Congo Red Method Research Articles

Identification of biofilm-producing microorganism’s and drug susceptibility pattern from diabetic foot ulcer patients at Puducherry

Diabetes mellitus is a significant health problem worldwide that affects approximately 171 million people; severe complications lead to the development of diabetic foot ulcers. Diabetic ulcer infections are mainly polymicrobial in nature and multidrug-resistant (MDR), which is capable of forming a biofilm, which is the important virulence factor results in treatment failure. The main objectives of this study to investigate the etiologic agents of diabetic foot infections, their antimicrobial resistance and biofilm formation. A total of 200 patient samples were taken from diabetic foot ulcer patients between September 2015 and February 2016. Isolation and identification of microorganism were made according to standard microbiological procedures. Antibiotic Susceptibility testing performed by Kirby Bauer disc diffusion method and the biofilm production was performed by the tube method and Congo Red Method. Out of 200 samples processed, 110 (55%) were polymicrobial, 50 (25%) monomicrobial and 40(20 %)culture Sterile. The most common organism isolated were 82(39%) Pseudomonas aeruginosa,45(21%) Staphylococcus aureus, 48(23%) Candida sp followed by others. Biofilm production was seen in 112 (53%) of the isolates. Antimicrobial drug resistance was higher among 92(82%) biofilm producers than non-biofilm 20(18%) producing microorganisms. Organisms isolated from chronic diabetic foot ulcers cases were multidrug-resistant and biofilm producers. Our study shows the importance of biofilm screening with the usual antibiogram, as a routine technique in diabetic foot ulcers patients for effective treatment.

Detection of bioflim production among Staphylococcus.aureus by Congo red method and tube method

: Bioflim is a mode of survival for various microbe by which they form aggregates during unfavourable conditions. is a major cause of nosocomial and community acquired infections.A total of 150 isolates were screened for biofilm production by Congored method and Tube method following standard guidelines.Of the 150 isolates, 85(56.6%) were MSSA and 65(43.4%) were MRSA. On Congored agar, 63 isolates showed black colonies with dry crystalline consistency indicating biofilm production. Out of 63 isolates 62% of isolates were MRSA and 38% of isolates were MSSA. Biofilm by tube method, 84isolates showed biofilm production. MSSA were 48.2% and MRSA were 81.53%.MRSA is the significant biofilm producer when compare to MSSA, Congored method is less accurate when compare to tube method as screening test for the detection of biofilm.

Evaluation of Polish wild Mushrooms as Beta-Glucan Sources.

Mushroom beta-glucans show immunomodulatory, anticancer and antioxidant features. Numerous papers have been published in the last years on fungal polysaccharides, especially beta-glucans, demonstrating their various biological activities. However substantial data about beta-glucan contents in many mushroom species, especially wild mushrooms, are still missing. Therefore, the main objective of the study was to evaluate β-glucans in 18 species of wild mushrooms and three species of commercial mushrooms for comparison purposes. The contents of β-glucans were determined by the Megazyme method and with the Congo red method, which differ in analytical procedure. Among wild mushrooms, the highest mean β-glucan content assessed with the Megazyme method was found in Tricholoma portentosum (34.97 g/100 g DM), whereas with the Congo red method in Lactarius deliciosus (17.11 g/100 g DM) and Suillus grevillei (16.97 g/100 g DM). The β-glucans in wild mushrooms assessed with the Megazyme method were comparable to commercial mushrooms, whereas β-glucans assessed with the Congo red method were generally higher in wild mushrooms, especially in Russula vinosa, L. deliciosus and S. grevillei. This study indicates wild mushrooms as interesting material for β-glucan extraction for food industry and medicinal purposes.

Antimicrobial Resistance Profile and Biofilm Production of Microorganisms Isolated from Oropharynx of Rupornis magnirostris (Gmelin, 1788) and Caracara plancus (Miller, 1777).

The aim of this preliminary study was to identify microorganisms with antimicrobial resistance profile and biofilm producers in oropharynx of Rupornis magnirostris and Caracara plancus. Six R. magnirostris and six C. plancus maintained in Triage Center for Wild Animals (CETAS) facilities were studied. Coagulase-positive staphylococci (CoPS), enterobacteria, and yeasts were identified by the biochemical analysis or MALDI-TOF mass spectrometry. The resistance profile of the microorganisms was analyzed according to CLSI. The biofilm production was evaluated by Congo red and violet crystal staining methods. Among the 12 birds, 10 presented strains of CoPS and/or enterobacteria with resistance profile, such as methicillin-resistant CoPS (MR-CoPS), vancomycin-resistant CoPS (VR-CoPS), extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL), and Klebsiella pneumoniae carbapenemase- (KPC-) producing bacteria. Regards the fungal analysis, Candida spp., Cryptococcus spp., Rhodotorula mucilaginosa, R. glutinis, and Trichosporon coremiiforme were identified. All the Trichosporon coremiiforme strains were resistant to amphotericin B, as well as all the Rhodotorula mucilaginosa exhibited resistance to fluconazole. Related to the biofilm production, among the 8 CoPS, 27 enterobacteria, and 10 yeasts isolates, 3, 16, and 7 strains were biofilm producers, respectively. Thus, the presence of these microorganisms in birds of prey is worrisome, highlighting its possible influence in the spread of infections in urban centers.

Effect of bile on growth and biofilm formation of non-typhoidal salmonella serovars isolated from seafood and poultry

Bacterial cells adopt various strategies to adapt themselves in diverse environmental conditions. Salmonella is one such bacteria with diverse mechanisms to survive, replicate and infect in wide host range. This study aims at investigating the biofilm-forming ability of multidrug-resistant and sensitive Salmonella serovars on exposure to bile. Antibiogram of all the isolates was determined by disk diffusion method and their biofilm-forming ability in the presence or absence of bile was assessed by microtiter plate assay. Biofilm results were validated by calcofluor, Congo red plate and test tube method. Few isolates were selected for further study of their expression of biofilm related genes on exposure to bile using real time PCR. Among the 59 isolates of Salmonella isolated from seafood and poultry, 30 isolates were multi-drug resistant (MDR). Under control conditions, 57% (n = 25) of the serovars were able to form biofilm. While, 86% (n = 51) of the serovars produced biofilm in the presence of bile. The relative gene expression study of the selected serovars for 8 different genes showed a striking difference in the expression levels, supporting the hypothesis that the presence of bile triggers biofilm formation in food associated strains of non-typhoidal Salmonella by upregulation of genes involved in biofilm production.

Occurrence of bacterial biofilm in leprosy plantar ulcers

Background: Plantar ulceration is the most common and serious disability in people affected by leprosy. There is a lack of evidence on biofilm formation among clinical isolates of bacteria in plantar ulcers in leprosy. Objectives: The study was undertaken to screen for bacterial biofilm production in plantar ulcers in leprosy and to examine the correlation between antibiotic resistance in biofilm-producers and non-biofilm-producers. Methods: Leprosy patients with plantar ulcers who were registered for care in LEPRA–Blue Peter Public Health and Research Center, were enrolled. Isolation and identification of bacteria were performed, based on culture and standard bacterial identification methods. Antimicrobial susceptibility testing was conducted using the disk diffusion test. Detection of biofilm production was carried out using the Congo Red method (CRA) and the Microtitre plate method (MTP). The pattern of antibiotic resistance exhibited by biofilm-producers was compared with non-biofilm-producers. Results: Out of 64 swabs tested for bacterial growth, 95.3% (61/64) were culture positive; of these, 90.1% (55/61) were mono-bacterial cultures. A total of 67 bacterial isolates were identified, 73% being gram-positive organisms and 27% being gram-negative. The most commonly isolated organism was S. aureus in 36/67 (53.7%), followed by P. aeruginosa in 8/67 (11.9%). A positive correlation between biofilm production and antibiotic resistance was observed. Conclusion: Biofilm producing bacterial isolates were widely distributed in plantar ulcers in leprosy patients; this suggests the need for further investigation into the correlation between biofilm on wounds with clinical outcomes of plantar ulceration.

A Comparative Proteomic Study of Thermobifida Cellulosilytica TB100T Secretome Grown on Carboxymethylcellulose and Rice Straw

Background: Cellulose, the major component of the plant cell wall, is the most abundant and cheap polymer on earth. It can be used by varieties of cellulolytic enzymes. Cellulases can hydrolyze cellulose to its glucose monomers, which can be fermented to many biotechnological products, such as biochemicals, bioplastics, and biofuels. Actinomycetes are potential sources of cellulases. Objective: The current study sheds light on the cellulolytic activity of Thermobifida cellulosilytica, a previously isolated thermophilic actinomycete, and the analysis of the lignocellulases produced in the secretome as a result of induction by different carbon sources. Methods: The cellulolytic activity was qualitatively confirmed by Congo red method showing a large halo zone around the colonies. The activity was also assayed using the 3,5-dinitrosalicylic acid (DNS) method. The secretome analysis was conducted by liquid chromatography-tandem mass spectroscopy (LC-MS/MS) based proteomic approach. Results: The cellulolytic activity increased by two folds upon the growth of T. cellulosilytica on rice straw (RS) as a complex substrate comparatively to Carboxymethylcellulose (CMC) as a simple one. These results were highly assured by LC-MS/MS. Where more proteins (n=31) were produced in the RS secretome, CMC produced only six proteins, including only one cellulase. Different classes of proteins produced in the RS secretome were cellulases (26%), hemicellulases (16%), proteases (10%), and others (48%). Conclusion: Lignocellulases are inducible enzymes. RS as a complex substrate induced T. cellulosilytica for the expression of more lignocellulolytic enzymes than CMC.

English

Escherichia coli has carved its niche in the urinary tract with the formation of a formidable matrix called biofilm. This biofilm is not only recalcitrant to the body’s immune system but also resistant to antibacterial agents. Senna siamea (Lam) Irwin and Barneby is a medicinal plant with established antibacterial effect against planktonic cells of many bacteria. An attempt was made herein to evaluate the effect of its leaf extract and fractions on biofilm of E. coli isolates. Crude extracts of leaf, stem bark and root of this plant were prepared using ethanol as the solvent for the cold extraction. Phytochemical screening was carried out on the three extracts. Two E. coli strains from different antenatal patients attending General Hospital, Kafanchan, Kaduna were donated to us by a researcher from Ahmadu Bello University, Zaria and the reference strain, E. coli, WDCM 00013 (from Germany) were tested for biofilm production using the Congo red method. Antimicrobial susceptibility testing of the crude extracts against the isolates was carried out using the agar diffusion method. The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) were determined for the leaf extract of the plant using micro broth dilution and agar diffusion methods respectively. In order to establish the antibiofilm activities of the leaf extract of the plant, sub-inhibitory concentrations (sub-MIC) were used against the test isolates in the remaining assays in the work. Column chromatography backed by thin layer chromatography (TLC) was used to fractionate leaf extract (having the best antibacterial activity) of the plant, using different ratios of a combination of hexane, ethyl acetate and n-butanol as fractionating solvents. MIC and MBC of the leaf extract were and 50 mg/ml respectively. High values of percentage biofilm inhibition were observed against all the bacterial isolates from the antibiofilm assay. Combination of solvents in the increasing order of polarity enhanced the antibiofilm activity of the various fractions of the leaf extract of Senna siamea. In conclusion, further fractionation of Senna siamea leaf extract increases its antibiofilm activities. Key words: Biofilm, Senna siamea, column chromatography, thin layer chromatography, Escherichia coli.

Isolation of Thermostable Lignocellulosic Bacteria From Chicken Manure Compost and a M42 Family Endocellulase Cloning From Geobacillus thermodenitrificans Y7.

The composting ecosystem provides a potential resource for finding new microorganisms with the capability for cellulose degradation. In the present study, Congo red method was used for the isolating of thermostable lignocellulose-degrading bacteria from chicken manure compost. A thermophilic strain named as Geobacillus thermodenitrificans Y7 with acid-resident property was successfully isolated and employed to degrade raw switchgrass at 60°C for 5 days, which resulted in the final degradation rates of cellulose, xylan, and acid-insoluble lignin as 18.64, 12.96, and 17.21%, respectively. In addition, GC-MS analysis about aromatic degradation affirm the degradation of lignin by G. thermodenitrificans Y7. Moreover, an endocellulase gene belong to M42 family was successfully cloned from G. thermodenitrificans Y7 and expressed in Escherichia coli BL21. Recombinant enzyme Cel-9 was purified by Ni-NTA column based the His-tag, and the molecular weight determined as 40.4 kDa by SDA–PAGE. The characterization of the enzyme Cel-9 indicated that the maximum enzyme activity was realized at 50°C and pH 8.6 and, Mn2+ could greatly improve the CMCase enzyme activity of Cel-9 at 10 mM, which was followed by Fe2+ and Co2+. Besides, it also found that the β-1,3-1,4, β-1,3, β-1,4, and β-1,6 glucan linkages all could be hydrolyzed by enzyme Cel-9. Finally, during the application of enzyme Cel-9 to switchgrass, the saccharification rates achieved to 1.81 ± 0.04% and 2.65 ± 0.03% for 50 and 100% crude enzyme, respectively. All these results indicated that both the strain G. thermodenitrificans Y7 and the recombinant endocellulase Cel-9 have the potential to be applied to the biomass industry.

Comparison of characterization, antioxidant and immunological activities of three polysaccharides from Sagittaria sagittifolia L.

To investigate and compare the preliminary structural characteristics and biological activity in vitro of polysaccharides from Sagittaria sagittifolia L. (SSs) by different extration methods, three polysaccharides (SSW, SSU, and SSP) were obtained with hot water, ultrasound-assisted, and subcritical water extraction. Their structural features were elucidated using High Performance Liquid Chromatography (HPLC), Gas Chromatography (GC), Scanning Electron Microscopy (SEM), Infrared Spectroscopy (IR), Atomic Force Microscopy (AFM), Zeta Potential and Congo red methods. Furthermore, the antioxidant activity and immunostimulatory effects were investigated in vitro. Molecular weight and monosaccharide composition analysis exhibited that SSW (2275.0 kDa), SSU (148.7 kDa), and SSP (1984.0 kDa) were heteropolysaccharide with dramatically different monosaccharide species and mole ratios. In addition, SSP exhibited stronger antioxidant activity in vitro and more potent immunomodulatory activity than SSW and SSU. SSP has greater potential to be explored as biologicalagents for use in complementary medicine or functional foods.

Trace Metals, Potassium Bromate and Nutritional Potentials in Bread from Bakeries in Uyo, Akwa Ibom State, Nigeria

This research assessed bread from bakeries in Uyo, Akwa Ibom State, Nigeria, for trace metals, nutritive value and potassium bromate using standard procedures. Samples were collected from four bakeries coded as: A, B, C and D. Alkaloids, glycosides, steroids and saponins were present in all samples. Flavonoids were only present in samples obtained from bakery D. Quantitatively, phytochemical composition ranged from 0.08±0.00 mg/100g glycoside in samples from bakery A to 3.76±0.00 mg/100g alkaloids also in samples from bakery A. Trace metal analysis indicated that copper was only detected in samples from bakery B, and that iron levels were less than 0.009 mg/100g in all the samples, while tin was < 0.001 mg/100g in samples from bakery A and was not detected in samples from bakeries B, C and D. The levels of potassium bromate analysed by the congo red oxidation method in all the samples ranged from 6.66±0.00 to 52.19±0.00 mg/kg. With crystal violet oxidation method, they ranged from 5.09±0.00 to 58.36±0.01 mg/kg. Proximate composition ranged from 0.03±0.00 fiber in samples from A, C and D to 73.38±0.00% CHO in samples from C, while the calorific values ranged from 280.11±0.00 to 359±0.00 kcal in all the samples. Anti-nutrient composition ranged from 0.21±0.00 mg/100g tannin in samples from B to 1.68±0.00 mg/100g HCN in samples from C. Essential mineral composition ranged from 0.01±0.00 mg/100g phosphorus and sodium in samples from A and D to 1.89±0.00 mg/100g calcium in samples from B. Vitamins composition ranged from 0.19±0.00 mg/100g vitamin B2 in samples from D to 4.62±0.00 mg/100g vitamin C in samples from C. The bread samples were to some extend nutritive. The anti-nutrients and trace metals in the bread were below the levels that could cause toxicity in humans, except that the bread contained potassium bromate (banned chemicals that are hazardous to human health).

Novel Fractional Purification Approach of Crude Polysaccharides via Boronic Acid-Tagged Thermoresponsive Triblock Copolymers

A novel fractional purification approach of crude polysaccharides with a boronic acid-tagged thermoresponsive triblock copolymer PEG113-b-PVBA49-b-PNIPAM105 was designed in this paper. The fractional precipitation of polysaccharides was realized through simple temperature regulation based on the difference in the lower critical solution temperatures of the triblock copolymer–dextran complex. The precipitated polysaccharides from the extracting solution was in a descending order of molecular weight. Under optimal conditions, four astragalus polysaccharide fractions (APS1, APS2, APS3, and APS4) with narrow distribution of molecular weights were obtained with the temperature-controlled fractional precipitation. The structural properties, monosaccharide compositions, and molecular weights of APS1, APS2, APS3, and APS4 were characterized by the Fourier transform infrared spectrum, Congo red spectrophotometric method, gas chromatography, and gel permeation chromatography. The differences in the biological activ...

Prevention of poly(vinyl chloride) degradation through organic terephthalamides generated from poly(ethylene terephthalate) waste

ABSTRACTThe use of organic compounds as thermal stabilizers for poly(vinyl chloride) (PVC) stabilization is the current state of art worldwide owing to their high efficiency and nontoxic residues after degradation. Terephthalamide, N,N′‐dimethylterephthalamide and N,N′‐dibutylterephthalamide have been prepared via depolymerization of poly(ethylene terephthalate) through an economical and environmentally friendly approach. These compounds have been examined as thermal stabilizers for PVC formulations and found to exhibit high thermal stabilizing effects. Thermal stability measurements were performed using conventional Congo red test method. Color change experiments were conducted by heating the samples at 200 °C in air, and the colored compounds formed were extracted and compared with the help of UV–visible spectroscopy. Fourier transform infrared spectroscopic studies of organic terephthalamides incorporated PVC samples have been performed in order to insight the probable mechanistic aspects involved in thermal stabilization process. Thermogravimetric analysis (TGA) thermograms of PVC sheets loaded with 10 wt % organic terephthalamides have been recorded and were found stable below 200 °C. SEM and energy dispersive X‐ray analysis of char residues of TGA samples were performed supporting the proposed thermal stabilizing action of the organic terephthalamides. Furthermore, specific gravity and mechanical performance of the PVC sheets have also been reported assisting in finding suitable commercial applications of PVC formulations. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019, 136, 48022.

Complete Genome Sequence of Paenibacillus sp. CAA11: A Promising Microbial Host for Lignocellulosic Biorefinery with Consolidated Processing.

Several bioprocessing technologies, such as separate hydrolysis and fermentation (SHF), simultaneous saccharification and fermentation (SSF), and consolidated bioprocessing (CBP), have been highlighted to produce bio-based fuels and chemicals from lignocellulosic biomass. Successful CBP, an efficient and economical lignocellulosic biorefinery process compared with other processes, requires microorganisms with sufficient cellulolytic activity and biofuel/chemical-producing ability. Here, we report the complete genome of Paenibacillus sp. CAA11, a newly isolated promising microbial host for CBP-producing ethanol and organic acids from cellulose. The genome of Paenibacillus sp. CAA11 comprises one 4,888,410bp chromosome with a G + C content of 48.68% containing 4418 protein-coding genes, 102 tRNA genes, and 39 rRNA genes. The functionally active cellulase, encoded by CAA_GH5 was identified to belong to glycosyl hydrolase family 5 (GH5) and consisted of a catalytic domain and a cellulose-binding domain 3 (CBM3). When cellulolytic activity of CAA_GH5 was assayed through Congo red method by measuring the size of halo zone, the recombinant Bacillus subtilis RIK1285 expressing CAA_GH5 showed a comparable cellulolytic activity to B. subtilis RIK1285 expressing Cel5, a previously verified powerful bacterial cellulase. This study demonstrates the potential of Paenibacillus sp. CAA11 as a CBP-enabling microbe for cost-effective biofuels/chemicals production from lignocellulosic biomass.

&lt;p&gt;Comparative study of antimicrobial resistance and biofilm formation among Gram-positive uropathogens isolated from community-acquired urinary tract infections and catheter-associated urinary tract infections&lt;/p&gt;

Background: Gram-positive cocci have emerged to be an important cause of urinary tract infection (UTI) both in community-acquired UTI (Com-UTI) and catheter-associated urinary tract infection (CA-UTI). The objective of this study was to investigate the frequency of Gram-positive cocci urinary tract infections, their susceptibility patterns to commonly used antimicrobial agents and the biofilm forming property with respect to catheter-associated UTI and community-acquired UTI.Methods: A total of 1,360 urine samples from indwelling catheter and 10,423 from mid-stream urine were obtained during a 6-month period and processed following standard microbiological guidelines. Biofilm formation was detected using congo red agar (CRA), tube method (TM) and tissue culture plate (TCP) method. Chi-square test and independent sample t-test were employed to calculate the significance. Statistical significance was set at P-value ≤0.05.Results: The infection rate was significantly higher in CA-UTI as compared to Com-UTI (25% vs 18%, p=0.0001). Among 2,216 organisms isolated, 471 were Gram-positive cocci; 401 were obtained from Com-UTI while 70 were from CA-UTI. Enterococcus faecalis was the most common organism isolated from Com-UTI, while Staphylococcus aureus was commonest among CA-UTI. Multi-drug resistance, methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococci were also significantly higher in CA-UTI as compared to Com-UTI. Biofilm-forming property was significantly higher in CA-UTI than Com-UTI. The sensitivity of congo red agar method and tube method was 79% and 81.9% respectively and specificity was 98.5% each. Antimicrobial resistance was significantly higher in biofilm-formers as compared to non-formers.Conclusion: Gram-positive bacteria are a significant cause of both CA-UTI and Com-UTI with Enterococcus faecalis and Staphylococcus aureus as common pathogen. Biofilm formation and multi-drug resistance is significantly higher in CA-UTI than Com-UTI. Routine surveillance of antimicrobial resistance and biofilm formation is necessary in all cases of UTI to ensure the proper management of patients.

Endo-xylanase enzyme from marine actinomycetes and its potential for xylooligosaccharide production

Marine microorganism was emerging as the great source for the discovery of novel enzyme. The search and discovery of new enzymes derived from actinomycetes have received much attention nowadays due to a growing need for applications in various industries including food industry. Endo-β-1,4-xylanases is considered one of critical enzyme involves in the degradation of xylan that is able to cleave the β-1,4-glycosidic linkages in the xylan backbone resulting in various xylooligosaccharides and xylose. Screening and characterization of novel actinomycetes that is capable of producing high xylanase are necessary to produce xylooligosaccharides from hemicellulose hydrolysis process. The objective of this research is screening and characterizing of novel marine actinomycetes from working culture of Research Center for Biotechnology, Biotechnology Culture Collection (BTCC) that are capable of producing high-level hemicellulase-degrading enzymes, especially endo-xylanase enzyme. Approximately, 70 strains from Indonesia marine Actinomycetes have been screened by using congo red, SDS-PAGE and zymogram methods. Of these 70 strains, 3 strains were successfully identified that is capable in producing high level of xylanase enzymes based on the diameter of clear zone more than 1.5 cm on xylan plate medium pH 5.0 and 6.0. Three strains from marine Actinomycetes were identified based on analysis a 16S rRNA gene sequence revealed that all of strains belonging to the genus Streptomyces. Strain 47 is closely related with Streptomyces variabilis (98%). All strains had ability to produce xylanase at optimum pH 5.0 and temperature range between 50-60 °C with range activity from 2.5 – 4.3 U/mL. Each isolate had a molecular weight from 20-50 kDa based on SDS-PAGE and zymogram analysis. Based on TLC analysis using beechwood xylan, each isolate had ability to produce xylooligosaccharides. These characterizations showed that these isolates are potentially used for xylooligosaccharides productions. Strain 47 was selected for the next analysis for molecular cloning based on the pattern and results of TLC clearer then strain 41 and strain 42.

Multi Drug Resistant and Biofilm Formation of Staphylococci Isolated from Patient with Diabetic Foot Infection in Basrah Province

Diabetic foot ulcers (DFUs) is characterized by a classical triad of neuropathy, ischemia and infection. Patient with Diabetic foot ulcers frequently required lower extremity amputations. Eighty one swab samples were collected from patient with Diabetic foot ulcers, from different hospitals in Basra governorate (Al-Fayhaa General Hospital). All samples were subjected to classical biochemical tests. Gram positive-vitek 2 card system was used for the definitive diagnosis of Gram-positive bacterial samples, 78 samples Gram-positive from 81 samples were diagnosed, and these isolates were found to be of the following species: Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus xylosus, Staphylococcus haemolyticus, Staphylococcus caprae, Staphylococcus lugdunensis, Staphylococcus warneri, Staphylococcus sciuri, Staphylococcus pseudintermedius, Staphylococcus lentus, Enterococcus faecalis, Streptococcus thoraltensis, Kytococcus sedentarius. Many bacterial species showed multiple-drug resistance, such as Staphylococcus aureus which were more resistant to the following antibiotics: -Oxacillin, Trimethoprim, Tetracycline. The results showed that Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus xylosus, Staphylococcus haemolyticus, Staphylococcus caprae, Staphylococcus lugdunensis, Staphylococcus warneri, Staphylococcus pseudintermedius, Staphylococcus sciuri, Staphylococcus lentus, Enterococcus faecalis, and Kytococcus sedentarius had the ability to produce biofilm which revealed by various methods, such as Congo red agar qualitative method, and tissue culture plate quantitative method.

Diagnostic histochemistry in medical diseases of the kidney.

The value of histochemical analysis in the diagnosis of medical renal diseases has long been known, and its use continues currently. Depending on the particular disorder in question, a variety of "special" stains may be applied to renal biopsies. These include the periodic acid-Schiff, Masson trichrome, Jones, von Kossa, Verhoeff-van Gieson, Congo Red, and toluidine blue methods, among others. This review considers the application of such techniques in the assessment of vascular, glomerular, and tubulointerstitial lesions of the kidney.

Investigate of the Ability of Cronobacter sakazakii Isolated from Clinical Samples of Children Under Two Years to Induce Swimming, Swarming and Biofilm

The study included 200 samples were collected from children under two years included (50 samples from each of Cerebrospinal fluid, Blood, Stool and Urine) from, Central Children Hospital and Children's Protections Educational Hospital. Isolates bacterial were obtained cultural, microscopic and biochemical examination and diagnosed to the species by using vitek2 system. The results showed there were contamination in 6.5% of clinical samples. The diagnosed colonies which gave pink color on the MacConkey agar , golden yellow color on the Trypton Soy agar and green color on the Birillent Enterobacter sakazakii agar and gave a probability of 99% in the vitek 2 and were identified as Cronobacter sakazakii. The identification revealed of thirteen isolates: 6(46.16%) isolated from Cerebrospinal fluid samples, 7(53.84%) isolated from blood samples and not isolated bacteria from stool and urine samples. The results of the investigation of some virulence factors showed that all bacteria isolates were able to swimming with a diameter ranging (1-9 mm) and swarming with a diameter ranging (1-40 mm) and their ability to biofilm formation by using three methods. The results show the ability of isolates to form biofilm by using Congo red media methods where it is 12 (92.30 %) out of 13 isolated bacteria belonging to C. sakazakii able to form biofilm on the Congo red media which is 3 (23.07%) were strong production biofilm , 8 (61.53%) were intermediate production biofilm and 1 (7.69% ) were weak biofilm formation , while the 1 (7.69%) unable to form biofilm. Tubes method were all isolates were able to form biofilm, it were found that 3 (23.07%) isolates strong, and 8 (61.53%) intermediate and 2( 15.38%) weak biofilm formation. Microtiter plate method gave 5 (38.46 %) isolates strong, 6 (46.15%) intermediate and 1 (7.69%) weak biofilm formation.

High Level of Biofilm Formation and Virulence Factors in Enterococci Species Isolated From Clinical and Normal Flora Samples

Abstract Enterococci are gram-positive bacteria, and they are members of normal intestinal microflora of humans as well as animals. However, recently, they evolved from common avirulent commensal bacteria into a multidrug-resistant health care–associated pathogen. In this study, 147 isolates were collected from clinical and healthy people. The presence of 7 genes including ace, ebp, efaA, ebrB, ecbA, fsrB, and sgrA were investigated by polymerase chain reaction method. Finally, for detection of biofilm formation, Congo red agar and microliter plate methods were used. Our results showed that ace (94.6%), efaA (94.6%), and ebp (92.5%) were present almost in all of the isolates, followed by fsrB (53.7%), sgrA (36%), ecbA (33.3%), and ebrB (13.6%). By Congo red method, 38% of our isolates formed black colonies on Congo red agar (biofilm producer), whereas others (62%) had red colonies (nonbiofilm producer). Finally, by using microliter plate method, 23.8% of isolates were able to form biofilm. These results indicate that normal flora isolates can constitute as a reservoir for virulent Enterococcus isolates.