ANNEXURE-II
PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION1 / nAME OF THE CANDIDATE AND ADDRESS (IN BLOCK LETTERS) / DR. THIPPERUDRASWAMY T.
POST GRADUATE STUDENT,
DEPT OF MICROBIOLOGY,
KIMS, HUBLI - 22
2 / nAME OF THE INSTITUTION / KARNATAKA INSTITUTE OF
MEDICAL SCIENCES (KIMS),
HUBLI-580 022.
3 / cOURSE OF STUDY AND SUBJECT / M. D. MICROBIOLOGY
4 / DATE OF ADMISSION TO COURSE / 30TH JUNE 2010
5 / tITLE OF THE TOPIC / “DETECTION OF AMP C AND METALLO - BETA LACTAMASES PRODUCING NON FERMENTING GRAM NEGATIVE BACILLI FROM CLINICAL SAMPLES.”
6 / BRIEF RESUME OF THE INTENDED WORK:
7 / 6.1 NEED FOR THE STUDY
Nonfermenting gram negative bacilli (NFGNB) are known to account for about 15%of all bacterial isolates from a clinical Microbiology laboratory. In recent years due to liberal and empirical use of antibiotics, NFGNB have emerged as important health care associated pathogens. They have been incriminated in infections such as, septicemia, meningitis, pneumonia, urinary tract infection
and surgical site infections.1
P.aeruginosa, one of the most common pathogens responsible for hospital infection, is intrinsically resistant to many antibiotics. It also shows an increasing pattern of resistance towards betalactam antibiotics, especially by production of class C chromosomal beta lactamases.2
Acquired metallo-betalactamases(MBL) have recently emerged as one of the most worrisome resistance mechanisms owing to their capacity to hydrolyse with the exception of aztreonam, all beta lactams including carbapenems. Also their genes are carried on highly mobile elements, allowing easy dissemination. Such strains are not susceptible to therapeutic serine beta lactamase inhibitors (such as clavulanate and sulfones ).3
MBL producing isolates arealso associated with a higher morbidity and mortality. Moreover, given that MBLs will hydrolyse virtually all class of beta lactams and that we are several years away from the development of a safe therapeutic inhibitor; their continued spread would be a clinical disaster. 3
NFGNB are innately resistant to many antibiotics and are known to produce Extended spectrum beta lactamases (ESBL) and Metallo beta lactamases(MBL).1
The present study is undertaken to identify the various nonfermenters isolated from patients samples, to assess antimicrobial susceptibility pattern, to knowthe prevalence of Amp C and MBL producing NFGNB.
6.2.REVIEW OF LITERATURE :
1. Malini A , Deepa E K , Gokul B N, Prasad S R in their study of “NFGNB infection in tertiary care hospital in Kolar” observed, P.aeruginosaas the most common isolate accounting for 53.8% followed by Acieitobacterboumanni(22.2%) P.flourescence(10.8%),
A. lwoffii(3.1%),Stenotrophomonasmaltophilia (2.6%). P.aeruginosa showed good sensitivity to imipenem(94%), cefaperazone(70%), amikacin(69%) and ticarcillin(63%).
A.bouwmanni showed 100% sensitivity to imipenem and70% sensitivity to piperacillin.1
2. Bhattacharjee A, Anuparba S, Gaur G, Sen M R. have reported that among 162 consecutive non repetitive isolates of P.aeruginosa, 36(22%) were suspected to be Amp C beta lactamase producers which are further confirmed by modified three dimensional test.
Antibiotic suspectibilitytesting showed piperacillin+tazobactam, imipenem
and cefeperazone +sulbactam to be the most effective.2
3. Behera B, Mathur P, Das A, Kapil A, Sharma V. have evaluated 4 different phenotypic methods for detection of MBL producing P.aeruginosa and concluded as MBL screening could be done in 63 isolates, of which 43 were MBL positive by combined disc test and 36 by double disc synergy test. For confirmation of MBL production MBL-E test was done.Imipenem - EDTA combined disc test and imipenem-EDTA MBL E test are equally effective for MBL detection, but given the cost-constraints, combined disk test can be used as a convenient screening method in clinical Microbiology laboratory.3
4ManoharanA ,Chaterjee S, Mathai D. SARI study group have summarised their study as 42.6% of P aeruginosa isolates were MBL producing. Combined disc diffusion test(CDDT) using IMP+EDTA had the highest sensitivity and specificity of 87.8% and 84.4% when compared to Etest, which was higher than the values obtained for (ceftazidime) CAZ+EDTA and (meropenem) MER+EDTA. CDDT using (imipenem) IMP+EDTA also compared very well with PCR.(specificity=90.9% sensitivity =93.3%) Carbapenem resistance among P.aeruginosa is mediated predominantly via MBL production.4
5. Chatterjee S S , Karmacharya R, Madhap S K . Gautham V, Das A, Ray P. studied on “High prevalence of co expression of beta lactamases in gram negative bacilli” and reported that ESBL , Amp C and MBL were determined in 338 GNB isolates, of these 80(23.7%) produced all three of the newer beta-lactamases.5
6. HalukVahaboglu, FigenCoskunkan,OzlemTansel et al. conducted a population based cohort study on clinical importance of ESBL (PER-1-Type) produced in Acinetobacter
species and P.aeruginosa strains and demonstrated the relationship of PER -1 type
ESBL producing Acinetobacter species and P.aeruginosa with poor clinical outcome.
7. Christophe DE Champs, Laurent Poirel, Richard Bonnet et al. carried prospective survey of beta-lactamases produced by Ceftazidime resistant P.aeruginosa isolated in Frenchhospital and reported 6.2% strains of P.aeruginosa were found to be Ceftazidime resistant. After genotyping, ceftazidime resistance was related solely to the overproduction of the cephalosporinases in 30 strains and showed 99% identity with the amp C gene of P aeruginosa PAO 1.
6.3Objectives of Study :
1. To know the prevalence of Extended spectrum beta lactamases producing NFGNB.
- Detection of Amp C beta lactamases producing NFGNB
- Detection of Metallo-beta lactamases producing NFGNB.
- Study of antibiotic resistance pattern of NFGNB,
7.1 Source of Data :
Clinical samples such as pus, urine, blood, body fluids etc. obtained from patients admitted in Karnataka Institute of Medical Sciences Hospital and received at the Department of Microbiology, Karnataka Institute of Medical Sciences, Hubli.
7.2Methods of collection of data
A Design of study: Cross sectional study.
B Size of the study sample:Approximately300NFGNB per year are isolated from different clinical samples at Department of Microbiology, Karnataka Institute of Medical Sciences, Hubli. Since it is a time bound study, 200 NFGNB isolates will be included in the study during the period from Dec 2010 to Nov 2011.
C Inclusion criteria:
Non repetitive, consecutive Nonfermenting gram negative bacilli isolated from
clinical samples obtained from hospitalized patients (IPD) received during study period.
D Exclusion criteria:
- Isolates other than NFGNB
- NFGNB isolated from samples of patients attending OPD.
The clinical samples are processed for bacterial culture as per the standard protocol.8
- NFGNB isolates are identified by conventional methods using relevant biochemical
fermentation test(Hugh Leifson`s medium ) for glucose, lactose, sucrose,
maltose, mannitol and xylose. Growthon 10% lactose agar, Lysine decarboxylase
test, Gelatin liquefaction test etc.
Antibiotic sensitivity test done by disc diffusion method (Kirby-Bauer) as per
CLSI guidelines.
- ESBL production is detected by phenotypic confirmatory Double disc diffusion
- Amp C betalactamase production is detected by using Disc antagonism test and
- Metallo beta lactamase(MBL)production is detected by Combined disc
Analysis of test results are done by using appropriate statistical methods such as
Chi square test.
7.3Does the study require any investigation to be conducted on patients (or) animals specify.
As the study involves the NFGNB isolates from the clinical samples sent for
diagnostic purpose it does not require any invasive procedures.
7.4Has ethical clearances been obtained from Ethical Committee of your institution in case of 7.3 ?
‘Yes’, ethical clearance has been obtained from Ethical Committee of Karnataka Instituteof Medical Sciences, Hubli.
8 / LIST OF REFERENCES :
1.Malini A, Deepa E K, Gokul B N,et al. NFGNB infection in tertiary care hospital in Kolar.J lab physicians 2009; 1(2): 62-66.
2.Battacharjee A, Anuparba S, Gaur G, Sen M R. Prevalence of inducible AMP C beta lactamases producingP. aeruginosa in a tertiary care hospital in North india. IndianJMed Microbiol 2008; 26 (1):89-98.
3.Behera B, Mathur P, Das A,et al. An evaluation of 4 different phenotypic methods for detection of MBL producing P.aeruginosa. Indian J MedMicrobiol 2008; 26 (3):233-237.
4.Manohar A, ChaterjeeS ,Mathai D, SARI study group. Detection and charecterisation of
MBL producing P.aeruginosa. Indian JMedMicrobiol 2010; 28 (3): 241-244.
5. Chaterjee S S, Karmacharya R, Madhup S K, et al. High prevalence of co expression of newerbeta lactamases (ESBL , MBL and AMP C) in gram negative bacilli, Indian JMedMicrobiol 2010; 28(3): 267-268.
6. Hauk Vahaboglu, FigenCoskunkan, OzlemTansel et al. Clinical importance of ESBL
(PER-1-Type) produced in Acinetobacter species and P.aeruginosastrains.
J Med Microbiol2000; 50 (2001): 642-645.
7. Christophe DE Champs, Laurent Poirel, Richard Bonnet et al. Prospective survey of beta-lactamases produced by Ceftazidime resistant P.aeroginosa isolated in French hospital in 2000. InternationalJ Anti-Microbial agentChemother 2002; 46(9):3031-3034.
8 Washington Winn, Jr., Stephen Allen, WilliumJanda, ElmerKoneman, Gary Procop, Paul Schreckenberger, Gail Woods. The nonfermentative gram negative bacilli. Koneman `s Colour atlas and text book ofdiagnostic microbiology. Sixth edition. Philadelphia. Lippincott Williams and Wilkins. 2006; 309-375.
9Black J A, Moland E S, Thamson K S. Amp C disc test for detection of plasmid mediated Amp C – beta lactamasesin Enterobacteriaceae.J clin microbial 2005; 43:3110-3.
9 / SIGNATURE OF THE CANDIDATE:
10 / REMARKS OF THE GUIDE:
11 /
NAME AND DESIGNATION OF
11.1 GUIDE
/DR.SHOBHA D. NADAGIR MD
PROFESSOR & HEAD, DEPARTMENT OF MICORBIOLOGY,KIMS, HUBLI
11.2 SIGNATURE:
11.3 CO-GUIDE:
11.4 SIGNATURE:
11.5 HEAD OF THE DEPARTMENT: /
DR.SHOBHA D. NADAGIR MD
PROFESSOR & HEAD, DEPARTMENT OF MICORBIOLOGY,KIMS, HUBLI
11.6 SIGNATURE:12 / 12.1 REMARKS OF THE CHAIRMAN & PRINCIPAL:
12.2 SIGNATURE:
1