Nonmucoid Strains Research Articles

Alginate lyase enhances antibiotic killing of mucoid Pseudomonas aeruginosa in biofilms

Once mucoid (alginate-producing) strains of Pseudomonas aeruginosa have become established in the respiratory tracts of cystic fibrosis patients they can rarely be eliminated by antibiotic treatment alone; we have investigated, in an in vitro biofilm system, the putative role of co-administration of alginate lyase with antibiotic. Biofilms were maintained in continuous flow culture in a medium resembling sputum from CF patients. Antibiotics and/or alginate lyase were added to some of the cultures. Biofilms of two mucoid CF strains of P. aeruginosa were, in most cases, not eradicated by a one-week course of treatment with 64 microg/ml of gentamicin; the same concentration of gentamicin, under the same conditions, led to the apparent elimination of all biofilms of non-mucoid derivatives of these strains. When alginate lyase and gentamicin were administered together the apparent elimination of mucoid bacteria from biofilms was achieved, whereas the mucoid bacteria in most control biofilms treated only with gentamicin persisted. Ceftazidime treatment of biofilms was more effective against those containing the non-mucoid strains than those with mucoid strains. These studies support the view that co-administration of antibiotics with alginate lyase, which degrades the exopolysaccharide produced by mucoid strains of P aeruginosa, might benefit CF patients by increasing the efficacy of antibiotic in the respiratory tract.

Cadmium uptake and nitrogen fixing ability in heavy-metal-resistant laboratory and field strains of Rhizobium leguminosarum biovar trifolii

Rhizobium leguminosarum bv. trifolii produced mucoid colonies on yeast mannitol agar, but non-mucoid colonies appeared with a frequency of 10−7 on agar additionally containing 4 mM ZnCl2. Cells producing non-mucoid colonies were also obtained from root nodules of white clover from experimental field plots treated with heavy-metal-contaminated sewage sludge. Compared to mucoid strains, non-mucoid strains produced approximately half of the extracellular polysaccharide, two to three fold higher levels of intracellular carbohydrate and possessed larger intracellular inclusions. They also showed up to two-fold times greater resistance to Cd2+, Cu2+, Ni2+ and Zn2+. The total uptakes of Cd2+ were similar between the laboratory strains and amongst the field strains. In non-mucoid strains, Cd2+ was not removed by washing in 0.1 M HNO3, suggesting Cd2+ was bound intracellularly. All strains were effective in fixing nitrogen in white clover.

Characterization of biochemical properties and biological activities of biosurfactants produced by Pseudomonas aeruginosa mucoid and non-mucoid strains isolated from hydrocarbon-contaminated soil samples

Biochemical and pharmacological properties of biosurfactants produced at 45 degrees C temperature by Pseudomonas aeruginosa mucoid (M) and non-mucoid (NM) strains, isolated from hydrocarbon-contaminated soil samples, were characterized. Both the strains secreted appreciable amount of biosurfactants (5.0-6.5 g/l), responsible for the reduction of surface tension of the medium from 68 to 29 +/- 0.5 mN/m post 96 h of growth. Maximum yield of biosurfactants was observed following the supplementation of NH(4)Cl and glycerol as nitrogenous source and carbon source, respectively. These thermostable biosurfactants exhibited strong emulsifying property and could release appreciable amount of oil from saturated sand-pack column. Pharmacological characterization of these biosurfactants revealed that they induced dose-dependent hemolysis and coagulation of platelet-poor plasma but were non-detrimental to chicken lung, liver, heart and kidney tissues. Our study has documented that biosurfactants from P. aeruginosa M and NM strains could be exploited for use in petroleum sectors as well in pharmaceutical industries.

NonmucoidPseudomonas aeruginosaExpresses Alginate in the Lungs of Patients with Cystic Fibrosis and in a Mouse Model

In patients with cystic fibrosis (CF), lung infection with mucoid Pseudomonas aeruginosa strains overexpressing the exopolysaccaride alginate is preceded by colonization with nonmucoid strains. We investigated the kinetics, impact of environmental signals, and genetics of P. aeruginosa alginate expression in a mouse model and in patients with CF. Using indirect immunofluorescence, microarray technology and real-time reverse-transcription polymerase chain reaction, we assessed alginate gene expression during aerobic and anaerobic growth of the nonmucoid strain PAO1 in vitro, in a mouse lung-infection model and in sputum specimens from patients with CF infected with nonmucoid or mucoid P. aeruginosa strains. Anaerobic conditions increased the transcription of alginate genes in vitro and in murine lungs within 24 h. Alginate production by PAO1 in murine lungs and by nonmucoid P. aeruginosa strains in patients with CF was reversible after in vitro culture under aerobic conditions. A subpopulation of P. aeruginosa clones revealing stable alginate production was detected in murine lungs 2 weeks after infection. Anaerobiosis and lung infection rapidly induce alginate production by gene regulation in nonmucoid P. aeruginosa. This trait may contribute to early persistence, leading to chronic P. aeruginosa infection once stable mucoid strains are generated.

De Novo Generation of Cationic Antimicrobial Peptides: Influence of Length and Tryptophan Substitution on Antimicrobial Activity

Comparison of human immunodeficiency virus lentiviral lytic peptide 1 with other host-derived peptides indicates that antimicrobial properties of membrane-active peptides are markedly influenced by their cationic, hydrophobic, and amphipathic properties. Many common themes, such as Arg composition of the cationic face of an amphipathic helix and the importance of maintaining the hydrophobic face, have been deduced from these observations. These studies suggest that a peptide with these structural properties can be derived de novo by using only a few strategically positioned amino acids. However, the effects of length and helicity on antimicrobial activity and selectivity have not been objectively evaluated in the context of this motif. To address these structure-function issues, multimers of a 12-residue lytic base unit (LBU) peptide composed only of Arg and Val residues aligned to form idealized amphipathic helices were designed. Bacterial killing assays and circular dichroism analyses reveal a strong correlation between antibacterial activity, peptide length, and propensity to form a helix in solvent mimicking the environment of a membrane. Increasing peptide length beyond two LBUs (24-residue peptides) resulted in no appreciable increase in antimicrobial activity. Derivatives (WLBU) of the LBU series were further engineered by substituting Trp residues in the hydrophobic domains. The 24-residue WLBU2 peptide was active at physiologic NaCl concentrations against Staphylococcus aureus and mucoid and nonmucoid strains of Pseudomonas aeruginosa. Further, WLBU2 displayed the highest antibacterial selectivity of all peptides evaluated in the present study by using a coculture model of P. aeruginosa and primary human skin fibroblasts. These findings provide fundamental information toward the de novo design of an antimicrobial peptide useful for the management of infectious diseases.

Human Monoclonal Antibodies toPseudomonas aeruginosaAlginate That Protect against Infection by Both Mucoid and Nonmucoid Strains

Two fully human mAbs specific for epitopes dependent on intact carboxylate groups on the C6 carbon of the mannuronic acid components of Pseudomonas aeruginosa alginate were found to promote phagocytic killing of both mucoid and nonmucoid strains as well as protection against both types of strains in a mouse model of acute pneumonia. The specificity of the mAbs for alginate was determined by ELISA and killing assays. Some strains of P. aeruginosa did not make detectable alginate in vitro, but in vivo protection against lethal pneumonia was obtained and shown to be due to rapid induction of expression of alginate in the murine lung. No protection against strains genetically unable to make alginate was achieved. These mAbs have potential to be passive therapeutic reagents for all strains of P. aeruginosa and the results document that alginate is a target for the proper type of protective Ab even when expressed at low levels on phenotypically nonmucoid strains.

The biological characteristics of a mucoid Pseudomonas aeruginosa strain with a newly discovered mutation of mucA gene

To compare the mucA gene sequence, biofilm formation, growth rate and antibiotic resistance of a mucoid Pseudomonas aeruginosa strain 'PA17' with those of the nonmucoid strain PAOI. The mucA genes of PA17 and PAOI were amplified and the PCR products were sequenced. Biofilm was established and detected with scanning electron microscope 8 h, 24 h, 3 d and 6 d later. Clones counting method was used to compare the growth curves of PA17 and PAOI. International standard plate dilution method was used to detect the susceptibility of common antibiotics for PA17 and PAOI in vitro. There was a 166-333 deletion mutation in mucA gene of PA17. The biofilm formation rate and growth rate of PA17 were noticeably lower than those of PAOI. The antibiotic resistance profile of PA17 was identical to that of PAOI. A mucoid Pseudomonas aeruginosa with a new type of mutation of mucA gene is discovered. The biologic characteristics of this strain are different from those of the mucoid Pseudomonas aeruginosa strain reported previously, which may be related with the newly discovered mutation of mucA gene.

The transcriptional regulator AlgR controls cyanide production in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic pathogen that causes chronic lung infections in cystic fibrosis (CF) patients. One characteristic of P. aeruginosa CF isolates is the overproduction of the exopolysaccharide alginate, controlled by AlgR. Transcriptional profiling analyses comparing mucoid P. aeruginosa strains to their isogenic algR deletion strains showed that the transcription of cyanide-synthesizing genes (hcnAB) was approximately 3-fold lower in the algR mutants. S1 nuclease protection assays corroborated these findings, indicating that AlgR activates hcnA transcription in mucoid P. aeruginosa. Quantification of hydrogen cyanide (HCN) production from laboratory isolates revealed that mucoid laboratory strains made sevenfold more HCN than their nonmucoid parental strains. In addition, comparison of laboratory and clinically derived nonmucoid strains revealed that HCN was fivefold higher in the nonmucoid CF isolates. Moreover, the average amount of cyanide produced by mucoid clinical isolates was 4.7 +/- 0.85 micromol of HCN/mg of protein versus 2.4 +/- 0.40 micromol of HCN/mg of protein for nonmucoid strains from a survey conducted with 41 P. aeruginosa CF isolates from 24 patients. Our data indicate that (i) mucoid P. aeruginosa regardless of their origin (laboratory or clinically derived) produce more cyanide than their nonmucoid counterparts, (ii) AlgR regulates HCN production in P. aeruginosa, and (iii) P. aeruginosa CF isolates are more hypercyanogenic than nonmucoid laboratory strains. Taken together, cyanide production may be a relevant virulence factor in CF lung disease, the production of which is regulated, in part, by AlgR.

Control of Pseudomonas aeruginosa algZ expression by the alternative sigma factor AlgT.

AlgZ controls Pseudomonas aeruginosa alginate synthesis by activating algD, yet algZ expression is not detectable in nonmucoid strains. Mobility shift and Western blot assays revealed that algZ expression requires the sigma factor AlgT. The mapped algZ transcription start site revealed a consensus AlgT-dependent promoter that, when mutated, substantially reduced algZ transcription.

Pseudomonas aeruginosa slime glycolipoprotein is a potent stimulant of tumor necrosis factor alpha gene expression and activation of transcription activators nuclear factor kappa B and activator protein 1 in human monocytes.

Pseudomonas aeruginosa, an opportunistic pathogen, causes infections associated with a high incidence of morbidity and mortality in immunocompromised hosts. Production of tumor necrosis factor alpha (TNF-alpha), primarily by cells of monocytic lineage, is a crucial event in the course of these infections. During in vivo infections with P. aeruginosa, both lipopolysaccharide (LPS) and extracellular slime glycolipoprotein (GLP) produced by mucoid and nonmucoid strains are released. In the present study, we sought to explore the relative contributions of these two bacterial products to TNF-alpha production by human monocytes. To this end, fresh human monocytes and THP-1 human monocytic cells were stimulated with P. aeruginosa LPS or GLP. GLP was found to be a more potent stimulus for TNF-alpha production (threefold higher) by human monocytes than LPS. Moreover, its effect was comparable to that of viable bacteria. Quantitative mRNA analysis revealed predominantly transcriptional regulation. Electrophoretic mobility shift assays and transfection assays demonstrated activation of NF-kappa B and activator protein 1 (AP-1). NF-kappa B activation by GLP was rapid and followed the same time course as that by viable bacteria, suggesting that bacteria could directly activate NF-kappa B through GLP. Moreover P. aeruginosa GLP induced the formation of AP-1 complex with delayed kinetics compared with NF-kappa B but much more efficiently than the homologous LPS. These results identify GLP as the most important stimulant for TNF-alpha production by human monocytes. Activation of NF-kappa B and AP-1 by P. aeruginosa GLP may be involved not only in TNF-alpha induction but also in many of the inflammatory responses triggered in the course of infection with P. aeruginosa.

Surfactant protein A and D differently regulate the immune response to nonmucoid Pseudomonas aeruginosa and its lipopolysaccharide.

We investigated the role of the surfactant proteins (SPs) A and D in the pulmonary immune defense of nonmucoid strains of Pseudomonas aeruginosa, the most etiologic agents of nosocomial Pseudomonas pneumonia. We first examined the interactions of recombinant human SP-D dodecamers and purified natural or recombinant human SP-A with two smooth, and two rough, clinical isolates of nonmucoid P. aeruginosa. SP-D bound to all four isolates, but agglutinated only one rough and one smooth strain. SP-D functioned as an opsonin to enhance the uptake of all four strains by the human monocytic cell line Mono Mac 6 (MM6). SP-D also enhanced tumor necrosis factor-alpha secretion by MM6 cells in response to purified lipopolysaccharide (LPS) isolated from the rough, but not the smooth, strains. Although SP-A bound to all four strains, it did not cause bacterial aggregation or enhance uptake. It showed small but statistically significant inhibitory effects on the cytokine response of MM6 cells to one strain of smooth organisms, but did not significantly alter the response to purified LPS. This study in combination with previously published data strongly suggests that SP-D may play important roles in the local innate pulmonary defense against nonmucoid P. aeruginosa of diverse LPS phenotypes, and preferentially augments the cellular response to rough P. aeruginosa endotoxin.

Early infection and progression of cystic fibrosis lung disease.

Chronic Pseudomonas aeruginosa infection is the major limitation in overall survival of patients with cystic fibrosis, and elective bronchoscopy shows that infection with this organism starts at a very early age. Early infection with nonmucoid P. aeruginosa gradually develops into chronic infection, characterized by the presence of microcolonies of alginate-producing (mucoid) P. aeruginosa in the bronchial tree. Chronic infection cannot be cured, but aggressive antimicrobial treatment will prolong life expectancy and decrease morbidity. It has, however, over the last decade become evident that early antimicrobial treatment of the initial infection with nonmucoid strains can prevent or at least postpone transition into chronic (mucoid) infection. This treatment strategy is likely to dramatically improve the prognosis of patients with cystic fibrosis in the immediate future.

Application of fluorescently labelled lectins for the visualization and biochemical characterization of polysaccharides in biofilms of Pseudomonas aeruginosa

Fluorescently labelled lectins were used in combination with epifluorescence microscopy and confocal laser scanning microscopy to allow the visualization and characterization of carbohydrate-containing extracellular polymeric substances (EPS) in biofilms of Pseudomonas aeruginosa. A mucoid strain characterized by an overproduction of the exopolysaccharide alginate, and an isogenic, non-mucoid strain were used. Model biofilms grown on polycarbonate filters were treated with lectins concanavalin A (ConA) and wheat germ agglutinin (WGA) that were fluorescently labelled with fluorescein isothiocyanate or tetramethyl rhodamine isothiocyanate. Fluorescently labelled ConA yielded cloud-like regions that were heterogeneously distributed within mucoid biofilms, whereas these structures were only rarely present in biofilms of the non-mucoid strain. The bacteria visualized with the fluorochrome SYTO 9 were localized both within and between the ConA-stained regions. In WGA-treated biofilms, the lectin was predominantly associated with bacterial cells. Alginate seemed to be involved in the interaction of ConA with the EPS matrix, since (i) pre-treatment of biofilms with an alginate lyase resulted in a loss of ConA biofilm staining, and (ii) using an enzyme-linked lectinsorbent assay (ELLA), ConA was shown to bind to purified alginate, but not to alginate that was degraded by alginate lyase. The application of fluorescently labelled lectins in combination with ELLA was found to be useful for the visualization and characterization of extracellular polysaccharide structures in P. aeruginosa biofilms.

Alginate overproduction affects Pseudomonas aeruginosa biofilm structure and function.

During the course of chronic cystic fibrosis (CF) infections, Pseudomonas aeruginosa undergoes a conversion to a mucoid phenotype, which is characterized by overproduction of the exopolysaccharide alginate. Chronic P. aeruginosa infections involve surface-attached, highly antibiotic-resistant communities of microorganisms organized in biofilms. Although biofilm formation and the conversion to mucoidy are both important aspects of CF pathogenesis, the relationship between them is at the present unclear. In this study, we report that the overproduction of alginate affects biofilm development on an abiotic surface. Biofilms formed by an alginate-overproducing strain exhibit a highly structured architecture and are significantly more resistant to the antibiotic tobramycin than a biofilm formed by an isogenic nonmucoid strain. These results suggest that an important consequence of the conversion to mucoidy is an altered biofilm architecture that shows increasing resistance to antimicrobial treatments.

Purification and crystallization of α-amylases from mucoid and non-mucoid B. amyloliquefaciens strains

The purified α-amylases from mucoid and non-mucoid B. amyloliquefaciens strains were crystallized in forms suitable for X-ray diffraction analysis. Crystals were grown by the hanging-drop vapor diffusion method. Very thin crystals of α-amylase of non-mucoid strain were obtained in the presence 15% propanol, 12% PEG 6000 and 0.1 M PIPES (pH: 7.1) at the end of 3 months and these were not suitable for X-ray diffraction analysis. Crystals of the mucoid strain of B. amyloliquefaciens α-amylase were obtained with 30% PEG 6000, 0.2 M (NH 4) 2SO 4 and 0.1 M PIPES (pH: 6.5) at the end of 3 months and these were suitable for X-ray diffraction analysis.

Comparison of two commercial systems (Vitek and MicroScan-WalkAway) for antimicrobial susceptibility testing of Pseudomonas aeruginosa isolates from cystic fibrosis patients

Antimicrobial susceptibility testing of cystic fibrosis (CF) isolates of Pseudomonas aeruginosa is difficult because the organisms are often mucoid and slow-growing. This study of 498 CF strains examined the correlation of results derived from two commonly used commercial systems (Vitek, MicroScan-WalkAway) with a reference method for 10 antimicrobials. Correlation to reference results was unacceptably low for all agents and both commercial systems had a high rate of very major (false-susceptible) errors. Although mucoid strains produced a 4.8% greater intermethod error, it was not markedly different than non-mucoid strains for the Vitek System. Overall, these tested commercial systems performed poorly for CF isolates in contrast to earlier reported, high correlations with the reference methods (broth microdilution frozen panels and agar dilution) of the National Committee for Clinical Laboratory Standards, the standardized disk diffusion test, and the Etest (AB BIODISK, Solna, Sweden).

In vitro pharmacodynamic properties of colistin and colistin methanesulfonate against Pseudomonas aeruginosa isolates from patients with cystic fibrosis.

The in vitro pharmacodynamic properties of colistin and colistin methanesulfonate were investigated by studying the MICs, time-kill kinetics, and postantibiotic effect (PAE) against mucoid and nonmucoid strains of Pseudomonas aeruginosa isolated from patients with cystic fibrosis. Twenty-three clinical strains, including multiresistant strains, and one type strain were selected for MIC determination. Eleven strains were resistant; MICs for these strains were >128 mg/liter. For the susceptible strains, MICs of colistin ranged from 1 to 4 mg/liter, while the MICs of colistin methanesulfonate were significantly higher and ranged from 4 to 16 mg/liter. The time-kill kinetics were investigated with three strains at drug concentrations ranging from 0.5 to 64 times the MIC. Colistin showed extremely rapid killing, resulting in complete elimination at the highest concentrations within 5 min, while colistin methanesulfonate killed more slowly, requiring a concentration of 16 times the MIC to achieve complete killing within 24 h. Colistin exhibited a significant PAE of 2 to 3 h at 16 times the MIC against the three strains after 15 min of exposure. For colistin methanesulfonate, PAEs were shorter at the concentrations tested. Colistin methanesulfonate had lower overall bactericidal and postantibiotic activities than colistin, even when adjusted for differences in MICs. Our data suggest that doses of colistin methanesulfonate higher than the recommended 2 to 3 mg/kg of body weight every 12 h may be required for the effective treatment of P. aeruginosa infections in cystic fibrosis patients.

Role of alginate and its O acetylation in formation of Pseudomonas aeruginosa microcolonies and biofilms.

Attenuated total reflection/Fourier transform-infrared spectrometry (ATR/FT-IR) and scanning confocal laser microscopy (SCLM) were used to study the role of alginate and alginate structure in the attachment and growth of Pseudomonas aeruginosa on surfaces. Developing biofilms of the mucoid (alginate-producing) cystic fibrosis pulmonary isolate FRD1, as well as mucoid and nonmucoid mutant strains, were monitored by ATR/FT-IR for 44 and 88 h as IR absorbance bands in the region of 2,000 to 1,000 cm(-1). All strains produced biofilms that absorbed IR radiation near 1,650 cm(-1) (amide I), 1,550 cm(-1) (amide II), 1,240 cm(-1) (P==O stretching, C---O---C stretching, and/or amide III vibrations), 1,100 to 1,000 cm(-1) (C---OH and P---O stretching) 1,450 cm(-1), and 1,400 cm(-1). The FRD1 biofilms produced spectra with an increase in relative absorbance at 1,060 cm(-1) (C---OH stretching of alginate) and 1,250 cm(-1) (C---O stretching of the O-acetyl group in alginate), as compared to biofilms of nonmucoid mutant strains. Dehydration of an 88-h FRD1 biofilm revealed other IR bands that were also found in the spectrum of purified FRD1 alginate. These results provide evidence that alginate was present within the FRD1 biofilms and at greater relative concentrations at depths exceeding 1 micrometer, the analysis range for the ATR/FT-IR technique. After 88 h, biofilms of the nonmucoid strains produced amide II absorbances that were six to eight times as intense as those of the mucoid FRD1 parent strain. However, the cell densities in biofilms were similar, suggesting that FRD1 formed biofilms with most cells at depths that exceeded the analysis range of the ATR/FT-IR technique. SCLM analysis confirmed this result, demonstrating that nonmucoid strains formed densely packed biofilms that were generally less than 6 micrometer in depth. In contrast, FRD1 produced microcolonies that were approximately 40 micrometer in depth. An algJ mutant strain that produced alginate lacking O-acetyl groups gave an amide II signal approximately fivefold weaker than that of FRD1 and produced small microcolonies. After 44 h, the algJ mutant switched to the nonmucoid phenotype and formed uniform biofilms, similar to biofilms produced by the nonmucoid strains. These results demonstrate that alginate, although not required for P. aeruginosa biofilm development, plays a role in the biofilm structure and may act as intercellular material, required for formation of thicker three-dimensional biofilms. The results also demonstrate the importance of alginate O acetylation in P. aeruginosa biofilm architecture.

Mucoid mutants of the biocontrol strain pseudomonas fluorescens CHA0 show increased ability in biofilm formation on mycorrhizal and nonmycorrhizal carrot roots.

Extracellular polysaccharides play an important role in the formation of bacterial biofilms. We tested the biofilm-forming ability of two mutant strains with increased production of acidic extracellular polysaccharides compared with the wild-type biocontrol strain Pseudomonas fluorescens CHA0. The anchoring of bacteria to axenic nonmycorrhizal and mycorrhizal roots as well as on extraradical mycelium of the arbuscular mycorrhizal fungus Glomus intraradices was investigated. The nonmucoid wild-type strain P. fluorescens CHA0 adhered very little on all surfaces, whereas both mucoid strains formed a dense and patchy bacterial layer on the roots and fungal structures. Increased adhesive properties of plant-growth-promoting bacteria may lead to more stable interactions in mixed inocula and the rhizosphere.

Carbohydrate sulfation effects on growth of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a key player in the pathology and morbidity of cystic fibrosis. Chronic obstructive pulmonary disease, which results from the most common and severe mutations in this genetic disorder, typically includes chronic infection with P. aeruginosa which, even with rugged antibiotic and physical therapy regimens, is rarely eradicated. It is not known whether the increased oligosaccharide sulfation characteristic of cystic fibrosis tracheobronchial mucins plays a role in the survival of P. aeruginosa in the airway. In this study, sulfated monosaccharides were synthesized and tested for their effects on the growth of clinical isolates and laboratory strains of this organism when supplied as the sole carbon source in vitro. Carbohydrate sulfation was observed to reduce, but not prohibit, growth of P. aeruginosa on carbohydrates normally utilized in their nonsulfated form. The various sulfated sugars employed as the sole carbon source gave characteristic and consistent growth profiles and maximum growth values across the strains tested. P. aeruginosa isolates from patients with cystic fibrosis often express a mucoid phenotype, which is thought to contribute to their ability to survive in harsh conditions. Carbohydrate sulfation effects on growth did not differ significantly between mucoid and nonmucoid strains. These results suggest that the additional sulfation of tracheobronchial mucin documented in cystic fibrosis may in fact contribute to the mucin's resistance to utilization by P. aeruginosa and potentially other pathogens, providing an additional level of host protection, and limiting the available nutrient pool and thereby bacterial growth.