Among the nonfermenters,
There are varieties of antibacterial resistance mechanisms which are involved in the resistance development in
A cross-sectional study was carried out from February to July 2015 at Chitwan Medical College, Bharatpur, Chitwan, Nepal. This 600-bed teaching hospital is a tertiary care center in the city of Bharatpur, Chitwan district of Nepal.
A total of 178 nonrepetitive and nonredundant clinical isolates of
Antimicrobial susceptibility testing was performed by the modified Kirby-Bauer disk diffusion method on standard Muller-Hinton agar medium using commercially prepared antibiotic disks according to the Clinical and Laboratory Standard Institute (CLSI) guidelines and interpretation of antibiotic susceptibility result was made according to the zone-interpretative chart provided by CLSI [
Screening of ESBLs production was carried out according to the NCCLS guidelines [
Screening of MBLs production was carried out by using imipenem, meropenem, or third-generation cephalosporin (ceftazidime) disks. The isolates that showed reduced susceptibility to imipenem, meropenem, or ceftazidime in Kirby-Bauer disk diffusion method were presumptively taken as MBLs producers and were confirmed by disk potentiation method. Briefly, inocula of test strains were prepared compared with 0.5 McFarland standards and inoculated on standard Muller-Hinton agar media to form a lawn culture. Two imipenem disks were placed on the plate at standard distance and 10
All strains with reduced susceptibility to cefoxitin in modified Kirby-Bauer disk diffusion method were confirmed for AmpC beta-lactamase production using the method described by Singhal et al. [
The isolates showing the positive result for at least two beta-lactamases methods described above (ESBLs, MBLs, and AmpC) were characterized as multitype enzyme producing isolates.
The bacterial strains used in this study were isolated from the routine clinical specimens and verbal consent was obtained from the patients. This study was approved by the Institutional Review Committee of Chitwan Medical College (IRC-CMC), Bharatpur, Chitwan, Nepal.
During the study period, 178
Gender- and department-wise distribution of positive specimens.
Specimens type (number of isolates) | Male | Female | IPD | OPD |
---|---|---|---|---|
Pus (wound infection) (32) | 16 | 16 | 26 | 6 |
Sputum (68) | 42 | 26 | 64 | 4 |
Urine (42) | 34 | 8 | 28 | 14 |
Blood (20) | 6 | 14 | 14 | 6 |
Endotracheal tube sample (8) | 6 | 2 | 8 | 0 |
Swab (other than wound infection) (4) | 0 | 4 | 4 | 0 |
Bronchoalveolar lavage (2) | 0 | 2 | 2 | 0 |
Pleural fluid (2) | 0 | 2 | 2 | 0 |
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In this study, we collected a variety of specimens such as pus (wound infections), sputum, urine, blood, endotracheal tube sample, swabs (other than wound infections), bronchoalveolar lavage, and pleural fluid. The majority of the isolates were from sputum (68) followed by urine (42) and pus (32), while a minor number of isolates were contributed by the bronchoalveolar lavage (2) and pleural fluid (2) samples (Table
Among tested antibiotics, the beta-lactams were found to be less effective in
Antibacterial resistance rates of
Antibiotics | Pus (32) |
Sputum (68) |
Urine (42) |
Blood (20) |
ET-tube (8) |
Swab (4) |
BAL (2) |
Pleural fluid (2) |
Total (178) |
---|---|---|---|---|---|---|---|---|---|
CAZ | 24 (75) | 56 (82.3) | 30 (71.4) | 9 (45.0) | 6 (75.0) | 3 (75.0) | 2 (100) | 0 | 130 (73.0) |
CTR | 12 (37.5) | 30 (44.1) | 26 (61.9) | 12 (60.0) | 6 (75.0) | 4 (100) | 2 (100) | 0 | 92 (51.7) |
CTX | 24 (75) | 26 (38.2) | 34 (80.9) | 8 (40.0) | 6 (75.0) | 4 (100) | 0 | 0 | 102 (57.3) |
CFM | 16 (50) | 59 (86.8) | 32 (76.2) | 16 (80.0) | 8 (100) | 3 (75.0) | 0 | 2 (100) | 134 (75.3) |
IPM | 12 (37.5) | 28 (41.2) | 18 (42.8) | 4 (20.0) | 8 (100) | 1 (25.0) | 0 | 0 | 71 (40.0) |
MRP | 12 (37.5) | 28 (41.2) | 16 (38.1) | 6 (30.0) | 8 (100) | 1 (25.0) | 0 | 0 | 71 (40.0) |
CB | 12 (37.5) | 18 (26.5) | 26 (61.9) | 4 (20.0) | 4 (50.0) | 2 (50.0) | 2 (100) | 0 | 68 (38.2) |
PI | 24 (75) | 55 (80.9) | 32 (76.2) | 13 (65.0) | 8 (100) | 4 (100) | 2 (100) | 2 (100) | 140 (78.6) |
PIT | 16 (50) | 21 (30.9) | 16 (38.1) | 6 (30.0) | 5 (62.5) | 2 (50.0) | 1 (50.0) | 2 (100) | 69 (38.8) |
CIP | 8 (25) | 30 (44.1) | 36 (85.7) | 11 (55.0) | 6 (75.0) | 0 | 0 | 0 | 91 (51.1) |
GEN | 12 (37.5) | 8 (11.8) | 14 (33.3) | 8 (40.0) | 6 (75.0) | 2 (50.0) | 0 | 0 | 50 (28.1) |
AK | 6 (18.75) | 6 (8.8) | 12 (28.6) | 4 (20.0) | 0 | 0 | 0 | 0 | 28 (15.7) |
TOB | 6 (18.75) | 8 (11.8) | 8 (19.0) | 6 (30.0) | 6 (75.0) | 1 (25.0) | 0 | 0 | 35 (19.7) |
PB | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 |
Enzyme mediated drug resistance was frequently observed in this study. ESBLs mediated resistance was observed in 59 (33.1%) isolates. The majority of ESBLs mediated resistant isolates were recovered from bronchoalveolar lavage sample (100%) followed by swab sample (50%) and pus sample (40.6%), while there was no ESBLs mediated resistance seen in isolates recovered from pleural fluid (Table
Distribution of enzyme producing isolates.
Specimen types (total number of isolates) | ESBLs |
MBLs |
AmpC |
---|---|---|---|
Pus (32) | 13 (40.6) | 6 (18.7) | 4 (12.5) |
Sputum (68) | 22 (32.4) | 22 (32.3) | 4 (5.9) |
Urine (42) | 12 (28.6) | 14 (33.3) | 8 (19.0) |
Blood (20) | 6 (30.0) | 6 (30.0) | 2 (10.0) |
Endotracheal tube (8) | 2 (25.0) | 6 (75.0) | 8 (100) |
Swab (4) | 2 (50.0) | 1 (25.0) | 2 (50.0) |
Bronchoalveolar lavage (2) | 2 (100) | 0 | 0 |
Pleural fluid (2) | 0 | 0 | 0 |
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Out of total isolates recovered, MBLs mediated resistance was observed in 55 (30.9%) isolates. The majority of MBLs positive isolates were recovered from endotracheal tube sample (75%) followed by urine sample (33.3%), while there was no MBL mediated resistance seen in isolates from bronchoalveolar lavage and pleural fluid (Table
AmpC mediated antibiotic resistance was reported in 15.7% of isolates, with the highest proportions of isolates being from endotracheal tube sample (100%) followed by swab sample (50%), while there was no AmpC positive isolate from bronchoalveolar lavage and pleural fluid samples (Table
In our study, no enzyme mediated drug resistance was found in isolates from pleural fluid specimens and this may be because of small sample size (Table
In current study, we also identified few isolates harboring multitypes of enzymes mediating antibacterial resistance. ESBLs together with MBLs and ESBLs together with AmpC combinations mediating resistance were identified from 6 (3.4%) isolates each, while the combination of MBLs together with AmpC mediated resistance from 14 (7.9%), whereas ESBLs together with MBLs and AmpC mediated resistance from 2 (1.1%) isolates (Table
Multitype enzymes production.
Specimen types (total number of isolates) | ESBL + MBL |
ESBL + AmpC |
MBL + AmpC |
ESBL + MBL + AmpC |
---|---|---|---|---|
Pus (32) | 0 | 0 | 4 | 0 |
Sputum (68) | 2 | 2 | 2 | 0 |
Urine (42) | 0 | 2 | 6 | 0 |
Blood (20) | 2 | 0 | 0 | 0 |
Endotracheal tube (8) | 2 | 2 | 2 | 0 |
Swab (4) | 0 | 0 | 0 | 2 |
Bronchoalveolar lavage (2) | 0 | 0 | 0 | 0 |
Pleural fluid (2) | 0 | 0 | 0 | 0 |
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The development of resistance to multiple antibiotics in microbes is in increasing trend which is the leading cause of treatment failure and is a serious problem of global magnitude. The organisms like
The immunosuppressed and chronic lung disease patients often develop hospital-acquired pneumonia.
Fluoroquinolones have been used extensively for the treatment of both minor and more serious infections. As a consequence, their role today is increasingly limited due to increasing resistance which poses a great therapeutic challenge in Nepal. We identified resistance rate to ciprofloxacin in as high as 85% of the isolates, higher than previous studies conducted in Nepal by Shankar et al. in 2005 [
In this study, one striking feature was that no isolates showed resistance to cytoplasmic membrane damaging agents (polymyxin), confirming that polymyxin B should be used as the empiric therapy for serious pseudomonal infections. Similarly, Agrawal et al. in 2008 [
In their natural habitats, most of
Carbapenems are mostly used as the last resort for treatment of multidrug resistance bacterial infections. However, MBLs mediated acquired resistance to this life saving antimicrobial has been increasingly reported in many types of Gram-negative bacteria including
The current CLSI guidelines do not describe a method for detection of AmpC beta-lactamase. AmpC disc test was originally introduced to detect plasmid-mediated AmpC beta-lactamases [
The combination of multitypes of beta-lactamase production in
In the present study, we found that almost all isolates were resistant to antibiotics. Most of the isolates were producers of beta-lactamases, which renders isolates resistant to cell wall inhibiting agents. However, this study could not omit certain limitations as we were unable to conduct the molecular characterization for various beta-lactamases production and we also failed in looking for other resistance mechanisms such as efflux pump system, as this also can mediate resistance in
This study reports the high prevalence of drug resistance and comparable rates of ESBLs, MBLs, and AmpC mediated resistance in
Clinical and Laboratory Standard Institute
Extended spectrum beta-lactamases
Metallo-beta-lactamases
AmpC beta-lactamase
Inpatient department
Outpatient department
American Society for Microbiology
National Committee for Clinical Laboratory Standards
Ethylene diamine tetra-acetic acid
Methicillin resistant
Vancomycin resistant
Glycopeptide intermediate
Glycopeptide resistant
Ceftazidime
Ceftriaxone
Cefotaxime
Cefixime
Imipenem
Meropenem
Carbenicillin
Piperacillin
Piperacillin/tazobactam
Ciprofloxacin
Gentamicin
Amikacin
Tobramycin
Polymyxin B.
The authors declare that they have no competing interests concerning the information reported in this paper.
The authors are deeply grateful to the subjects participating in this study. The authors also thank the laboratory staff of the Microbiology Department of Chitwan Medical College Teaching Hospital (CMCTH) for their kind support in the collection of data and performing the necessary laboratory tests during the study.