Get Permission Bhonsle, Vyas, and Vyas: Isolation, identification and frequency of isolated uropathogens with their antibiotic susceptibility pattern causing urinary tract infections in patients of Ujjain M.P. (India)


Introduction

Urinary tract infection (UTI), is a major health related problem affecting large number of people all over the world and causes clinical and financial burden on human society. It is an inflammatory disorder which occurs in one or more part of urinary tract and causes cystitis, pyelonephrities and urethrietis.1 The major symptoms of UTI are fever, burning and pain in urination, increased frequency of urination, pain in kidney and tissue damage.2 UTI’s are the second most common infection and several types of microorganisms such as bacteria, viruses, fungi and parasites causes UTI but bacteria are most common cause of UTI. Most of the UTI cases are mainly caused by Gram negative bacteria such as Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter spp. and some Gram positive bacteria like Enterocoocus faecalis and Staphylococcus aureus. Among these bacteria, E. coli is most predominant agent of UTI.3, 4, 5, 6

Urinary tract infections are found in both male and female patients and in all age groups but females are more prone to UTI due to their anatomical and physiological factors of female urinary tract system. UTI may be Asymptomatic or Symptomatic and can occur in uncomplicated or complicated types. The uncomplicated UTI occur in a normal urinary tract while complicated UTI occur in abnormal urinary tract. If complicated UTI is not diagnosed and treated properly then it may spread and causes serious problems such as bacterimia, renal failure and premature delivery due to acute pyelonephritis.7, 8

The proper treatment of UTI is done by timely identification of causative agent of UTI and detecting its susceptibility to various antibiotics but now a days resistance of uropathogens towards antibiotics is increases worldwide due to non-judicious, random and uncontrolled use of antibiotics, self-medication and incorrect investigations.9 [9] The increasing drug resistance among uropathogens has made treatment of UTI difficult and it has became a serious problem so research is needed to identify causative agent of UTI and their antibiotic susceptibility patterns among patients of UTI in Ujjain , which may help in proper antibiotic therapy and prevent development of resistant microbes.9, 10

Materials and Methods

The present study was conducted for one-year period starting from November 2018 to October 2019 to identify common uropathogenic bacteria causing UTI and antibiotic susceptibility of isolated uropathogens were determined against different antibiotics. The urine samples of suspected patients were collected in sterile container from different hospitals of Ujjain and processed in SRL laboratory Ujjain centre. Urine samples were examined physically for observing color and turbidity of urine sample and microscopically for detecting pus cells, epithelial cells, RBCs and bacteria.

After examination urine samples were inoculated and cultured on different types of culture media blood agar, Chrome agar and Mac’Conkey agar plate and incubated at 37oC for 24 hrs. After incubation if the growth of bacterial culture was> 105 (CFU)/ ml then it was considered as a positive sample. All the bacterial isolates were further identified and confirmed by their motility, morphological and biochemical characters as per the standard procedures. 11

Antibiotic susceptibility test

Antibiotic susceptibility test was performed on Muller Hinton Agar by using Kirby Bauer Disc Diffusion method. This test and interpretation of results was done according to Clinical Laboratory Standards Institute (CLSI) guidelines for determination of susceptibility of uropathogens causing UTI against antimicrobial agents. Bacterial colony was selected then inoculated in peptone water broth and incubated at 370C for 2 hrs. After adjustingto 0.5 McFarlands standard this test organism was spread on Muller Hinton Agar by sterile swab using lawn method and commercially available antibiotic Himedia discs were placed on this lawn and plate was incubated at 37oc for 24-48 hours. The incubated plates were observed for size of zone of clearance and the size was interpreted as sensitive, intermediate and resistant according to CLSI guidelines. 12

The following antibiotic discs (drugs concentrations in mcg) were applied for observing susceptibility pattern of uropathogens: Amikacin (30mcg), Gentamicin (10mcg), Tobramycin (10mcg), Ampicillin (10mcg), Amoxicillin-Clavulanate (20/10mcg), Piperacillin (100mcg), Piperacillin/Tazobactam (100mcg), Ciprofloxacin (5mcg), Norfloxacin (10mcg), Tetracycline (30mcg), Doxycycline (30mcg), Nitrofurantoin (300mcg), Cotrimoxazole (1.25/23.75mcg), Ceftazidime (30mcg), Imipenem (10mcg), Meropenem (10mcg), Doripenem (10mcg) and Aztreonam (30mcg) were used for gram negative bacteria. In addition to this Teicoplanin (30mcg), Vancomycin (30mcg), Penicillin (10), Cefoxitin (30mcg), Rifampin (5mcg) and Lenizolid (30mcg) were used for gram positive bacteria.

Quality control strains used were:

  1. Staphylococcus aureus ATCC 25923

  2. Escherichia coli ATCC 25922

  3. Pseudomonas aeruginosa ATCC 27853

Results

A total of 770 midstream clean catch urine samples were collected from patients during the studies and 486 urine samples were found positive and prevalence of UTI in patients was (63.1%). The percentage of 430(88.4%) gram negative bacteria and 56 (11.5%) gram positive bacteria was detected. Out of 321 positive urine samples, 349(71.8%) were from female and 137(28.1%) were from male patients found UTI (Table 1).

Table 1

Prevalence of UTI in different genders

Gender

Number of UTI patients

Percentage of UTI (%)

Male

137

28.1

Female

349

71.8

Total

486

63.1

Table 2

The frequency of isolated uropathogenic bacteria from urine culture

Isolated Bacteria

Number of isolated bacteria

Percentage of occurence (%)

Escherichia coli

222

45.6

Klebsiella pneumoniae

132

27.1

Pseudomonas aeruginosa

76

15.6

Enterococcus faecalis

36

7.4

Staphylococcus aureus

20

4.1

Total

486

Table 3

Antibiotic susceptibility pattern of isolated gram negative bacteria. (N=430)

Tested antibiotics

E. coli n=222

K. pneumoniae n=132

P. aeruginosa n=76

No. of sensitive bacteria

(%)

No. of sensitive bacteria

(%)

No. of sensitive bacteria

(%)

Amikacin

186

(83.7)

111

(84.0)

56

(73.6)

Gentamicin

162

(72.9)

108

(81.8)

60

(78.9)

Tobramycin

170

(76.5)

96

(72.7)

52

(68.4)

Ampicillin

21

(9.4)

15

(11.3)

NA

NA

AMC

26

(11.7)

21

(15.9)

NA

NA

Piperacillin

69

(31.0)

109

(82.5)

50

(65.7)

PTZ

182

(81.9)

111

(84.0)

61

(80.2)

Ciprofloxacin

38

(17.1)

23

(17.4)

22

(28.9)

Norfloxacin

67

(30.1)

27

(20.4)

39

(51.3)

Tetracycline

138

(62.1)

72

(54.5)

NA

NA

Doxycycline

158

(71.7)

68

(51.5)

NA

NA

Nitrofurantoin

188

(84.6)

70

(53.0)

NA

NA

COT

88

(39.6)

54

(40.9)

NA

NA

Ceftazidime

58

(26.1)

82

(62.1)

44

(57.8)

Imipenem

202

(90.9)

109

(82.5)

64

(84.2)

Meropenem

205

(92.3)

119

(90.1)

68

(89.4)

Doripenem

209

(94.1)

124

(93.9)

71

(93.4)

Aztreonam

168

(75.6)

104

(78.7)

54

(71.0)

[i] Value in columns indicates number of sensitive bacteria & values in parenthesis indicate percentage of sensitive bacteria.

[ii] N- Total no. of bacteria, n- Number of sensitive bacteria, NA-Not Applicable

[iii] AMC-Amoxicillin/clavulanate, PTZ- Piperacillin/Tazobactam, COT- Co-trimoxazole

Table 4

Antibiotics sensitivity pattern of isolated gram positive bacteria. N=(56)

Tested Antibiotics

E. faecalis n=36

S. aureus n =20

No. of sensitive bacteria

(%)

no. of sensitive bacteria

(%)

Amikacin

NA

NA

15

(75.0)

Gentamicin

NA

NA

14

(70.0)

Teicoplanin

32

(88.8)

18

(90.0)

Vancomycin

33

(91.6)

19

(95.0)

Tetracycline

03

(8.3)

15

(75.0)

Doxycycline

05

(13.8)

18

(90.0)

Penicillin

02

(5.5)

06

(30.0)

Ampicillin

19

(52.7)

08

(40.0)

AMC

NA

NA

10

(50.0)

Ciprofloxacin

04

(11.1)

06

(30.0)

Norfloxacin

03

(8.3)

10

(50.0)

Nitrofurantoin

29

(80.5)

16

(80.0)

Co-trimaxazole

NA

NA

08

(40.0)

Rifampin

30

(83.3)

18

(90)

Linezolid

34

(94.4)

19

(95)

[i] Value in columns indicates number of sensitive bacteria & values in parenthesis indicate percentage of sensitive bacteria.

[ii] N- Total no. of bacteria, n- Number of sensitive bacteria, NA-Not Applicable

[iii] AMC-Amoxicillin/clavulanate

[iv] NA- Not applicable

The most commonly isolated gram negative uropathogens included Escherichia coli (45.6%) followed by Klebsiella pneumoniae (27.1%), Pseudomonas aeruginosa (15.6%) and gram positive uropathogens included Enterococcus faecalis (7.4%) and Staphylococcus aureus (4.1%) (Table 2).

The study of antibiotic susceptibility patterns of isolated uropathogens showed that E. coli showed highest sensitivity against carbapenems such as Doripenem (94.1%) and Meropenem (92.3%) closely followed by Imipenem(90.9%) beside these antibiotics more than 70% sensitivity was found against Amikacin, Gentamicin, Tobramycin, Doxycycline, Nitrofurantoin and Aztreonam antibioticsandleast sensitivity found against to Ampicillin (9.4%). Similarly, K. pneumoniae showed highest sensitivity to Doripenem (93.9%), Meropenem (90.1%) while susceptibility to Ampicillin was (12.9%). It was also found that P. aeruginosa was highly sensitive against Doripenem, and Meropenem and their susceptibility rates are (93.4%) and (89.4%) respectively and least sensitivity was seen for Ciprofloxacin (28.9%). In case of gram positive bacteria E. faecalis was predominant uropathogen and it was highly sensitive for Teicoplanin, Vancomycin and Linezolid, for these antibiotics its sensitivity rate was (88.8%), (91.6%) and (94.4%) respectively which was followed by Rifampin (83.3%) and Nitrofurantoin (80.5%) and it showed lower sensitivity for Penicillin (5.5%). Similarly S. aureus was highly susceptible for Tiecoplanin, Vancomycin, Rifampin and Linezolid and least susceptible to Ciprofloxacin and Penicillin (Table 3, Table 4).

Discussion

The present study represents prevalence and antibiotics susceptibility patterns of uropathogens which can differ in same country with time and place. In this study prevalence of UTI was found (63.1%). This findings similar with other studies done in India. 13, 14, 15 The frequency of occurrence of UTI was found higher (71.6%) in female than (28.4%) in male patients. The findings with this regard are in consonance with the studies done in India and Ethiopia.4, 15 In the present study among gram negative bacteria E. coli has been reported as the predominant uropathogen, responsible for causing UTI and its frequency of occurrence was (45.6%) which was followed by K. pneumoniae, P. aeruginosa and gram positive bacteria E. faecalis and S. aureus uropathogens. These results are in accordance with the studies done in Pakistan, Ethiopia and Saudi Arabia.4, 8, 7 In this study E. coli showed highest sensitivity for Doripenem (94.1), Meropenem (92.3%) and Imipenem (90.9%) and these findingsare same as with the study done in India.5, 14, 16 However study done in Pakistan showed opposite results to our study and they reported low susceptibility to Imipenem (39.5%). 17 E. coli also showed high susceptibility for Amikacin, Nitrofurantoinand Piperacillin/Tazobactam and their percentage of susceptibility was (83.7%), (84.6%), (81.9) respectively. These findings are similar to the study done in Southern Iran 18 while E. coli showed least susceptibility for Ampicillin (9.4), Ciprofloxacin (17.1%). These results of susceptibility are correlate with the study done in Gujarat, India.5 In case of K. pneumoniae uropathogen it shows higher susceptibility to Carbapenems, Piperacillin /Tazobactam, Gentamicin and Amikacin and very low susceptibility for Ampicillin these results are same as the studies done in India and Southern Iran. 5, 19, 18 In this study P. aeruginosa shows higher sensitivity towards Carbepenems, Piperacillin / Tazobactam, Aztreonam, Amikacin and Gentamicin and their sensitivity was more than 70 percent while for Norfloxacin it showed 51.3% percent sensitivity and low sensitivity to Ciprofloxacin (27.2%). These results are same to the studies performed in India and Ethiopia.5, 20, 21 In this study it was found that gram positive bacteria E. faecalis and S. aureus were highly susceptible to Vancomycin, Linezolid, Rifampin and Teicoplanin and their percentage of sensitivity was in between 80-95% while low sensitivity was demonstrated against Ciprofloxacin, Norfloxacin and Penicillin. These findings are in accordance with the previous studies done in India. 1, 22

Conclusion

The findings of this study showed that UTI’s have become a major health related problem and affect large number of people through the World. In this study E. coli isolates were the predominant uropathogen and all isolates are sensitive for carbapenems, aminoglycoside, antibiotic groups while these isolates are resistant to commonly prescribed antibiotics so very few options of drugs are available for physicians for the proper treatment of UTIs. Therefore, to ensure appropriate therapy current knowledge related to the organisms that cause UTI and their antibiotic susceptibility is mandatory.

Source of Funding

None.

Conflicts of Interest

None.

Acknowledgements

We would like to thanks the laboratory staff of SRL lab Ujjain and Deshumukh Hospital Ujjain (M.P.) for their support in completing my work.

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Article History

Received : 12-05-2023

Accepted : 22-06-2023


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https://doi.org/ 10.18231/j.ijmmtd.2023.022


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