Aeromonas punctataderived depolymerase that disrupts the integrity ofKlebsiella pneumoniaecapsule: optimization of depolymerase production

Abstract

Formation of dense, highly hydrated biofilm structures pose a risk for public and environmental health. Extracellular polymeric substances encompassing biofilms offer 1000-fold greater resistance as compared to the planktonic cells. Using enzymes as anti-biofouling agents, will improve penetration of antimicrobials and increase susceptibility of biofilms to components of immune system. The challenge of using enzymes derived from unrelated bacteria for the degradation of capsular matrix of Klebsiella pneumoniae has not been dealt in the past. Thus, statistical optimization was done to enhance depolymerase production by Aeromonas punctata, directed against the exopolysaccharide matrix of Klebsiella pneumoniae B5055, capable of substituting the available phage borne depolymerase enzyme. Optimization via central composite design (CCD) resulted in 16-fold enhancement in depolymerase yield (166.65 µmoles ml(-1) min(-1) ) over unoptimized medium. Out of the 19 variables, media composition giving maximum expression levels of the enzyme consisted of 1 mg ml(-1) galactose and ammonium chloride, 1.5 mg ml(-1) each of capsular polysaccharide (CPS) and magnesium sulfate. Tryptic peptide analysis of the purified 29 kDa band by Matrix assisted laser desorption ionization-time of flight (MALDI-TOF) showed a high homology with a protein of unknown function from Aeromonas cavaie Ae398. Further improvements in the enzyme can lead to its successful development as prophylactic and/or a therapeutic agent.

Optimization of the biological synthesis of silver nanoparticles using Penicillium oxalicum GRS-1 and their antimicrobial effects against common food-borne pathogens

“…A second-order model like central composite design (CCD) is widely used in RSM because it can take on a wide variety of functional forms, and such flexibility allows it to predict the true response surface more closely. This approach has already been successfully employed to maximize the productivities of various enzymes [42][43][44][45][46][47][48]. The major environmental factors that affect the production of Ag nanoparticles are pH and temperature.…”

Applications of bacteriophages versus phage enzymes to combat and cure bacterial infections: an ambitious and also a realistic application?

“…Moreover, LPS-degrading enzymes are also able to loosen biofilm structure as LPS-containing outer membrane vesicles are embedded within the matrix (Olszak et al 2017 ). There are many reports confirming the efficacy of depolymerases in eradication of biofilms formed by both Gram-positive and Gram-negative bacteria (Mushtaq et al 2004 , 2005 ; Cornelissen et al 2011 ; Gutiérrez et al 2012 , 2015 ); Bansal et al 2014 ; Pan et al 2015 ). Besides depolymerases, also phage lysins have been successfully used in the removal of bacterial biofilms.…”

Pathogens in Ornamental Waters: A Pilot Study

“…The production of depolymerise enzymes by A. punctata is responsible for disruption of K. pneumoniae capsule integrity causing biofilm dispersion. Although this fact has been only demonstrated for K. pneumoniae strains harbouring a K2 capsule, we cannot exclude the hypothesis of its occurence for other capsular phenotypes [ 20 ]. Furthermore the absence of K. pneumoniae after fish introduction in the fountain also supports the speculation that either the fish or their transport water could account for microbiota changes and appearance of Aeromonas spp.…”