Introduction

Urinary tract infection (UTI) is one of the most common microbial infections worldwide, with a global prevalence of 10 per 1,000 peoples. UTI is associated with considerable societal cost, significant morbidity, and even increasing antibiotic resistance, a current challenge for infection control1,2.

The route of infection is mainly through ascending infection from the distal urethra colonized by intestinal source microbes. The principal pathogens are Gram-negative bacteria, particularly the species of Escherichia coli accounting for 65~75% of all urinary tract infections, and the remaining are caused by Gram-positive bacteria or fungi2.

UTIs are categorized into two forms: uncomplicated UTI and complicated UTI, owing to different pathophysiology and management. Uncomplicated UTI is defined as individuals with UTI who are otherwise healthy and without any structural or neurological urinary tract abnormality that predisposes them to infection. In contrast, complicated UTI is associated with underlying factors that compromise urinary tract or host defense giving rise to predisposition to UTI, such as urinary obstruction caused by calculus or tumor, urinary retention by neurological disease, decreased urine output due to renal failure, renal transplantation, immune compromise or suppression, pregnancy, and indwelling catheter2.

Interestingly, recent studies found that the strains of E. coli causing UTIs, called extra-intestinal pathogenic E. coli (ExPEC), were distinct from the intestinal pathogenic strains and normal commensal strains, except some UTIs caused by the commensal E. coli in the complicated form (Table 1)3,4. Furthermore, analyses showed the isolates of E. coli from urine specimens of UTI patients were closely related to, if not indistinguishable from the pathogenic E. coli of poultry and pork5,6,7,8. This implicates that food reservoirs and foodborne transmission may play a role in the dissemination of ExPEC. As vegetarians avoid meat (including poultry and pork), we hypothesize that a vegetarian diet may be associated with a lower risk of UTI.

Table 1 The pathogenic potential of the three main different groups of E. coli strains3,4.
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Materials and Methods

Study design and population

The Tzu Chi Vegetarian Study (TCVS) is a prospective cohort study that recruited 12062 Tzu Chi volunteers in 2005 throughout communities in Taiwan. Tzu Chi volunteers are devoted Buddhists who participate in a variety of charity works and disaster relief organized by the Buddhist Compassion Relief Tzu Chi Foundation. About one-third of this population became vegetarians in order to protect the environment and animals. Certified Tzu Chi volunteers are also required to quit smoking and alcohol-drinking.

The Buddhists Compassion Relief Tzu Chi Foundation has many liaison sites throughout Taiwan. In 2005, community leaders in all sites were asked to distribute research questionnaires to their local community volunteers, and send back these questionnaires upon completion. The questions included personal data, demographic information, educational level, medical history, and lifestyle habit including smoking, alcohol-drinking, physical activities, and diet. All participants had signed the informed consent on the first page of the questionnaire. This study was approved by the Institutional Review Board for ethics at Buddhist Dalin Tzu Chi Hospital.

Assessment of disease status

We followed the TCVS participants from 2005 to 2014 by linking the baseline data to the National Health Insurance Research Database (NHIRD) and the death records at the National Health and Welfare Data Science Center (HWDC), Ministry of Health and Welfare.

In Taiwan, the National Health Insurance Program is a governmental universal health insurance program covering nearly 100% of the population. Individuals’ medical claim data including diagnosis from both inpatient and outpatient are available. In order to protect the privacy of individuals as required by local law, personal identification number is masked and all analyses must be performed within the HWDC. Only summarized research results (not individual data), could be released.

Dietary assessment

The diet section in the questionnaire was adopted from the food frequency questionnaire (FFQ) used in Nutrition and Health Survey in Taiwan, and included 57 items on food and food groups, and a section on cooking methods and cooking oil. A similar questionnaire had been validated later and showed good reliability and validity among Tzu Chi volunteers9.

Participants were categorized as vegetarians if they (1) self-identified as vegetarians in a question asking vegetarian status, and (2) reported “no eating” in frequency questions for all individual meat and fish items in the FFQ. Those who self-identified as vegetarians but reported eating meat or fish in the FFQ were categorized as non-vegetarians.

Exclusion criteria

Individuals with age < 20 years old (n = 85), no available data on NHIRD (n = 677), incomplete data on the questionnaire (n = 950), and prior history of UTI (n = 626) were excluded from this study (Fig. 1).

Figure 1
figure 1

Chart showing the flow from TCVS to eligible study population and UTI case identification.

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Case ascertainment

Incident cases of UTI were identified by the International Classification of Diseases, Ninth Revision (ICD-9) for UTIs and the codes for the execution of bacterial culture and prescription of antibiotics against UTI within the HWDSC database during follow-up.

The ICD-9 codes used include 590 for infections of the kidney, 595 for cystitis, and 599.0 for UTI, site not specified. The code for the execution of bacterial culture is 13007C. We identified UTI cases by the above ICD-9 diagnosis in either inpatient or outpatient, accompanied by the execution of bacterial culture and prescription of antibiotic against UTI10,11 (Fig. 1).

Statistical analysis

Baseline characteristics of vegetarians and non-vegetarians were compared using Chi-square (χ2) test for categorical variables and independent sample t test for continuous variables.

Cox regression was used to analyze the association between different factors and risks of UTI, presented as hazard ratio (HR) with 95% confidence interval (CI). Model 1 adjusted for age and/or sex. Model 2 additionally adjusted for educational level and lifestyle factors (smoking and alcohol-drinking). Model 3 additionally adjusted for hypertension, diabetes mellitus, and hyperlipidemia. Model 4 additionally adjusted for disease conditions that predispose to complicated UTIs, including urinary tract obstruction caused by hyperplasia or tumor, urolithiasis (calculus), urine retention, renal failure, immune dysfunction, pregnancy, and urinary catheter indwelling. Kaplan-Meier curves of crude data were plotted to show the disease-free survival throughout follow-up period. Subgroup analyses by the above factors and test of interactions were performed. In addition to total UTI-combined, we also examined subtypes of UTI (complicated and uncomplicated) separately.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee (Institutional Review Board [IRB] for ethics at Buddhist Dalin Tzu Chi Hospital with IRB approval document numbers: B09401003 and B10403020, grant numbers: TCMMPSP104-08-02, TCMMP105-13-05, TCMMP106-04-01) and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Results

Baseline characteristics

Table 2 shows the baseline characteristics of both vegetarian and non-vegetarian groups. The ratio of non-vegetarians to vegetarians was about 2:1. The vegetarian group was older, had a higher proportion of female, received less education, was less likely to have a history of smoking or alcohol-drinking, and had a significantly lower proportion of individuals with hypertension, diabetes, hyperlipidemia, and disease conditions that predispose to UTI.

Table 2 Baseline characteristics of vegetarian and non-vegetarian groups.
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Risk factors associated with UTI

In the 10 years of follow-up, 661 individuals were diagnosed with UTIs. Table 3 shows the Cox regression analysis. Overall, UTI risk in vegetarians was significantly lower than non-vegetarians (HR = 0.84, 95% CI: 0.71–0.99, p = 0.038). Age, female sex, diabetes, urine retention, and renal failure were associated with significant increases in UTI risks. Hypertension, urinary obstruction (hyperplasia/tumor of the urinary tract), urolithiasis, and immune dysfunction were associated with non-significant higher UTI risks.

Table 3 UTI risks associated with vegetarian diet and other factors by Cox regression.
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Subgroup analysis

Subgroup analysis of the association between vegetarian diet and UTI by sex in various models were shown in Table 4. Vegetarian diet was significantly associated with a reduced risk of UTI mainly in female, particularly after adjustment for education and lifestyle (smoking and alcohol-drinking) in Model 2. Additional adjustments for hypertension, diabetes, hyperlipidemia, and conditions predisposing to UTI (as in Model 3 and 4) yielded similar results. Kaplan-Meier curves of UTI-free survival over the ten-year period for vegetarians and non-vegetarians (Fig. 2) highlight the protection of vegetarian diet in female, while no distinct difference was found in male.

Table 4 Hazard ratios (95% confidence interval) of UTI in different dietary and sex groups with different adjusted models by Cox regression.
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Figure 2
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Kaplan-Meier curve of crude data over the ten-year period for vegetarians and non-vegetarians.

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Additional subgroup analyses and the test for interactions are presented in Table 5. The test of interaction between dietary patterns and smoking was significant (P-interaction = 0.029), with the protection mainly among never smokers. No significant test of interaction was found between diet and other factors (sex, educational level, alcohol-drinking, hypertension, diabetes, hyperlipidemia, and disease conditions predisposing to UTI).

Table 5 Subgroup analysis of UTI risks in vegetarians versus non-vegetariansSubgroups.
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The association between vegetarian diet and subtypes of UTI is shown in Table 6. Vegetarian diet was associated with significantly reduced risk of mainly the uncomplicated UTI but not complicated UTI.

Table 6 UTI risks of vegetarian diet in uncomplicated and complicated categories compared with non-vegetarian group by Cox regression.
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Discussion

In this prospective cohort study, vegetarian diet (versus non-vegetarian diet) is associated with a lower risk of UTI particularly in female, non-smokers, and mainly for the uncomplicated-subtype of UTI. To the best of our knowledge, this is the first study that evaluated the prospective association between a vegetarian diet and UTI.

Previous research suggested that meat-related foodborne ExPECs may be the main cause of uncomplicated UTIs and a half of complicated UTIs (Table 1)2,3,4,5,6,7,8. Consistent with this finding, we found that the protective association between vegetarian diet and UTI risk was mainly in uncomplicated rather than complicated UTI, and this association is independent of diseases and risk factors predisposing conditions to complicated UTI including hypertension12, diabetes13, and hyperlipidemia14. This suggests that the effect of vegetarian diet is not mediated through these related diseases. The test of interaction showed that vegetarian diet (versus non-vegetarian diet) was associated with protection among the never smoking group, but not the ever smoking group (P-interaction = 0.029, Table 5). It is possible that smoking compensates the benefit of vegetarian diet by increasing blood pressures (which predisposing to renal failure)12,15, renal stone formation16, and urinary tract cancers17. All of which are also the disease conditions predisposing to complicated UTI2,3,4. However, the number of smokers in our study is small and the significant interaction could arise from chance that further confirmation from other studies is needed.

The principal pathogenic strains causing UTIs are ExPECs (accounting for 65~75% UTIs), a group of strains distinct from the intestinal pathogenic and normal commensal E. coli3,4. ExPECs express many characteristic virulent factors which are not present in other strains of non-ExPECs2,3,4,18. Many studies have illustrated that meat, including poultry and pork, as the major reservoirs for ExPECs5,6,7,8,19. Vincent et al. identified the E. coli strains by using O:H serotype and sequence type (ST) and discovered the O25:H4-ST131 and O114:H4-ST117 strains acquired from retail chicken were very similar to, if not indistinguishable from E. coli of UTI patients5. Jacobsen et al. performed the phylogenetic analysis by triplex polymerase chain reaction using three DNA markers showed the UTI-associated B2 and D phylogroups are found among isolates from broiler chicken meat, broiler chickens, pork, and pigs. Further, antimicrobial resistance phenotypes of E. coli from meat were very similar to isolates from UTI patients7. In addition, sequencing analysis of ExPECs showed UTI-causing ST10, ST69, ST95, ST117, and ST131 are associated with poultry and retail poultry meat8,19. Vegetarians do not consume meat and therefore avoid uptake of animal-food-associated ExPECs.

Recent studies discovered the gut and fecal microbiotas were different between vegetarians and non-vegetarians20,21,22,23. The high fiber contents in vegetarian diet may modulate the intestinal microbiota in humans20,21. Fiber is metabolized by intestinal anaerobic microflora to produce short-chain fatty acids, which decrease intestinal pH20,21. Vegans or vegetarians were shown to have lower total counts of E. coli and Enterobacteriaceae spp. and lower stool pH than non-vegetarians, and the lower stool pH could prevent the growth of E. coli and Enterobacteriaceae22. The shifts in the gut and fecal microbiotas in vegetarians may potentially contribute to the protection from UTI.

Vegetarians consume predominantly plant foods and with more frequent consumption of vegetables, fruits, and nuts than non-vegetarians in most cohorts24,25, as in our cohort study, published previously26. Briefly, vegetarians in this present study had previously been shown to have a distinct overall dietary pattern, with a more frequent consumption of a variety of vegetables, fruits, nuts, soy products and beans, while avoiding all types of meat, fish, and seafood, compared with non-vegetarians26. Plant foods contain phytochemicals (major classes including terpenoids, phenolics, alkaloids) which have been demonstrated to have antibacterial activities27,28,29,30,31,32,33,34,35,36,37,38,39,40, in addition to anti-carcinogenic, anti-mutagenic, anti-inflammatory, and anti-oxidative factors27. Among the phytochemicals, phenolics compounds are related to the antibacterial activities27,28,29,30,31,32,33,34,35,36,37,38,39,40, and are abundant in vegetables, whole grains, fruits, and nuts28,29, and some of which had been demonstrated with antimicrobial effect27,30,31,32,33,34,35,36,37,38,39,40. Cranberry (Vaccinium macrocarpon), belong to the Ericaceae family, is one of the most well-known fruit against UTI as evidenced in many trials30,31,32,33,34,35,36. Besides cranberry, fresh juice (berry or fruit) have been was associated with a decreased risk of recurrence of UTI (odds ratio: 0.66, 95% CI: 0.48–0.92, per 2 dL juice)33. Proanthocyanidins (PAC), a member of tannins belong to phenolics found in cranberry and other fruits may defend against microbes30. PAC inhibits the adhesion of P fimbriae in vitro via conformational changes and decrease in adhesion forces30,34,36, and this in vitro anti-adherence effect is dose-dependent30. Moreover, fructose found in fruits may also inhibit the adherence of type 1 fimbriae30. These fimbriae-mediated adhesions are the key step for infection2. In addition to ExPEC, in vitro studies have shown that cranberry may inhibit adherence of other common UTI pathogens including Klebsiella pneumoniae, Staphylococcus aureus, Proteus spp., Pseudomonas aeruginosa, and Enterococcus faecalis30,36. Clinical research suggests the dose of cranberry as follows30: (1) cranberry juice cocktail 240–300 ml daily (preventing 50% of recurrences of UTIs and reducing bacteriuria), (2) dried, concentrated juice extract 600 to 1,200 mg per day divided into two or three daily doses, or (3) total 36 mg of PAC in twice-daily manner. In addition to cranberry, other berries and fruits, such as elderberry (Sambucus nigra), blueberry (Vaccinium corymbosum), strawberries (Fragaria ananassa), blackberries (Rubus spp.), red raspberry (Rubus idaeus), blackcurrant (Ribes nigrum), and redcurrants (Ribes rubrum), also contain phenolics and may have similar properties against p-fimbriated E. coli37. Another plant with studies showing an effect against UTI is roselle (Hibiscus sabdariffa)38,39,40. Roselle, commonly consumed in the form of tea or jam in Taiwan, belongs to the Malvaceae family and is rich in phytochemical phenolics especially anthocyanins38. Alshami et al. had demonstrated the antimicrobial activity of roselle extract against uropathogenic E. coli, K. pneumoniae, and Candida albicans isolated from recurrent urinary tract infection, through inhibiting the biofilm forming capacity39,40. Moreover, commonly consumed nuts including almond, Brazil nut, cashew, chestnut, hazelnut, heartnut, macadamia, peanut, pecan, pine nut, pistachio, and walnut also contain a variety of phytochemicals including PAC, although there is no study demonstrating the antibacterial effect so far41. Studies about medicinal plants had shown many of them with broad spectrum antimicrobial activity against uropathogenic E. coli and other UTI pathogens42,43. However, the component contributing to the effect remains to be investigated. In sum, vegetarian diets comprise abundant phytochemicals, which may contain anti-microbial properties and protect against UTI.

ExPECs isolated from animal food products and UTI patients are very similar in antibiotic resistant patterns and virulence factor profiles5,6,8,44,45. Antibiotics use in animal agriculture may contribute to the increasing antibiotic resistance in humans44. The extended spectrum β-lactamase (ESBL)-producing and fluoroquinolone-resistant ExPECs, most of which are globally disseminated E. coli strain O25:H4-ST131 (accounting for 78%), are a major problem with resistance to penicillins, cephalosporins, and fluoroquinolones2,8,46. Moreover, the ESBLs encoded on plasmids typically carry other antibiotic resistance genes against aminoglycosides, sulfonamides, and quinolones, leading to multidrug resistant2. The high recurrence rates and increasing multidrug resistance in ExPECs2 make antibiotic treatment of UTI challenging. Therefore, alternative non-antibiotic management for UTI is important. High phytochemical content in a healthy vegetarian diet may provide an alternative prophylaxis from and bactericidal effect against UTI27,30,31,32,33,34,35,36,37,38,39,40, and a study had demonstrated the PAC from cranberry also exhibited the anti-adherence property in the multidrug resistant strains of uropathogenic P-fimbriated E. coli47.

Several limitations must be noted in our study. (1) Our case definition is based on ICD-9 code rather than the gold standard, which should be clinical symptoms and laboratory examination including pyuria and urine culture with a more than 100,000 colony forming units per milliliter48,49. (2) Residual confounding may still remain as several known confounders, including water intake50, sexual intercourse33,51, intake of honeydew melon (which may also contain ExPEC5, though evidence is insufficient), were not measured. (3) Although ExPEC is found mainly from meat sources, one study reported the detection of ExPEC isolates from a honeydew melon sample of restaurant/ready-to-eat foods in North America5. To the best of our knowledge, E. Coli or ExPEC had never been reported in plant based foods in Taiwan. The sporadic case of ExPEC in non-meat product may be due to contamination and will require further research. However, if plant-based foods were a major source of ExPEC contributing to UTI, the protective association between vegetarian diet and UTI would have been attenuated, strengthening the null association. (4) Another limitation lies in our FFQ, which did not ask specifically about intake of honeydew melon, cranberries, or roselle. Thus, we were unable to conduct further analysis on the association between UTI and these plant foods. In addition, the FFQ was self-administered with some degree of missing values, that it is difficult to calculate the exact intake of specific phytochemicals known to contribute to UTI-protection.

In conclusion, vegetarian diet is protectively associated with UTI particularly in female and for uncomplicated UTI. Further study with identification of pathogens from urine culture is needed to clarify the relationship among UTI risk, pathogens, and vegetarian diet.