Journal of Clinical and Diagnostic Research, ISSN - 0973 - 709X

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On Aug 2018




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Dr. Arundhathi. S
MBBS, MD (Pathology),
Sanjay Gandhi institute of trauma and orthopedics,
Bengaluru.
On Aug 2018




Dr. Mamta Gupta,
"It gives me great pleasure to be associated with JCDR, since last 2-3 years. Since then I have authored, co-authored and reviewed about 25 articles in JCDR. I thank JCDR for giving me an opportunity to improve my own skills as an author and a reviewer.
It 's a multispecialty journal, publishing high quality articles. It gives a platform to the authors to publish their research work which can be available for everyone across the globe to read. The best thing about JCDR is that the full articles of all medical specialties are available as pdf/html for reading free of cost or without institutional subscription, which is not there for other journals. For those who have problem in writing manuscript or do statistical work, JCDR comes for their rescue.
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Dr. Mamta Gupta
Consultant
(Ex HOD Obs &Gynae, Hindu Rao Hospital and associated NDMC Medical College, Delhi)
Aug 2018




Dr. Rajendra Kumar Ghritlaharey

"I wish to thank Dr. Hemant Jain, Editor-in-Chief Journal of Clinical and Diagnostic Research (JCDR), for asking me to write up few words.
Writing is the representation of language in a textual medium i e; into the words and sentences on paper. Quality medical manuscript writing in particular, demands not only a high-quality research, but also requires accurate and concise communication of findings and conclusions, with adherence to particular journal guidelines. In medical field whether working in teaching, private, or in corporate institution, everyone wants to excel in his / her own field and get recognised by making manuscripts publication.


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Salient features of the JCDR: It is a biomedical, multidisciplinary (including all medical and dental specialities), e-journal, with wide scope and extensive author support. At the same time, a free text of manuscript is available in HTML and PDF format. There is fast growing authorship and readership with JCDR as this can be judged by the number of articles published in it i e; in Feb 2007 of its first issue, it contained 5 articles only, and now in its recent volume published in April 2011, it contained 67 manuscripts. This e-journal is fulfilling the commitments and objectives sincerely, (as stated by Editor-in-chief in his preface to first edition) i e; to encourage physicians through the internet, especially from the developing countries who witness a spectrum of disease and acquire a wealth of knowledge to publish their experiences to benefit the medical community in patients care. I also feel that many of us have work of substance, newer ideas, adequate clinical materials but poor in medical writing and hesitation to submit the work and need help. JCDR provides authors help in this regards.
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Every one of us: authors, reviewers, editors, and publisher are responsible for enhancing the stature of the journal. I wish for a great success for JCDR."



Thanking you
With sincere regards
Dr. Rajendra Kumar Ghritlaharey, M.S., M. Ch., FAIS
Associate Professor,
Department of Paediatric Surgery, Gandhi Medical College & Associated
Kamla Nehru & Hamidia Hospitals Bhopal, Madhya Pradesh 462 001 (India)
E-mail: drrajendrak1@rediffmail.com
On May 11,2011




Dr. Shankar P.R.

"On looking back through my Gmail archives after being requested by the journal to write a short editorial about my experiences of publishing with the Journal of Clinical and Diagnostic Research (JCDR), I came across an e-mail from Dr. Hemant Jain, Editor, in March 2007, which introduced the new electronic journal. The main features of the journal which were outlined in the e-mail were extensive author support, cash rewards, the peer review process, and other salient features of the journal.
Over a span of over four years, we (I and my colleagues) have published around 25 articles in the journal. In this editorial, I plan to briefly discuss my experiences of publishing with JCDR and the strengths of the journal and to finally address the areas for improvement.
My experiences of publishing with JCDR: Overall, my experiences of publishing withJCDR have been positive. The best point about the journal is that it responds to queries from the author. This may seem to be simple and not too much to ask for, but unfortunately, many journals in the subcontinent and from many developing countries do not respond or they respond with a long delay to the queries from the authors 1. The reasons could be many, including lack of optimal secretarial and other support. Another problem with many journals is the slowness of the review process. Editorial processing and peer review can take anywhere between a year to two years with some journals. Also, some journals do not keep the contributors informed about the progress of the review process. Due to the long review process, the articles can lose their relevance and topicality. A major benefit with JCDR is the timeliness and promptness of its response. In Dr Jain's e-mail which was sent to me in 2007, before the introduction of the Pre-publishing system, he had stated that he had received my submission and that he would get back to me within seven days and he did!
Most of the manuscripts are published within 3 to 4 months of their submission if they are found to be suitable after the review process. JCDR is published bimonthly and the accepted articles were usually published in the next issue. Recently, due to the increased volume of the submissions, the review process has become slower and it ?? Section can take from 4 to 6 months for the articles to be reviewed. The journal has an extensive author support system and it has recently introduced a paid expedited review process. The journal also mentions the average time for processing the manuscript under different submission systems - regular submission and expedited review.
Strengths of the journal: The journal has an online first facility in which the accepted manuscripts may be published on the website before being included in a regular issue of the journal. This cuts down the time between their acceptance and the publication. The journal is indexed in many databases, though not in PubMed. The editorial board should now take steps to index the journal in PubMed. The journal has a system of notifying readers through e-mail when a new issue is released. Also, the articles are available in both the HTML and the PDF formats. I especially like the new and colorful page format of the journal. Also, the access statistics of the articles are available. The prepublication and the manuscript tracking system are also helpful for the authors.
Areas for improvement: In certain cases, I felt that the peer review process of the manuscripts was not up to international standards and that it should be strengthened. Also, the number of manuscripts in an issue is high and it may be difficult for readers to go through all of them. The journal can consider tightening of the peer review process and increasing the quality standards for the acceptance of the manuscripts. I faced occasional problems with the online manuscript submission (Pre-publishing) system, which have to be addressed.
Overall, the publishing process with JCDR has been smooth, quick and relatively hassle free and I can recommend other authors to consider the journal as an outlet for their work."



Dr. P. Ravi Shankar
KIST Medical College, P.O. Box 14142, Kathmandu, Nepal.
E-mail: ravi.dr.shankar@gmail.com
On April 2011
Anuradha

Dear team JCDR, I would like to thank you for the very professional and polite service provided by everyone at JCDR. While i have been in the field of writing and editing for sometime, this has been my first attempt in publishing a scientific paper.Thank you for hand-holding me through the process.


Dr. Anuradha
E-mail: anuradha2nittur@gmail.com
On Jan 2020

Important Notice

Original article / research
Year : 2009 | Month : August | Volume : 3 | Issue : 4 | Page : 1653 - 1656 Full Version

AmpC Beta Lactamases Among ESBL Producing Escherichia Coli And Klebsiella Pneumoniae- If You Don’t Look, You Won’t Find.


Published: August 1, 2009 | DOI: https://doi.org/10.7860/JCDR/2009/.547
VANDANA K E *, HONNAVAR P**

*(MD)Asso. Prof.,**Final Yr MSc student, Dept. of Microbiology, Kasturba Medical College, Manipal Udupi-576104 Karnataka. (India)

Correspondence Address :
Dr.K.E.Vandana,(M.D.)
Asso. Prof., Dept. of Microbiology,
Kasturba Medical College, Manipal,
Udupi, 576104 Karnataka, (India).
Ph:0820-2922322,9902206009
Email:vandanake@yahoo.co.in

Abstract

Amp C beta lactamases confer resistance to a wide variety of beta lactams and pose both diagnostic and therapeutic challenges, as their presence goes undetected in the presence of ESBLs. We evaluated 52 ESBL positive clinical isolates of E.coli (n=41) and K.pneumoniae (n=11) for AmpC production by a phenyl boronic acid disc (PBA) and the 3 dimensional enzyme test (3DET). The PBA method detected 24 (58.5%) and 9 (82%) E.coli and K.pneumoniae isolates as AmpC positive, while detection by the 3DET increased the numbers to 29 (70.7%) and 10 (91%) isolates, respectively. We conclude that a large number (75%) of ESBL producers are also found to produce AmpC and that the PBA disc method is a very useful and reliable method for routine use in laboratories.

Keywords

Amp C beta lacatamase, ESBL, phenyl boronic acid, 3 dimensional enzyme test.

Introduction
The production of extended spectrum beta lactamases (ESBLs) has been reported in virtually all species of Enterobacteraceae, which greatly complicates the therapy for infections caused by these organisms. However, the frequency of the isolates producing AmpC beta lactamases is largely unknown due to difficulties in the phenotypic detection and confusion about the appropriate reporting convention (1),(2). Amp C beta lactamase is often misidentified as ESBL. Detecting Amp C isolates is clinically important, not only because of their broader cephalosporin resistance, but also because carbapenem resistance can arise in such strains by further mutations, resulting in reduced porin expression (3),(4) Here, we studied the prevalence of Amp C beta lactamases in E.coli and Klebsiella isolates that produce ESBLs.

Material and Methods

A total of 52 clinical isolates of ESBL producing E.coli and Klebsiella from a tertiary care hospital were analyzed for the production of Amp C beta lactamase. ESBL production was tested by the disc approximation method using Co-amoxiclav and cefotaxime (5). Amp C production was detected by the boronic acid disc enhancement method (6) and compared with the three dimensional enzyme test (3DET) as a gold standard (7). In the former method, 2 cefoxitin discs (30µg) were placed on a Mueller Hinton Agar plate lawn inoculated with a 0.5 McFarland turbidity adjusted suspension of the test strain. To one of the discs, 400µg of phenyl boronic acid (Sigma-Aldrich) was added. After overnight incubation at 37° C, the zones of inhibition were measured. Enhancement of zone of inhibition by 5mm around a cefoxitin disc with PBA, in comparison with a disc with cefoxitin alone, was taken as a positive result for Amp C production.

For 3DET, crude enzyme extracts were prepared by freeze thawing centrifuged cell pellets of broth cultures of bacterial isolates. The enzyme extracts were then inoculated into wells on an MHA plate lawn inoculated with E.coli, ATCC 25922 and a cefoxitin disc was placed on it. Linear slits were made from the well directing towards the disc, 3mm away from it. The plates were read after overnight incubation at 37° C. Any enhanced growth of the indicator strain decreasing the radius of the cefoxitin inhibition zone at the end point of the slit, was considered as positive for Amp C (Table/Fig 1)

Antimicrobial susceptibility of test isolates against aminoglycosides, cephalosporins, monobactums, beta lactam and beta lactamase inhibitor combinations, fluoroquinolones and carbapenems, were tested using the Kirbey Bauer disc diffusion method according to CLSI standards (8).

Results

Of the total 52 isolates, E.coli (n=41) and K.pneumoniae(n=11) were obtained from urine (50) and pus ( 2 ). Out of 41 E.coli isoaltes, 24 (58.5%) were found to produce Amp C by the PBA disc method, while the 3DET method detected 5 more Amp C positive isolates (n=29, 70.7%). Of 11 K.pneumoniae isolates, 10 (91%) were found to be positive by the 3DET method that detected one extra isolate as compared to the PBA method (82%) (Table/Fig 2).All the 52 isolates were resistant to cefoxitin. The sensitivity and specificity of the PBA method were found to be 85% and 100% respectively, when compared to the 3DET method. AST results of these strains are given in (Table/Fig 3) . Meropenem was the only drug which was found to be effective against all these isolates, followed by cefoperazone sulbactum, piperacillin tazobactum and amikacin. Three isolates were susceptible to cefipime, all of which were positive for Amp C. All were uniformly resistant to fluoroquinolones. The overall prevalence of Amp C in ESBL producing isolates was 75%.

Discussion

Organisms overexpressing Amp C beta lactamases are a major clinical concern because these are usually resistant to all beta lactam drugs, except for cefepime, cefpirome and carbapenems (9), (10). In contrast to ESBLs, they hydrolyse cephamycins and are not inhibited by beta lactamase inhibitors. Costitutive overexpression of Amp C occurs, either by deregulation through the mutation of the Amp R gene in the chromosome or by acquisition of a transferable AmpC gene on a plasmid or on another transferable element commonly called as plasmid mediated amp C beta lactamase (10),(11). The origin of Amp C in E.coli is chromosomal, although recently, plasmidic Amp C also has been isolated. K.pneumoniae harbours only plasmid mediated Amp C. Detection of any type of Amp C beta lactamase is a challenge to clinical microbiologists. There are no guidelines in place for efficient detection by CLSI guidelines. Phenotypic variations in the bacterial expression of the enzymes make the task of laboratory detection more complicated. Clinical implications of plasmid encoded amp C mediated resistance have to be addressed more cautiously. There are no guidelines for detection of this resistance mechanism and yet, there is as much need for clinical laboratories to address this issue as there is for the detection of ESBLs. The accurate detection of plasmid mediated amp C is important to improve the clinical management of infection and to provide sound epidemiological data (2).

There is a paucity of data from Indian laboratories on the coexistence of multiple beta lactamases in individual isolates. Studies from various parts of India have reported the prevalence of Amp C in clinical isolates of Enterobacteria as varying from 2.2% to 20.7 % (12), (13). However, these studies were designed to estimate the prevalence of AmpC among all the clinical isolates of Enterobacteria. Our study highlights the importance of appropriate detection methods for Amp C enzymes in those isolates which are already designated to be ESBL positives, as the coexistence of different classes of beta lactamases in a single bacterial isolate poses a challenge, both in diagnosis and therapy. Use of a cefoxitin disc is useful in screening for Amp C. However, we observed that 13 cefoxitin resistant isolates did not produce Amp C, which may be attributed to other resistance mechanisms like porin channel alterations in these isolates. Though the isolates showed in vitro susceptibility to cefoperazone sulbactum and piperacillin tazobactum combinations, their clinical usefulness is doubtful as AmpC is not inbibited by beta lactamase inhibitors.

Conclusion

In the present study, we found a large number of ESBL producing E.coli and K.pneumoniae strains, also produced AmpC beta lactamases (75%). Though the 3DET test is the gold standard for Amp C detection, it is labour intensive and cannot be performed routinely on all clinical isolates. The PBA disc method is simple and reliable for detection of this resistance mechanism. Microbiology laboratories should proceed to detect multiple beta lactamases in individual isolates which are already designated as ESBL producers, so that the appropriate therapy can be chosen for patient management, and sound data can be generated on resistance mechanisms and hospital infection epidemiology.

References

1.
. da Silva Dias RC, Borges-Neto AA, D'Almeida Ferraiuoli GI, de-Oliveira MP, Riley LW, Moreira BM. Prevalence of AmpC and other beta-lactamases in enterobacteria at a large urban university hospital in Brazil. Diagn Microbiol Infect Dis. 2008; 60:79-87.
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. Hanson ND AmpC beta-lactamases: what do we need to know for the future? J Antimicrob Chemother, 2003; 52:2–4.
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. Bradford PA, Urban C, Mariano N, Projan SJ, Rahal JJ, Bush K. Imipenem resistance in Klebsiella pneumoniae is associated with the combination of ACT-1, a plasmid-mediated AmpC beta-lactamase, and the loss of an outer membrane protein. Antimicrob. Agents Chemother.1997; 41:563– 69.
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. Martínez-Martínez L, Pascual A, Hernández-Allés S, Alvarez-Díaz D, Suárez AI, Tran J, Benedí VJ et al. Roles of beta-lactamases and porins in activities of carbapenems and cephalosporins against Klebsiella pneumoniae. Antimicrob. Agents Chemother.1999; 43:1669–73.
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. Thomson KS, Sanders CC. Detection of extended-spectrum -lactamases in members of the family Enterobacteriaceae: Comparison of the double-disk and three-dimensional tests. Antimicrob. Agents Chemother.1992; 36:1877-82
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. Crompton IE, Cuthbert BK, Lowe G, Waley SG. Beta –lactamase inhibitors. The inhibition of serine beta -lactamases by specific boronic acids. Biochem.J. 1988; 251:453–59.
7.
. Coudron PE, Moland ES, Thomson KS. Occurrence and detection of AmpC beta-lactamases among Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis isolates at a veterans medical center. J Clin Microbiol 2000; 38: 1791-6.
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. Clinical Laboratory Standards Institute. Performance standards for antimicrobial disc susceptibility tests. 2006 Wayne PA; M100-S15.
9.
. Girlich D, Naas T, Bellais S, Poirel L, Karim A, Nordmann P. Heterogeneity of AmpC cephalosporinases of Hafnia alvei clinical isolates expressing inducible or constitutive ceftazidime resistance phenotypes Antimicrob. Agents Chemother.2000; 44:3220–3223.
10.
. Thomson KS, Smith Moland E.Version 2000:The new betalactamases of Gram-negative bacteria at the dawn of the new millennium. Microbes Infect. 2000; 2:1225–35.
11.
. Petit A, Ben-Yaghlane-Bouslama H, Sofer L, Labia R. Characterization of chromosomally encoded penicillinases in clinical isolates of Klebsiella pneumoniae. J. Antimicrob. Chemother. 1992; 29:629–38.
12.
. Manchanda V, Singh NP. Occurrence and detection of AmpC beta-lactamases among Gram-negative clinical isolates using a modified three-dimensional test at Guru Tegh Bahadur Hospital, Delhi, India. J Antimicrob Chemother 2003; 51 : 415-8.
13.
. Ratna AK, Menon I, Kapur I, Kulkarni R. Occurrence & detection of AmpC beta-lactamases at a referral hospital in Karnataka. Indian J Med Res 2003; 118 : 29-32.

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