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Table of Contents
ORIGINAL ARTICLE
Year : 2020  |  Volume : 9  |  Issue : 4  |  Page : 200-205

A study of bacterial pathogens obtained from various samples in a tertiary care hospital and their antimicrobial susceptibility patterns


Department of Microbiology, Sri Venkateswara Institute of Medical Sciences, Tirupathi, Andhra Pradesh, India

Date of Submission14-Nov-2018
Date of Decision22-May-2020
Date of Acceptance13-Aug-2020
Date of Web Publication5-Jan-2021

Correspondence Address:
R Jayaprada
Assistant Professor, Department of Microbiology, Sri Venkateswara Institute of Medical Sciences, Tirupati 517 507, Andhra Pradesh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/JCSR.JCSR_27_18

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  Abstract 


Background: Antimicrobial resistance (AMR) is of great concern in recent years as it is posing a global threat. The present study was undertaken to identify the prevalence of common bacterial isolates and their antimicrobial susceptibility patterns and to tackle the AMR by strict implementation of antibiotic stewardship programme (AMSP).
Methods: We prospectively studied the microbiome and its antibiotic susceptibility patterns from various clinical samples at our tertiary care hospital, from January to June 2018. All samples were processed as per standard protocols. Antibiotic susceptibility testing was performed using Kirby–Bauer disc diffusion method as per the Clinical and Laboratory Standards Institute 2016 guidelines.
Results: Escherichia coli was the predominant organism followed by Klebsiella spp, Acinetobacter spp and Pseudomonas spp. More than 50% resistance was observed to third generation cephalosporins and quinolones in all Gram negative isolates. Methicillin-resistant Staphylococcus aureus and methicillin-resistant coagulase negative Staphylococcus constituted 41% and 54% respectively. Vancomycin intermediate-sensitive S. aureus/vancomycin resistance S. aureus were not evident. Vancomycin-resistant enterococci (n = 10) were also isolated, but all were sensitive to linezolid.
Conclusions: The present study highlights the alarming development of multidrug-resistance to almost all the drugs with a very few exceptions. Continuous and periodic evaluation of bacteriological profile and its susceptibility patterns are helpful in formulating appropriate antibiotic policy to tackle the AMR.

Keywords: Antimicrobial stewardship, bacterial, drug resistance, multiple


How to cite this article:
Ramakrishna N, Jayaprada R, Sharma K K, Yamini S, Srikala V S. A study of bacterial pathogens obtained from various samples in a tertiary care hospital and their antimicrobial susceptibility patterns. J Clin Sci Res 2020;9:200-5

How to cite this URL:
Ramakrishna N, Jayaprada R, Sharma K K, Yamini S, Srikala V S. A study of bacterial pathogens obtained from various samples in a tertiary care hospital and their antimicrobial susceptibility patterns. J Clin Sci Res [serial online] 2020 [cited 2021 Jan 27];9:200-5. Available from: https://www.jcsr.co.in/text.asp?2020/9/4/200/306190




  Introduction Top


In spite of vast advances made by medical science, bacterial infection remains a major cause of concern. Throughout the world, bacterial infections are one of the leading causes of morbidity, mortality, responsible for increased healthcare cost and account for major burden on patients and public health system of any country.[1],[2]

The increased risk of bacterial infection is further compounded by rising trends of antibiotic resistance in commonly implicated organisms all over the world.[3] Antibiotic resistance among bacteria is becoming a more and more serious problem throughout the world. This is particularly true in the case of members of Enterobacteriaceae group such as Escherichia coli and Klebsiella spp. and non-fermenter group of bacteria such as Pseudomonas spp. and Acinetobacter spp.[3],[4]

Increasing resistance among Gram-positive organisms is also a matter of concern, and high rates of methicillin resistance Staphylococcus aureus (MRSA) in clinical samples have been noted. Similarly, resistance to glycopeptide antibiotics such as vancomycin and teicoplanin among clinical isolates of Enterococcus spp. is also increasing.[5],[6]

The pattern of bacteria causing infections and their antibiogram vary widely from one country to another, as well as from one hospital to another and even among intensive care units with one hospital to another.[2],[4],[5] There also appears to be a significant lack of studies highlighting susceptibility patterns of locally prevalent organisms.

Knowledge of predominantly isolated bacterial microorganisms and their sensitivity to available drugs is of immense value to the rational selection of antimicrobial agents and for development of appropriate antibiotic policies. Therefore, the present study was undertaken to identify the prevalence of common bacterial isolates and their antimicrobial susceptibility pattern of various clinical samples from patients attending to a tertiary care hospital.


  Material and Methods Top


The present prospective study and was carried out at the Department of Microbiology, Sri Venkateswara Institute of Medical Sciences (SVIMS), Tirupathi, Andhra Pradesh, a tertiary care hospital, from January to June 2018, after obtaining approval from the Institutional Ethics Committee.

The samples collected were mainly urine, blood, pus, sputum, cerebrospinal fluid, throat swab, stool, endotracheal tips/aspirates and other body fluids such as pericardial fluid, pleural fluid, peritoneal fluid, peritoneal dialysis fluid, synovial fluid, percutaneous nephrostomy fluid, bone marrow, wound aspirate, lymph node aspirate, lymph node biopsy, cyst fluid, bone and tissues. The samples were received and processed within 1 h of collection for culture and sensitivity.

The samples were then inoculated on nutrient agar, blood agar, chocolate agar and MacConkey agar plates and incubated aerobically at 37°C temperature for 24–48 h. Growth was processed according to standard microbiological techniques which include Gram staining, colony characteristics, biochemical properties and antibiotic susceptibility testing.[7],[8],[9]

Criteria for antimicrobial sensitivity testing were performed as per the Clinical and Laboratory Standards Institute (CLSI) 2016 guidelines.[10] Antimicrobial sensitivity testing was done on Mueller Hinton agar (MHA) by Kirby–Bauer's disc diffusion method. Commercially available discs (HiMedia) were used.

For Enterobacteriaceae and Acinetobacter groups, first-line drug susceptibility testing was done for ampicillin (10 μg), amoxycillin-clavulanate (20/10 μg), cefotaxime (30 μg), cefoperazone-sulbactam (75/10 μg), imipenem (10 μg), ciprofloxacin (5 μg), amikacin (30 μg), gentamicin (10 μg), piperacillin-tazobactam (100/10 μg) and co-trimoxazole (25 μg). Second line drug susceptibility testing was done for cefepime (30 μg), cefoxitin (30 μg), ceftazidime (30 μg), chloramphenicol (30 μg), tetracycline (30 μg), netilmicin (30 μg), meropenem (10 μg), polymyxin B (300 U) and tigecycline (15 μg).

For Pseudomonas species, first line drug susceptibility testing was done for ampicillin, amoxycillin-clavulanate and co-trimoxazole were replaced with ceftazidime (30 μg), netilmicin (30 μg) and polymyxin B (300 U). Second line drug susceptibility testing was done for aztreonam (30 μg), carbenicillin (100 μg), cefepime (30 μg), meropenem (10 μg) and tobramycin (10 μg). The media and antibiotic discs were procured from HiMedia (India).

For polymyxin B, no CLSI guidelines are available for interpretation of disc diffusion technique as regards to Enterobacteriaceae and Acinetobacter spp. For these isolates, the recommendation of Galani et al.[11] was adopted. Nitrofurantoin (300 μg) and nalidixic acid (30 μg) were additionally used in case of urine isolates. Methicillin resistance in S. aureus was tested using MHA with cefoxitin disc (30 μg) by Kirby–Bauer disc diffusion method as per the CLSI guidelines 2016.[10]

Methicillin resistance was determined using cefoxitin (30 μg) disk on MHA as per the CLSI guidelines, and results were read after 18 h of incubation at 35 °C. The S. aureus isolates which showed zone size =21 mm were considered as MRSA and coagulase-negative staphylococci (CoNS) which showed zone size =24 mm were considered as methicillin-resistant CoNS (MRCoNS).

A standard bacteriological procedure was followed to perpetuate correct laboratory test results. American Type Culture Collection (ATCC) standard reference strains S. aureus (ATCC 25923), E. coli (ATCC25922) and Pseudomonas aeruginosa (ATCC 27853) were used as quality control throughout the study for culture and antimicrobial susceptibility testing.

The study was carried out during the period of January 2018–July 2018; a total of 8189 clinical samples were evaluated. The clinical samples received from various departments of the hospital were included in the study.

Statistical analysis

Descriptive statistics were done by measuring proportion of each species of bacteria from the clinical specimens. The proportion of bacteria which were susceptible/resistant to each antibiotic was done. The above analysis was done using Microsoft Excel 2016 (Microsoft Corp., Redmond, WA).


  Results Top


During the study period, 3865 samples tested culture positive. Of these, 2380 (61.57%) were from male patients. The positive isolates were obtained from the following samples: urine (1621), blood (575), pus (439), sputum (144), body fluids (65), endotracheal tips/aspirates (345), throat swab (110), peritoneal dialysis fluid (11), wound tissue (35) and others (531). The most common Gram-negative isolates were E. coli, Klebsiella spp., Acinetobacter spp. and Pseudomonas spp. [Table 1].
Table 1: Sample-wise distribution of isolates

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Most of the E. coli were isolated from urine samples followed by pus, blood, endotracheal tips/aspirates and from other samples. Klebsiella species were predominantly isolated from urine samples followed by endotracheal tips/aspirates, blood, pus, sputum and from other samples. Majority of Acinetobacter spp. were isolated from endotracheal tips/aspirates followed by pus, blood, sputum and from other samples and Pseudomonas sp. were predominantly isolated from endotracheal tips/aspirates followed by pus, blood, urine and from other samples.

Antimicrobial resistance (AMR) pattern of E. coli isolates had shown high resistance to cefotaxime (77%), ciprofloxacin (52%) and co-trimoxazole (57%), and E. coli isolates were highly sensitive to amikacin (80%), cefoperazone + sulbactam (75%), gentamicin (82%), piperacillin + tazobactam (80%), imipenem (85%) and colistin/polymyxin B (100%).

Among Klebsiella spp., isolates were highly resistance to cefotaxime (58%), ciprofloxacin (62%), amikacin (56%), gentamicin (52%) and co-trimoxazole (67%) and were sensitive to cefoperazone + sulbactam (50%), piperacillin + tazobactam (59%), imipenem (62%) and colistin/polymyxin B (100%).

Similar to the above-mentioned Gram-negative bacteria, Acinetobacter spp. isolates were highly resistance to cefotaxime (86%), ciprofloxacin (58), co-trimoxazole (54%), amikacin (54%) and gentamicin (53%) and sensitive to piperacillin + tazobactam (55%), imipenem (77%), cefoperazone + sulbactam (73%) and colistin/polymyxin B (100%).

In contrast to the other Gram-negative bacilli, Pseudomonas spp. isolates were highly sensitive to ceftazidime (65%), imipenem (88%), amikacin (78%), gentamicin (78%), piperacillin + tazobactam (76%), cefoperazone + sulbactam (57%) and colistin/polymyxin B (100%) and resistance to ciprofloxacin was 49% [Figure 1].
Figure 1: Organism-wise antimicrobial resistance pattern in Gram-negative bacilli (%). AK = Amikacin; CTX = Cefotaxime; CFS = Cefoperazone + sulbactam; CF = Ciprofloxacin; COT = Co-trimoxazole; G = Gentamicin; I = Imipenem; PTZ = Piperacillin + tazobactam; Pb = Polymyxin-b; CTZ = Ceftazidime

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Because of strict implementation of antibiotic stewardship programme (AMSP) and infection control practices, percentage has declined from 68% in January to 25% in December [Figure 2].
Figure 2: Month-wise distribution of Klebsiella pneumoniae carbapenemases
KPC = Klebsiella pneumoniae carbapenemase


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Screening of healthcare workers (HCWs) for MRSA should be done as MRSA percentage was 41 and methicillin resistance coagulase-negative Staphylococcus percentage was 54, and these isolates are predominantly from emergency and nephrology departments. HCWs were treated intranasally with 2% mupirocin for 5 days. Vancomycin resistance in Enterococcus (VRE) was 2%. All VRE (n = 10) were isolated from urine samples except one which was isolated from blood sample, but all were sensitive to linezolid. Most of the MRSA were isolated from pus followed by blood, endotracheal tips/aspirates, urine, sputum and from other samples [Figure 3].
Figure 3: Antimicrobial resistance patterns of Staphylococcus aureus, coagulase-negati ve staphylococci and E n t e r o c o c c u s MRSA = Methicillin resistance Staphylococcus aureus ; MRCoNS = Methicillin-resistant coagulase-negative Staphylococcus; VRSA = Vancomycin resistance Staphylococcus aureus; VISA = Vancomycin intermediate-sensitive Staphylococcus aureus; VRE = Vancomycin-resistant Enterococcus spp.

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  Discussion Top


In our prospective study from January to June 2018, E. coli was the predominant organism followed by Klebsiella, Acinetobacter and Pseudomonas. Most of the E. coli were isolated from urine samples followed by pus, blood, endotracheal tips/aspirates and from other samples. Similar results were reported in a study[13] from a tertiary care hospital in South India, Chennai. A high proportion of E. coli isolates had shown resistance to cefotaxime (77%), ciprofloxacin (52%), co-trimoxazole (57%) and sensitive to amikacin (80%), cefoperazone + sulbactam (75%), gentamicin (82%), piperacillin + tazobactam (80%), imipenem (85%) and colistin/polymyxin B (100%), which was similar to other studies.[12],[13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24]

In our study, MRSA percentage was 41% and MRCoNS were 54% and vancomycin intermediate-sensitive S. aureus/vancomycin resistance S. aureus (VISA/VRSA) were nil. The high incidence of MRSA in the present study was comparable to other studies conducted in different areas including 55% in Banaras Hindu University, 43% in Davangere, 39% in AIIMS and 31% in a Chennai hospital.[16],[22],[25],[26] As prevalence of MRSA is on rise and the treatment of option for MRSA being vancomycin this may lead to emergence of vancomycin intermediate-resistant S. aureus and vancomycin-resistant S. aureus. VISA and VRE have been reported from, South India, during 2004–2006.[26] To prevent emergence of VISA, VRSA and VRE, it requires continuous monitoring of minimum inhibitory concentration levels of vancomycin and its restricted judicious use in clinical practice is of great concern. As a part of AMSP, we have initiated a high-end antibiotic monitoring sheet as per the National Centre for Disease Control,[27] so that monitoring of vancomycin use in clinical practice can be made more vigilant.

In our study, multidrug-resistant (MDR) Pseudomonas isolates were reported in pus and endotracheal tips/aspirates samples, suggesting that these isolates are common healthcare-associated infections (HAIs) as has been documented in other studies.[15],[28],[29] Majority of these Pseudomonas isolates contributed by pus samples, which was similar to other studies.[17] In the present study, an increased sensitivity to amikacin when compared to quinolones, and increasing trends of resistance were observed against third-generation cephalosporins; similar observations were reported in another study.[16]

In the present study, Klebsiella isolates were highly resistance to cefotaxime (58%), ciprofloxacin (62%), amikacin (56%), gentamicin (52%) and co-trimoxazole (67%) and sensitive to cefoperazone + sulbactam (50%), piperacillin + tazobactam (59%), imipenem (62%) and colistin/polymyxin B (100%). This observation is mimicking the results of many studies.[14],[15],[16],[17],[29],[30],[31] Vancomycin-resistant enterococci (n = 10) were isolated from urine samples and also from blood culture of one patient, but all were sensitive to linezolid.

Acinetobacter spp. were isolated from endotracheal tips/aspirates, pus, blood, sputum and from other samples, which were highly resistance to cefotaxime (86%), ciprofloxacin (58), co-trimoxazole (54%), amikacin (54%) and gentamicin (53%) and sensitive to piperacillin + tazobactam (55%), imipenem (77%), cefoperazone + sulbactam (73%) and colistin/polymyxin B (100%). Similar sensitivity patterns were reported from South India.[16]

In the present study, most of the Gram-negative isolates developed resistance against cephalosporins and it is due to the widespread usage of these antibiotics empirically for all suspecting infections. Restriction of the usage of third- and fourth-generation cephalosporins by replacing with other alternatives would be recommended for rapid development of resistance against these antibiotics. Increasing trends of resistance to co-trimoxazole and quinolones were also reported in our present study. In a study,[32] they reported that the increased use of quinolones might be the result of increased resistance to this antibiotic. In our study, a low level of resistance was reported for drugs such as aminoglycoside, carbapenems and beta-lactamase inhibitor combinations.

Empirical use of antibiotics in hospitalised patients has been focussed in some studies,[15],[27],[32],[33] which was also observed in our present study. Inappropriate usage of antimicrobial agents is one of the important factors which develop AMR. However, recent therapy with antibiotics was not considered as a risk factor for the development of MDR organisms.[34]

Organisms isolated from HAI were more resistant to antibiotics than the community-acquired isolates as evidenced by many studies.[15],[21] In a study[20] reported that nasal carriage of MRSA was identified without prior antimicrobial therapy.

The results of the present study highlight the alarming development of resistance to almost all the drugs with a very few exceptions of drugs in our study. Microbial resistance patterns related to area-wise and time-based variations were observed in our study on comparing with previous studies. Continuous and periodic evaluation of bacteriological profile and its susceptibility patterns expedites the successful treatment of infection and is of significant help in formulating appropriate antibiotic policy to tackle the AMR. Pandemic AMR surveillance and strict implementation of AMSP are the key essential elements to combat the emergence of multidrug resistance.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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