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 Table of Contents  
LETTER TO EDITOR
Year : 2019  |  Volume : 3  |  Issue : 2  |  Page : 131-133

Comparison of two methods for direct susceptibility testing of Salmonella typhi and Salmonella paratyphi a from blood cultures in a high-burden laboratory setting


1 Department of Pathology and Laboratory Medicine, Aga Khan University, Karachi, Pakistan
2 Department of Pediatrics and Child Health, Aga Khan University, Karachi, Pakistan
3 Department of Pathology and Laboratory Medicine; Department of Pediatrics and Child Health, Aga Khan University, Karachi, Pakistan

Date of Submission15-Mar-2019
Date of Decision15-Apr-2019
Date of Acceptance25-Apr-2019
Date of Web Publication17-Jun-2019

Correspondence Address:
Dr. Yusra Shafquat
Department of Pathology and Laboratory Medicine, Aga Khan University, P. O. Box: 3500, Karachi 74800
Pakistan
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/bbrj.bbrj_70_19

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How to cite this article:
Shafquat Y, Shaheen G, Qamar F, Shakoor S. Comparison of two methods for direct susceptibility testing of Salmonella typhi and Salmonella paratyphi a from blood cultures in a high-burden laboratory setting. Biomed Biotechnol Res J 2019;3:131-3

How to cite this URL:
Shafquat Y, Shaheen G, Qamar F, Shakoor S. Comparison of two methods for direct susceptibility testing of Salmonella typhi and Salmonella paratyphi a from blood cultures in a high-burden laboratory setting. Biomed Biotechnol Res J [serial online] 2019 [cited 2019 Nov 20];3:131-3. Available from: http://www.bmbtrj.org/text.asp?2019/3/2/131/260488



Blood cultures are the cornerstone of enteric fever diagnostics and are the only tools available for reliable detection and monitoring of antimicrobial resistance (AMR).[1] Despite major advancements in rapid susceptibility testing, many automated methods remain inaccessible in resource-poor settings where AMR emerges and spreads. In view of rising AMR in enteric fever in resource-poor settings, validation of rapid and reliable methods is essential. Disk diffusion is still the most widely used and least expensive method employed in high-burden countries.[2] Although reliability of direct susceptibility testing from blood culture broths is well known, methods are not standardized, and no published studies have focused on typhoidal  Salmonella More Detailse. The American Society for Microbiology (ASM) endorses the application of susceptibility testing procedures to blood culture broth diluted in Tryptic soy broth (TSB).[3] We have evaluated direct susceptibility testing in  Salmonella typhi Scientific Name Search ratyphi A from blood cultures with and without dilution in TSB against the reference 0.5 McFarland susceptibility testing from bacterial colonies. The study was approved by the Ethics Review Committee of the Aga Khan University (approval number 4184-Ped-ERC-16).

Susceptibility testing was performed by three methods on blood cultures received at our laboratory and positive for Gram-negative bacilli on smears [Figure 1]. The first two “direct” methods were performed when blood culture bottles flagged positive in the automated BacT/Alert system (bioMerieux, l'Etoile France). The direct broth (DB) method was performed by inoculating 3 drops of blood culture broth directly onto 100-mm Mueller-Hinton agar (MHA) plates (a drop represents one- or two-twentieth of a milliliter). This was followed by “lawning” with sterile cotton swabs and inoculation of Oxoid Filter paper discs (6 discs per plate). Plates were incubated in 37°C for 18 h, following which zone diameters were read in reflected light, for isolates identified as Salmonella Typhi and Salmonella Paratyphi A by agglutination with standard antisera (Difco, BD Diagnostics, USA) and API 20E biochemical panel (bioMerieux, France). The ASM-recommended TSB method was performed as described previously[3] onto MHA plates followed by procedures similar to the DB method. Reference 0.5 McFarland disk diffusion method was performed from 18-h colonies on MacConkey agar. Zone diameters for all three methods and categorical susceptibility interpretations as per Clinical and Laboratory Standards Institute breakpoints[4] were recorded in MS Excel and analyzed for categorical agreement with the reference method. Quality control was performed with the American Type Culture Collection (ATCC) controls on each day of testing with  Escherichia More Details coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853. [Table 1] shows the results of disk diffusion testing with the direct blood (DB) and ASM TSB methods for 130 isolates of Salmonella Typhi and their categorical agreements. Fifty (n = 50) of 130 blood cultures tested grew ceftriaxone-resistant extensively drug-resistant (XDR) Salmonella Typhi, against which azithromycin and imipenem results were also recorded. Categorical agreement for ampicillin, cefixime, ceftriaxone, chloramphenicol, and co-trimoxazole was 100% for both methods. Categorical agreement for azithromycin and imipenem for 50 XDR Salmonella Typhi was also 100%. Minor errors were observed for ciprofloxacin, however. For both methods, results for 8 isolates were discrepant (6 for each method). Interestingly, such discrepancies were not observed in XDR Salmonella Typhi which is likely to be due to high-level resistance to fluoroquinolones owing to the presence of the additional qnr plasmid-borne gene.[5] The overall categorical agreement was >90% for ciprofloxacin.
Figure 1: Study methods and protocol for processing and direct susceptibility testing from blood cultures as well as reference susceptibility testing of blood culture isolates of Salmonella Typhi and Salmonella Paratyphi A

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Table 1: Categorical agreements and error rates of the direct disk diffusion method and the American Society for Microbiology method from blood cultures for Salmonella Typhi and Salmonella Paratyphi A

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For Salmonella Paratyphi A, among 20 cultures tested, categorical agreements were 100% each for ampicillin, ceftriaxone, ciprofloxacin, co-trimoxazole, and chloramphenicol. For the direct method, one isolate tested intermediate to cefixime but tested sensitive on the TSB and the reference method [Table 1].

Although this is not a multicenter study, the blood cultures processed were not restricted to a single health-care center or setting but include a large population denominator owing to the wide network of diagnostic laboratory services provided by our center in Pakistan.[6]

In conclusion, performance of the DB method for cultures growing typhoidal salmonellaeis acceptable, with no very major or major errors observed and very few minor errors only for ciprofloxacin and cefixime. Since this method shortens reporting turnaround time and avoids unnecessary processing and inoculation in TSB from blood, we recommend that high-volume laboratories in typhoid endemic regions use this method after a small verification study on local Salmonella isolates under recommended biosafety conditions.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Parry CM, Ribeiro I, Walia K, Rupali P, Baker S, Basnyat B. Multidrug resistant enteric fever in South Asia: Unmet medical needs and opportunities. BMJ 2019;364:k5322.  Back to cited text no. 1
    
2.
Ochiai RL, Acosta CJ, Danovaro-Holliday MC, Baiqing D, Bhattacharya SK, Agtini MD, et al. Astudy of typhoid fever in five Asian countries: Disease burden and implications for controls. Bull World Health Organ 2008;86:260-8.  Back to cited text no. 2
    
3.
York MK, Henry M, Gilligan P. General detection and interpretation of blood cultures. In: Garcia LS, editor. Clinical Microbiology Procedures Handbook. American Society for Microbiology Press; 2010. p. 3.4.1.10.  Back to cited text no. 3
    
4.
Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing; CLSI Approved Standard. 28th ed., Vol. M100. Wayne, PA: Clinical and Laboratory Standards Institute; 2018.  Back to cited text no. 4
    
5.
Klemm EJ, Shakoor S, Page AJ, Qamar FN, Judge K, Saeed DK, et al. Emergence of an extensively drug-resistant Salmonella enterica serovar typhi clone harboring a promiscuous plasmid encoding resistance to fluoroquinolones and third-generation cephalosporins. MBio 2018;9. pii: e00105-18.  Back to cited text no. 5
    
6.
The Aga Khan University. Aga Khan University Hospitals Clinical Laboratories. Available from: https://hospitals.aku.edu/pakistan/medical-and-diagnostics/clinical-labs/Pages/default.aspx. [Last updated on 2019 Jan 01].  Back to cited text no. 6
    


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