How to isolate, identify and determine antimicrobial susceptibility of anaerobic bacteria in routine laboratories

Nagy, Elisabeth; Boyanova, Lyudmila; Justesen, Ulrik Stenz

doi:10.1016/j.cmi.2018.02.008

Cited by 99 publications

(110 citation statements)

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“…In vivo studies highlight links between the long-term microbiota compositional changes and host dysbiosis, including the development of allergic, metabolic, immunological and inflammatory diseases 5-8,10,11,19-21 . While uncovering the direct effects of different antibiotics on our gut flora is critical to improve general health, technical difficulties hamper routine testing of antibiotic susceptibility in anaerobes 22,23 . Currently available data on bacterial susceptibility to antibiotics is focused on invasive pathogens and offers little to no resolution in the diversity of the human gut microbiota 24 .…”

Section: Main Textmentioning

confidence: 99%

Dissecting the collateral damage of antibiotics on gut microbes

Maier

Goemans

Pruteanu

et al. 2020

Preprint

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24Antibiotics are used for fighting pathogens, but also target our commensal bacteria as a side 25 effect, disturbing the gut microbiota composition and causing dysbiosis and disease 1-3 . 26Despite this well-known collateral damage, the activity spectrum of the different antibiotic 27 classes on gut bacteria remains poorly characterized. Having monitored the activities of 28 >1,000 marketed drugs on 38 representative species of the healthy human gut microbiome 4 , 29 we here characterize further the 144 antibiotics therein, representing all major classes. We 30 determined >800 Minimal Inhibitory Concentrations (MICs) and extended the antibiotic 31 profiling to 10 additional species to validate these results and link to available data on 32 antibiotic breakpoints for gut microbes. Antibiotic classes exhibited distinct inhibition spectra, 33including generation-dependent effects by quinolones and phylogeny-independence by β-34 lactams. Macrolides and tetracyclines, two prototypic classes of bacteriostatic protein 35 synthesis inhibitors, inhibited almost all commensals tested. We established that both kill 36 different subsets of prevalent commensal bacteria, and cause cell lysis in specific cases. 37This species-specific activity challenges the long-standing divide of antibiotics into 38 bactericidal and bacteriostatic, and provides a possible explanation for the strong impact of 39 macrolides on the gut microbiota composition in animals 5-8 and humans 9-11 . To mitigate the 40 collateral damage of macrolides and tetracyclines on gut commensals, we exploited the fact 41 that drug combinations have species-specific outcomes in bacteria 12 and sought marketed 42 drugs, which could antagonize the activity of these antibiotics in abundant gut commensal 43 species. By screening >1,000 drugs, we identified several such antidotes capable of 44 protecting gut species from these antibiotics without compromising their activity against 45 relevant pathogens. Altogether, this study broadens our understanding of antibiotic action on 46 gut commensals, uncovers a previously unappreciated and broad bactericidal effect of 47 prototypical bacteriostatic antibiotics on gut bacteria, and opens avenues for preventing the48 collateral damage caused by antibiotics on human gut commensals.49 3 MAIN TEXT 50 Medication is emerging as major contributor for changes in the composition of the human gut 51 microbiota 4,13-15 . Such severe and long-lasting changes are associated, and in some cases 52 causatively linked, to dysbiosis and a wide range of diseases 16 . Although several non-53 antibiotic drugs may also have a previously unappreciated impact on the gut microbiome 54 composition 4,16,17 , antibiotics, developed to have broad spectra and thereby target very 55 diverse pathogens, are long known to take a heavy toll on our gut flora, causing a variety of 56 gastrointestinal side-effects 18 , including Clostridioides (former Clostridium) difficile infections. 57Recently more attention has been given to this collateral damage of antibiot...

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Section: Main Textmentioning

confidence: 99%

Dissecting the collateral damage of antibiotics on gut microbes

Maier

Goemans

Pruteanu

et al. 2020

Preprint

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“…The miscellaneous colonies on the three-culture media at the primary isolation were sub cultured and characterized by phenotypic features, cultural and cellular morphology by gram stain depending on the rules of Nagy et al, (2018) for the identification of anaerobic clinical isolates. Molecular amplification, LAMP technique was used to confirm bacterial species.…”

Section: Identification Of the Bacterial Isolatesmentioning

confidence: 99%

“…Rapid identification kits e.g. API systems and VITEK MS-bioMérieux system require high dense inoculum which is difficult with the slow growing periodontal bacteria, also many anaerobes are non-reactive in biochemical tests and there is no available data base for each type of periodontal pathogens (Nagy et al, 2018;Garner et al, 2014). Therefore, most laboratory identification methods shifted from classical biochemical methods to the most potent identifier one, 16SrRNAtargeted molecular technique that overcomes limitations of culture depended-identification methods (wade, 2013).…”

Section: Introductionmentioning

confidence: 99%

Identification of Red Complex Pathogens Group from Chronic Periodontitis Patients in Mosul City

Al-Hamdoni¹,

Al-Rawi²

2020

RJS

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This study is concerned with the isolation of red complex pathogens, identifying them by a new molecular method as the first locally used and described procedure and characterizing these pathogens by their phenotypic features and biofilms. Gingival fluids were sampled from chronic periodontitis and inoculated into three types of culture media, Schaedler Anaerobe Agar, Tannerella forsythia (TF) agar, and Trypton Yeast extracts Gelatin Volatile fatty acids and Serum (TYGVS) agar. The different appearing colonies were purified and identified by Loop-Mediated Isothermal Amplification protocol (LAMP) for the detection of red complex species. Bacterial biofilm was estimated in term of mono-and polytypic biofilm by measuring the absorbance of a crystal violet-stained biofilm formed in a microtiter plate. Different forms of colonies appeared at the primary isolation. LAMP method was of a significant value for perfect rapid identification of the target species of extracted DNA or intact cells within half an hour. The three types of red complex pathogens were simultaneously detected in the same gingival fluid sample. They formed mono-and polymicrobial biofilms in a synergistic manner particularly the two intimates P. gingivalis and T. denticola. In conclusion, the updated LAMP molecular protocol was attractive method for the diagnosis of red complex pathogens which showed great morphological variations. P. gingivalis fortified the growth of the other two to establish polymicrobial biofilms in a synergistic manner.

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“…There are many reports on anaerobic infections worldwide, and it has been demonstrated that antibiotics in the current market have reduced the efficacy against anaerobic bacterial infection (5). Antimicrobial resistance among anaerobes is increasing dramatically due to changes in the drug resistance mechanisms (6). Metronidazole is a broad-spectrum antibiotic for anaerobic infections.…”

Section: Introductionmentioning

confidence: 99%

Identification and Antimicrobial Susceptibility of Clinically Isolated Anaerobic Bacteria: A Retrospectively Study in a Jiangxi Tertiary-Care Hospital

Zou

Xiao-ying

Zeng

et al. 2019

Jundishapur J Microbiol

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Background: Anaerobic infections have been reported for many years, and there is an increasing trend in these infections worldwide, but anaerobic infections have not received sufficient attention. Rapid identification is important for the treatment of anaerobes because of their different antibiotic-resistance profiles. Objectives: This study aimed to analyze the hospital's present condition to improve anaerobic culture detection rates and enhance the monitoring of anaerobes in hospitals. Methods: This study retrospectively analyzed sterile body fluids sent to the First Affiliated Hospital of Nanchang University in the form of culture bottles in 2017. Finally, 28 strains of obligate anaerobes were isolated, then combined 16S rRNA gene sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to identify the strains and performed separate antimicrobial susceptibility testing. Results: The results showed that these two methods are highly consistent. There were 17/28 (61%) Gram-negative and 11/28 (39%) Gram-positive bacteria. The predominant bacteria were Bacteroides fragilis (15/28). Ten strains were isolated from the Obstetrics and Gynecology Department. The next most frequently affected departments were General Surgery (17.86%) and the ICU (17.86%). We analyzed the resistance to penicillin, cefoxitin, clindamycin, metronidazole, meropenem, piperacillin/tazobactam and amoxicillin/clavulanic acid by using the agar dilution method. The resistance rates to clindamycin were relatively high but relatively sensitive to metronidazole. Conclusions: The results of this research indicate that we should pay attention to the cultivation of anaerobic bacteria, especially in certain high-risk departments.

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How to isolate, identify and determine antimicrobial susceptibility of anaerobic bacteria in routine laboratories

Abstract: The present review aims to update and improve anaerobic microbiology in laboratories with optimal conditions as well as encourage its routine implementation in laboratories with restricted resources.

Cited by 99 publications

References 54 publications

Dissecting the collateral damage of antibiotics on gut microbes

Dissecting the collateral damage of antibiotics on gut microbes

Identification of Red Complex Pathogens Group from Chronic Periodontitis Patients in Mosul City

Identification and Antimicrobial Susceptibility of Clinically Isolated Anaerobic Bacteria: A Retrospectively Study in a Jiangxi Tertiary-Care Hospital

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