since the new surveillance system under the new law began , all laboratory - confirmed vtec cases are reported and counted in japan . under this system , stool samples or rectal swabs are obtained from patients when the clinician suspects hemorrhagic enterocolitis due to pathogenic e. coli based on clinical symptoms such as hemorrhagic colitis . these specimens are sent to laboratories at the hospital , private companies , national institutions in each prefecture , or the niid . to maintain high levels of sensitivity and specificity to detect vtec , the protocol and training in these laboratories fall under the guidance of the niid . at these laboratories , the specimens are cultured on specific media such as chromagar o157 ( kanto co. ltd . , tokyo , japan ) or cefixime - tellurite sorbitol macconkey agar ( oxoid , unipath ltd . , hampshire , uk ) ; specific antibodies against each serotype of e. coli are used ( 3,4 ) . if the existence of pathogenic e. coli is confirmed , the ability to produce verocytotoxin from isolates is investigated by using reversed passive latex agglutination or a multiplex polymerase chain reaction assay ( 3,5,6 ) . if the production of verocytotoxin is confirmed by the laboratory , the case is considered symptomatic vtec . persons associated with the initial case ( e.g. , family members ) may be further examined for vtec at the doctor 's discretion . when a doctor diagnoses either symptomatic or asymptomatic vtec infection , he or she has to report this event to the local health center immediately and manage the cases to prevent further spread of the disease . the number of vtec cases in japan is totaled for each prefecture weekly . in this study , we used this surveillance data reported from april 1999 to october 2004 ( 287 weeks ) , which were retrieved from the infectious agents surveillance report published by niid . climate variables , which were summarized weekly , were retrieved from meteorologic agencies in the capitals of the 47 prefectures . considering the incubation period between infection and reporting a diagnosis of vtec , we used the climatic conditions from the 2-week period before each case was reported . we also used annual socioeconomic data for each of the 47 prefectures ( 7 ) , including the following information : population density , percentage of children ( < 15 years of age ) , percentage of elderly ( > 65 years of age ) , average number of persons in the household , number of livestock ( beef cattle , dairy cattle , hogs , and chickens ) per person in the prefecture , and average income . the highest number that occurred in a prefecture was 63 vtec cases per 1,000,000 during 1 week in a single prefecture . more than 16 vtec cases were observed in 10% of 13,489 weeks ( 287 weeks  47 prefectures ) , and no cases were reported in 57.6% of 13,489 weeks . age distribution of patients indicates that the number of vtec cases was highest in children < 5 years of age and fewer cases were reported in older age groups . a total of 65 outbreaks , defined as > 11 laboratory - confirmed vtec cases in a certain time frame and area , were reported during the study . the biggest outbreak occurred during september 2003 in kanagawa prefecture and included 252 symptomatic and 197 asymptomatic cases of vtec . the change in vtec cases over time is shown in figure 1 , on which the average air temperature ( > 25c ) during each week of the summer season is overlaid . though the annual incidence showed no clear tendency to increase or decrease during this study , a marked seasonal oscillation pattern with peaks centered in july and august was shown . the annual oscillation of verocytotoxin - producing escherichia coli ( vtec ) cases during the study period . in addition to the vtec cases , the average air temperature ( > 25c ) during each week of the summer season is overlaid in the graph . the geographic distribution of vtec cases per 100,000 per year in each of the 47 prefectures indicated that a relatively higher incidence of vtec was clustered in western sections of several japanese prefectures and northeastern sections of 2 japanese prefectures ( figure 2 ) . the 4 prefectures with the highest annual incidences were rural areas : saga ( 9.2/100,000 ) , ishikawa ( 7.9/100,000 ) , akita ( 5.8/100,000 ) , and iwate ( 5.8/100,000 ) . conversely , the prefectures with the lowest incidences were near urban areas : yamanashi ( 1.3/100,000 ) , ibaraki ( 1.1/100,000 ) , niigata ( 0.9/100,000 ) , and shizuoka ( 1.4/100,000 ) . average number of vtec cases per 100,000 population per year in each of 47 prefectures from 1999 to 2004 , japan . the association of climate and socioeconomic factors with the fluctuation of vtec cases was estimated by using multiple regression analyses ( table ) . within the climate variables , average air temperature of the day , wind speed , and the number of sunny days were significantly associated with the incidence of vtec cases per 100,000 per week per prefecture . by adjusting for these 3 climate variables as well as calendar months , associations between 7 socioeconomic variables and vtec incidence/100,000 population per week per prefecture were analyzed . results indicated that the following population - related factors were strong risk factors for vtec incidence : a higher percentage of elderly people in the prefecture , higher population density , higher number of persons in a household of the prefecture , and higher percentage of children . the following socioeconomic factors in the prefecture showed a positive association with vtec incidence : lower average income in the prefecture and greater number of beef cattle per person . on the other hand , , this multiple regression model showed that these population , socioeconomic , and climate factors could statistically explain 31% of the variability of vtec incidence . * 5,580 of 13,489 weeks ( 287 weeks  47 prefectures ) or 41.4 % of the weeks were included in the analysis as no cases of e. coli were reported during 7,909 weeks ( 58.6% ) . r = 0.31 : calculated based on the multiple linear regression model using the 7 socioeconomic variables , 3 climate variables , and calendar months . data for the 2-week period before the week e. coli was reported were used to approximate the period between infection and diagnosis . persons in a household , and percentage of elderly was 0.9 , 0.4 , and 0.6 , respectively .  * we can not determine a causal relationship because of the nature of the ecologic study that we used in this research . however , the results imply that higher beef cattle density , higher population density , and more persons per household might increase the risk of developing vtec infection . because our surveillance data were collected from different regions of japan , we compared them on the assumption that 1 ) people seek care with the same frequency in all regions when they are ill , 2 ) doctors request stool specimens with the same frequency in all regions , and 3 ) laboratories test for vtec with the same standards in all regions . thus , some degree of observation bias may exist even under control of the law , which is a limitation of this study . in addition , the number of cases includes not only symptomatic but also asymptomatic vtec , which may also raise the incidence rate in the japanese surveillance system . in conclusion , we showed a high annual incidence of vtec of 2.74 per 100,000 that was associated with climate , socioeconomic , and population factors . however , because this was an ecologic study , further longitudinal studies are necessary to address these complicated associations .
in 1999 , an infectious disease prevention law was enacted in japan that affected the nationwide infectious surveillance system . 
 a total of 19,304 laboratory - confirmed verocytotoxin - producing escherichia coli cases were reported through 2004 . 
 the annual incidence was 2.74/100,000 population ; its fluctuation over time and space was associated with climate , socioeconomic , and population factors .