Helicobacter pylori infection in rural China: demographic, lifestyle and environmental factors

Linda Morris Browna, Terry L Thomasa, Jun-Ling Mab, Yun-Sheng Changb, Wei-Cheng Youa, Wei-Dong Liuc, Lian Zhangc, David Peed and Mitchell H Gaila

a Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
b Beijing Institute for Cancer Research and School of Oncology, Beijing University, Beijing, China 100034.
c Linqu Public Health Bureau, Linqu, Shandong Province, China 262600.
d IMS, Inc., Rockville, MD 20852, USA.

Linda Morris Brown, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Executive Plaza South, Room 8026, 6120 Executive Blvd. MSC 7244, Bethesda, MD 20892–7244, USA. E-mail: BrownL{at}mail.nih.gov


    Abstract
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Background Although Helicobacter pylori is one of the most common human bacterial infections worldwide, its mode of transmission is unclear.

Methods To investigate possible associations between H. pylori infection and demographic, lifestyle, and environmental factors in a rural Chinese population, a cross-sectional survey was administered to 3288 adults (1994 seropositive, 1019 seronegative, 275 indeterminate) from 13 villages in Linqu County, Shandong Province, China.

Results Helicobacter pylori prevalence was elevated for: infrequent handwashing before meals (OR = 1.7, 95% CI: 1.0–3.0), crowding (i.e. sharing a bed with >2 people [OR = 2.3, 95% CI: 1.3–4.2]), washing/bathing in a pond or ditch (OR = 1.5, 95% CI: 1.0–2.4), and medium (OR = 1.6, 95% CI: 1.3–2.0) and low (OR = 2.3, 95% CI: 1.9–2.9) compared to high village education level, and reduced for never being married or divorced (OR = 0.4, 95% CI: 0.2–1.0). There was also a suggestion that source of drinking water, especially water from a shallow village well might be related to H. pylori seropositivity. There was no evidence of an association between H. pylori prevalence and alcohol or tobacco use, raw fruit and vegetable intake, or individual social class measures.

Conclusions The results of this study suggest that person-to-person transmission is the most plausible route of H. pylori infection in this rural Chinese population, but waterborne exposures deserve further investigation.

Keywords Helicobacter pylori, risk factors, aetiology, transmission

Accepted 22 October 2001


    Introduction
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Helicobacter pylori is one of the most common human bacterial infections worldwide, but its mode of transmission is unclear. A cross-sectional survey of the 3288 adults aged 35–69 enrolled in an intervention trial in Linqu County, Shandong Province, China was conducted in 1997–1998 to assess possible risk factors that may be associated with H. pylori infection in this rural area of China, which has one of the highest rates of gastric cancer in the world,1 and a greater than 60% prevalence of H. pylori.1,2 Although a high percentage of the children in this area are known to be infected with H. pylori (50% by age 3 and 67% by age 11),3 preliminary data indicate that seroconversions have occurred over the course of the intervention trial suggesting the possibility of new or continued infection in this cohort. A previous analysis of data from the cross-sectional study found no association between H. pylori prevalence and exposure to animals during childhood or adulthood and no indication of zoonotic transmission.4 This paper explores other determinants of transmission in this high-risk area of rural China and whether an association exists between H. pylori seropositivity and certain demographic, environmental, and lifestyle factors.


    Methods
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Study population
This study is a cross-sectional survey of 3288 H. pylori infected and uninfected adults from Linqu County, Shandong Province, China enrolled in a joint US National Cancer Institute (NCI)/ Beijing Institute for Cancer Research (BICR) double-blinded population-based randomized intervention trial to inhibit progression of precancerous gastric lesions. A description of the intervention trial study population and methodology have been presented in detail elsewhere.5 In brief, all residents aged 35–69 in 1994, from 13 Linqu villages were invited to participate in a screening programme that included interview, gastroscopy, gastric biopsy, and phlebotomy. Of the 4035 possible participants, 210 subjects were excluded from the study because they were not medically eligible for endoscopy and 226 subjects were excluded because they refused endoscopy. A total of 3599 individuals completed the procedures and were eligible to participate in the intervention trial. Thirty-nine subjects refused to participate and 149 subjects were excluded because they were deceased (14), ‘out-of-scope’ for age or village (42), had a history of allergy to antibiotics (67), or they were not tested for antibodies to H. pylori (26). The remaining 3411 subjects (93% of all eligible adults in the 13 villages) were enrolled in the trial in September 1995.

Description of area
Shandong Province is one of the least economically developed provinces in China and Linqu County is one of the poorest counties in Shandong, with three-quarters of the land located in mountainous regions resulting in infertile soil and periodic severe droughts. The annual stomach cancer mortality rates for Linqu County are 70 per 100 000 for men and 26 per 100 000 for women.1 The 13 rural villages range in size from 642 to 1845 inhabitants (mean 800). Farming is the major occupation and source of income for residents and variation in the socioeconomic status of the villages is due to differences in the ability of the land to support farming. There is also variation in the source of water, with one village using a public well, other villages using private wells, and still others using running water piped in from a village water tower. Most of the villages consist of 3–5 room brick houses built with floors of either dirt, cement, or brick. Food is stored in one room of the house. All homes have electricity, most have a small black and white TV set, and a few have telephones; however, they lack other modern conveniences such as refrigerators, washing machines, and indoor plumbing. The cooking area and pit privies/outhouses are located outside the main house.

Interview and questionnaire
To study the aetiology of H. pylori, in-person home interviews lasting approximately 15 minutes were conducted from October 1997 to May 1998 by trained BICR field staff. Informed consent to participate in the study was obtained from each subject prior to interview. Interviews were completed with 3288 (96%) of the 3411 study subjects enrolled in the intervention trial, and 91% of all eligible adults in the 13 villages. Reasons for non-response included death (54 subjects; 1.6%), dropped out of trial (33 subjects; 1.0%), lost to follow-up (10 subjects; 0.3%), and refusal (26 subjects; 0.8%). Neither the interviewers nor the interviewees were aware of the H. pylori sero-status of the study participants. The questionnaire sought information on adult socioeconomic factors, personal hygiene, and exposures during adulthood related to food, water, tobacco, alcohol, and animals. The questionnaire also collected information on certain childhood exposures related to domestic animals, household composition, type of flooring, and oral contact (eating premasticated food and being kissed by their parents on the lips). Data on village education were ascertained separately and used to group the 13 villages into three categories based on the average number of years of schooling residents had completed: high (range 3.99–3.83 years, 4 villages), medium (range 3.74–3.37 years, 4 villages), and low (range 3.27–2.32 years, 5 villages). Questionnaire data were merged with H. pylori serology test results (used as the measure of H. pylori infection) and information on intake of garlic and other allium vegetables obtained from the same subjects in a separate survey. The study was approved by Institutional Review Boards at BICR, Westat, Inc., and NCI.

Blood collection and Helicobacter pylori serological assays
In 1994, a 5-ml blood sample was collected from eligible subjects. Serum was separated and aliquoted in the field, stored immediately at –20°C, and then transferred to a –70°C freezer at BICR. One aliquot of serum (0.5 ml) was tested by one of us (LZ) for IgG and IgA antibodies to H. pylori.2 Helicobacter pylori strains cultured from gastric biopsies of two patients in Linqu County were used to provide the antigenic preparation for serology because it had been suggested that assays based on indigenous strains had higher sensitivity and specificity.6 Based on data from the intervention trial pilot study, the sensitivity and specificity of the two-strain Chinese assay were 90% and 87%, respectively, compared to the 13C urea breath test ‘gold standard’. All assays were performed on coded samples in duplicate and then repeated. The averages of the duplicate values for each of the two repeated assays were recorded. Cutoff values were based on examination of the distribution of readings in relation to a group of uninfected people and reference sera.2 A subject was considered seropositive if both IgG optical density readings were >=1.1 and seronegative if both IgG values were <=0.9 and neither IgA value was >=1.0. Otherwise, subjects were categorized as indeterminate. The H. pylori serostatus of the participants was positive for 1994 subjects (60.6%), negative for 1019 subjects (31.0%), and indeterminate for 275 (8.4%).

Data analysis
The measure of association used in this analysis is the prevalence odds ratio comparing the odds of being exposed to a factor given the subject was H. pylori seropositive to the odds of being exposed to the same factor given the subject was H. pylori seronegative. Maximum likelihood estimates were computed for seropositive and seronegative subjects using logistic regression.7 Subjects with an indeterminate H. pylori serostatus were excluded from the analysis. Standard calculations of variances based on the logistic model that assumes that responses are independent, conditional on covariates, may be misleading in this application because 62% of the study subjects came from households with more than one participating member. Of the 2026 households represented in the study, 1110 had one participating member, 856 had two members, 49 had three members, and 11 had four members. Therefore 100 bootstrap replicates based on resampling households with replacement were used to estimate the needed variances and covariances. To control for potential confounding, age was included as a continuous variable in all logistic models. Multivariate analyses were also conducted and included age plus selected variables with significantly increased or decreased risks in the simpler models. Only the crowding-related variable ‘number of people shared a bed with’ was included in the multivariate model because it was highly correlated with the variable ‘number of children in the household’. For each variable we present the age-adjusted prevalence odds ratio (OR), the 95% CI computed directly from the logistic regression, and/or the bootstrap 95% CI that allows for additional familial correlation. A total of 1514 males and 1499 females are included in the analyses presented. Approximately 23% of the population was aged 35–39, 26% was 40–44 years, 27% was 45–54 years, and 23% was aged 55–69.


    Results
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
The per cent H. pylori seropositive decreased somewhat by age ranging from 68.8% in those aged <40 years to 64.9% in those aged 45–54 to 63.3% in those aged >=60. In addition, there was significant variation in H. pylori serostatus by village, ranging from 51.3% to 79.6% ({chi}2 = 120.76, d.f. = 12, P < 0.001).

The association between social factors and prevalence of H. pylori infection is presented in Table 1Go. Helicobacter pylori seroprevalence was significantly associated with village education level. The odds of being H. pylori positive increased with decreasing village education level, from 1.0 for high (referent category), to 1.7 (95% CI: 1.4–2.1) for medium, to 2.4 (95% CI: 2.0–3.0) for low. There was a corresponding increase in per cent seropositive, from 54.4% to 67.2% to 74.3%. The percentage of residents with greater than a 6th grade education was 27%, 22%, and 15% for villages classified as high, medium, and low, respectively. No associations were seen for subject's level of education or annual family income. Median annual family income was 4000 yuan or approximately US$500.


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Table 1 Association between prevalence of Helicobacter pylori infection and social factors
 
Table 2Go depicts the association between H. pylori infection and crowding/density factors and hygiene practices. The odds of being H. pylori positive were significantly reduced for subjects who were never married (OR = 0.4, 95% CI: 0.4–0.9), and there was a similar but non-significant reduction for the small number who were divorced. The per cent seropositive was 46.9% for never married and 40.0% for divorced compared to 66.5% for never married. Having more than one child in the household (OR = 2.0, 95% CI: 1.0–3.7) and sharing a bed with >2 people (OR = 2.1, 95% CI: 1.1–3.8) were significantly associated with the odds of being H. pylori seropositive. The per cent H. pylori seropositive was almost 80% for each of these factors. The OR was also elevated for subjects who reported washing their hands before eating less than half the time (OR = 1.6, 95% CI: 1.0–2.5; % seropositive = 74.4%) or never (OR = 3.8, 95% CI: 0.5–31.0; % seropositive = 87.5%). The odds of being H. pylori positive were reduced for subjects who shared cups with their family less than half the time, but no consistent patterns were seen with frequency of washing hands or body with soap, or frequency of cup washing after use (data not shown). No significant associations were noted between prevalence of H. pylori infection as an adult and childhood exposures (i.e. crowding or density factors, type of flooring, and oral contact) (data not shown).


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Table 2 Association between prevalence of Helicobacter pylori infection and crowding/density factors and hygiene practices
 
Source of drinking water was found to vary between H. pylori positive and negative subjects (Table 3Go) with the highest risk (OR = 1.8, 95% CI: 1.4–2.3) and per cent seropositive (72.2%) observed for subjects who obtained their water from a shallow village well. An elevated OR was also associated with washing or bathing in a pond or ditch when the weather was warm (OR = 1.6, 95% CI: 1.0–2.4; % seropositive = 71.5%).


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Table 3 Association between prevalence of Helicobacter pylori infection and water-related factors
 
Table 4Go presents prevalence OR for H. pylori infection and smoking, drinking, and dietary-related factors. The odds of being H. pylori seropositive among subjects who ever smoked cigarettes was slightly decreased (OR = 0.9, 95% CI: 0.7–1.0) compared to that for never-smokers, but there were no trends with lifetime number of packs smoked. There was also no association between infection status and alcohol use overall or with any measure of alcohol consumption including number of times the subject drank per week. There was a pattern of decreasing OR with increased consumption of allium vegetables (ranging from 1.0 for <5.5 kg per year to 0.8 for >10.5 kg per year), but the trend was not significant. There was no apparent association between H. pylori infection and the number of times per month raw fruits and vegetables were consumed. Overall, there was no association between ever having gastric reflux and prevalence of H. pylori infection. However, the OR and the per cent seropositive were slightly but non-significantly reduced for subjects who had reflux nine or more times per month (OR = 0.8, 95% CI: 0.6–1.1; % seropositive = 61.4%).


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Table 4 Association between prevalence of Helicobacter pylori infection and smoking, drinking, and dietary factors
 
Table 5Go presents OR for variables of interest adjusted for one another in multivariate logistic models using bootstrap techniques to calculate CI. The OR for subjects who obtained their water from a shallow village well was significantly elevated in Model I (OR = 1.8, 95% CI: 1.4–2.3), but was no longer significantly elevated in Model II (OR = 1.2, 95% CI: 0.9–1.6) when village education level was included. The adjusted OR for village education level was 1.6 (95% CI: 1.3–2.0) for subjects living in a village classified as medium and 2.3 (95% CI: 1.9–2.9) for those living in a village classified as low. Odds ratios in Model II were also elevated for subjects who shared a bed with >2 people (OR = 2.3, 95% CI: 1.3–4.2), for those who washed their hands before meals less than half the time (OR = 1.7, 95% CI: 1.0–3.1), and for those who washed in a pond or ditch (OR = 1.5, 95% CI: 1.0–2.4). The OR was reduced for those who never married or were divorced (OR = 0.4, 95% CI: 0.2–1.0).


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Table 5 Association between prevalence of Helicobacter pylori infection and selected factors in a multivariate logistic model
 

    Discussion
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
This large cross-sectional survey in Shandong Province, China provided us the opportunity to study everyone in the population aged 35–69 to determine what factors were related to H. pylori positivity. Social class factors have been associated with H. pylori infection status in a number of studies throughout the world.8–18 In this study, we found a strong inverse relationship between village education level and prevalence of H. pylori infection, but no association with subject's education level and annual family income. It is possible that average village education level in rural China may reflect general lifestyle or hygiene practices that vary substantially between villages. For example, there was some suggestion in our data that source of drinking water may be related to H. pylori infection, with the greatest risk observed for subjects who obtained their water from a shallow village well. However, since water source is highly correlated with village education level (e.g. the poorest and least educated villages obtain their water from a shallow village well) it is possible that some of the excess risk seen for medium and low village education level may be due to differences in water source. Evidence that waterborne transmission may be important, especially in areas of the world with high rates of H. pylori infection and less than adequate water quality comes from studies conducted in Colombia, China, Peru, Japan, and Inuit communities in Canada.2,19–23

We found elevated OR associated with measures of crowded living conditions, particularly number of children in the household and the number of people sharing a bed, and a reduced risk for subjects who never married or were divorced. These findings are consistent with other recent studies10,11,19,24–30 and suggest that household transmission contributes to H. pylori seropositivity.

We sought information about the frequency as an adult of handwashing and found elevated OR associated with infrequent handwashing before meals, but no protective effect of washing hands or body with soap or detergent. A recent study in rural Guatemala found evidence of H. pylori DNA under the fingernails of infected subjects, suggesting that the hand may play a role in transmission.31 Several studies found the prevalence of H. pylori infection to be higher in subjects who did not have a bathroom, indoor toilet, or running water.26,32,33 These modern necessities were not found in any of the homes of our study subjects. Since all of our study subjects ate with chopsticks, we also could not confirm the excess risk (OR = 2.5) reported among Chinese immigrants in Australia who used chopsticks.12

Analysis of the relationship between smoking and H. pylori infection in recent studies has provided conflicting results. While some studies have reported significantly elevated risks of H. pylori infection with smoking8,30,34–38 and one reported significantly reduced risks,39 most, similar to ours, found no significant association with current smoking or any other measure of cigarette use.2,12,14,14,17,24,40–45 Recent epidemiological studies that looked at the relation between alcohol use and H. pylori infection reported either no association11,12,14,14,18,36,41 or a reduction in risk8,17,37,39,46–48 that was stronger for wine than beer and more apparent at moderate to high levels of alcoholic beverage consumption. Similarly, our study found no association between H. pylori infection and ever/never use of alcohol, and only a slight (OR = 0.9) non-significant reduction in the OR for subjects who drank alcohol >=7 times per week. The relative homogeneity among drinkers in our study, including the lack of heavy drinkers, may have limited our ability to detect a protective effect from alcohol's antimicrobial properties.49 The use of serostatus as a marker of infection could also mask a potential association of alcohol consumption and H. pylori infection as antibodies are still circulating in the blood after accidental H. pylori loss. Increased consumption of allium vegetables has been suggested as a protective factor for stomach cancer50,51 and its precursor lesions,52 and garlic has been shown to inhibit H. pylori in vitro,53,54 but not in vivo.55 In our study, we found a pattern of decreasing OR with increased consumption of all allium vegetables combined, although the trend was not significant. Several studies have reported significantly reduced OR and negative gradients in risk of H. pylori infection with increased consumption of fruits and/or vegetables, vitamin C, and beta-carotene.19,37,56,57 In contrast, consumption of raw/uncooked vegetables was associated with elevated risk of H. pylori infection.19,58 In our data, there was no apparent association between H. pylori infection and the number of times per month raw fruits and vegetables were eaten.

Some studies have suggested that H. pylori infection appears to be lower in people with gastroesophageal reflux disease (the major risk factor for Barrett's oesophagus which is strongly associated with adenocarcinoma of the oesophagus) than in controls.59,60 In our study, we found slight non-significant protective effects for subjects who reported the occurrence of gastric reflux nine times or more per month.

The major strengths of this cross-sectional investigation are that the data were sought from all eligible individuals age 35–69 in 13 rural Chinese villages, participation rates were high, and H. pylori status (unknown by study investigators, interviewers, and subjects) was determined objectively based on the ELISA optical density test utilizing an indeterminate zone (± 10% of the cutoff value of 1.0). In addition, the same structured questionnaire was administered to all study subjects by trained interviewers and bootstrap CI were calculated to avoid underestimation of the standard errors due to possible intrafamilial correlations. Although there may be some degree of misclassification, inaccuracies in data acquisition are not likely to differ systematically according to H. pylori serostatus because of the blinded design.

There are also several limitations of this study. The outcome was measured as prevalence of disease and thus reflects H. pylori infections that could have occurred from early childhood to several weeks before the blood sample was drawn; the suboptimal sensitivity and specificity (90% and 87%, respectively) of the serological test would tend to attenuate the risk estimates making it more difficult to find associations; the outcome and the exposure were assessed simultaneously, making it impossible to determine temporal relationships; and the high degree of homogeneity in this rural Chinese population may have limited the ability to detect differences between H. pylori positive and negative subjects.

In summary, our data from this rural Chinese population suggest a possible association between H. pylori and village education level, marital status, crowding, lack of handwashing, washing/bathing in a pond or ditch, and source of drinking water, but no evidence of an association between H. pylori and alcohol or tobacco use, raw fruit and vegetable intake, or individual social class measures. Based on these findings, person-to-person transmission appears to be the most plausible route for H. pylori infection in this rural Chinese population, but waterborne exposures deserve further investigation.


    Acknowledgments
 
This work was supported in part by National Cancer Institute Contract NO1-CP-71103. The authors wish to thank John Heinrich of Westat, Inc., Rockville, MD, USA and Stella Semiti of Information Management Systems, Rockville, MD, USA for technical assistance and Drs David Cruess, Andre Dubois, and Gary Gacksteller of the Uniformed Services University of the Health Sciences, Bethesda, MD, USA for their valuable guidance in the analysis of data from this investigation.


    References
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
1 You WC, Blot WJ, Li JY et al. Precancerous gastric lesions in a population at high risk of stomach cancer. Cancer Res 1993;53:1317–21.[Abstract]

2 Zhang L, Blot WJ, You WC et al. Helicobacter pylori antibodies in relation to precancerous gastric lesions in a high-risk Chinese population. Cancer Epidemiol Biomarkers Prev 1996;5:627–30.[Abstract]

3 Ma JL, You WC, Gail MH et al. Helicobacter pylori infection and mode of transmission in a population at high risk of stomach cancer. Int J Epidemiol 1998;27:570–73.[Abstract]

4 Brown LM, Thomas TL, Ma JL et al. Helicobacter pylori infection in rural China: exposure to domestic animals during childhood and adulthood. Scand J Infect Dis 2001;33:686–91.[CrossRef][ISI][Medline]

5 Gail MH, You WC, Chang YS et al. Factorial trial of three interventions to reduce the progression of precancerous gastric lesions in Shandong, China: design issues and initial data. Control Clin Trials 1998;19:352–69.[CrossRef][ISI][Medline]

6 Bodhidatta L, Hoge CW, Churnratanakul S et al. Diagnosis of Helicobacter pylori infection in a developing country: comparison of two ELISAs and a seroprevalence study. J Infect Dis 1993;168:1549–53.[ISI][Medline]

7 Engelman L. Stepwise logistic regression. In Dixon W (ed.). BMDP Statistisal Software Manual, Vol. 2. Berkeley, CA: University of California Press, 1990, pp. 1013–46.

8 Lin SK, Lambert JR, Nicholson L, Lukito W, Wahlqvist M. Prevalence of Helicobacter pylori in a representative Anglo-Celtic population of urban Melbourne. J Gastroenterol Hepatol 1998;13:505–10.[ISI][Medline]

9 Souto FJ, Fontes CJ, Rocha GA, de Oliveira AM, Mendes EN, Queiroz DM. Prevalence of Helicobacter pylori infection in a rural area of the state of Mato Grosso, Brazil. Mem Inst Oswaldo Cruz 1998;93: 171–74.

10 Rothenbacher D, Bode G, Peschke F, Berg G, Adler G, Brenner H. Active infection with Helicobacter pylori in an asymptomatic population of middle aged to elderly people. Epidemiol Infect 1998;120:297–303.[CrossRef][ISI][Medline]

11 Peach HG, Pearce DC, Farish SJ. Helicobacter pylori infection in an Australian regional city: prevalence and risk factors. Med J Aust 1997; 167:310–13.[ISI][Medline]

12 Chow TK, Lambert JR, Wahlqvist ML, Hsu-Hage BH. Helicobacter pylori in Melbourne Chinese immigrants: evidence for oral-oral transmission via chopsticks. J Gastroenterol Hepatol 1995;10:562–69.[ISI][Medline]

13 Replogle ML, Glaser SL, Hiatt RA, Parsonnet J. Biologic sex as a risk factor for Helicobacter pylori infection in healthy young adults. Am J Epidemiol 1995;142:856–63.[Abstract]

14 Epidemiology of, and risk factors for, Helicobacter pylori infection among 3194 asymptomatic subjects in 17 populations. The EUROGAST Study Group. Gut 1993;34:1672–76.[Abstract]

15 Malaty HM, Evans DG, Evans DJJ, Graham DY. Helicobacter pylori in Hispanics: comparison with blacks and whites of similar age and socioeconomic class. Gastroenterology 1992;103:813–16.[ISI][Medline]

16 Torres J, Leal-Herrera Y, Perez-Perez G et al. A community-based seroepidemiologic study of Helicobacter pylori infection in Mexico. J Infect Dis 1998;178:1089–94.[ISI][Medline]

17 Rosenstock SJ, Jorgensen T, Andersen LP, Bonnevie O. Association of Helicobacter pylori infection with lifestyle, chronic disease, body-indices, and age at menarche in Danish adults. Scand J Public Health 2000;28:32–40.[ISI][Medline]

18 Russo A, Eboli M, Pizzetti P et al. Determinants of Helicobacter pylori seroprevalence among Italian blood donors. Eur J Gastroenterol Hepatol 1999;11:867–73.[ISI][Medline]

19 Goodman KJ, Correa P, Tengana AH et al. Helicobacter pylori infection in the Colombian Andes: a population-based study of transmission pathways. Am J Epidemiol 1996;144:290–99.[Abstract]

20 Klein PD, Graham DY, Gaillour A, Opekun AR, Smith EO. Water source as risk factor for Helicobacter pylori infection in Peruvian children. Gastrointestinal Physiology Working Group. Lancet 1991;337: 1503–06.[ISI][Medline]

21 Hulten K, Han SW, Enroth H et al. Helicobacter pylori in the drinking water in Peru. Gastroenterology 1996;110:1031–35.[ISI][Medline]

22 Sasaki K, Tajiri Y, Sata M et al. Helicobacter pylori in the natural environment. Scand J Infect Dis 1999;31:275–79.[CrossRef][ISI][Medline]

23 McKeown I, Orr P, MacDonald S et al. Helicobacter pylori in the Canadian arctic: seroprevalence and detection in community water samples. Am J Gastroenterol 1999;94:1823–29.[CrossRef][ISI][Medline]

24 Kikuchi S, Kurosawa M, Sakiyama T. Helicobacter pylori risk associated with sibship size and family history of gastric diseases in Japanese adults. Jpn J Cancer Res 1999;89:1109–12.[ISI]

25 Rothenbacher D, Bode G, Winz T, Berg G, Adler G, Brenner H. Helicobacter pylori in out-patients of a general practitioner: prevalence and determinants of current infection. Epidemiol Infect 1997;119:151–57.[CrossRef][ISI][Medline]

26 Breuer T, Sudhop T, Hoch J, Sauerbruch T, Malfertheiner P. Prevalence of and risk factors for Helicobacter pylori infection in the western part of Germany. Eur J Gastroenterol Hepatol 1996;8:47–52.[ISI][Medline]

27 Malaty HM, Graham DY. Importance of childhood socioeconomic status on the current prevalence of Helicobacter pylori infection. Gut 1994;35:742–45.[Abstract]

28 Hammermeister I, Janus G, Schamarowski F, Rudolf M, Jacobs E, Kist M. Elevated risk of Helicobacter pylori infection in submarine crews. Eur J Clin Microbiol Infect Dis 1992;11:9–14.[ISI][Medline]

29 Mendall MA, Goggin PM, Molineaux N et al. Childhood living conditions and Helicobacter pylori seropositivity in adult life. Lancet 1992;339:896–97.[ISI][Medline]

30 Woodward M, Morrison C, McColl K. An investigation into factors associated with Helicobacter pylori infection. J Clin Epidemiol 2000; 53:175–81.[CrossRef][ISI][Medline]

31 Dowsett SA, Archila L, Segreto VA et al. Helicobacter pylori infection in indigenous families of Central America: serostatus and oral and fingernail carriage. J Clin Microbiol 1999;37:2456–60.[Abstract/Free Full Text]

32 Luzza F, Imeneo M, Maletta M et al. Suggestion against an oral-oral route of transmission for Helicobacter pylori infection: a seroepidemiological study in a rural area. Dig Dis Sci 1998;43:1488–92.[CrossRef][ISI][Medline]

33 Webb PM, Knight T, Greaves S et al. Relation between infection with Helicobacter pylori and living conditions in childhood: evidence for person to person transmission in early life. BMJ 1994;308:750–53.[Abstract/Free Full Text]

34 Zober A, Schilling D, Ott MG, Schauwecker P, Riemann JF, Messerer P. Helicobacter pylori infection: prevalence and clinical relevance in a large company. J Occup Environ Med 1998;40:586–94.[CrossRef][ISI][Medline]

35 Hamajima N, Inoue M, Tajima K et al. Lifestyle and anti-Helicobacter pylori immunoglobulin G antibody among outpatients. Jpn J Cancer Res 1997;88:1038–43.[ISI][Medline]

36 Murray LJ, McCrum EE, Evans AE, Bamford KB. Epidemiology of Helicobacter pylori infection among 4742 randomly selected subjects from Northern Ireland. Int J Epidemiol 1997;26:880–87.[Abstract]

37 Fontham ET, Ruiz B, Perez A, Hunter F, Correa P. Determinants of Helicobacter pylori infection and chronic gastritis. Am J Gastroenterol 1995;90:1094–101.[ISI][Medline]

38 Bateson MC. Cigarette smoking and Helicobacter pylori infection. Postgrad Med J 1993;69:41–44.[Abstract]

39 Ogihara A, Kikuchi S, Hasegawa A et al. Relationship between Helicobacter pylori infection and smoking and drinking habits. J Gastroenterol Hepatol 2000;15:271–76.[CrossRef][ISI][Medline]

40 Rudi J, Toppe H, Marx N et al. Risk of infection with Helicobacter pylori and hepatitis A virus in different groups of hospital workers. Am J Gastroenterol 1997;92:258–62.[ISI][Medline]

41 Shinchi K, Ishii H, Imanishi K, Kono S. Relationship of cigarette smoking, alcohol use, and dietary habits with Helicobacter pylori infection in Japanese men. Scand J Gastroenterol 1997;32:651–55.[ISI][Medline]

42 Martin-de-Argila C, Boixeda D, Canton R et al. Helicobacter pylori infection in a healthy population in Spain. Eur J Gastroenterol Hepatol 1996;8:1165–68.[ISI][Medline]

43 Fraser AG, Scragg R, Metcalf P, McCullough S, Yeates NJ. Prevalence of Helicobacter pylori infection in different ethnic groups in New Zealand children and adults. Aust N Z J Med 1996;26:646–51.[ISI][Medline]

44 Gasbarrini G, Pretolani S, Bonvicini F et al. A population based study of Helicobacter pylori infection in a European country: the San Marino Study. Relations with gastrointestinal diseases. Gut 1995;36:838–44.[Abstract]

45 Gilboa S, Gabay G, Zamir D, Zeev A, Novis B. Helicobacter pylori infection in rural settlements (Kibbutzim) in Israel. Int J Epidemiol 1995;24:232–37.[Abstract]

46 Brenner H, Rothenbacher D, Bode G, Adler G. Relation of smoking and alcohol and coffee consumption to active Helicobacter pylori infection: cross sectional study. BMJ 1997;315:1489–92.[Abstract/Free Full Text]

47 Brenner H, Rothenbacher D, Bode G, Adler G. Inverse graded relation between alcohol consumption and active infection with Helicobacter pylori. Am J Epidemiol 1999;149:571–76.[Abstract]

48 Brenner H, Berg G, Lappus N, Kliebsch U, Bode G, Boeing H. Alcohol consumption and Helicobacter pylori infection: results from the German National Health and Nutrition Survey. Epidemiology 1999;10: 214–18.[CrossRef][ISI][Medline]

49 Klontz KC. Does imbibing alcohol protect against enteric pathogens? Epidemiology 1999;10:207–09.[ISI][Medline]

50 Gao CM, Takezaki T, Ding JH, Li MS, Tajima K. Protective effect of allium vegetables against both esophageal and stomach cancer: a simultaneous case-referent study of a high-epidemic area in Jiangsu Province, China. Jpn J Cancer Res 1999;90:614–21.[ISI][Medline]

51 You WC, Blot WJ, Chang YS et al. Allium vegetables and reduced risk of stomach cancer. J Natl Cancer Inst 1989;81:162–64.[Abstract]

52 You WC, Zhang L, Gail MH et al. Helicobacter pylori infection, garlic intake and precancerous lesions in a Chinese population at low risk of gastric cancer. Int J Epidemiol 1998;27:941–44.[Abstract]

53 Sivam GP, Lampe JW, Ulness B, Swanzy SR, Potter JD. Helicobacter pyloriin vitro susceptibility to garlic (Allium sativum) extract. Nutr Cancer 1997;27:118–21.[ISI][Medline]

54 Cellini L, Di Campli E, Masulli M, Di Bartolomeo S, Allocati N. Inhibition of Helicobacter pylori by garlic extract (Allium sativum). FEMS Immunol Med Microbiol 1996;13:273–77.[CrossRef][ISI][Medline]

55 Graham DY, Anderson SY, Lang T. Garlic or jalapeno peppers for treatment of Helicobacter pylori infection. Am J Gastroenterol 1999;94: 1200–02.[CrossRef][ISI][Medline]

56 Goodman KJ, Correa P, Tengana AH, DeLany JP, Collazos T. Nutritional factors and Helicobacter pylori infection in Colombian children. J Pediatr Gastroenterol Nutr 1997;25:507–15.[CrossRef][ISI][Medline]

57 Jarosz M, Dzieniszewski J, Dabrowska-Ufniarz E, Wartanowicz M, Ziemlanski S, Reed PI. Effects of high dose vitamin C treatment on Helicobacter pylori infection and total vitamin C concentration in gastric juice. Eur J Cancer Prev 1998;7:449–54.[ISI][Medline]

58 Hopkins RJ, Vial PA, Ferreccio C et al. Seroprevalence of Helicobacter pylori in Chile: vegetables may serve as one route of transmission. J Infect Dis 1993;168:222–26.[ISI][Medline]

59 Blaser MJ. Hypothesis: the changing relationships of Helicobacter pylori and humans: implications for health and disease. J Infect Dis 1999; 179:1523–30.[CrossRef][ISI][Medline]

60 Wu JC, Sung JJ, Ng EK et al. Prevalence and distribution of Helicobacter pylori in gastroesophageal reflux disease: a study from the East. Am J Gastroenterol 1999;94:1790–94.[CrossRef][ISI][Medline]