Screening of antibiofilm and anti-quorum sensing activty of Actinomycetes isolates extracts against aquaculture pathogenic bacteria

Abstract

Background: Indonesia is the third largest producer of fish and other aquaculture products in the world, making this industry a major contributor in the economy of Indonesia. However, this industry continually overcome challenges, one of them are bacterial outbreaks. In addition, the emergence of these bacterial outbreaks were worsen due to the biofilm produced by many significant pathogenic bacteria and the impact of increased antibiotic resistance. These issues have become a global concern, because antibiotics are currently one of the main treatments available to overcome this problems. Therefore, studies aimed at finding and characterizing bioactive compounds to combat these issues. In this study actinomycetes isolates were screened and characterized for their bioactive compounds produced which have inhibitory and destructive activity and also QS inhibitors against biofilm structure of aquatic pathogenic bacteria, such as Vibrio harveyi, A. hydrophila, and S. agalactiae.

Result: Extracts (20 mg/mL) produced by sixteen Actinomycetes isolates showed anti-quorum sensing activity towards reporter stain Chromobacterium violaceum wild-type. Most of these extracts showed better inhibitory activity on all of the pathogenic bacteria biofilm structure tested than the destructive activity on the preformed of those biofilm structure. Subsequently, we also performed characterization of bioactive compound and found that in this study, polysaccharide is the most common antibiofilm agents, which were responsible to their antibiofilm activity. Finally, we found that the value of LC₅₀ of all extracts tested were more than 1 mg/mL, thereby all of extracts tested did not show cyto-toxic effect against Artemia salina.

Conclusion: All of the extracts of Actinomycetes isolates showed promising inhibitory activity towards biofilm structure of pathogenic bacteria tested. So far, all of the extracts are potential to be QS inhibitors and antibiofilm agents of all pathogenic bacteria tested.

Keywords: Actinomyces; Antibiofilm; Aquatic pathogen; Biofilm; Quorum sensing.

Fig. 1

Result of screening of anti-quorum sensing activity of several Actinomycetes isolates (a: 18 PM; b: 20 PM; c: SW03)

Fig. 2

Result of detection of anti-quorum sensing activity of a) Isolate 1 AC b) Isolate 20 PM against C. violaceum wild-type.*The top well is a positive-control; well in the middle of the plate containing the extract of Actinomycetes isolates; and the lowest well is a negative-control

Fig. 3

Result of antibacterial assay a) Isolate SW17 against Vibrio harveyi b) Isolate 16 PM against Streptococcus agalactiae c) Isolate CW01 against Streptococcus agalactiae .*The top well is a positive-control; well in the middle of the plate containing the extract of Aktinomycetes isolates; and the lowest well is a negative-control

Fig. 4

Characterization of extracts produced by isolates 20 PM (a, b, c) and TB12 (d, e, f) which is responsible for the inhibitory activity towards biofilm structure of A. hydrophila by treating the extract with NaIO₄ (20 mM), proteinase K (1 mg/mL), and DNase I (100 mg/mL). *Cr = Crude extracts

Fig. 5

Characterization of extracts produced by isolates 11 AC (a, b, c) and TB12 (d, e, f) which is responsible for the inhibitory activity towards biofilm structure of S. agalactiae by treating the extract with NaIO₄ (20 mM), proteinase K (1 mg/mL), and DNase I (100 mg/mL)

Fig. 6

Characterization of extracts produced by isolates CW01 (a, b, c) which is responsible for the inhibitory activity towards biofilm structure of S. agalactiae by treating the extract with NaIO₄ (20 mM), proteinase K (1 mg/mL), and DNase I (100 mg/mL)