1Al-Quds University, Israel
2Al-Quds Public Health Society, Israel
Introduction: Staphylococcus aureus ability to produce biofilm and adhesion makes them resistant to antimicrobial therapy. The current study aims to characterize MRSA strains and to determine the prevalence of genes encoding adhesion factors and biofilm formation, also their correlation with drug resistance.
Methodology: A total of 248 isolates of MRSA were collected from Palestinian patients, during 2015 to 2017. Biofilm formation was studied by microtiter plate assay, the strains characterized by staphylococcal chromosome cassette mec (SCCmec) typing, and screened for the bbp, cna, ebpS, eno, fib, fnbA, fnbB, clfA, and clfB genes that encode microbial surface components recognizing adhesive matrix molecules and icaD/icaA, bap and the staphylococcal accessory regulator (sarA) and agr group genes that associated with biofilm formation.
Results: The majority of isolates harbored SCCmec type IV, which is common in community-acquired MRSA strains. Most isolates also showed resistance to more than four of the tested antimicrobials. The results demonstrated that all (100%) of isolates were biofilm producers by the quantitative microtitre plate assays. The distributions of biofilm formation between isolates were 21%, 46.4%, 32.7% as high, moderate and weak, respectively.
All of the strains harbored icaD/icaA genes and produced biofilm (P<0.05). None of the isolates harbored bap. Furthermore, 94.8% of isolates were positive for eno, 80.2% for each clfA, clfB, 78.2%, for fnbA, 76.2% for ebps, 62.2% for fib, 39.9% for cna and 29.0% for fnbB. Further, nearly 69.8% of the isolates were found positive for the gene sarA. All four agr groups were present; agr group 1 was predominant (39.5%) but agr group 2 strains carried more toxin genes and were more frequent toxin producers.
Conclusions: These results are important for epidemiological studies involving MRSA infections. Our findings showed that the MRSA carriage is high and the genetic variations of adhesion genes require further investigation.
Biography:
Kifaya Azmi has completed her PhD from Charite University School of Medicine, Berlin. She is one of the staff members of Al-Quds University, faculty of medicine, Teaching biochemistry and molecular biology. Recently working on MRSA project, published more than 35 papers in reputed journals.
1Cardiff University, UK
2Northeastern University, USA
Several theories have been proposed to explain the “Great Plate Anomaly”, that is, the observed phenomenon by which most of the environmental microorganisms seen under the microscope, cannot currently be grown under laboratory conditions. A possible explanation could combine the commonly known bet-hedging strategies of dormancy and bistability: Many microorganisms seem to enter a dormant state of low metabolic activity when environmental conditions are unfavorable and bistability is observed when gene expression within a bacterial population of genetically identical bacteria bifurcates into phenotypically distinct subpopulations. In this context, we wanted to determine if the entry into dormancy (in the form of sporulation) was able to erase the phenotypic memory acquired through vertical inheritance. Bacillus subtilis spores were germinated into different media, and several passages were performed to favor selection of individuals adapted to each specific growth medium. After this acclimatization process, bacteria were induced to sporulate and the spores were germinated in the different media. Monitorization of colony growth through the ScanLag technology showed different growth patterns for germinated spores before acclimatization, after acclimatization, and vegetative cells after growth passages, suggesting that the phenotypic memory acquired through passages was partially removed during sporulation. This observation could provide a first hint at another aspect of bet-hedging, in which environmental bacteria could use dormancy to reset their acquired phenotypic memory to be more adaptable to different environments.
Biography:
Dr. Epstein is a recognized expert in Environmental Microbiology and a Professor of Biology at Northeastern University, in Boston. Together with Dr. Lewis, Dr. Epstein co-invented the iChip, a method to grow previously uncultured microorganisms and which allowed for the discovery of teixobactin. Dr. Espina is currently doing a postdoc in Cardiff University in conjunction with Neem Biotech, working in a project to increase culturability of environmental bacteria in order to increase the possibilities of finding antimicrobial-producing species. Dr. Espina was invited to a research stay in Dr. Epsteinʼs laboratory to try to elucidate the role of dormancy in bacterial culturability.
University of Sheffield, UK
Burkholderia thailandensis is a non-pathogenic bacterium and is widely used as a model of Burkholderia pseudomallei, the causative agent of melioidosis. One of the histopathological features of melioidosis is multinucleate giant cell (MNGC) formation, potentially a protective structure for the bacteria. However, the molecular mechanisms of this pathogenic process have not been elucidated. It is possible that the bacteria can regulate this process by affecting the function and expression of host cell membrane molecules, including tetraspanins. Tetraspanins are known to be involved in MNGC formation and other cell fusion events, probably indirectly through the formation of tetraspanin-enriched-microdomains that organize fusogenic proteins at the membrane. This work aims to investigate the role of tetraspanins in MNGC formation during Burkholderia infection using the mouse macrophage J774.2 and human lung epithelial A549 cell lines, as they represent two of the cell types commonly infected in melioidosis. We used a B. thailandensis type VI secretion system mutant strain (Δtssk-5 which can infect cells without stimulating MNGC formation) to distinguish between tetraspanins involved in infection from those with a role in cell fusion infection. We first measured tetraspanin expression changes during B. thailandensis infection using qPCR. All 33 TSPANs had detectable mRNA in uninfected cells; 10 TSPANs were found to change post-infection but only 5 TSPANs showed consistent changes in both host cell types and no change with Δtssk-5. Of these, TSPAN-2, TSPAN-5, TSPAN-13 and CD9 mRNA levels appeared to correlate most closely with the onset of fusion in both cell types. Flow cytometry confirmed changes at the protein level for TSPANs (TSPAN-2 and TSPAN-13). The functional roles of these tetraspanins in bacterial infection and cell fusion assays were investigated by treating cells with antibodies, recombinant EC2 proteins and peptides derived from CD9 EC2. TSPANs (CD9, TSPAN-2 and TSPAN-13) were shown to have roles in infection and cell fusion, respectively. Finally, 10 adhesion molecules known to interact with TSPANs had detectable mRNA levels in uninfected cells; 5 of these were found to change after infection but only 3were affected in both host cell types and not with Δtssk-5 (ADAM10, CD172a and CD98).
1Al-Quds Nutrition and Health Research Institute, Faculty of Medicine, Al-Quds University, Palestine, Israel
2Biochemistry and Molecular Biology Department, Faculty of Medicine, Al-Quds University, Palestine, Israel 3Al-Quds Public Health Society, Palestine, Israel
Background: Methicillin-Resistant Staphylococcus aureus (MRSA) is a major cause of hospital-acquired and community-acquired infections. This study aimed to investigate the epidemiological and genetic diversity of clinical MRSA isolates in healthcare settings in Palestine, from 2015 to 2017.
Methods: All MRSA isolates from nine major hospitals in Palestine were collected. Each strain was characterized by: antibiotic susceptibility pattern, staphylococcal chromosomal cassette mec (SCCmec), Staphylococcus aureus protein A (spa) accessory gene regulator (agr) groups and a panel of toxin genes (including: Panton-Valentine Leukocidin (PVL), arcA, Toxic Shock Syndrome Toxin-1 (TSST-1) and Exfoliative Toxin A (ETA)), and genes that encode microbial surface components recognizing adhesive matrix molecules (MSCRAMMs): bbp, eno, cna, ebpS, fib, fnbA, fnbB, clfA and clfB.
Results: All the 112 isolates of MRSA were susceptible to Vancomycin. Resistance was observed to erythromycin, ciprofloxacin, clindamycin, gentamicin and trimethoprim sulfamethoxazole (SXT), 63.4%, 39.3%, 34.8%, 23.2% and 18.7., respectively. Of all the isolates, 32 isolates (28.6%) were MDR (i.e., were resistant to at least three different non-β-lactam antibiotic groups). The majority of the isolates were identified as SCCmec type IV (86.6%). Molecular typing identified 29 spa types representing 12 MLST clonal complexes (CC). The most prevalent spa types were t386 (CC01) (12.5%), spa t044 (CC80) (10.7%) and t008 (CC8) (10.7%). All four agr groups were present; agr group 1 and 3 were predominant (40.2%, 43.7%, respectively). PVL genes were detected in (29.5%) of all isolates, while ArcA genes were present in 18.8 % of all isolates and 23.2% had the TSST-1 toxin gene. The two most common spa types among TSST-1-positive isolates were the spa type t223 (CC22) and t021 (CC30). All spa type t991 was ETA positive (5.4%). In this study, USA300 clone (positive for PVL and ArcA genes) was found in 9 isolates (8.0%). None of the isolates harbored bap. Moreover, 92.8% isolates were positive for eno, 85.7% for clfA and clfB, 83.9% for fnbA, 66.9% for ebpS, 71.4% for fib, 48% for can and 29.5% for fnbB.
Conclusions: Our results provide insights into the epidemiology of MRSA strains in Palestine. We report a diversity of MRSA strains in hospitals in Palestine, with frequent SCCmecIV carriage and the genetic variations of adhesion genes require further investigation.
Keywords: MRSA, spa, SCCmec, PVL, ETA, TSST1, arcA, Biofilm
Biography:
Kifaya Azmi has completed her PhD from Charite University School of Medicine, Berlin. She is one of the staff members of Al-Quds University, faculty of medicine, Teaching biochemistry and molecular biology. Recently working on MRSA project, published more than 35 papers in reputed journals.
USIM, Malaysia
Psoriasis is a complex genetic disease involving both autoimmune and inflammatory components. It has worldwide prevalence and affects both men and women over a wide age range. In Malaysia, about 2% of the population are affected. The typical aetiology of psoriasis is unknown, but it is evident that an immune-mediated inflammatory disorder associated with complex genetic susceptibility is involved and treatments are basically symptomatic and non curative. The use of complementary and alternative medicines is common among psoriasis patients. In Malaysia, sweet potato leaves (SPL) of different cultivars have been used as a complementary or alternative therapy for psoriasis with a strong belief being effective. The mouse model of psoriasis was induced chemically by topical Imiquimoid application onto shaved skin of the examined BALB/c mice, following the same protocol described by van der fits et al. (2009). It was confirmed histologically that the resulting lesions were typical to psoriasis vulgaris as confirmed by dermatologist. Flow cytometry showed increased number of CD3+, CD4+, CD19 cells in the peripheral blood which is a sign of inflammatory reaction seen in acute psoriasis. Evaluation of the induced psoriasis lesionʼs response to the SPL concentrated extracts depended on clinical observation based on modified PASI (Psoriasis Area and Severity Index) score, histopathological examination of skin sections and molecular assays of TNF-alpha and INF-gamma as sensitive bio-markers of psoriasis. The examined lesions responded to topical lipophilic sweet potato leave (SPL) extract treatment with significant histopathological, immunological and molecular improvement compared to hydrophilic-SPL treated and negative control groups. The results showed clinical improvement in the lipophilic treated group, as scored by modified PASI. Histopathological changes showed significant reduction of epidermal thickness, reduced rete ridges size and less vascular formation in mice treated with lipophilic SPL extract. Immunological and molecular parameters showed significant decrease in the inflammatory biomarkers namely TNF-alpha and gamma IFN and the percentage of double negative T cells in both treated groups, though more pronounced in lipophilic SPL group. Overall, substantial evidence of the anti-inflammatory properties of SPL lipophilic extract in the examined mouse model of acute psoriasis thus should encourage further studies in human models.
Cardiff University, UK
Infectious diseases are considered a public health problem in most countries. The occurrence of antibiotic resistant pathogens is increasing worldwide. As antibiotics are presently losing their effectiveness, there is an urgent need to develop safe alternatives for treating bacterial infections especially those of the skin. One of these alternatives is “Probiotics”. Health-enhancing properties of probiotics should be performed by both in-vitro and in-vivo techniques. The greater wax moth Galleria mellonella is used as an in vivo model for host-pathogen interactions. A group of Lactobacillus type strains was obtained from culture collections, while the other group was isolated from fermented food products: yogurt and olives. Pathogens were isolated from skin infections patients. All the bacterial species were identified by 16S rRNA gene sequencing. In-vitro antagonistic activity of lactobacilli was investigated on the major causes of skin infections (Staphylococcus aureus and Streptococcus pyogenes) by performing both overlay and well diffusion assays during different incubation times and conditions.
In-vivo susceptibility of the wax moth larvae to both lactobacilli and pathogenic species was assessed by injection with serial dilutions of three preparations of each Lactobacillus food isolates: Bacterial suspension, the supernatant and washed cells. Serial dilutions of pathogenic bacteria suspensions were injected inside the larvae as well. All the tested pathogens were sensitive to the antibacterial effect of lactobacilli. The maximum antagonism was achieved after 72h under anaerobic incubation. Injection of the larvae with both of lactobacilli and pathogens displayed differences in the survival percentages of larvae.
Biography:
Duaa Al-Dulaimy graduated from Department of Biology/School of Biosciences/Al-Mustansiriyah University/Baghdad/Iraq. Duaa Al-Dulaimy started to work as a laboratory assistant at the same department. At this point, Al-Dulaimy has done a demonstration for undergraduate students in a couple of modules such as General Microbiology, Food Microbiology, Industrial Microbiology and Medical Microbiology. After this position, Al-Dulaimy studied a Master Degree in Microbiology at the same university and got degree to start working as an assistant lecturer for more than ten years. This academic and research position has developed scientific knowledge to supervise final year students and her research ability to perform several studies to produce articles and publish them in different journals. By this, she got a high level in job to be a lecturer in the mentioned university. Then, Al-Dulaimy got a scholarship from government to study PhD in the UK at the school of Biosciences/Cardiff University. Nowadays, she is at the third year of my project, which is investigating the antibacterial activity of Lactobacillus on skin infectionsʼ Pathogens: In-vitro and In-Vivo Studies.
1Shahid Sadoughi University of Medical Sciences, Iran
2Tehran University of Medical Sciences, Iran
Leishmaniasis is caused by protozoan of Leishmania parasite. The pentavalent antimonials compounds remain the first-line treatment. J-binding protein encoded by JBP1 involves in starting synthesis of J base that is unique in kinetoplastida. In this study, we were assessed the JBP1 gene expression in exposure of different doses of glucantime. L. major (MRHO/IR/75/ER) promastigotes were distributed in groups for exposure with glucantime with end concentrations of 5, 10, 15, and 20 mg/ml. Then, RNA was extracted and cDNA synthesis was performed. Gene expression of JBP1 was assessed using SYBR Green Real Time PCR by ΔΔCT analysis. Gene expression of all groups exposure with different concentrations of glucantime were the same but the JBP1 gene expression showed 1.4 fold decreasing in groups with promastigotes exposed with glucantime with the end concentration of 5 mg/ml. The base J is synthesized with JBP1. More synthesis of J base is resulted in decreasing RNA polymerase II that could affect the gene expression of other genes. In this study, we showed that in high concentrations of glucantime, the JBP1 was increased resulted in increasing in J base synthesis. Therefore, it seems that expression regulation of the genes involving in drug exposure would be in the other mechanisms.
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