Selective synthetic growth medium “Acinetobacter phenylalanine agar” for isolation and identification of Acinetobacter calcoaceticus — Acinetobacter baumannii complex species

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Abstract

The aim of the study was to carry out clinical and microbiological testing of selective growth medium Acinetobacter phenylalanine agar to isolate and identify bacterial species belonging to the Acinetobacter calcoaceticus — Acinetobacter baumannii complex (ACB complex). For this, 400 samples of clinical material (wound discharge, blood, urine, bronchoalveolar lavage) were examined in 2018 in the Clinical and Bacteriological Laboratory of the Military Medical Academy by using routine assays (seeding on blood agar growth medium, pure culture isolation and identification by using biochemical assays as well as VITEK 2 microbial identification system, bioMerieux) and plating together with growth medium Acinetobacter phenylalanine agar selective to Acinetobacter spp. owing to L-phenylalanine as a sole nitrogen and carbon source additional selected with trimethoprim. ACB complex Acinetobacters were identified 18–24 hours later after incubation at 37°C by emergence of typical colonies on selective medium. Control tests for cytochrome oxidase as well as oxidative/fermentation (OF)-glucose test by using peroxide-hydrogen microvolume method (for 1 h) allowed to rapidly distinguish ACB complex Acinetobacters from other bacteria as well as from Acinetobacter spp. unable to glucose oxidation. Next, species identity for all isolated Acinetobacter strains was established by using matrix-activated laser desorption/ionization with time-of-flight mass spectrometry (MALDI-TOF MS). It was found that by using novel vs. routine assay Acinetobacter spps. were isolated in 26 (18 samples — in monoculture, 8 — in association with Pseudomonas aeruginosa or Klebsiella pneumoniae subsp. pneumoniae with tiny associate colonies) vs. 20 (7 — in monoculture, 13 — in association with Pseudomonas aeruginosa, Klebsiella, Escherichia, Citrobacter, Providencia, Staphylococcus, Enterococcus, C. albicans). Using MALDI-TOF MS method revealed that 25 out of 26 Acinetobacter strains isolated on selective growth medium belonged to the ACB complex (A. baumannii — 23, A. pittii — 2), whereas one strain (A. baylyi) did not belong to ACB complex. Hence, the diagnostic specificity of the Acinetobacter phenylalanine agar synthetic growth medium for isolation and identification of the A. calcoaceticus — A. baumannii complex species comprised 96.2%, with diagnostic sensitivity exceeding that one for routine assay by 25%. Use of selective growth medium accelerates research by allowing to isolate and identify ACB complex Acinetobacter spp. 18–24 h later after plating clinical material. Selectivity of growth medium was potentially stable, as its major trophic selection factor did not depend on acquired bacterial antibiotic resistance, which is also suitable as a synthetic growth medium for standardized studies.

About the authors

E. P. Sivolodskii

S.M. Kirov Military Medical Academy;
St. Petersburg Pasteur Institute

Email: es279@yandex.ru

PhD, MD (Medicine), Professor of the Department of Microbiology, S.M. Kirov Military Medical Academy; Senior Researcher, St. Petersburg Pasteur Institute

St. Petersburg

Russian Federation

G. V. Gorelova

S.M. Kirov Military Medical Academy

Email: elenagorelova@mail.ru

Bacteriologist, Microbiological Laboratory, Central Clinical Diagnostic Laboratory

St. Petersburg

Russian Federation

S. P. Bogoslovskaya

S.M. Kirov Military Medical Academy

Email: telola@mail.ru

Bacteriologist, Microbiological Laboratory, Central Clinical Diagnostic Laboratory

St. Petersburg

Russian Federation

E. V. Zueva

St. Petersburg Pasteur Institute

Author for correspondence.
Email: elenazueva9@gmail.com

Zueva Elena V., PhD (Medicine), Senior Researcher, Laboratory of Molecular Immunology

197101, St. Petersburg, Mira str., 14

Russian Federation

References

  1. Патент № 2660567 Российская Федерация, МПК C12Q 1/04 (2006.01); C12N 1/20 (2006.01); C12R 1/02 (2006.01). Способ выделения и идентификации бактерий комплекса Acinetobacter calcoaceticus — Acinetobacter baumannii: № 2017130632; заявлено 29.08.2017: опубликовано 06.07.2018 / Сиволодский Е.П. Патентообладатель: Сиволодский Е.П. 8 с.
  2. Сиволодский Е.П. Пероксидводородный микрообъемный метод ОФ-теста: экспрессное определение окисления и ферментации глюкозы грамотрицательными бактериями // Журнал микробиологии, эпидемиологии и иммунобиологии. 1991. № 1. С. 14–17.
  3. Сиволодский Е.П., Зуева Е.В. Таксономическое и прикладное значение профилей утилизации белковых аминокислот бактерий Acinetobacter baumannii, Acinetobacter pittii, Acinetobacter nosocomialis // Вестник Российской военно-медицинской академии. 2018. № 4 (64). С. 113–116.
  4. Cosgaya C., Mari-Almirall M., Van Assche A., Fernandez-Orth D., Mosqueda N., Telli M., Huys G., Higgins P., Seifert H., Lievens B., Roca I., Vila J. Acinetobacter dijkshoorniae sp. nov., a member of the Acinetobacter calcoaceticus — Acinetobacter baumannii complex mainly recovered from clinical samples in different countries. Int. J. Syst. Evol. Microbiol., 2016, vol. 66, pp. 4105–4111. doi: 10.1099/ijsem.0.001318
  5. Gerner-Smidt P., Tjernberg I., Ursing J. Reliablity of phenotypic tests for identification of Acinetobacter species. J. Clin. Microbiol., 1991, vol. 29, no. 2, pp. 277–282.
  6. Jawad A., Hawkey P.M., Heritage J., Shnelling A.M. Description of Leeds Acinetobacter medium, a new selective and differential medium for clinically important Acinetobacter spp. and comparison with Herella agar and Holton’s agar. J. Clin. Microbiol., 1994, vol. 32, no. 10, pp. 2353–2358.
  7. Nemec A., Krizova L., Maixnerova M., Sedo O., Brisse S., Higgins P.G. Acinetobacter seifertii sp. nov., a member of the Acinetobacter calcoaceticus — Acinetobacter baumannii complex isolated from human clinical specimens. Int. J. Syst. Evol. Microbiol., 2015, vol. 65, pp. 934–942. doi: 10.1099/ijs.0.000043
  8. Nemec A., Krizova L., Maixnerova M., Van der Reijden T.J., Deschaght P., Passet V., Veneechoutte M., Brisse S., Dijkshoorn L. Genotypic and phenotypic characterization of the Acinetobacter calcoaceticus — Acinetobacter baumannii complex with the proposal of Acinetobacter pittii sp. nov. (formeily Acinetobacter genomic species 3) and Acinetobacter nosocomialis sp. nov. (formerly Acinetobacter genomic species 13 TU). Res. Microbiol., 2011, vol. 162, pp. 393–404.

Copyright (c) 2020 Sivolodskii E.P., Gorelova G.V., Bogoslovskaya S.P., Zueva E.V.

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