Keywords: Respiratory tract infections; Drug resistance; Microbial; Antimicrobial stewardship; Enterobacteriaceae; Gram positive bacteria; Pneumonia

Introduction

Respiratory tract infections (RTI) represent the highest burden of infectious diseases in the world, accounting for a substantial proportion of morbidity and mortality especially in developing nations [1-4]. RTI contribute to over 500 million cases and about 50 million annual deaths accounting to 13.4% of all disability adjusted life years across the globe [5-8]. The incidence of RTI and associated mortality/morbidity vary on several factors such as age, geographical location, seasons, local antimicrobial prescription practices and the prevailing antimicrobial resistance (AMR) patterns [9-14].

RTI affecting the upper and lower respiratory tract are caused by a diverse group of pathogens, including bacteria, viruses, fungi, and parasites [15,16]. The scary six bacteria (Klebsiella pneumoniae, Escherichia coli, Streptococcus pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Staphylococcus aureus) are responsible for the majority of deaths in pneumonia. Majority of these organisms are multi drug resistant or extensively drug-resistant. RTIs are the most common cause of antibiotic abuse leading to anti microbial resistance (AMR) and also contribute to majority of deaths caused by multi drug resistant organisms [4,17].

India, as a developing nation, having the world’s largest population and not so well regulated antibiotic practices is uniquely placed in the global AMR campaign [18]. Indian Council of Medical Research (ICMR) has initiated nationwide “Antimicrobial Resistance Surveillance and Research Network” (AMRSN) [19,20]. After successfully consolidating anti-microbial stewardship program (AMSP) at various tertiary care centres, AMRSN has envisaged extending to secondary care centres across India as a hub-spoke model. The state of Madhya Pradesh in India has developed state action plan for containment of antimicrobial resistance (MPSAPCAR) in 2019 on the guidelines of National Action Plan on AMR (NAP AMR).

The present study was conceived to identify the pattern of antibiogram for RTIs in secondary care hospitals (district hospitals / nursing homes) at central India with support from ICMRAMRSN and MPSAPCAR.

Methodology

Study Setting and Ethical Clearance: This prospective longitudinal observational chart review study was conducted in the state of undivided Madhya Pradesh in central India.  Among the nominated 10 secondary care centres, two hospitals were government district level hospitals and the remaining eight study-sites were private nursing homes, located in urban/semi-urban areas. These sites were chosen based on the availability of in-house accredited microbiology laboratory and a full-time microbiologist. The location of these cities is presented in Figure 1.

 Figure 1: AIIMS Bhopal ICMR-AMRSN initiated network.

The study was carried out as part of ICMR-AMRSN, with Institute Human Ethics Committee (IHEC) approval Letter No. LOP/2020/EF0157 dated February 24, 2020. As the study was only observational and chart review without any patient identifier, hence waiver of consent was granted by IHEC. The present data set was collected from 1st April 2022 to 30 September 2022. The study procedure was in accordance with the principles of the Declaration of Helsinki. Training protocols provided by ICMR-AMRSN were customised at All India Institute of Medical Sciences, Bhopal, for these secondary care centres.

Sample size and sampling: Formal sample size was not calculated. Consecutive and feasible sampling was adopted. Antibiograms were generated only for those organisms with a cumulative frequency of more than 30 samples.

Sample Collection:  Sputum (spontaneous or induced) samples and endo-tracheal (ET) tube aspirates from symptomatic patients (fever and/or cough and/or clinical chest sign and infiltration in X-ray) were sent for aerobic culture as per CLSI guidelines. Routine nasopharyngeal swabs and routine endo-tracheal tube aspirates from asymptomatic patients were not taken into consideration. The collected specimens were promptly transported to the laboratory as soon as possible (preferably within 1 hour).

Microbiological isolation and reporting: Clinical data were collected by the nursing officers, and the microbiological data were collected by the laboratory technician and verified by the microbiologist. Prior to processing for culture, the sputum specimens were checked for appropriateness of collection by Murray & Washingtoncriteria. Appropriate sputum samples were cultured on Chocolate agar, Sheep blood agar and MacConkey agar and incubated at 35±2ºC under aerobic conditions with 5-10% CO₂ for overnight incubation. Plates were examined each day for up to 72 hours for colonies of interest. The identification of colonies of interest was based on their cultural and morphological characteristics followed by conventional biochemical tests and susceptibility testing by Kirby-Bauer disk-diffusion method. Results of antimicrobial susceptibility were interpreted as per CLSI-M100.

Antimicrobial Resistance Patterns: Third-generation cephalosporin susceptibility for the Enterobacteriaceae family was reported when susceptible to ceftriaxone and for Pseudomonas using ceftazidime. 3rd generation cephalosporin resistance (3rd GCR) was calculated by 100 minus the susceptibility percentage of ceftriaxone/ceftazidime. Carbapenem resistance was calculated by 100 minus the susceptibility percentage of meropenem. Methicillin resistance was calculated by 100 minus the susceptibility percentage of oxacillin.

Data Analysis: Cleaned data were entered in a spreadsheet, and the data were summarized as frequencies and percentage up to one decimal value.

Result

Out of the 1315 respiratory samples, 651 (49.5%) showed significant pathological growth. Among the positive cultures (n=651), 626 (96.1%) showed bacterial growth, and 25 (3.8%) were Candida spp. Among the bacterial growth (n=626), 590 (94.2%) were Gram-negative bacteria, and 36 (5.7%) were Grampositive bacteria.

Among the Gram-negative isolates (n=590), the predominant isolate was Klebsiella pneumoniae (266, 45%) followed by Pseudomonas aeruginosa (143, 24.2%) and Acinetobacter spp (91, 15.4%). The prevalence of other isolates is given in detail in Table-1. The susceptibility patterns of the identified pathogens to different antibiotics were analysed and provided below.

Table 1: Spectrum of culture positive isolates.

Klebsiella pneumoniae: Resistance to 3rd generation cephalosporin was noticed in 81.1% cases, piperacillin resistance was in 56.6% cases, carbapenem resistance in 37.2% and colistin resistance in 7.9%. The detailed antibiogram of Klebsiella pneumoniae is provided in Figure-2A.

  Escherichia coli: Resistance to 3rd generation cephalosporin was noticed in 84.6% cases, piperacillin resistance in 61.3% cases, carbapenem resistance in 45.6 % and colistin resistance in 2.9 % of cases. The detailed antibiogram of Escherichia coli is provided in Figure- 2C.

Acinetobacter spp: Resistance to piperacillin was noticed in 83.5% cases, carbapenem resistance in 79.1% and colistin resistance in 6.6% of cases. The detailed antibiogram of Acinetobacter spp isolates is provided in Figure-2B.

Pseudomonas aeruginosa: Resistance to 3^rd generation cephalosporins was observed in 46.6% of cases, piperacillin resistance in 38.2% cases, carbapenem resistance in 35.7% and colistin resistance in 9.8% of cases. The detailed antibiogram of Pseudomonas aeruginosa is provided in Figure-2(D).

Staphylococcus aureus: Vancomycin resistance was seen in 9.7% of cases and linezolid resistance in 16.7% of cases. The detailed antibiogram of Staphylococcus aureus isolates is provided in Figure-2(E).

Discussion

Approximately 1.2 million individuals succumb to AMR annually, and an estimated 10 million more may face the same fate by the year 2050 [4,21]. According to economic predictions, the global economy may experience a decline of 2-3.5% in gross domestic product (GDP) by 2050, and a drop of 3-8% in livestock as a result of AMR, with the potential cost of USD 100 trillion [17]. Even after an extensive literature review, we found scanty literature that analysed the prevalence of AMR among primary and secondary care centres of respiratory pathogens in India. Most of these studies focus mainly in intensive care units of tertiary care centres. A comparative analysis of observed global AMR patterns in non-tertiary care centres is presented below in Table-2. This study serves as a projection as to how far the menace of AMR has percolated down.