| Streptococcus suis is an important zoonotic pathogen,not only can lead to meningitis and septicemia in pigs,but can also cause human infected with meningitis and toxic shock syndrome.S.suis often be identified and classified by its morphological,biochemical characteristics and serological features.To date,33 serotypes(type 1 through 31,33,and 1/2)of S.suis have been identified based on antigenic differences in the capsular polysaccharide.There were another parts of the undifferentiated strains exist.In different parts of the world,the different clinical disease caused by serotypes of S.suis were different,strains isolated from diseased human and pigs primarily belong to serotype 2 in most countries,followed by serotypes 1,2,3,4,7,9.Healthy pigs can harbor S.suis in tonsils and upper respiratory tract,with the diseases pigs play an important role in disse minating S.suis.Based on the strength of its pathogenic S.suis can be divided into highly-virulent strain,weakly-virulent and non-virulent.Now we generally deter mined the virulence of the strain based on the distribution of the virulence factors,but up to now,we still not have a clear identification method,animal testing is still an unique way to evaluate the virulence of S.suis.Therefore,we developed an epidemiological investigation of S.suis in Guangdong province,and provide a basis for research on the pathogenic mechanism of Streptococcus suis.1.Deter mination of the single PCR assay for the identification of 33 serotypes and A multiplex PCR assay for main pathogenic serotypes 2,7,9 of Streptococcus suisThe primers of 33 serotypes of S.suis were synthesized according to the related references articles.Specificity of the PCR was using the genome of 31 serotype reference strains test 33 pairs of specific primers,by optimizing the annealing temperature,we deterninated an no cross reaction PCR assay of the 33 serotypes successfully(serotype 2 and 1/2,serotype 1 and 14 can not be separated by PCR).The 19 serotypes positive reference plasmid were constructed successfully in the end.According to the epidemic situation of Guangdong region of Streptococcus suis,this research established a multiplex PCR assay of the major pathogenic serotype 2,7,9 of S.suis,The identified results of specificity and sensitivity show that the multiplex PCR assay is highly efficient and sensitivity can be achieved 104CFU/ml,the coincidence of accuracy and repeatability test has reached 100%.Because of low cost,high efficiency,this assay is suitable for the diseases surveillance and epidemiological diagnosis of Streptococcus suis.2.Epidemiology investigation of Streptococcus suis in Guangdong province488 samples of nasal swabs and tonsil were collected from healthy pigs in this research;143 samples(39.72%)showed positive when detected 360 samples,the positive rate of nasal swab samples(68%)of the piglets was the highest.68 strains of SS were isolated from 488 healthy pig samples,50 strains were identified and serotyped in 16 serotypes among them,mainly in serotype 16(11.76%),2(8.82%),29(7.35%)and et.40 strains were identified and serotyped in 10 serptypes in 58 SS strains collected from diseased pigs stroed in Haida laboratory,mainly in serotype 3(13.79%),7 and 9 possessed(12.72%)and et.These results suggested SS is widespread in pigs in Guangzhou and the surrounding areas,and widely distribution in seroptypes too(covered 18 serotypes).Except the serotype 2,7,9,the 16,19 and 29 are also popular in Guangdong province.There are different in predo minant serotype betwen the healthy pigs and pathogenic pigs.It proved that the complexity popularation of Streptococcus suis in Guangzhou and the surrounding areas.3.The distribution of the main virulence factors of Streptococcus suis and characteristics analysis of isolates SS2PCR detection of the distribution of virulence related genes mrp,epf,sly in 126 clinical isolated SS strains.In 68 SS isolated from healthy pigs,1 strain showed mrp-/epf+/sly-,6 strains showed mrp-/ef-/sly+;when in 58 SS isolates from pathogenic pigs,mrp+/ef+/sly+has 2 strains,mrp-/ef+/sly+ 2 strains,16 strains showed mrp-/ef-/sly+,1 strains showed mrp-/epf+/sly-.Two serotype 2 strains YN28 and SS22 isolated from diseased pigs,and the PCR results showed YN28(mrp+/epf+/sly+/sbp2'+/gdh+);SS22(mrp-epf+/sly+/sbp2'+/gdh+).Two serotype 2 strains ZY05719 and T15 were used as the reference strains of highly-virulent strain,and weakly-virulent strain(internal standard).Zebrafish model was introdued to evaluate the virulence of SS22 and YN28,and the virulence level was that ZY05719>YN28>T15>SS22.Growth analysis showed that strains YN28 and SS22 switched into logarithmic phase more previously than reference strains T15 and ZY05719;and their situation wouled be stable at the 6th h;however,possessed lower value of the highest OD600 than references. |
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