目的 构建以人细胞毒性T淋巴细胞抗原4（cytotoxic T lymphocyte-associated antigen 4， CTLA4）为引导的抗变形链球菌（Streptococcus mutans， S1mutans）葡糖基转移酶（glucosyltransferase， GTFs）和表面蛋白PAc的靶向融合防龋DNA疫苗pGJA-P，检测其在真核细胞中的表达。方法 利用RT-PCR方法从外周淋巴细胞中获得CTLA4胞外区和Igγ1恒定区基因并进行T2A克隆，获得携带CTLA4-Ig融合基因的重组质粒pGJ，选择合适的酶切位点，再克隆入融合防龋DNA疫苗pGLUA-P中，构建靶向融合防龋DNA疫苗p GJA2P，并转染CHO细胞系，用蛋白质免疫印迹实验检测其表达。结果 重组质粒pGJ、pGJA-P分别含有CTLA4-Igγ1融合基因和CTLA42、Igγ1、pac、glu基因序列。蛋白质免疫印迹结果显示，pGJA-P表达的抗原可以与特异性抗PAc抗体发生免疫反应。结论 成功构建了靶向融合防龋DNA 疫苗pGJA2P，并可在真核细胞中正确表达。

目的 体外检测靶向融合防龋DNA疫苗pGJA-P的免疫反应性；与非靶向融合防龋DNA疫的pGJA-P进行比较，评价其增强疫苗免疫效能的能力。方法 将pGJA-P转染CHO细胞，蛋白质免疫印迹实验检测重组蛋白抗体免疫反应性。分5组免疫兔：pGJA-P经肌肉注射组（A组）；pGJA-P经鼻黏膜免疫组（B组）；pGJA-P经肌肉注射（C组）；pGJA-P和pCI栽体经肌四头肌注射组（E组）。酶联免疫吸附实验检测血清唾液中的特异性抗体。用收集的血清进行葡糖转移酶（GTF）合成水不溶性葡聚糖抑制实验。结果 pGJA-P表达的重组蛋白可以与抗GTF抗体反应。A组血清特异性IgG抗体水平均数远高于C组（P<0.01）；B组唾液特异性lgA抗体水平显著高于D组（P<0.01）；A组同样诱导了高水平的唾液特异性lgA抗体。A组的免疫血清抑制GTF活性的能力最强。结论 pGJA-P具有GTF的免疫反庆性；并较pGJA-P有更强的诱异系统和黏膜免疫反应及抑制GTF的能力。

Salivary secretory immunoglobulin A (S-IgA) antibodies act as the first line of defense against dental caries by blocking of adherence of Streptococcus mutans to tooth surfaces. This study focused on finding proper mucosal immunization route and delivery system to induce higher level of specific anti-S. mutans saliva S-IgA and inhibit dental caries in animal model. By immunizing rats with an anti-caries DNA vaccine, pCIA-P, via different mucosal routes, we found that intranasal (i.n.) immunization with pCIA-P/bupivacaine DNA complexes elicited the highest specific anti-S. mutans saliva S-IgA mucosal antibody responses compared with naked DNA and other routes. Correspondingly, rats immunized with pCIA-P/bupivacaine DNA complex via i.n. displayed the least carious lesions. Our findings suggested that DNA vaccination via intranasal immunization with bupivacaine delivery system be a promising approach against dental caries.

Streptococcus mutans and Streptococcus sobrinus are associated with the development of dental caries. The purpose of this study was to differentiate these bacteria by morphology, biochemical characteristics and PCR, and to compare their occurrence with the prevalence of dental caries in the Mosuo people. Plaque samples were collected from the permanent first molar in 126 Mosuo people (83 females, 43 males, aged 25-55 years, average age 36.1 B 7.73). Dental status was recorded as DMFT by WHO caries diagnostic criteria. Males had a significantly lower prevalence of caries and DMFT than females: 11.4 vs. 86.9% and 1.65 vs. 6.95, respectively (p ! 0.001). Morphological and biochemical tests gave unreliable results. The prevalence of S. mutans and S. sobrinus was 75.4 and 57.1%, respectively. 26.5% of females and 53.5% of males were positive for S. mutans alone, 18.1% of females and 16.3% of males were positive for S. sobrinus alone, while 50.6% of females and 18.6% of males were positive for both S. mutans and S. sobrinus and 4.8% of females and 11.6% of males were negative for both species. The DMFT scores of subjects positive for both S. mutans and S. sobrinus were significantly higher than of those positive for either S. mutans or S. sobrinus alone. These results indicate that subjects harboring both S. mutans and S. sobrinus have a significantly higher prevalence of dental caries than those with S. mutans or S. sobrinus alone.