Abstract:Factors like temperature,

The 16S rRNA gene of Ehrlichia genus was examined from the blood and spleen of 26 wild small mammals that were captured at a canine feed lot where the first case of canine ehrlichiosis was found in south of China in 1998. Samples (3 blood and 1 spleen) from Rattus norvegicus were positive by polymerase chain reaction. The amplimers were then cloned and sequenced. All of the 16S rRNA gene sequence of these samples were found to be same and unique when compared with the other members of genus EHRLICHIA: The most closely related species was Ehrlichia-like sp. Schotti variant (99.03% related), which was detected and identified in Dutch Ixodes ricinus ticks by Schouls in 1999. The organism found on these animals was obviously not the species of E. muris, which was isolated and identified from a wild mouse in Japan in 1995, since the relatedness of this gene between the former and the latter was 93.28%. It is unclear whether this organism can cause disease in other mammals, including human beings.

To identify the causative agent of canine ehrlichiosis that has occurred in the suburbs of Guangzhou, China, since 1998, the 16S rRNA gene was amplified and sequenced. Two sequences of 1,482 and 1,483 base pairs were obtained and named as Gzh981 and Gzh982, respectively. The level of similarity of these two was 91.50%, and Gzh981 closely resembled the 16S rRNA gene of Ehrlichia canis, whereas Gzh982 resembled Ehrlichia platys. We therefore conclude that E. canis and E. platys together caused recent outbreaks of canine ehrlichiosis in China.

Melioidosis caused by Burkholderia pseudomallei is regarded as both a tropical and subtropical disease, and has received considerable attention in Southeast Asia and Northern Australia. During the last decade, reviewers' attention seems to have focused on Thailand, where melioidosis was once reported to be a major cause of community-acquired septicemia (2), and yearly case reports reached to more than 800 (8). Hainan Island, now an isolated province having the same latitude as Thailand, was bacteriologically and serologically confirmed to be an endemic area for the pathogen in 1981 (6). Animal melioidosis was first discovered in 1986 (12), however human cases failed to be identified till the first case was discovered in 1991 (7). Seeing the island so close and so similar in natural conditions to the epidemic countries around it, we conducted this study in an attempt to reveal the actual conditions of human melioidosis prevalence on this island.Ashdown medium (ASH) was prepared as previously described (10). Briefly, one liter of ASH contained 30 g nutrient agar, 40 ml glycerol, 5 ml selective solution (0.1% crystal violet and 1% neutral red, w/v), and distilled water to a final volume of 1,000 ml. The mixture was autoclaved at 121 C for 15 min. Before plates were made, gentamicin was added aseptically to a final concentration of 5 mg/liter.Clinical isolation of B. pseudomallei was conducted in Hainan People's Hospital, a provincial hospital with 1,200 beds, an annual 26,100 inpatients and 600,000 outpatients. The study lasted one year from October 1995 to October 1996. Specimens like swabs from wounds, skin lesions and abscesses were directly inoculated into ASH plate. Blood cultures were made in nutrient broth. B. pseudomallei was first suspected by its special morphology on ASH, and then identified by conventional methods. The isolates were confirmed by the Sceptor system (Becton Dickinson, U.S.A.), and antibiotic susceptibility tests were done simultaneously.Indirect hemagglutination (IHA) tests were done as previously described (11). The IHA tests were performed using V-bottomed microtiter plates. First, 0.05 ml of the sera to be tested was inactivated at 56 C for 30 min, and then absorbed with 0.45 ml of 10% unsensitized erythrocytes at room temperature for 3 hr. Doubled dilutions of the absorbed sera were made at 0.025 ml/well in 0.01 M PBS, pH 7.2, with a commencing dilution of 1:20. Then, 0.025 ml of 1% sensitized erythrocytes with 0.5% rabbit serum supplemented was added to each well, giving the starting well a dilution of 1:40; positive and negative controls were included in each plate.