Bacterial profile and antimicrobial susceptibility pattern of asymptomatic urinary tract infections among pregnant women attending antenatal clinic at tertiary care hospital in central India

Introduction: It is estimated that 2 to 10% of pregnant woman suffer from any form of urinary tract infections. These infections complicate up to 20% ofpregnancies and are responsible for the majority of ante-partum admissions to the maternal-fetal medicine units. Material and methods: It was a cross sectional study conducted in the Department of Microbiology, Peoples College of Medical Sciences & Research Center, Bhopal (M.P) during period of July 2015 to March 2016. A total of 210 pregnant women having asymptomatic bacteriuria during the period of July2015 to March 2016 attending antenatal clinic were included (all trimester) in this study. Results: Women in age group 21-30years showed the highest incidence of asymptomatic bacteriuria (70.58%), incidence of asymptomatic bacteriuria was generally high throughout pregnancy, but relatively highest during the second trimester. Escherichia coli was the most common 9 (52.94%) followed by Staphylococcus aureus 4 (23.52%), Klebisella pneumonia 2(11.76%) among the isolates. E.coli was found to be sensitive to cefepime and norfloxacin followed by ceftazidime ceftriaxone amoxicillin + clavulanic acid, nitrofurantoin, gentamycin and amikacin. Conclusion: This study revealed 8.09% prevalence of asymptomatic bacteriuria among pregnant women. This is of concern because UTI in pregnancy may have serious consequences for both the mother and the child.


Introduction
It is estimated that several hundred million women suffer from urinary tract infection annually, with costs to health care providers amounting to over $6 billion annually worldwide, a figure that may even be an underestimate [1].
It is estimated that 2 to 10% of pregnant woman suffer from any form of urinary tract infections [2]. These infections complicate up to 20% of pregnancies and are responsible for the majority of ante-partum admissions to the maternal-fetal medicine units [3].
Urinary tract infection (UTI) occurs frequently in pregnancy with a large number of infections being asymptomatic. The frequency of asymptomatic bacteriuria (ASB) in pregnancy accounts for the rationale of urinalysis throughout pregnancy period [4]. It can be associated with varieties of adverse obstetric outcome and medical conditions such as the development of acute and chronic pyelonephritis, preterm labour, low birth weight, pre eclampsia, chronic renal disease, and prenatal mortality [5]. Undiagnosed and untreated bacteriuria in pregnancy represents a considerable risk factor for acute pyelonephritis in approximately 5% of pregnant women with increased risk of low birth weight and prenatal fatal death [6].
Detection and treatment of bacteriuria has been reported to prevent up to 80% cases of pyelonephritis [7]. Bacteriuria in pregnancy usually reflects prior colonization e.g. at the time of onset of sexual activity rather than acquisition during pregnancy itself [8]. Furthermore, there is obviously a substantial risk that the infecting pathogens are becoming resistant to empirically prescribed antimicrobial agents normally used in the community.  Inclusion criteria: Pregnant women who had asymptomatic bacteriuria were included.

Original Research Article
Exclusion criteria: Diabetes mellitus/hypertension, fever and known congenital anomalies of urinary tract will be excluded from this study.
Sample collection: About 30ml of clean catch midstream urine sample was collected in 100ml sterile, dry, leak-proof container [9]. The patients were instructed to discard first 10-30 ml of the voided urine and collect about 30ml of the second midstream urine. The pregnant woman was instructed to adequately clean the peri-urethral area with water and soap solution without antibacterial activity [10].
Transport: Urine specimen was transported to the laboratory immediately after obtaining and was processed within one hour or in case of delay they were refrigerated at 4°c (upto 24 hours), until culture could be performed [11].
Microscopic examination: At first, urine was examined microscopically as a wet preparation to detect significant pyuria, Urine is mixed carefully and 0.05ml is transferred on to the middle of a microscope slide and applies cover slip. The preparation was examined microscopically using the 10x and 40x to look for bacteria, pus cells, epithelial cells, crystals.
Semi quantitative Urine culture was done using calibrated wire loop (0.005ml) of 28 SWG which had internal diameter of 3.28 mm [12], and urine samples were inoculated into Cystine Lactose Electrolyte Deficient medium. Culture plate was incubated in aerobic atmosphere at 37°C for 18-24 hours [13]. Colonies were counted to check the presence of significant bacteriuria Colony count yielding bacterial growth of 10 5 CFU/ml of urine was regarded as significant bacteriuria. If the colony forming unit (CFU) is less than 10 5 , it was considered as non significant bacteriuria or negative [14].
All positive cultures with significant bacteriuria were then identified to species level by their colony characteristics, gram staining reaction and by the pattern of biochemical profiles using standard procedures.
Biochemical reactions: Following biochemical test were put up to identify bacteria to species level such as catalase, coagulase for gram positive and oxidase, indole, citrate, urea, triple sugar iron, lysine decarboxylase, arginine and ornithine and motility testfor Gram negative bacteria following standard procedure.

Results
Out of 210 urine samples of ANC Women examined for asymptomatic bacteriuria, 17 were having significant bacteriuria (10 5 cfu/ml) giving frequency rate of 8.09%.
The frequency of asymptomatic bacteriuria according to age distribution showed that majority of the subjects screened were in the age group of 21-30 years, Women in age group 21-30 years showed the highest incidence of asymptomatic bacteriuria (70.58%) is shown in Table 1. Frequency of asymptomatic bacteriuria was generally high among the three trimesters, but relatively highest during the second trimester as shown in Table 2.  Our study showed that Escherichia coli was the most common -9 (52.94%) followed by Staphylococcus aureus 4(23.52%), Klebsiella pneumonia 2 (11.76%) among the isolates as shown in Table 3. In our study, E.coli was found to be sensitive to cefepime and norfloxacin followed by ceftazidine ceftriaxone amoxicillin + clavulanic acid, nitrofurantoin Gentamycin and amikacin as shown in table no. 4.

Original Research Article
* (The isolate in which antibiotic sensitivity was not performed is shown by -)

Discussion
In the present study, the frequency of asymptomatic bacteriuria was 8.09% (17 cases) among the sampled population of 210 pregnant women attending ante-natal clinic. Similar observations were found by other authors [15] although R Sujata et al found it to be 3% which is less than our study [16]. This variation may be explained by the differences in the environment, social habits ofthe community, the standard of personal hygiene and education. In the present study, the subjects in the age groups of 21-30 years showed the highest frequency of infection (70.58%). Study conducted by Lavanya et al [17] reported higher frequency in subjects  [16]. There was no difference in frequency of urinary tract infection in pregnant women with previous history of urinary tract infection and those without.
In the present study, it was observed that the women in their second and third trimester had higher frequency of asymptomatic bacteriuria (52.94%), and this could be attributed to the anatomical and physiological changes experienced by pregnant women during these stage of pregnancy, for instance the uterus expands and also there are increased hormonal effects which together offset normal homeostatic balance making conditions favorable for microbial invasion.
Though frequency of UTI in the first trimester was low it could only be that it served as incubation period for most of the microbes, only for microbial invasion to be manifest in the second and third trimester [19].
In the present study it was observed that Escherichia coli was the most common isolated organism in 9 (52.94%) subjects followed by Staphylococcus aureus 4 (23.52%), Klebsiella pneumonia 2(11.76%) among the isolates.
There were one isolate of Proteus mirabilus and Pseudomonas aeroginosa each. Other studies have also reported that Escherichia coli were often the most common isolated organism in asymptomatic bacteriuria [20][21][22].
In the present study, E.coli was found to be sensitive to cefepime and norfloxacin followed by ceftazidine, ceftriaxone amoxicillin + clavulanic acid, nitrofurantoin Gentamycin and amikacin. This was in accordance to the study done by Andabati G and Byamugisha et al in Africa in 2010 [23].

Conclusion
High prevalence of asymptomatic bacteriuria among pregnant women is of concern because UTI in pregnancy may have serious consequences for both the mother and the child. The most predominant organisms were Escherichia coli and Staphylococcus aureus. Norfloxacin, Ceftriaxone, Cefepime and Amoxycillin + clavulanic acid were very effective against most of the urinary isolates.
What the study adds in the existing knowledge?
Routine urine culture test should be carried out on all antenatal women to detect asymptomatic bacteriuria. Screening should be done regularly for the presence of asymptomatic bacteriuria in pregnancy and testing antimicrobial susceptibility with safe drugs in pregnant women is also necessary, so that these can be used for the treatment.