Serratiopeptidase Affects the Physiology of Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients

Abstract

Pseudomonas aeruginosa is frequently involved in cystic fibrosis (CF) airway infections. Biofilm, motility, production of toxins and the invasion of host cells are different factors that increase P. aeruginosa’s virulence. The sessile phenotype offers protection to bacterial cells and resistance to antimicrobials and host immune attacks. Motility also contributes to bacterial colonization of surfaces and, consequently, to biofilm formation. Furthermore, the ability to adhere is the prelude for the internalization into lung cells, a common immune evasion mechanism used by most intracellular bacteria, such as P. aeruginosa. In previous studies we evaluated the activity of metalloprotease serratiopeptidase (SPEP) in impairing virulence-related properties in Gram-positive bacteria. This work aimed to investigate SPEP’s effects on different physiological aspects related to the virulence of P. aeruginosa isolated from CF patients, such as biofilm production, pyoverdine and pyocyanin production and invasion in alveolar epithelial cells. Obtained results showed that SPEP was able to impair the attachment to inert surfaces as well as adhesion/invasion of eukaryotic cells. Conversely, SPEP’s effect on pyocyanin and pyoverdine production was strongly strain-dependent, with an increase and/or a decrease of their production. Moreover, SPEP seemed to increase swarming motility and staphylolytic protease production. Our results suggest that a large number of clinical strains should be studied in-depth before drawing definitive conclusions. Why different strains sometimes react in opposing ways to a specific treatment is of great interest and will be the object of future studies. Therefore, SPEP affects P. aeruginosa’s physiology by differently acting on several bacterial factors related to its virulence.

Virulence of Pseudomonas aeruginosa in Cystic Fibrosis: Relationships between Normoxia and Anoxia Lifestyle

“…Pyocyanin production was determined as previously described [31]. Briefly, bacterial cells were inoculated in BHI broth and aerobically incubated for different times at 37 • C. The cell-free supernatant, recovered after centrifugation at 10,000 rpm for 15 min, was used for pyocyanin extraction.…”

“…For pyoverdine quantification, bacteria were grown in King's B Medium supplemented with 0.5% (w/v) of Casamino Acid (CAA) at 37 • C in aerobic and anaerobic conditions [31]. Pyoverdine was quantified by reading 100 µL of each P. aeruginosa cell-free supernatant into a black 96-well plate (Greiner, Stonehouse, UK) at excitation and emission wavelengths of 400/460 nm, as previously reported [32], on an Infinite 200 PRO (Molecular Devices, San Jose, CA, USA) fluorescence microplate reader.…”

Secreted Metabolites from Pseudomonas, Staphylococcus, and Borrelia Biofilm: Modulation of Immunogenicity by a Nutraceutical Enzyme and Botanical Blend

“…Non-pharmaceutical enzymatic treatments have been researched to disrupt biofilm formation by inhibiting new biofilm formation, decreasing bacterial viability, and breaking down the biofilm matrix [29,30]. Pa biofilm can be disrupted by various enzymes such as lipase [31], cellulase [32], and serratiopeptidase [33]. Lysozyme is an antimicrobial enzyme, part of our innate immune system [34].…”

Scale-Up of the Fermentation Process for the Production and Purification of Serratiopeptidase Using Silkworm Pupae as a Substrate

“…In this context, serratiopeptidase, a protease (hydrolytic enzyme), is obtained from bacteria such as Serratia marcescens [ 17 , 18 , 19 ]. This enzyme is renowned for its anti-inflammatory, fibrinolytic, antibiofilm, and analgesic effects [ 20 , 21 , 22 , 23 , 24 , 25 ]. The production of serratiopeptidase has been reported using various agro-industrial residues, such as wheat husk and rice bran [ 7 ].…”