East-West (1999)

Asymptomatic Bacteriuria And Sensitivity Pattern In Children With Sickle Cell Anaemia In A Tertiary Health In Enugu, South East Nigeria

Introduction

 

Urinary tract infection (UTI) is a common cause of renal disorder in the tropics 1 and causes significant morbidity 2, 3 and mortality 4 in children, especially when it is asymptomatic, not detected and promptly treated.

        Children with sickle cell anaemia have increased susceptibility to developing UTI because of slugging of sickled cells in the renal vasculature which causes papillary necrosis and loss of urinary concentrating and acidifying ability of the nephrons resulting in abnormally dilute and alkaline urine which favours bacterial proliferation.5 This predisposes them to recurrent UTI and subsequent renal damage. Studies 6, 7 have noted that children with sickle cell anaemia are more prone to developing UTI and other bacterial infections than their counterparts with normal haemoglobin.

        Children with sickle cell anaemia may have compromised kidney function from repeated vaso-occlusive episodes and recurrent UTI 8 These factors tend to hasten their development of CKD. However, this trend can be forestalled if the presence of asymptomatic bacteriuria can be detected early and appropriate therapy instituted.

        The current study, thus examines asymptomatic bacteriuria in children with sickle cell anaemia compared to their counterparts with normal haemoglobin. Findings from this study will be useful for making recommendations on measures to curtail the development of UTI among patients with sickle cell anaemia and thus reduce the burden and consequent morbidity and mortality arising thereby.

 

                                   

 

 

 

 

 

 

 

 

 

 

 

                               

 

 

 

 

 

 

 

 

 

 

Subjects and Methods

        It was a prospective study in which sickle cell anaemia children aged two years to 12 years who attended the weekly haemoglobinopathy clinic of the University of Nigeria Teaching Hospital (UNTH), Enugu were examined over an eight month period (December 2007 to July, 2008). A total of 100 children (57 males and 43 females) were selected consecutively as they presented to the clinic. Another 100 school children of the same age group with normal haemoglobin were also selected from day-care centres, nursery and primary schools in Enugu urban to act as controls. Children who had fever or may have had antibiotics in the past two weeks or had symptoms suggestive of UTI were excluded from the study.

        Ethical approval   for the study was obtained from the ethical and research committee of UNTH, Enugu while consent was obtained from parents and care-givers before commencing the study.

        Spot midstream urine specimens were collected into sterile boric acid bottles and transported in an ice containing box for analyses. Venous blood samples were collected from the controls for determination of their genotype using cellulose acetate electrophoresis (pH=8.6).

        Urinalysis was done using Combur-9 ® test strips. Urine sediments of each child’s urine after centrifugation at 2000rpm for five minutes was examined for red blood cells, leukocytes, casts, crystals and bacteria.

         The urine samples were cultured in cystine lactose electrolyte deficient(CLED) and blood agar media within one hour of urine collection by employing the semi-quantitative method as described by Guttmann and Stokes.9 A well calibrated standard wire loop of internal diameter 3mm and delivering 0.003ml of urine per loopful was sterilized over a Bunsen burner flame before immersing in well mixed uncentrifuged urine and then streaked into well dried plates of CLED and blood agar media (which were earlier incubated) as described by Uquarhart and Gould.10 Cultures were incubated aerobically at 370C for 24 hours and the colonies counted by using a colony counter. Only samples that yielded pure bacterial growth of 105 or more colony forming units (cfu) per milliliter were regarded as yielding significant bacteriuria. Counts between 104 and 105 were repeated while counts≤ 104 CFU were regarded as negative. Mixed growths were regarded as contaminants and therefore disregarded. Second urine samples were collected from children with significant bacteriuria and those whose second urine samples yielded significant bacteriuria were regarded as having asymptomatic bacteriuria.

        Organisms were identified using standard identification techniques.11 Antibiotic sensitivity disc (Abtek biologicals)® containing ampicillin, cotri-moxazole , gentamicin , nitrofurantoin, colistin, tetracycline, nalidix acid, streptomycin and single discs of Drovid (ofloxacin), Siprosan (ciprofloxacin) and Avicef (ceftriaxone) were used and sensitivity pattern determined by the Stoke’s method  of comparing the zones of inhibition of the test organism.12

                                   

                                      Data analyses

Data was analyzed using the Statistical Package for the Social Sciences (SPSS) version 15.0. Proportions were tested using chi-squared test while means were compared with t-tests. Statistical significance was taken as p

                                     

                                             

                                                    

 

 

 

 

 

 

 

 

 

                                                                Results

        Out of the 100 urine specimens from the subjects, 6(6%) had significant bacterial growth on two consecutive cultures showing a prevalence of asymptomatic bacteriuria of 6% among children with sickle cell anaemia. Five (83.3%) of these were females while one(16.3%) was a male giving female to male ratio of 5:1 which is statistically significant(χ2=4.2, df=1, p=0.04). On the other hand, only two of 100 urine specimens from the female controls yielded significant bacteriuria on two consecutive cultures showing a prevalence of 2%.

        Both Gram positive and Gram negative organisms were isolated from the subjects with Escherichia coli, a Gram negative enterobacteria  isolated from two (33.3%) of the six subjects with asymptomatic bacteriuria while proteus spp, Staphylococcus albus, Streptococcus faecalis, Staphylococcus aureus constituted 16.7% each. Staphylococcus aureus and Streptococcus faecalis constituted 50% each of the isolates from the controls. Table I shows the Gram stain distribution of isolated organisms from subjects and contols while Table II shows the distribution of organisms isolated.

        Sensitivity pattern of the isolated organisms from the subjects showed that 4(66.7%) were sensitive to gentamicin and ciprofloxacin, 3(50%) were sensitive to nitrofurantoin, nalidixic acid and colistin sulphate while 6(100%) were sensitive to ofloxacin and ceftriaxone. All were resistant to co-trimoxazole, ampicillin, tetracycline and streptomycin. Among the controls, the two (100%) isolated organisms were sensitive to gentamicin, ofloxacin and nitrofurantoin while the two (100%) were resistant to co-trimoxazole, ampicillin and streptomycin. Tables III and IV depict the sensitivity of the isolated organisms in subjects and controls respectively.

        All the urine samples from subjects and controls with asymptomatic bacteriuria were acidic and none was positive for nitrite test. There was no correlation between pyuria and significant bacteriuria.

                                     

 

 

 

 

 

 

Discussion

Studies on asymptomatic bacteriuria in children with sickle cell anaemia are very scanty. However, in this study, the prevalence of asymptomatic bacteriuria in children with sickle cell anaemia is 6% and is comparable with the result obtained from Lagos by Ajasin et al13 who documented 5.8%. The proportion of children with sickle cell anaemia who had significant bacteriuria in the current study is quite high when compared with the prevalence (2%) among children with normal haemoglobin genotype. At same time it is higher than the figures obtained by Abdulrahman14 (1%) and Okafor15 (2.1%) among children with normal haemoglobin in Kaduna and Enugu respectively. The current study further supports the greater susceptibility of children with sickle cell anaemia to urinary tract infection (UTI) with a threefold increase in asymptomatic bacteriuria when compared to their normal haemoglobin genotype controls. This higher risk is due to the defect in urine concentrating and acidifying abilities of the kidneys of children with sickle cell anaemia. This produces an abnormally dilute urine which favours bacterial proliferation.5

       Of the children with sickle cell anaemia who had significant bacteriuria in the current study, females out-numbered their male counterparts in a ratio of 5:1.  This ratio agrees with that obtained by Tarry et al16 (10:1) as well as Ajasin et al13 (3:2). This higher risk in the females has been attributed to short course of the female urethra and its proximity to the anal region. 

     Escherichia coli, a Gram negative enterobacteria was isolated from two (33.3%) of the six subjects with asymptomatic bacteriuria while proteus spp, Staphylococcus albus, Streptococcus faecalis, Staphylococcus aureus constituted 16.7% each. Previous studies of asymptomatic bacteriuria both in HbSS7 and HbAA subjects 17.18,19,20 have identified Gram negative organisms particularly E. coli and Klebsiella species as the most prevalent pathogens causing UTI in children. The Gram negative organisms have also been implicated as the most common cause of symptomatic UTI both in children with sickle cell anaemia16, 21 and their counterparts with normal haemoglobin.18 The range of pathogens in the current study is similar to that reported by earlier workers 13,,16,20 except that Klebsialla species, the second most commonly reported pathogen causing UTI was not isolated in the current study. Thus organisms like E.coli, Klebsiella species, Proteus species, Streptococcus faecalis and Staphylococcus aureus are frequently isolated in subjects with asymptomatic bacteriuria irrespective of haemoglobin genotype. In developed countries of the world, E. coli is responsible for 80-90% of all organisms isolated from the urinary tract of children with UTI. The frequency with which this organism causes UTI in the developing countries including Nigeria is low as organisms such as Staphylococcus aureus, Streptococcus faecalis and proteus species have larger representation of causative agents in UTI in these less developed countries. This may be due to poor environmental and personal hygiene in these less developed countries. It also seems that sickle cell anaemia has some effect in the pattern of distribution of the organisms responsible for UTI allowing a greater representation of some other organisms such as Proteus and staphylococcus species. This may be due to the general impairment of the immune system in patients with sickle cell anaemia. The organisms isolated from the two controls with positive culture were both Gram positive organisms. This is at variance with what was obtained by Okafor et al20 in which Gram negative organisms accounted for 59% of the 17 cases of asymptomatic bacteriuria among pre-school children. This variance may be due to the smaller number of control children used in the current study.

        The sensitivity of the isolated organisms indicates that most of the organisms were resistant to the older antibiotics such as cotrimoxazole, ampicillin, streptomycin and tetracycline (contraindicated in children less than 8 years) both in subjects and controls. This high resistance to the older antibiotics was also noted in the study by Okafor et al20 though that by Ajasin et al 13 showed varied resistance to these older antibiotics. The reason for this high resistance may be due to self medication and/or sub-therapeutic (drug pressure) prescription by health workers. It may also be due to intrinsic drug resistance developed by the pathogens.

         Urinalysis in both subjects and controls indicated that the urine samples were acidic in all the children with asymptomatic bacteruria. It has been stated that children with sickle cell anaemia have urine acidifying defect but this has not been so in the current study. It may be that the kidneys of these subjects still retain their ability to acidify urine. Nitrite test was negative in all the children with asymptomatic bacteriuria and this may be due to the low sensitivity of this test in detecting bacteriuria.22    

        The urine microscopy among the sickle cell anaemia patients showed pyuria ranging from 1-6/hpf but only one with significant bacteriuria had significant pyuria. The sensitivity and specificity of significant pyuria as a determinant of significant bacteriuria in centrifuged urine sample is 61% and 43% respectively.23 This relatively low sensitivity may explain the presence of significant pyuria in only one of the subjects with asymptomatic bacteriuria. Pyuria is an indication of active inflammation and in cases of asymptomatic bacteriuria as in the current study, significant pyuria may not be detected.

        In view of the increased incidence of asymptomatic bacteriuria in children with sickle cell anaemia, we recommend routine urine screening in the clinics using at least the fast and economical tests for detecting bacteriuria such as nitrite and leukocyte esterase tests and that quinolones may be considered in the empirical treatment of UTI in children.

 

                                                     Acknowledgement                                    

We wish to acknowledge Merss  Obi and Paul of the department of microbiology, UNTH, Enugu for their immense technical assistance as well as Dr Ohanu, the head of microbiology department of UNTH for approving the use of the laboratory for this study.

References

 

1              Eke FU and Eke NN. Renal disorder in children: a Nigerian Study: Pediatr Nephrol 1994; 8:383-386. 

                

2              Stockland E, Hellstrom M, Jackobson B, Judal U and Sixt R. Renal        damage one year after       first urinary tract infection: Role of   dimecarptosuccinic acid scintigrapahy. J Pediatr 1999;              129:815-820.

3              Disk PT and Foldman W. Routine diagnostic imaging for childhood urinary tract                infection. J Pediatr 1996; 128:15-22.

4              Neuman CG and Pryles CR. Pyelonephritis in infants and children; Autopsy experience at               Boston City Hospital, 1933-1960. Am J Dis     Child 1962; 104: 90-102.

5              Smith CH. Blood diseases of infancy and childhood. 3rd ed. St. Louis: C. V. Mosby Company;       1972: 376-377.

6              Konotey-Ahulu FID. Sickle cell disease patient. Tetteh A’Domeno company; 1996: 376-377.

7              Gendrel O, Richard-lenoble D, Valette H, Kombila M, MakangaH, Toure R, et al. Salmonella         infections and Haemoglobin S. JPediatr 1982 ; 101 : 68-69.

8              Ronald JF and Charles JJ. Renal disease. In: Stephen HE., Robert PH, Narla M and           Martin HE (editors). Sickle cell disease: basic principles and clinical practice, New York:          Raven press ltd; 1996:673-680.

9              Guttman DE and Stokes J. Diagnosis of urinary tract infection: Comparison of a pour plate            method with a routine method. BMJ 1963; 25: 1384-1387.

10           Uqurhart GED and Gould JC. Simplified technique of counting bacteria in urine and other               fluids. J Cln Path 1965; 18: 480.       

  

11           Monica Cheesbrough. Laboratory examination of urine. In: District laboratory practice in               tropical countries; part 2. London: Cambridge University press 2000: 107-113.

               

12           Baker FJ, Silverton RF and pallister CJ. In: Baker and Silverton’sIntroduction to medical                 laboratory technology. 4th ed. London: Butterworth-Heinemann 1998: 313-314

 

13           Ajasin MA and Adegbola RA. Asymptomatic Bacteriuria in children with sickle cell           anaemia. Nig J Paediatr 1988; 15: 19-25.

 

14           Abdurrahman MB, Chakrabarty DP, and Ochoga SA. Bacteriuria and other urinary          abnormalities among primary school children in Kaduna. Nig J Paediatr 1978; 5: 21-24

 

.15          Okafor HU, Okoro BA, Ibe BC. and Njoku Obi NU. Prevalence of Asymptomatic Bacteriuria         among nursery school children. Nig J Paediatr 1993; 20:84-88

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16           Tarry WF, Dukket JW, and Mc Synder. Urological complications of sickle cell disease in a                paediatric population. J Urol 1987; 138:592-594

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17           Newcastle Asymptomatic Bacteriuria Research group. Arch Dis Child 1975; 50: 90-102.

18           Kunnin CM, and DeGroot JE. Self screening for significant bacteriuria. JAMA 1975; 231:                1349-1353.

19           Wemambu SNC. Bacteriological profile and sensitivity pattern in childhood urinary tract                 infection in Benin City. J Trop Paediatr 1983; 29: 85-86.

20           Okafor HU, Okoro BA, Ibe BC, and Njoku Obi NU. Bacteriology of asymptomatic bacteriuria       in preschool children in Enugu. Orient J Med 2005; 17: 37-42.

21           Robinson MG, and Halpera C. Infections, Escherichia coli and sickle cell anaemia. JAMA               1975; 230: 1145-1148.

               

22           Wammanda RD, Aikhionbare HA and Ogalla WN. Use of nitrite dipstick test in the           screening for urinary tract infection in children. W. Afr J Med 2000; 19:206-208.

23           Pryles CV and Eliot CR. Pyuria and bacteriuria as a diagnostic criterion of urinary tract    infections. Am J Dis Child 1965; 110:628-               635.

               

 

 

 

 

 

 

 

 

 

 

 

 

 

Table I: Gram stain distribution of isolated organisms in subjects and controls

 

 

Gram stain

Subjects

Controls

Total

Positive

3

2

5

Negative

3

0

3

 

 

Table II:                Frequency/distribution of organisms isolated from subjects and controls

Organisms

Subjects

Controls

E. coli

2(33.3%)

0

Staph. aureus

1(16.7%)

1(50.0%)

Staph. albus

1(16.7%)

0

Proteus spp

1(16.6%)

0

Strep. Faecalis

1(16.7%)

1(50.0%)

Total

6(100.0%)

2(100.0%)

 

 

 

 

 

 

 

 

 

 

 

Table III: Positive cultures, organisms and sensitivity pattern in subjects.

S/N

Sex

Organism

Sensitivity

20

F

E. coli

S/G,NT,NA,CL,CF,OF,CP R/AM,CT,ST,TET

44

F

Staph. albus

S/CF,OF,CP, R/G,NT,NA,AM,CT,CL,ST,TET

63

M

Staph. aureus

S/G,NT,NA,CF,OF

R/CT,CL,ST,TET,AM,CP

72

F

E. coli

S/CF,CL,NT,OF. R/G,NA,CP,AM,CT,ST,TET

82

F

Proteus

S/G,NA,CF,CP,OF,CL

R/NT,AM,CT,ST,TET

88

F

Strep. Faecalis

S/G,CF,CP,OF, R/NA,NT,CT,CL,AM,ST,TET

 

S=Sensitive, R=Resistant

G=Gentamicin, NA=Nalidixic acid, CL=colistin sulphate, OF=Ofloxacin

AM=Ampicillin, TET=Tetracycline, CF=Ceftriaxone, ST=Streptomycine

CT=Cotrimoxazole, NT=Nitrofurantoin, CP=Ciprofloxacine

S/N=Serial number

 

Table IV: Positive cultures, Organisms and sensitivity pattern in controls.

S/N

Age in years

Sex

Social class

Organism

Sensitivity

30

10

F

2

Strept.faecalis

S/G,NT,CF,CP,OF R/NA,CT,CL,TET,AM,ST

60

8

F

4

Staph.aureus

S/G,NT,OF,TET R/ST,CL,CT,CP,CF,NA,AM

 

S=Sensitive, R=Resistant, AM=Ampicillin

G=Gentamicin, NT=Nitrofurantoin, CF=Ceftriaxone, CP=Ciprofloxacine

NA=Nalidixic acid, CT=Cotrimoxazole, CL=Colistin sulphate

TET=Tetracycline, ST=Streptomycine, Of=Ofloxacine

S/N=Serial number.

 

 

About the Author

CHUKWU BF, OKAFOR HU, IKEFUNA AN.

 

 

Department of Paediatrics, University of Nigeria Teaching Hospital, Enugu.

 

 

Correspondence: Dr BF Chukwu, Department of Paediatrics, University of Nigeria Teaching                    Hospital, Enugu.

 

E-mail: chizzy_bath@yahoo.com

 

 

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