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RESEARCH NOTE

Risk-factors for human infection with studies concerning scrub have, to date, tsutsugamushi: a case-control study involved case reports, clinical findings and diag- in Korea nosis, serological diagnosis, and treatment regi- mens [3–5], and no epidemiological studies have D.-M. Kim1, K. Y. Kim2,H.S.Nam2, been conducted to examine the risk-factors for S. S. Kweon3, M.-Y. Park4 and S. Y. Ryu5 developing . The present report describes a case-control study designed to iden- 1Division of Infectious Diseases, Department of tify such risk-factors. Internal Medicine, Chosun University College of Patients were selected according to the follow- Medicine, 2Department of Preventive Medicine, ing eligibility criteria: (i) a diagnosis of scrub Chungnam National University Medical School, typhus reported to the Korean Centers for Disease 3Department of Preventive Medicine, Seonam Control and Prevention (KCDC) during 2005; University College of Medicine, 4Korea Center (ii) residency in south-west Korea (Gwangsan-gu, for Disease Control and Prevention and Gwangju City; Gurye-gun or Suncheon City, 5Department of Preventive Medicine, Chosun Chonnam Province; or Namwon City, Chonbuk University Medical School, Gwangju, Korea Province); (iii) an age of 40–75 years; (iv) an ability to communicate effectively; and (v) informed consent to participate in the study. In total, 6756 ABSTRACT patients with scrub typhus were registered with A case-control study was conducted involving 156 the KCDC during 2005. Of these, 279 had a patients with scrub typhus and 130 controls. Three residence in south-west Korea, and 271 of these factors were associated significantly with the risk patients fulfilled the age criteria. A letter was sent of developing scrub typhus: engaging in fruit to these patients to explain the objective of the farming (OR 2.44; 95% CI 1.04–5.69), gathering present study and to request their participation. chestnuts (OR 2.05; 95% CI 1.09–3.87) and taking Trained examiners collected the patients’ baseline breaks in areas adjacent to agricultural operations data. In total, 156 (71.9%) patients were selected (OR 3.06; 95% CI 1.50–6.22). In contrast, receiving as cases. A control group with a socio-economic information or educational materials concerning status similar to that of the patient group was the prevention of scrub typhus had a protective obtained using the patients’ neighbours as con- % effect (OR 0.45; 95 CI 0.24–0.83). These results trols (n = 130). Statistical analysis was performed suggest that a health education programme will using SPSS v.12.0 for Windows (SPSS Inc., lower the risk of developing scrub typhus when Chicago, IL, USA). applied to high-risk groups. There were no significant differences in age, Keywords Education, epidemiology, Orientia tsutsu- gender or socio-economic parameters between the gamushi, risk-factors, scrub typhus two groups, although there were significant dif- ferences between cases and controls with respect Original Submission: 16 March 2007; Revised Submission: 23 August 2007; Accepted: 28 September to fruit farming (p <0.05), outdoor agricultural 2007 activities (p <0.05) and gathering chestnuts (p <0.05) (Table 1). In total, 72% of the cases took Clin Microbiol Infect 2008; 14: 174–177 lunch ⁄ snack breaks at home or elsewhere (rather 10.1111/j.1469-0691.2007.01901.x than in proximity to agricultural activities) as compared to 86.5% of the controls, and 25.2% of Scrub typhus is a -borne infectious disease the cases and 39.8% of the controls used a rug or caused by Orientia tsutsugamushi, which is trans- other covering as a mat while outside (Table 1). mitted by chigger mite larvae Chronic disease, e.g., hypertension, diabetes, liver belonging to the family [1,2]. Most disease or renal disease, had been present for >3 months among 18.6% of the patients and 13.8% of the controls (p >0.05, data not shown). Corresponding author and reprint requests: S. Y. Ryu, Depart- Using statistically significant parameters, and ment of Preventive Medicine, Chosun University Medical those that were known previously to be essential School, 375 Seosuk-dong, Dong-gu, Gwangju 501-759, Korea in preventing scrub typhus, a multiple logistic E-mail: [email protected]

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Table 1. Demographical and socio- Cases (n = 156) Controls (n = 130) p value economic data and risk-factors for scrub typhus Demographical and socio-economic data Gender (male ⁄ female) 55 ⁄ 101 (35.3% ⁄ 64.7%)49⁄ 81 (37.7% ⁄ 62.3%) 0.762 Mean age (years) 60.6 ± 10.4 61.0 ± 11.8 0.736 Marital status 132 (84.6%) 101 (78.3%) 0.222 Educational level Minimal education 45 (29.6%) 36 (27.9%) 0.877 Primary school graduates 59 (38.8%) 50 (38.8%) Middle school graduates 26 (17.1%) 19 (14.7%) High school graduates and above 22 (14.5%) 24 (18.6%) Type of medical insurance coverage Local medical insurance 100 (65.8%) 81 (62.3%) 0.429 Medical insurance for employees 44 (28.9%) 37 (28.5%) Medical aid 8 (5.3%) 12 (9.2%) Occupation and agricultural activities Occupation Agriculture-related 109 (69.9%) 88 (67.7%) 0.692 Non-agriculture-related 47 (30.1%) 42 (32.3%) Agricultural activities Cultivating a rice field 90 (57.7%) 76 (58.5%) 0.991 Dry field farming 53 (34.0%) 34 (26.2%) 0.193 Fruit farming 32 (20.5%) 12 (9.2%) 0.014 Growing crops for special use 6 (2.6%) 9 (6.9%) 0.140 Growing greenhouse vegetables 7 (4.5%) 5 (3.8%) 1.000 Raising livestock 17 (10.9%) 10 (7.7%) 0.472 Outdoor agricultural activities and other outdoor activities Agricultural History of agricultural activities 127 (81.4%) 91 (70.0%) 0.034 Removing weeds 81 (51.9%) 67 (51.5%) 1.000 Gathering a harvest 93 (59.6%) 77 (59.2%) 1.000 Sowing 90 (57.7%) 72 (55.4%) 0.785 Rice reaping 72 (46.2%) 70 (53.8%) 0.239 Fruit harvesting 47 (30.1%) 25 (21.5%) 0.131 Non-agricultural Mowing 90 (57.7%) 72 (55.4%) 0.785 Gathering herbs 10 (6.4%) 10 (7.7%) 0.849 Gathering chestnuts 59 (37.8%) 32 (24.6%) 0.024 Climbing 25 (16.0%) 25 (19.2%) 0.579 Freshwater fishing 5 (3.2%) 6 (4.6%) 0.759 Visiting ancestral tombs 76 (48.7%) 62 (47.7%) 0.957 Outdoor places for rest Places for lunch and a snack At home or elsewhere 108 (72.0%) 109 (86.5%) 0.003 Near agricultural ⁄ outdoor activities 42 (28.0%) 17 (13.5%) Seating arrangements Use of a rug or other covering 37 (25.2%) 47 (39.8%) 0.035 Use of paper covering 38 (25.9%) 27 (22.9%) Sitting on bare ground 72 (49.0%) 44 (37.3%) Personal hygiene, protection and education Mean frequency of taking a shower Almost every day 55 (36.2%) 59 (45.7%) 0.066 3–4 times weekly 43 (28.3%) 22 (17.1%) Less than twice weekly 54 (35.5%) 48 (37.2%) Frequency of wearing long trousers and sleeves Always 109 (72.2%) 99 (79.2%) 0.300 Depending on weather 42 (27.9%) 26 (20.8%) Carrying gloves Always 96 (63.6%) 84 (66.7%) 0.539 Sometimes 38 (25.2%) 25 (19.8%) Seldom 17 (11.3%) 17 (13.5%) Wearing socks Always 133 (87.5%) 107 (84.9%) 0.598 Sometimes 16 (10.5%) 14 (11.1%) Seldom 3 (2.0%) 5 (4.0%) Use of insect repellent Yes 14 (9.0%) 19 (14.8%) 0.184 No 141 (91.0%) 109 (85.2%) Received health education Yes 41 (26.5%) 46 (35.4%) 0.103 No 114 (73.5%) 84 (64.6%)

Total 156 (100.0%) 130 (100.0%) regression analysis was performed to identify risk- in gathering chestnuts, and 3.06-fold higher factors (Table 2). The risk of developing scrub (95% CI 1.50–6.22) for individuals who had typhus was 2.44-fold higher (95% CI 1.04–5.69) for lunch ⁄ snack breaks in proximity to agricultural individuals engaged in fruit farming, 2.05-fold operations. In addition, the risk of developing scrub higher (95% CI 1.09–3.87) for individuals engaged typhus was 0.45-fold lower (95% CI 0.24–0.83) for

2007 The Authors Journal Compilation 2007 European Society of Clinical Microbiology and Infectious Diseases, CMI, 14, 174–189 176 Clinical Microbiology and Infection, Volume 14 Number 2, February 2008

Table 2. Risk-factors for scrub Univariate analysis Adjusted results typhus according to logistic regres- sion analysis Category Variables OR 95% CI OR 95% CI

Occupation Agriculture 1.00 1.00 Others 0.90 0.55–1.49 1.64 0.76–3.54 Fruit farming No 1.0 1.00 Yes 2.54 1.25–5.16 2.44 1.04–5.69 Agricultural activities No 1.00 1.00 Yes 1.88 1.08–3.26 2.21 0.99–4.93 Gathering chestnuts No 1.00 1.00 Yes 1.86 1.11–3.11 2.05 1.09–3.87 Places for lunch ⁄ snack breaks At home or elsewhere 1.00 1.00 Near outdoor ⁄ agricultural activities 2.49 1.34–4.65 3.06 1.50–6.22 Seating arrangements Use of a rug or other covering 1.00 1.00 The bare ground 1.62 0.98–2.65 1.49 0.84–2.63 Mean frequency of Almost every day 1.00 1.00 taking a shower Less than four times weekly 1.49 0.92–2.40 1.56 0.88–2.78 Changing clothes after Yes 1.00 1.00 outdoor activities No 1.62 0.95–2.75 1.79 0.95–3.35 Use of insect repellant No 1.00 1.00 Yes 0.57 0.27–1.19 0.50 0.21–1.16 Receipt of health education No 1.00 1.00 Yes 0.66 0.40–1.09 0.45 0.24–0.83 individuals who were informed or given educa- insect repellent must be sprayed on the clothing tional materials concerning the prevention of scrub and skin. A health education programme was not typhus. a significant factor in univariate analysis, but was Leptotrombidium pallidum and Leptotrombidium significant following multivariate analysis. This scutellare are common types of mite in Korea. indicates that information or a health education They are usually found in locations in which programme is an essential step in helping to wild mice thrive, e.g., moist and humid loca- prevent scrub typhus. tions with tall grass. L. pallidum and L. scutellare In conclusion, these results indicate that fruit are usually found 4–8 cm from the soil surface farming, gathering chestnuts, and taking breaks [6,7]. L. scutellare larvae are active and take a in areas adjacent to agricultural operations are blood meal in cool dry autumn conditions [8]. associated with a high risk of developing scrub For these reasons, the could transmit typhus. In contrast, individuals who are given O. tsutsugamushi to humans during chestnut information concerning the avoidance of scrub harvesting and orchard farming in the autumn, typhus have a decreased risk of developing the around the time of Chusok (Korean Thanksgiv- disease. ing Day) [9]. The frequency of having outdoor lunch or ACKNOWLEDGEMENTS snack breaks adjacent to agricultural activities was significantly higher in the patient group, This study was supported financially by the Korean Centers implying that individuals are at increased risk of for Disease Control and Prevention (KCDC). The authors declare that they have no conflicts of interest in relation to this being exposed while having lunch or resting in work. areas where the chigger mites live. Consequently, individuals should have breaks indoors or, if this is not possible, they should be instructed to use a REFERENCES mat or rug outdoors. 1. Watt G, Parola P. Scrub typhus and tropical rickettsioses. In regions where scrub typhus is common, Curr Opin Infect Dis 2003; 16: 429–436. Korean public health centres dispense supplies of 2. Chang WH. Scrub typhus. J Kor Med Assoc 1994; 26: 387–393. insect repellent and use health education pro- 3. Ogawa M, Hagiwara T, Kishimoto T et al. Scrub typhus in Japan: epidemiology and clinical features of cases reported grammes that provide the following advice: (i) in 1998. Am J Trop Med Hyg 2002; 67: 162–165. clothing must not be taken off while working 4. Kim DM, Yun NR, Yang TY et al. Usefulness of nested PCR outdoors, and sitting or lying down in the grass for the diagnosis of scrub typhus in clinical practice: a should be avoided; (ii) clothes worn during work prospective study. Am J Trop Med Hyg 2006; 75: 542–545. 5. Kim DM, You KD, Lee JH, Kim HK, Lee SH. Controlled outdoors must be dusted-off and cleaned and, in trial: 5-day course of telithromycin versus for addition, daily showers must be taken; and (iii)

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the treatment of mild to moderate scrub typhus. Antimicrob Original Submission: 19 April 2007; Revised Submis- Agents Chemother 2007; 51: 2011–2015. sion: 8 August 2007; Accepted: 27 September 2007 6. Goddard J. Chigger mites and scrub typhus. Infect Med 2000; 17: 236–239. Clin Microbiol Infect 2008; 14: 177–180 7. Gentry JW, Yueh CS, Wah PO. Preliminary observations on 10.1111/j.1469-0691.2007.01897.x Leptotrombidium (Leptotrombidium) akamushi and Leptotreom- bidium (Leptotrombidium) deliensis in their natural habitat in Malaya: (Acarina, Trombiculidae). Am J Hyg 1963; 78: 181– Early initiation of adequate antibiotic treatment in 190. cases of sepsis significantly reduces mortality, 8. Yi KS, Chong Y, Covington SC et al. Scrub typhus in Korea: importance of early clinical diagnosis in this newly recog- morbidity and hospital costs [1]. Rapid species nized endemic area. Mil Med 1993; 158: 269–273. identification could help to correct inadequate 9. Strickman D, Tanskul P, Eamsila C, Kelly DJ. Prevalence of empirical antimicrobial treatment. Rapid molecu- antibodies to rickettsiae in the human population of lar-based species identification methods have suburban Bangkok. Am J Trop Med Hyg 1994; 51: 149– 153. been developed, but these are expensive, are labour-intensive and require special equipment [2,3]. The present study describes the develop- ment of a method for rapid preliminary species RESEARCH NOTE identification from positive blood culture vials. Significant growth of from blood culture vials was obtained within a few hours on agar plates. Rapid culture and identification: a practical Data were collected prospectively at Karolinska method for early preliminary laboratory University Hospital, Huddinge, Sweden, during diagnosis of sepsis 6 months in 2006. In total, 541 positive blood V. O¨ zenci, K. Tegmark-Wisell, C. Lundberg cultures were included in the study, with only and B. Wretlind one culture for each patient. The method was also tested using nine samples from sterile body sites Division of Clinical Microbiology, Karolinska other than blood. The Bactec 9240 automated Institutet, Karolinska University Hospital, blood culture system with both aerobic and Huddinge, Stockholm, Sweden anaerobic vials (Becton Dickinson, Sparks, MD, USA) was used in the study. The results were compared with those obtained by conventional microbiological methods. ABSTRACT In preliminary experiments to determine the This study describes the development of a optimal incubation time and amount of inoculum method for rapid preliminary species identifica- for rapid culture, reference strains of Escherichia tion of bacteria from positive blood culture vials. coli and Staphylococcus aureus, as well as eight The method yielded preliminary identification clinical samples, were inoculated in blood culture results for 496 (92%) of 541 positive blood vials together with 5 mL of horse blood. When the cultures within 5 h. The method was capable of automated blood culture system signalled identifying the most frequently isolated bacteria growth, samples of the blood culture were anal- (i.e., Staphylococcus aureus, coagulase-negative ysed. The results revealed that five drops yielded staphylococci, , Streptococcus pneu- optimal growth and provided the best discrimi- moniae and Enterococcus spp.) to the species level. nation of single colonies after a 4-h incubation The method can be established easily, with a period. materials cost of 2–5 Euros per sample. Accordingly, five drops of blood culture Keywords Blood cultures, diagnosis, identification, (n = 541) were then subcultured in duplicate on rapid, sepsis, spot tests blood agar, chocolate blood agar with haematin, and cysteine lactose electrolyte-deficient agar, followed by incubation at 37C for 4 h in both Corresponding author and reprint requests: V. O¨ zenci, Karo- an aerobic and a CO2 5% v ⁄ v atmosphere. When linska University Hospital, Clinical Microbiology, F72, Hudd- anaerobic bacteria were suspected, similar inge, 141 86 Stockholm, Sweden E-mail: [email protected] amounts of culture were subcultured on brucella

2007 The Authors Journal Compilation 2007 European Society of Clinical Microbiology and Infectious Diseases, CMI, 14, 174–189