**4. ASB and renal function decline and hypertension in patients with DM**

Women with DM have an increased prevalence of ASB, but also an increased risk on symptomatic UTI's and developing complications of UTI's such as renal abscesses. It was also shown that at short term follow-up treatment of ASB in women with DM did not appear to reduce complications. *E. coli* is the leading uropathogen in non-diabetic as well as in diabetic patients. Ninety percent of *E. coli* possesses type 1 fimbriae, the adhesive organelles found at the outer bacterial membrane. We have shown in vitro that type 1 fimbriated *E. coli* have an increased adherence to uroepithelial cells voided by women with DM. Others demonstrated that UTI's with type 1-fimbriated *E. coli* can lead to scar formation in the renal parenchyma of infected rats.At present, conclusive and prospective data with a long follow-up period directly relating ASB (with *E. coli*) to long-term risk of renal failure in diabetic patients are lacking. Taken together, we hypothesized that ASB in women with DM could lead to a faster decline in renal function, and decided to enlarge our cohort of diabetic women and to prolong the follow-up period. Besides the effects on renal function, we also studied the influence of ASB on the development of hypertension.

The association between ASB and renal function decline (and hypertension) in patients with DM was investigated in a prospective study with women with DM type 1 (n=296) and type 2 (n=348). All patients were interviewed and their medical records were reviewed at baseline and at study closure to collect all relevant information. All patients were asked to provide 1 or 2 midstream urine specimens. The women were followed up for a mean (SD) duration of 6.1 (1.9) years. Women with DM type 1 were younger, but had a longer duration of DM, than women with DM type 2. At baseline, 201 women with DM type 2 (58%) were treated with insulin only, 97 (28%) with oral hypoglycemic medication only, 41 (12%) with a combination of both, and five women (2%) were on a diet only (data were incomplete for 4 women). Because the Cockcroft-Gault formula for the estimation of the creatinine clearance includes age, adjusting for age in a multivariate model is not possible. Therefore patients were stratified into 3 age strata to assess the impact of age on the association between ASB and the (relative increase in the) creatinine clearance (respectively 18 to 36, 37 to 55, and 56 to 75 years old). All analyses were performed on the entire study population and on women with DM type 1 and DM type 2 separately.The prevalence of ASB was 17% in the study population, lower in

Asymptomatic Bacteriuria (ASB), Renal Function and Hypertension 383

type 2 DM. As shown, women with ASB at baseline had a lower creatinine clearance at study end point, a faster relative decrease in creatinine clearance, and hypertension more often when compared univariately with women without ASB. However, the differences were mainly explained by differences in age and duration of DM, and all differences

Comparable results were found in a small Polish study (25 patients with DM, including both men and women), in which no differences in the incidence of hypertension and renal function decline were demonstrated between patients with and those without ASB after 14 years.

In a recent Canadian study it was investigated whether successive isolates of urinary *E. coli* from the same diabetic woman were genetically similar. It was shown that untreated diabetic women with ASB may carry a genetically unique *E. coli* strain for up to 13 months. Women who received treatment for ASB had bacteriuria for a shorter duration and carried a single strain of *E. coli* for a shorter period compared with women who did not receive treatment. However, treatment was followed by recurrent infections for most women, usually with a new strain of *E. coli*. The ASB-causing *E. coli* from diabetic women did not have virulence characteristics typical of UTI-causing strains. This non-virulent microorganism might be an explanation of the low number of patients who have also

Because in the above mentioned prospective study no evidence was found that ASB in itself can lead to a decline in renal function, either in women with type 1 DM or in women with type 2 DM, it is not likely that treatment of ASB will lead to a decrease in the incidence of diabetic nephropathy. This is in accordance with a recent study of women with DM with ASB in which a comparison was made between women who received antibiotic therapy and women who received placebo. In that study, no difference was seen in serum creatinine

In conclusion, the hypothesis that ASB will lead to renal function deterioration in women with DM can be rejected because no difference in renal function development, in either women with type 1 DM or those with type 2 DM were found. Also, the incidence of hypertension was not increased when comparing women with ASB versus women without ASB. Therefore, at this time, screening andsubsequent treatment for ASB are not indicated in patients withDM.

It has been found that up to 50% of renal transplant recipients have ASB and UTIs. Many risk factors contribute to the high incidence of UTIs and ASB, which can undermine graft function and survival. In a retrospective study the impact of ASB on renal transplant outcome was analysed in 189 renal transplant recipients. Screening resulted into 298 episodes of ASB in 96 recipients (follow-up 36 months). Significant risk factors included female gender, glomerulonephritis as the disease that led to transplantation, and double renal transplant. There were no differences in serum creatinine, creatinine clearance, or proteinuria between patients with and without bacteriuria. The incidence of pyelonephritis in these patients was 7.6 episodes per 100 patient-years compared with 1.1 in those without ASB. A total of 2-5 ASB episodes were independent factors associated with pyelonephritis whereas more than 5 episodes was a factor associated with rejection. Studies show contradictory results whether

antibiotic treatment results into a lower prevalence of ASB in these patients.

leukocyturia, as a result of the absence of a host response to this.

disappeared in the multivariate analyses.

levels after a mean follow-up of 2 years.

**5. ASB in renal transplant recipients** 

women with type 1 DM (12%) compared to women with type 2 DM (21%), but multivariate analysis revealed that this was due to the difference in age. *E. coli* was cultured in 74 (67%) of the 110 women with ASB. Other isolated microorganisms included *enterococci (9%), group B streptococci (8%), Klebsiella pneumoniae (6%), Staphylococcus aureus (3%), Proteus mirabilis (2%), Enterobacter* species (2%). The prevalence of leukocyturia (5 or more leukocytes per highpower field) was 15% in women with ASB, suggesting that bacteria were present without resulting into an inflammatory response. The creatinine clearance decreased from 87 at baseline to 76 mL/min at study endpoint in diabetic women with ASB, and from 97 mL/min to 88 mL/min in those without ASB (Figure 2). In the univariate analysis, ASB was associated with a higher relative decrease in creatinine clearance (14 ± 22% and 9 ± 23% in women with versus women without ASB, respectively, p = 0.03), but not with the absolute decrease in creatinine clearance (12 ± 19 and 9 ± 20 mL/min, respectively, p = 0.12). Using univariate analysis, age, the length of follow-up, the duration of DM and microalbuminuria were identified as possible confounding factors when studying the influence of ASB on renal function development. Therefore, a multivariate analysis was done, according to age strata, and including the length of follow-up, duration of DM, and microalbuminuria at baseline. In the multivariate analysis no association was found between ASB and the relative or the absolute decrease in creatinine clearance. Also when women with DM type 1 and those with DM type 2 were analyzed separately, no association was found (data not shown). Finally, also no association with a faster decline in renal function was found when only the urines with *E. coli* as the cultured microorganism were included in the analysis

Fig. 2. Differences in creatinine clearance between women WITH DM with and without ASB. (Meiland R, Geerlings SE, Stolk RP, Netten PM, Schneeberger PM, Hoepelman AIM. Asymptomatic bacteriuria in women with diabetes mellitus. Arch Intern Med 2006; 166: 2222-7.)

Diabetic women with ASB developed hypertension more often than women without ASB (54% vs 37%; p=.045). However, in the multivariate analysis, including age, duration of DM, and length of follow-up, the association between ASB and hypertension disappeared (p>.20); a higher age was the strongest predictor for hypertension. In conclusion, in this prospective study after 6 years of follow-up, no association was found between ASB and a decline in renal function or the development of hypertension in women with type 1 DM or

women with type 1 DM (12%) compared to women with type 2 DM (21%), but multivariate analysis revealed that this was due to the difference in age. *E. coli* was cultured in 74 (67%) of the 110 women with ASB. Other isolated microorganisms included *enterococci (9%), group B streptococci (8%), Klebsiella pneumoniae (6%), Staphylococcus aureus (3%), Proteus mirabilis (2%), Enterobacter* species (2%). The prevalence of leukocyturia (5 or more leukocytes per highpower field) was 15% in women with ASB, suggesting that bacteria were present without resulting into an inflammatory response. The creatinine clearance decreased from 87 at baseline to 76 mL/min at study endpoint in diabetic women with ASB, and from 97 mL/min to 88 mL/min in those without ASB (Figure 2). In the univariate analysis, ASB was associated with a higher relative decrease in creatinine clearance (14 ± 22% and 9 ± 23% in women with versus women without ASB, respectively, p = 0.03), but not with the absolute decrease in creatinine clearance (12 ± 19 and 9 ± 20 mL/min, respectively, p = 0.12). Using univariate analysis, age, the length of follow-up, the duration of DM and microalbuminuria were identified as possible confounding factors when studying the influence of ASB on renal function development. Therefore, a multivariate analysis was done, according to age strata, and including the length of follow-up, duration of DM, and microalbuminuria at baseline. In the multivariate analysis no association was found between ASB and the relative or the absolute decrease in creatinine clearance. Also when women with DM type 1 and those with DM type 2 were analyzed separately, no association was found (data not shown). Finally, also no association with a faster decline in renal function was found when only the

urines with *E. coli* as the cultured microorganism were included in the analysis

0 1 to 5 5 to 7 7 to 9

Fig. 2. Differences in creatinine clearance between women WITH DM with and without ASB. (Meiland R, Geerlings SE, Stolk RP, Netten PM, Schneeberger PM, Hoepelman AIM. Asymptomatic bacteriuria in women with diabetes mellitus. Arch Intern Med 2006; 166:

Diabetic women with ASB developed hypertension more often than women without ASB (54% vs 37%; p=.045). However, in the multivariate analysis, including age, duration of DM, and length of follow-up, the association between ASB and hypertension disappeared (p>.20); a higher age was the strongest predictor for hypertension. In conclusion, in this prospective study after 6 years of follow-up, no association was found between ASB and a decline in renal function or the development of hypertension in women with type 1 DM or

lenght of follow-up, yr

no ASB ASB

0

20

40

60

80

creatinine clearance, mL/min

2222-7.)

100

120

type 2 DM. As shown, women with ASB at baseline had a lower creatinine clearance at study end point, a faster relative decrease in creatinine clearance, and hypertension more often when compared univariately with women without ASB. However, the differences were mainly explained by differences in age and duration of DM, and all differences disappeared in the multivariate analyses.

Comparable results were found in a small Polish study (25 patients with DM, including both men and women), in which no differences in the incidence of hypertension and renal function decline were demonstrated between patients with and those without ASB after 14 years.

In a recent Canadian study it was investigated whether successive isolates of urinary *E. coli* from the same diabetic woman were genetically similar. It was shown that untreated diabetic women with ASB may carry a genetically unique *E. coli* strain for up to 13 months. Women who received treatment for ASB had bacteriuria for a shorter duration and carried a single strain of *E. coli* for a shorter period compared with women who did not receive treatment. However, treatment was followed by recurrent infections for most women, usually with a new strain of *E. coli*. The ASB-causing *E. coli* from diabetic women did not have virulence characteristics typical of UTI-causing strains. This non-virulent microorganism might be an explanation of the low number of patients who have also leukocyturia, as a result of the absence of a host response to this.

Because in the above mentioned prospective study no evidence was found that ASB in itself can lead to a decline in renal function, either in women with type 1 DM or in women with type 2 DM, it is not likely that treatment of ASB will lead to a decrease in the incidence of diabetic nephropathy. This is in accordance with a recent study of women with DM with ASB in which a comparison was made between women who received antibiotic therapy and women who received placebo. In that study, no difference was seen in serum creatinine levels after a mean follow-up of 2 years.

In conclusion, the hypothesis that ASB will lead to renal function deterioration in women with DM can be rejected because no difference in renal function development, in either women with type 1 DM or those with type 2 DM were found. Also, the incidence of hypertension was not increased when comparing women with ASB versus women without ASB. Therefore, at this time, screening andsubsequent treatment for ASB are not indicated in patients withDM.
