Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria
Background Bacterial biofilms have become a major threat to human health. The objective of this study was to isolate amylase-producing bacteria from soil to determine the overall inhibition of certain pathogenic bacterial biofilms. Methods We used serial dilution and the streaking method to obtain a...
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PeerJ Inc.
2020-11-01
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| _version_ | 1827605937471356928 |
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| author | Rokaia Elamary Wesam M. Salem |
| author_facet | Rokaia Elamary Wesam M. Salem |
| author_sort | Rokaia Elamary |
| collection | DOAJ |
| description | Background Bacterial biofilms have become a major threat to human health. The objective of this study was to isolate amylase-producing bacteria from soil to determine the overall inhibition of certain pathogenic bacterial biofilms. Methods We used serial dilution and the streaking method to obtain a total of 75 positive amylase isolates. The starch-agar plate method was used to screen the amylolytic activities of these isolates, and we used morphological and biochemical methods to characterize the isolates. Optimal conditions for amylase production and purification using Sephadex G-200 and SDS-PAGE were monitored. We screened these isolates’ antagonistic activities and the purified amylase against pathogenic and multi-drug-resistant human bacteria using the agar disk diffusion method. Some standard antibiotics were controlled according to their degree of sensitivity. Finally, we used spectrophotometric methods to screen the antibiofilm 24 and 48 h after application of filtering and purifying enzymes in order to determine its efficacy at human pathogenic bacteria. Results The isolated Bacillus species were Bacillus megaterium (26.7%), Bacillus subtilis (16%), Bacillus cereus (13.3%), Bacillus thuringiesis (10.7%), Bacillus lentus (10.7%), Bacillus mycoides (5.3%), Bacillus alvei (5.3%), Bacillus polymyxa (4%), Bacillus circulans (4%), and Micrococcus roseus (4%). Interestingly, all isolates showed a high antagonism to target pathogens. B. alevi had the highest recorded activity (48 mm) and B. polymyxa had the lowest recorded activity (12 mm) against Staphylococcus aureus (MRSA) and Escherichia coli, respectively. On the other hand, we detected no antibacterial activity for purified amylase. The supernatant of the isolated amylase-producing bacteria and its purified amylase showed significant inhibition for biofilm: 93.7% and 78.8%, respectively. This suggests that supernatant and purified amylase may be effective for clinical and environmental biofilm control. Discussion Our results showed that soil bacterial isolates such as Bacillus sp. supernatant and its purified amylase are good antibiofilm tools that can inhibit multidrug-resistant former strains. They could be beneficial for pharmaceutical use. While purified amylase was effective as an antibiofilm, the isolated supernatant showed better results. |
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| language | English |
| last_indexed | 2024-03-09T06:29:53Z |
| publishDate | 2020-11-01 |
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| spelling | doaj.art-4e5541828bd74ccdaef68bb379281c672023-12-03T11:06:27ZengPeerJ Inc.PeerJ2167-83592020-11-018e1028810.7717/peerj.10288Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteriaRokaia Elamary0Wesam M. Salem1Department of Botany and Microbiology, South Valley University, Qena, EgyptDepartment of Botany and Microbiology, South Valley University, Qena, EgyptBackground Bacterial biofilms have become a major threat to human health. The objective of this study was to isolate amylase-producing bacteria from soil to determine the overall inhibition of certain pathogenic bacterial biofilms. Methods We used serial dilution and the streaking method to obtain a total of 75 positive amylase isolates. The starch-agar plate method was used to screen the amylolytic activities of these isolates, and we used morphological and biochemical methods to characterize the isolates. Optimal conditions for amylase production and purification using Sephadex G-200 and SDS-PAGE were monitored. We screened these isolates’ antagonistic activities and the purified amylase against pathogenic and multi-drug-resistant human bacteria using the agar disk diffusion method. Some standard antibiotics were controlled according to their degree of sensitivity. Finally, we used spectrophotometric methods to screen the antibiofilm 24 and 48 h after application of filtering and purifying enzymes in order to determine its efficacy at human pathogenic bacteria. Results The isolated Bacillus species were Bacillus megaterium (26.7%), Bacillus subtilis (16%), Bacillus cereus (13.3%), Bacillus thuringiesis (10.7%), Bacillus lentus (10.7%), Bacillus mycoides (5.3%), Bacillus alvei (5.3%), Bacillus polymyxa (4%), Bacillus circulans (4%), and Micrococcus roseus (4%). Interestingly, all isolates showed a high antagonism to target pathogens. B. alevi had the highest recorded activity (48 mm) and B. polymyxa had the lowest recorded activity (12 mm) against Staphylococcus aureus (MRSA) and Escherichia coli, respectively. On the other hand, we detected no antibacterial activity for purified amylase. The supernatant of the isolated amylase-producing bacteria and its purified amylase showed significant inhibition for biofilm: 93.7% and 78.8%, respectively. This suggests that supernatant and purified amylase may be effective for clinical and environmental biofilm control. Discussion Our results showed that soil bacterial isolates such as Bacillus sp. supernatant and its purified amylase are good antibiofilm tools that can inhibit multidrug-resistant former strains. They could be beneficial for pharmaceutical use. While purified amylase was effective as an antibiofilm, the isolated supernatant showed better results.https://peerj.com/articles/10288.pdfAmylase Bacillus Soil bacteria AntibiofilmPathogenic bacteriaSDS-PAGE profile |
| spellingShingle | Rokaia Elamary Wesam M. Salem Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria PeerJ Amylase Bacillus Soil bacteria Antibiofilm Pathogenic bacteria SDS-PAGE profile |
| title | Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria |
| title_full | Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria |
| title_fullStr | Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria |
| title_full_unstemmed | Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria |
| title_short | Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria |
| title_sort | optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm forming bacteria |
| topic | Amylase Bacillus Soil bacteria Antibiofilm Pathogenic bacteria SDS-PAGE profile |
| url | https://peerj.com/articles/10288.pdf |
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