Abstract

Background and objectives: Coagulase-negative staphylococci (CoNS) are considered important causative agents of hospital acquired infection. These organisms are found in various clinical specimens from hospitalized patients. Present study was carried out to determine the species distribution and antimicrobial susceptibility pattern of CoNS isolated from clinical specimens at a tertiary care hospital.

Materials and methods: CoNS isolated from various clinical samples were included in this study. The isolates were identified by colony morphology, Gram’s staining, catalase and coagulase tests. Further differentiation of species was performed by susceptibility to novobiocin, urease activity and ornithine decarboxylase test. Antibiotic susceptibility testing was performed according to the Clinical and Laboratory Standard Institute (CLSI) guidelines.

Results: Total 108 isolates of CoNS were included and analysed. Out of 108 CoNS, S. epidermidis was the most common species (36.1%) followed by S. saprophyticus (23.1%), S. hemolyticus  (17.6%), S. hominis (13%) and S. lugdunensis (10.2%). Most of the isolates showed resistance to penicillin, oxacillin, amoxycillin, erythromycin, ciprofloxacin and ofloxacin. All the isolates were sensitive to vancomycin.

Conclusions: CoNS emerged as an important nosocomial pathogen and should not be neglected as contaminant. High rate of antimicrobial resistance warrants susceptibility testing prior to the treatment of CoNS.

IMC J Med Sci. 2024; 18(1):010. DOI: https://doi.org/10.55010/imcjms.18.010

*Correspondence: Sabiha S Tamboli, Department of Microbiology, Parbhani Medical College and RP Hospital Research Institute, Pathri Road, Parbhani, Maharashtra, India. Email: [email protected]

Introduction

Coagulase-negative staphylococci (CoNS) are part of normal flora of skin [1]. Previously, they were considered non-pathogenic with low virulence. But since 1950, cases of CoNS associated infections have been reported with increased frequencies. The predisposing factors for CoNS infections are patients with catheter, prosthetic implants, dialysis, oncological diseases, compromised immunity and neonatal state [2]. CoNS survive on synthetic medical devices and equipment such as intravenous catheters, prosthetic heart valves and various implants [3]. Currently more than fifty different CoNS species have been described. Out of this, S. epidermidis, S. saprophyticus, S. hemolyticus, S. hominis and S. lugdunensis have higher clinical significance [4].

The main challenge in the diagnosis is to correctly identify the cases in which CoNS are causative agents for infection rather than contaminants. This leads to under treatment (i.e., delayed or withheld antibiotics) and thereby contributing to increased morbidity and mortality [5]. Due to increasing clinical significance of CoNS infection, accurate species identification and determination of antibiotic resistance are of paramount importance to treat CoNS infections. The aim of the study was to investigate species distribution and antimicrobial resistance pattern of CoNS isolated from clinical specimens at a tertiary care centre.

Material and method

The study was conducted at the Department of Microbiology in a tertiary care teaching hospital of Marathwada region of Maharashtra state, India. Study was done over a period of one year from January 2015 to December 2015. The study was approved by the institutional ethical committee.

CoNS isolated from different clinical samples such as pus, urine, blood, sputum, vaginal swab, wound swab, suction tip, pleural fluid and nasal swab were included. Same strain of CoNS isolated twice in pure culture from an infected site or body fluid was considered clinically significant. Samples were cultured on nutrient and blood agar plates for bacterial isolation. Plates were incubated aerobically overnight at 37°C for 48 hours [6]. Isolates were identified by colony morphology, Gram’s stain, catalase and coagulase tests. Bacitracin susceptibility was performed to exclude micrococci and Stomatococcus species [7]. The speciation of CoNS was done by ornithine decarboxylase, urease, mannose fermentation and novobiocin (5 µg) sensitivity tests [8,9].

The antimicrobial susceptibility of all the isolates was performed by Kirby-Bauer disc diffusion method using Muller-Hinton agar plates as per the recommendation of Clinical Laboratory Standard Institute guidelines [10]. S. aureus ATCC 25923 was used as a standard control strain for antimicrobial susceptibility testing.

Antimicrobial discs used were penicillin (10 µg), amoxicillin-clavulanic acid (20/10 µg), oxacillin (1 µg), erythromycin (15 µg), linezolid (30 µg), gentamicin (10 µg), and vancomycin (30 µg). The various antibiotic discs used were purchased from HiMedia Laboratories Private Limited, India. 

Results

A total of 108 CoNS isolates were included in the study. Out of 108 isolates, 65 (60.2%) and 43 (39.8%) were from male and female patients respectively. Maximum numbers of isolates (44.4%) were from age group 21-40 years followed by 18.5% from age group 41-60 years [Table-1].

Table-1: Age and sex distribution (n=108)

Majority of CoNS were from pus sample (44.4%) followed by urine (23.1%), blood (9.3%), suction tip (6.5%), sputum (4.6%), vaginal swab (3.7%) and, 2.8% each from wound swab, nasal swab and pleural fluid [Table-2]. The commonest CoNS species isolated was S. epidermidis 39 (36.1%) followed by S. saprophyticus 25 (23.1%), S. hemolyticus 19 (17.6%), S. hominis 14 (13%) and S. lugdunensis 11 (10.2%) [Table-2]. Antimicrobial sensitivity test of the isolates showed maximum resistance to oxacillin (88%), ciprofloxacin (80%), penicillin (78.7%), amoxicillin-clavulanic acid (75%), erythromycin (60.2%) and  ofloxacin (60.2%). None of the CoNS species showed resistance to vancomycin while only 9.3% were resistant to linezolid [Table-3].

Table-2: Distribution of CoNS and their species in different clinical samples (n=108)

Table-3: Species wise antibiotic resistance pattern of isolated CoNS

Discussion

CoNS formerly considered as contaminant bacteria have now emerged as a major cause of nosocomial infections. CoNS are the common agents of nosocomial bloodstream infections as well as other type of infections. Factors helpful in identification of true infections by CoNS include repeated isolation of same strain of CoNS in pure culture from infected site or specimen over the course of an infection plus presence of clinical evidence of infection [11,12]. Recent studies have shown that CoNS are one of the important causative agents of human infection, especially in immunocompromised patients, premature newborns and patients with indwelling medical devices [13]. In our study, majority of CoNS were isolated from male patients (60.2%). Similar findings are also reported by Usha et al and Asangi et al [14,15]. On the other hand, Goudarzi M et al found maximum number of CoNS from female patients [16].

In this study, out of 108 isolates, most of the isolates were from pus (44.4%) and urine (23.2%). The results differ from studies by Bhatt P et al and Parashar [17,18]. In their studies maximum numbers of CoNS were isolated from blood samples. This difference could be due to types of patients and hospital settings.

In the laboratory, staphylococci other than S. aureus are reported as CoNS without speciation. As various species of CoNS are associated with different diseases, CoNS should be identified to the species level by simple, reliable and inexpensive method [19]. In the present study we have identified CoNS species by slide and tube coagulase, ornithine decarboxylase, urease, mannose fermentation and novobiocin sensitivity tests. These tests are inexpensive and affordable and can be practiced in most of the diagnostic laboratories.

In the present study, S. epidermidis constituted the predominant species (36.1%) followed by S. saprophyticus, S. hemolyticus , S. hominis and S. lugdunensis. This is in concurrence with other reported studies from India and adjoining region [20-22]. Those studies have reported S. epidermidis as the most common species (41% - 46.84%) among the isolated CoNS. The second most common species in our study was S. saprophyticus which is similar to other studies who also found S. saprophyticus as second most common species [7,23].

In the present study, majority of the isolates showed resistance to oxacillin, ciprofloxacin, amoxicillin-clavulanic acid, penicillin, erythromycin and ofloxacin. So, these antibiotics could not be recommended for empiric treatment of CoNS. All the isolates in our study were sensitive to vancomycin and only 9.3% were resistant to linezolid. Similar results were also observed by others [24-26]. Vancomycin and linezolid are the most effective drugs in treating infections caused by CoNS species.

Prolonged hospital stays, widespread antibiotic use and the ability of CoNS to form multi-layered bio-films on artificial surfaces are the potential causes of high resistance rate to multiple antimicrobial agents especially for the species that are isolated from catheter tips and blood cultures. This high resistance to multiple antimicrobial agents poses a significant challenge in the clinical management of infections caused by CoNS.

Therefore, continued surveillance, prudent use of antibiotics and emphasis on infection prevention measures in hospitals are imperative in curbing the rise of antibiotic resistant CoNS strains. In addition to increased vigilance, advanced diagnostic approaches and an understanding of the antibiogram profiles are essential for effective clinical management and the prevention of CoNS associated infections in healthcare settings.

Conflict of interest:

The author declares no conflict of interest.

Funding: None

References

1.     Hall KK, Lyman JA. Updated review of blood culture contamination.ClinMicrobiol Rev. 2006; 19(4): 788–802. doi:10.1128/CMR.00062-05.

2.     Michalik M, Samet A, Podbielska-Kubera A, Savini V, Międzobrodzki J, Kosecka-Strojek M. Coagulase negative Staphylococci (CoNS) as a significant etiological factor of laryngological infections: a review. Ann Clin Microbiol Antimicrob. 2020; 19(26). doi: 10.1186/s12941-020-00367-x.

3.     Sharma V, Jindal N, Devi P. Prevalence of methicillin resistant coagulase negative staphylococci in a tertiary care hospital. Iran J Microbiol.2010; 2(4): 185-188.

4.     Becker K, Both A, Weißelberg S, Heilmann C, Rohde H. Emergence of coagulase-negative staphylococci. Expert Rev Anti Infect Ther. 2020; 18(4): 349–366. doi: 10.1080/14787210.2020.1730813.

5.     Taylor SP, Anderson WE, Beam K, Taylor B, Ellerman J, Kowalkowski MA. The association between antibiotic delay intervals and hospital mortality among patients treated in the emergency department for suspected sepsis. Crit Care Med. 2021; 49(5): 741-747. doi: 10.1186/s13054-021-03736-w.

6.     Winn WS, Allen SD, Janda WM, Koneman EW, Procop GW, Schreckenberger PC, et al. Charts. In: Darcy P, Peterson N, Montalbano J, Editors. Koneman’s Color Atlas and Textbook of Diagnostic Microbiology. 6^th Eds. Philadelphia: Lippincott; 1997; 1442-1535. doi: 10.1097/01.prs.0000358868.74684.60.

7.     Singh S, Banerjee G, Agarwal SK, Kumar M, Singh RK. Simple method for speciation of clinically significant coagulase negative staphylococci and its antibiotic sensitivity/ resistant pattern in NICU of tertiary care centre. Biomed Res. 2008; 19(2): 97-101.

8.     De Paulis AN, Predari SC, Chazarreta CD, Santoianni JE. Five test simple scheme for the species level identification of clinically significant coagulase negative staphylococci. J Clin Microbiol. 2003; 41(3): 1219-24. doi: 10.1128/JCM.41.3.1219-1224.2003.

9.     Baird D. Staphylococcus: cluster forming Gram-positive cocci. In: Colle JG, Fraser AG, Marimom BP, Simmons A, editors. Mackie and McCartney Practical Medical Microbiology. 14th eds. New York: Churchill Living Stone; 1996; 245-261.

10.  Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Fourth Informational Supplement. CLSI M100-S24. Wayne, PA: Clinical and Laboratory Standards Institute; 2014.

11.  Kloos WE, Bannerman TL. Update on clinical significance of coagulase-negative staphylococci. Clin Microbiol Rev. 1994; 7(1): 117–140. doi: 10.1128/CMR.7.1.117.

12.  Beekmann SE, Diekema DJ, Doern GV. Determining the clinical significance of coagulase-negative staphylococci isolated from blood cultures. Infect Control Hosp Epidemiol. 2005; 26(6): 559–566. doi: 10.1086/502584.

13.  Nzeako BC, Al Rusheidi S, Neilson F, Al-Balkhair A. Types of bacteria on some medical devices used in Sultan Qaboos University hospital wards. Middle-East J SciRes. 2010; 5(6): 449– 53.

14.  Usha MG, Shwetha DC, Vishwanath G. Speciation of coagulase negative Staphylococcal isolates from clinically significant specimens and their antibiogram. Indian J Pathol Microbiol. 2013; 56(3): 258-260. doi:10.4103/0377-4929.120383.

15.  Asangi SY, Mariraj J, Sathyanarayan MS, Nagabhushan R. Speciation of clinically significant coagulase negative staphylococci and their antibiotic resistant pattern in a tertiary care hospital. Int J Biol Med Res. 2011; 2(3): 735-9.

16.  Goudarzi M, Seyedjavadi SS, Goudarzi H, Boromandi S, Ghazi M, Azad M, et al. Characterization of coagulase-negative staphylococci isolated from hospitalized patients in Tehran, Iran. J Paramedical Sci. 2014; 5(2): 44–50. https://doi.org/10.22037/jps.v5i2.5841.

17.  Bhatt P, Tandel K, Singh A, Mugunthan M, Grover N, Sahni AK. Species distribution and antimicrobial resistance pattern of coagulase-negative staphylococci at a tertiary care centre. Med J Armed Forces India. 2016; 72(1): 71-74. doi: 10.1016/j.mjafi.2014.12.007.

18.  Parashar S. Significan ce of coagulase negative staphylococci with special reference to species differentiation and antibiogram. Ind Med Gaz. 2014; CXLVII(7): 255-258.

19.  Ieven M, Verhoeven J, Pattyn SR, Goossens H. Rapid and economical method for species identification of clinically significant coagulase-negative staphylococci. J Clin Microbiol.1995; 33(5): 1060-3. doi: 10.1128/jcm.33.5.1060-1063.1995.

20.  Kavitha Y, Shaik KM. Speciation and antibiogram of clinically significant coagulase negative staphylococci. Int J Health Sci Res. 2014; 4(12): 157-161.

21.  Goyal R, Singh NP, Kumar A, Kaur I, Singh M, Sunita N, et al. Simple and economical method for speciation and resistotyping of clinically significant coagulase negative staphylococci. Indian J Med Microbiol. 2006; 24(3): 201-4. https://doi.org/10.1016/S0255-0857(21)02350-1.

22.  Sheikh AF, Mehdinejad M. Identification and determination of coagulase-negative staphylococci species and antimicrobial susceptibility pattern of isolates from clinical specimens. Afr J Microbiol Res. 2012; 6(8): 1669–1674. doi: 10.5897/AJMR11.076.

23.  Singh S, Banerjee G, Agarwal SK, Rajput A, Tripathi P, Kumar M, et al. Prevalence of MecA gene positive coagulase negative staphylococci in NICU of a tertiary care hospital. Biomed Res. 2009; 20(2): 94-8.

24.  Al Tayyar IA, Al-Zoubi MS, Hussein E, Khudairat S, Sarosiekf K. Prevalence and antimicrobial susceptibility pattern of coagulase negative staphylococci (CoNS) isolated from clinical specimens in Northern of Jordan. Iran J Microbiol. 2015; 7(6): 294–301.

25.  Asaad AM, Qureshi MA, Hasan SM. Clinical significance of coagulase-negative staphylococci isolates from nosocomial bloodstream infections. Infect Dis (Lond). 2016; 48(5): 356–360. doi: 10.3109/23744235.2015.1122833.

26.  Pedroso SHSP, Sandes SHC, Filho RAT, Nunes AC, Serufo JC, Farias LM, et al. Coagulase-negative staphylococci isolated from human bloodstream infections showed multidrug resistance profile. Microb Drug Resist. 2018; 24(5): 635–47. doi: 10.1089/mdr.2017.0309.

Cite this article as:

Tamboli SS, Tamboli SB, Species distribution and antimicrobial susceptibility pattern of coagulase-negative staphylococci isolated from clinical specimens at a tertiary care hospital. IMC J Med Sci. 2024; 18(1):010. DOI: https://doi.org/10.55010/imcjms.18.010