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The Microbiology Laboratory
Published in Keith Struthers, Clinical Microbiology, 2017
The first step in the classification of streptococci is their haemolytic nature as exhibited on blood agar. Many bacteria produce haemolysins, which are extra-cellular proteins secreted by the cells that degrade lipid membranes. The membrane of red blood cells is also degraded and the lysis of these cells can be seen on blood agar plates. There are two types of haemolysis, α and β. α-haemolysis produces a green discolouration of the agar as a result of incomplete haemolysis of blood cells; β-haemolysis produces a clear zone of haemolyzed cells around the bacterial colony (Figure 5.19). α-haemolytic streptococci are further subdivided on the basis of the optochin test. Streptococcus pneumoniae is sensitive to the chemical optochin, while the remainder of the α-haemolytic streptococci are resistant to this compound. This test is also shown in Figure 5.19 and is another example of a simple method to classify an organism to species level.
Streptococcus pneumoniae
Published in Peter M. Lydyard, Michael F. Cole, John Holton, William L. Irving, Nino Porakishvili, Pradhib Venkatesan, Katherine N. Ward, Case Studies in Infectious Disease, 2010
Peter M. Lydyard, Michael F. Cole, John Holton, William L. Irving, Nino Porakishvili, Pradhib Venkatesan, Katherine N. Ward
S. pneumoniae is lysed by exposure to bile or optochin (ethylhydrocupreine dihydrochloride) while commensal α-hemolytic streptococci in the nasopharynx are resistant to both. An optochin disc placed on a blood agar plate inoculated with S. pneumoniae will show a wide zone of growth inhibition (Figure 7).
The impact of vaccination on the burden of invasive pneumococcal disease from a nationwide surveillance program in Lebanon: an unexpected increase in mortality driven by non-vaccine serotypes
Published in Expert Review of Vaccines, 2022
Lina Reslan, Nour Youssef, Celina F. Boutros, Aia Assaf-Casals, Danielle Fayad, Sarah Khafaja, Fata Akl, Marc Finianos, Amena A. Rizk, Rouba Shaker, Alissar Zaghlout, Mireille Lteif, Bassam El Hafi, Mohammad Bahij Moumneh, Rita Feghali, Soha Ghanem, Tamima Jisr, Gilbert Karayakoupoglou, Malak Naboulsi, Monzer Hamze, Salam Samad, Elie Khoury, Ricardo Sarraf, Marwan Osman, Elie Bou Raad, Hadi El Amin, Ibrahim Abadi, Hicham Abdo, Marwan Chedid, Fatima Chamseddine, Angelique Barakat, Mohammad Houmani, Antoine Haddad, Georges Abdel Nour, Jacques E. Mokhbat, Ziad Daoud, Mohamad El-Zaatari, Elie Salem Sokhn, Nada Ghosn, Walid Ammar, Randa Hamadeh, Ghassan M. Matar, George F. Araj, Ghassan S. Dbaibo
Bacterial isolates were cultured on Mueller-Hinton blood sheep agar (MHSB) plates and incubated overnight in 5% CO2 at 37°C. S. pneumoniae isolates were identified by colony morphology and optochin susceptibility. Antibiotic susceptibility testing was determined using disk diffusion to oxacillin, erythromycin, clindamycin, trimethoprim-sulfamethoxazole (TMP-SMX), chloramphenicol, tetracycline, vancomycin, levofloxacin, as well as E-test (Biomerieux, France) for both penicillin (PEN) and ceftriaxone (CRO) following the Clinical and Laboratory Standards Institute (CLSI) guidelines [33]. For PEN, non-meningitis breakpoints were considered susceptible (MIC ≤ 2 µg/ml), intermediate (MIC = 4 µg/ml) or resistant (MIC ≥ 8 µg/ml). In meningitis cases, MIC ≤ 0.06 µg/ml were categorized as susceptible and ≥ 0.12 µg/ml as resistant. For CRO, non-meningitis breakpoint was considered susceptible (MIC ≤ 1 µg/ml), intermediate (MIC = 2 µg/ml) or resistant (MIC ≥ 4 µg/ml). For meningitis, MIC ≤ 0.5 µg/ml were categorized as susceptible, whereas MIC ≥ 2 µg/ml was considered resistant. Non-susceptible isolates include all isolates with an intermediate and/or resistant phenotype.
Bacterial biofilm in adenoids of children with chronic otitis media. Part II: a case–control study of nasopharyngeal microbiota, virulence, and resistance of biofilms in adenoids
Published in Acta Oto-Laryngologica, 2020
Hervé Jacquier, Pierre Vironneau, Huong Dang, Benjamin Verillaud, Gerda Lamers, Philippe Herman, Eric Vicaut, Natacha Tessier, Philippe Bidet, Emmanuelle Varon, Thierry Van Den Abbeele, Emmanuelle Cambau, Béatrice Bercot, Romain Kania
Identification of colonies was performed using by Matrix-Assisted Laser Desorption Ionisation Time of Flight Mass Spectrometry (MALDI-TOF-MS). Briefly, replated colonies were harvested in 20 μL of sterile water, and cocrystallized with 1 μL of matrix solution DHB (2,5-dihydroxybenzoic acid, 80 mg/mL; 30% acetonitrile; 0.1% trifluoroacetic acid) in duplicate. Samples were processed in the MALDI-TOF-MS spectrometer (Microflex Bruker Daltonics/BioTyper™ version 2.0). Identification isolates at the species level were fixed at ≥1.9. When a score was lower and all proposals converged on the same genus, identification at the genus level was retained. In the other cases, colonies were subjected to 16S rRNA sequencing. All presumptive Streptococcus pneumoniae results obtained by MALDI-TOF-MS analysis were confirmed by optochin susceptibility testing. When necessary an additional test targeting the 16S rDNA by PCR/sequencing was performed on colonies by using A2/S15 primers for identification at the species level as previously described [9]. The sequences of the corresponding amplicons were submitted to the NCBI (http://www.ncbi.nlm.nih.gov) and BIBI database (http://umr5558-sud-str1.univ-lyon1.fr/lebibi/lebibi.cgi).
Comparative in vitro activity of ceftaroline and comparator agents against nosocomial Gram-negative and Gram-positive clinically significant bacterial isolates from patients in a teaching hospital in Kuwait
Published in Journal of Chemotherapy, 2018
Wafaa Jamal, Hayfaa Abdulkareem, Vincent O. Rotimi
The bacterial isolates were identified by VITEK II (bioMérieux, Marcy, L’Etoile, France) using VITEK 2 GN ID cards, VITEK 2 NH ID cards and VITEK 2 GP ID cards for the identification of Gram-negative bacilli, Haemophilus influenzae and Gram-positive bacteria, respectively, according to the manufacturer’s instructions. Isolates with low scores on VITEK II were subjected to further identification via VITEK MS, a matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry (MALDI-TOF; bioMérieux, Marcy L’Etoile, France). Additional biochemical tests were used for the final identification of certain organisms. These were, Optochin (Sigma-Adrich Company Ltd, Gillingham, Dorset, UK) susceptibility test for S. pneumoniae and Coagulase (Sigma-Adrich Company Ltd) test for the differentiation of Staphylococcus spp.