Oral immunization of humans with dehydrated liposomes containing Streptococcus mutans glucosyltransferase induces salivary immunoglobulin A2 antibody responses

Seven healthy adult volunteers each ingested an enteric-coated capsule containing 500 fig of Streptococcus mutans glucosyltransferase (GTF) in dehydrated liposomes for 3 consecutive days. The immunization regimen was repeated 28 days later. Parotid saliva and plasma were collected prior to and at a weekly interval for 8 weeks following the first immunization for analysis of anti-GTF activity by enzyme-linked immunosorbent assay. The levels of immunoglobulin A, (IgA), and IgA: anti-GTF activities increased in the parotid saliva from 5 of 7 individuals after immunization. Increases in the mean level of IgA, and lgA: anti-GTF responses peaked on day 35 (77% and 175% increase over baseline, respectively), although variation was noted in the kinetics and subclass of responses between individuals. No salivary IgG or IgM responses were observed. Low plasma IgM, IgG and IgA anti-GTF responses were seen in immunized subjects. Oral immunization with a dehydrated liposome-protein vaccine was effective in inducing a secretory IgA antibody response, which was primarily of the IgA: subclass. These results provide the first evidence for the use an oral dehydrated liposome-protein vaccine in humans.     

Nasal immunization of humans with dehydrated liposomes containing Streptococcus mutans antigen

Five healthy female adult volunteers were intranasally immunized twice (7-day interval) with 250 μg of a crude glucosyltransferase (GTF) preparation from Streptococcus mutans in liposomes. Parotid saliva, nasal wash, and serum were collected prior to and at weekly intervals for 6 weeks following the first immunization for analysis of anti-GTF activity by enzyme-linked immunosorbent assay. The levels of IgA1 anti-GTF activity increased in the nasal wash from all five individuals after immunization. Increases in salivary IgA1 and IgA2 anti-GTF activities were observed to a lesser extent. Increased serum IgM and IgA (but not IgG) anti-GTF activities were seen in immunized subjects. Nasal immunization with a dehydrated liposome-protein vaccine was effective in inducing an apparent secretory IgA antibody response, which was primarily of the IgAl subclass. These results provide the first evidence of the effective use of a nasal liposome-protein vaccine in humans.     

Intranasal immunization of humans with Streptococcus mutans antigens

To evaluate the effectiveness of a low dose of soluble or liposomal (L) glucosyltransferase-enriched preparation (E-GTF) in inducing mucosal immune responses after intranasal immunization, 12 adults were immunized on days 0 and 7 by the IN route with 62.5 microg of soluble E-GTF or L-E-GTF. An increase in the mean salivary IgA anti-E-GTF response (P < 0.03) was seen in the L-E-GTF but not the soluble E-GTF group. A significant increase (P < 0.05) in the mean specific IgA antibody activity was also seen in nasal wash from both groups. Although the nasal wash responses were higher in the L-E-GTF than in the soluble E-GTF group, they were not significantly different. The soluble E-GTF immunized group showed a higher serum IgG response than the L-E-GTF immunized group on day 90 (P < 0.05). These results indicate that as little as 62.5 microg of E-GTF, when given by the intranasal route, induced an IgA response in secretions.     

Immune response in humans to a nasal boost with Streptococcus mutans antigens

We previously reported that a Streptococcus mutans enriched-glucosytransferase (E-GTF) preparation induces an immune response following intranasal, but not tonsillar, immunization of humans. In this study, we determined whether intranasal immunization of these subjects 2 years later resulted in augmented immune responses compared to those seen in control subjects. Subjects previously immunized via the intranasal (IN, n = 7) or tonsillar (IT, n = 7) route and control (n = 12) subjects were immunized via the intranasal route with E-GTF. Nasal wash, saliva, and serum were collected before immunization and then weekly for 3 months after immunization. Significant (P < 0.05) mucosal and serum immunoglobulin A (IgA) anti-E-GTF responses were observed in all three groups. Nasal and serum IgA anti-E-GTF responses were significantly higher (P < 0.05) in the IN group. The salivary responses in the three groups were, in general, similar. These results indicate that intranasal immunization primes the immune system for a localized secondary response to S. mutans antigens.     

Anti-cell-associated glucosyltransferase immunoglobulin Y suppression of salivary mutans streptococci in healthy young adults

BackgroundThe authors evaluated the suppressive effects of lozenges containing egg yolk antibodies (that is, immunoglobulin Y [IgY]) against Streptococcus mutans cell-associated glucosyltransferase (CA-gtf) on oral colonization by mutans streptococci (MS) in healthy young adults.MethodsIn a five-day double-masked placebo-controlled trial, young adult participants self-administered lozenges containing anti–CA-gtf IgY (Ovalgen DC, GHEN, Gifu-City, Japan) or a placebo at prescribed times each day. On the basis of bacterial colony counts of saliva cultures, the authors analyzed the pretrial and posttrial differences in levels of MS and total anaerobic bacteria among participants in the treatment (anti–CA-gtf IgY) and placebo groups and a control group.ResultsSalivary MS scores in participants in the treatment group decreased significantly (P < .001), and the mean anaerobic bacterial count in the treatment group was not statistically different before and after the trial. In the placebo and control groups, posttrial changes in median MS scores and total salivary anaerobic bacterial counts were not statistically significant.ConclusionsThe results of the study show that lozenges containing anti–CA-gtf IgY can suppress oral colonization by MS in healthy young adults.Clinical ImplicationsLozenges containing anti–CA-gtf IgY may help reduce dental caries risk in humans.     

Maternal caries incidence and salivary close-contacts with children affect antibody levels to Streptococcus mutans in children

Serum and salivary immunoglobulins and antibodies reactive with Streptococcus mutans were determined in 67 5-8-year-old children. The children of mothers whose caries incidence rates exceeded the median value had significantly more serum IgG antibodies to S. mutans than those with a lower maternal caries activity (p less than 0.05). The children (n = 14) who had been exposed to frequent maternal salivary close-contacts in their first year had significantly higher (p less than 0.05) serum total specific IgG antibodies but significantly lower (p less than 0.01) high-avidity IgG antibodies to S. mutans than the other children (n = 53) with less frequent close-contacts. The maternal caries incidence rates were significantly positively associated with children's dfs/DFS-indices (r = 0.41; p less than 0.001). However, the high maternal caries incidence did not increase the risk of caries in those children whose mothers' saliva contained high amounts of lactobacilli during the first nursing year, when compared with matched children with a low maternal level of lactobacilli. This was possibly due to the fact that the former children had significantly (p less than 0.05) more anti-S. mutans IgG antibodies in their sera than the latter.     

Streptococcus mutans in plaque and saliva after mouthrinsing with SnF2

Abstract – Mouthrinses with SnF2 in 11 subjects significantly reduced ( P <0.01) the total colony forming units (CFU) count and the numbers of Streptococcus mutans and Streptococcus sanguis in plaque. The numbers of S. mutans and S. sanguis were significantly more reduced than was the total CFU count. After rinsing with SnF2 S. sanguis was present in 97% and S. mutans in only 42% of plaque samples from tooth surfaces where they were detected after rinsing with water. SnF2 also significantly reduced ( P <0.01) the number of S. mutans in saliva. Mouthrinses with NaF did not markedly affect the number of S. mutans either in plaque or in saliva.     

Effect of an essential oil-containing antiseptic mouthrinse on plaque and salivary Streptococcus mutans levels

BACKGROUND: Clinical studies in which antimicrobial mouthrinses were shown to have significant antiplaque activity most frequently have used gingivitis as the clinically relevant endpoint. However, there is evidence to suggest that mouthrinses containing active agents effective against Streptococcus mutans, such as chlorhexidine, may also have a role in inhibiting dental caries. This clinical study was conducted to determine the effect of 2x-daily rinsing with an essential oil-containing antiseptic mouthrinse (Listerine Antiseptic) on levels of recoverable S. mutans and total streptococci in supragingival interproximal plaque and in saliva. Additionally, a follow-up in vitro study is reported which determined whether a differential susceptibility to the antiseptic mouthrinse exists among different strains of streptococci. METHOD: Following baseline saliva and plaque sampling for quantification of recoverable S. mutans and total streptococci, 29 qualifying subjects were randomly assigned either the essential oil mouthrinse or a sterile water control. They rinsed with 20 ml for 30s 2 x daily for 11 days and once on the 12th day, in addition to their usual oral hygiene procedures. On day 12, saliva and plaque samples were again collected and microbiological quantification performed. The procedures were repeated with the alternate rinse after a 1-week washout period. RESULTS: The essential oil mouthrinse produced respective reductions of 69.9% and 75.4% in total recoverable streptococci and in S. mutans in plaque, and corresponding reductions of 50.8% and 39.2% in saliva. The in vitro study revealed that streptococci from the mutans group were more susceptible to the bactericidal activity of the essential oil mouthrinse than streptococci from the mitis group. CONCLUSIONS: As antimicrobial mouthrinses are most frequently recommended to patients whose mechanical oral hygiene procedures are not adequate for the control of supragingival plaque and gingivitis, this study provides an additional rationale for the inclusion of the essential-oil mouthrinse as an adjunct to daily oral hygiene procedures.     

Effect of SnF2, administered as mouthrinses or topically applied, on Streptococcus mutans, Streptococcus sanguis and lactobacilli in dental plaque and saliva

Abstract – Mouthrinsing with SnF₂ reduced the Streptococcus mutans population in plaque and saliva and the proportion of Streptococcus sanguis in plaque. The effect was of short duration: 2 weeks after treatment the values of S. mutans in plaque and saliva were even higher than the pretreatment values. Topical SnF₂ applications reduced the S. mutans population in plaque and saliva but did not reduce the proportion of S. sanguis in plaque. The eflect was more prolonged: 4 weeks after treatment the S. mutans population in interproximal plaque remained significantly reduced and the salivary levels of the organism had not fully returned to pretreatment levels. Both SnF₂ treatments significantly increased the salivary levels of lactobacilli. The values of laclobacilli in saliva remained signilicantly increased 4 weeks after the SnF₂ mouthrinsing but had almost returned to pretreatment levels within 2 weeks after the topical SnF₂ applications. The findings suggest that the cariogenic potential of dental plaque is differently affected depending on whether a drug is administered as a mouthrinse or is applied topically.     

树脂充填前后唾液中变异链球菌的变化

目的 观察使用树脂进行龋齿治疗前后唾液中变异链球菌数量的变化.方法 招募志愿者20名,对其口腔内所有龋齿去净龋坏组织后使用不含氟树脂材料进行充填,分别收集充填前及充填后第1、2、3、4周的非刺激性唾液样本,使用轻唾-杆菌肽琼脂培养基厌氧培养48 h后检测变异链球菌数量,比较其在龋齿充填治疗前后的变化.结果 龋齿行树脂材料充填后1周唾液中变异链球菌数量较充填前下降明显(P＜0.05),充填后2、3、4周较充填前无明显变化(P＞0.05).结论 对患龋个体进行龋齿备洞后的树脂充填能在短期内降低唾液中变异链球菌的含量,但随时间推移,唾液中变异链球菌数量会逐渐恢复到治疗前的状态.单纯依靠龋齿充填治疗并不能改变唾液中变异链球菌的数量.     

Serum antibody responses to Streptococcus mutans antigens in humans systemically infected with oral streptococci

Sera from patients with subacute bacterial endocarditis (SBE) due to Streptococcus mutans or other oral streptococci and from normal subjects were assayed by enzyme-linked immunosorbent assay for antibodies to defined S. mutans antigens. Antibodies of IgG and IgA isotypes to Ag I/II and Ag III were greatly elevated in S. mutans-SBE sera, and the IgA antibodies in 3 sera included both polymeric and monomeric forms. Elevated IgM and IgG anti-lipoteichoic acid and IgG and IgA anti-serotype c polysaccharide antibodies were also found. The sera of 4 of 6 patients infected with other oral streptococci also displayed antibodies to S. mutans Ag I/II. Sera of 3 patients infected with Streptococcus mitis or Streptococcus oralis, but none of the S. mutans-infected cases, showed elevated antibodies to human heart sarcolemma, and all SBE sera had elevated rheumatoid factor. These results suggest that the known surface protein antigens of S. mutans are immunodominant in humans, and are not likely to be heart cross-reactive.     

Primary reservoirs of Streptococcus mutans and their relationship to caries experience in adults with good oral hygiene

Primary reservoirs of Streptococcus mutans were identified in a group of 21 adults with intact dentitions and good oral hygiene by obtaining multiple plaque samples from all available tooth surfaces. Routine cultural methods and a rapid plate scoring method employing a 7-point ordinal scale were used to assess S. mutans levels. Oral hygiene status (DI-S) and caries experience (DMFT and DMFS) were determined clinically. Primary S. mutans reservoirs (high scores) were restricted essentially to the posterior interproximal areas and occasionally in the occlusal pits and fissures of partially erupted mandibular third molars. An anterior to posterior gradient of increasing S. mutans scores was observed for interproximal sites. Oral hygiene scores correlated poorly with DMFT, DMFS and the S. mutans scores obtained for different anatomic locations.     

变异链球菌葡糖基转移酶植物表达质粒p2355-gtfB的构建

目的构建含变异链球菌葡糖基转移酶多个免疫显性抗原表位的植物表达质粒p2355-gtfB,为其转化植物研究奠定物质基础,为可食防龋疫苗研究提供条件。方法以真核表达质粒pcDNA3-gtfB为模板,通过聚合酶链反应(polymerase chain reaction,PCR)扩增含编码变异链球菌葡糖基转移酶抗原表位的目的基因gtfB。将回收纯化的PCR产物经T-A克隆技术克隆于中间载体pMD18-T,双酶切鉴定插入方向后,将目的基因从中间载体释放,再克隆至高效的植物表达载体p2355,用电转化法转化根癌农杆菌EHA105,对重组质粒阳性克隆株进行筛选与鉴定。结果通过对重组质粒T-gtfB进行酶切图谱分析,获得2.9 kb和3.7 kb的2个片断,将重组质粒p2355-gtfB进行酶切、PCR及测序分析,显示p2355-gtfB中插入基因长度为3 675 bp,基因序列与双脱氧链终止法的测定结果相同,证明植物表达质粒p2355-gtfB构建成功,开放阅读框架正确。结论本研究成功构建了含变异链球菌多个葡糖基转移酶免疫显性抗原表位编码基因的植物表达质粒p2355-gtfB,为可食性防龋疫苗的研究奠定了基础。     

Crevicular IgG antibodies and recovery of locally implanted Streptococcus mutansR in humans

This study was undertaken with the aim of examining the effect of highly different IgG antibody activities in gingival crevicular fluid (GCF) on the elimination of a streptomycin-resistant strain of Streptococcus mutans (S. mutans) topically implanted on the tooth. Two groups of people were selected for the study, one with high and one with low specific IgG antibody activity (IgG antibody activity against S. mutans) in GCF. During an experimental period of 15 d the specific IgG antibody activity in GCF in the same individual stayed on a relatively constant level. Between individuals, however, the specific IgG antibody activity varied considerably. The number of indigenous mutans streptococci and the number of implanted bacteria on the same tooth surface were strongly correlated. The implanted S. mutans strain was rapidly eliminated in all subjects. No difference in the elimination of the implanted strain of S. mutans could be demonstrated between the groups of high and low specific IgG antibody activity.     

变形链球菌GbpB融合蛋白免疫SD大鼠实验研究

目的 研究变形链球菌GbpB融合蛋白免疫别大鼠时，其所诱导的血清和唾液的特异性免疫反应。方法 用纯化的变形链球菌GbpB融合蛋白皮下免疫别大鼠，免疫后间隔一定时间收集大鼠的血清及唾液，间接ELISA检测血清及唾液中特异性抗GbpB抗体。结果 GbpB融合蛋白免疫大鼠，可引起血清中IgG及唾液中IgA的显著提高。另外，GbpB融合蛋白免疫大鼠，38天采血，血清中抗GbpB抗体效价最高，以后逐渐降低。结论 变形链球菌GbpB融合蛋白具有免疫原性，可供研制防龋疫苗。     

变形链球菌GbpA的GBD基因疫苗动物免疫防龋研究

目的：研究变形链球菌GbpA的GBD基因疫苗免疫SD大鼠时,其所诱导的血清和唾液特异性免疫反应及防龋效果。方法：实验分2组,实验组用纯化的变形链球菌GbpA的GBD真核表达质粒pcDNA3．1-GBD颌下腺周围皮下免疫SD大鼠,其中实验1组免疫后间隔一定的时间收集大鼠的血清及唾液,间接ELISA检测血清及唾液中特异性抗GBD抗体,实验2组免疫同时喂致龋饲料—Keyes改良的高糖Diet 2000,并于第一次注射20d时连续3d大鼠口腔中接种S．mutans Ingbritt,在接种S．mutans后第77 d处死大鼠,收集大鼠颌骨标本用于龋齿记分分析。对照组2只大鼠颌下腺周皮下注射1×PBS(pH7．4)。结果：用纯化的pcDNA3．1-GBD基因疫苗免疫大鼠,血清中IgG及唾液中IgA明显升高,大鼠高糖饮食及口腔接种S．mutans Ingbritt,实验组的龋患率明显低于对照组,说明GBD基因疫苗具有免疫防龋作用。结论：变形链球菌GbpA的GBD基因疫苗具有免疫原性,是一种有效的免疫防龋疫苗。     

Oral glucose clearance in subjects with high or low salivary levels of Streptococcus mutans and lactobacili

Abstract — The ability to eliminate sugar from the oral cavity was studied in subjects with high or low numbers of salivary Streptococcus mutans and lactobacilli. The glucose concentration in saliva was determined after consumption of biscuits using the Gloxtest, and the logarithms of the glucose values were plotted against time. The sugar clearance time was measured where the slope of the plot intersected with a glucose level of I mg/ml. Subjects with a high level of S, mutans had a significantly longer clearance time than subjects with a low level. More salivary lactobacilli were found in a subject group- with a long sugar clearance time than in a group with a fast clearance rate.     

不同龋敏感儿童口腔变异链球菌不同基因型临床分离株产酸力的实验研究

目的比较不同龋敏感儿童口腔变异链球菌不同基因型临床分离株对蔗糖进行酵解产生有机酸的能力。方法从不同龋敏感的3~5岁学龄前儿童口腔变异链球菌中选取66株变异链球菌临床分离株,用气相色谱法测定高龋、中龋和无龋儿童口腔变异链球菌不同基因型临床分离株产生各有机酸的量。结果不同龋敏感儿童不同基因型变异链球菌临床分离株产生有机酸的量的差异有统计学意义（P<0.05）,同一龋敏感儿童变异链球菌携带不同基因型菌株产酸量的差异也有统计学意义（P<0.05）,携带基因型数目越多的菌株其产酸量越多。变异链球菌临床分离株产生乳酸、乙酸、甲酸的量的差异有统计学意义（P<0.05）,乳酸多于乙酸和甲酸。结论不同龋敏感儿童口腔变异链球菌不同基因型菌株产酸力不同,携带基因型数目越多的菌株其产酸力越强。     

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目的    探讨2岁儿童口腔菌斑中变形链球菌（变链菌）的检出率和蔗糖依赖性黏附能力。方法    选取广州市郊花都区2008年8—9月于入托前在妇幼保健院儿保科体检的2岁高龋（龋失补牙数≥ 5）及无龋（龋失补牙数 = 0）儿童各32名,形态学、生理生化学方法对其牙菌斑进行分离鉴定;采用分光光度比浊法,检测高龋儿童和无龋儿童变链菌临床分离株对玻壁的黏附情况。结果    64名儿童中变链菌的检出率为56.25%,高龋组儿童变链菌检出率（75.00%）显著高于无龋组儿童（37.50%）,差异有统计学意义（P < 0.05）。生化鉴定获得高龋儿童75株和无龋儿童34株变链菌临床分离株。在含1%蔗糖的培养基中,高龋组变链菌分离株对玻壁的黏附比为0.58 ± 0.15,略高于无龋组0.56 ± 0.13,但两组间差异无统计学意义。结论    2岁儿童变链菌检出率与高发龋相关,而未见其蔗糖依赖性黏附力对龋病高发有影响。     

Comparison of effect of SnF2 and NaF mouthrinses on caries incidence, salivary S. mutans and gingivitis in high caries prevalent adults

Abstract – The effect of SnF₂ and NaF mouthrinses were compared in subjects initially selected because of their high caries prevalence and high levels of salivary S. mutans. Subjects rinsed twice a day with either SnF₂ or NaF (200 ppm F⁻). Subjects in both groups continued to be caries active during the course of the study. However, at the first year examination subjects rinsing with SnF₂ had less S. mutans/ml saliva, a lower caries incidence and less gingivitis. After 2 yr the S. mutans levels in the SnF₂ rinsing group remained significantly less. Both caries incidence and gingivitis were lower in the SnF₂ group after 2 yr but the difference was not significant.