Biofilm formation by Aspergillus flavus and Aspergillus niger: Influence of cultural conditions and their controls
Aspergillus organisms have biofilm-forming abilities which make their control with some routine food-contact sanitizers or antifungal agents, more complicated. Studies on biofilm forming abilities of Aspergillus spp and its control are limited. This study was designed to investigate biofilm development by isolates of Aspergillus flavus and Aspergillus niger from meat tables in meat markets and abattoirs in Ibadan. Also, variations in cultural conditions, Lactobacillus rhamnosus GG, and sodium hypochlorite were assessed as measures for their control. Laboratory stock cultures of A. flavus and A. niger from abattoir environment were cultured by pour-plate method at 105 dilution on Sucrose-Potato dextrose agar, supplemented with streptomycin, and incubated at 26oC for five days, and organisms were enumerated using pre-described standard microbiological methods. The influence of two sugars (sucrose and fructose), at 0.2% and 0.4% concentrations; sodium hypochlorite (NaOCl) at 0.05% and 0.5% concentrations, and Lactobacillus rhamnosus GG at 108 and 104 concentrations were investigated on biofilm development by A. niger (n=2) and A. flavus (n=2) isolates, using Crystal Violet Binding Assay. Biofilm development using overnight grown culture was done in 96-well micro-titer plates, while the biofilm mass was determined by measuring the absorbance at 600 nm. An un-inoculated broth served as control. The experiment was done in at least three replicates. Data were analysed using ANOVA at α0.05. A. flavus (0.10±0.02) significantly produced a higher biofilm mass than A. niger (0.09±0.01). There was significant increase in biofilm mass with the addition of sucrose and fructose, with more biofilm mass (0.14±0.08 for sucrose and 0.11±0.03 for fructose) observed at 0.2% concentration. Lactobacillus rhamnosus GG significantly decreased biofilm mass of A. flavus (0.07±0.01) and A. niger (0.06±0.00) when compared to the inoculated broth culture without lactic acid bacteria (0.09±0.01). There was a significant reduction in biofilm mass with both 0.05% and 0.5% sodium hypochlorite, with the highest reduction seen with 0.5% concentration at 24 hr. A significant decrease in biofilm mass was observed for A. niger (106) with 0.5% concentration of NaOCl at 24 hr (0.07±0.01) when compared to control (0.09±0.01). Similar results were obtained across species at 24 and 72 hr of incubation. A lower biofilm mass was also formed at 11oC (0.07±0.01) when compared with 28oC (0.10±0.01). Sodium hypochlorite suppressed biofilm development, thus showing potentials for eliminating Aspergillus biofilms from meat tables in commercial markets, while Lactobacillus rhamnosus GG may be employed as an additive in meat and meat products processing.