Accumulation-Associated Protein Enhances Staphylococcus epidermidis Biofilm Formation under Dynamic Conditions and Is Required for Infection in a Rat Catheter Model

Biofilm formation is the primary virulence factor ofStaphylococcus epidermidis.S. epidermidisbiofilms preferentially form on abiotic surfaces and may contain multiple matrix components, including proteins such as accumulation-associated protein (Aap). Following proteolytic cleavage of the A domain, which has been shown to enhance binding to host cells, B domain homotypic interactions support cell accumulation and biofilm formation. To further define the contribution of Aap to biofilm formation and infection, we constructed anaapallelic replacement mutant and anicaADBC aapdouble mutant. When subjected to fluid shear, strains deficient in Aap production produced significantly less biofilm than Aap-positive strains. To examine thein vivorelevance of our findings, we modified our previously described rat jugular catheter model and validated the importance of immunosuppression and the presence of a foreign body to the establishment of infection. The use of our allelic replacement mutants in the model revealed a significant decrease in bacterial recovery from the catheter and the blood in the absence of Aap, regardless of the production of polysaccharide intercellular adhesin (PIA), a well-characterized, robust matrix molecule. Complementation of theaapmutant with full-length Aap (containing the A domain), but not the B domain alone, increased initial attachment to microtiter plates, as did intransexpression of the A domain in adhesion-deficientStaphylococcus carnosus. These results demonstrate Aap contributes toS. epidermidisinfection, which may in part be due to A domain-mediated attachment to abiotic surfaces.

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A Single B-Repeat of Staphylococcus epidermidis Accumulation-Associated Protein Induces Protective Immune Responses in an Experimental Biomaterial-Associated Infection Mouse Model

Clinical and Vaccine Immunology ◽

10.1128/cvi.00306-14 ◽

2014 ◽

Vol 21 (9) ◽

pp. 1206-1214 ◽

Cited By ~ 12

Author(s):

Lin Yan ◽

Lei Zhang ◽

Hongyan Ma ◽

David Chiu ◽

James D. Bryers

Keyword(s):

Staphylococcus Epidermidis ◽

Biofilm Formation ◽

Porous Scaffold ◽

The United States ◽

Protein Vaccine ◽

Weight Changes ◽

Biofilm Inhibition ◽

Content Type ◽

Accumulation Associated Protein

ABSTRACTNosocomial infections are the fourth leading cause of morbidity and mortality in the United States, resulting in 2 million infections and ∼100,000 deaths each year. More than 60% of these infections are associated with some type of biomedical device.Staphylococcus epidermidisis a commensal bacterium of the human skin and is the most common nosocomial pathogen infecting implanted medical devices, especially those in the cardiovasculature.S. epidermidisantibiotic resistance and biofilm formation on inert surfaces make these infections hard to treat. Accumulation-associated protein (Aap), a cell wall-anchored protein ofS. epidermidis, is considered one of the most important proteins involved in the formation ofS. epidermidisbiofilm. A small recombinant protein vaccine comprising a single B-repeat domain (Brpt1.0) ofS. epidermidisRP62A Aap was developed, and the vaccine's efficacy was evaluatedin vitrowith a biofilm inhibition assay andin vivoin a murine model of biomaterial-associated infection. A high IgG antibody response againstS. epidermidisRP62A was detected in the sera of the mice after two subcutaneous immunizations with Brpt1.0 coadministered with Freund's adjuvant. Sera from Brpt1.0-immunized mice inhibitedin vitroS. epidermidisRP62A biofilm formation in a dose-dependent pattern. After receiving two immunizations, each mouse was surgically implanted with a porous scaffold disk containing 5 × 106CFU ofS. epidermidisRP62A. Weight changes, inflammatory markers, and histological assay results after challenge withS. epidermidisindicated that the mice immunized with Brpt1.0 exhibited significantly higher resistance toS. epidermidisRP62A implant infection than the control mice. Day 8 postchallenge, there was a significantly lower number of bacteria in scaffold sections and surrounding tissues and a lower residual inflammatory response to the infected scaffold disks for the Brpt1.0-immunized mice than for of the ovalbumin (Ova)-immunized mice.

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Oral Administration of the Broad-Spectrum Antibiofilm Compound Toremifene Inhibits Candida albicans and Staphylococcus aureus Biofilm FormationIn Vivo

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03869-14 ◽

2014 ◽

Vol 58 (12) ◽

pp. 7606-7610 ◽

Cited By ~ 16

Author(s):

Kaat De Cremer ◽

Nicolas Delattin ◽

Katrijn De Brucker ◽

Annelies Peeters ◽

Soña Kucharíková ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Candida Albicans ◽

Broad Spectrum ◽

Staphylococcus Epidermidis ◽

Biofilm Formation ◽

Candida Krusei ◽

Content Type ◽

And Staphylococcus Aureus

ABSTRACTWe here report on thein vitroactivity of toremifene to inhibit biofilm formation of different fungal and bacterial pathogens, includingCandida albicans,Candida glabrata,Candida dubliniensis,Candida krusei,Pseudomonas aeruginosa,Staphylococcus aureus, andStaphylococcus epidermidis. We validated thein vivoefficacy of orally administered toremifene againstC. albicans and S. aureusbiofilm formation in a rat subcutaneous catheter model. Combined, our results demonstrate the potential of toremifene as a broad-spectrum oral antibiofilm compound.

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Antibacterial Effects of Phage Lysin LysGH15 on Planktonic Cells and Biofilms of Diverse Staphylococci

Applied and Environmental Microbiology ◽

10.1128/aem.00886-18 ◽

2018 ◽

Vol 84 (15) ◽

Cited By ~ 9

Author(s):

Yufeng Zhang ◽

Mengjun Cheng ◽

Hao Zhang ◽

Jiaxin Dai ◽

Zhimin Guo ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Staphylococcus Epidermidis ◽

Biofilm Formation ◽

Multidrug Resistant ◽

Planktonic Cells ◽

Content Type ◽

Resistant Strains ◽

Staphylococcus Hominis ◽

Phage Lysin

ABSTRACT Treatment of infections caused by staphylococci has become more difficult because of the emergence of multidrug-resistant strains as well as biofilm formation. In this study, we observed the ability of the phage lysin LysGH15 to eliminate staphylococcal planktonic cells and biofilms formed by Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, and Staphylococcus hominis. All these strains were sensitive to LysGH15, showing reductions in bacterial counts of approximately 4 log units within 30 min after treatment with 20 μg/ml of LysGH15, and the MICs ranged from 8 μg/ml to 32 μg/ml. LysGH15 efficiently prevented biofilm formation by the four staphylococcal species at a dose of 50 μg/ml. At a higher dose (100 μg/ml), LysGH15 also showed notable disrupting activity against 24-h and 72-h biofilms formed by S. aureus and coagulase-negative species. In the in vivo experiments, a single intraperitoneal injection of LysGH15 (20 μg/mouse) administered 1 h after the injection of S. epidermidis at double the minimum lethal dose was sufficient to protect the mice. The S. epidermidis cell counts were 4 log units lower in the blood and 3 log units lower in the organs of mice 24 h after treatment with LysGH15 than in the untreated control mice. LysGH15 reduced cytokine levels in the blood and improved pathological changes in the organs. The broad antistaphylococcal activity exerted by LysGH15 on planktonic cells and biofilms makes LysGH15 a valuable treatment option for biofilm-related or non-biofilm-related staphylococcal infections. IMPORTANCE Most staphylococcal species are major causes of health care- and community-associated infections. In particular, Staphylococcus aureus is a common and dangerous pathogen, and Staphylococcus epidermidis is a ubiquitous skin commensal and opportunistic pathogen. Treatment of infections caused by staphylococci has become more difficult because of the emergence of multidrug-resistant strains as well as biofilm formation. In this study, we found that all tested S. aureus, S. epidermidis, Staphylococcus haemolyticus, and Staphylococcus hominis strains were sensitive to the phage lysin LysGH15 (MICs ranging from 8 to 32 μg/ml). More importantly, LysGH15 not only prevented biofilm formation by these staphylococci but also disrupted 24-h and 72-h biofilms. Furthermore, the in vivo efficacy of LysGH15 was demonstrated in a mouse model of S. epidermidis bacteremia. Thus, LysGH15 exhibits therapeutic potential for treating biofilm-related or non-biofilm-related infections caused by diverse staphylococci.

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Streptococcus pneumoniae Biofilm Formation Is Strain Dependent, Multifactorial, and Associated with Reduced Invasiveness and Immunoreactivity during Colonization

mBio ◽

10.1128/mbio.00745-13 ◽

2013 ◽

Vol 4 (5) ◽

Cited By ~ 68

Author(s):

Krystle Blanchette-Cain ◽

Cecilia A. Hinojosa ◽

Ramya Akula Suresh Babu ◽

Anel Lizcano ◽

Norberto Gonzalez-Juarbe ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Biofilm Formation ◽

Lavage Fluid ◽

Cytokine Response ◽

Host Cells ◽

Nasal Lavage ◽

Content Type ◽

Nasal Lavage Fluid

ABSTRACT Biofilms are thought to play an important role during colonization of the nasopharynx by Streptococcus pneumoniae, yet how they form in vivo and the determinants responsible remain unknown. Using scanning electron microscopy, we show that biofilm aggregates of increasing complexity form on murine nasal septa following intranasal inoculation. These biofilms were highly distinct from in vitro biofilms, as they were discontiguous and appeared to incorporate nonbacterial components such as intact host cells. Biofilms initially formed on the surface of ciliated epithelial cells and, as cells were sloughed off, were found on the basement membrane. The size and number of biofilm aggregates within nasal lavage fluid were digitally quantitated and revealed strain-specific capabilities that loosely correlated with the ability to form robust in vitro biofilms. We tested the ability of isogenic mutants deficient in CbpA, pneumolysin, hydrogen peroxide, LytA, LuxS, CiaR/H, and PsrP to form biofilms within the nasopharynx. This analysis revealed that CiaR/H was absolutely required for colonization, that PsrP and SpxB strongly impacted aggregate formation, and that other determinants affected aggregate morphology in a modest fashion. We determined that mice colonized with ΔpsrP mutants had greater levels of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), IL-1β, and KC in nasal lavage fluid than did mice colonized with wild-type controls. This phenotype correlated with a diminished capacity of biofilm pneumococci to invade host cells in vitro despite enhanced attachment. Our results show that biofilms form during colonization and suggest that they may contribute to persistence through a hyperadhesive, noninvasive state that elicits a dampened cytokine response. IMPORTANCE This work demonstrates the first temporal characterization of Streptococcus pneumoniae biofilm formation in vivo. Our results show that the morphology of biofilms formed by both invasive and noninvasive clinical isolates in vivo is distinct from that of formed biofilms in vitro, yet propensity to form biofilms in vivo loosely correlates with the degree of in vitro biofilm formation on a microtiter plate. We show that host components, including intact host cells, influence the formation of in vivo structures. We also found that efficient biofilm formation in vivo requires multiple bacterial determinants. While some factors are essential for in vivo biofilm formation (CiaRH, PsrP, and SpxB), other factors are less critical (CbpA, LytA, LuxS, and pneumolysin). In comparison to their planktonic counterparts, biofilm pneumococci are hyperadhesive but less invasive and elicit a weaker proinflammatory cytokine response. These findings give insight into the requirements for and potential role of biofilms during prolonged asymptomatic colonization.

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Candida albicans Als3, a Multifunctional Adhesin and Invasin

Eukaryotic Cell ◽

10.1128/ec.00279-10 ◽

2010 ◽

Vol 10 (2) ◽

pp. 168-173 ◽

Cited By ~ 159

Author(s):

Yaoping Liu ◽

Scott G. Filler

Keyword(s):

Candida Albicans ◽

Host Cell ◽

Surface Protein ◽

Host Cells ◽

Content Type ◽

Multifunctional Protein ◽

Multiple Functions ◽

Abiotic Surfaces ◽

Disseminated Disease

ABSTRACTCandida albicansis part of the normal human flora, and it grows on mucosal surfaces in healthy individuals. In susceptible hosts, this organism can cause both mucosal and hematogenously disseminated disease. ForC. albicansto persist in the host and induce disease, it must be able to adhere to biotic and abiotic surfaces, invade host cells, and obtain iron. TheC. albicanshypha-specific surface protein Als3 is a member of the agglutinin-like sequence (Als) family of proteins and is important in all of these processes. Functioning as an adhesin, Als3 mediates attachment to epithelial cells, endothelial cells, and extracellular matrix proteins. It also plays an important role in biofilm formation on prosthetic surfaces, both alone and in mixed infection withStreptococcus gordonii. Als3 is one of two knownC. albicansinvasins. It binds to host cell receptors such as E-cadherin and N-cadherin and thereby induces host cells to endocytose the organism. Als3 also binds to host cell ferritin and enablesC. albicansto utilize this protein as a source of iron. Because of its multiple functions and its high expression levelin vivo, Als3 is a promising target for vaccines that induce protective cell-mediated and antibody responses. This review will summarize the multiple functions of this interesting and multifunctional protein.

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Pneumococcal Interactions with Epithelial Cells Are Crucial for Optimal Biofilm Formation and ColonizationIn VitroandIn Vivo

Infection and Immunity ◽

10.1128/iai.00488-12 ◽

2012 ◽

Vol 80 (8) ◽

pp. 2744-2760 ◽

Cited By ~ 100

Author(s):

Laura R. Marks ◽

G. Iyer Parameswaran ◽

Anders P. Hakansson

Keyword(s):

Epithelial Cells ◽

Biofilm Formation ◽

Invasive Disease ◽

Respiratory Epithelial Cells ◽

Symptomatic Infection ◽

Content Type ◽

Abiotic Surfaces ◽

Horizontal Spread ◽

Respiratory Epithelial

ABSTRACTThe human nasopharynx is the main reservoir forStreptococcus pneumoniae(the pneumococcus) and the source for both horizontal spread and transition to infection. Some clinical evidence indicates that nasopharyngeal carriage is harder to eradicate with antibiotics than is pneumococcal invasive disease, which may suggest that colonizing pneumococci exist in biofilm communities that are more resistant to antibiotics. While pneumococcal biofilms have been observed during symptomatic infection, their role in colonization and the role of host factors in this process have been less studied. Here, we show for the first time that pneumococci form highly structured biofilm communities during colonization of the murine nasopharynx that display increased antibiotic resistance. Furthermore, pneumococcal biofilms grown on respiratory epithelial cells exhibited phenotypes similar to those observed during colonizationin vivo, whereas abiotic surfaces produced less ordered and more antibiotic-sensitive biofilms. The importance of bacterial-epithelial cell interactions during biofilm formation was shown using both clinical strains with variable colonization efficacies and pneumococcal mutants with impaired colonization characteristicsin vivo. In both cases, the ability of strains to form biofilms on epithelial cells directly correlated with their ability to colonize the nasopharynxin vivo, with colonization-deficient strains forming less structured and more antibiotic-sensitive biofilms on epithelial cells, an association that was lost when grown on abiotic surfaces. Thus, these studies emphasize the importance of host-bacterial interactions in pneumococcal biofilm formation and provide the first experimental data to explain the high resistance of pneumococcal colonization to eradication by antibiotics.

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Monitoring of Fluconazole and Caspofungin Activity against In Vivo Candida glabrata Biofilms by Bioluminescence Imaging

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01555-18 ◽

2018 ◽

Vol 63 (2) ◽

pp. e01555-18 ◽

Cited By ~ 3

Author(s):

Aranka Persyn ◽

Ona Rogiers ◽

Matthias Brock ◽

Greetje Vande Velde ◽

Mohamed Lamkanfi ◽

...

Keyword(s):

Real Time ◽

Biofilm Formation ◽

Candida Glabrata ◽

Bioluminescence Imaging ◽

Firefly Luciferase ◽

Antibiofilm Activity ◽

Limited Information ◽

Content Type ◽

Abiotic Surfaces

ABSTRACT Candida glabrata can attach to various medical implants and forms thick biofilms despite its inability to switch from yeast to hyphae. The current in vivo C. glabrata biofilm models only provide limited information about colonization and infection and usually require animal sacrifice. To gain real-time information from individual BALB/c mice, we developed a noninvasive imaging technique to visualize C. glabrata biofilms in catheter fragments that were subcutaneously implanted on the back of mice. Bioluminescent C. glabrata reporter strains (lucOPT 7/2/4 and lucOPT 8/1/4), free of auxotrophic markers, expressing a codon-optimized firefly luciferase were generated. A murine subcutaneous model was used to follow real-time in vivo biofilm formation in the presence and absence of fluconazole and caspofungin. The fungal load in biofilms was quantified by CFU counts and by bioluminescence imaging (BLI). C. glabrata biofilms formed within the first 24 h, as documented by the increased number of device-associated cells and elevated bioluminescent signal compared with adhesion at the time of implant. The in vivo model allowed monitoring of the antibiofilm activity of caspofungin against C. glabrata biofilms through bioluminescent imaging from day four after the initiation of treatment. Contrarily, signals emitted from biofilms implanted in fluconazole-treated mice were similar to the light emitted from control-treated mice. This study gives insights into the real-time development of C. glabrata biofilms under in vivo conditions. BLI proved to be a dynamic, noninvasive, and sensitive tool to monitor continuous biofilm formation and activity of antifungal agents against C. glabrata biofilms formed on abiotic surfaces in vivo.

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Host factors abolish the need for polysaccharides and extracellular matrix-binding protein in Staphylococcus epidermidis biofilm formation

Journal of Medical Microbiology ◽

10.1099/jmm.0.001287 ◽

2021 ◽

Author(s):

Sandra M. Skovdal ◽

Liva Kjær Hansen ◽

Diana Malskær Ivarsen ◽

Guanghong Zeng ◽

Henning Büttner ◽

...

Keyword(s):

Extracellular Matrix ◽

Human Plasma ◽

Staphylococcus Epidermidis ◽

Biofilm Formation ◽

Type Species ◽

Host Factors ◽

Content Type ◽

Link Type ◽

Matrix Components

Introduction. Staphylococcus epidermidis is predominant in implant-associated infections due to its capability to form biofilms. It can deploy several strategies for biofilm development using either polysaccharide intercellular adhesin (PIA), extracellular DNA (eDNA) and/or proteins, such as the extracellular matrix-binding protein (Embp). Hypothesis/Gap Statement. We hypothesize that the dichotomic regulation of S. epidermidis adhesins is linked to whether it is inside a host or not, and that in vitro biofilm investigations in laboratory media may not reflect actual biofilms in vivo. Aim. We address the importance of PIA and Embp in biofilm grown in ‘humanized’ media to understand if these components play different roles in biofilm formation under conditions where bacteria can incorporate host proteins in the biofilm matrix. Methodology. S. epidermidis 1585 WT (deficient in icaADBC), and derivative strains that either lack embp, express embp from an inducible promotor, or express icaADBC from a plasmid, were cultivated in standard laboratory media, or in media with human plasma or serum. The amount, structure, elasticity and antimicrobial penetration of biofilms was quantified to describe structural differences caused by the different matrix components and growth conditions. Finally, we quantified the initiation of biofilms as suspended aggregates in response to host factors to determine how quickly the cells aggregate in response to the host environment and reach a size that protects them from phagocytosis. Results. S. epidermidis 1585 required polysaccharides to form biofilm in laboratory media. However, these observations were not representative of the biofilm phenotype in the presence of human plasma. If human plasma were present, polysaccharides and Embp were redundant for biofilm formation. Biofilms formed in human plasma were loosely attached and existed mostly as suspended aggregates. Aggregation occurred after 2 h of exposing cells to plasma or serum. Despite stark differences in the amount and composition of biofilms formed by polysaccharide-producing and Embp-producing strains in different media, there were no differences in vancomycin penetration or susceptibility. Conclusion. We suggest that the assumed importance of polysaccharides for biofilm formation is an artefact from studying biofilms in laboratory media void of human matrix components. The cell–cell aggregation of S. epidermidis can be activated by host factors without relying on either of the major adhesins, PIA and Embp, indicating a need to revisit the basic question of how S. epidermidis deploys self-produced and host-derived matrix components to form antibiotic-tolerant biofilms in vivo.

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Vaccination with SesC Decreases Staphylococcus epidermidis Biofilm Formation

Infection and Immunity ◽

10.1128/iai.00104-12 ◽

2012 ◽

Vol 80 (10) ◽

pp. 3660-3668 ◽

Cited By ~ 37

Author(s):

Mohammad Shahrooei ◽

Vishal Hira ◽

Laleh Khodaparast ◽

Ladan Khodaparast ◽

Benoit Stijlemans ◽

...

Keyword(s):

Staphylococcus Epidermidis ◽

Biofilm Formation ◽

Inhibitory Effect ◽

Antigenic Determinants ◽

Infection Model ◽

Content Type ◽

Protein Size ◽

Promising Target

ABSTRACTThe increased use of medical implants has resulted in a concomitant rise in device-related infections. The majority of these infections are caused byStaphylococcus epidermidisbiofilms. Immunoprophylaxis and immunotherapy targetingin vivo-expressed, biofilm-associated, bacterial cell surface-exposed proteins are promising new approaches to prevent and treat biofilm-related infections, respectively. Using anin silicoprocedure, we identified 64 proteins that are predicted to beS.epidermidissurface exposed (Ses), of which 36 were annotated as (conserved) hypothetical. Of these 36 proteins, 5 proteins—3 LPXTG motif-containing proteins (SesL, SesB, and SesC) and 2 of the largest ABC transporters (SesK and SesM)—were selected for evaluation as vaccine candidates. This choice was based on protein size, number of antigenic determinants, or the established role inS. epidermidisbiofilm formation of the protein family to which the candidate protein belongs. Anti-SesC antibodies exhibited the greatest inhibitory effect onS. epidermidisbiofilm formationin vitroand on colonization and infection in a mouse jugular vein catheter infection model that includes biofilms and organ infections. Active vaccination with a recombinant truncated SesC inhibitedS. epidermidisbiofilm formation in a rat model of subcutaneous foreign body infection. Antibodies to SesC were shown to be opsonic by anin vitroopsonophagocytosis assay. We conclude that SesC is a promising target for antibody mediated strategies againstS. epidermidisbiofilm formation.

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Biofilm Formation on Biotic and Abiotic Surfaces in the Presence of Antimicrobials by Escherichia coli Isolates from Cases of Bovine Mastitis

Applied and Environmental Microbiology ◽

10.1128/aem.01953-14 ◽

2014 ◽

Vol 80 (19) ◽

pp. 6136-6145 ◽

Cited By ~ 20

Author(s):

Vitor O. Silva ◽

Larissa O. Soares ◽

Abelardo Silva Júnior ◽

Hilário C. Mantovani ◽

Yung-Fu Chang ◽

...

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Expression Profiles ◽

Bovine Mastitis ◽

Pcr Analysis ◽

Content Type ◽

E Coli ◽

Biofilm Production ◽

Abiotic Surfaces

ABSTRACTEscherichia coliis a highly adaptive microorganism, and its ability to form biofilms under certain conditions can be critical for antimicrobial resistance. The adhesion of fourE. coliisolates from bovine mastitis to bovine mammary alveolar (MAC-T) cells, biofilm production on a polystyrene surface, and the expression profiles of the genesfliC,csgA,fimA, andluxSin the presence of enrofloxacin, gentamicin, co-trimoxazole, and ampicillin at half of the MIC were investigated. Increased adhesion ofE. coliisolates in the presence of antimicrobials was not observed; however, increased internalization of some isolates was observed by confocal microscopy. All of the antimicrobials induced the formation of biofilms by at least one isolate, whereas enrofloxacin and co-trimoxazole decreased biofilm formation by at least one isolate. Quantitative PCR analysis revealed that all four genes were differentially expressed when bacteria were exposed to subinhibitory concentrations of antimicrobials, with expression altered on the order of 1.5- to 22-fold. However, it was not possible to associate gene expression with induction or reduction of biofilm formation in the presence of the antimicrobials. Taken together, the results demonstrate that antimicrobials could induce biofilm formation by some isolates, in addition to inducing MAC-T cell invasion, a situation that might occurin vivo, potentially resulting in a bacterial reservoir in the udder, which might explain some cases of persistent mastitis in herds.

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