광주에서 포획된 설치류에서 라임병 원인균 검출

Abstract

Background: Lyme disease is a vector-borne disease that occurs mainly in North America, Europe and Asia, human gets infected by being bitten by a tick infected with a bacterium Borrelia species. Symptoms include erythema migrans, fatigue, fever, headache, neck stiffness, myalgia, arthralgia and swelling of the lymph nodes. Borrelia species such as Borrelia burgdorferi, B. afzelii and B. garinii have been reported as the pathogens of Lyme disease, and ticks such as Ixodes scapularis and I. persulcatus have been reported as their carriers. For the diagnosis of Lyme disease, either indirect immunofluorescence or enzyme-linked immunosorbent assay is performed first and then positive specimen is confirmed by Western blot test, to isolate and identify the bacteria in a patient sample (blood, etc.) culture or molecular method targeting the Borrelia gene can be used. However, there are few reports on the prevalence of Lyme disease among tick - borne diseases in Korean wild rodents. This study was carried out to investigate the infection rate of Borellia bacterium, a causative organism of Lyme disease, and to investigate the different Borellia species by using wild mouse specimens captured in the suburbs of Gwangju Metropolitan City in southwestern Korea. Experimental method: In October and November 2014, a total of 27 rodents were captured at two different sites Gwangsan-gu and Buk-gu in the Gwangju Metropolitan City, South Korea. In October, 12 wild mice were caught, and in November 15 wild mice were caught. After the anesthesia, the spleen, kidney, liver, heart and lung tissues were collected. Tissues were homogenized, genomic DNA was extracted and PCR were performed. Nested PCR were performed targeting the pyrG gene and ospA gene specific to Borrelia. The products obtained after PCR were purified and sequenced using N-PCR primers to identify the bacteria. Result: After PCR, positive specimens were sequenced. N-PCR targeting the pyrG gene specific to Borrelia confirmed the PCR positive specimen in 8 out of 27 wild mice. As a result of pyrG N-PCR sequence analysis, the positive rate of Borrelia in 27 wild mice captured in October and November was 29.63% (8 positive / 27 total) and only detected in spleen, kidney and heart tissues. The highest detection rate was found in the heart tissue that is 25.9% (7 positive / 27 total). The kidney and spleen showed 7.4% positivity (2 positive / 27 total) and 14.8% positivity (4 positive / 27 total), respectively. The infection rate of B. afzelii was 25.9% (7 positive / 27 total) and showed highest detection for B. afzelii in heart tissue. The kidney and spleen showed 7.4% (2 positive / 27 total) and 14.8% (4 positive / 27 total) positivity, respectively. In the case of B. garinii, the infection rate was 7.4% (2 positive / 27 total) in wild mice, while only the spleen tissue showed positive results among the five organs used in the experiment. When ospA N-PCR was performed, B. afzelii positive rate was 7.4%, detected in only 2 heart tissue among the 27 wild mice captured in October and November, all other tissues did not showed positive result. A phylogenetic tree was constructed using the sequences of pyrG DNA fragment from Borrelia-positive tissue samples and pyrG gene from various Borrelia strains obtained from GenBank. All of B. afzelii positive specimens lied in the same clade of with B. afzelii. It was confirmed that B. garinii, which was detected in two wild mice spleen, was also located in the same clade of B. garinii. The pyrG DNA identified as B. afzelii positive in each organ showed the 99.1% or more similarity to pyrG gene of B. afzelii isolate from the Chinese patient HLJ01 strain. The B. garinii DNA positive for two mice spleen tissue were confirmed to have 98.3% or more similarity to B. garinii strains SCCH-7 CTP synthase gene isolated from US rodents. Conclusion: In this study, B. afzelii, one of the causative bacteria of Lyme disease, was found to be highly positive 29.6% in wild mice caught in Buk-gu and Gwangsan-gu, Gwangju Metropolitan City. Infection was confirmed in spleen, kidney and especially in heart tissues. B. garinii positive rate was 7.4% but only the spleen tissues showed positive result. These results indicate that B. afzelii, a Lyme spirochete, shows a high tropism for the heart, and B. garinii has a high tropism for the spleen in mice. Therefore, in order to diagnose Lyme disease in rodents, detection in these tissues, heart and spleen should be conducted.