Associate professor Department of Microbiology BLDEU’s Shri B M Patil Medical College Bijapur –Karnataka 586103
Email: basucm@rediffmail.com
Received: 25 Aug 2014 Revised and Accepted: 24 Sep 2014
ABSTRACT
Objectives: Urinary tract infection (UTIs) caused by Citrobacter species have been described in 5 to 12% of bacterial urine isolates in adults. The genus Citrobacter is a distinct group of aerobic Gram-negative bacilli from the Enterobacteriaceae family. We report here the emergence of Citrobacter as an increasingly common urinary pathogen in patients attending this medical college.
Methods: The study was carried out over a period of 2 years from January 2010 to December 2011. Urine specimens from both outpatients and inpatients of our hospital were processed. Urine samples which yielded the growth Citrobacter spp. were included in the study
Results: E. coli was the most common organism isolated followed by Klebsiella spp. and Citrobacter spp. The most active antimicrobial agent against Citrobacter isolates was piperacillin-tazobactum (55% sensitive), closely followed by amikacin (47%).
Conclusion: The emergence of this usually rare organism as the third most common urinary pathogen, which is resistant to commonly available antibiotics is alarming. Such studies will guide clinicians to choose accurate empirical treatment options and will help to reduce the mortality and morbidity rates from infections.
Keywords: Anti-microbial resistance, Antibiotics, Citrobacter, Drug resistance, Urinary tract infections.
INTRODUCTION
Urinary tract infection (UTI) continues to be the commonest nosocomial infection according for approximately 40% of all hospital acquired infections and it is one of the most important causes of morbidity and mortality [1, 2]. UTIs caused by Citrobacter species have been described in 5 to 12% of bacterial urine isolates in adults [3]. The genus Citrobacter is a distinct group of aerobic Gram-negative bacilli from the Enterobacteriaceae family [4]. Citrobacter species are primary inhabitants of intestinal tract, often found in human feces [5]. They are also found in soil, sewage, food, and animal feces [6, 7].
These organisms are isolated from variety of clinical specimens like urine, pus, blood, and cerebrospinal fluid [8]. Organisms of genus Citrobacter are Gram-negative straight rods, found singly or in pairs, and are motile by peritrichous flagellae. They are facultative anaerobes, oxidase-negative, and typically utilize citrate as sole source of carbon. The genus Citrobacter comprises 11 different species. Among these, Citrobacter koseri (previously known as C. diversus) and C. freundii are the commonest species implicated in infections. Citrobacter species can cause variety of infections like, respiratory tract infections, urinary tract infection, blood stream infections, wound and burns infections, meningitis, endocarditis, and peritonitis [9]. We report here the emergence of Citrobacter as an increasingly common urinary pathogen in patients attending this medical college.
MATERIALS AND METHODS
Study population, design, and setting
The current study was conducted in the Department of Microbiology, Shri B M Patil Medical College, Bijapur, from January 2010 to December 2011.
Ethical clearance and consent
As it was a retrospective study, ethical clearance and consent was not obtained.
Patient evaluation
A total of 1434 urine specimens from both outpatients and inpatients of our hospital having one or more urinary symptoms, like burning during micturition, fever, pyuria, frequency of urine, dysuria, hematuria, flank pain, suprapubic discomfort, etc., were processed.
Sample collection and isolation, identification of the organism
Mid-stream urine sample in early morning was collected in wide mouth sterile container [10]. All urine samples were examined by routine microscopic examination by wet mount of urine sediment after centrifuging urine for 10 minutes at 1000 revolution per minute (rpm).
Presence of pus cells, red blood cells (RBCs), epithelial cells, casts, and crystals were noted as supportive findings of urinary infection. Simultaneously all urine samples were cultured over routine culture media; Mac Conkey agar and Cysteine lactose electrolyte deficient (CLED) agar with a sterile standard loop. These plates were incubated at 37°C for 2 consecutive days.
Culture results were interpreted according to the standard criteria and a growth of ≥105 colony forming units/ml was considered as significant bacteriuria [11][11]. Cultures with more than three colonies were discarded, as contaminants and their antibiotic susceptibility were not tested. Species were identified by conventional biochemical tests according to standard microbiological techniques [10].
Antimicrobial susceptibility testing
All isolates were tested for antimicrobial susceptibility on Mueller Hinton agar by the standard disc diffusion method recommended by the Clinical and Laboratory Standards Institute (CLSI)[12]. agents (disks) were obtained from Hi Media laboratories, Pvt Ltd, Mumbai. Appropriate quality control strains were used to validate the results of the antimicrobial disk. E. coli, ATCC 25922, and Pseudomonas aeruginosa, ATCC 27853, were used as quality control strains [12].
Statistical analysis
Statistical analysis was performed with SPSS 14 software. Categorical data was analyzed using Chi-Square test with a 0.05 significance level.
RESULTS AND DISCUSSION
The education and application of personal hygiene are important in that Citrobacter strains are excreted as fecal wastes. Epidemics occurring in hospitals are closely related to the fact that hospital staff carry the bacterium in their hands and gastrointestinal systems. Citrobacter strains are mostly isolated from the infections of urinary and respiratory systems as nosocomial infection causes [13].
Urinary tract infection by C. koseri has been reported to be 12.0% in 1969 and the prevalence rate is rising. Invasive procedures like, catheterization helps them to colonize urinary bladder and during intensive chemotherapy this bacterium disseminates to the blood stream to cause severe bacteremia. Intact immunity helps to control the pathogen to certain extent but when the patients are immunocompromised, the situation is grave. The problem is further intensified by the emergence of multidrug resistance Citrobacter sp. resulting into treatment failure [14].
In the present study, Citrobacter infections were high among elderly people; this is because of the fact that these groups constitute large proportion of our hospital populations and reduced immunity in these people to fight against infection in general. Similar results were seen in the study conducted by Shih et al [15].
In this study piperacillin-tazobactum was the most active drug, closely followed amikacin. Least active antibiotics were ampicillin and amoxyclav. Amoxycillin and ampicillin are often used as oral therapy for gram –negative UTIs, but the high rate of in vitro resistance demonstrated in this study and others suggests that they should not be used. Trimethoprim and amoxyclav are also often prescribed; of concern is the increasing rate of resistance to trimethoprim over the last 10 years and the more recent increase in resistance to amoxyclav, presumably as a result of mechanisms other than production of beta –lactamase [16].
Table 1: Frequency of different organism isolated from UTI
| Organism | Number | Percentage |
| E. coli | 314 | 45.6 |
| Klebsiella spp. | 124 | 17.9 |
| Citrobacter spp. | 108 | 15.7 |
| Other Gram negatives bacilli | 38 | 5.5 |
| Gram positive cocci | 61 | 8.9 |
| Fungi | 44 | 6,4 |
| Total | 689 | 100 |
Of the 1434 urine samples received for culture during the study period, 12 samples contained more than one species, significant bacteriuria was found in 48.1 % samples. Among these E. coli was the most common organism isolated (Table 1) followed by Klebsiella spp. and Citrobacter spp.
Table 2: Age and sex wise distribution of Citrobacter spp from UTI patients
Age in years |
Male |
Female |
Total |
|||
|
Number |
% |
Number |
% |
Number |
% |
1-20 |
6 |
8.1 |
4 |
10.8 |
10 |
9.26 |
21-50 |
24 |
35.1 |
23 |
62.2 |
47 |
43.5 |
> 51 |
41 |
57.7 |
10 |
27.0 |
51 |
47.2 |
In all age groups, except those aged more than 51 years, females were more frequently affected than males. Among females, frequency of UTI was more among 21-50 years age group and among males elderly patients were more commonly affected.
Table 3: Antimicrobial susceptibility of uropathogenic Citrobacter isolates
| Antibiotics | Sensitive | Sensitive(%) |
| Ampicillin | 3 | 2.9 |
| Co –trimoxazole | 20 | 18.5 |
| Norfloxacin | 28 | 25.9 |
| Ciprofloxacin | 40 | 37 |
| Gentamicin | 25 | 23.1 |
| Nalidixic acid | 10 | 9.26 |
| Amoxyclav | 8 | 7.41 |
| Amikacin | 51 | 47.2 |
| Cephalexin | 8 | 7.41 |
| Cefoparazone –salbactam | 35 | 32.4 |
| Piperacillin-tazobactum | 59 | 54.6 |
| Ofloxacin | 46 | 42.6 |
| Nitrofurantoin | 39 | 36.1 |
The most active antimicrobial agent against Citrobacter isolates were piperacillin-tazobactum (55% sensitive), closely followed by amikacin (47 %). Amikacin was more effective aminoglycoside than others. Least active antibiotics were ampicillin (3%) and cephalexin.(7%)
Majority of the isolates showed multidrug resistence in the current study and were found to be resistant to cephalaxin, norfloxacin, ciprofloxacin, and the aminoglycosides [Table 3]. This has important implications as patients in a tertiary care hospital like ours receive cephalosoprins, aminoglycosides, fluoroquinolone, or a combination of these drugs as empirical therapy or as definitive treatment. Since good in vitro activity was shown by piperacillin-tazobactum it may be considered as first line therapy for ambulatory patients. The high rates of antibiotic resistance observed in the present study may be due to the fact that ours is a tertiary care hospital with widespread usage of broad spectrum antibiotics leading to selective survival advantage of pathogens.
In our study, most of the isolates were from inpatients, similar observations have been made by previous studies [14,17], the probable reason for this are as follows. The hospital staff carry the bacterium in their hands and gastrointestinal systems and invasive procedures like catheterization helps these organisms to colonize urinary bladder [13,14].
CONCLUSION
The Citrobacter isolates resistant to multiple anti-microbial agents have emerged, making it an emerging nosocomial pathogen. The emergence of this usually rare organism as the third most common urinary pathogen, which is resistant to commonly available antibiotics is alarming. The indiscriminate use of antimicrobial agents is possibly the main reason responsible for this situation. Therefore, such studies will guide clinicians to choose accurate empirical treatment options and will help to reduce the mortality and morbidity rates from infections. It is more significant to prevent the resistance development in micro-organisms and to lend the accurate information to clinicians in terms of the use of antibiotics in appropriate period.
CONFLICT OF INTERESTS
Declared None
REFERENCES