Detection of biofilm-forming genes in Staphylococcus aureus clinical isolates by PCR assay (original) (raw)
Related papers
JOURNAL OF CLINICAL AND DIAGNOSTIC RESEARCH, 2018
Introduction: Superbug known as Staphylococcus aureus possess a tendency to form biofilm, which has a significant role in causing infection and abating host defense response. Amongst many mechanisms, biofilm formation depends on the icaABCD operon involved in the synthesis of a polysaccharide intercellular adhesion. Aim: To investigate biofilm forming ability of S. aureus isolates by phenotypic and genotypic methods. Materials and Methods: Of the 97 S. aureus clinical isolates collected, the quantitative biofilm formation was determined by microtiter plates. All S. aureus isolates were examined for detection of the icaABCD genes and mecA gene by using PCR method. Statistical analysis was performed with SPSS program version 17.0. Results: Among 97 S. aureus isolates from blood, wound, skin, surgery, internal, burn and infectious wards, urine and body fluids specimens, five isolates appeared as strong biofilm producer, while 28 displayed moderate biofilm formation, and 55 showed weak biofilm formation. Nine isolates did not reveal biofilm production on microtiter plates. The frequency of icaA, icaB, icaC and icaD genes in S. aureus isolates was 81 (83.5%), 71 (73.2%), 51 (52.5%) and 97 (100%), respectively. There was no relation between presence of icaABCD genes and biofilm formation (p=0.74). Conclusion: The presence of biofilm genes may not coincide with the ability to produce biofilm or vice versa. At the results, S. aureus clinical isolates possess different capacity to produce biofilm and adhesion. Methicillin resistance and susceptible isolates may not differ in their capacity to form biofilm.
Open Journal of Medical Microbiology
This study evaluated biofilm formation and antibiotic susceptibility in 36 clinical S. aureus isolates recovered from orthopaedic patients and detected the presence of intercellular adhesion and adhesin genes. Staphylococcus aureus was isolated from nasal swab, wound and urine specimens collected from orthopaedic patients in National Orthopaedic Hospital Dala, Kano over a period of three months. The isolates were identified using rapid identification kit for Staphylococcus species. The antibiotics susceptibility of the isolates was determined using modified disc diffusion method. Phenotypically, the biofilm formation was assessed using the Congo red agar method and microtitre plate assay. Polymerase chain reaction (PCR) analysis was used to detect biofilm-associated genes and characterize the isolates. The isolation rate of S. aureus from the samples (n = 134) was 26.8%, mainly from nasal swab (36%) and wound swab (36%). A total of 19 (52.7%) of the isolates showed positive for slime production. Majority of the isolates 29/36 (81.6%) were biofilm positive with only 2 (5.5%) and 5 (13.8%) as strong biofilm-formers and moderate biofilm-formers respectively. Molecular evaluation of the biofilm-associated genes in 12 S. aureus isolates revealed the prevalence of bbp genes (25%), clfA genes (16.6%) and the icaA (8.3%). None of the isolates harboured the fnbA and cna genes. There is no significant difference (P > 0.05) in the antibiotic resistance pattern between biofilm-positive and biofilm-negative S. aureus isolates. This result revealed that phenotypically most of the S. aureus isolates were biofilm formers but few of them chromosomally harbour the biofilm-associated genes.
2019
Background Among hospitalized patients, Staphylococcus aureus (S. aureus) infections pose a serious health threat. The present study investigated the frequency of biofilm forming genes among clinical isolates S. aureus and their susceptibility to antibiotics. Methods The clinical samples were analyzed via standard biochemical assays for identifying different bacterium, which was then confirmed using the multiplex colony PCR method. Those samples identified as S. aureus were examined for the presence of the cna, fnbA, fnbB and pvl genes. The antibiotic susceptibility of the S. aureus isolates was then tested. Results Using the standard biochemical assay approach, 54 S. aureus strains were identified. However, when using the multiplex PCR method 50 S. aureus strains were identified among the clinical samples. The in vitro biofilm formation assays determined 3 (6%) strains of S. aureus to be strong biofilm forming, 15 (30%) of the isolates were determined to be moderate biofilm forming...
Introduction: Biofilm producing Staphylococcus aureus is known as one of the major causative agents of infections, failure of implanted devices and persistent infection among hospitalized patients. The aim of the present study was to determine the frequency of biofilm producing S. aureus isolates amongst the clinical specimens. Methods: This cross-sectional study was conducted during 2012 to 2013 in two teaching hospitals in Shiraz, southwest of Iran. Totally, 345 S. aureus isolates from various clinical specimens were included. Biofilm producing isolates were phenotypically detected using Congo Red Agar (CRA) and genotypically by PCR assay for the icaA and icaD genes. Results: Of the 345 S. aureus isolates, 42.3% were methicillin-resistant S. aureus (MRSA) and subsequently 57.7% were methicillin susceptible isolates. The results of CRA plates showed that 77 (52.7%) and 74 (37.2%) of MRSA and MSSA were biofilm producing isolates. The frequency of icaA/D genes among MRSA and MSSA iso...
Detection of genes involved in biofilm formation in Staphylococcus aureus isolates
GMS Hygiene and Infection Control, 2016
Staphylococcus aureus is one of the Gram-positive pathogens causing a wide range of nosocomial infections. The present study investigates genotypic and phenotypic aspects involved in biofilm formation in methicillin-resistant Staphylococcus aureus strains isolated from nosocomial infections in Isfahan. A total of 110 S. aureus strains were collected from three major hospitals in Isfahan, the center of Iran. The antibiotic resistance pattern, phenotypes, and biofilm formation genes were studied using Congo red agar (CRA) and multiplex PCR (M-PCR). We found that 103 out of 110 samples (93.6%) were MRSA. The highest frequency of resistance was found to penicillin (89%), ciprofloxacin (87.4%), and erythromycin (86.1%). Phenotypic results showed that 53.5% were high biofilm producers, while 33.3% and 13.2% were intermediate and low biofilm producers, respectively. icaC (69.3%) had the highest frequency in comparison to other intercellular adhesion (ica) genes, icaD (54.8%) was second mos...
Molecular detection of biofilm coding genes in Staphylococcus aureus
In accordance with epidemic COVID-19, the elevated infection rates, disinfectant overuse and antibiotic misuse what led to immune suppression in most of the population in addition to genotypic and phenotypic alterations in the microorganisms, so a great need to reevaluate the genetic determinants that responsible for bacterial community (biofilm) has been raised. A total of 250 clinical specimens were obtained from patients in Baghdad hospitals and streaked on Mannitol salt agar medium. The results revealed that 156 isolates appeared as round yellow colonies, indicating that they were mostly identified as Staphylococcus aureus from 250 specimens. The antibiotic resistance pattern of the isolates for methicillin 37.17% (n=58), Amoxicillin-Clavulanate 58.9% (n=92), chloramphenicol 6.4% (n=10), Tetracyclin 62.8% (n=98), ceftriaxone 53.8% (n=84), Ciprofloxacin 6.4% (n=10), Gentamicin 42.3% (n=66), levofloxacin 28.2% (n=44), Penicillin 33.3% (n=52). The results demonstrated that 49 isolates were multidurg resistance. The biofilm formation ability of MDR was detected and total of 120 S. aureus isolates (76.92 %) were found to be adherent to varied degrees. Only fifty isolates (32.05% of the total) were classified as strong biofilm producers. Twenty-three (14.75%) were moderate producers, and forty seven isolates (30.12%) were found to be weak producers. A total of 36 isolates (23.08%) exhibited no biofilm production. Molecular detection of four biofilm coding genes icaA, icaD, icaR and eno was applied using doublex PCR technique. The current study revealed that 72%, 78%, 70% and 84% of isolates that carried icaA, icaD, icaR and eno genes respectively, eno is the predominant in the Iraqi isolates
Detection of icaA Gene Expression in Clinical Biofilm-Producing Staphylococcus Aureus Isolates
Iraqi Journal of Science, 2020
The pathogenicity resulting from Staphylococcus aureus infection has remarkable importance as one of the community-associated bacterial infections, due to the virulent ability of these bacteria to produce biofilms. This study was designed to detect biofilm production in clinical isolates from samples of wounds and urinary tract infections. The expression levels of the icaA gene that is responsible of slime layer production in biofilms was compared in isolates with different biofilm producing capabilities. Fifty seven samples that included 32 samples from urine and 25 samples from wounds were collected from Alwasti Hospital, Al-Kindi Teaching Hospital, and Alzahraa Clinic, Baghdad, Iraq. The bacteria was identified according to biochemical tests, API20 strip test, and PCR assay. The results of 16S rRNA PCR detection revealed that nine isolates were identified as S. aureus. The biofilm assay showed that 46.15% of the isolates were strong biofilm producers, 46.15% had moderate ability ...
Evaluation of Biofilm Formation in Staphylococcus aureus Clinical Isolates
medicallaboratory journal, 2019
Background and Objectives: Staphylococcus aureus is a common cause of nosocomial infections. The ability of S. aureus to form biofilm and acquire antimicrobial resistance has made this organism a major health problem. In this study, we investigate the biofilm-forming ability of S. aureus isolates from clinical samples. Methods: Sixty S. aureus isolates from clinical specimens were collected from the 5th Azar Hospital of Gorgan (Iran) in 2018. The isolates were identified using conventional methods including Gram staining and biochemical tests (catalase and coagulase). Biofilm formation by S. aureus isolates was evaluated using a microplate-based method. Results: Of 60 S. aureus isolates, 47 (78.3%) strains were identified as biofilmforming and 13 (21.7%) strains were non-biofilm-forming. Conclusion: The high prevalence of biofilm-producing S. aureus isolates in the 5 th Azar hospital of Gorgan could pose a major health challenge with serious consequences for hospitalized patients. Therefore, it is crucial to disinfect and sterilize hospital surfaces and equipment effectively to minimize the risk of contamination and spread of bacteria in the hospital settings.
Pharmaceutical Sciences, 2021
Background: The success of Staphylococcus aureus is as an important human pathogen is probably due to possession of various virulence determinants. Attachment and biofilm formation is considered the main step in any infection. The present study aimed to determine the presence of S. aureus surface (sas) genes and their association with biofilm formation and antibiotic resistance. Methods: S. aureus isolates collected were analyzed for biofilm formation using polystyrene microtitre plates. All S. aureus isolates were also examined for the determination of sas genes by PCR assays and antibiotic susceptibility assay by disk diffusion method. Results: Biofilm formation assay revealed that 29 S.aureus isolates were weak biofilm producers, 57 had moderate biofilm production, while only five isolates showed strong biofilm formation. The biofilm production was not revealed among nine isolates. The frequency of sas genes were 95 (88%), 94 (87%), 94 (87%), 92 (85.2%), 98 (90.7%), 93 (86.1%), 9...
Periodic monitoring of Staphylococcus aureus characteristics in a locality is imperative as their drug-resistant variants cause treatment problem. In this study, antibiograms, prevalence of toxin genes (sea-see, seg-ser, seu, tsst-1, eta, etb, and etd), PFGE types, accessory gene regulator (agr) groups, and ability to form biofilm of 92 S. aureus Thailand clinical isolates were investigated. They were classified into 10 drug groups: groups 1-7 (56 isolates) were methicillin resistant (MRSA) and 8-10 (36 isolates) were methicillin sensitive (MSSA). One isolate did not have any toxin gene, 4 isolates carried one toxin gene (seq), and 87 isolates had two or more toxin genes. No isolate had see, etb, or tsst-1; six isolates had eta or etd. Combined seg-sei-sem-sen-seo of the highly prevalent egc locus was 26.1%. The seb, sec, sel, seu, and eta associated significantly with MSSA; sek was more in MRSA. The sek-seq association was 52.17% while combined sed-sej was not found. Twenty-three PFGE types were revealed, no association of toxin genes with PFGE types. All four agr groups were present; agr group 1 was predominant (58.70%) but agr group 2 strains carried more toxin genes and were more frequent toxin producers. Biofilm formation was found in 72.83% of the isolates but there was no association with antibiograms. This study provides insight information on molecular and phenotypic markers of Thailand S. aureus clinical isolates which should be useful for future active surveillance that aimed to control a spread of existing antimicrobial resistant bacteria and early recognition of a newly emerged variant. BioMed Research International cytokines and T-cell stimulating factors leading to toxic shock syndrome which may be fatal [8, 9]. The enterotoxicity and superantigenicity are distinct properties of the toxin molecule [6]. SEs are classified into two types based on their emetic activity in the toxin fed modeled primate. Toxins that induce vomiting in the primate are placed in the classical SE type while those that lack the emetic activity or have not been tested are allocated in the SE-like (SEls) type [10, 11]. Members of the classical SEs are SEA-SEE and the more recently recognized SEG, SEH, SEI, SER, SES, and SET. The SEls members include SElJ, SElK, SElL, SElM, SElN, SElO, SElP, SElQ, SElU, SElU2 or SEW, and SElV [11]. The staphylococcal enterotoxin F (SEF) which lacks emetic activity but is associated with toxic shock syndrome is presently called toxic shock syndrome toxin-1 (TSST-1) [12]. The SEs and the TSST-1 as well as the bacterial resistance to drugs are encoded by genes on the mobile genetic elements including prophages, plasmids, pathogenicity islands, genomic islands, and antibiotic resistance cassette [13]; thus they are transmitted horizontally rather easily. Expression of S. aureus virulence factors and metabolism of metabolic pathways during growth are coordinated/regulated by a quorumsensing operon named accessory gene regulator (agr) [14, 15]. Based on the amino acid sequence polymorphisms of the agr-encoding autoinducing peptides and their responding receptors, S. aureus strains can be divided into four major agr groups (groups 1-4) [16]. During the last five decades, S. aureus clones that resist methicillin (methicillin-resistant S. aureus, MRSA) disseminated and caused medical and public health problem worldwide [17, 18]. These strains are not only resistant to methicillin, but also resistant to all other-lactams, such as cephalosporin [18, 19]. In Thailand, MRSA infections were reported from 23 hospitals from 1988 to 1998 [20, 21]. The proportions of MRSA to MSSA in the northeast, central, and southern regions of the country during the studied period increased from 11 to 23.4%, 16 to 30.5%, and 21 to 30.3%, respectively [22]. Moreover, methicillin-resistant S. aureus with reduced susceptibility to vancomycin was recognized [23]. However, data on genotypic characteristics and other attributes of the S. aureus isolates in Thailand are relatively rare. Therefore, this study investigated the prevalence of virulence toxin genes coding for enterotoxins (sea-see, seg-ser, and seu), toxic shock syndrome toxin-1 (tsst-1), and exfoliative toxins (eta, etb, and etd) among S. aureus Thailand clinical isolates. Molecular diversity of the isolates regarding their endonuclease-restricted patterns of genomic DNA (PFGE), agr types, and antimicrobial susceptibility as well as their ability to produce biofilm were also investigated.