Detection Of Biofilm Formation Research Articles

Analysis of biofilm formation by intestinal lactobacilli.

In this study, the biofilm-forming potential of intestinal Lactobacillus reuteri strains under different culture conditions was characterized by microtiter plate biofilm assays. Moreover, the spatial organization of exogenously applied L. reuteri L2/6 (a pig isolate) at specific locations in gastrointestinal tract of monoassociated mice was investigated by fluorescence in situ hybridization. We did not detect biofilm formation by tested strains in nutrient-rich de Man-Rogosa-Sharpe (MRS) medium. On the contrary, a highly positive biofilm formation was observed in medium with lower accessibility to the carbon sources and lack of salts. The results obtained confirmed the significant role of Tween 80 and the quantity and nature of the sugars in the growth medium in biofilm formation. The omission of Tween 80 in MRS medium favored the formation of biofilm. Abundant biofilm formation was detected in the presence of lactose, galactose, and glucose. However, a gradual increase in sugar concentration triggered a significant decrease in biofilm formation. In addition, conditions related to the gastrointestinal environment, such as low pH and the presence of bile and mucins, highly modulated biofilm production. This effect seems to be dependent on the specificity and properties of the medium used for cultivation. From the evidence provided by this study we conclude that the biofilm formation capacity of L. reuteri is strongly dependent on the environmental factors and culture medium used.

FISH DETECTION OF CAMPYLOBACTER AND ARCOBACTER ADHERED TO STAINLESS STEEL COUPONS

This study focuses on detecting biofilm and planktonic bacterial cells of the genera Arcobacter and Campylobacter. This study is, to our knowledge, the first study deals with application of FISH procedure to detect biofilm formation on stainless steel coupons of Arcobacter-isolates from real-world environments. These bacteria can cause a lot of diseases. Especially, in the last decade, arcobacters have been increasingly isolated from feces of clinically healthy and ill animals, foods of animal origin and various types of water. Fluorescence in situ hybridization was used to detect biofilm and planktonic cells of selected microorganisms. This method was optimized and subsequently applied to biofilm samples prepared on stainless steel coupons. The study results indicate that fluorescence in situ hybridization is suitable for detecting biofilm and planktonic cells of the studied bacteria.

Concomitant Detection of Biofilm Formation and MBL Production in Meropenem Resistant Isolates of Pseudomonas aeruginosa

The purpose of this study was to detect biofilm formation and to examine the correlation between biofilm and Metallo-β-lactamases (MBL) production in Pseudomonas aeruginosa. A total of 64 P. aeruginosa isolates were identified using standard microbiological methods and antimicrobial susceptibility testing (AST) was performed on them according to Clinical and Laboratory Standards Institute (CLSI) guidelines. The isolates were screened for biofilm production using both qualitative and quantitative methods. The presence of MBL genes were checked by multiplex PCR assay. Out of all 30 meropenem resistant P. aeruginosa , 2 isolates were found producing all the three genes (i.e. bla IMP , bla VIM , bla SIM ) for MBL production and they were found to produce biofilm. Resistant to four antibiotics such as aztreonam (85.7% vs 11.1%, P< 0.000), Cefepime (82.1% vs 2.8%, P<0.000) gentamycin (82.1% vs 27.8%, P< 0.000) and Pipercillin/Tazobactum was also high

Analysis of biofilm formation and antibiotic susceptibility pattern of uropathogens in patients admitted in a tertiary care hospital in India

Background: Microorganisms attach to surfaces and produce polysaccharides resulting in the formation of biofilms and providing an ideal niche for the exchange of genetic material leading to the emergence of drug-resistant pathogens. Biofilms can develop on anatomical surfaces and implants producing chronic and intractable infections. Aims: Detection of biofilm formation and comparison of antibiotic resistance between biofilm producers and nonproducers. Study Design: Prospective study in which urine specimens from adult patients with urinary tract infection (UTI) during the period of the study were analyzed (1 year). Materials and Methods: Mid-stream clean catch urine from noncatheterized and urine aspirated from in-dwelling urinary catheter in catheterized patients were taken for microbiological processing. Wet mounts, Gram-staining, and urine culture were done. Biofilm formation was detected by tissue culture plate method (TCPM). Statistical Analysis: Chi-square test and mid P test were used to analyze the data. A value ofP<0.05 was taken as significant. Results: Gram-negative organisms predominated (89%). Biofilm production was detected in 27% isolates. Maximum biofilm production was seen in Enterococcus spp. (71%), followed by Escherichia coli (26%). Biofilm-producing isolates demonstrated higher antibiotic resistance. All the biofilm-producing Enterococcus spp. showed high-level aminoglycoside resistance. The biofilm-producing isolates of Pseudomonas aeruginosa and Klebsiella pneumoniae demonstrated multi-drug resistance. Conclusions: TCPM is an economical phenotypic method which can be used routinely to detect biofilm formation. Biofilms contribute to an increased resistance to antibiotics used for the treatment of UTIs. Therefore, detection of biofilms is recommended for all patients presenting with chronic or recurrent disease.

Detection of Biofilm Formation in Uropathogenic Bacteria = الكشف عن تكون البيوفيلم في البكتيريا المسببة للأمراض البولية

Urinary tract infection (UTI) is the most commonly acquired bacterial infection. Biofilm formation being the prime cause for antibiotic resistance by uropathogenes. The purpose of this study was to detect biofilm formation by uropathogenes isolated from UTIs, their antibiotic susceptibility pattern and detection of genes responsible for biofilm production in some isolates. The isolated bacteria were tested for biofilm production by tube adherence (TA), congo red agar (CRA) and micro titer plate (MtP) methods. Antibiotic susceptibility pattern of uropathogens was done by Kirby-Bauer disc diffusion method and detection of genes responsible for biofilm production in some isolates by PCR. Out of 320 cultures positive cases 146 (46.5 %) isolates were biofilm producers Ecoli was the commonest uropathogen followed by Staphylococcus aureus and Enterococcus feacalis. The antibiotic resistance was higher among biofilm producers to commonly used antibiotics as compared to non biofilm producers. The detection of icaD gene of Staph. areus and cup gene ofpseudomonas by PCR was siginfiacntly related to its detection by congo red agar method, while there no significant relation between detection of ESP gene by PCR and the three used conventional methods. It was concluded that the ability of slime production, adherence and biofilm formation of uropathogenic strains had significant role in antibiotic resistance. Biofilm formation is the major virulence determinant of uropathogen, so it is necessary to screen all urinary isolates for biofilm production.

Effects on quinolone resistance due to the biofilm formation activity in Ureaplasma urealyticum.

To detect the effects on quinolone resistance due to biofilm formation activity in Ureaplasma urealyticum. An improved checkerboard dilution susceptibility test was used to analyze the quinolone resistance of clinical Ureaplasma isolates and a postbiofilm formation susceptibility assay was used to compare the effects on the quinolone susceptibility between pre- and postbiofilm formations. Several important functional gene expressions were detected to evaluate their roles in the process of the quinolone resistance mechanism. The quinolone-resistant isolates produced more biofilms than the sensitive isolates. In both the quinolone-resistant and -sensitive groups, the minimal inhibitory concentrations after biofilm formation were higher than those before biofilm formation. The expression of the metabolism-related gene ureC in postbiofilm formation was higher than those in prebiofilm formation. It seems that biofilm formation is quite important in the generation of quinolone resistance in Ureaplasma. It is very useful to detect biofilm formation activity as well as to analyze other laboratory parameters for Ureaplasma.

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Detection of Biofilm Production and Antibiotic Resistance Pattern in Clinical Isolates from Indwelling Medical Devices

Microbial biofilms pose great threat for patients requiring indwelling medical devices (IMDs) as it is difficult to remove them. It is, therefore, crucial to follow an appropriate method for the detection of biofilms. The present study focuses on detection of biofilm formation among the isolates from IMDs. We also aimed to explore the antibiogram of biofilm producers. This prospective analysis included 65 prosthetic samples. After isolation and identification of bacteria following standard methodology, antibiogram of the isolates were produced following Kirby-Bauer disc diffusion method. Detection of biofilms was done by tube adherence (TA), Congo red agar and tissue culture plate (TCP) methods. Out of 67 clinical isolates from IMDs, TCP detected 31 (46.3 %) biofilm producers and 36 (53.7 %) biofilm non-producers. Klebsiella pneumoniae, Pseudomonas aeruginosa and Burkholderia cepacia complex were found to be the most frequent biofilm producers. The TA method correlated well with the TCP method for biofilm detection. Higher antibiotic resistance was observed in biofilm producers than in biofilm non-producers. The most effective antibiotics for biofilm producing Gram-positive isolates were Vancomycin and Tigecycline, and that for biofilm producing Gram-negative isolates were Polymyxin-B, Colistin Sulphate and Tigecycline. Nearly 46 % of the isolates were found to be biofilm producers. The antibiotic susceptibility pattern in the present study showed Amoxicillin to be an ineffective drug for isolates from the IMDs. For the detection of biofilm production, TA method can be an economical and effective alternative to TCP method.

Microbiological profile and biofilm formation on removed intrauterine contraceptive devices from a sample of Egyptian women.

The aim of this study was to investigate the presence of biofilm formation around intrauterine contraceptive devices (IUCD) and to correlate the microbiological profile of the IUCD-associated genital infections to the microbiological profile of specimens retrieved from vaginal discharge. Samples of the vaginal discharge in the posterior fornix were collected from 50 women attending the Family Planning Clinic in Ain Shams University Hospital using two high vaginal swabs. Swabs were immediately sent for Gram staining as well as microbiological culture. The IUCD was then removed. A 0.5-cm piece of the removed IUCD was cut and sent for culture. Growing colonies were tested for their abilities to form a biofilm (colorimetric method). Another 0.5-cm piece of the removed IUCD was examined by electron microscopy (EM) for detection of biofilm formation. Among the included 50 women, 24 (48%) women showed biofilm formation (via colorimetric methods). EM scanning was able to detect biofilm formation in the prepared pieces of the removed IUCD of 48 (96%) women. There was no significant agreement between the isolated microorganisms on the removed IUCD and the vaginal swab (proportion of agreement was 14 [11.4%]; κ = -0.089, P = 0.892). Scanning EM is a useful tool in detection of biofilm formation on removed IUCD.

Characteristics of biofilms formed on non-tunneled hemodialysis catheters

BackgroundMicrobial biofilms are mechanisms used by microorganisms that cause chronic infections in humans. In hemodialysis patients with catheter-related bacteremia, Staphylococcus aureus is an independent risk factor for both infectious complications and failure of bacteremia treatment. We analyzed the characteristics of biofilms formed by these Staphylococcus species on non-tunneled hemodialysis catheters. Patients and methodsA total of 50 adult patients with end-stage renal disease receiving hemodialysis through non-tunneled catheters, whose catheters were removed for catheter-related bacteremia, were studied. ResultsCatheter cultures were positive in only 32 patients and staphylococcal biofilm was found in 25 patients. All biofilm producers were S. aureus. In tissue culture plate method, 2 were strong biofilm producers, 15 were moderate biofilm producers and 5 isolates were considered as weak biofilm producers. In tube method, there were no strong biofilm producers, 12 were moderate biofilm producers and 13 were weak biofilm producers. In Congo red agar method there were no strong biofilm producers, 10 were moderate biofilm producers and 15 isolates were weak biofilm producers. ConclusionOur study shows that S. aureus is the most common bacteria isolated from patients with catheter-related bacteremia. S. aureus is the predominant microorganism responsible for biofilm formation in the non-tunneled HD catheters. Tissue culture plate method is more sensitive to detect biofilm formation by S. aureus.

Exoenzymes Activity and Biofilm Production in Candida Species Isolated from Various Clinical Specimens in Benha University Hospital, Egypt

Background: The frequency of severe systemic fungal diseases has increased in the last few decades. Aims: This study was done to speciate the candida isolates, to determine their antifungal susceptibility pattern and to detect biofilm formation and exoenzymes (phospholipase and proteinase) production. Place and Duration of Study: This is a Six-months Cross sectional study conducted in ICU and Microbiology & Immunology departments, Benha University, Egypt Methodology: The study was conducted on 75 Candida spp. isolated from various clinical samples of patients admitted in ICU. The Candida isolates were identified upto species level. Antifungal susceptibility testing was done by disc diffusion method. The biofilm formation was assessed by inoculating the isolates in conical polystyrene test tube containing Sabouraud’s dextrose broth supplemented with glucose. Proteinase activity was detected by using plates containing bovine serum albumin (BSA) agar. Phospholipase activity was detected by using egg yolk agar. Results: Seventy five Candida spp. were isolated from different clinical samples. C. Original Research Article British Microbiology Research Journal, 4(6): 654-667, 2014 655 albicans was isolated from 39(52%) samples. Non-albicans Candida (NAC) spp. were isolated from 36 (48%) clinical specimens. Forty one (54.7%) out of 75 Candida species isolates obtained from the clinical isolates produced biofilm. Out of 39 C. albicans isolates 20 (51.3%) produced biofilm, while out of 36 NAC species isolates 21 (58.3%) produced biofilm. The number of total proteinase positive isolates were 50 (66.7%). C. albicans was higher than that of the NAC isolates (29 [66.7%] versus 21 [58.3%]). Phospholipase positive isolates of C. albicans was higher than that of the NAC isolates (32[82.1%] versus 37[49.3%]). All isolates were susceptible to amphotericin B and ketoconazole. Resistance to fluconazole was found in 8 isolates (22.2%) of NAC spp. and 2 isolates (5.1%) of C. albicans isolates. Conclusion: The isolation of C. albicans were 39 (52%) in different clinical samples and isolation of NAC spp. were 36(48%). So NAC spp. is no longer overlooked as these organisms are emerging pathogens. The number of NAC producing proteinase, phospholipase and biofilm are more than the number of C. albicans producing these virulence factors. The C. albicans and NAC showed 100% susceptiblity to amphotericin B and ketokonazole while fluconazole showed resistance in 22.2% of NAC spp. and 5.1% of C. albicans isolates. All resistant Candida species to fluconazole were biofilm producers.

Detection of Biofilm Formation in some Pathogenic Bacteria Using Tube and Congo Red Agar Methods

Detection of Biofilm Formation in some Pathogenic Bacteria Using Tube and Congo Red Agar Methods

Association of Biofilm Formation with Antimicrobial Resistance Among the Acinetobacter Species in A Tertiary Care Hospital in Bangladesh

Purpose: The purpose of this study was to detect biofilm formation in clinical isolates of Acinetobacter species and to observe correlation between biofilm formation and antimicrobial resistance among Acinetobacter isolates. Methods: Two hundred fifty six clinical samples collected from patients who were admitted in Intensive Care Unit (ICU) and on device, patients from Surgery, Medicine, Gynae &amp; Obs and Urology department of Bangabandhu Sheikh Mujib Medical University (BSMMU) and from Burn unit of Dhaka Medical College Hospital were included in this study. Biofilm formation and antibiotyping were performed for the isolates of Acinetobacter species recovered from clinical samples including tracheal aspirates, blood, urine, wound swab, pus, throat swab, endotracheal tubes, burn samples, ascitic fluid, sputum, aural swab, oral swab, cerebrospinal fluid, and catheter tip. Correlation of biofilm formation with antimicrobial resistance pattern among Acinetobacter isolates were also observed in this study. Result: A total of 256 various specimens were studied of which 95 Intensive Care Unit (ICU) and 161 Non ICU samples. Out of 95 ICU and 161 Non ICU samples, Acinetobacter species were isolated from 32 (33.7%) and 20(12.4%) respectively. From 32 ICU and 20 Non ICU Acinetobacter isolates, 28 (87.5%) and 11 (55%) were biofilm producers. Biofilm forming capacity of Acinetobacter species was significantly (p&lt;0.008) greater in ICU than in Non ICU isolates. In both ICU and Non ICU isolates, biofilm forming Acinetobacter species were 100% resistant to amoxicillin, ceftriaxone, ceftazidime, cefotaxime, cefuroxime, and aztreonam. Resistance to antibiotics such as gentamicin, amikacin, netilmicin, ciprofloxacin and imipenem was higher among biofilm forming Acinetobacter isolates in ICU than Non ICU isolates. Susceptibility to colistin was 100% in Non ICU isolates but in ICU it showed 7.1% resistance. Conclusions: This investigation showed that most of the clinical isolates of Acinetobacter species were biofilm producers especially from ICU samples and they were multidrug resistant. Even polymixin resistant Acinetobacter isolates are slowly emerging. This is very alerming for us that biofilm forming multidrug resistant Acinetobacter species represents a severe threat in the treatment of hospitalized patients. So, antibiotic policy and guidelines are essential to eliminate major outbreak in future.DOI: http://dx.doi.org/10.3329/jom.v14i1.14533 J MEDICINE 2013; 14 : 28-32

Detection of Micrococcus luteus biofilm formation in microfluidic environments by pH measurement using an ion-sensitive field-effect transistor.

Biofilm formation in microfluidic channels is difficult to detect because sampling volumes are too small for conventional turbidity measurements. To detect biofilm formation, we used an ion-sensitive field-effect transistor (ISFET) measurement system to measure pH changes in small volumes of bacterial suspension. Cells of Micrococcus luteus (M. luteus) were cultured in polystyrene (PS) microtubes and polymethylmethacrylate (PMMA)-based microfluidic channels laminated with polyvinylidene chloride. In microtubes, concentrations of bacteria and pH in the suspension were analyzed by measuring turbidity and using an ISFET sensor, respectively. In microfluidic channels containing 20 μL of bacterial suspension, we measured pH changes using the ISFET sensor and monitored biofilm formation using a microscope. We detected acidification and alkalinization phases of M. luteus from the ISFET sensor signals in both microtubes and microfluidic channels. In the alkalinization phase, after 2 day culture, dense biofilm formation was observed at the bottom of the microfluidic channels. In this study, we used an ISFET sensor to detect biofilm formation in clinical and industrial microfluidic environments by detecting alkalinization of the culture medium.

Biofilm Formation and Antifungal Susceptibility of Candida Isolates from Various Clinical Specimens

Background: Candida species are now recognized as major causative agents of hospitalacquired infection. Aims: To evaluate the species distribution, biofilm formation,and antifungal susceptibility (amphotericin B, ketoconazole, and fluconazole) of Candida isolates. Place and Duration of Study: This is a Six-months Cross sectional study conducted in Alansar hospital, Al-Madinah, Saudi Arabia. Methodology: One hundred and three isolates of Candida spp. were cultured on Sabouraud dextrose agar (SDA). Candida spp. were identified by four standard methods, CHROMagar candida, cornmeal agar, germ tube test and API 20C. Detection of Biofilm formation was done by microtitre plate and antifungal susceptibility testing was done by disc diffusion. Results: C. albicans was the most common species 61%, followed by C. tropicalis 25%, C. lusitanaie 5%, C. parapsilosis 4%, C. glabrata 4% and C. famata 1%. Biofilm formation was found to occur most frequently among non-albicans spp.(70%) than C. albicans (46%). All Research Article British Microbiology Research Journal, 3(4): 590-601, 2013 591 isolates were sensitive to amphotericin B and ketoconazole. Resistance to fluconazole was found in 22.5% of non-albicans spp. and 5% of C. albicans isolates. Conclusion: The present study proved that C. albicans is still the major isolate from urinary, vaginal and respiratory samples but non-albicans spp. predominate in the blood samples and from plastic devices. The non-albicans spp. were more biofilm producers compared to C. albicans and C. tropicalis showed the highest score of biofilm intensity (grade 4+). The species isolated are less susceptible to fluconazole.

Modification of the Congo red agar method to detect biofilm production by Staphylococcus epidermidis

Staphylococcus epidermidis in immunocompromised patients can cause bacteremia related to the use of catheter due to biofilm production. There are different phenotypic methods to detect biofilm formation. One method is based on culture in brain heart infusion agar (BHIA) containing sucrose and red Congo dye (original Congo red agar). Our group created a new CRA formula and we have confirmed its capacity to detect biofilm production in 210 S. epidermidis strains, including 76 (36.2%) icaAB gene–positive strains. Other parameters were also evaluated. The new CRA formula that gave the best results was BHIA with sucrose (5%), Congo red (0.08%), NaCl (1.5%), glucose (2%), and vancomycin (0.5 mg/mL) (vancomycin-modified CRA—CRAmod). The CRAmod plus vancomycin may be a promising tool and can help to determine the real participation of S. epidermidis in the infectious process.

New Paradigms in the Pathogenesis of Otitis Media in Children

Acute otitis media (AOM) is a multifactorial disease with a significant socioeconomic impact. The pathogenesis of AOM is attributed to a variety of well-established internal and extrinsic factors. Recent evidence strongly points to bacterial biofilm formation as an important contributor to this disease entity. The nasopharynx is a likely reservoir for infection with subsequent seeding of pathogens to the middle ear via planktonic shedding. Various modalities have been used to directly detect biofilm formation in the middle ear mucosa of children with AOM. Further insights into this disease may lead to new strategies for prevention and treatment.

In vitro biofilm formation by uropathogenic Escherichia coli and their antimicrobial susceptibility pattern

ObjectiveTo detect in vitro biofilm formation of uropathogenic Escherichia coli (E. coli) (UPEC) strains isolated from urine specimens and also to determine their antimicrobial susceptibility pattern using 13 commonly used antibiotics. MethodsThe present study comprised of 166 urine specimens collected from tertiary care hospitals in and around Coimbatore, South India. All the specimens were subjected to gram staining, bacterial culture and the E. coli strains were screened for biofilm formation using Tube Method (TM), Congo Red Agar (CRA) and Tissue Culture Plate method (TCP) respectively. Subsequently, the antimicrobial susceptibility test was performed by Kirby Bauer-disk diffusion method for the biofilm and non-biofilm producing E. coli strains. ResultsOf the 100 (60.2 %) E. coli strains, 72 strains displayed a biofilm positive phenotype under the optimized conditions in the Tube Method and the strains were classified as highly positive (17, 23.6%), moderate positive (19, 26.3 %) and weakly positive (36, 50.0 %), similarly under the optimized conditions on Congo Red agar medium, biofilm positive phenotype strains were classified as highly positive (23, 23 %), moderate positive (37, 37 %) and weakly positive (40, 40%). While in TCP method, the biofilm positive phenotype strains were also classified as highly positive (6, 6 %), moderate positive (80, 80 %) and weakly positive (14, 14 %), it didn't not correlate well with the tube method for detecting biofilm formation in E. coli. The rates of antibiotic resistance of biofilm producing E. coli were found to be 100 % for chloramphenicol and amoxyclav (amoxicillin and clavulanic acid), 86% for gentamicin and cefotaxime, 84% for ceftazidime, 83% for cotrimoxazole and piperacillin/tazobactam, 75% for tetracycline and 70% for amikacin. ConclusionsThis study reveals the prevalence and antimicrobial susceptibility pattern of biofilm and non-biofilm producing uropathogenic E. coli strains.

Detection of biofilm formation in bacteria from cases of urinary tract infections, septicemia, skin and soft tissue infections and post-operative infections by Congo Red Agar method

Detection of biofilm formation in bacteria from cases of urinary tract infections, septicemia, skin and soft tissue infections and post-operative infections by Congo Red Agar method

Detection of biofilm formation among the mastitis isolates of Staphylococci by evaluation of three different screening methods

Internalization and biofilm formation is an important step in staphylococcal mastitis pathogenesis . Twenty five staphylococcal isolates of coagulase negative and coagulase positive were collected from clinical mastitis in cows, and biofilm production were studied for these isolate by three different methods. Tube method , tissue culture plate and Congo red agar method. The first two methods were used to study effect of glucose addition as source of polysaccharides that important in biofilm formation. The results showed significant increase in biofilm formation by addition of glucose in staphylococci coagulase negative diagnosed by tissue culture plate method. Congo red agar method results showed increase number of positive isolates by increasing incubation time. The results showed that tissue culture plate method (Spectrophotometric method) was the best method for the diagnosis of biofilm formation in comparison with both tube method and Congo red agar method.