CROSSMARK_Color_Square https://doi.org/10.35845/kmuj.2024.23499                                  ORIGINAL ARTICLE

Antimicrobial susceptibility patterns of Linezolid-resistant coagulase-negative staphylococci in Pakistan

Arooj Sattar  1, Namra Mahmood  1 , Zahra Riaz  1, Iram Yousaf 1 , Rubeena Hafeez 1, Sakina Jamil 2

 

1: Department of Pathology, Central Park Medical College, Lahore, Pakistan

2: Department of Pathology, Pak Red Crescent Medical and Dental College, Lahore, Pakistan

 

Email Description: Description: Description: C:\Users\Asghars\Downloads\email corrspondence.gif: draroojsattar@gmail.com
Contact #: +92-321-9881893

 

Date Submitted:  September 26, 2023

Date Revised:      June 16, 2024

Date Accepted:   July 03, 2024

THIS ARTICLE MAY BE CITED AS: Sattar A, Mahmood N, Riaz Z, Yousaf I, Hafeez R, Jamil S. Antimicrobial susceptibility patterns of Linezolid resistant coagulase-negative staphylococci in Pakistan. Khyber Med Univ J 2024;16(3):182-86. https://doi.org/10.35845/kmuj.2024.23499

ABSTRACT

OBJECTIVE: To determine the antimicrobial resistance pattern in Linezolid-resistant coagulase negative staphylococcus (CoNS) isolates in Pakistan.

METHODS: This cross-sectional study was conducted from February and July 2022, at a private laboratory in Lahore, with samples collected from various regions across Pakistan. The study included microbiological samples such as blood and wound specimens, from which 4153 isolates of CoNS were identified. Of these, only 64 isolates that were both coagulase-negative and Linezolid-resistant were selected for further investigation. The antimicrobial susceptibility patterns of these Linezolid-resistant isolates were analyzed. The antibiotics tested included Amikacin, Ciprofloxacin, Clindamycin, Doxycycline, Erythromycin, FusidicAacid, Gentamicin, Oxacillin,Tteicoplanin, Trimethoprim-Sulfamethoxazole, and Vancomycin. Data analysis was performed using SPSS version-22.

RESULTS: Among 4153 samples, 64 (1.54%) were Linezolid-resistant CoNS isolates, predominantly from adults (aging 18-59 years) and senior adults (aging >59 years), with a higher prevalence in males (56.3%). Resistance was observed in 51.6% of blood and 48.4% of pus samples. Most isolates were also resistant to Teicoplanin (100%), Ciprofloxacin (92.2%), Oxacillin (89.1%), and Fusidic Acid (84.4%), While Sensitivity Was Highest for Vancomycin (85.9%) and Doxycycline (79.7%). Statistically significant differences were noted for all antibiotics except erythromycin.

CONCLUSION: Our findings highlight the importance of monitoring the evolution of Linezolid resistance in CoNS as Linezolid resistant isolates also showed high resistance to some other major antimicrobial drugs (e.g. Teicoplanin, Ciprofloxacin, Oxacillin etc.). Linezolid resistance must be closely monitored, especially when frequent and prolonged Linezolid therapy is indicated to implement control measures and reduce the risk of CoNS spreading in the community.

KEY WORDS: Linezolid (MeSH); Multidrug Resistance (MeSH); Drug Resistance, Microbial (MeSH); Drug Resistances, Microbial (MeSH); Antimicrobial drug resistance (Non-MeSH); Microbial Sensitivity Test (MeSH); Coagulase (MeSH); Staphylococcal Infections (MeSH); Bloodstream Infection (Non-MeSH); Vancomycin (MeSH); Infections, Nosocomial (MeSH); Cross Infection (MeSH).

 

INTRODUCTION

 

Coagulase negative Staphylococci (CoNS) are a significant component of the normal cutaneous and mucous flora. However, with the increasing use of implanted medical devices and procedure-related changes, they have become one of the leading causes of nosocomial infections.1  Managing CoNS infections is challenging, as these isolates often show resistance to various antimicrobials, particularly Penicillins and Oxacillin/Methicillin.2

Methicillin-resistant Staphylococcus aureus (MRSA) and CoNS are among the Gram-positive bacteria effectively treated by the Oxazolidinone antibiotic, linezolid.3 Despite Linezolid being a cornerstone in the treatment of multidrug-resistant Gram-positive infections, resistance to this drug has emerged.4  It works by binding to the 50S ribosomal subunit, preventing the assembly of the initiation complex and thereby inhibiting protein synthesis with its bacteriostatic properties.5 Linezolid resistance is often linked to mutations in domain V of 23rRNA with G2576T substitution and acquisition of methyltransferase gene cfr.6,7

Linezolid is a very desirable antibiotic for the therapy of possible or proven staphylococcal infections due to its great oral absorption.8 However, the earliest Linezolid resistance strain was claimed in 2001 in the USA.6 Since then, Linezolid resistant strains start to appear worldwide.  In many regions such as North America, Ireland, China, Brazil, Greece, Spain, Italy and France, Linezolid resistance in CoNS infections has been reported.2 In Pakistan, a tertiary care cardiac facility reported the earliest case of coagulase-negative, Linezolid-resistant Staphylococcus in 2018.9 There is relatively little research done in Pakistan, despite the fact that Linezolid resistance in CoNS infections constitutes a significant health risk.

METHODS

This cross-sectional study was conducted from February 2022 to July 2022 in a private laboratory in Lahore, which received samples from various regions across Pakistan. Ethical approval for the study was granted on February 3, 2022, with IRB number CIP/IRB/1099. The study included 4153 isolates of CoNS derived from microbiological samples such as blood and wound cultures. CoNS were identified based on positive gram staining (cocci in clusters) and catalase tests, with negative tube coagulase tests. Among these, only 64 isolates that were resistant to Linezolid were included in this study, while CoNS isolates that were Linezolid-sensitive were excluded.

The isolates were further identified and characterized using MALDI-TOF and Vitek2 techniques. Antimicrobial susceptibility was assessed using the agar dilution method and disc diffusion method. Minimal inhibitory concentrations (MICs) were determined using the E test and Vitek. Staphylococcus aureus ATCC 25923 was used as a quality control for susceptibility testing, following the Clinical Laboratory Standards Institute (CLSI) guidelines.10 The study evaluated susceptibility to Linezolid, Amikacin, Ciprofloxacin, Clindamycin, Doxycycline, Erythromycin, Fusidic Acid, Gentamicin, Oxacillin, Teicoplanin, Trimethoprim-Sulfamethoxazole, and Vancomycin.

Data were entered and analyzed using SPSS version 22. The binomial test of proportion was employed to detect statistical differences between resistant and sensitive isolates, with statistical significance defined as a p-value ≤0.05.

RESULTS

Among the 4153 samples, 64 (1.54%) were Linezolid resistant. Among those 64 Linezolid resistant CoNS isolates, it was observed that most of the isolates were of adult patients followed by senior adults. The resistance of Linezolid was relatively more common among male patients. Out of those 64 samples, Linezolid resistance was observed in 51.6% blood samples followed by 48.4 % pus samples (Table I). 

Table I: Baseline features of patients with Linezolid resistant isolate

Variables

Frequency

Percentage

Age (years)

<12 (Children)

11

17.2

12-17 (Adolescents)

02

3.1

18-59 (Adults)

34

53.1

> 59 (Senior Adults)

17

26.6

Gender

Male

36

56.3

Female

28

43.8

Location

Punjab

51

79.7

Sindh

05

7.8

Khyber Pakhtunkhwa

08

12.5

Specimen

Blood

33

51.6

Wound

31

48.4

Antimicrobial sensitivity patterns showed that Linezolid-resistant isolates also demonstrated resistance to teicoplanin. About 54 (84.4%) isolates were resistant to fusidic acid, 59 (92.2%) were resistant to ciprofloxacin and 57 (89.1%) were resistant to oxacillin. However, most of the isolates were sensitive to vancomycin followed by doxycycline as shown in Table II.

Table II: Antimicrobial sensitivity pattern among Linezolid resistant isolates

Antibiotics

Resistant

Sensitive

p-value

Amikacin

17 (26.6%)

47 (73.4%)

0.000*

Ciprofloxacin

59 (92.2%)

5 (7.8%)

0.000*

Clindamycin

47 (73.4%)

17 (26.6%)

0.000*

Doxycycline

13 (20.3%)

51 (79.7%)

0.000*

Erythromycin

36 (56.3%)

28 (43.8%)

0.382

Fusidic Acid

54 (84.4%)

10 (15.6%)

0.000*

Gentamicin

47 (73.4%)

17 (26.6%)

0.000*

Oxacillin

57 (89.1%)

7 (10.9%)

0.000*

Teicoplanin

64 (100.0%)

0 (00.0%)

0.000*

Trimethoprim Sulfamethoxazole

46 (71.9%)

18 (28.1%)

0.001*

Vancomycin

9 (14.1%)

55 (85.9%)

0.000*

      *Statistically Significant Difference i.e. p-value ≤ 0.05

The binomial test of proportion was employed to assess the statistical differences between resistant and sensitive isolates. Statistically significant differences were observed among the resistant and sensitive isolates for Amikacin, Ciprofloxacin, Clindamycin, Doxycycline, Fusidic Acid, Gentamicin, Oxacillin, Teicoplanin, Trimethoprim-Sulfamethoxazole, and Vancomycin. However, at the 5% level of significance, no statistically significant difference was found between erythromycin-resistant and erythromycin-sensitive isolates.

Binomial test of proportion was used to observe the statistical difference between resistant and sensitive isolates. Among resistant and sensitive isolates of Amikacin, Ciprofloxacin, Clindamycin, Doxycycline, Fusidic Acid, Gentamicin, Oxacillin, Teicoplanin, Trimethoprim Sulfamethoxazole and Vancomycin; statistically significant difference was demonstrated. At the 5% level of significance, there was no statistically significant difference between the erythromycin isolates that were resistant and those that were sensitive.

DISCUSSION

In this study on 4153 samples, 64 (1.54%) were linezolid-resistant CoNS, predominantly affecting males (56.3%) and adults aged 18-59 years and over 59 years. Resistance was noted in 51.6% of blood and 48.4% of pus samples. The isolates were universally resistant to Teicoplanin (100%), highly resistant to Ciprofloxacin (92.2%), Oxacillin (89.1%), and Fusidic acid (84.4%), while showing higher sensitivity to Vancomycin (85.9%) and Doxycycline (79.7%). Statistically significant resistance differences were observed for all antibiotics tested except Erythromycin.

Many researchers have highlighted the growing significance of identifying CoNS as they are increasingly recognized as key pathogens in healthcare-associated infections. 11 The antibiotic-resistance profiles of CoNS isolates were examined in this study. Reports from clinical samples of hospitalized patients have indicated resistance to several critical antimicrobial agents, including linezolid. While over 98% of Staphylococcus strains remain susceptible to Linezolid, resistance has been documented in 1.4% of coagulase-negative Staphylococcus isolates (n = 73/5202). 12 A large multicenter study conducted in Greece from 2011 to 2013 showed a notable increase in linezolid resistance among CoNS, rising from 6.9% to 9%.13 In our study, 1.54% of CoNS isolates (n=64/4153) were resistant to Linezolid, which may reflect an increased resistance rate linked to the clinicians' preference for this antibiotic.

Recent studies have highlighted an increase in the isolation rates of CoNS from bloodstream infections. For instance, out of 581 CoNS isolates from a hospital in Pakistan, 311 (53.5%) were from blood samples, and 204 (35.1%) were from pus/swabs.14 Another study reported that bloodstream infections accounted for 51% to 78% of CoNS infections among newborns with very low birth weights.15 In our study, 51.6% of CoNS isolates were from blood, while 48.4% were from wounds.

A study conducted in Jordan assessed the prevalence and antimicrobial susceptibility of CoNS isolates from clinical specimens. According to CLSI 2009 criteria, CoNS isolates exhibited high sensitivity to Vancomycin, Linezolid, Rifampin, and Nitrofurantoin but showed significant resistance to Penicillin, Ampicillin, and other antibiotics. 16 Another study from India in 2019 reported that all Linezolid-resistant CoNS isolates were also resistant to Erythromycin, Cefoxitin, Clindamycin, and Trimethoprim/Sulfamethoxazole, with 80% resistant to Gentamicin and 90% resistant to Chloramphenicol and Ciprofloxacin. While all isolates were susceptible to Vancomycin (MIC range: 1-4 µg/ml), only one-third were susceptible to Teicoplanin. 17 In our study, linezolid-resistant strains were also resistant to Teicoplanin, and we observed high resistance to Ciprofloxacin, Fusidic Acid, Oxacillin, and Trimethoprim-sulfamethoxazole, with sensitivity noted for Vancomycin, Amikacin, and Doxycycline. The ongoing development of antibiotic resistance highlights the importance of selecting appropriate antimicrobial therapies.

A surveillance study in Italy from January 2016 to October 2018 evaluated the susceptibility of 828 CoNS isolates and found no resistance to Teicoplanin, Vancomycin, or linezolid. However, varying resistance levels were observed for other antibiotics, including Ampicillin (87%), Penicillin (86%), Oxacillin (70%), Erythromycin (69%), Ciprofloxacin (54%), Gentamicin (47%), Trimethoprim-Sulfamethoxazole (30%), and Clindamycin (28%).18 In our study, of the 4153 CoNS isolates analyzed, 64 (1.54%) were identified as Linezolid-resistant.

CONCLUSION

Our findings emphasize the critical need to monitor the development of Linezolid resistance in CoNS. Linezolid-resistant isolates have also demonstrated significant resistance to other key antimicrobial agents, such as Teicoplanin, Ciprofloxacin, and Oxacillin. It is essential to closely monitor Linezolid resistance, particularly when prolonged or frequent Linezolid therapy is required. This vigilance will help implement effective control measures and minimize the risk of resistant CoNS spreading within the community.

ACKNOWLEDGEMENT

We are grateful to friends and colleagues for their invaluable assistance throughout various stages of this study.

REFERENCES

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AUTHORS' CONTRIBUTIONS

Following authors have made substantial contributions to the manuscript as under:

 

AS: Study design, acquisition of data, critical review, approval of the final version to be published

NM: Acquisition and  analysis of data, drafting the manuscript, approval of the final version to be published

ZR: Concept and study design, analysis and interpretation of data, critical review, approval of the final version to be published

IY: Concept and study design, critical review, approval of the final version to be published

RH: Conception, drafting the manuscript, critical review, approval of the final version to be published

SJ: Acquisition of data, drafting the manuscript, critical review, approval of the final version to be published

 

Authors agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

 

CONFLICT OF INTEREST

Authors declared no conflict of interest, whether financial or otherwise, that could influence the integrity, objectivity, or validity of their research work.

 

GRANT SUPPORT AND FINANCIAL DISCLOSURE

Authors declared no specific grant for this research from any funding agency in the public, commercial or non-profit sectors

 

DATA SHARING STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request

 

https://mirrors.creativecommons.org/presskit/buttons/88x31/png/by.png This is an Open Access article distributed under the terms of the Creative Commons Attribution 4.0 International License.

 

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