Q fever from Unknown Febrile Patients in Korea Young Sill Choi*, Kyu Jam Hwang, Sang Hee Park, Soo Kyoung Shim, and Mi Yeoun Park   Division of Zoonoses, Center for Immunology & Pathology, National institute of Health, Korea Center for Disease Control & Prevention, Seoul, Korea Introduction Table 2. Results of diagnosis of acute Q fever patients after the onset of clinical signs 1:2048 1:1024 IgG 1:10 + 1 Patients Bloods IgA IgM Days com-1 gene Phase II antibody titer (MIFA) 75 40 27 10 NT 1:512 1:320 2 1:256 1:160 14 7 _ 3 12 4 1:80 23 5 <1:10 1:40 1:640 > 1:1024 95 67 90 >1:4096 6 >1:2048 16 8 30 1:16 9 20 13 11 21 : The point of antibiotic treatment <1:16 1;2048 100 70 Table 3. Characteristics of Q fever patients 11 M 46 52 Sing and Symptoms 71 F 63 Sex Ages Patients NO 45 10 12 73 41 66 55 31 27 fever, chills, fibrin granuloma, granulomatous hepatitis fever, chills, FUO fever, chills, hepatosplenomegaly, granulomatous hepatitis fever, chills, icterohepatitis fever, chills, hepatosplenomegaly, icterohepatitis, granulomatous hepatitis endocarditis fever, chills, headache, diarrhea, hepatosplenomegaly fever, chills, headache, myalgia, sweats, cough, rash, hepatosplenomegaly fever, chills, headache, myalgia, diarrhea, rash, hepatosplenomegaly 2 1 4 3 5 8 7 9 6 Coxiella burnetii, an obligate intracellular parasite with a worldwide distribution, is the causative agent of Q fever in humans and animals. Human is generally infected by inhalation of infected aerosols, usually associated with direct or indirect contact with infected animals, or by ingestion of un-pasteurized milk from infected farm animals. In Western countries, acute Q fever has been well documented but only recently identified in Korea. It was classified and controlled as type 4 notifiable disease in Communicable Disease Prevention Act in January 2006. The aim of this work is to illustrate results of serological confirmation and detection of C. burnetii DNA from 240 unknown febrile patients ,during the period of 3 years from 2004 to 2006. Materials and Methods Fig. 1. Detection of C. burnetii com-1 gene in blood by nested PCR Lane M: molecular size marker(100-bp DNA ladder), P: DNA of C. burnetii, N: negative control , lane 1~ 12 specimens M P N 1 2 3 4 5 6 7 8 9 10 11 12 438bp→ Bacterial strains and cell line * Coxiella burnetii Nine Mile Strain ATCC VR 615 * Cultured in L929 mouse fibroblast in DMEM ( 5% FBS, antibiotic-free) Clinical specimens * 240 serum and blood from acute febrile patients MIFA & Cross–test (Mycoplasma, Legionella,Chlamydia, Borrelia, Brucella, Bartonella) Nested PCR and Sequence * Genomic DNA was extracted with Gentra DNA kit * Amplification of the omp 27kDa encoding gene (com-1, 438bp) and the amplified DNA were sequenced with an ABI 3100 automated PCR-RFLP * PCR amplified DNA was digested with Sal I Fig. 2. RFLP of C. burnetii com-1 (438bp) gene by nested PCR. The amplification products were digested with Sal I, electrophoresed on 2% agarose gel . : Lane M; size standards Marker(1Kb); lanes 1, 11 were C. burnetii com-1; lanes 2 - 11 were un-cutting com-1 gene of specimens; lanes 21-111 were cutting com-1 gene of specimens. 313bp 125bp → 438bp→ M 1 11 2 21 3 31 4 41 5 51 6 61 7 71 8 81 9 91 10 101 11 111 Table 1. DNA sequence of primers for the detection of C. burnetii used in this study Target gene OMP! OMP2 OMP3 OMP4 27kDa C. burnetii Primer set Oligoneucleotide sequence Amplified product Target species ' 5' TGC CTG CTA GCT GTA ACG ATT G 3' 5' GAA GCG CAA CAA GAA GAA CAC 3' 5' TTG GAA GTT ATC ACG CAG TTG 3' 5' AGT AGA AGC ATC CCA AGC ATT G 3' 501 438 Results The results of serology tests, 12 cases were evaluated from 240 unknown febrile patients during the 2004-2006. All of the 12 cases, IgG antibody titers for phase II antigen showed from 1:256 to 1:4096. IgM titers of 1 case showed less then 1: 16 the other 11 cases showed from 1: 80 to 1:1024, and IgA titers of 1 case was conformed 1: 80 The 10 cases were amplified to the 27kDa encoding gene (com-1) of C. burnetii PCR- RFLP patterns by digested with Sal I showed that identical patterns to that of C. burnetii Nine Mile Strain. Conclusions This study conformed Q fever from unknown febrile patients. These cases were lack in epidemiological data, it is necessary for epidemiology evidence of Q fever. The further study needs to strength of sentinel surveillance as zoonosis and shared information with veterinary field.