Characterization of Bordetella pertussis growing as biofilm by chemical analysis and FT-IR spectroscopy

Abstract

Although Bordetella pertussis, the etiologic agent of whooping cough, adheres and grows on the ciliated epithelium of the respiratory tract, it has been extensively studied only in liquid cultures. In this work, the phenotypic expression of B. pertussis in biofilm growth is described as a first approximation of events that may occur in the colonization of the host. The biofilm developed on polypropylene beads was monitored by chemical methods and Fourier transform infrared (FT-IR) spectroscopy. Analysis of cell envelopes revealed minimal differences in outer membrane protein (OMP) pattern and no variation of lipopolysaccharide (LPS) expression in biofilm compared with planktonically grown cells. Sessile cells exhibited a 2.4- to 3.0-fold higher carbohydrate/protein ratio compared with different types of planktonic cells. A 1.8-fold increased polysaccharide content with significantly increased hydrophilic characteristics was observed. FT-IR spectra of the biofilm cells showed higher intensity in the absorption bands assigned to polysaccharides (1,200-900 cm(-1) region) and vibrational modes of carboxylate groups (1,627, 1,405, and 1,373 cm(-1)) compared with the spectra of planktonic cells. In the biofilm matrix, uronic-acid-containing polysaccharides, proteins, and LPS were detected. The production of extracellular carbohydrates during biofilm growth was not associated with changes in the specific growth rate, growth phase, or oxygen limitation. It could represent an additional virulence factor that may help B. pertussis to evade host defenses.

Proteome approaches combined with Fourier transform infrared spectroscopy revealed a distinctive biofilm physiology in Bordetella pertussis

“…Biofilms were grown on flat polypropylene beads (300 g; f = 4.2 mm, h = 1.5 mm, with an average density of 0.901 g/cm 3 , Petroken, Argentina) in glass column reactors (f = 6 cm, h = 45 cm, IVA SA, Argentina). Experiments were carried out essentially as described previously by Bosch et al [21]. Briefly, a planktonic culture of B. pertussis (200 mL, OD 650 = 1.0) grown in SS broth was used as inoculum for each column and incubated for 5 h at 377C to allow cell attachment to the beads.…”

“…In B. pertussis physiologic and pathogenic aspects have been extensively studied focusing on the planktonic mode of growth, while only a few works so far have considered the biofilm lifestyle of this bacterial pathogen [2,[20][21][22]. Therefore, generally only limited data is available on the nature of B. pertussis biofilm and information on a proteomic level is completely missing.…”

Can Near-infrared Spectroscopy Detect and Differentiate Implant-associated Biofilms?

“…In a proposed translational application we hope to identify not only the presence or absence of bacteria but also to identify a particular strain thus increasing the importance of improving Type II error. Previous studies have used spectroscopy to analyze biologic specimens [2,10,12,18,21]. Ngo-Thi et al [18] reported the ability of conventional FT-NIR spectroscopy in the MIR region and multivariate data analysis to correctly classify different nonbiofilm-associated bacteria in groups.…”

“…Specifically, we asked: (1) Can principal component analysis differentiate the spectral curves for biofilm obtained from Staphylococcus aureua, Staphylococcus epidermidis, and Pseudomonas aeruginosa? (2) What is the accuracy of FT-NIR/multivariate data analysis in identifying the specific species associated with biofilm?…”

“…Questions/purposes We asked: (1) Can principal component analysis explain variation in spectral curves for biofilm obtained from Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa? (2) What is the accuracy of Fourier transformed-near infrared (FT-NIR)/multivariate data analysis in identifying the specific species associated with biofilm? Methods Three clinical isolates, S aureus, S epidermidis, and P aeruginosa were cultured to create biofilm on surgical grade stainless steel.…”