|Year : 2015 | Volume
| Issue : 2 | Page : 75-80
What is happening to our Pseudomonas? Trends of susceptibilities of culture-confirmed Pseudomonas aeruginosa infections in a tertiary care hospital
Seema Oommen1, PM Sivan Pillai1, Rajeev Arivandakshan2, Kavitha Nair1, Sreeja Nair1
1 Department of Microbiology, Pushpagiri Institute of Medical Sciences and Research Centre, Tiruvalla, Kerala
2 Department of Community Medicine, Pushpagiri Institute of Medical Sciences and Research Centre, Tiruvalla, Kerala
|Date of Web Publication||15-Dec-2015|
Department of Microbiology, Pushpagiri Institute of Medical Sciences and Research Centre, Tiruvalla, Kerala
Source of Support: None, Conflict of Interest: None
Background and Objectives: Infections due to Pseudomonas aeruginosa, especially healthcare-associated infections, present a significant health problem worldwide. Drug resistance especially multidrug resistance in P. aeruginosa is on the rise. Thus the objective of this study was to determine the sensitivity pattern among clinical isolates obtained from the patients admitted to a tertiary care centre. Materials and Methods: A total of 484 non-duplicate P. aeruginosa isolates in 2014, 444 isolates in 2013, 350 isolates in 2012 and 460 isolates in 2007 were characterized according to their antibiotic susceptibility, and the trends over time were analyzed. Results: There were statistically significant increases in sensitivities of isolates P. aeruginosa from 2007 to 2014 to most of the routinely used antibiotics including Ciprofloxacin. Conclusions: A detailed genomic- and/or proteomic-level study for our isolates is much desired to understand this loss of resistance so that the same may be replicated in other bacteria.
Keywords: Antibiotic resistance, Pseudomonas aeruginosa
|How to cite this article:|
Oommen S, Sivan Pillai P M, Arivandakshan R, Nair K, Nair S. What is happening to our Pseudomonas? Trends of susceptibilities of culture-confirmed Pseudomonas aeruginosa infections in a tertiary care hospital. J Acad Clin Microbiol 2015;17:75-80
|How to cite this URL:|
Oommen S, Sivan Pillai P M, Arivandakshan R, Nair K, Nair S. What is happening to our Pseudomonas? Trends of susceptibilities of culture-confirmed Pseudomonas aeruginosa infections in a tertiary care hospital. J Acad Clin Microbiol [serial online] 2015 [cited 2019 Nov 15];17:75-80. Available from: http://www.jacmjournal.org/text.asp?2015/17/2/75/171869
| Introduction|| |
The latter half of the 20 th century witnessed the introduction of antibiotics in the treatment of infections; judging from the number, variety and effectiveness of antibiotics available for therapeutic use, it was widely accepted that the role of infectious disease specialists in clinical management of patients would be on the decline. However, the ability of bacterial pathogens to evolve and to overcome the challenges of clinically available antibiotics in the environment has been nothing short of impressive. The misplaced faith in and overdependence on antibiotics proved to be counterproductive, and the world's medical community is anxious about a post-antibiotic era in medical history. A rising population of pan-resistant bacteria, commonly described by Rice  as the 'ESKAPE' pathogens, including Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter species, are an emerging threat globally. Data from the Centers for Disease Control and Prevention (CDC) show that the ESKAPE pathogens currently cause the majority of infections in USA hospitals and effectively 'escape' the effects of antibacterial drugs; thus the rapidly increasing rates of infection due to methicillin-resistant Staphylococcus aureus (MRSA), Vancomycin-resistant Enterococcus faecium (VRE) and fluoroquinolone-resistant P. aeruginosa.
Of these, P. aeruginosa, belonging to the family Pseudomonadaceae is a ubiquitous bacterial pathogen present in diverse environmental settings, including plants, animals and humans. P. aeruginosa requires only minimal nutritional requirements to survive and tolerate a variety of physico-chemical conditions, having unique properties that allow the organism to persist in both community and hospital settings. In the community, the reservoirs of this organism include swimming pools, whirlpools, hot tubs, contact lens solution, home humidifiers, soil and vegetables.  In the hospital, P. aeruginosa can be isolated from a variety of sources, including respiratory therapy equipment, antiseptics, soap, sinks, mops, medicines, and physiotherapy and hydrotherapy pools. 
The CDC reported that the overall incidence of P. aeruginosa infections in USA hospitals averages about 0.4% (4 per 1,000 discharges), and the bacterium is the fourth most commonly-isolated nosocomial pathogen, accounting for 10.1% of all hospital-acquired infections.  Rates of infection due to resistant P. aeruginosa continue to increase globally, as does resistance to both quinolones and carbapenems. Aminoglycoside resistance is emerging as a significant problem. , Patients at risk include those in intensive care units (ICUs), particularly those who are ventilator-dependent, and individuals with cystic fibrosis.  There is little hope of new antibiotics, making polymyxins, a class of drug that was forgotten for quite some time, the first line in the management of infections caused by carbapenem-resistant bacteria.
Susceptibility trends of P. aeruginosa isolates from Kerala are few and far between. In this scenario, we analysed the antibiotic resistance patterns of P. aeruginosa isolated from our hospital in the past few years to analyse the susceptibility trends over the past few years.
| Materials and methods|| |
The study was retrospective and observational. Clinically relevant P. aeruginosa isolates were obtained from blood, body fluids, urine, tissue, pus and respiratoryspecimens during 2007, 2012, 2013 and 2014. When multiple isolations were available, the first isolate per person per 365-day period (non-duplicate isolates) only were included. P. aeruginosa-in-pus specimens were considered as clinically significant only if obtained by deep aspirations or repeated isolations.
Antimicrobial susceptibility testing was done according to the Clinical Standards Laboratory Institute (CLSI) standards for 2007, 2012, 2013 and 2014 by disc diffusion method for the following antibiotics: Amikacin (30 μg), Ceftazidime (30 μg), Ciprofloxacin (5 μg), Gentamicin (10 μg), Imipenem (10 μg) and Piperacillin + Tazobactam (100/10 μg). Comparison of P. aeruginosa isolates was carried out with Escherichia coli and K. pneumoniae for the year 2014 to observe the current trends in antibiotic susceptibility in other commonly isolated Gram-negative bacteria.
Proportions of resistant and sensitive pseudomonad strains were calculated and compared for the four years under study. All variables were tabulated using frequency tables, along with pattern of susceptibility to commonly used antibiotics. Chi-square tests were done to determine the statistical significance. A P value of <0.05 was considered as statistically significant.
| Results|| |
We received a total of 484 non-duplicate P. aeruginosa isolates in 2014, 444 isolates in 2013, 350 isolates in 2012 and 460 isolates in 2007. Analysis of these isolates showed [Table 1] that most of them were from pus, followed by urine and lower respiratory specimens.
|Table 1: Various sites from which P. aeruginosa was isolated during the study period|
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There were statistically significant increases in the sensitivities of P. aeruginosa from 2007 to 2014 [Figure 1]: Piperacillin/Tazobactam (chi-square = 153.84; P = 0.000); Ceftazidime (chi-square = 92.73; P = 0.000); Imipenem (chi-square = 104.69; P = 0.000); Amikacin (chi-square = 199.02; P = 0.000); Gentamicin (chi-square = 260.31; P = 0.000); and Ciprofloxacin (chi-square = 259.56; P = 0.000).
|Figure 1: Comparison of sensitivity patterns of Pseudomonas aeruginosa in 2007, 2012, 2013 and 2014|
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Considering the increasing susceptibility of P. aeruginosa in the past few years, the susceptibilities of commonly isolated Gram-negative bacteria such as E. coli and K. pneumoniae were compared for the same year, that is, 2014 [Table 2]. The sensitivities of P. aeruginosa versus E. coli with regard to Piperacillin + Tazobactam (chi-sq. = 18.28; P = 0.000); Ceftazidime (chi-sq. = 224.02; P = 0.000); Imipenem (chi-sq. = 5.18; P = 0.023); Amikacin (chi-sq. = 9.07; P = 0.003); Ciprofloxacin (chi-sq. = 257.35; P = 0.000) were statistically much higher for the same period except for Gentamicin (chi-sq. = 0.065; P = 0.799). P. aeruginosa also had higher sensitivities versus Klebsiella with regard to Piperacillin + Tazobactam (chi-sq. = 69.36; P = 0.000); Ceftazidime (chi-sq. = 198.81; P = 0.000); Imipenem (chi-sq. = 5.18; P = 0.023); Amikacin (chi-sq. = 10.98; P = 0.001); Gentamicin (chi-sq. = 8.524; P = 0.004) and Ciprofloxacin (chi-sq. = 62.86; P = 0.000).
|Table 2: Comparison of susceptibility of P. aeruginosa with E. coli and Klebsiella pneumoniae for the year 2014|
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Resistance to carbapenems is on the rise worldwide in most enterobacteriaceae, including K. pneumoniae, in the last few years. Imipenem resistance [Figure 2] in K. pneumoniae in 2007 was lower at 3% for our isolates but moved up to 17% in 2013 and to 15% in 2014. In stark contrast, the resistance in P. aeruginosa was as high as 27% in 2007 for Imipenem but fell to 10% in 2014 (chi-sq. = 114.88; P = 0.000).
|Figure 2: Comparison of resistance to Imipenem between 2007 and 2012, 13 and 14|
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Analysis of trends of Pseudomonas aeruginosa
Considering the peculiar trend of increasing susceptibility in P. aeruginosa at a time when other commonly isolated Gram-negative bacteria isolated from the same hospital showed decreased sensitivity, we analysed whether most of our strains of P. aeruginosa differed between various locations [Table 3] within the hospital, such as ICUs, inpatients (IP) and outpatients (OP). The numbers of specimens of P. aeruginosa did not vary in terms of contribution from OP (chi-sq. = 0.39; P = 0.942), whereas the contribution from IP (chi-sq. = 12.22; P = 0.007) and ICU were altered only in 2012 (chi-sq. = 11.19; P = 0.011). In 2012, the number of the isolates of Pseudomonas from the IP category was higher than from ICUs. The sensitivities of various antibiotics among these three locations were analysed for 2014 [Table 4].
|Table 3: Distribution of Pseudomonas in the various locations of the hospital|
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|Table 4: Sensitivities of P. aeruginosa between various locations in 2014|
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The isolates were further analysed across the years from the various sites it was obtained. Pus isolates, lower respiratory tract specimens and urine specimens were analysed over the years to observe the difference in trends, if any. Moreover, the proportion of pan-sensitive isolates were also analysed among these specimens for 2013 and 2014 [Table 5].
There were increases in the sensitivities of P. aeruginosa from pus isolates [Figure 3] between 2007 and 2014 with regard to Piperacillin + Tazobactam (chi-sq. = 79.98; P = 0.000); Ceftazidime (chi-sq. = 27.89; P = 0.000); Imipenem (chi-sq. = 88.01; P = 0.000); Amikacin (chi-sq. = 74.77; P = 0.000); Gentamicin (chi-sq. = 122.76; P = 0.000) and Ciprofloxacin (chi-sq. = 66.76; P = 0.000).
|Figure 3: Comparison of sensitivity patterns of Pseudomonas aeruginosa in pus specimens|
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There were increases in sensitivities of P. aeruginosa from lower respiratory tract infection (LRTI) specimens such as sputum and broncho-alveolar lavage [Figure 4] between 2007 and 2014 with regard to Piperacillin/Tazobactam (chi-sq. = 103.49; P = 0.000); Ceftazidime (chi-sq. = 58.35; P = 0.000); Imipenem (chi-sq. = 39.55; P = 0.000); Amikacin (chi-sq. = 65.98; P = 0.000); Gentamicin (chi-sq. = 158.71; P = 0.000) and Ciprofloxacin (chi-sq. = 132.74; P = 0.000).
|Figure 4: Comparison of sensitivity patterns of Pseudomonas aeruginosa in lower respiratory tract specimens|
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There were increases in the sensitivities of P. aeruginosa from urine specimens [Figure 5] between 2007 and 2014 with regard to Ceftazidime (chi-sq. = 13.77; P = 0.003); Piperacillin + Tazobactam (chi-sq. 8.36; P 0.0391), Imipenem (chi-sq. = 16.78; P = 0.001); Amikacin (chi-sq. = 132.65; P = 0.000); Gentamicin (chi-sq. = 132.47; P = 0.000) and Ciprofloxacin (chi-sq. = 90.24; P = 0.000).
|Figure 5: Comparison of sensitivity patterns of Pseudomonas aeruginosa in urine specimens|
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| Discussion|| |
0P. aeruginosa presents a serious therapeutic challenge for the treatment of both community-acquired and hospital-acquired infections, and selection of the appropriate antibiotic is essential for an optimal outcome. Unfortunately, the selection of the most appropriate antibiotic is complicated by the ability of P. aeruginosa to develop resistance to multiple classes of antibacterial agents, even during the course of treating an infection. Epidemiological outcome studies have shown that infections caused by drug-resistant P. aeruginosa are associated with significant increases in morbidity, mortality, need for surgical intervention, length of hospital stay and overall cost of treating the infection.  Aminoglycosides, fluoroquinolones, Cephalosporins and carbapenems have been used for the treatment of infections caused by P. aeruginosa. However, a decreased susceptibility rate of P. aeruginosa to β-lactams, carbapenems, quinolones and aminoglycosides has been reported worldwide. Not only are rates of resistance to individual drugs or drug classes a concern, but the prevalence of multidrug-resistant strains (resistant to three or more drug classes) is an even more serious therapeutic challenge.
During the years, 2001-2006, rates of non-susceptibility among P. aeruginosa isolates in Brooklyn, NY, USA ranged 27-29% for Cefepime, 30-31% for Imipenem, 23% for Meropenem and 41-44% for Ciprofloxacin.  Closer to home at Salem, (MA, USA,) Mohanasoundaram  has reported resistance to Ceftazidime (63%), Ciprofloxacin (62.5%), Gentamicin (67.8%), Amikacin (54.4%), Piperacillin + Tazobactam (34%) and Imipenem (16%) in 2010.
In the present study, the resistance of P. aeruginosa to the antibiotics used in 2007 is comparable to these studies referred above, but the data for 2012 and 2013 were radically different, and the decreases in resistance to several antibiotics studied were statistically significant [Figure 1] and [Figure 2]. The decreases in resistance to carbapenems (from 27% in 2007 to 10% in 2014), Piperacillin + Tazobactam (from 47% to 16%), Gentamicin (from 69% to 25.5%), Amikacin (from 49% to 16%) and Ciprofloxacin (from 70% to 24%) deserve special attention.
Isolates of Pseudomonas were also analysed across the years for common specimens such as the pus, urine and Lower respiratory tract from which it was isolated. All the specimens showed a consistent increase in sensitivity to all the antibiotics tested over the years [Figure 3],[Figure 4] and [Figure 5]. Specimens from the lower respiratory tract were generally more sensitive than from urine and pus [Figure 3],[Figure 4] and [Figure 5] and [Table 5].
In our hospital, other Gram-negative bacilli such as E. coli and K. pneumoniae are far more resistant to the same antibiotics compared to P. aeruginosa in 2014 [Table 2]. Sensitivities of P. aeruginosa to Imipenem, Piperacillin + Tazobactam, Ciprofloxacin and Amikacin were 90%, 84%, 76% and 84% respectively in 2014 when compared to K. pneumoniae (85%, 62%, 54% and 76%) and E. coli (93%, 74%, 33% and 89%) respectively. P. aeruginosa strains that were sensitive to all the tested antibiotics (pan-sensitive strains) were 62% in 2013 and 67% 2014 [Table 5]. In comparison, pan-sensitive isolates to the equivalent antibiotics of E. coli and K. pneumoniae were 6% and 22% in the same period.
It is to be considered that the bulk of our pseudomonad isolates in the study came from the ICU admissions or from IP (86%) and in contrast to OP, which contributed to approximately 15% of the study population [Table 3]. Isolates from the ICU in 2014 [Table 4] were predictably more resistant to Ceftazidime, Imipenem and Ciprofloxacin (30%, 17%, 31%) than the OP (22%, 9%, 13%).
We tried to reason out this falling trend of resistance seen only in P. aeruginosa isolates. Looking through the pharmacy records, the most common drug sold in 2014 was Ciprofloxacin (32223 prescriptions), followed by Piperacillin+Tazobactam (23985 prescriptions), while in 2007 the most common drug was Ciprofloxacin (31244 prescriptions), followed by Gentamicin (9160 prescriptions). Though this explains the increased resistance to Ciprofloxacin and Gentamicin in 2007, it does not effectively explain the increased sensitivity in 2014 to the same drugs, especially Ciprofloxacin. Antibiotic policy based on susceptibility trends in our hospital was constituted and implemented from 2013 and it recommended Piperacillin + Tazobactam as an empiric choice for most Gram-negative bacterial infections pending culture reports. This probably explains the increased prescriptions of Piperacillin+Tazobactam in our hospital in 2014. But again, despite the increased use of Piperacillin+Tazobactam since 2013, increase in sensitivity to P. aeruginosa is a contradiction. Meanwhile, we also went through a random assortment of 60 case sheets of patients with P. aeruginosa infection, 20 each in 2010, 2013 and 2014. We could not access the 2007 medical records as they were unavailable due to the institutional policy of keeping records for only the past five years. Almost all cases were treated successfully with Ceftazidime in 2010, while in 2013 and 2014 there was a shift in treatment to Piperacillin + Tazobactam. All the 60 patients completed treatment successfully and were discharged on recovery except for one patient in 2013, who had P. aeruginosa pneumonia and sepsis. This patient had a metallo-β-lactamase (MBL)-producing strain which was resistant to all known antibiotics except polymyxins. She expired in spite of aggressive therapy with colistin. A P. aeruginosa infection with a similar antibiogram has not been isolated from our hospital after this.
Interestingly, the fall in resistance of P. aeruginosa occurred during a period of increased antibiotic consumption globally, which could have led to increase in selection pressure and enhancement of known or evolution of new resistance mechanisms of bacteria; this needs to be studied further, mainly by sequencing. Mechanisms of resistance  in P. aeruginosa include overexpression of efflux pump mechanism for resistance to carbapenems, acquisition of extended-spectrum β-lactamases (ESBLs) to third- and fourth-generation Cephalosporins and MBLs, conferring resistance to all known beta-lactams including carbapenems; target site or outer membrane modification that confer resistance to beta-lactams, aminoglyocsides and quinolones are predominant. P. aeruginosa develops resistance to antibacterials, either through the acquisition of resistance genes on mobile genetic elements (that is, plasmids) or through mutations that alter the expression and/or function of chromosomally encoded mechanisms.
Loss of these plasmids and/or mutations most likely explains the decrease in the resistance characteristics in P. aeruginosa leading to the circulation of a sensitive strain. Reasons for the loss of resistance need to be carefully looked into. One reason could be the presence of pseudomonad-specific plasmid-dependant bacteriophages in the local environment. These phages notably reduce the development of multidrug resistance by infecting the bacteria harboring multidrug-resistant plasmids and killing them.  In a study at Sankara Nethralaya, Chennai,  three strains isolated in the hospital, two resistant and one sensitive, were sequenced and it was found that all three had similar genome size. However, it was observed that the resistant strains had higher numbers of RNA-coding genes compared to the sensitive ones, probably resulting in a higher expression of proteins among drug-resistant strains.
Two other mechanisms that have been published are the counter-transcript RNA (ctRNA)-based control mechanisms and the plasmid addiction systems.  For both these mechanisms, small drug-like organic compounds bind to speciﬁc macromolecules involved in plasmid replication and induce plasmid elimination, thus leading to sensitive bacteria. 
A detailed genomic- and/or proteomic-level study for our isolates is much desired to understand this loss of resistance so that the same may be replicated in other bacteria. This would offer a new weapon in the war we are currently losing against bacterial infections.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]