The present study was a retrospective cross-sectional study which was conducted on the basis of the records of 200 pus swab samples from discharging ear submitted for culture and antibiotic susceptibility testing to the Department of Microbiology in the Government Medical College, Kannauj, Uttar Pradesh, India from January 2019 to October 2019 and the analysis of data was done in February 2020. Since the study was a retrospective study ethical clearance and consent was not required.

Sample size calculation: With 95% confidence level or power and taking margin of error as 5%, the sample size calculated was 121 samples, but to be on better side, 200 samples were included for which the power of the study was 100%.

The study participants were all clinically suspected cases of otitis media that provided pus swab from discharging ears at the Microbiology Department during the study period. Age, sex profiles, results of bacterial isolates and antimicrobial susceptibility testing of patients who had ear infection were retrieved from the Microbiology laboratory using a standard data recording format.

The samples were labeled with date, time, patient’s name, age and sex. Two pus samples were collected; the first swab was used for direct smear stain examination [Table/Fig-1] and the second swab for culture on 5% Sheep Blood Agar (SBA), Mac-conkey Agar (MA) and Chocolate Agar (CA). Different types of colonies in Nutrient Agar (NA) and MA is shown in [Table/Fig-2]. The SBA, MA were incubated at 37°C for 24 hours, whereas CA was incubated in candle jar generating 5% Carbon Dioxide (CO₂) at 37°C for 48 hours [10]. The isolates identification was done on the basis of standard microbiological techniques like microscopic stain examination, colony morphology and biochemical testing [10]. The antibiotic susceptibility testing was done in-vitro on Muller Hinton agar using Kirby Bauer disc diffusion method using 0.5 McFarland culture suspension of isolate as per Clinical and Laboratory standard Institute (CLSI) recommendation [11]. The antibiotics used for testing were ampicillin (10 mcg), amikacin (30 mcg), amoxycillin/clavulanic acid (20/10 mcg), ceftriaxone (30 mcg), gentamicin (10 mcg), levofloxacin (5 mcg), meropenem (10 mcg) and vancomycin (30 mcg).

A standard procedure was followed to maintain the quality control in the laboratory test results. American Type Culture Collection (ATCC) control strains of Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853) were used for the quality control purpose [11,12].

The result were expressed as percentage and proportion for distribution of CSOM cases according to age and gender using Microsoft Excel version 2007.

A total of 200 samples were studied, of which 147 (73.5%) samples showed growth and 53 (26.5%) samples were sterile. Out of 147, 136 were pure aerobic growth and 11 samples showed mixed growth [Table/Fig-3,4]. Out of the 200 cases studied, 114 (57%) were males and 86 (43%) were females. Among the 147 cases showing microbial growth, 82 (55.8%) were males and 65 (44.2%) were females. The maximum numbers of patients were in age group 0-10 years followed by age group 11-20 years [Table/Fig-5].

[Table/Fig-1]:

Direct gram stain of ear swabs of patients with CSOM showing gram positive cocci (X1000).

[Table/Fig-2]:

a) Nutrient agar culture plate showing colonies of Staphylococcus growth; b) Nutrient agar culture plate showing colonies of Pseudomonas growth; c) Mac-conkey agar culture plate showing colonies of Klebsiella species.

In this study, it was found that unilateral infection (69%) was more common than bilateral infection (31%) and the disease was more prevalent in younger age group and among rural population (71%).

Out of the 158 (136 samples showed pure aerobic monomicrobial growth, 11 samples showed aerobic polymicrobial growth with each having two different microbial growth) bacterial isolates, 110 were gram negative bacteria, of which Pseudomonas aeruginosa 67 (42.40%) was the most predominant followed by Klebsiella spp. 23 (14.55%) and Escherichia coli 15 (9.49%). Among the gram-positive bacteria, Staphylococcus aureus was isolated in 48 (30.37%) cases [Table/Fig-6].

Pseudomonas aeruginosa was the predominant organism isolated in this study, which was highly sensitive to meropenem and aminoglycoside and least sensitive to fluoroquinolone group of drugs. Staphylococcus aureus isolates were 100% sensitive to vancomycin followed by aminoglycosides and fluoroquinolone. Gram negative isolates showed high degree sensitivity to meropenem and aminoglycosides. They showed least sensitivity to penicillin, amoxicillin-clavulanic acid and cephalosporins [Table/Fig-7].

Otitis media is a worldwide disease with higher prevalence in developing countries, particularly among lower socio-economic section of the society [4]. In this study, males were more affected as compared to females (M:F=1.26:1), which is similar to the study done by Falagas ME et al., [13]. In present study, patients in the age group 0-10 years had high prevalence of CSOM. Other studies that have also shown the similar results [14,15]. This may be explained by short eustachian tube in small children as the infected material from nose and sinuses may trickle along the eustachian tube to the tympanic cavity [16]. In this study, 110 gram negative and 48 gram positive organisms were isolated. The frequency of bacterial isolates were Pseudomonas aeruginosa 67 (42.40%), Staphylococcus aureus 48 (30.37%), Klebsiella species 23 (14.55%). [Table/Fig-8] shows comparative chart of various studies done in the past [17-22]. The difference in the isolated organism may be due to geographical variation.

In the present study, gentamicin (85%) and amikacin (83%) had high level of antibacterial activity on all the isolates. In Worku M and Bekele M study, amikacin (90%) and gentamicin (89%) had shown good results to both gram positive and gram negative isolates [23]. The most effective drug against Pseudomonas aeruginosa was meropenem (91%), followed by gentamicin (81%) and amikacin (78%). In a study done by Harshika YK et al., they found Pseudomonas aeruginosa isolates susceptible to imipenem and amikacin (93.6%) followed by gentamicin (89.3%) [24]. Staphylococcus aureus isolates were found to be maximum sensitive to vancomycin (100%) followed by good sensitivity to aminoglycosides (93%) and levofloxacin (88%). Pseudomonas species invade middle ear through defect in the tympanic membrane from an acute episode of otitis media. The high isolation of Staphylococcus aureus in CSOM cases can be due to its ubiquitous nature and high colonisation rate in external auditory canal and upper respiratory tract.

Otitis media is an important cause of hearing impairment and is a serious issue in pediatric age group. If it is not managed well in time it could led to serious effects later in the life in language development, communication skill and educational process. In current scenario, microbial profile and their antimicrobial susceptibility pattern in CSOM cases changes with time to time due to geographical and environmental variations in study population [18]. Along with that many people self medicate themselves with antibiotics which change their microbial flora and susceptibility to antibiotics. This has led to emergence of resistance in common microbes to the basic antibiotics and shifting treatment towards higher antibiotics, making the treatment more expensive and unaffordable. So, it is the urgency in today’s time to monitor the changing trends in microbial profile and their susceptible pattern to control the indiscriminate use of antibiotics, ultimately controlling the emergence of antimicrobial resistance. Poor hygiene, pond/river bath, homemade therapy for treatment i.e use of oil, honey and unconventional ear drops should also be avoided.

In the present study, 53 samples were culture sterile, which may be due to either fungal infection or some anaerobic aetiology. Further studies should be done to overcome these limitations.

In Chronic Suppurative Otitis Media (CSOM), gram negative isolates were higher compared to gram positive isolates. Pseudomonas aeruginosa was the most predominant organism followed by Staphylococcus aureus. Thus, this study concludes that CSOM remains an ongoing problem. Time to time evaluation of microbial pattern and antibiotic susceptibility of isolates is necessary for rational antibiotic usage and successful treatment. Proper selection of topical antibiotics on the basis of culture and sensitivity with keeping the ear dry is the main modality of treatment for CSOM. The patient should be advised to take complete course of antibiotics which will prevent complications as well as emergence of drug resistance.