Temporal Expression of Genes in Biofilm-Forming Ocular Candida albicans Isolated From Patients With Keratitis and Orbital Cellulitis

Abstract

Citation: Ranjith K, Kalyana Chakravarthy S, Adicherla H, Sharma S, Shivaji S. Temporal expression of genes in biofilm-forming ocular Candida albicans isolated from patients with keratitis and orbital cellulitis. Invest Ophthalmol Vis Sci. 2018;59:528-538. https://doi.org/ 10.1167/iovs.17-22933 PURPOSE. To study antibiotic susceptibility and biofilm-forming potential of ocular isolates of Candida albicans along with gene expression. METHODS.Seven clinical isolates of C. albicans (keratitis-6 and orbital cellulitis-1) were evaluated. Biofilm formation in one isolate was monitored by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Expression of 27 genes (real-time PCR) associated with biofilm formation and virulence was compared between biofilm-positive and biofilm-negative ocular C. albicans isolates. The temporal expression (4 to 72 hours) of the 27 overexpressed genes was also determined. Similar studies were also done with biofilmpositive and biofilm-negative nonocular C. albicans.RESULTS. Four of seven ocular C. albicans isolates exhibited the potential to form biofilm, one of which was resistant to three antifungals, whereas three were susceptible to all. SEM studies indicated that biofilm increased from two to three adherent layers of cells at 24 hours to multiple layers by 72 hours. CLSM showed that biofilm thickness increased from 5.2 lm at 24 hours to 17.98 lm at 72 hours. Upregulation of 27 genes involved in virulence and biofilm formation was observed both in the ocular and nonocular C. albicans positive for biofilm formation and compared to the respective non-biofilm-forming C. albicans. The results also indicated similarity in expression of genes between biofilm-forming ocular and nonocular pathogenic C. albicans. Temporal expression of the 27 genes (involved in adhesion, initiation, maturation, and dispersal stages of biofilm) in the biofilm-positive ocular isolate indicated that expression pattern followed four different patterns.CONCLUSIONS. This is the first study showing similarity in expression of genes in biofilmforming ocular and nonocular isolates of C. albicans, suggesting that upregulated genes could serve as a potential target for developing therapeutic strategies.

The role of fungi in fungal keratitis

“…Biofilms promote fungal adhesion and structural stability, whilst protecting the fungi from external threats. When compared to freeliving cells, biofilms are exhibit different phenotypic behaviors in growth rate, changes in gene expression and are often highly resistant to antifungal treatments and the host immune system (Hirota et al, 2017;Mukherjee and Chandra, 2004;Sandai et al, 2016, Ranjith et al, 2018.…”

“…Interestingly, not all FK fungal isolates are able to form biofilms. Recent studies by Ranjith et al (Ranjith et al, 2017(Ranjith et al, , 2018 determined that 42-47% of Candida FK isolates were unable to form biofilm, underscoring the heterogeneity of pathogens causing FK. In a study of 7 C. albicans isolates, four were able to form biofilm and only one exhibited multi-antifungal resistance, whereas the other three remained susceptible (Ranjith et al, 2018).…”

“…Recent studies by Ranjith et al (Ranjith et al, 2017(Ranjith et al, , 2018 determined that 42-47% of Candida FK isolates were unable to form biofilm, underscoring the heterogeneity of pathogens causing FK. In a study of 7 C. albicans isolates, four were able to form biofilm and only one exhibited multi-antifungal resistance, whereas the other three remained susceptible (Ranjith et al, 2018). 27 genes involved in virulence and biofilm formation were found to be temporally upregulated in the biofilm-forming Candida compared to the non-biofilm formers, thus targeting of these genes across the stages of biofilm development could serve as a therapeutic strategy.…”

Ocular Surface Infection Mediated Molecular Stress Responses: A Review

“…ZAG breaks down lipid in adipocytes but the exact role of ZAG in tears has not been studied intensively [ 82 ]. Biofilm-forming capability and antibiotic susceptibility of the ocular isolates of Candida albicans were studied and gene expression has been reported [ 83 ] in six keratitis and one orbital cellulitis clinical isolate. Biofilm formation was monitored by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM).…”

Amphotericin B Polymer Nanoparticles Show Efficacy against Candida Species Biofilms

“…Candida biofilms are usually associated with oral and vaginal candidiasis as well as with implanted medical devices such as vascular and urethral catheters, prosthetic heart valves, and dentures [3]. However biofilms have been implicated in ocular fungal infections, with an ocular biofilm reported in a patient with refractive infectious crystalline keratopathy due to C. albicans [11], biofilm forming C. albicans species isolated from patients [12], and biofilm forming keratitis patient isolates of Fusarium solani, C. sphaerospermum, and Acremonium Implicatum reported [13]. It is safe to assume that fungal biofilms are a feature of ocular fungal infections, especially those showing drug resistance.…”