For conventional biochemical tests (Kent, 1985 ; Babady & Wengenack, 2012 ) and rapid biochemical profiling using API strips (bioMerieux, France), the inoculum used was a suspension of each isolate in normal saline, made up to McFarland 0.5 turbidity. For the Biolog Phenotype Microarray analysis, a turbidimeter was used to check the turbidity of the suspension and bacterial cells were added to achieve 81% T (transmittance). Microplates PM1, PM2, PM9 and PM10 were used with the OmniLog system (Biolog, USA). These microplates are 96-well microtiter plates containing different kinds of compounds. PM1 and PM2 test for carbon-utilization patterns while PM9 and PM10 test for tolerance to a wide range of osmolytes and pH (http://www.biolog.com/pmMicrobialCells.html ). M. tuberculosis H37Ra was used as the control in all these tests.
Api strips
Manufactured by bioMérieux
Sourced in France
API strips are a type of laboratory equipment used for the identification and biochemical characterization of microorganisms. They provide a standardized and reproducible method for the rapid identification of a wide range of bacterial species.
Automatically generated - may contain errors
Lab products found in correlation
Omnilog system, by Biolog (1 mentions)
Etest, by bioMérieux (1 mentions)
Half fraser broth, by Thermo Fisher Scientific (1 mentions)
Full fraser broth, by Thermo Fisher Scientific (1 mentions)
Vitek 2 compact, by bioMérieux (1 mentions)
Vitek 2 cards, by bioMérieux (1 mentions)
Vitek 2 compact system, by bioMérieux (1 mentions)
9 protocols using api strips
1
Biochemical Profiling of M. tuberculosis
2
Characterization of Hot Spring Bacteria
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One sediment sample (10 g of sediment soil) and two water samples (10 ml each) were collected from different sites of Tatta Pani hot spring of Kotli AJK in sterile poly bags and bottles. The samples were immediately brought into the laboratory. Two water samples of about 100 μl were spreaded on LB agar plates, whereas, sediment sample was spreaded on LB agar by serial dilution method. After 24 h of incubation at 70 °C bacterial strains, presenting clear morphological difference, were purified by further streaking on LB agar media. Cell and colony morphology of purified isolates were examined. For physiological characterization, isolates were grown in LB broth at different pHs, temperature, incubation periods and inocula volumes. By API strips (bioMérieux) biochemical analysis was done by following the procedure of manufacturer.
3
Comprehensive Antibiotic Resistance Profiling
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Nasal swabs or tracheal aspirate, urine, wound swabs and blood samples will be taken from patients who have consented to the study for bacteriological analysis. Samples will be subjected to standard bacteriological analysis to isolate the culprit bacteria. Bacterial species will be confirmed by use of biochemical test and analytical profiles index API strips (bioMérieux France). Antimicrobial susceptibility tests will be performed on isolated bacteria as per the Kirby-Bauer method following manufacturer’s instruction. Results will be interpreted using the Clinical and Laboratory Standards Institute tables.24
Any bacteria isolate found to be resistant to third generation cephalosporins will bE tested for production of extended spectrum beta-lactamase (ESBL) using the synergy disk diffusion test. Vancomycin-resistant Enterococci (VRE) will be identified using disc diffusion tests. Methicillin-resistant Staphylococcus aureus (MRSA) will be detected by testing isolates resistant to cefoxitin using the E test (AB Biodisk, Solna, Sweden) on Mueller-Hinton agar supplemented with 2% NaCl and incubated at 37°C for 24 hours. The identified ESBL, VRE and MRSA samples will be analysed by PCR and sequencing to identify the resistance genotype. In vitro conjugation tests will be performed to determine if resistance in bacteria is transferable.
Any bacteria isolate found to be resistant to third generation cephalosporins will bE tested for production of extended spectrum beta-lactamase (ESBL) using the synergy disk diffusion test. Vancomycin-resistant Enterococci (VRE) will be identified using disc diffusion tests. Methicillin-resistant Staphylococcus aureus (MRSA) will be detected by testing isolates resistant to cefoxitin using the E test (AB Biodisk, Solna, Sweden) on Mueller-Hinton agar supplemented with 2% NaCl and incubated at 37°C for 24 hours. The identified ESBL, VRE and MRSA samples will be analysed by PCR and sequencing to identify the resistance genotype. In vitro conjugation tests will be performed to determine if resistance in bacteria is transferable.
4
Isolation and Identification of Listeria monocytogenes
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L. monocytogenes was isolated and identified, according to ISO 11290-1 [18 ]. Briefly, the samples were inoculated into a selective primary enrichment medium (half Fraser broth, Oxoid) and incubated at 30°C for 24 h, followed by inoculation of 0.1 μL of the incubated broth into a full-strength secondary liquid enrichment medium (full Fraser broth, Oxoid). After incubation at 37°C for 24 h, the broth was streaked on Agar Listeria according to Ottaviani and Agosti (ALOA) medium and Oxford agar (LabM and Oxoid) and incubated at 37°C for 24-48 h. The typical colonies were blue-green surrounded with or without an opaque halo on ALOA agar and olive green with a black halo on Oxford agar. Then, the Listeria strains were identified using API strips (BioMérieux, France).
5
Screening Phage Cocktails for Staphylococcus spp. PJI
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Three commercial preparations of phage cocktails, produced by Microgen, Russia, were screened: Purified polyvalent pyobacteriophage, Sextaphage® polyvalent pyobacteriophage, and “Staphylococcal bacteriophage”. Before treatment, the PJI pathogen was isolated from two or more separate pre-operative punctures or intra-operative biopsies of periprosthetic tissues and identified by a “VITEK 2 Compact” automatic analyzer (Biomerieux, Craponne, France) or API® strips (Biomerieux, Craponne, France). The susceptibility of the isolated Staphylococcus spp. to the above phage preparations was tested using a spot-assay method [40 ]. In each case, only the “Staphylococcal bacteriophage” was found to be active against the isolated Staphylococcus spp. So, next, phage batches were used: H143 and H178 produced by the Perm branch of Microgen (Perm Biomed, Perm, Russia), as well as batches H182 and H184 produced by the Nizhny Novgorod branch of Microgen (IMBIO, Nizhny Novgorod, Russia).
6
Bacterial Identification in Sepsis Patients
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All patients with suspected sepsis had blood culture collected per clinical discretion. Blood cultures collected at community hospitals were transported at 15–30°C for centralized testing at the Nakhon Phanom provincial hospital microbiology laboratory within 24 hours.24 (link) Those specimens were incubated in the BactT/ALERT® 3D automated blood culture system (BioMérieux, Marcy-l’Étoile, France). Bottles which signaled positive growth were subcultured onto sheep blood, chocolate, and MacConkey agar plates and then incubated for further gram staining, biochemical tests, and commercial bacterial identification strips (API strips; BioMérieux) to identify positive bacterial cultures25 including B. pseudomallei. From 2010 to 2013, antibiotic susceptibility testing was performed using minimum inhibitory concentration strips (E-test; BioMérieux) for ceftazidime and trimethoprim/sulfamethoxazole, antibiotics typically used to treat melioidosis patients in this province. Isolates were classified as resistant or susceptible using resistance breakpoints per Clinical and Laboratory Standards Institute recommendations.26
7
Streptococcal Infection Evaluation Protocol
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Biological parameters including blood C-reactive protein and renal and hepatic functions were collected during the episode of infection and during the treatment period. Per-operative specimens were recorded for each patient: number of positive specimens and susceptibility profile to antibiotics. Streptococci yielded from intraoperative sample cultures on standard medium and enriched broth were identified by automated techniques [API® strips (Biomérieux, Marcy l Marcy, France) and VITEK2® cards (Biomérieux, Marcy l Marcy, France). The antimicrobial susceptibility tests were also performed on the VITEK2® automate (Biomérieux, Marcy l, France).
The diffusion agar technique was used in each case, and the procedure and interpretation of the susceptibility tests were performed in accordance with the Comité de l’Antibiogramme de la Société Française de Microbiologie; annual guides from 2001 to 2011) recommendations (http://www.sfm-microbiologie.org ).
The diffusion agar technique was used in each case, and the procedure and interpretation of the susceptibility tests were performed in accordance with the Comité de l’Antibiogramme de la Société Française de Microbiologie; annual guides from 2001 to 2011) recommendations (
8
Isolation and Identification of Bacteria from Retail Sausages
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Seven different types of retail sausages were collected in Kampar, Perak between January and November, 2014. The selected sausages were chopped into small pieces and weighed to approximately 2 grams before they were transferred into sterile Luria-Bertani (LB) enrichment broths (Laboratories CONDA, Spain). These broths were incubated at 37°C for 16 to 18 hours with agitation. Afterwards, a loop full of bacteria from LB enrichment broth was streaked onto MacConkey agar petri dishes (Oxoid, England) and incubated at 37°C for 24 hours. MacConkey agar was used to selectively grow the Gram negative bacteria and differentiate the lactose fermentation. The purified isolates were preserved and suspended in 80% (v/v) glycerol and stored at -80°C. Further identification of the isolates was carried out using API strips (bioMerieux®, France).
9
Bacterial Identification and Antibiotic Susceptibility
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Between January 2008 and May 2010, bacterial species were identified using the BBL Crystal TM Identification Systems (BD Diagnostics, Sparks, USA) and API Ⓡ strips (BioMérieux). The susceptibility of bacteria to antibiotics was performed using ATB TM test strips (BioMérieux). After June 2010, bacterial identification and anti-microbial susceptibility testing were performed using the VITEK 2 Compact System (BioMérieux), a fully automated microbial analysis system. Quality controls were performed according to the manufacturer's instructions and the CISI criteria. The diagnosis of coagulase-negative staphylococcal sepsis required at least two positive bloodcultures, as previously described (12) .
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