Isolation of obligate anaerobes from clinical samples received for routine bacterial culture and sensitivity: a cross sectional study

Abstract

Background and objectives: Obligate anaerobic bacteria are known to cause various infections in human beings. We aimed to determine the prevalence and spectrum of obligate anaerobes encountered in pus aspirate, sterile fluids and tissue samples received for routine bacterial culture and sensitivity.

Materials and methods: A total of 160 samples including tissue, sterile body fluids and pus aspirate were collected, analysed for prevalence and spectrum of obligate anaerobes. Identification of obligate and facultative anaerobes was done by automated MALDI-TOF and Vitek 2 method.

Results: Among 160 samples, 75 samples (46.8%) yielded obligate anaerobes out of which 41 samples (26%) yielded obligate anaerobes along with facultative anaerobes which was significant (p value=0.031) and 34 samples (21%) yielded only obligate anaerobes. 90 obligate anaerobes were isolated from 75 samples among which only 34 (37.7%) samples yielded only obligate anaerobes and 56 (62.2%) yielded both obligate and facultative anaerobes. Gram stain with polymicrobial appearance (p value 0.02) was found to be significantly associated with growth of obligate anaerobes. Clinical conditions where obligate anaerobes were commonly associated were appendicular abscess, empyema, fournier's gangrene, diabetic foot, ludwigs angina and deep abscess. Out of 75 positive samples 30 (40%) patients had predisposing conditions like diabetes mellitus, hypertension etc. Total of 90 obligate anaerobes and 49 facultative anaerobes were isolated. The common obligate anaerobes were Bacteroides fragilis 18 (20%), Prevotella spp. 20 (22.2%), and Clostridium spp. 8 (8.88%). Facultative anaerobes like Escherichia coli 25 (34.7%), Klebsiella species 15 (20.8%), Enterococcus faecalis 19 (26.3%) were isolated. Antibiotic sensitivity was performed for facultative anaerobes by Kirby bauer disc diffusion method. Out of 15 Escherichia coli isolates resistance was commonly seen for ampicillin 13 (86.6%), cephalosporins 11 (73.3%), ciprofloxacin 10 (66.6%) and Piperacillin tazobactum 8 (53.3%). In Klebsiella species resistance were commonly seen to Ampicillin 6 (100%), cephalosporins 2 (33.3%) and ciprofloxacin 2 (33.3%).

Conclusion: There was significant isolation of obligate anaerobes along with facultative anaerobes in clinical samples received for aerobic culture and sensitivity. There is a need for isolation of these bacteria routinely and a scope for doing antibiotic susceptibility testing, which will help in evidence-based medicine and a better clinical outcome by giving appropriate therapy.

Keywords: Facultative anaerobes; Obligate anaerobes; Polymicrobial appearance.

Fig. 2.

Flow chart showing the algorithm of sample processing and identification of bacteria.

Fig. 3.

Growth of different types of organisms in culture and their direct Gram stain showing polymicrobial appearance.

Fig. 4.

Pie chart showing clinical samples yielding different categories of bacteria in culture.

Fig. 1.

(a) Showing black pigmented colonies of Prevotella melaninagenica on Brucella Blood Agar. (b) Gram stained colony smear of Prevotella melaninagenica showing Gram negative coccobacilli (GNCB) under 1000×. (c) Gram stained colony smear of Fusobacterium nucleatum showing slender gram negative bacilli (GNB) with tapering ends under 1000×