Antibacterial, Antibiofilm, and Antioxidant Activity of Polysaccharides Obtained from Fresh Sarcotesta of Ginkgo biloba: Bioactive Polysaccharide that Can Be Exploited as a Novel Biocontrol Agent

Staphylococcus aureus (S. aureus) biofilm plays an important role in the persistence of chronic infection due to its resistance to antibiotics. Because of their functional diversity, active polysaccharide is increasingly being applied as a biocontrol agent to inhibit the formation of biofilm by pathogens. In this study, a new polysaccharide, GBSPII-1, isolated from the fresh sarcotesta of Ginkgo biloba L. (G. biloba) was characterized and its effect on antibiofilm formation of S. aureus was examined in vitro. High-Performance Liquid Chromatography (HPLC) analysis showed that GBSPII-1 is an acidic heteropolysaccharide composed of mannose, rhamnose, glucose, glucuronic acid, and galacturonic acid. GBSPII-1 demonstrated a molecular weight of 34 kDa and may affect the accumulation of polysaccharide intercellular adhesion (PIA) by inhibiting icaA, icaB, icaC, and icaD gene expression at subinhibitory concentrations. Under 10 g/L, GBSPII-1 showed an antioxidant effect on the inhibition rate of H2O2-induced erythrocyte hemolysis and the scavenging rate of DPPH radicals was 76.5 ± 0.5% and 89.2 ± 0.26%, respectively. The findings obtained in this study indicate that GBSPII-1 has antibacterial effect, is a possible source of natural antioxidants, and may be a potential biocontrol agent for the design of new therapeutic strategies for biofilm-related S. aureus infections.

The Correlation between icaA and icaD Genes with Biofilm Formation Staphylococcus epidermidis In Vitro

This study was conducted to identify the presence of icaA and icaD genes in S. epidermidis and to analyze the relationship between the presence of icaA and icaD genes with the ability of in vitro biofilm formation in S. epidermidis. S. epidermidis isolates from patients and healthy people were collected and PCR was examined to detect icaA and icaD genes. which then continued to examine the ability of biofilm formation by the method of Congo Red Agar. The results of this genotypic and phenotypic examination were then tested for correlation with statistical tests using SPSS 23.0. A total of 40 S. epidermidis isolates were collected, consisting of 20 clinical isolates and 20 isolates of normal flora. The icaA gene was positive in 5 isolates (12.5%), and 8 isolates (20%) were positive for the icaD gene, 3 isolates with icaA and icaD were both positive. One hundred percent of isolates with icaA or icaD positively formed biofilms, but there were 15 isolates (42.9%) who did not have the icaA gene but showed the ability to form biofilms, while 12 isolates (37.5%) who did not have the icaD gene also formed biofilms. Fifty percent of S. epidermidis isolates showed the ability to form biofilms at CRA. The Fisher Exact test showed a significant relationship between the icaA gene and the ability of biofilm formation (p=0.047 (p<0.05)) as well as the icaD gene (p=0.03 (p<0.05)). The icaA and icaD genes have a significant relationship to biofilm formation in S. epidermidis. There was another mechanism in the formation of biofilms that are not dependent on the ica gene.

The Correlation between icaA and icaD Genes with Biofilm Formation Staphylococcus epidermidis In Vitro

This study was conducted to identify the presence of icaA and icaD genes in S. epidermidis and to analyze the relationship between the presence of icaA and icaD genes with the ability of in vitro biofilm formation in S. epidermidis. S. epidermidis isolates from patients and healthy people were collected and PCR was examined to detect icaA and icaD genes. which then continued to examine the ability of biofilm formation by the method of Congo Red Agar. The results of this genotypic and phenotypic examination were then tested for correlation with statistical tests using SPSS 23.0. A total of 40 S. epidermidis isolates were collected, consisting of 20 clinical isolates and 20 isolates of normal flora. The icaA gene was positive in 5 isolates (12.5%), and 8 isolates (20%) were positive for the icaD gene, 3 isolates with icaA and icaD were both positive. One hundred percent of isolates with icaA or icaD positively formed biofilms, but there were 15 isolates (42.9%) who did not have the icaA gene but showed the ability to form biofilms, while 12 isolates (37.5%) who did not have the icaD gene also formed biofilms. Fifty percent of S. epidermidis isolates showed the ability to form biofilms at CRA. The Fisher Exact test showed a significant relationship between the icaA gene and the ability of biofilm formation (p=0.047 (p<0.05)) as well as the icaD gene (p=0.03 (p<0.05)). The icaA and icaD genes have a significant relationship to biofilm formation in S. epidermidis. There was another mechanism in the formation of biofilms that are not dependent on the ica gene.

Biofilm Formation and Detection of icaD Gene in Staphylococcus aureus Isolated from Clinical Specimens

Background: S. aureus is found to be a major source of community as well as hospital acquired infections. The increase in antimicrobial resistance and emergence of multidrug resistance has become a big threat worldwide. The biofilm formation of S. aureus influenced the survival and persistence in both environment and host. Aim: The study was conducted with the aim to evaluate in-vitro biofilm formation and the presence of icaD gene in S. aureus from clinical isolates of S. aureus. Methods: A total of 570 wound/pus samples were processed by standard microbiological techniques. Colony morphology, Gram’s staining and biochemical tests were used for the identification of S. aureus. Antimicrobial susceptibility test was performed by Kirby-Bauer disc diffusion technique and methicillin-resistant S. aureus was detected by using cefoxitin antibiotics. The production of biofilm was screened by Congo Red Agar and finally, the presence of icaD gene was determined by PCR. Results: Out of 570 samples, a total 19.3% (110/570) samples showed the growth of S. aureus. Among which 59.1% (65/110) were multi-drug resistant. Similarly, 26.4% (29/110) isolates were methicillin-resistant S. aureus. Among MRSA isolates 93.1% (27/29) were MDR with more than 3 classes of antibiotics. Biofilm production was shown by 95.45% (105/110) and 77.3% (85/110) isolates on Congo Red Agar and presence of icaD gene respectively. Conclusion: In this study, the significant association was observed in phenotypic production of biofilm and the presence of icaD gene for the genotypic expression of biofilm. There were also increasing rates of MRSA and multidrug resistance S. aureus.

MINIMUM BACTERICIDAL CONCENTRATION OF COMMERCIAL DISINFECTANTS ON STAPHYLOCOCCUS SPP. ISOLATED FROM MASTITIS IN GOATS AND DETECTION OF THE icaD GENE

This study aimed to evaluate the in vitro efficacy of commercial disinfectants on Staphylococcus spp., isolated from mastitis cases in goats, and to associate the observed resistance with the presence of the icaD gene. Broth microdilution was employed to evaluate the in vitro antimicrobial activity of the disinfectants, whereas the Congo Red technique was used for the evaluation of biofilm production and amplification of the icaD gene. All evaluated samples were sensitive to disinfectants, with the following ranges of activity: quaternary ammonium (0.13 - 21.33 µg/ml), chlorhexidine (4.00 - 313.00 µg/ml) and iodine (190.00 - 12500.00 µg/ml); however, the sodium hypochlorite-based disinfectant showed no bactericidal activity in the concentration range from 15.0 to 0.03 µg/ml. The icaD gene presented a frequency of 14.7% in the isolate samples. Fisher’s exact test showed a significant effect of the relation between the minimum bactericidal concentration value of the quaternary ammonium-based disinfectant and the presence/absence of the icaD gene (P <0.01). Commercial disinfectants with quaternary ammonium, chlorhexidine and iodine active ingredients presented in vitro activity even at concentrations lower than those recommended by the manufacturers. Therefore, the periodic evaluation of the sensitivity profile of the disinfectants must be performed.

icaA/D Genes and Biofilm Formation of Methicillin-Resistant Staphylococcus aureus in Dr. Soetomo Hospital, Surabaya

Methicillin-resistant Staphylococcus aureus (MRSA) is a global health concern. One of the factors causing hospital infection is related to the ability of MRSA bacteria to form biofilms. Polysaccharide intercellular adhesin (PIA), encoded by ica gene, have an important role in S. aureus intracellular accumulation and aggregation. The aims of this study was to analyze the relationship between icaA, icaD genes and biofilm production in MRSA carrier and clinical isolate in Dr. Soetomo Hospital Surabaya. This study was an observational study using cross sectional approach. The sample was 47 MRSA isolates is as follow 28 isolates from carrier and 19 were clinical isolates. All of MRSA isolates carried mecA gene. PCR was performed to detect icaA and icaD genes. Biofilm formation was detected using microtiter plate assay (MTP). icaA gene was detected in all isolates whereas icaD gene in 96,4% carrier isolates and all (100%) of clinical isolates. Positive MTP results showed in all (100%) of carrier isolates and 57,9% of clinical isolates. Statistic result was significantly different in biofilm formation between carrier and clinical MRSA isolates. The proportion of positive biofilm formation in isolate with positive icaA/D genes was 82.6%. There was not any association between icaA and icaD gene with biofilm production.

Characterization of virulence and antibiotic profile and agr typing of Staphylococcus aureus from milk of subclinical mastitis bovine in State of Rio de Janeiro

ABSTRACT This study aims to detect the main virulence and antimicrobial resistance genes in Stapylococcus aureus from bovine mastitic milk as well as classifying them according to agr typing. A total of 55 strains from six dairy unities in the state of Rio de Janeiro were selected, of these 27.3% presented fbnA and 78,2% for fbnB genes, respectively. None of the strains tested were positive for cap5 gene, 3.6% were positive for cap8 gene. Additionally, 94.5% of strains had hlA gene and 89.1% had hlB gene while 67.3% of the strains had icaA gene and 87.3% had icaD gene. From these results it was possible to establish 12 different virulence profiles. Prevalence of agrII type was detected in 81.8% of the isolates. Concerning antimicrobial resistance evaluation, the studied strains were susceptible to all antibiotics tested except penicillin, 83.6% being resistant strains. None of the strains had mecA gene, however, 40% of the strains had blaZ gene. Associating virulence and resistance data made it possible to obtain 23 different profiles. This great diversity of strains shows wide array of bacterial strategies and the challenge of mastitis prevention in cattle. Despite antimicrobial susceptibility, these strains presented certain genes that allow its persistence in the herd.

Biofilm-forming and antimicrobial resistance traits of staphylococci isolated from goat dairy plants

Introduction: Biofilm-associated antimicrobial resistance is of increasing importance to the maintenance and spread of foodborne pathogens in the food industry. This study aimed to investigate the ability to form biofilm and the antimicrobial resistance of staphylococci contaminating small-scale goat milk dairy plants. Methodology: Sixty isolates were tested for antimicrobial resistance against 20 drugs by the microdilution method. Biofilm-forming traits were assessed by the microtiter plate method (MtP), Congo red agar method (CRA), and icaD gene detection by polymerase chain reaction (PCR). Results: High antimicrobial resistance to ampicillin (60/60; 100%), penicillin G (21/60; 35%), and erythromycin (15/60; 25%) was observed, but all isolates were susceptible to amoxicillin/K-clavulanate, ceftriaxone, ciprofloxacin, gentamicin, levofloxacin, linezolid, and moxifloxacin. No resistance to oxacillin or vancomycin was seen among Staphylococcus aureus. Twenty-seven isolates (27/60; 45%) were considered to form biofilm according to MtP, and similar biofilm-producing frequencies were observed in coagulase-negative staphylococci (CoNS) (20/44; 45.4%) and S. aureus (7/16; 43.7%). The icaD gene was observed only in S. aureus isolates. There was no association between biofilm production and antimicrobial resistance. A higher frequency of biofilm-producing staphylococci was found in isolates from bulk tank milk and hand swabs. On the other hand, isolates from pasteurized milk showed lower frequency of biofilm formation. Conclusions: Staphylococci contaminating goat dairy plants are potential biofilm producers. The results suggest no association between the ability to form biofilm and antimicrobial resistance.

Genes encoding adhesion factors and biofilm formation in methicillin-resistant Staphylococcus aureus in Morocco

Introduction: Infections involving methicillin-resistant Staphylococcus aureus (MRSA) remain a serious threat to hospitalized patients worldwide. MRSA is characterized by recalcitrance to antimicrobial therapy, which is a function not only of widespread antimicrobial resistance, but also the capacity to form biofilms. The present study evaluated the presence of genes encoding adhesion factors and the biofilm-forming capacity in MRSA. Methodology: In this study, 53 isolates of MRSA, recovered from December 2010 to May 2014 in a mother and child hospital, CHU Mohamed VI in Marrakech, Morocco, were screened for the presence of bap and ica genes associated with biofilm formation, and for bbp, cna, ebpS, eno, fib, fnbA, fnbB, clfA, and clfB genes that encode microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). The biofilm formation assay was performed in 96-well microtiter polystyrene plates. The presence of genes was determined by polymerase chain reaction (PCR). Results: An association was found between icaD gene detection and biofilm formation; 100% of the strains harbored icaD and produced biofilm. None of the isolates harbored bap or bbp. Furthermore, 96.23% isolates were positive for fnbA, 60.37% for eno, 43.39% for clfA and clfB, 11.32% for cna, 9.34% for ebpS, 5.66% for fib, and 1.89% for fnbA. Conclusions: Our findings showed that the MRSA carriage in Marrakech children was high. The genetic variations of adhesion genes require further investigation.