diffusion assay
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2021 ◽  
pp. 913-920
Author(s):  
Ľ Janovičová ◽  
B. Gromová ◽  
D. Drobná ◽  
B. Konečná ◽  
E. Renczés ◽  
...  

Extracellular DNA (ecDNA) activates immune cells and is involved in the pathogenesis of diseases associated with inflammation such as sepsis, rheumatoid arthritis or metabolic syndrome. DNA can be cleaved by deoxyribonucleases (DNases), some of which are secreted out of cells. The aim of this experiment was to describe plasma DNase activity in relation to extracellular DNA in adult rats, to analyse potential sex differences and to prove whether they are related to endogenous testosterone. Adult Lewis rats (n=28) of both sexes were included in the experiment. Male rats were gonadectomized or sham-operated and compared to intact female rats. Plasma ecDNA and DNase activity were measured using fluorometry and single radial enzyme diffusion assay, respectively. Concentrations of nuclear ecDNA and mitochondrial ecDNA were determined using real-time PCR. Females had 60% higher plasma DNase activity than males (p=0.03). Gonadectomy did not affect plasma DNase in males. Neither the concentration of total ecDNA, nor nuclear or mitochondrial DNA in plasma differed between the groups. No significant correlations between DNase and ecDNA were found. From previous studies on mice, it was expected, that male rats will have higher DNase activity. In contrast, our study in rats showed the opposite sex difference. This sex difference seems not to be caused by endogenous testosterone. Interestingly, no sex differences were observed in plasma ecDNA suggesting a complex or missing association between plasma ecDNA and DNase. The observed sex difference in plasma DNase should be taken into account in animal models of ecDNA-associated diseases.


Author(s):  
Anita Rana

Microorganisms and helminthes can cause serious diseases in humans as well as in animals. The use of antimicrobial and antihelminthic drugs have created selective pressure and caused resistance to antibiotics used against them, thus it necessitates the use of honey bee’s derived natural products. One such bee derived product is pollen, collected by worker honey bees from the flowering plants and modify it by adding its salivary secretions. The present study embodies use of pollen as antimicrobial and antihelminthic substance. Among microorganisms 4 Gram (+ve) bacteria; (Bacillus subtilis, Staphylococcus epidermidis, Staphylococcus aureus, Streptococcus pneumoniae) and 3 Gram (-ve) bacteria; (Escherichia Coli, Pseudomonas aeruginosa, Salmonella enteric) and 2 yeasts (Candida albicans and Saccharomyces cerevisiae) were used and the methodology used disc diffusion assay and broth dilution method. The antihelminthic effect was observed among amphistomes via bioassay method under in vitro conditions. For observations three types of pollen extracts (ethanolic, methanolic and water extract) were prepared and positive controls used were; Ampicillin for antibacterial, Amphotericin B for antifungal and Albendazole for anti-helminthes. The antimicrobial activities were determined by measuring the zones of inhibition diameters in millimeters after 24 hours of incubation at optimum temperature for each microbe and also by broth dilution method. Results obtained showed that the water extract of pollen was found to be most effective against bacteria used in the present study where; Gram (+ve) bacteria were more susceptible as compared to the Gram (-ve) bacteria. It was also observed that among yeasts; Saccharomyces cerevisiae was more susceptible towards ethanolic extract of pollen while Candida albicans showed more inhibitions towards water extract of pollen. Results also demonstrated that none of the extracts of pollen was found to be effective against Helminthes (amphistomes) used in the present study.


Pathogens ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1574
Author(s):  
Munaf AL-Dulaimi ◽  
Ammar Algburi ◽  
Alyaa Abdelhameed ◽  
Maria S. Mazanko ◽  
Dmitry V. Rudoy ◽  
...  

Acinetobacter spp., the nosocomial pathogen, forms strong biofilms and is resistant to numerous antibiotics, causing persistent infections. This study investigates the antibacterial and anti-biofilm activity of polymyxin E alone and in combination with the cell-free supernatants (CFS) of the tested probiotic bacilli, Bacillus subtilis KATMIRA1933 and Bacillus amyloliquefaciens B-1895 against the selected Acinetobacter spp. starins. Three isolates of Acinetobacter spp., designated as Acinetobacter spp. isolate 1; Acinetobacter spp. isolate 2, and Acinetobacter spp. isolate 3, were collected from patients with burns, wounds, and blood infections, respectively. Bacterial identification and antibiotic susceptibility testing were conducted using the VITEK2 system. Auto-aggregation and coaggregation of the tested bacilli strains with the selected Acinetobacter spp. isolates were evaluated. A disk diffusion assay was used to identify the microorganism’s susceptibility to the selected antibiotics, alone and in combination with the CFS of the bacilli. The MIC and MBIC (minimum inhibitory and minimum biofilm inhibitory concentrations) of polymyxin E combined with bacilli CFS were determined. Acinetobacter spp. isolates were (i) sensitive to polymyxin E, (ii) able to form a strong biofilm, and (iii) resistant to the tested antibiotics and the CFS of tested bacilli. Significant inhibition of biofilm formation was noticed when CFS of the tested bacilli were combined with polymyxin E. The bacilli CFS showed synergy with polymyxin E against planktonic cells and biofilms of the isolated pathogens.


2021 ◽  
Vol 2 (4) ◽  
pp. 306-319
Author(s):  
Mostafa Essam Eissa ◽  
Engy Refaat Rashed ◽  
Dalia Essam Eissa

Till nowadays microbiological assay is still widely used with several antibiotics that are composed of a mixture of related active compounds. However, obtaining a reasonably valid determination of the potency is dependent on the validity and the suitability of the assay design. The present work aimed to validate an assay design of an aminoglycoside antibiotic (Gentamicin Sulfate) using a two-dose Parallel Line Model agar diffusion assay in a large 8×8 rectangular plate. All preparatory procedures were done following United States Pharmacopeia and the Inhibition Zones were measured using a digital caliper to the nearest 0.01 mm. Analysis of variance of compendial requirements of regression and parallelism were found to be satisfactorily meeting the acceptance criteria. Specificity was achieved for the product under investigation with no detectable IZ that could be found for all components except the antibiotic. The validation method showed acceptable linearity of r2≥0.98. Accuracy and precision parameters showed RSD (%)<2. All relative error value estimates were below 4%. The proposed validation design for 32×32 cm antibiotic plates yielded valid results and can be projected for the routine Quality Control analysis of the antibiotic material, especially which is incorporated into a finished medicinal dosage form. Doi: 10.28991/HIJ-2021-02-04-04 Full Text: PDF


2021 ◽  
Vol 8 (12) ◽  
pp. 295
Author(s):  
Salem Djebala ◽  
Julien Evrard ◽  
Fabien Gregoire ◽  
Calixte Bayrou ◽  
Linde Gille ◽  
...  

The aim of this study was to identify the species and antimicrobial susceptibility of bacteria involved in parietal fibrinous peritonitis (PFP). We studied 156 peritoneal fluid samples from cows presenting PFP after caesarean section. Bacteria were cultured in selective media and their antimicrobial susceptibility was tested by disk diffusion assay. Bacteria were isolated in the majority (129/156; 83%) of samples. The majority (82/129; 63%) of positive samples contained one dominant species, while two or more species were cultured in 47/129 (36%) samples. Trueperella pyogenes (T. Pyogenes) (107 strains) was the most identified species, followed by Escherichia coli (E. coli) (38 strains), Proteus mirabilis (P. mirabilis) (6 strains), and Clostridium perfringens (C. perfringens) (6 strains). Several other species were sporadically identified. Antimicrobial susceptibility was tested in 59/185 strains, predominantly E. coli (38 strains) and P. mirabilis (6 strains). Antibiotic resistance, including resistance to molecules of critical importance, was commonly observed; strains were classified as weakly drug resistant (22/59; 37%), multidrug resistant (24/59; 41%), extensively drug resistant (12/59; 20%), or pan-drug resistant (1/59; 2%). In conclusion, extensive antibiotic resistance in the isolated germs might contribute to treatment failure. Ideally, antimicrobial therapy of PFP should be based upon bacterial culture and susceptibility testing.


2021 ◽  
Vol 71 (1) ◽  
Author(s):  
Emmanuel Edoghogho Imade ◽  
Solomon Esharegoma Omonigho ◽  
Olubukola Oluranti Babalola ◽  
Ben Jesuorsemwen Enagbonma

Abstract Purpose Incidence of foodborne diseases and growing resistance of pathogens to classical antibiotics is a major concern in the food industry. Consequently, there is increasing demand for safe foods with fewer chemical additives but natural products which are not harmful to the consumers. Bacteriocins, produced by lactic acid bacteria (LAB), is of interest because they are active in a nanomolar range, do not have toxic effects, and are readily available in fermented food products. Methods In this research, LAB were isolated from fufu, gari, kunu, nono, and ogi using De Mann, Rogosa, and Sharpe agar. Cell-free supernatants were prepared from 18-24 h LAB culture grown on MRS broth. Effect of organic acid was eliminated by adjusting the pH of the supernatants to 7.0 with 1M NaOH while the effect of hydrogen peroxide was eliminated by treating with Catalase enzyme. The supernatant was then filter-sterilized using a membrane filtration unit with a 0.2-μm pore size millipore filter and subjected to agar well diffusion assay against foodborne antibiotic-resistant bacteria. Result A total of 162 isolates were obtained from the food samples. The antimicrobial sensitivity test yielded positive results for 45 LAB isolates against Staphylococcus aureus ATCC 25923 while 52 LAB isolates inhibited Escherichia coli ATCC 25922. On confirmation of the bacteriocinogenic nature of the inhibitory substance, 4 of the LAB isolates displayed a remarkable degree of inhibition to Leuconostoc mesenteroides, Salmonella typhimurium, and Bacillus cereus. Agar well diffusion assay was also performed against antibiotic-resistant foodborne pathogens using the cell-free supernatant (CFS) obtained from Lactobacillus fermentum strain NBRC15885 (Limosilactobacillus fermentum), Lactobacillus fermentum strain CIP102980 (Limosilactobacillus fermentum), Lactobacillus plantarum strain JCM1149 (Lactiplantibacillus garii), and Lactobacillus natensis strain LP33 (Companilactobacillus nantensis). The foodborne pathogens exhibited a notable level of resistance to antibiotics, with B. cereus exhibiting a resistance profile of 40%, S. aureus (50%), K. pnuemoniae (70%), E. coli (60%), and S. typhi (40%). The (CFS) was able to inhibit the growth of B. cereus, Klebsiella pneumonia, S. typhimurium, S. aureus, and E. coli. Conclusion Therefore, it portends that the bacteriocins produced by the LAB isolated from these food products could act as probiotics for effective inhibition of the growth of antibiotic-resistant foodborne pathogens.


2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Suranat Phonghanpot ◽  
Faongchat Jarintanan

Abstract Background Bauhinia strychnifolia Craib is an herb in Thai traditional medicine. Its decoction is traditionally used as an anticancer, antidiarrheal, and hangover remedy for centuries. Several studies described bioactivities of its organic solvent extracts, however, only few demonstrated the usefulness of the decoction. Here, we aimed to determine the bioactivities of Bauhinia strychnifolia Craib root and stem aqueous extracts in gut and liver perspective. Methods To achieve the goal, we performed MTT test, microscopic analyses, disc diffusion assay, broth microdilution assay, free radicals scavenging assays, and LC-MS analysis. Results We found that the extracts inhibited the growth of human hepatocellular carcinoma (HepG2) and colon adenocarcinoma (HT-29) cell lines. Moreover, they also inhibited the growth of gram-positive bacteria Staphylococcus aureus and Bacillus cereus but not inhibited the growth of gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa. Furthermore, the extracts exhibited moderate antioxidant activity and increased GSH production in HepG2 cell line when compared with untreated. Our LC-MS analysis confirmed the existence of anticancer and antioxidant; 3,5,7,3′,5′-pentahydroxyflavanonol-3-O-α-L-rhamnopyranoside and β-sitosterol, in the extracts. Conclusion The results from our study supported that the administration of Bauhinia strychnifolia Craib root and stem decoction would really aid colon or liver cancer patients and detoxify the alcoholic drunkard as it is claimed in Thai traditional medicine.


Pharmaceutics ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 1783
Author(s):  
Alessandra Cataldo Cataldo Russomando ◽  
Ronit Vogt Vogt Sionov ◽  
Michael Friedman ◽  
Irith Gati ◽  
Ron Eliashar ◽  
...  

The aim of the study was to develop a sustained-release varnish (SRV) containing chlorhexidine (CHX) for sinonasal stents (SNS) to reduce bacterial growth and biofilm formation in the sinonasal cavity. Segments of SNS were coated with SRV-CHX or SRV-placebo and exposed daily to bacterial cultures of Staphylococcus aureus subsp. aureus ATCC 25923 or Pseudomonas aeruginosa ATCC HER-1018 (PAO1). Anti-bacterial effects were assessed by disc diffusion assay and planktonic-based activity assay. Biofilm formation on the coated stents was visualized by confocal laser scanning microscopy (CLSM) and high-resolution scanning electron microscopy (HR-SEM). The metabolic activity of the biofilms was determined using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) method. Disc diffusion assay showed that SRV-CHX-coated SNS segments inhibited bacterial growth of S. aureus subsp. aureus ATCC 25923 for 26 days and P. aeruginosa ATCC HER-1018 for 19 days. CHX was released from coated SNS segments in a pH 6 medium up to 30 days, resulting in growth inhibition of S. aureus subsp. aureus ATCC 25923 for 22 days and P. aeruginosa ATCC HER-1018 for 24 days. The MTT assay showed a reduction of biofilm growth on the coated SNS by 69% for S. aureus subsp. aureus ATCC 25923 and 40% for P. aeruginosa ATCC HER-1018 compared to the placebo stent after repeated exposure to planktonic growing bacteria. CLSM and HR-SEM showed a significant reduction of biofilm formation on the SRV-CHX-coated SNS segments. Coating of SNS with SRV-CHX maintains a sustained delivery of CHX, providing an inhibitory effect on the bacterial growth of S. aureus subsp. aureus ATCC 25923 and P. aeruginosa ATCC HER-1018 for approximately 3 weeks.


Author(s):  
Lijo John ◽  
Lijo John ◽  
Lijo John ◽  
Lijo John ◽  
Lijo John

Pigeon breeding has transformed from being a mere hobby to becoming established as an industry. The increased trade of pigeons inadvertently invites the risk of dissemination of infections including zoonoses like salmonellosis. Pigeons once infected remain carriers for life. This coupled with the ability of the organism to acquire antimicrobial resistance makes salmonellosis, particularly from pigeons an important, public health risk for pigeon handlers. Cloacal swabs from a total of 200 exotic pigeons belonging to 24 lofts from Northern districts of Kerala were collected and attempted to isolate Salmonella and understand its antimicrobial resistance profile. Five isolates of salmonella could be obtained from four of the lofts studied. A prevalence of 2.5 per cent was identified for salmonellosis with 16.67 per cent of the lofts affected. Antimicrobial sensitivity based on disk diffusion assay revealed that all the five isolates were sensitive to amoxicillin-clavulanate and all were resistant to tetracycline and streptomycin. Sixty per cent of the isolates were sensitive to co-trimoxazole, chloramphenicol, ampicillin, cefoperazone, amikacin and gentamicin.


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