Yulong Tan

Antibiofilm activity of carboxymethyl chitosan on the biofilms of non-Candida albicans Candida species

Although most cases of candidiasis have been attributed to Candida albicans, non-C. albicans Candida species have been isolated in increasing numbers in patients. In this study, we determined the inhibition of carboxymethyl chitosan (CM-chitosan) on single and mixed species biofilm of non-albicans Candida species, including Candida tropicalis, Candida parapsilosis, Candida krusei and Candida glabrata. Biofilm by all tested species in microtiter plates were inhibited nearly 70%. CM-chitosan inhibited mixed species biofilm in microtiter plates and also on medical materials surfaces. To investigate the mechanism, the effect of CM-chitosan on cell viability and biofilm growth was employed. CM-chitosan inhibited Candida planktonic growth as well as adhesion. Further biofilm formation was inhibited with CM-chitosan added at 90min, 12h or 24h after biofilm initiation. CM-chitosan was not only able to inhibit the metabolic activity of Candida cells, but was also active upon the establishment and the development of biofilms.

Inhibition of mixed fungal and bacterial biofilms on silicone by carboxymethyl chitosan

Mixed biofilms with fungi and bacteria are the leading cause for the failure of medical silicone devices, such as voice prostheses in laryngectomy. In this study, we determined the effect of carboxymethyl chitosan (CM-chitosan) on mixed biofilm formation of fungi and bacteria on silicone which is widely used for construction of medical devices. Mixed biofilm formations were inhibited 72.87% by CM-chitosan. Furthermore, CM-chitosan significantly decreased the metabolic activity of the biofilms using 2, 3-bis (2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5 carboxanilide (XTT) reduction assay. The examination using confocal laser scanning microscopy and scanning electron microscope confirmed that CM-chitosan inhibited the mixed biofilm and damaged the cells. Effects of CM-chitosan on different stages of biofilms were also evaluated. CM-chitosan inhibited the adhesion of fungi and bacteria with an efficiency of >90%. It prevented biofilm formation at efficiencies of 69.86%, 50.88% and 46.58% when CM-chitosan was added at 90min, 12h and 24h after biofilm initiation, respectively. Moreover, CM-chitosan inhibited Candida yeast-to-hyphal transition. CM-chitosan was not only able to inhibit the metabolic activity of biofilms, but also active upon the establishment and development of biofilm. Therefore, CM-chitosan may serve as a possible antibiofilm agent to limit biofilm formation on voice prostheses.

Influence of culture conditions for clinically isolated non-albicans Candida biofilm formation

Non-albicans Candida species have been isolated in increasing numbers in patients. Moreover, they are adept at forming biofilms. This study analyzed biofilm formation of clinically isolated non-albicans Candida, including Candida tropicalis, Candida krusei and Candida parapsilosis under the influence of different growth media (RPMI 1640, YPD and BHI) and several culture variables (inoculum concentration, incubation period and feeding conditions). The results showed that culture conditions strongly influenced non-albicans Candida species biofilm formation. YPD and BHI resulted in larger amount of biofilm formation with higher metabolic activity of biofilms. Furthermore, the growth media seems to have varying effects on adhesion and biofilm development. Growth conditions may also influence biofilm formation, which was enhanced when starting the culture with a larger inoculum, longer incubation period and using a fed-batch system. Therefore, the potential influences of external environmental factors should be considered when studying the non-albicans Candida biofilms in vitro.

Long-term antibiofilm activity of carboxymethyl chitosan on mixed biofilm on silicone.

Silicone voice prostheses are most frequently used in voice rehabilitation of laryngectomized patients. However, the functional device lifetimes are limited due to formation of mixed biofilms. Existing in vitro models simulating biofilm formation are restricted to only short‐term periods.

In vitro biofilm growth on modern voice prostheses

Biofilm formation on voice prostheses in laryngectomized patients usually limits the lifetime of the device. The purpose of this study was to compare the biofilm resistance of different valve flaps of modern voice prostheses in an in vitro simulation of an oropharyngeal biofilm.

β-1,3-glucanase disrupts biofilm formation and increases antifungal susceptibility of Candida albicans DAY185

β-1,3-glucan plays a role in Candida biofilm formation and survival of biofilm-forming Candida to stresses. In this study, we evaluated the antibiofilm activity of β-1,3-glucanase, which can degrade poly-β(1→3)-glucose of Candida albicans biofilms. Biofilm was dispersed by 55.96%. β-1,3-glucanase had no effect on Candida planktonic growth as well as adhesion. β-1,3-glucanase markedly enhanced the antifungal susceptibility of fluconazole and amphotericin B. The examination using confocal laser scanning microscopy and scanning electron microscope confirmed the antibiofilm activity of β-1,3-glucanase. Our findings demonstrate that β-1,3-glucanase may be useful as an antibiofilm agent.

Efficacy of carboxymethyl chitosan against Candida tropicalis and Staphylococcus epidermidis monomicrobial and polymicrobial biofilms

Polymicrobial biofilms with fungi and bacteria are the leading cause for the failure of medical devices and related infections. In this study, antibiofilm activities of carboxymethyl chitosan (CM-chitosan) on monomicrobial and polymicrobial biofilms of Staphylococcus epidermidis and Candida tropicalis in vitro were evaluated. CM-chitosan was effective as a sole agent, inhibiting both monomicrobial and polymicrobial biofilms in microplates and also on the silicone surface in short- and long-term periods. Biofilm architecture was investigated by scanning electron microscopy and confocal laser scanning microscopy was used to examine living/dead organisms within biofilm. CM-chitosan inhibited planktonic growth as well as adhesion. Further biofilm formation was inhibited by CM-chitosan added at 90min or 12h after biofilm initiation. CM-chitosan may serve as a possible antibiofilm agent to limit monomicrobial and polymicrobial biofilm.

Evaluation of combined growth media for in vitro cultivation of oropharyngeal biofilms on prosthetic silicone

AbstractIn the upper aerodigestive tract, biofilm deposits by oropharyngeal microbes can cause failure of medical polymer devices like voice prostheses. Previous studies on testing of inhibitive strategies still lack of comparability due to varying study protocols concerning growth media, microbial species and growth conditions. Goal of the study was therefore to test cultivation of a mixed biofilm of isolated oropharyngeal microbes under in vitro growth conditions using mixtures of common growth media. Mixtures of yeast peptone dextrose medium (YPD), fetal bovine serum (FBS), RPMI 1640, Yeast nitrogen base medium (YNB) and brain heart infusion (BHI) were tested to grow mixed biofilm deposits of Candida albicans, Candida tropicalis, Staphylococcus aureus, Streptococcus epidermidis, Rothia dentocariosa and Lactobacillus gasseri on medical grade silicone. Periodic assessment of living biofilm was performed over 22 days by a digital microscope and the cultivated biofilm structures were analyzed by scanning electron microscopy after completion of the study. Mixtures of BHI, YPD and FBS improved microscopic growth of multispecies biofilm deposits over time, while addition of RPMI and YNB resulted in reduction of visible biofilm deposit sizes. A mixtures of FBS 30% + YPD 70% and BHI 30% + YPD 70% showed enhanced support of permanent surface growth on silicone. Growth kinetics of in vitro multispecies biofilms can be manipulated by using mixtures of common growth media. Using mixtures of growth media can improve growth of longterm multispecies oropharyngeal biofilm models used for in vitro testing of antibiofilm materials or coatings for voice prostheses.

Characterization of a new endo-type polysaccharide lyase (PL) family 6 alginate lyase with cold-adapted and metal ions-resisted property

Alginate lyase played an important role in brown algae degradation, and its enzymatic degradation products showed various biological activities. Although many alginate lyases have been characterized, the enzymes with special characterizations are still rather rare. In this study, a new alginate lyase gene, tsaly6A, has been cloned from marine bacterium Thalassomonas sp. LD5, and expressed in Escherichia coli. The deduced alginate lyase, TsAly6A, belonged to the polysaccharide lyase (PL) family 6 and showed the highest amino acid identity (63%) with an exo-type oligoalginate lyase AlyGC. However, this study showed that TsAly6A was an endo-type enzyme yielding alginate trisaccharides (64.5%) as the main products. Compared with other alginate lyases, TsAly6A showed high trisaccharide-yielding levels. Meanwhile, TsAly6A showed the specific activity of 15,960 U/μmol at its optimal pH (pH 8.0) and temperature (35 °C). In addition, TsAly6A was a cold-adapted, salt-activated and metal ions-resisted alginate lyase, which will enable it to perform high activity in the solution containing various ions. Its cold-adaptation, metal ions-tolerance and high trisaccharides yields make TsAly6A an excellent candidate for industrial applications.

Enhancing antibiofilm activity with functional chitosan nanoparticles targeting biofilm cells and biofilm matrix

Bacterial biofilms play a key role during infections, which are associated with an increased morbidity and mortality. The classical antibiotic therapy cannot eradicate biofilm-related infections because biofilm bacteria display high drug resistance due to biofilm matrix. Thus, novel drug delivery to overcome biofilm resistance and eliminate biofilm-protected bacteria is needed to be developed. In this study, positively charged chitosan nanoparticles (CSNP) loaded with oxacillin (Oxa) and Deoxyribonuclease I (CSNP-DNase-Oxa) were fabricated. The antibiofilm activity was evaluated against Staphylococcus aureus biofilms. Biofilm architecture on silicone surfaces was investigated by scanning electron microscopy (SEM). Confocal laser scanning microscopy (CLSM) was used to examine live/dead organisms within biofilm. CSNP-DNase-Oxa exhibited higher antibiofilm activity than Oxa-loaded nanoparticles without DNase (CSNP-Oxa) and free Oxa (Oxa and Oxa + DNase) at each concentration in all in-vitro tests. CSNP-DNase-Oxa inhibited biofilm formation in-vitro and eradicated mature biofilm effectively. CSNP-DNase-Oxa could disrupt the biofilm formation through degradation of eDNA, reduced biofilm thickness and the amount of viable cells on silicone. Repeated treatment with CSNP-DNase-Oxa for two days resulted in 98.4% biofilm reduction. Moreover, CSNP-DNase-Oxa was not only able to affect the biofilm of a standard S. aureus strain, but also showed the highest eradication of biofilms of clinical isolates compared with control groups. These results suggest the potential applicability of NPs for the treatment of biofilm-related infections and provide a platform for designing novel drug delivery with more functions.