinically relevant bacteria.

scholarly journals Privatization of Biofilm Matrix in Structurally Heterogeneous Biofilms

mSystems ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Simon B. Otto ◽  
Marivic Martin ◽  
Daniel Schäfer ◽  
Raimo Hartmann ◽  
Knut Drescher ◽  
...  
Keyword(s):  
Gene Expression ◽  
Shear Stress ◽  
Content Type ◽  
Matrix Gene ◽  
Starting Point ◽  
The Matrix ◽  
Matrix Components ◽  
Biofilm Development ◽  
The Impact

ABSTRACT The self-produced biofilm provides beneficial protection for the enclosed cells, but the costly production of matrix components makes producer cells susceptible to cheating by nonproducing individuals. Despite detrimental effects of nonproducers, biofilms can be heterogeneous, with isogenic nonproducers being a natural consequence of phenotypic differentiation processes. For instance, in Bacillus subtilis biofilm cells differ in production of the two major matrix components, the amyloid fiber protein TasA and exopolysaccharides (EPS), demonstrating different expression levels of corresponding matrix genes. This raises questions regarding matrix gene expression dynamics during biofilm development and the impact of phenotypic nonproducers on biofilm robustness. Here, we show that biofilms are structurally heterogeneous and can be separated into strongly and weakly associated clusters. We reveal that spatiotemporal changes in structural heterogeneity correlate with matrix gene expression, with TasA playing a key role in biofilm integrity and timing of development. We show that the matrix remains partially privatized by the producer subpopulation, where cells tightly stick together even when exposed to shear stress. Our results support previous findings on the existence of “weak points” in seemingly robust biofilms as well as on the key role of linkage proteins in biofilm formation. Furthermore, we provide a starting point for investigating the privatization of common goods within isogenic populations. IMPORTANCE Biofilms are communities of bacteria protected by a self-produced extracellular matrix. The detrimental effects of nonproducing individuals on biofilm development raise questions about the dynamics between community members, especially when isogenic nonproducers exist within wild-type populations. We asked ourselves whether phenotypic nonproducers impact biofilm robustness, and where and when this heterogeneity of matrix gene expression occurs. Based on our results, we propose that the matrix remains partly privatized by the producing subpopulation, since producing cells stick together when exposed to shear stress. The important role of linkage proteins in robustness and development of the structurally heterogeneous biofilm provides an entry into studying the privatization of common goods within isogenic populations.

scholarly journals Cross Talk between MarR-Like Transcription Factors Coordinates the Regulation of Motility in UropathogenicEscherichia coli

2018 ◽  
Vol 86 (12) ◽  
Author(s):  
Courtney L. Luterbach ◽  
Harry L. T. Mobley
Keyword(s):  
Urinary Tract ◽  
Cross Talk ◽  
Flagellar Gene ◽  
Upec Strain ◽  
Content Type ◽  
Dose Effects ◽  
Regulator Genes ◽  
Strain Cft073 ◽  
Tract Infections ◽  
The Impact

ABSTRACTThe MarR-like protein PapX represses the transcription of the flagellar master regulator genesflhDCin uropathogenicEscherichia coli(UPEC), the primary cause of uncomplicated urinary tract infections (UTIs). PapX is encoded by thepapoperon, which also encodes the adherence factors termed P fimbriae. Both adherence and motility are critical for productive colonization of the urinary tract. However, the mechanisms involved in coordinating the transition between adherence and motility are not well characterized. UPEC strain CFT073 carries bothpapXand a homolog,focX, located in thefocoperon encoding F1C fimbriae. In this study, we characterized the dose effects of “X” genes on flagellar gene expression and cross talk betweenfocXandpapX. We found that both FocX and PapX repressflhDtranscription. However, we determined that the ΔpapXmutant was hypermotile, while the loss offocXdid not affect motility. We further investigated this phenotype and found that FocX functions as a repressor ofpapX. Additionally, we identified a proximal independent promoter upstream of bothfocXandpapXand assessed the expression offocXandpapXduring culture in human urine and on LB agar plates compared to LB medium. Finally, we characterized the contributions of PapX and FocX to fitness in the ascending murine model of UTI and observed a subtle, but not statistically significant, fitness defect in colonization of the kidneys. Altogether, these results expand our understanding of the impact of carrying multiple X genes on the coordinated regulation of motility and adherence in UPEC.

scholarly journals Importance of Pyruvate Sensing and Transport for the Resuscitation of Viable but NonculturableEscherichia coliK-12

2018 ◽  
Vol 201 (3) ◽  
Author(s):  
Cláudia Vilhena ◽  
Eugen Kaganovitch ◽  
Alexander Grünberger ◽  
Magdalena Motz ◽  
Ignasi Forné ◽  
...  
Keyword(s):  
Response Regulator ◽  
Bacterial Species ◽  
Microfluidic System ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Content Type ◽  
E Coli ◽  
High Affinity ◽  
Vbnc Bacteria ◽  
K 12

ABSTRACTEscherichia coliand many other bacterial species can enter into a viable but nonculturable (VBNC) state, which is a survival strategy adopted by cells exposed to adverse environmental conditions. Pyruvate is known to be one factor that promotes resuscitation of VBNC cells. Here we studied the role of a pyruvate-sensing network, composed of the histidine kinase-response regulator systems BtsS/BtsR and YpdA/YpdB and the target genebtsT, encoding the high-affinity pyruvate/H+symporter BtsT, in the resuscitation of VBNCE. coliK-12 cells after exposure to cold for 120 days. Analysis of the proteome of VBNC cells revealed upregulation, relative to exponentially growing cells, of BtsT and other proteins involved in pyruvate metabolism. Provision of pyruvate stimulated protein and DNA biosynthesis, and thus resuscitation, in wild-type but notbtsSR ypdABmutant VBNC cells. This result was corroborated by time-dependent tracking of the resuscitation of individual VBNCE. colicells observed in a microfluidic system. Finally, transport assays revealed that14C-labeled pyruvate was rapidly taken up into VBNC cells by BtsT. These results provide the first evidence that pyruvate is taken up as a carbon source for the resuscitation of VBNCE. colicells.IMPORTANCEViable but nonculturable (VBNC) bacteria do not form colonies in standard medium but otherwise retain their metabolic activity and can express toxic proteins. Many bacterial genera, includingEscherichia,Vibrio, andListeria, have been shown to enter the VBNC state upon exposure to adverse conditions, such as low temperature, radiation, and starvation. Ultimately, these organisms pose a public health risk with potential implications for the pharmaceutical and food industries, as dormant organisms are especially difficult to selectively eliminate and VBNC bacteria can be resuscitated if placed in an environment with appropriate nutrition and temperature. Here we used a microfluidic system to monitor the resuscitation of single VBNC cells over time. We provide new molecular insights into the initiation of resuscitation by demonstrating that VBNCE. colicells rapidly take up pyruvate with an inducible high-affinity transporter, whose expression is triggered by the BtsSR-YpdAB sensing network.

scholarly journals Impact of the MIC of Piperacillin-Tazobactam on the Outcome of Patients with Bacteremia Due to Extended-Spectrum-β-Lactamase-Producing Escherichia coli

2013 ◽  
Vol 57 (7) ◽  
pp. 3402-3404 ◽  
Author(s):  
Pilar Retamar ◽  
Lorena López-Cerero ◽  
Miguel Angel Muniain ◽  
Álvaro Pascual ◽  
Jesús Rodríguez-Baño ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Urinary Tract Infections ◽  
Bloodstream Infections ◽  
Content Type ◽  
Extended Spectrum ◽  
Tract Infections ◽  
The Impact

ABSTRACTWe investigated the impact of the piperacillin-tazobactam MIC in the outcome of 39 bloodstream infections due to extended-spectrum-β-lactamase-producingEscherichia coli. All 11 patients with urinary tract infections survived, irrespective of the MIC. For other sources, 30-day mortality was lower for isolates with a MIC of ≤2 mg/liter than for isolates with a higher MIC (0% versus 41.1%;P= 0.02).

scholarly journals Stress Resistance and Pathogenicity of Nonthermal-Plasma-Induced Viable-but-Nonculturable Staphylococcus aureus through Energy Suppression, Oxidative Stress Defense, and Immune-Escape Mechanisms

2020 ◽  
Vol 87 (2) ◽  
Author(s):  
Xinyu Liao ◽  
Weicheng Hu ◽  
Donghong Liu ◽  
Tian Ding
Keyword(s):  
Oxidative Stress ◽  
Stress Resistance ◽  
Immune Escape ◽  
Nonthermal Plasma ◽  
Host Cells ◽  
Viable But Nonculturable ◽  
Content Type ◽  
Oxidative Stress Defense ◽  
Vbnc Bacteria ◽  
Potential Risks

ABSTRACT The occurrence of viable-but-nonculturable (VBNC) bacteria poses a potential risk to food safety due to failure in conventional colony detection. In this study, induction of VBNC Staphylococcus aureus was conducted by exposure to an atmospheric-pressure air dielectric barrier discharge-nonthermal-plasma (DBD-NTP) treatment with an applied energy of 8.1 kJ. The stress resistance profiles and pathogenicity of VBNC S. aureus were further evaluated. We found that VBNC S. aureus showed levels of tolerance of heat, acid, and osmosis challenges comparable to those shown by culturable S. aureus, while VBNC S. aureus exhibited enhanced resistance to oxidative and antibiotic stress, relating to the mechanisms of cellular energy depletion, antioxidant response initiation, and multidrug efflux pump upregulation. Regarding pathogenicity, NTP-induced VBNC S. aureus retained the capacity to infect the HeLa host cells. Compared with the culturable counterparts, VBNC S. aureus caused reduced immune responses (Toll-like receptor [TLR], nucleotide-binding oligomerization domain [NOD]) in HeLa cells, which was attributed to suppression of biosynthesis of the recognized surface ligands (e.g., peptidoglycan). Additionally, the proteomic analysis revealed that upregulation of several virulence factors (ClfB, SdrD, SCIN, SasH, etc.) could ensure that VBNC S. aureus would adhere to and internalize into host cells and avoid the host attack. The camouflaged mechanisms described above led to VBNC S. aureus causing less damage to the host cells, and their activity might result in longer intracellular persistence, posing potential risks during NTP processing. IMPORTANCE The consumer demand for freshness and nutrition has accelerated the development of mild decontamination technologies. The incomplete killing of nonthermal (NT) treatments might induce pathogens to enter into a viable-but-nonculturable (VBNC) status as a survival strategy. The use of nonthermal plasma (NTP) as a novel food decontamination technology received increased attention in food industry during recent decades. Our previous work confirmed that the foodborne pathogen S. aureus was induced into VBNC status in response to NTP exposure. This work further revealed the development of stress resistance and virulence retention of NTP-induced VBNC S. aureus through the mechanisms of energy suppression, oxidative stress defense, and immune escape. The data provide fundamental knowledge of the potential risks posed by NTP-induced VBNC S. aureus, which require further parameter optimization of the NTP process or combination with other techniques to avoid the occurrence of VBNC bacteria.

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