Antimicrobial sensitivity trends and bacterial profile of bloodstream infections: A retrospective study

INTRODUCTION

Blood plays a crucial role in maintaining physiological homeostasis by transporting O₂, nutrients, hormones, and metabolic waste products, along with providing immunity against invading pathogens.^[1] Bloodstream infections (BSIs) represent a significant clinical concern, and they are caused by a wide range of microorganisms, including bacteria, fungi, viruses, and protozoa.^[2] Among these bacteria are the highest frequently isolated etiological agents. Entry of micro-organisms into the bloodstream can lead to systemic dissemination, resulting in severe clinical consequences as circulating pathogens affect multiple organs.^[3] If it is not diagnosed and treated promptly, BSIs may progress to life-threatening complications such as septic shock, disseminated intravascular coagulation, multiple organ dysfunction syndrome, and death [Figure 1].^[2,3] Bloodstream infections remain a major global health concern because of their associated morbidity and mortality, particularly in hospitalized patients.^[4,5] Even though BSI occurs worldwide, its incidence and fatality rates are considerably mostly higher in the developing countries.^[5]

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Figure 1: Bloodstream infection; causes symptoms and mortality.

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BSI management largely depends on early identification of the causative organisms and determination of antimicrobial susceptibility patterns. Initial empirical therapy is usually guided by local epidemiological data and resistance trends, while targeted therapy is initiated after isolation and susceptibility testing of the pathogens.

Conventional blood culture methods are time-consuming because they depend on microbial growth. In contrast, automated blood culture systems such as the BD BACTEC™ FX40 enable earlier detection of bloodstream pathogens with improved sensitivity and shorter turnaround time. The BD BACTEC™ FX40 system allows early detection of bloodstream pathogens with improved sensitivity and recovery rates. Therefore, the present study was undertaken to identify the prevalent bacterial pathogens causing bloodstream infections in this region and to evaluate their antimicrobial susceptibility patterns.

MATERIAL AND METHODS

Specimen collection and processing

Timing

Blood samples were collected from clinically suspected cases of BSIs admitted to various inpatient departments of the hospital for routine blood culture and sensitivity testing. Samples were collected before the initiation of empirical antimicrobial therapy. In patients who were already receiving antibiotics, blood samples were collected just before the administration of the next dose.

Volume

The volume of blood collected was considered critical due to the low concentration of organisms in bacteria, particularly in patients receiving antimicrobial therapy. A higher blood volume increases the yield of bacterial isolation. BD BACTEC blood culture bottles were used with recommended volumes of 8–10 mL/bottle for adults (BD BACTEC Plus Aerobic/F Culture vials) and 1–3 mL/bottle for pediatrics (BD BACTEC Plus^™/F Culture vials). Only aerobic bottles were used in the study.^[6]

Collection method

All blood samples were collected by venipuncture before the collection of any other blood samples. Blood culture bottles were maintained in an upright or slightly inclined position to prevent contamination of the bottle neck during venipuncture.^[6]

Preparation of site and antiseptics

Blood collection was performed under strict aseptic conditions using sterile disposable syringes. The venepuncture site was first cleaned with 70% isopropyl alcohol, followed by application of a second antiseptic (tincture iodine or chlorhexidine). The rubber septum of the blood culture bottle was disinfected with alcohol and allowed to dry. After blood collection, any residual iodine on the skin was removed using alcohol.^[6]

Labeling and transport

All blood culture bottles were appropriately labeled and transported to the microbiology laboratory at room temperature. In case of transport delay, samples were not refrigerated as per standard guidelines.^[6]

Laboratory procedure

Blood culture bottles were immediately loaded into the BD BACTEC^™ FX40 [Figure 2] automated blood culture system. Positive bottles were indicated automatically by the system upon sufficient bacterial growth. Bottles showing no growth after 5 days of incubation were reported as sterile for aerobic bacterial pathogens.^[6]

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Figure 2: Use of an automated blood culture system.

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Identification of bacterial isolates

Positive blood culture bottles indicated by the BD BACTEC^™ FX40 system were subcultured on Selective media – blood Agar and indicator media – MacConkey Agar plates [Figure 3]. The inoculated plates were incubated aerobically at 37°C for 18–24 h and examined for colony morphology. Identification of bacterial isolates was performed using standard microbiological techniques.^[7] This included assessment of colony morphological characteristics, Gram staining, and biochemical testing such as indole, methyl red, VogesProskauer, citrate utilization, urease, triple sugar iron, oxidase, and mannitol motility test for Gram-positive cocci, which were further identified based on colony morphology, Gram stain, enzymatic test, catalase, and coagulase test [Figure 4].

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Figure 3: Isolation of organisms by conventional culture method using the streak culture technique on (a) blood agar and (b) macConkey agar.

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Figure 4: Current workflow of microbiological diagnosis in bloodstream infection.

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Antimicrobial susceptibility testing

Antimicrobial susceptibility testing was carried out using the Kirby–Bauer disk diffusion method [Figure 5] in accordance with the Clinical and Laboratory Standards Institute guidelines, and antibiotic discs (HIMEDIA Laboratories) were used as listed in Tables 1 and 2. Quality control for antimicrobial susceptibility testing was ensured using the following American Type Culture Collection (ATCC) reference strains:

• Escherichia coli (ATCC 25922)

• Staphylococcus aureus (ATCC 25923)

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Figure 5: Antibiotic susceptibility test by Kirby–Bauer disk diffusion method.

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Table 1: Antibiotics disk for Gram-positive cocci. Antibiotic Disk

Antibiotic Disk
	Cotrimoxazole	Azithromycin	Teicoplanin	Gentamicin	Doxycycline	Moxifloxacin	Ciprofloxacin
Symbol	COT	AZM	TEI	GEN	DO	MO	CIP
Potency (μg)	5	15	30	10	30	5	5
Antibiotic Disk
	Cefoxitin	Erythromycin	Linezolid	Penicillin G	Vancomycin	Amoxy/clav. acid	Clindamycin
Symbol	CX	E	LZ	P	VA	AMC	CD
Potency (μg)	30	15	30	10 units	30	20/10	2

RESULTS

The present study was conducted between March 2022 and April 2022, and a total of 181 non-repetitive blood samples were collected from patients clinically suspected of BSIs at the JNUIMSRC, Jaipur National University, Jaipur. Patient details such as hospital identification number, laboratory accession number, age, sex, and site of sample collection were recorded using a patient’s pro forma. Out of the 181 blood samples processed, 37 (20.44%) showed bacterial growth while 144 (79.55%) samples were reported as no growth by the BD BACTEC™ FX40 automated blood culture system. The highest blood culture positivity was observed among patients in the 0–10-year age group, and the lowest positivity was noted in patients aged 50–60 years and above 61 years [Figure 5]. Among the 37 culture-positive cases, the majority of patients were admitted to ICUs, followed by pediatric wards, general medicine, surgery, and respiratory medicine. Gram-positive cocci were the most frequently isolated organisms, with methicillin-sensitive S. aureus (MSSA) being the predominant pathogen [Figure 6]. Among Gram-negative bacilli, Klebsiella species were the most commonly isolated organisms [Figure 7].

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Figure 6: Age-wise distribution of blood culture-positive patients.

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Figure 7: Distribution of Gram-positive cocci isolated from the blood cultures.

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DISCUSSION

BSIs continue to pose a significant clinical challenge due to their high morbidity and mortality, particularly in hospitalized patients. Early diagnosis, prompt initiation of appropriate antimicrobial therapy, and continuous monitoring of resistance trends are essential for effective patient management.

In the present study, the blood culture positive rate was 20.44% which is comparable to findings reported in earlier studies conducted in similar tertiary care settings in India.^[8-12] Variations in positivity rate across studies may be attributed to differences in patient population, volume of blood collected, number of culture sets, and prior antibiotic exposure. Lower positivity in some studies may reflect early empirical antibiotic use before sample collection.^[13-15]

The highest (37.83%) incidence of BSIs was observed among pediatric patients (0–10 years), indicating increased susceptibility in this age group. This may be due to an immature immune system, frequent hospital visits, and increased exposure to environmental pathogens. Male predominance was observed in this study is consistent with previous reports and may be associated with higher exposure to external risk factors.^[12]

Gram-positive organisms were predominant (54.05%) and slightly less than the second highest Gram-negative organisms constituted (45.94%) in the present study. The predominance of Gram-positive organisms in BSIs has also been documented by several earlier studies.^[9,12,16,17] Among Gram-positive isolates, Staphylococcus species was the most common with S. aureus accounting for 85% of cases. Among Gram-positive isolates, Staphylococcus aureus accounted for 85% of cases. MSSA was the predominant isolate, constituting 50% of cases, followed by coagulase-negative staphylococci (CoNS), which accounted for 30% of cases. The increasing prevalence of MSSA BSIs has been highlighted in previous reports.^[12] Enterococcus species constituted approximately (15%), Gram-positive isolates, and demonstrated resistance to multiple commonly used antibiotics, emphasizing the need for careful antimicrobial selection.

Among Gram-negative organisms, Klebsiella species were the leading cause of BSIs (52.94%), followed by Acinetobacter species (35.30%) [Figure 8]. These findings are clinically significant as both organisms are known to exhibit high levels of antimicrobial resistance and are frequently associated with hospital-acquired infection.^[12,18-20]

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Figure 8: Distribution of Gram-negative bacilli isolated from the blood cultures. X-axis = Gram-negative Isolates, Y-axis = Number of Isolates (%).

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Antimicrobial susceptibility pattern observed in the study revealed that vancomycin and teicoplanin were highly effective against Gram-positive isolates, whereas colistin and amoxiclav demonstrated good activity against Gram-negative isolates. These results provide valuable insight into local antimicrobial resistance patterns and may assist clinicians in selecting appropriate empirical and targeted therapy for the management of BSIs.

CONCLUSION

The current study aims to highlight the prevalence of BSIs and the antimicrobial susceptibility patterns of bacterial isolates at a tertiary care hospital in Jaipur, Rajasthan, India. Gram-positive organisms, particularly S. aureus, were the most common causative agents, while Klebsiella species predominated among Gram-negative isolates. A high proportion of multidrug-resistant organisms, including MSSA and Gram-negative bacilli, was observed.

Knowledge of local pathogen distribution and antimicrobial susceptibility trends is essential for the selection of appropriate empirical therapy. Early identification of causative organisms and timely initiation of targeted antimicrobial treatment can significantly reduce morbidity, mortality, treatment duration, and healthcare costs associated with BSIs. Continuous surveillance of antimicrobial resistance patterns is strongly recommended to guide effective infection control and antibiotic stewardship programs.