Biofilms in Otolaryngology

Definition: 3-Dimentional Communities of sessile bacteria embedded in a matrix of extracellular polymeric substances of their own synthesis that adhere to a foreign body or a mucosal surface with impaired host defense
Within the biofilm population, there appears to be a division of job. This may occur between bacteria of different species, as well as with fungi, protozoa. Intercellular signalling between individual members of the biofilm population allows the colony to respond rapidly to changing environmental circumstances. Through the process of quorum sensing, pathogenic bacteria can coordinate their virulence to evade immune capture and successfully infect their host. Once a mature biofilm has been established, planktonic showers of free-floating organisms may be rereleased periodically from the mature colony, completing a life cycle Slow-growing cells have been shown to be less susceptible to antimicrobial agents. Alternatively, the bacteria may adopt a new, resistantphenotype, which may involve the transfer of genetic material, including virulence and resistance coding between monospecies and polyspecies of bacteria

The question is, does always a biofilm cause a clinical disease

biofilms exist in normal mucosa as well as infected sinuses the organism virulence and the quantity of the organism allows the biofilm to become pathogenic

The problem of biofilms Growing resistance to our treatment of infections in otolaryngology. Disease process is both more chronic and more recurrent Direct bacterial evolution of resistance biofilms play rule in COM,sinusitis,adenoiditis and implants failure

Physiologic Limitation of Antimicrobial Efficiency semistarved cells within the nutrient-depleted centre of the biofilm grow at a slower rate. Slow-growing cells have been shown to be less susceptible to antimicrobial agents. Alternatively, the bacteria may adopt a new, resistant phenotype, which may involve the transfer of genetic material, including virulence and resistance coding between monospecies and polyspecies of bacteria. Inaddition to these resistance mechanisms, consideration should be given to the limitations of traditional laboratory tests of antibiotic susceptibility. Most antibiotics have been developed by testing their efficacy against a suspension of planktonic cells

Two Distinct Biofilm Characteristics

1) Biofilms are highly resistant to immune killing and clearance and to treatment with antimicrobial agents. Even the correct antibiotic from in vitro culture will not kill the bacteria hiding under the matrix in vivo.
2) Biofilms might be capable of shedding individual bacteria to the surrounding tissues and into the circulatory system, thus causing bouts of infection, which may recur despite intensive antimicrobial treatment. Antibiotics only treat the circulating bacteria, not those hiding and waiting until the antimicrobial pressure is off. This is the Reason why longer term antibiotics needed for chronic sinusitis and tonsillitis.

5 Step Process to become a Biofilm

1.Bacterial adherence to mucosa electrostatically (Planktonic form)----(Sec) 2.Irreversible adhesion by chemical bonds------(Sec to Min) 3.Aggregation and formation of extracellular polysach substances(matrix)(hrs-days) 4. Maturation of the biofilm structure------(hrs-days) Secondary colonizers like fungi or other bacteria can attach to the biofilm at this stage 5. Bacteria detachment(days-months)

Specific ENT Diseases

Ear Infections 1) (AOM) is associated with planktonic bacteria Susceptible to antibiotics 2)OME and CSOM may be associated with biofilm formation when the biofilm is present culture will be difficult PCR can detect DNA of bacteria such as H.influenza Refractory to antibiotic treatment
Intermittent shedding of planktonic middle ear pathogens from the nasopharynx may help explain the recalcitrance and recurrence of otitis media, the clinical benefit seen with adenoidectomy in RAOM may be due to mechanical debridement of these resistant nasopharyngeal biofilms

outcomes from studies on otitis media (OM) that looked at the presence of biofilms on the adenoids of children who had undergone adenoidectomy for various conditions. One study showed that for the children with recurrent acute OM, the average adenoid was 98 percent covered with biofilm on its surface, while 28 percent of children with chronic middle ear fluid had biofilms on their adenoids, and less than 1 percent of those who had their adenoids removed because of obstruction alone had biofilms on their adenoids. In addition, the biofilms identified from otitis-prone children appeared to be multiorganismal and mature

Cholesteatoma is ideal for biofilm formation and associated with pseudomonal firm attachment on microscopy. Infections of implanted devices seen by electron microscopy myringotomy tubes,Cochlear implants , BAHA or artificial ossicles

biofilms might explain the behaviour of cholesteatoma-related ear infection. Bouts of active otorrhea that appear to respond to topical antibiotic therapy might correlate with the release of motile planktonic forms. In this state, the organisms are more liable to antimicrobial therapy. Unfortunately, the resistant biofilm remains within the cholesteatoma matrix and ensures further episodes of otorrhea and the chronicity of the condition. Bacterial endotoxin release and the binding of sessile bacteria to epithelial cells are thought to leadto regulation of signals that contribute to increased epithelial cell turnover and keratin production. Microsurgical removal of the cholesteatoma matrix and associated biofilm is likely to remain the standard of care for this condition.

Otitis Media with Effusion and Postventilation Tube Otorrhea Difficult to culture and resistant to antibiotic treatment, chronic middle ear effusions have traditionally been considered to be a sterile inflammatory process. The recent discovery of (DNA) in culture-negative effusions has offered the alternative explanation that otitis media with effusion (OME) might be a biofilm disease. Evidence to support the theory that OME is an infective condition relies on the common chronologic association with acute suppurative otitis media and the fact that similar organisms have been identified in each condition. Acute effusions are often positive to culture, but with chronicity, this culturability is lost and the effusions appear to be sterile. A partial, short-term response of OME to antibiotic treatment usually occur. This may reflect the partial success of an antimicrobial assault on the biofilm. superficial bacteria may be killed, correlating with a degree of clinical improvement. However, antibiotic treatment may indirectly promote the resistant biofilm phenotype, leading to recurrence and chronicity. If biofilm formation is ndeed involved in OME, then the efficacy of tube insertion is probably related to the drying effect of reestablishing middle ear ventilation. Removal of adenoidal tissue by curettage or diathermy has been shown to impact on the resolution of OME, possibly by removing the biofilm residing in the nasopharynx. Biofilm formation has also been implicated as a cause for chronic post-tympanostomy tube otorrhea. Biofilm has be directly visualized on tubes removed from affected individuals

The most important step in treating a biofilm infection is first to have a high index of suspicion that it actually exists. Cases of chronic or recurrent infection apparently resistant to conventional treatment, particularly in the presence of implanted devices or other foreign bodies, must raise concern. Attempts to identify the infective agents often prove to be the most challenging, potentially expensive, and most impractical part of the puzzle. Electron microscopy and polymerase chain reaction typing are inaccessible to most in clinical practice. Traditional antiseptic techniques, the removal of infected foreign bodies, and the provision of meticulous physical dbridement still have a fundamental role to play in our armamentarium against the biofilm. However, the development of our understanding of how bacteria interact to form a biofilm offers novel therapeutic opportunities for the future. Specific enzymes are known to be capable of digesting the biofilms protective mucoid extracellular polysaccharide (EPS) coat. With Pseudomonas, the EPS is an alginate, and treatment of the Pseudomonasbiofilm with alginase has been shown to increase the efficacy of simultaneous antibiotic therapy

Infections of the nose and throat

Two strains of bacteria, P. aeruginosa and S. aureus, were clinically isolated from CRS patients who had poor outcomes following functional endoscopic sinus surgery. Persistence of viable bacteria was assessed quantitatively by assessment of colony-forming units and by confocal scanning laser microscopy. Treatment with static application of saline alone or topical antibiotic solutions at standard concentrations produced no reduction in bacterial counts

Chronic rhinosinusitis (CRS) Biofilms found on mucosa from all sinuses when undergoing functional endoscopic sinus surgery for CRS No biofilms on patients undergoing nasal surgery not for CRS when adenotonsillectomy done in p with CRS both tonsils and adenoids where covered with biofilm this goes with the theory of shedding of planktonic cells and there implantation at distant site as biofilm colony

bacteria in the biofilm are substantially more resistant to conventional antimicrobial therapy and host immune defense pathways than free bacteria and thus are hypothesized to contribute to medical Rx resistant rhinosinusitis conventional therapies are of little use against biofilm infections and support the idea that delivery of a soaplike surfactant with a calcium ion-sequestering agent combined with the use of hydrodynamic force can break down bacterial biofilms possibly associated with CRS. Eg: Nasal lavage with sodium hypochlorite(0.05% NaOCl solution) in Staphylococcus aureus persistent rhinosinusitis is suitable for long-term use and seems to be a good alternative to lavage with saline alone in the management of symptomatic RS

Virtually all microbes can live in biofilm communities. These communities cause chronic and device-associated infections. Within the specialist field of otorhinolaryngology, biofilms have been shown to play a role in many infections, including: chronic otitis media, cholesteatoma, chronic tonsillitis, chronic sinusitis, and infections of tracheostomies, endotracheal tubes and cochlear implants,BAHA. most common Microrganisms encountered in ENT are p.aeruginosa , S.aureus , candida albicans,haemophilus and pneumococci to a lesser extent but usually mature biofilms are multimicrobial

Treatment of Biofilm Infections

Biofilms are 500 times more resistant than planktonic bacteria to antibiotics

multidrug efflux pumps stress-response regulons reduced rates of cellular growth- multiple rates in a colony and respiration of biofilm bacteria protection conferred by biofilm matrix polymers Avoid macrophage phagocytosis and oxidative burst mechanisms Heterogenic population so one stategy does not kill all

Treatment of Biofilm Infections

Now understand a little about biofilms can create treatment strategies

Interruption of quorum sensing Interference with transcription-genes for biofilm formation Interference with genes for attachment Disruption of the EPS mechanically or chemically to expose the bacteria to the antimicrobials Probiotic colonization to prevent space for biofilm formation

probiotics or good biofilms may be developed to combat pathogenic species. This approach has been used with some success in combating tracheoesophageal valve colonization Other bacteria produce biosurfactants, which impair adhesion, damage the biofilm matrix and release free bacteria, which may be more susceptible to the host's immune system and conventional antimicrobial agents

Treatment of Biofilm Infections

1)Antibiotics Fluoquinolones kill non-growing bacteria B-lactams kill only growing cells Macrolides inhibit biofilm formation, possibly because of properties other than bactericidal activity
Studies have shown that some macrolide antibiotics (i.e. clarithromycin and erythromycin) may be effective in killing sessile bacteria within biofilms. Furthermore, some macrolides inhibit biofilm formation, possibly because of properties other than bactericidal activity. macrolides have immunomodulatory action
2)Indwelling devices Cover the surfaces with albumin or ion bombardment of silicone
iron inhibits the expression of certain genes essential for biofilm production. Since these iron salts are well tolerated biologically and inhibit biofilms of Pseudomonasspp. at low concentrations, they may be an efficient way of treating biofilm diseases in vivo
single modal therapy will most likely not be effective to treat biofilms in patients with CRS and that most will need multi-modality treatment that includes high-dose topical antibiotics with a surfactant after mechanical disruption such as endoscopic sinus surgery or hydrodynamic shearing of the biofilm in the sinuses

Summary

Biofilm formation is a dynamic five-step process guided by interbacterial communicating systems. Bacteria in biofilms express different genes and have markedly different phenotypes from their planktonic counterparts. Detachment of cells, production of endotoxin, increased resistance to the host immune system, and provision of a niche for the generation of resistant organisms are biofilm processes that could initiate the infection process.

Effective prevention and management strategies include interruption of intrabiofilm signalling, inhibition of related genes, disruption of the protective extrapolymer matrix, macrolides (clarithromycin and erythromycin), and mechanical debridement of the biofilm-bearing tissues.

Bacteria within a biofilm differ from bacteria growing in suspension in many ways, including reduced growth rate, distinct gene expression and increased resistance to antibiotics. the difficulty of culturing bacteria in biofilms in the lab, and their resistance to antibiotics, may explain the conflicting features of OM in a patient with symptoms of chronic OM who responds poorly to antibiotics and in whom their culture is negative. Since biofilms are resistant to many antibiotics, we suspect that biofilm formation may explain why infections by Streptococcus pneumoniae and other organisms may be so difficult to treat in some children, So new therapies are needed to overcome these hardy infections.