Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia

Streptococcus pyogenes or group A streptococcus (GAS) is one type of bacterium that causes a wide variety of infectious and immunologically-related diseases. Hyaluronidase enzyme produced by GAS degrades hyaluronic acid, a major component in extracellular matrix of human connective tissues allowing...

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Main Author:	Wan Mansor, Wan Muhammad Zamir
Format:	Thesis
Language:	English
Published:	2019
Subjects:	Streptococcus pyogenes - cytology Hyaluronoglucosaminidase - adverse effects
Online Access:	http://psasir.upm.edu.my/id/eprint/98050/1/FPSK%28m%29%202020%2032%20UPMIR.pdf
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spelling	my-upm-ir.980502022-07-13T06:32:02Z Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia 2019-07 Wan Mansor, Wan Muhammad Zamir Streptococcus pyogenes or group A streptococcus (GAS) is one type of bacterium that causes a wide variety of infectious and immunologically-related diseases. Hyaluronidase enzyme produced by GAS degrades hyaluronic acid, a major component in extracellular matrix of human connective tissues allowing the pathogen to enter. Meanwhile, biofilm formation in GAS is associated with therapeutic failure due to the robustness of biofilms against antibiotics. Till date, little is known on GAS hyaluronidase, its gene (Hyl) and biofilms. The aims of this study were to determine the production of bacterial hyaluronidase and its molecular characteristics and biofilm formation as well as emm typing among GAS clinical isolates. A total of 45 S. pyogenes clinical isolates were obtained from the previous stock cultures which were collected from Kuala Lumpur and Serdang hospitals. Phenotypic hyaluronidase production was assessed by a solid plate assay. Hyl genes (HylA, HylP1, HylP2, HylP3) were detected through multiplex PCR using established primers. emm typing was done by emm gene amplification and sequencing according to established protocols. The enzymatic activity of hyaluronidase from different GAS isolates was determined through hyaluronic acid turbidity reduction assay. Biofilm formation was detected using Congo Red Agar (CRA) and Crystal Violet Assay (CVA). Forty isolates (88.9%) exhibited hyaluronidase production despite the presence of Hyl genes (chromosomal and phage-associated) in all isolates. Hyaluronidase exhibited the maximum activity at 10th minute time point with appropriate conditions that include 3% of bovine serum albumin (BSA), pH 3.5 and 0.1M NaCl. Positive correlation between bacterial growth and enzymatic activity was observed (p value < 0.05). Most of the strains exhibited moderate biofilm production, (30, 66.66%) in CRA and (23, 51.11%) in CVA. A total of 29 different emm types were detected. The most prevalent emm types were emm1, 18, 28, 97 and 102 with (6.7%) of the isolates each, while emm types 63, 71, 76, 89 and 91 were seen in 4.4% of the isolates respectively. The rest of the emm types 12, 15, 44, 56, 65, 81, 98, 101, 105 and 120 were found about 2.2% each. No new emm types were detected. In general, there was no strong association between hyaluronidase production, emm type, invasiveness and biofilm formation as p values were more than 0.05). Hyaluronidase and production of biofilms in GAS may be involved in its pathogenesis as hyaluronidase able to break hyaluronic acid (HA) which is the layer wall on the host cells and the development of biofilm aid bacteria to become antimicrobial resistance towards antibiotics. Diverse emm types in the study may signify the heterogeneity of the local GAS strains. More research is warranted in future to establish to link between GAS virulence factors and its biofilms. Streptococcus pyogenes - cytology Hyaluronoglucosaminidase - adverse effects 2019-07 Thesis http://psasir.upm.edu.my/id/eprint/98050/ http://psasir.upm.edu.my/id/eprint/98050/1/FPSK%28m%29%202020%2032%20UPMIR.pdf text en public masters Universiti Putra Malaysia Streptococcus pyogenes - cytology Hyaluronoglucosaminidase - adverse effects Awang Hamat, Rukman
institution	Universiti Putra Malaysia
collection	PSAS Institutional Repository
language	English
advisor	Awang Hamat, Rukman
topic	Streptococcus pyogenes - cytology Hyaluronoglucosaminidase - adverse effects
spellingShingle	Streptococcus pyogenes - cytology Hyaluronoglucosaminidase - adverse effects Wan Mansor, Wan Muhammad Zamir Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
description	Streptococcus pyogenes or group A streptococcus (GAS) is one type of bacterium that causes a wide variety of infectious and immunologically-related diseases. Hyaluronidase enzyme produced by GAS degrades hyaluronic acid, a major component in extracellular matrix of human connective tissues allowing the pathogen to enter. Meanwhile, biofilm formation in GAS is associated with therapeutic failure due to the robustness of biofilms against antibiotics. Till date, little is known on GAS hyaluronidase, its gene (Hyl) and biofilms. The aims of this study were to determine the production of bacterial hyaluronidase and its molecular characteristics and biofilm formation as well as emm typing among GAS clinical isolates. A total of 45 S. pyogenes clinical isolates were obtained from the previous stock cultures which were collected from Kuala Lumpur and Serdang hospitals. Phenotypic hyaluronidase production was assessed by a solid plate assay. Hyl genes (HylA, HylP1, HylP2, HylP3) were detected through multiplex PCR using established primers. emm typing was done by emm gene amplification and sequencing according to established protocols. The enzymatic activity of hyaluronidase from different GAS isolates was determined through hyaluronic acid turbidity reduction assay. Biofilm formation was detected using Congo Red Agar (CRA) and Crystal Violet Assay (CVA). Forty isolates (88.9%) exhibited hyaluronidase production despite the presence of Hyl genes (chromosomal and phage-associated) in all isolates. Hyaluronidase exhibited the maximum activity at 10th minute time point with appropriate conditions that include 3% of bovine serum albumin (BSA), pH 3.5 and 0.1M NaCl. Positive correlation between bacterial growth and enzymatic activity was observed (p value < 0.05). Most of the strains exhibited moderate biofilm production, (30, 66.66%) in CRA and (23, 51.11%) in CVA. A total of 29 different emm types were detected. The most prevalent emm types were emm1, 18, 28, 97 and 102 with (6.7%) of the isolates each, while emm types 63, 71, 76, 89 and 91 were seen in 4.4% of the isolates respectively. The rest of the emm types 12, 15, 44, 56, 65, 81, 98, 101, 105 and 120 were found about 2.2% each. No new emm types were detected. In general, there was no strong association between hyaluronidase production, emm type, invasiveness and biofilm formation as p values were more than 0.05). Hyaluronidase and production of biofilms in GAS may be involved in its pathogenesis as hyaluronidase able to break hyaluronic acid (HA) which is the layer wall on the host cells and the development of biofilm aid bacteria to become antimicrobial resistance towards antibiotics. Diverse emm types in the study may signify the heterogeneity of the local GAS strains. More research is warranted in future to establish to link between GAS virulence factors and its biofilms.
format	Thesis
qualification_level	Master's degree
author	Wan Mansor, Wan Muhammad Zamir
author_facet	Wan Mansor, Wan Muhammad Zamir
author_sort	Wan Mansor, Wan Muhammad Zamir
title	Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
title_short	Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
title_full	Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
title_fullStr	Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
title_full_unstemmed	Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia
title_sort	characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive streptococcus pyogenes clinical isolates in two hospitals in malaysia
granting_institution	Universiti Putra Malaysia
publishDate	2019
url	http://psasir.upm.edu.my/id/eprint/98050/1/FPSK%28m%29%202020%2032%20UPMIR.pdf
_version_	1747813832751841280

Characterisation of hyaluronidase, biofilm formation and emm type among invasive and non-invasive Streptococcus pyogenes clinical isolates in two hospitals in Malaysia

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