Evaluation of the Role of Staphylococci in the Pathomechanism of Conjunctivitis

2021 ◽  
Author(s):  
Ewa Jasińska ◽  
Agnieszka Bogut ◽  
Agnieszka Magryś ◽  
Alina Olender
Keyword(s):  
Antibiotic Resistance ◽  
Epithelial Cells ◽  
Biofilm Formation ◽  
Methicillin Resistance ◽  
Microtiter Plate ◽  
Host Cells ◽  
Diffusion Method ◽  
Microtiter Plate Assay ◽  
Low Prevalence

Abstract Purpose: Determination of the association between ica genes and phenotypic biofilm formation in staphylococcal isolates involved in conjunctivitis, their antibiotic resistance as well as detection of selected virulence characteristics: adhesion to epithelial cells and in vitro cytotoxicity.Methods: The study included 26 Staphylococcus aureus (SA) and 26 Staphylococcus epidermidis (SE) isolates. The presence of icaAD genes and ica operon was determined by the PCR assay. Phenotypic biofilm formation was verified using the microtiter plate assay. Antibiotic resistance was performed using the disc diffusion method. Staphylococcal ability to attach to host cells was assessed by flow cytometry. Cytotoxicity on epithelial cells was evaluated by LDH assay.Results: The ica genes were detected in 26.9% of SE and in 42.3% of SA isolates. Only 15.3% of isolates (SE) were positive for both the icaAD and the ica operon. Phenotypically, 19.2% of SE isolates were strong biofilm producers, among which three were both icaAD- and ica operon-positive. 26.9% of SA isolates were strong biofilm producers. Methicillin resistance (MR) was detected in 34.6% of SE and 26.9% of SA isolates. 75% of MR isolates were multidrug resistant. SA isolates adhered to host cells more extensively than SE. SA isolates released higher level of LDH than SE.Conclusions: Adherence abilities were commonly observed in staphylococci associated with conjunctivitis. However, low prevalence of isolates positive for a complete and functional ica locus and low prevalence of strong biofilm producers was detected. SA adhered to a greater extent to eukaryotic cells than SE and were more cytotoxic.

scholarly journals Evaluation of the role of staphylococci in the pathomechanism of conjunctivitis

2021 ◽  
Author(s):  
Ewa Jasińska ◽  
Agnieszka Bogut ◽  
Agnieszka Magryś ◽  
Alina Olender
Keyword(s):  
Antibiotic Resistance ◽  
Epithelial Cells ◽  
Biofilm Formation ◽  
Methicillin Resistance ◽  
In Vitro Cytotoxicity ◽  
Host Cells ◽  
Diffusion Method ◽  
Microtitre Plate Assay ◽  
Low Prevalence

Abstract Purpose Determination of the association between ica genes and phenotypic biofilm formation in staphylococcal isolates involved in conjunctivitis, their antibiotic resistance as well as detection of selected virulence characteristics: adhesion to epithelial cells and in vitro cytotoxicity. Methods The study included 26 Staphylococcus aureus (SA) and 26 Staphylococcus epidermidis (SE) isolates. The presence of icaAD genes and ica operon was determined by the PCR assay. Phenotypic biofilm formation was verified using the microtitre plate assay. Antibiotic resistance was performed using the disc diffusion method. Staphylococcal ability to attach to host cells was assessed by flow cytometry. Cytotoxicity on epithelial cells was evaluated by LDH assay. Results The ica genes were detected in 26.9% of SE and in 42.3% of SA isolates. Only 15.3% of isolates (SE) were positive for both the icaAD and the ica operon. Phenotypically, 19.2% of SE isolates were strong biofilm producers, among which three were both icaAD- and ica operon-positive. About 26.9% of SA isolates were strong biofilm producers. Methicillin resistance (MR) was detected in 34.6% of SE and 26.9% of SA isolates. About 75% of MR isolates were multidrug resistant. SA isolates adhered to host cells more extensively than SE. SA isolates released higher level of LDH than SE. Conclusions Adherence abilities were commonly observed in staphylococci associated with conjunctivitis. However, low prevalence of isolates positive for a complete and functional ica locus and low prevalence of strong biofilm producers was detected. SA adhered to a greater extent to eukaryotic cells than SE and were more cytotoxic.

scholarly journals Antibiofilm and antivirulence potential of silver nanoparticles against multidrug-resistant Acinetobacter baumannii

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Helal F. Hetta ◽  
Israa M. S. Al-Kadmy ◽  
Saba Saadoon Khazaal ◽  
Suhad Abbas ◽  
Ahmed Suhail ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Antibacterial Activity ◽  
Acinetobacter Baumannii ◽  
Biofilm Formation ◽  
Microtiter Plate ◽  
Diffusion Method ◽  
Resistance Pattern ◽  
Infection Model ◽  
The Impact

AbstractWe aimed to isolate Acinetobacter baumannii (A. baumannii) from wound infections, determine their resistance and virulence profile, and assess the impact of Silver nanoparticles (AgNPs) on the bacterial growth, virulence and biofilm-related gene expression. AgNPs were synthesized and characterized using TEM, XRD and FTIR spectroscopy. A. baumannii (n = 200) were isolated and identified. Resistance pattern was determined and virulence genes (afa/draBC, cnf1, cnf2, csgA, cvaC, fimH, fyuA, ibeA, iutA, kpsMT II, PAI, papC, PapG II, III, sfa/focDE and traT) were screened using PCR. Biofilm formation was evaluated using Microtiter plate method. Then, the antimicrobial activity of AgNPs was evaluated by the well-diffusion method, growth kinetics and MIC determination. Inhibition of biofilm formation and the ability to disperse biofilms in exposure to AgNPs were evaluated. The effect of AgNPs on the expression of virulence and biofilm-related genes (bap, OmpA, abaI, csuA/B, A1S_2091, A1S_1510, A1S_0690, A1S_0114) were estimated using QRT-PCR. In vitro infection model for analyzing the antibacterial activity of AgNPs was done using a co-culture infection model of A. baumannii with human fibroblast skin cell line HFF-1 or Vero cell lines. A. baumannii had high level of resistance to antibiotics. Most of the isolates harbored the fimH, afa/draBC, cnf1, csgA and cnf2, and the majority of A. baumannii produced strong biofilms. AgNPs inhibited the growth of A. baumannii efficiently with MIC ranging from 4 to 25 µg/ml. A. baumannii showed a reduced growth rate in the presence of AgNPs. The inhibitory activity and the anti-biofilm activity of AgNPs were more pronounced against the weak biofilm producers. Moreover, AgNPs decreased the expression of kpsMII , afa/draBC,bap, OmpA, and csuA/B genes. The in vitro infection model revealed a significant antibacterial activity of AgNPs against extracellular and intracellular A. baumannii. AgNPs highly interrupted bacterial multiplication and biofilm formation. AgNPs downregulated the transcription level of important virulence and biofilm-related genes. Our findings provide an additional step towards understanding the mechanisms by which sliver nanoparticles interfere with the microbial spread and persistence.

scholarly journals Bacterial Profile of Urinary Tract Infections: Evaluation of Biofilm Formation and Antibiotic Resistance Pattern of Uropathogenic Escherichia coli

2020 ◽  
Vol 14 (4) ◽  
pp. 2577-2584
Author(s):  
Tariq Ahmad Shah ◽  
P. Preethishree ◽  
Ashwini ◽  
Vidya Pai
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Urinary Tract ◽  
Biofilm Formation ◽  
Urine Samples ◽  
Diffusion Method ◽  
E Coli ◽  
Biofilm Production ◽  
Tract Infections

Urinary tract infection (UTI) is one of the most common complaints in the outpatient clinic and a major health problem owing to the emergence of antibiotic resistance and biofilm formation. The objective of this study was to isolate and identify the causative bacterial agent of UTI and detect in vitro biofilm formation by Escherichia coli and investigate its correlation with antibiotic resistance. Urine samples from 519 patients with suspected UTIs were collected and processed by conventional microbiological procedures. Antimicrobial susceptibility testing for E. coli isolates was performed on Mueller Hinton agar (MHA) plates using the Kirby-Bauer disk diffusion method. Biofilm production was evaluated using the tissue culture plate method. Of 519 urine samples, 115 (22.1%) showed significant bacteriuria. The most common isolate was E. coli (n=57, 49.6%), followed by Klebsiella spp. (n=23, 20%). All E. coli isolates were evaluated for their ability to form biofilms in vitro. Of 57 isolates, 50 (87.7%) were biofilm producers and 7 (12.3%) were non-biofilm producers. Antibiogram of E. coli isolates revealed the highest resistance to ampicillin (96.5%) and nitrofurantoin (91.2%), followed by amoxyclav (82.5%), ceftazidime (73.7%), cefepime (71.9%), and tetracycline (71.9%). A significant association (p<0.05) was observed between biofilm formation and resistance to amoxyclav, ceftazidime, cefepime, imipenem, and nitrofurantoin. A significant correlation was noted between biofilm production and antibiotic resistance. Hence, screening of all isolates of uropathogenic E. coli for biofilm production and studying their antibiogram would allow appropriate choice of antibiotic therapy.

scholarly journals Analysis of the bacterial biofilm formation in different models of the in vitro culture

2021 ◽  
Vol 19 (1) ◽  
pp. 40-45
Author(s):  
Agnieszka Bogut ◽  
◽  
Agnieszka Magryś ◽  
Keyword(s):  
Biofilm Formation ◽  
Culture Media ◽  
Microtiter Plate ◽  
Bacterial Biofilm ◽  
Atmospheric Conditions ◽  
Experimental Conditions ◽  
Microtiter Plate Assay ◽  
Biofilm Production ◽  
Strain Type

Introduction. Microtiter plate assay (MPA) remains one of workhorses of in vitro biofilm research but it requires optimization of experimental conditions to fulfill the biofilm formation requirements of different bacterial pathogens. Aim. The aim was to determine the effect of TSB and RPMI1640 culture media and selected culture variables (O2 vs. 5% CO2, extended incubation time) on the biofilm production by bacteria commonly involved in biofilm-related infections: Enterococcus faecalis (EF), Escherichia coli (EC), Staphylococcus aureus (SA), Pseudomonas aeruginosa (PA), Klebsiella pneumoniae (KP). Material and methods. The investigation was performed using the MPA with crystal violet. Results. Statistically significant (p<0.05) increase in biofilm production between 24h and 72h time points was observed for EF (TSB o2, RPMIo2 and RPMIco2), EC (TSBo2), SA (TSBo2, TSBco2), KP (TSBo2, TSBco2), PA (RPMIco2, TSBco2). The TSB caused a significantly greater stimulation of biofilm production compared to RPM1640. It outcompeted RPMI1640 irrespective of the atmospheric conditions for SA and KP and under aerobic conditions for EF. Conclusion. Although the TSB provided the most optimal conditions for biofilm production, the process was influenced by the strain type, atmospheric conditions and period of cultivation which limits the ability to design a single universal model of the in vitro biofilm investigation.

scholarly journals The biofilm formation ability of Listeria monocytogenes isolated from meat, poultry, fish and processing plant environments is related to serotype and pathogenic profile of the strains

2012 ◽  
Vol 2 (1) ◽  
pp. 12 ◽  
Author(s):  
Domenico Meloni ◽  
Roberta Mazza ◽  
Francesca Piras ◽  
Sonia Lamon ◽  
Simonetta Gianna Consolati ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Biofilm Formation ◽  
Target Genes ◽  
Microtiter Plate ◽  
Food Sources ◽  
Processing Plant ◽  
Pathogenic Potential ◽  
Microtiter Plate Assay ◽  
Processing Plants

In the present study, the relationships between serotype, pathogenic profile and <em>in vitro</em> biofilm formation of 106 <em>Listeria monocytogenes</em> strains, having no epidemiological correlation and isolated from different environmental and food sources, were analyzed. The quantitative assessment of the <em>in vitro</em> biofilm formation was carried out by using a microtiter plate assay with spectrophotometric reading (OD620). The isolates were also submitted to serogrouping using the target genes <em>lmo0737</em>, <em>lmo1118, ORF2819, ORF2110, prs</em>, and to the evaluation of the presence of the following virulence genes: <em>prfA, hlyA, rrn, inlA, inlB, iap, plcA, plcB, actA</em> and <em>mpl</em>, by multiplex PCRs. The 62% of the strains showed weak or moderate <em>in vitro</em> ability in biofilm formation, in particular serotypes 1/2b and 4b, frequently associated with sporadic or epidemic listeriosis cases. The 25% of these isolates showed polymorphism for the <em>actA</em> gene, producing a fragment of 268-bp instead of the expected 385-bp. The deletion of nucleotides in this gene seems to be related to enhanced virulence properties among these strains. Strains belonging to serotypes associated with human infections and characterized by pathogenic potential are capable to persist within the processing plants forming biofilm.

Influence of Temperature, Source, and Serotype on Biofilm Formation of Salmonella enterica Isolates from Pig Slaughterhouses

2015 ◽  
Vol 78 (10) ◽  
pp. 1875-1878 ◽  
Author(s):  
FRANCESCA PIRAS ◽  
FEDERICA FOIS ◽  
SIMONETTA GIANNA CONSOLATI ◽  
ROBERTA MAZZA ◽  
RINA MAZZETTE
Keyword(s):  
Optical Density ◽  
Salmonella Enterica ◽  
Biofilm Formation ◽  
Microtiter Plate ◽  
Cross Contamination ◽  
Meat Processing ◽  
Microtiter Plate Assay ◽  
Antimicrobial Resistance Profile ◽  
Density Values

Quantitative assessment of in vitro biofilm formation by 40 Salmonella enterica isolates isolated in pig abattoirs from animal and environmental sources (surfaces in contact and not in contact with meat) and classified in eight seroytpes was carried out by using a microtiter plate assay with spectrophotometric reading (optical density at 620 nm). Biofilm-forming ability was statistically correlated with the temperature of incubation (22 and 35°C), the source of the isolates, and the antimicrobial resistance profile. After incubation at 35°C, 9 isolates (22.5 %) were classified as weak biofilm producers. After incubation at 22°C, 25 isolates (62.5%) were classified as weak producers and 3 (7.5%) as moderate producers. The quantity of biofilm formed after incubation at 22°C was significantly higher (P &lt; 0.01) than at 35°C. This result is notable because 22°C is a common temperature in meat processing facilities and in slaughterhouses. At 35°C, isolates detected from surfaces in contact with meat showed significantly higher (P &lt; 0.1) optical density values compared to isolates from other samples, highlighting the risk of cross-contamination for carcasses and offal. No correlation was detected between quantity of biofilm and serotype or between biofilm formation and resistance to antimicrobials.

scholarly journals The Anti-Biofilm Activity of Oregano Essential Oil Against Dental Plaque-Forming Streptococcus mutans In Vitro and In Vivo

2020 ◽  
Vol 24 (3) ◽  
Author(s):  
Fatemeh Hejazinia ◽  
Leila Fozouni ◽  
Nasrin Sadat Azami ◽  
Seyedgholamreza Mousavi
Keyword(s):  
Essential Oil ◽  
Elementary School Students ◽  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Microtiter Plate ◽  
School Students ◽  
Microtiter Plate Assay ◽  
Oregano Essential Oil

Background: The oral and dental infections that are mainly caused by bacterial biofilms are among the most prevalent human infections worldwide. Objectives: The present study aimed to investigate the in vitro and in vivo inhibitory and anti-biofilm effects of oregano essential oil on the Streptococcus mutans isolates obtained from elementary school students. Methods: This experimental study was conducted on 150 samples collected from the buccal and lingual surfaces of the posterior teeth of elementary school students. S. mutans strains were identified using conventional microbiological and biochemical tests, and biofilm formation was assessed using the microtiter plate assay. The minimum inhibitory concentration (MIC) of the oregano essential oil against the isolates was determined using the broth microdilution method. In addition, the effective constituents of the essential oil were measured via gas chromatography-mass spectroscopy. The in vitro and in vivo anti-biofilm activities of the oregano essential oil were also evaluated using the modified microtiter plate assay and on the tooth surfaces of male NMRI mice, respectively. Results: The frequency of S. mutans was 15.3%, 87% of which were capable of biofilm formation. The MIC of the oregano essential oil was 50 µl/ml against the S. mutans isolates, and 82% of the isolates did not grow at the concentrations of ≥ 512 µl/ml. However, none of the isolates were capable of biofilm formation at the MIC and sub-MIC concentrations of the essential oil. Limonene and myrcene were the most effective constituents of the essential oil. Furthermore, a significant correlation was observed between treatment with the oregano essential oil and biofilm formation by the streptococci isolates (P = 0.05). Conclusions: According to the results, the presence of biofilm and incidence of dental caries were significantly correlated. Moreover, the essential oil of oregano and its main constituents had potent anti-biofilm and antibacterial properties and could be utilized for the production of new plant-based mouthwashes.

scholarly journals Genotypic and virulence characteristics of Listeria monocytogenes recovered from food items in Lebanon

2016 ◽  
Vol 10 (07) ◽  
pp. 712-717 ◽  
Author(s):  
Nathaline Haidar-Ahmad ◽  
Kohar Annie B Kissoyan ◽  
Sukayna M Fadlallah ◽  
Rima El-Hajj ◽  
Majd Saleh ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Biofilm Formation ◽  
Food Selection ◽  
Pcr Amplification ◽  
Food Products ◽  
Microtiter Plate ◽  
Pathogenic Potential ◽  
Microtiter Plate Assay ◽  
Imported Food

Introduction: Listeria monocytogenes is the agent of listeriosis, a life threatening foodborne disease for immunocompromised patients and pregnant women. This bacterium is not routinely screened for in Lebanon and there is lack of data about the prevalent strains and their potential pathogenicity. To that purpose, this study was undertaken to characterize L. monocytogenes from various food products, by assessing the in vitro biofilm forming ability, detecting their virulence potential, and characterizing them at the strain level. Methodology: Fifty-nine isolates were obtained from the Lebanese Agriculture Research Institute (LARI). They were collected in 2012-2013 from local and imported food products in the Lebanese market. Biofilm formation was measured using the Microtiter Plate Assay. PCR amplification was performed for three main virulence genes; hly, actA, and inlB. Pulsed field gel electrophoresis (PFGE) and BIONUMERICS analysis were carried out. Results: Lebanese isolates from cheese and raw meat showed higher biofilm formation than imported and Lebanese seafood isolates. A total of 100% of the isolates were PCR positive for hly and actA genes and 98.3% for inlB gene. PFGE analysis demonstrated the prevalence of 13 different subtypes with 100% similarity. Detected subtypes were grouped into 6 clusters of 90% genomic similarity. Clustered subtypes were particular to the country of origin. Conclusion: This study highlights the presence of L. monocytogenes in the Lebanese food market with high pathogenic potential and stresses the importance of enhanced surveillance and the implementation of strict regulations on local and imported food. Future investigations may be conducted on a larger food selection.

scholarly journals Investigation on Antibiotic-Resistance, Biofilm Formation and Virulence Factors in Multi Drug Resistant and Non Multi Drug Resistant Staphylococcus pseudintermedius

2019 ◽  
Vol 7 (12) ◽  
pp. 702 ◽  
Author(s):  
Gabriele Meroni ◽  
Joel F. Soares Filipe ◽  
Lorenzo Drago ◽  
Piera A. Martino
Keyword(s):  
Antibiotic Resistance ◽  
Virulence Factors ◽  
Biofilm Formation ◽  
Microtiter Plate ◽  
Drug Resistant ◽  
Virulence Determinants ◽  
Commensal Bacterium ◽  
Staphylococcus Pseudintermedius ◽  
Microtiter Plate Assay ◽  
Human Agent

Staphylococcus pseudintermedius is a commensal bacterium frequently isolated from canine skin and recognized as a zoonotic agent especially for dog-owners. This study focused on (a) the antibiotic-resistance phenotypes; (b) the ability to produce biofilm (slime); and (c) the dissemination of virulence factors in S. pseudintermedius strains. Seventy-three S. pseudintermedius strains were screened for antibiotic-resistance against 22 different molecules by means of Kirby-Bauer assay. The ability to produce biofilm was investigated using the microtiter plate assay (MtP) and the amplification of icaA and icaD genes. Virulence factors such as cytotoxins (lukI), enterotoxins (seC), and exfoliative toxins (siet, expA, and expB) were evaluated. The antibiotic-resistance profiles revealed 42/73 (57%) multi-drug resistant (MDR) strains and 31/73 (43%) not-MDR. All the MDR strains and 8/31 (27%) of not-MDR resulted in biofilm producers. Leukotoxin LukI was found in 70/73 (96%) of the isolates. Moreover, the enterotoxin gene seC was detected in 47/73 (64%) of the strains. All the isolates carried the siet gene, whereas expA and expB were found in 3/73 (4%) and 5/73 (7%), respectively. In conclusion, S. pseudintermedius should be considered a potential zoonotic and human agent able to carry different virulence determinants and capable of producing biofilm which facilitates horizontal gene transfer.

scholarly journals Effect of sodium chloride and temperature on biofilm formation and virulence of Flavobacterium columnare isolated from striped catfish (Pangasianodon hypophthalmus)

2020 ◽  
Vol 12 (3) ◽  
Author(s):  
Nguyen Thi Kim My ◽  
Tu Thanh Dung ◽  
Channarong Rodkhum ◽  
Dong Thanh Ha
Keyword(s):  
Biofilm Formation ◽  
In Vitro Study ◽  
Microtiter Plate ◽  
Flavobacterium Columnare ◽  
Immersion Method ◽  
Microtiter Plate Assay ◽  
Pangasianodon Hypophthalmus ◽  
Striped Catfish

This research was conducted to investigate the biofilm formation ability at various salt concentrations and temperatures of Flavobacterium columnare isolated from striped catfish (Pangasianodon hypophthalmus) at Can Tho University. Microtiter plate assay and the in vivo challenge were used to test the virulence of this strain of F. columnare for 10 days by immersion method at different salt concentrations (0, 3, 6, 9, 12 and 15 ppt). Results showed that biofilm formation of F.columnare was inhibited at 3 and 6 ppt and stronger reductions were recorded at 9, 12 and 15 ppt. In the same trend, the higher temperature the lower biofilm formation, the highest biofilm formation was at 25°C treatment, then it was reduced at 28 and 31°C and at 35°C the formed biofilm was greatly reduced. Interestingly, there were no statistically significant differences between 28 and 31°C (P>0.05). The virulent study found that 100% fish died after 1 day post challenge at 0 ppt. There were 10% and 25% of fish died at 3 and 6 ppt respectively. No dead fish was found at 9 and 12 ppt. In conclusion, biofilm formation was inhibited at 3 ppt, was almost controlled at 9, 12 and 15 ppt and was also mostly reduced at 31°C at least in the in-vitro study. Furthermore, the virulence of this bacterial strain was controlled 90% at 3 ppt and completely controlled (100%) at 9, 12 and 15 ppt.

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