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Journal of the association of physicians of india • november 2013 • VOL. 61

Original Article

Clostridium difficile Infection at a Tertiary Care

Hospital in South India
Shashidhar Vishwanath1, Arpita Singhal1, Annet D’Souza1,
Chiranjay Mukhopadhyay1, Muralidhar Varma2, Indira Bairy1

Abstract
Objective: The objective of this study was to detect C. difficile in patients presenting with Antibiotic
Associated Diarrhoea.
Methods: Stool samples from twenty-five patients collected over a period of four months were processed
for C. difficile by culture and the isolates were identified following standard methods. C. difficile toxins
A and B and C. perfringens enterotoxin were detected by ELISA performed directly on stool specimens.
Results: Four patients (16%) were found positive for C. difficile infection. All patients with C. difficile
infection received prior treatment with third-generation cephalosporins or β-lactam / β-lactamase inhibitor
antibiotics. C. perfringens enterotoxin was found in two (8%) patients. Severe colitis was seen in one (25%)
of the four patients who had co-infection with C. difficile and C. perfringens.
Conclusion: This study demonstrated a significant occurrence of C. difficile infection in this hospital
population. There is a need to further evaluate the role of C. perfringens in causing antibiotic associated
diarrhoea. Good clinical and laboratory studies to generate local epidemiological data are essential to
increase awareness among the treating clinicians about C. difficile infection. Also limited and rational use
of broad spectrum antibiotics is recommended.

Introduction CDI in patients with diarrhoea and

recent antimicrobial exposure and

C lostridium difficile is recognised as hospitalisation. 3 Along with cytotoxin

the primary pathogen responsible
for antibiotic-associated colitis and
accounts for 15–25% of cases of
nosocomial antibiotic-associated
diarrhoea.1 Clinical manifestations
of C. difficile infection (CDI) range
from mild or moderate diarrhoea to
fulminant, pseudomembranous colitis,
sepsis, multi-organ failure and death. 1,2
Widespread regional outbreaks of C.
difficile strains involving more severe and
refractory disease have been reported,
with greater numbers of complications,
colectomies, and deaths than previously
1
Department of Microbiology
described. 3 Early diagnosis is the key
and 2Medicine, Kasturba
to prevent complications from severe
Medical College, Manipal
CDI and to prevent transmission. Rapid
University, Manipal 576 104,
Karnataka diagnosis depends on maintaining a
Received: 03.05.2012; high degree of clinical suspicion for
Accepted: 08.11.2012
detection, culture has been a mainstay
in the laboratory diagnosis of CDI which
is also essential for the epidemiologic
study of isolates. 1 We report our initial
experience with CDI in an Indian tertiary
care hospital setting.
Material and Methods
A p r o s p e c t i v e , p i l o t s t u d y wa s
conducted from May to August 2010
with consecutive 25 patients clinically
suspected with antibiotic associated
diarrhoea (AAD) were included. This
study was approved by our Institutional
Ethical Committee. Informed consent
was obtained at enrolment. Patients were
suspected as having AAD if significant
diarrhoea was noted along with a history

804  © JAPI • november 2013 • VOL. 61

Journal of the association of physicians of india • november 2013 • VOL. 61 31

Table 1 : Shows the clinical profile and microbiological factors in patients with Clostridium difficile infection
Duration Culture ELISA for ELISA for
Reason for
Sl Age (years) Prior antibiotic of prior for Clostridium Clostridium Severe
Diagnosis choosing the
No. / Sex therapy Antibiotic use Clostridium difficile Toxins perfringens Disease
Antibiotic
(days) difficile A and B Enterotoxin
Urinary tract
Based on
65 / infection Cefoperazone-
1 Susceptibility 12 + + - -
Female (Escherichia Sulbactum
results
coli)
Bilateral
+
55 / lower lobe Piperacillin-
2 Empirical 24 - + + Severe
Male pneumonia, Tazobactam
colitis
Sepsis
Septic
3 60 / Female Ceftriaxone Empirical 7 - + - -
arthritis
Cefoperazone-
Urosepsis Sulbactum;
Based on 3 months
4 76 / Female (Escherichia Cefixime- + + - -
Susceptibility (on and off)
coli) Clavulanic acid;
results
Ceftazidime

of treatment with antimicrobial or antineoplastic
agents within the previous 8 weeks. Patients were
considered to have CDI, if AAD was present along
with a positive toxigenic culture and / or a positive
result for C. difficile toxin by ELISA. 1 Children
below two years of age and those with other proven
aetiological agents of diarrhoea or diarrhoea inducing
factors were excluded from the study. Stool samples
collected in sterile, wide mouth containers were sent
to microbiology laboratory for further processing
without delay.
Stool specimens were processed for culture of
C. difficile following standard methods 4 and tested
for C. difficile toxins A and B (Premier toxins A and
B, Meridian Bioscience, Ohio, USA). Clostridium
perfringens is one of the most frequently cited
alternative causes of AAD. 5 Hence, the stool samples
were also tested for C. perfringens enterotoxin
(Ridascreen, R-Biopharm AG, Germany) by ELISA.
Culture for C. perfringens was not performed as it is
not recommended as a diagnostic method, given the
relative ubiquity of the bacterium in human faeces
as normal flora, and because not all isolates will
be enterotoxigenic. Also, C. perfringens enterotoxin
production from cultured isolates can be difficult to
demonstrate because it occurs during sporulation
and it is difficult to make this bacterium sporulate
in vitro. 5
Results
The mean age of study group was 40.7 years (range:
4 - 76 years, M:F = 1.5:1). Age group of 51 - 60 years had
the highest cases of AAD (36%). Most of the patients
(18, 72%) with AAD were from general medical wards,
followed by oncology (3, 12%); surgery (2, 8%), and
paediatric wards (2, 8%). CDI was diagnosed in four
(16%) patients with suspected AAD, all from general
medical wards. These four patients had received prior
treatment with beta-lactam antibiotics. The clinical
profile and microbiological factors of patients with
CDI are summarised in Table 1. None of them received
prior treatment with clindamycin or fluoroquinolones.
Mean age of patients with CDI was 64 years. Mean
duration of antibiotics use prior to onset of illness
was 18 and 11 days respectively in patients with CDI
and non CDI illness.
Among the risk factors which were analysed, prior
antibiotic usage, advanced age, history of contact
with child < 2 yrs of age and gastric acid suppression
therapy were seen among all the four CDI cases.
Three patients (75%) were hospitalised in the prior 6
months. None of them had a previous gastrointestinal
surgery or a prior C. difficile infection or nasogastric
intubation.
Discussion
ELISA (4, 16%) had a higher positivity rate
compared to culture (2, 8%). Both the cultured isolates
were toxigenic. The lesser sensitivity of culture may
be explained by sampling errors inherent to uneven
distribution of C. difficile in the faecal samples. 6 Also
inclusion of glutamate dehydrogenase detection
as screening test will be helpful to rule out false
positivity of ELISA in such instances. In comparison
with the positivity rates in Indian setups, this is in
agreement with the earlier studies of Dhawan B et
al. (15%), 7 Gogate A et al. (18%) 8 and Joshy L et al.
(12.1%). 9 Considering other geographical regions,
Heimesaat MM et al. 10 and Jamal W et al. 11 have
found the prevalence rates of CDI to be 11.4% and
8% respectively. None of the four patients diagnosed
with CDI in the present analysis were on prior
metronidazole or vancomycin. It is essential to know
the risk factors associated with CDI, as some of them

© JAPI • november 2013 • VOL. 61  805

32 Journal of the association of physicians of india • november 2013 • VOL. 61
may be modified to reduce the occurrence; or the
presence of risk factors in a patient with AAD can
raise the clinical suspicion of CDI, so that appropriate
treatment and control measures can be adopted.
Prevention of symptomatic C. difficile infections
requires a change in antibiotic policy with prudent
use of antibiotics and implementation of standard
infection control measures. 12
Clostridium perfringens enterotoxin (CPEnt) was
detected in 2 (8%) specimens. C. perfringens is reported
to account for 2-15% of all cases of AAD. 5 Asha NJ
et al. 5 and Vaishnavi C et al. 13 have reported similar
positive rates of 8% and 8.77% for CPEnt. However,
Joshy L et al. 14 have reported lower incidence of C.
perfringens AAD (2.6%). Both C. difficile toxin and
CPEnt were detected in one patient, who had severe
colitis 15 as evidenced by leucocytosis (20.3 x 10 9/L)
and radiographic findings (CT abdomen - Long
segment bowel thickening involving colon and
rectum with pericolonic fat stranding). It is likely
that both C. perfringens and C. difficile might work in
synergy for production of AAD 13 and thus may also
be responsible for causing severe disease. The inciting
antibiotics were withdrawn following the diagnosis
of CDI and all four patients improved significantly
with introduction of metronidazole therapy. There
was no mortality reported among CDI cases in this
study. The reported mortality rates in CDI vary from
0.6% to 83%. 16
Conclusion
Avoiding empirical therapy with β-lactam and
β-lactamase inhibitors (BL-BLIs) and choosing
antibiotics from the susceptibility panel with less risk
for developing AAD may reduce the occurrence of CDI.
In conclusion, this study demonstrated significant
presence of CDI in this hospital population. BL-BLIs
and third generation cephalosporins were the most
common antibiotics associated with development of
CDI. Being a pilot study, this study had limitation
of small sample size, however good clinical and
laboratory studies to generate local epidemiological
data are essential to increase awareness among the
treating clinicians about CDI. Limited and rational use
of broad spectrum antibiotics is recommended. There
is a need to further evaluate the role of C. perfringens
in causing antibiotic associated diarrhoea.
806  © JAPI • november 2013 • VOL. 61
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