|Year : 2017 | Volume
| Issue : 2 | Page : 121-125
Antibiotic susceptibility profiles of non-fermenting gram-negative Bacilli at a Tertiary Care Hospital in Patiala, India
Udhayvir Singh Grewal1, Rupinder Bakshi1, Geeta Walia1, Parth Rajeshbhai Shah2
1 Department of Microbiology, Government Medical College, Patiala, Punjab, India
2 Department of Microbiology, Smt. NHL Municipal Medical College, Ahmedabad, Gujarat, India
|Date of Web Publication||24-Jul-2017|
Udhayvir Singh Grewal
Department of Microbiology, Government Medical College, Patiala, Punjab
Source of Support: None, Conflict of Interest: None
Background: Non-fermenting Gram-negative bacilli (NFGNB) have emerged as a major cause of healthcare-associated infections and are innately resistant to many antibiotics. Aim: The aim of this study was to determine the prevalence of NFGNB isolated from various clinical specimens and evaluate their antimicrobial susceptibility profiles. Materials and Methods: This retrospective study was done at our Department of Microbiology from December 2015 to December 2016. NFGNB were isolated from a variety of clinical specimens, plated on blood agar and MacConkey agar and incubated at 37°C for 18–24 h under aerobic conditions. Appropriate biochemical tests were done to identify the organisms isolated. Antibiotic susceptibility test was performed using the modified Kirby–Bauer disc diffusion method using commercially available discs on Mueller–Hinton agar. Data was analyzed using SPSS IBM version 20. Results: Out of 19065 clinical samples, cultures were positive in 1854 samples. Out of 1854 culture-positive samples, 216 (11.6%) yielded NFGNB. Pseudomonas aeruginosa was the most common NFGNB, isolated in 190/216 (87.96%) samples, followed by Acinetobacter baumannii (17/216, 7.87%). Overall, most of the NFGNB isolates were susceptible to polymyxin B (88.4%), imipenem (82.9%) and cefoperazone + sulbactam (50.9%), and a total of 11 (64.71%) multidrug-resistant A. baumannii (MDRAB) strains were isolated in the study. Conclusion: Our study showed a significantly high prevalence of NFGNB. Isolation of multidrug-resistant P. aeruginosa and MDRAB in the present study raises the concern of rapidly emerging antibiotic resistance in this group of bacteria in our region.
Keywords: Antibiotic susceptibility, multidrug resistance, non-fermenting Gram-negative bacilli, nosocomial infections
|How to cite this article:|
Grewal US, Bakshi R, Walia G, Shah PR. Antibiotic susceptibility profiles of non-fermenting gram-negative Bacilli at a Tertiary Care Hospital in Patiala, India. Niger Postgrad Med J 2017;24:121-5
|How to cite this URL:|
Grewal US, Bakshi R, Walia G, Shah PR. Antibiotic susceptibility profiles of non-fermenting gram-negative Bacilli at a Tertiary Care Hospital in Patiala, India. Niger Postgrad Med J [serial online] 2017 [cited 2020 Apr 6];24:121-5. Available from: http://www.npmj.org/text.asp?2017/24/2/121/211464
| Introduction|| |
Non-fermenting Gram-negative bacilli (NFGNB) are a taxonomically diverse group of organisms that either do not use carbohydrates as a source of energy or utilise it oxidatively. NFGNB have emerged as a major cause of healthcare-associated infections, especially in immunocompromised hosts. Injudicious and empirical use of antibiotics has caused the emergence of this group as important healthcare-associated pathogens. They have been commonly found on the skin of healthcare workers, instruments such as ventilator machines, humidifiers and mattresses  and are known to account for nearly 12%–16% of all bacterial isolates from a clinical microbiology laboratory. Most commonly isolated NFGNB are Pseudomonas aeruginosa and Acinetobacter baumannii. Infections usually caused by these bacteria are septicemia, meningitis, pneumonia, urinary tract infections and surgical site infections. NFGNB are innately resistant to many antibiotics and have been documented to produce extended spectrum β-lactamases and metallo-β-lactamases. Multidrug resistance is common and increasing amongst Gram-negative non-fermenters, and a number of strains have now been identified that exhibit resistance to essentially all commonly used antibiotics. We undertook this study to characterise the prevalence of NFGNB isolated from various clinical specimens, evaluate their antimicrobial susceptibility profiles and note the prevalence of multidrug resistance in P. aeruginosa and A. baumannii.
| Materials and Methods|| |
This retrospective study was done at the Department of Microbiology, Government Medical College, Patiala (Punjab), India, from December 2015 to December 2016. Institutional Ethics Committee approval was obtained before initiating the study. The study participants were patients who were referred to our department for culture and antibiotic susceptibility testing and were found to be culture positive for non-fermenters. NFGNB were isolated from a variety of clinical specimens such as pus swabs, blood and urine. These samples were plated on blood agar and MacConkey agar and incubated at 37°C for 18–24 h under aerobic conditions. Appropriate biochemical tests were done to identify the organisms isolated. Organisms showing growth on triple sugar iron agar and producing an alkaline reaction were provisionally considered to be NFGNB, and further identification was done using a standard protocol. Characters assessed include morphology, motility, oxidase, catalase, indole, urease, nitrate, citrate tests and oxidation-fermentation reactions of glucose, lactose, xylose, arginine decarboxylase test and gelatin liquefaction test. Antibiotic susceptibility test was performed with the help of the Kirby–Bauer disc diffusion method using commercially available discs on Mueller–Hinton agar. Interpretation was done according to the Clinical and Laboratory Standards Institute (2013) guidelines.Escherichia coli ATCC 25922 and P. aeruginosa ATCC 27853 were used as control strains. Isolates of P. aeruginosa intermediate or resistant to at least three drugs in the following classes: beta-lactams, carbapenems, aminoglycosides and fluoroquinolones were labelled as multidrug-resistant P. aeruginosa(MDRPA). Moreover, isolates of A. baumannii resistant to at least two specific representatives of at least two classes of antibiotic categories: aminoglycosides, antipseudomonal penicillins, carbapenems, 3rd or 4th generation cephalosporins and fluoroquinolones were labelled as multidrug-resistant A. baumannii (MDRAB). Statistical analysis was done using SPSS 20 (IBM Corp. Released 2011. IBM SPSS Statistics for Windows, Version 20.0. Armonk, NY: IBM Corp.).
| Results|| |
Out of 19,065 clinical samples, cultures were positive in 1854 samples. Out of 1854 culture-positive samples, 216 (11.6%) yielded NFGNB. The mean of our study participants was found to be 42.22 ± 12.46 years, with a male: female ratio 2.7:1. P. aeruginosa was isolated in 190/216 (87.96%) samples, followed by A. baumannii (17/216, 7.87%) and Alcaligenes faecalis (5/216, 2.31%). Burkholderia cepacia, Pseudomonas stutzeri (distinguished from P. aeruginosa mainly by the absence of pigment production and negative gelatin liquefaction test) and Stenotrophomonas maltophilia were rarely isolated, accounting together for 1.85% of the isolates.
Clinical sources of various NFGNB isolates are shown in [Table 1]. Out of 216 clinical samples positive for NFGNB, pus swab accounted for 91 (42.13%) samples, followed by urine culture 53 (24.53%), blood culture 45 (20.83%), sputum culture 15 (6.94%) and tracheal swab 8 (3.7%). One sample each of chest tube, endotracheal tube, common bile duct stent and stool culture yielded NFGNB isolates.
|Table 1: Clinical sources of various non-fermenting Gram-negative bacilli isolates|
Click here to view
Clinico-microbiological correlation of NFGNB isolates in our study is shown in [Figure 1].
|Figure 1: Clinico-microbiological correlation of non-fermenting Gram-negative bacilli isolate|
Click here to view
Overall, most of the NFGNB isolates were susceptible to polymyxin B (88.4%), imipenem (82.9%) and cefoperazone + sulbactam (50.9%). Percentage antibiotic susceptibility of the various isolates is shown in [Table 2].
|Table 2: Percentage antibiotic susceptibilities of non-fermenting Gram-negative bacilli to major antibiotics|
Click here to view
Out of a total of 190 P. aeruginosa isolates, 46 (24.2%) were labelled as multidrug resistant (MDRPA). About 100% of the MDRPA isolates were found to be susceptible to polymyxin B and 18 (39.1%) isolates showed susceptibility to imipenem. A total of 11 (64.71%) MDRAB strains were isolated in the study, out of which 9 (81.81%) and 6 (54.55%) were found to be susceptible to imipenem and cefoperazone + sulbactam, respectively.
| Discussion|| |
NFGNB, which were only considered to be contaminants in the past, have now emerged as important nosocomial pathogens. In our study, isolation rate of NFGNB was 11.6%, which is in parallel to other studies by Rit et al. and Benachinmardi et al. that reported isolation rates of 12.8% and 10%, respectively. The most common NFGNB isolated in our study was P. aeruginosa (87.96%), followed by A. baumannii (7.87%) which is similar to the results obtained by Malini et al. who reported P. aeruginosa as the most common isolate accounting for 104/189 (53.8%) isolates, followed by A. baumannii (43/189, 22.2%). Similarly, the study done by Rit et al. also found P. aeruginosa to be the predominant isolate (101/201, 50.24%), followed by A. baumannii (50/201, 24.8%). Other Gram-negative non-fermenters such as B. cepacia, P. stutzeri and S. maltophilia that were rarely isolated by us (1.85%) vary from study to study both in terms of their genera and prevalence. However, their role as opportunistic pathogens in immunocompromised and debilitated individuals has been invariably established.
In our study, the highest number of isolates was isolated from pus swabs (40%), which is in accordance with the observations made by Rit et al. and Gokale and Metgud  who also reported pus swabs as the source of maximum percentage of the isolates i.e., 27.86% and 58.4%, respectively. As evident from [Figure 1], NFGNB were majorly found to cause urinary tract infections (24.53%) and would infections (23.7%).
P. aeruginosa isolates in our study were found to be most susceptible to polymyxin B (100%), which is not routinely used to treat infections caused by P. aeruginosa and is only tried as a last resort in case of severe multidrug-resistant Gram-negative bacterial infections. Nearly 83.7% of the P. aeruginosa isolates were found to be sensitive to imipenem. Similarly, Malini et al. and Rit et al. documented 94.2% and 91.08% susceptibility to imipenem, respectively. In contrast with the studies done by Benachinmardi et al. and Naqvi et al. that showed higher susceptibility to quinolones, only 45.8%, 34.7% and 28.9% of P. aeruginosa isolates in the present study showed susceptibility to the quinolones such as ciprofloxacin, ofloxacin and norfloxacin, respectively. In our study, P. aeruginosa showed least susceptibility to cefepime (7.3%) and amoxicillin + clavulanic acid (7.3%).
Almost 24.2% of the isolates of P. aeruginosa in our study were labelled as multidrug resistant (MDRPA), comparable to the findings of Jayakumar and Appalaraju who reported 22% isolation rate of MDRPA in their study. About 100% of the MDRPA isolates were found to be susceptible to polymyxin B, which is similar to the results obtained by Ramakrishnan et al. who also reported 100% susceptibility to imipenem. Nearly 60.9% of the MDRPA isolates in our study showed resistance to imipenem, which is usually the preferred therapeutic choice for treating the infections caused by them. As carbapenems are a potent antimicrobial weapon against MDRPA, this bacterium has developed resistance even against this group of drugs by producing metallo-beta-lactamases (carbapenemase). Goossens  and Ramakrishnan et al. showed 44.9% and 40% resistance of MDRPA isolates to imipenem in their studies, respectively [Figure 2]. Imipenem resistance in MDRPA may possibly be influenced by the amount and duration of utilisation of the antibiotic used to treat these infections.
|Figure 2: Comparison of the present study with other studies reporting imipenem resistance in multidrug-resistant Pseudomonas aeruginosa (a) and multidrug-resistant Acinetobacter baumannii (b) isolates|
Click here to view
Isolates of A. baumannii in our study showed maximum susceptibility to imipenem (88.2%), followed by cefoperazone + sulbactam (76.4%). Results obtained by other studies show variable results. Rit et al. documented 90% and 16% susceptibility of A. baumannii isolates to imipenem and cefoperazone + sulbactam, respectively. Tunyapanit et al. have reported 100% susceptibility to imipenem and 47% susceptibility to cefoperazone + sulbactam in A. baumannii isolates. Highest resistance amongst these isolates in our study was recorded against aztreonam (susceptibility = 11.7%). Similarly, Juyal et al. reported least susceptibility of A. baumannii isolates to aztreonam (8.33%) in their study.
A total of 11 (64.71%) of A. baumannii isolates showed multidrug resistance (MDRAB) in the present study which is in accordance with Cai et al. who reported 72.23% prevalence of MDRAB isolates. Fortunately, MDRAB isolates in our study showed good susceptibility to imipenem (85.7%), which is usually the most common therapeutic choice for MDRAB bacteraemia. This is, however, in contrast with the findings of Tunyapanit et al. and Cai et al. [Figure 2] who documented only 12% and 9.27% susceptibility to imipenem, respectively. Nearly 54.55% of the MDRAB isolates in our study were found to be susceptible to cefoperazone + sulbactam, which is comparable to Tunyapanit et al. who reported 47% susceptibility to cefoperazone + sulbactam combination.
A. faecalis was the third most commonly isolated NFGNB (7.87%) in our study. Sidhu et al. reported a prevalence of 2.31%. The isolate of A. faecalis showed maximum susceptibility (60%) to imipenem, amikacin and ciprofloxacin. Sidhu et al. reported 100%, 50% and 75% susceptibility of A. faecalis isolates to imipenem, amikacin and ciprofloxacin, respectively, in their study. There is a lack of substantial data regarding the prevalence and antibiotic susceptibility profile of A. faecalis due to its limited pathogenic role and rare isolation.
The other less commonly isolated NFGNB, namely, B. cepacia, P. stutzeri and S. maltophilia showed high resistance to almost most of the antibiotics tested for susceptibility, as shown in [Table 2].
In our study, B. cepacia was isolated from urine culture and showed maximum (100%) susceptibility to imipenem, in accordance with Sidhu et al. who also reported 100% susceptibility to imipenem. Similarly, in the study done by Rit et al., B. cepacia isolates showed excellent susceptibility to imipenem (92.85%). Therefore, it can be inferred that imipenem offers excellent therapeutic effect in infections caused by B. cepacia, which is known to be resistant to many first-line therapeutics of choice against serious pseudomonal infections, such as beta-lactam drugs, polymyxin B and aminoglycosides.
S. maltophilia, isolated from a pus swab, showed 100% susceptibility to some of the antibiotics, notably ciprofloxacin. Similar were the results obtained by Malini et al. and Chawla et al. who reported 100% and 93.3% susceptibility of S. maltophilia to ciprofloxacin, respectively. S. maltophilia was found to be 100% resistant to majority of the antibiotics in our study, including imipenem, which could be attributed to the production of a zinc-dependent β-lactamase by this bacterium.
P. stutzeri was isolated from a tracheal swab and was found to be resistant to all the antibiotics as shown in the antibiogram [Table 2], except the combination of cefoperazone + sulbactam. However, the antibiograms of P. stutzeri isolates in various other studies by Rit et al. and Benachinmardi et al. show greater susceptibility towards most of the antibiotics. According to Carvalho-Assef et al., multidrug resistance in P. stutzeri could be attributed to IMP-16, a type of metallo-β-lactamase.
| Conclusion|| |
Our study showed a significantly high prevalence of NFGNB, the most common being P. aeruginosa and A. baumannii. P. aeruginosa isolates showed good susceptibility to polymyxin B and imipenem whereas the isolates of A. baumannii showed good susceptibility to imipenem and cefoperazone + sulbactam. Isolation of MDRPA and MDRAB in the present study raises the concern of rapidly emerging antibiotic resistance in this group of bacteria in our region. Proper screening of non-fermenters in nosocomial settings, regular assessment of their antibiotic susceptibility profiles and judicious use of antibiotics are suggested for effective management of the infections caused by them and limiting the emergence of multidrug resistance.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Winn W Jr., Allen S, Janda W, Koneman E, Procop G, Schreckenberger P, et al
., editors. Nonfermenting Gram-negative bacilli. Koneman's Colour Atlas and Textbook of Diagnostic Microbiology. 6th
ed. USA: Lippincott Williams and Wilkins Company; 2006. p. 305-91.
Malini A, Deepa E, Gokul B, Prasad S. Nonfermenting gram-negative bacilli infections in a tertiary care hospital in Kolar, Karnataka. J Lab Physicians 2009;1:62-6.
] [Full text]
McGowan JE Jr. Resistance in nonfermenting gram-negative bacteria: Multidrug resistance to the maximum. Am J Med 2006;119 6 Suppl 1:S29-36.
Mellmann A, Bimet F, Bizet C, Borovskaya AD, Drake RR, Eigner U, et al.
High interlaboratory reproducibility of matrix-assisted laser desorption ionization-time of flight mass spectrometry-based species identification of nonfermenting bacteria. J Clin Microbiol 2009;47:3732-4.
Rit K, Nag F, Raj HJ, Maity PK. Prevalence and susceptibility profiles of nonfermentative gram-negative bacilli infection in a tertiary care hospital of Eastern India. Indian J Clin Pract 2013;24:451-55.
Gales AC, Jones RN, Forward KR, Liñ ares J, Sader HS, Verhoef J. Emerging importance of multidrug-resistant Acinetobacter
species and Stenotrophomonas maltophilia
as pathogens in seriously ill patients: Geographic patterns, epidemiological features, and trends in the SENTRY Antimicrobial Surveillance Program (1997-1999). Clin Infect Dis 2001;32 Suppl 2:S104-13.
Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing. 23rd
Informational Supplement, CLSI Document M100-S23. Wayne PA: Clinical and Laboratory Standards Institute; 2013.
Obritsch MD, Fish DN, MacLaren R, Jung R. Nosocomial infections due to multidrug-resistant Pseudomonas aeruginosa
: Epidemiology and treatment options. Pharmacotherapy 2005;25:1353-64.
Chen CH, Huang CC. Tracing the emergence of multidrug-resistant Acinetobacter baumannii
in a Taiwanese hospital by evaluating the presence of integron gene intI1. J Negat Results Biomed 2014;13:15.
Benachinmardi KK, Padmavathy M, Malini J, Naveneeth BV. Prevalence of non-fermenting gram-negative bacilli and their in vitro
susceptibility pattern at a tertiary care teaching hospital. J Sci Soc 2014;41:162-6. [Full text]
Chawla K, Vishwanath S, Munim FC. Nonfermenting gram-negative bacilli other than Pseudomonas aeruginosa
Spp. Causing respiratory tract infections in a tertiary care center. J Glob Infect Dis 2013;5:144-8.
Gokale S, Metgud SC. Characterization and antibiotic susceptibility pattern of nonfermenting gram-negative bacilli from various clinical samples in a tertiary care hospital. Belgaum J Pharm Biomed Sci 2012;17:1-5.
Evans ME, Feola DJ, Rapp RP. Polymyxin B sulfate and colistin: Old antibiotics for emerging multiresistant gram-negative bacteria. Ann Pharmacother 1999;33:960-7.
Naqvi ZA, Hashmi K, Rizwan QM, Kharal SA. Multi drug resistant Pseudomonas aeruginosa:
A nosocomial infection threat in burn patients. Pak J Pharmacol 2005;22:9-15.
Jayakumar S, Appalaraju B. Prevalence of multi and pan drug resistant Pseudomonas aeruginosa
with respect to ESBL and MBL in a tertiary care hospital. Indian J Pathol Microbiol 2007;50:922-5.
Ramakrishnan K, Rajagopalan S, Nair S, Kenchappa P, Chandrakesan SD. Molecular characterization of metallo β-lactamase producing multidrug resistant Pseudomonas aeruginosa
from various clinical samples. Indian J Pathol Microbiol 2014;57:579-82.
] [Full text]
Andradel SS, Jones RN, Gales AC, Sader HS. Increasing prevalence of antimicrobial resistance among Pseudomonas aeruginosa
isolates in Latin American medical centers: 5 year report of the SENTRY Antimicrobial Surveillance Program (1997-2001). J Antimicrob Chemother 2003;52:140-1.
Goossens H. Susceptibility of multi-drug-resistant Pseudomonas aeruginosa
in Intensive Care Units: Results from the European MYSTIC study group. Clin Microbiol Infect 2003;9:980-3.
Tunyapanit W, Pruekprasert P, Laoprasopwattana K, Chelae S. Antimicrobial susceptibility of Acinetobacter baumannii
isolated from hospital patients. Sci Asia 2014;40:28-34.
Juyal D, Prakash R, Shankarnarayan SA, Sharma M, Negi V, Sharma N. Prevalence of non-fermenting gram-negative bacilli and their in vitro
susceptibility pattern in a tertiary care hospital of Uttarakhand: A study from foothills of Himalayas. Saudi J Health Sci 2013;2:108-12. [Full text]
Cai XF, Sun JM, Bao LS, Li WB. Risk factors and antibiotic resistance of pneumonia caused by multidrug resistant Acinetobacter baumannii
in pediatric Intensive Care Unit. World J Emerg Med 2012;3:202-7.
Kuo LC, Lai CC, Liao CH, Hsu CK, Chang YL, Chang CY, et al.
Multidrug-resistant Acinetobacter baumannii
bacteraemia: Clinical features, antimicrobial therapy and outcome. Clin Microbiol Infect 2007;13:196-8.
Sidhu S, Arora U, Devi P. Prevalence of nonfermentative gram-negative bacilli in seriously ill patients with bacteraemia. JK Sci 2010;12:168-71.
Govan JR, Brown AR, Jones AM. Evolving epidemiology of Pseudomonas aeruginosa
and the Burkholderia cepacia complex
in cystic fibrosis lung infection. Future Microbiol 2007;2:153-64.
Paton R, Miles RS, Amyes SG. Biochemical properties of inducible beta-lactamases produced from Xanthomonas maltophilia
. Antimicrob Agents Chemother 1994;38:2143-9.
Carvalho-Assef AP, Gomes MZ, Silva AR, Werneck L, Rodrigues CA, Souza MJ, et al.
IMP-16 in Pseudomonas putida
and Pseudomonas stutzeri
: Potential reservoirs of multidrug resistance. J Med Microbiol 2010;59(Pt 9):1130-1.
[Figure 1], [Figure 2]
[Table 1], [Table 2]
|This article has been cited by|
||Long-Term Surveillance of Antibiotic Prescriptions and the Prevalence of Antimicrobial Resistance in Non-Fermenting Gram-Negative Bacilli
| ||Chia-Huei Chou,Yi-Ru Lai,Chih-Yu Chi,Mao-Wang Ho,Chao-Ling Chen,Wei-Chih Liao,Cheng-Mao Ho,Yu-An Chen,Chih-Yu Chen,Yu-Tzu Lin,Chia-Der Lin,Chih-Ho Lai |
| ||Microorganisms. 2020; 8(3): 397 |
|[Pubmed] | [DOI]|