Bacterial resistance towards antibiotics is a clinical threat because it increases the problem of infectious disease. Concern regarding multidrug resistant (MDR) bacteria, especially nosocomial pathogens is attracting more interest because new drugs to overcome resistant bacteria in the drug development pipeline are not readily available [1]. Morbidity and mortality because of gram-negative MDR nosocomial pathogens is high [2, 3]. Because of irrational use of antibiotics pathogens can develop and share resistance to common antimicrobials and the development of new drugs appears distant [4]. This growing resistance has rekindled interest in colistin, one of the oldest antibiotics [5]. Colistin is in the polymyxin group of antibiotics, and was available for clinical use from 1959, although it was not always the first preferred drug for many years [6, 7]. The use of colistin against panresistant nosocomial infections caused especially by
Colistin resistant organisms are reported in various parts of the world, including resistance of
All 94 clinical isolates included in the present study were from two different regions in Tamil Nadu; Chennai and Trichy, and were collected between March and July 2014. All the isolates identified to be multidrug resistant using a disk diffusion method in clinical centers were received in vials from both Chennai (Hitech Diagnostic Centre) and Trichy (Doctor’s Diagnostic Centre). Anonymized isolates (unlinked to patient data) were received at the Antimicrobial Resistant Laboratory, School of Bio Sciences and Technology in VIT University, Vellore, India. All isolates were subcultured onto brain-heart infusion agar (HiMedia Laboratories, Mumbai, India) and stored at –80°C for further analysis.
Minimal inhibitory concentrations (MICs) for colistin and meropenem were determined using a microbroth dilution method. Briefly, Mueller Hinton (MH) broth No. 2 with controlled cations (HiMedia Laboratories) was prepared and 100 μl was aliquoted into each of the 96 wells in a microtiter plate. A total of 100 μl of broth with antibiotic with concentrations ranging from 0.25 μg/ml to 256 μg/ml were made and a single well in each row was allocated as a growth control without antibiotic. Then 100 μl of MH broth with bacteria (to have final count of 5 × 104 ml) prepared from an overnight culture of a single colony inoculated into MH broth No. 2 with controlled cations was added. Microtiter plates were incubated at 37°C for 18 h. An MIC value ≥ 8 μg/mL was considered to be the resistance breakpoint.
DNA was extracted from all the isolates using a boiling method. Briefly, 500 μ
Identification of all 94 isolates included in the present study was determined by 16S rRNA gene sequencing. The results showed that among 94 isolates, 48 were
All the isolates sent to our laboratory were multidrug resistant as determined by disk diffusion. All multidrug resistant isolates tested against colistin with concentrations ranging from 0.12 μg/mL to 128 μg/mL showed MIC50 1 μg/mL and MIC90 >128 μg/mL, respectively. Colistin resistant isolates (
Comparison of the total number of isolates from each bacterial species with colistin resistance, and carbapenem and colistin resistant isolates. Numbers at the top the bars indicate the number of isolates.Figure 1
Among 27 colistin resistant isolates 14 (52%) were resistant also to meropenem. A total of 15% isolates were found to be resistant to both meropenem and colistin. Isolates with dual resistance were
MICs of 27 colistin resistant isolates including 14 isolates resistant to both colistin and meropenem
Strain No. | Organism | MIC (μg/ml) | Strain No. | Organism | MIC (μg/ml) | ||
---|---|---|---|---|---|---|---|
Colistin | Meropenem | Colistin | Meropenem | ||||
1. | >128 | 32 | 19. | >128 | 0.12 | ||
2. | >128 | 0.12 | 20. | 16 | 64 | ||
3. | >128 | <0.06 | 21. | 16 | 0.25 | ||
4. | 32 | >128 | 22. | 8 | 32 | ||
5. | 16 | 16 | 23. | >128 | 0.12 | ||
6. | 16 | 16 | 24. | >128 | 0.12 | ||
7. | 8 | 16 | 25. | 8 | <0.06 | ||
8. | 8 | 16 | 26. | 16 | 32 | ||
9. | 16 | 16 | 27. | >128 | 0.12 | ||
10. | >128 | 8 | |||||
11. | 16 | 16 | |||||
12. | 16 | 8 | |||||
13. | 16 | 0.12 | |||||
14. | 16 | <0.06 | |||||
15. | 8 | 16 | |||||
16. | >128 | 0.25 | |||||
17. | >128 | 0.25 | |||||
18. | >128 | 0.12 |
MIC, minimal inhibitory concentration
Knowledge of antibiotics that bacteria are susceptible to is necessary to overcome the problem of developing bacterial resistance towards common antibiotics. Interest in colistin has reemerged because of its antibacterial activity that finds use against many carbapenem resistant bacteria [8-10, 12]. However, the development of resistance is becoming a problem again. Colistin is used mainly against gram-negative bacteria including
Reports on colistin resistant bacteria from various parts of the world suggest that there is a developing resistance towards colistin among gram-negative bacteria, although the mechanism of resistance is not clear. A study of