Papers by Luis E. Chávez de Paz
Non-typeable Haemophilus influenzae (NTHi) is a common pathogen associated with diseases such as ... more Non-typeable Haemophilus influenzae (NTHi) is a common pathogen associated with diseases such as acute otitis media or exacerbations in patients with chronic obstructive pulmonary disease. The biofilmforming capability substantially contributes to the persistence of NTHi. However, the regulation of biofilm formation is not completely understood. Quorum sensing regulated by autoinducer-2 produced by luxS is until now the only described regulatory mechanism. In this study, we show that the two-component signalling system QseB/C is involved in the biofilm formation of NTHi in vitro. An isogenic NTHi mutant of qseC (Hi3655KR2) showed a significant decrease in biofilm formation under static and semi-static conditions as assessed by crystal violet staining. In addition, under constant flow conditions, Hi3655KR2 formed less biofilm after 48 h. The biofilm defects were irrespective of autoinducer-2 levels. Hence, here we suggest for the first time a regulatory circuit in NTHi, which controls biofilm formation by mechanisms other than or in addition to luxS-dependent factors.
BMC Oral Health, 2013
Background: Mucosal surfaces are coated with layers of mucus gel that protect the underlying tiss... more Background: Mucosal surfaces are coated with layers of mucus gel that protect the underlying tissues and promote colonization by members of the commensal microflora. Lactobacillus fermentum is a common inhabitant of the oral cavity, gastrointestinal and reproductive tracts and is one of the most important lactic acid bacteria contributing to the formation of a healthy intestinal microflora. We have investigated the proteolytic activity in L. fermentum in response to interactions with the MUC5B mucin, which is a major component of mucus gels at sites colonized by this micro-organism. Methods: Biofilms of Lactobacillus fermentum were established in mini-flow cells in the presence or absence of human salivary MUC5B. The proteolytic activity of biofilm cells was examined in a confocal scanning laser microscope with a fluorescent protease substrate. Degradation of MUC5B by L. fermentum was analysed using SDS-PAGE followed by Western blotting with antisera raised against the MUC5B peptide. Cell surface proteins differentialy expressed in a MUC5B-rich environment were identified with the aid of comparative two-dimensional electrophoresis followed by LC-MS/MS.
Journal of Endodontics, 2007
Current concepts suggest that persisting infections subsequent to endodontic therapy are caused b... more Current concepts suggest that persisting infections subsequent to endodontic therapy are caused by one or two bacterial species that are "too robust" to be eliminated by conventional treatment measures. As a consequence, numerous studies are exploring the characteristics of these "most" resistant organisms to define an effective treatment strategy to eradicate them from root canals. By taking an ecological perspective, the main objective of this review is to present evidence that the nature of persisting endodontic infections depends not on the robustness of the organisms in the infected site, but on their capability of adapting their physiology to the new environmental conditions set by the treatment. Changes in the environment, such as an increase in pH by calcium hydroxide or the effect of antimicrobials, are capable of triggering genetic cascades that modify the physiological characteristics of bacterial cells. Surface adherence by bacteria to form biofilms is a good example of bacterial adaptation and one that is pertinent to endodontic infections. Increasing information is now available on the existence of polymicrobial biofilm communities on root canal walls, coupled with new data showing that the adaptive mechanisms of bacteria in these biofilms are significantly augmented for increased survival. This ecological view on the persisting infection problem in endodontics suggests that the action of individual species in persisting endodontic infections is secondary when compared to the adaptive changes of a polymicrobial biofilm community undergoing physiological and genetic changes in response to changes in the root canal environment. (J Endod 2007; 33:652-662)
Caries Research, 2011
In this study we evaluated the effect of chitosan nanoparticles on the acid tolerance response (A... more In this study we evaluated the effect of chitosan nanoparticles on the acid tolerance response (ATR) of adhered Streptococcus mutans. An ATR was induced by exposing S. mutans to pH 5.5 for 2 h and confirmed by exposing the acid-adapted cells to pH 3.5 for 30 min, with the majority of cells appearing viable according to the LIVE/DEAD® technique. However, when chitosan nanoparticles were present during the exposure to pH 5.5, no ATR occurred as most cells appeared dead after the pH 3.5 shock. We conclude that the chitosan nanoparticles tested had the ability to hinder ATR induction in adhered S. mutans.
International Endodontic Journal, 2003
Cha¤ vez de Paz LE, Dahle¤ n G, Molander A, Mo « ller -, Bergenholtz G. Bacteria recovered from t... more Cha¤ vez de Paz LE, Dahle¤ n G, Molander A, Mo « ller -, Bergenholtz G. Bacteria recovered from teeth with apical periodontitis after antimicrobial endodontic treatment. International Endodontic Journal, 36, 500^508, 2003.
Oral Surgery Oral Medicine Oral Pathology Oral Radiology and Endodontology, 2005
Objectives. The object of this study was to investigate the diversity among streptococcal species... more Objectives. The object of this study was to investigate the diversity among streptococcal species isolated from root canals in conjunction with endodontic therapy and to characterize their production of extracellular proteins.
BMC Oral Health, 2011
Background: The soft tissue around dental implants forms a barrier between the oral environment a... more Background: The soft tissue around dental implants forms a barrier between the oral environment and the periimplant bone and a crucial factor for long-term success of therapy is development of a good abutment/soft-tissue seal. Sol-gel derived nanoporous TiO 2 coatings have been shown to enhance soft-tissue attachment but their effect on adhesion and biofilm formation by oral bacteria is unknown. Methods: We have investigated how the properties of surfaces that may be used on abutments: turned titanium, sol-gel nanoporous TiO 2 coated surfaces and anodized Ca 2+ modified surfaces, affect biofilm formation by two early colonizers of the oral cavity: Streptococcus sanguinis and Actinomyces naeslundii. The bacteria were detected using 16S rRNA fluorescence in situ hybridization together with confocal laser scanning microscopy.
Oral Surgery Oral Medicine Oral Pathology Oral Radiology and Endodontology, 2002
Objective. The extent to which Fusobacterium nucleatum is recovered from root canals of teeth tha... more Objective. The extent to which Fusobacterium nucleatum is recovered from root canals of teeth that present with an interappointment flare-up following endodontic instrumentation was investigated.
International Endodontic Journal, 2007
Aim To determine whether bacteria isolated from infected root canals survive alkaline shifts bet... more Aim To determine whether bacteria isolated from infected root canals survive alkaline shifts better in biofilms than in planktonic cultures.Methodology Clinical isolates of Enterococcus faecalis, Lactobacillus paracasei, Olsenella uli, Streptococcus anginosus, S. gordonii, S. oralis and Fusobacterium nucleatum in biofilm and planktonic cultures were stressed at pH 10.5 for 4 h, and cell viability determined using the fluorescent staining LIVE/DEAD BacLight bacterial viability kit. In addition, proteins released into extracellular culture fluids were identified by Western blotting.Results Enterococcus faecalis, L. paracasei, O. uli and S. gordonii survived in high numbers in both planktonic cultures and in biofilms after alkaline challenge. S. anginosus, S. oralis and F. nucleatum showed increased viability in biofilms compared with planktonic cultures. Alkaline exposure caused all planktonic cultures to aggregate into clusters and resulted in a greater extrusion of cellular proteins compared with cells in biofilms. Increased levels of DnaK, HPr and fructose-1,6-bisphosphate aldolase were observed in culture fluids, especially amongst streptococci.Conclusions In general, bacteria isolated from infected roots canals resisted alkaline stress better in biofilms than in planktonic cultures, however, planktonic cells appeared to use aggregation and the extracellular transport of specific proteins as survival mechanisms.
International Endodontic Journal, 2004
Chá vez de Paz LE, Molander A, Dahlé n G. Gram-positive rods prevailing in teeth with apical peri... more Chá vez de Paz LE, Molander A, Dahlé n G. Gram-positive rods prevailing in teeth with apical periodontitis undergoing root canal treatment. International Endodontic Journal, 37, 579-587, 2004.
Applied and Environmental Microbiology, 2009
The novel image analysis software package bioImage_L was tested to calculate biofilm structural p... more The novel image analysis software package bioImage_L was tested to calculate biofilm structural parameters in oral biofilms stained with dual-channel fluorescent markers. By identifying color tonalities in situ, the software independently processed the color subpopulations and characterized the viability and metabolic activity of biofilms.
Fems Immunology and Medical Microbiology, 2010
Pseudomonas aeruginosa is often found in chronic infections, including cystic fibrosis lung infec... more Pseudomonas aeruginosa is often found in chronic infections, including cystic fibrosis lung infections and those related to chronic wounds and venous ulcers. At the latter sites, P. aeruginosa can be isolated together with Staphylococcus epidermidis, and we have therefore explored the effect of clinical isolates and laboratory strains of P. aeruginosa strains on colonization by S. epidermidis in dual-species biofilms. Biofilm formation was assayed using 16S rRNA FISH and confocal laser scanning microscopy. Among the six P. aeruginosa strains tested, one particular strain, denoted 14:2, exerted a significant inhibitory effect, and even after 6 h, S. epidermidis levels in dual-species biofilms were reduced by >85% compared with those without P. aeruginosa. Interestingly, strain 14:2 was found to be negative for classical virulence determinants including pyocyanin, elastase and alkaline protease. Therefore, we suggest that less virulent phenotypes of P. aeruginosa, which may develop over time in chronic infections, could counteract colonization by S. epidermidis, ensuring persistence and dominance by P. aeruginosa in the host micro-habitat. Further studies are required to explain the inhibitory effect on S. epidermidis, although extracellular polysaccharides produced by P. aeruginosa might play a role in this phenomenon.
Fems Immunology and Medical Microbiology, 2010
Pseudomonas aeruginosa and Staphylococcus epidermidis are common opportunistic pathogens associat... more Pseudomonas aeruginosa and Staphylococcus epidermidis are common opportunistic pathogens associated with medical device-related biofilm infections. 16S rRNA-FISH and confocal laser scanning microscopy were used to study these two bacteria in dual-species biofilms. Two of the four S. epidermidis strains used were shown to form biofilms more avidly on polymer surfaces than the other two strains. In dual-species biofilms, the presence of P. aeruginosa reduced biofilm formation by S. epidermidis, although different clinical isolates differed in their susceptibility to this effect. The most resistant isolate coexisted with P. aeruginosa for up to 18 h and was also resistant to the effects of the culture supernatant from P. aeruginosa biofilms, which caused dispersal from established biofilms of other S. epidermidis strains. Thus, different strains of S. epidermidis differed in their capacity to withstand the action of P. aeruginosa, with some being better equipped than others to coexist in biofilms with P. aeruginosa. Our data suggest that where S. epidermidis and P. aeruginosa are present on abiotic surfaces such as medical devices, S. epidermidis biofilm formation can be inhibited by P. aeruginosa through two mechanisms: disruption by extracellular products, possibly polysaccharides, and, in the later stages, by cell lysis.
Journal of Endodontics, 2010
Introduction: In the present study, confocal microscopy, a miniflow cell system, and image analys... more Introduction: In the present study, confocal microscopy, a miniflow cell system, and image analysis were combined to test in situ the effect of antimicrobials and alkali on biofilms of Enterococcus faecalis, Lactobacillus paracasei, Streptococcus anginosus, and Streptococcus gordonii isolated from root canals with persistent infections. Methods: Biofilms formed for 24 hours were exposed for 5 minutes to alkali (pH = 12), chlorhexidine digluconate (2.5%), EDTA (50 mmol/L), and sodium hypochlorite (1%). The biofilms were then characterized by using fluorescent markers targeting cell membrane integrity (LIVE/DEAD) and metabolic activity (5-cyano-2,3ditolyl tetrazolium chloride and fluorescein diacetate). In addition, the biofilm architecture and the extent to which coating of the substrate surface with collagen influenced the resistance pattern to the chemicals were also analyzed. Results: NaOCl (1%) affected the membrane integrity of all organisms and removed most biofilm cells. Exposure to EDTA (50 mmol/L) affected the membrane integrity in all organisms but failed to remove more than a few cells in biofilms of E. faecalis, L. paracasei, and S. anginosus. Chlorhexidine (2.5%) had a mild effect on the membrane integrity of E. faecalis and removed only 50% of its biofilm cells The effects were substratumdependent, and most organisms displayed increased resistance to the antimicrobials on collagen-coated surfaces. Conclusions: The biofilm system developed here was sensitive and differences in cell membrane integrity and removal of biofilm cells after exposure to antimicrobials commonly used in endodontics was discernible. (J Endod 2010;36:70-77)
Microbiology-sgm, 2008
The ability of oral bacteria to enter a non-growing state is believed to be an important mechanis... more The ability of oral bacteria to enter a non-growing state is believed to be an important mechanism for survival in the starved micro-environments of the oral cavity. In this study, we examined the reactivation of nutrient-deprived cells of two oral bacteria in biofilms, Streptococcus anginosus and Lactobacillus salivarius. Non-growing cells were generated by incubation in 10 mM potassium phosphate buffer for 24 h and the results were compared to those of planktonic cultures. When both types of cells were shifted from a rich, peptone-yeast extract-glucose (PYG) medium to buffer for 24 h, dehydrogenase and esterase activity measured by the fluorescent dyes 5-cyano-2,3ditolyl-tetrazolium chloride (CTC) and fluorescein diacetate (FDA), respectively, was absent in both species. However, the membranes of the vast majority of nutrient-deprived cells remained intact as assessed by LIVE/DEAD staining. Metabolic reactivation of the nutrient-deprived biofilm cells was not observed for at least 48 h following addition of fresh PYG medium, whereas the nongrowing planktonic cultures of the same two strains were in rapid growth in less than 2 h. At 72 h, the S. anginosus biofilm cells had recovered 78 % of the dehydrogenase activity and 61 % of the esterase activity and the biomass mm "2 had increased by 30-35 %. With L. salivarius at 72 h, the biofilms had recovered 56 % and 75 % of dehydrogenase and esterase activity, respectively. Reactivation of both species in biofilms was enhanced by removal of glucose from PYG, and S. anginosus cells were particularly responsive to yeast extract (YE) medium. The data suggest that the low reactivity of non-growing biofilm cells to the introduction of fresh nutrients may be a survival strategy employed by micro-organisms in the oral cavity.
Current concepts suggest that persisting infections subsequent to endodontic therapy are caused b... more Current concepts suggest that persisting infections subsequent to endodontic therapy are caused by one or two bacterial species that are "too robust" to be eliminated by conventional treatment measures. As a consequence, numerous studies are exploring the characteristics of these "most" resistant organisms to define an effective treatment strategy to eradicate them from root canals. By taking an ecological perspective, the main objective of this review is to present evidence that the nature of persisting endodontic infections depends not on the robustness of the organisms in the infected site, but on their capability of adapting their physiology to the new environmental conditions set by the treatment. Changes in the environment, such as an increase in pH by calcium hydroxide or the effect of antimicrobials, are capable of triggering genetic cascades that modify the physiological characteristics of bacterial cells. Surface adherence by bacteria to form biofilms is a good example of bacterial adaptation and one that is pertinent to endodontic infections. Increasing information is now available on the existence of polymicrobial biofilm communities on root canal walls, coupled with new data showing that the adaptive mechanisms of bacteria in these biofilms are significantly augmented for increased survival. This ecological view on the persisting infection problem in endodontics suggests that the action of individual species in persisting endodontic infections is secondary when compared to the adaptive changes of a polymicrobial biofilm community undergoing physiological and genetic changes in response to changes in the root canal environment. (J Endod 2007; 33:652-662)
Uploads
Papers by Luis E. Chávez de Paz