Objective To investigate the molecular epidemiological characteristics of human brucellosis in Alxa league, Inner Mongolia autonomous region (Inner Mongolia), China. Methods The clinical isolates of suspected Brucella with a positive blood culture result during 2015 to 2019 were identified by conventional biological methods and multiplex PCR assay. The molecular epidemiological characteristics of Brucella were investigated using multiple-locus variable-number tandem repeat analysis (MLVA). Results Twenty-three suspected Brucella strains were isolated from the confirmed cases of brucellosis in Alxa league from 2015 to 2019, and were identified as B. melitensis biovar 3 using conventional biological methods and multiplex PCR assay. Among the 23 cases, herdsmen accounted for 30.43% (7/23) and farmers accounted for 39.13% (9/23), both of whom had a history of sheep contact. The positive rate of Brucella antibody in serum was 78.26% (18/23). The MLVA showed that the 23 strains had genotypes of the main epidemic B. melitensis strains in China. According to the three loci with differences, the strains isolated from Bayanmuren Sumu belonged mainly to one group, and the two strains from Jilantai town belonged to another group. The 23 strains were associated with the isolates from Ningxia, Liaoning, Shandong, Zhejiang, Shanghai, and Guangdong (provinces, autonomous regions, and municipalities directly under the central government), and were not associated with isolates from its neighboring province Gansu. Conclusion The Brucella strains from Alxa league, Inner Mongolia are consistent with the epidemic strains of brucellosis in China, and are associated with the isolates from Ningxia, Liaoning, Shandong, Zhejiang, Shanghai, and Guangdong (provinces, autonomous regions, and municipalities directly under the central government).

Objective To compare the phenol water method and the iTron kit in extracting lipopolysaccharides(LPS) from Brucella. Methods LPS were extracted from B. melitensis strain 16M using phenol water method and kit method. The purity and structure of LPS was compared by SDS-gel electrophoresis and silver stain. The activity of the extracted LPS was detected by limulus amebocyte lysate agglutination test, and the characteristics of the two methods were compared. Results LPS extracted from Brucella by both methods had a high purity. There was no significant difference in activity of LPS extracted by phenol water method and the iTron kit ( t'=1.270, P=0.332), which was (4.926±0.051) and (5.015±0.037) EU/ng, respectively. LPS extracted by phenol water method might lose some LPS at 17×10 ³ and 26×10 ³, while using the iTron kit we can get intact LPS in a convenient and safe way. Conclusion The iTron kit has advantages in extracting LPS from Brucella compared with the phenol water method.

Objective Three suspected brucellosis cases were investigated in Suizhou city of Hubei province, and three suspected Brucella strains were identified. Methods Field investigation was applied to finish Brucella epidemiological survey, used Brucella conventional appraisal classification method and the phage cracking test for testing. BCSP31-PCR method (Based on Brucella genus specific gene BCSP31 target gene detection method), AMOS-PCR (based on electrophoresis banding identify B. abortus biovar 1, 2, 4 of Brucella, B. melitensis, B. ovis and B. suis biovar 1) and Real-time PCR, three molecular classification method were compared. Results Three patients were infected by close contact with sick animals and their meat, conventional identification and a variety of polymerase chain reaction (PCR) had the same results, B. melitensis biovar 3. Conclusion Brucella melitensis biovar 3 was identified as the pathogen of the three cases from Suizhou city and cases were mainly caused by direct contact with sick animals and their meat.

Objective To explore the feasibility of dilution rose bengal plate agglutination test (RBPT) for the diagnosis of brucellosis. Methods Serum samples from 93 brucellosis patients who came from Inner Mongolia, China in 2013 were tested by both dilution RBPT and standard tube agglutination test (SAT). According to the correlation between the results of the two tests and using the results of SAT as criteria, the sensitivity and specificity of RBPT were analyzed at different dilutions as cut-off values to evaluate the diagnostic accuracy. Results The area under the receiver operating characteristic curve for dilution RBPT was 0.946. When taking1∶2 as the cut-off value, the Youden index was 0.893, the sensitivity was 89.30% , and the specificity was 100%; when taking1∶4 as the cut-off value, the Youden index was 0.631, the sensitivity was 63.10%, and the specificity was 100%; when taking1∶8 as the cut-off value, the Youden index was 0.571, the sensitivity was 57.14%, and the specificity was 100%; when taking1∶16 as the cut-off value, the Youden index was 0.191, the sensitivity was 19.05%, and the specificity was 100%. Conclusion Dilution RBPT has the highest diagnostic accuracy at a dilution of1∶2, which provides a reference for the diagnosis of brucellosis.

【Abstract】 Objective To study the method to identify Brucella suis biovar 1 wild strain and S2 vaccine strain.  Methods A total of 21 strains of B.suis biovar 1 wild strain were analyzed by AMOS-PCR, PFGE and multiple-locus variable-number tandem-repeat analysis (MLVA).   Results B.suis biovar 1 and S2 could be discriminated by MLVA Bru9 and Bru16. Conclusion The MLVA assay can be applied to identification of wild strains and vaccine strain and can be proposed to as a complement of classical biotyping methods.