ICUs are associated with considerable morbidity, mortality, and cost worldwide. Infection and related sepsis are the leading cause of death in non-cardiac ICUs, with mortality rate that can reach up to 60% and account for approximately 40% of the total ICU expenditure [1].

Even though ICU represents only 5% of total hospital beds, they account for almost 20-25% of all health care associated infections.2 The rate of ICU infections worldwide is 23.7 infections per 1,000 patient days, with the rate of nosocomial infections being 5-30% amongst the ICU patients [3-4].

In the ICUs there is increased use of invasive devices such as peripheral vascular line and central venous line catheters which can be linked to blood stream infections and sepsis. The blood stream infections in ICUs are also secondary to other common ICU infections such as respiratory infections and urinary tract infections [5-6].

The pattern of clinical specimens, their positivity and isolated organisms pattern varies widely one country to the other, one hospital to another and even amongst different ICUs of the same hospital [3-4].

The present study attempts to evaluate the pattern of various clinical specimens from the patients admitted in intensive care units of a tertiary care hospital, their positivity and isolated organisms 

Aims and Objectives

To evaluate the pattern of various clinical specimens from the patients admitted in intensive care units of a tertiary care hospital, their positivity and isolated organisms

• Study Setting: The present study was conducted in the department of Microbiology, Indira Gandhi Medical College and Hospital Shimla

• Study Design: It was a descriptive cross sectional study

• Study Period: The study period was of one year from 14^th January 2019 to 13^th January 2020

• Selection of Cases: In this study all clinical samples received in the department of Microbiology for bacterial culture and susceptibility testing from patients admitted in the intensive care units of this institution during the study period

Samples received in the department of Microbiology and processed using standard procedures were of Bloodm,Sterile Body fluids (cerebrospinal fluid & pleural fluid), Pus samples, Sputum,Swabs (throat and wound) ,Respiratory secretions,Urine and Central line catheter tip.

The demographic profile of the patient whose samples were included in the study was also recorded.

Processing of Samples

The samples of blood and other sterile body fluids received in BACTEC blood culture vials were first cultured using the BD BACTEC™ FX system for five days. The positive blood culture vials were further processed within 8hrs. For obtaining pure growth for further processing, the positive blood culture vials were inoculated on blood agar and MacConkey agar plates and incubated aerobically at 37^oC overnight for 18-24 hrs. The positive cultures detected in the BD BACTEC™ FX system were subjected to identification and antibiotic susceptibility testing by BD Phoenix™ FX Automated Microbiology system. Negative report was given only after 5days of incubation in BD BACTEC ™ FX automated system.

Body fluid such as pus, sputum, respiratory secretions, pleural fluid were received in sterile containers, inoculated onto blood agar and MacConkey agar plates, and were incubated at 37^oC for 24 hrs. Cerebrospinal fluid was received in sterile container and inoculated on blood agar and chocolate agar plates and incubated at 37⁰ C for 24 hrs.Uncentrifuged urine but well-mixed urine was inoculated by a calibrated loop onto plates of cystine lactose electrolyte deficient (CLED) agar. Throat swab was inoculated onto blood agar plates and incubated at 37⁰C for 24 hrs.

Morphology and Characteristics

The pure growth obtained after 24 hours of incubation on solid media was identified on the basis of colony characters, gram staining and preliminary biochemical reactions. When pure growth is not obtained on primary plating then it was subcultured on blood agar and MacConkey agar to get pure growth.

Preliminary Identification of Bacteria

• Colony Characters: The growth of the organism was observed on Blood Agar and MacConkey Agar. The characters of the colony were studied like size, shape, surface, edge, elevation, consistency, colour of the colony, odour, and effect on media

• Gram Staining: Gram staining of the growth was done using modified Huckers method. Based on the gram staining of organism whether gram positive or gram negative, the panels for antimicrobial sensitivity were selected

• Preliminary Biochemical Reactions: Preliminary biochemical reactions such as catalase, coagulase and oxidase were performed to identify the organism

The isolates were then processed in BD Phoenix™ FX automated system for further identification. The final report indicating organism identified was communicated to respective ICUs and also recorded in the performa.

Quality Control

The quality check was performed using standard strains as per the manufacturer’s instructions.

Exclusion Criteria

The samples where BD Phoenix™ FX automated system gave inconclusive results or failed to identify the isolates were excluded from the study.

Statistical Analysis

The Data collected was entered in Excel spreadsheet and accuracy of data entered was checked by cross-verification of the data entered. Categorical variables were expressed as frequencies and percentages. 

In the present study clinical samples were received from general ICU, neonatal ICU, paediatric ICU, HDU, respiratory ICU, sick neonatal care unit and CTVS-ICU. The bacterial isolates were studied in detail in the present study and Candida spp. was also documented. In the present study 2458 samples were processed out of which 766(31.16%) were found to be culture positive, sterile were 1463(59.52%) and contaminants were found in 229(9.32%). The number of isolates in all culture positive specimens was 840, as more than one isolates were also obtained in some specimens Figure 1.

Figure 1: Culture Positivity among clinical samples

Among all isolates gram negative bacteria 477 56.78% were most common, followed by Candida spp. 266 31.66% and gram positive bacteria 97 11.54% Figure 2.

Figure 2: Pattern of Organisms Isolated in All Clinical Specimens

Table 1 shows the distribution of various clinical specimens. Maximum number of samples obtained was of blood (52.31%).Most common culture positive clinical specimen in the present study was pus (63.49%) followed by endotracheal secretions (59.31%).

Table 1: Distribution of Various Clinical Specimens

Gram negative bacteria (91.96%) were predominant in endotracheal secretions, gram positive bacteria (100%) predominated in CSF, and Candida (53.58%) was predominant in blood samples Table 2. 

Table 2: Pattern of Organisms Isolated in Different Clinical Specimens

The most common bacteria isolated in the present study were Klebsiella pneumoniae followed by Non fermenter group of organism. Klebsiella pneumoniae was also found to be the most common isolate from blood, endotracheal secretions, pus and drain whereas in urine Enterococcus faecium and Escherichia coli were found to be predominant isolates (Table 3).

Table 3: Pattern of Bacteria Isolated from Different Clinical Specimen 

In the present study various types of clinical samples were received from different ICUs. Maximum number of samples obtained was of blood (52.31%). Several studies documented similar findings [5-7]. The reason for more blood samples in this study can probably be explained by the fact that in the ICU there is increased use of invasive devices such as peripheral vascular line and central venous line catheters which can be linked to blood stream infections and sepsis. The blood stream infections in ICUs are also secondary to other common ICU infections such as respiratory infections and urinary tract infections. 

Most common culture positive clinical specimen in the present study was pus (63.49%) followed by endotracheal secretions (59.31%). The results are in acccordance to the several other studies done in the recent past [8-10]. Unlike the present study maximum culture positivity was reported from sputum (83.6%) and urine (51.1%) by Pattanayak C. et al. and Sarvepalli AK.et al. [11-12].

The most common bacteria isolated in the present study were Klebsiella pneumoniae followed by Non fermenter group of organism. Klebsiella pneumoniae was also found to be the most common isolate from blood, endotracheal secretions, pus and drain. Whereas in urine Enterococcus faecium and Escherichia coli were found to be predominant isolates. Klebsiella pneumoniae was also found to be predominant isolate from ICU infections in several other studies [13-15]. Unlike the present study Pradhan et al. found Acinetobacter baumannii as predominant pathogen from endotracheal secretions and Pseudomonas spp. from urine. Richards M. et al. found CONS as predominant isolate from blood, Pseudomonas aeruginosa from respiratory secretions, Escherichia coli from urine and Pseudomonas aeruginosa from pus. Jain et al., found Citrobacter koseri from blood, Escherichia coli from urine, Pseudomonas aeruginosa from sputum and Staphylococcus aureus from pus [8,16]. Klebsiella pneumoniae was found to be predominant isolate in this study this can probably be explained by the fact that the members of Enterobacteriaceae are found as normal commensal flora of human body, which make them one of the most important endogenous bacteria causing infections in immune-compromised patients. Increased colonization of Enterobacteriaceae organisms in the respiratory tract of patients on prolonged mechanical ventilation also contributes to higher infections due to these bacterias in ICUs [17]. 

Non fermenter gram negative bacteria have emerged as important pathogen in the hospitals particularly in ICUs, probably due to wide spread and injudicious use of antibiotics. These bacteria are normally inhabitants of water and soil. In the hospital environment they may be isolated from instruments, hospital linen and also from skin. Most of these are MDR pathogen. Hence pose a challenge to the treating physician. In the present study Non fermenter gram negative bacteria were next common group isolated from several clinical specimens. They were increasingly reported from endotracheal secretions. Pseudomonas and Acinetobacter baumannii were predominant Non fermenter bacteria isolated from endotracheal secretions. Pseudomonas and Acinetobacter baumannii are reported as predominant isolates from various ICUs in literature [18-19].

In the present study Candida spp. were isolated and reported from different clinical samples. Maximum number of Candida spp. were isolated from blood (85.33%), followed by urine (13.90%), endotracheal secretions (3.00%), pus, wound, sputum and drain with equal no of isolates (0.37%). The Candida spp. were predominantly isolated from NICU (64.28%) followed by PICU (60.14%). In the present study the predominant Candida isolated were non albicans Candida(51.12%) followed by Candida utilis (22.55%), Candida albicans (13.53%), Candida tropicalis (6.01%), Candida pelliculosa (4.51%), Candida melibiosica (1.12%), Candida parapsilosis (0.75%) and Candida glabrata (0.37%). The findings are similar to a study done from north India by Chakrabarti A. et al. [20] Whereas another study from India reported higher number of Candida albicans isolates from ICU specimen [21]. In the present study Candida spp. were predominantly isolated from pediatric ICU than adult ICU as children are known to be at a high risk of candidemia because of more aggressive and invasive therapies, such as indwelling central lines, mechanical ventilation, parenteral hyperalimentation, longer hospital stay and probably immunocompromised state of neonates [22].

In the present study, maximum number of samples obtained was of blood and the most common culture positive clinical specimen in the present study was pus followed by endotracheal secretions. Gram negative bacteria were predominant in endotracheal secretions, gram positive bacteria predominated in CSF, and Candid was predominant in blood samples. The most common bacteria isolated in the present study were Klebsiella pneumoniae followed by Non fermenter group of organism. Klebsiella pneumoniae was also found to be the most common isolate from blood, endotracheal secretions, pus and drain whereas in urine Enterococcus faecium and Escherichia coli were found to be predominant isolates.

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