Recrudescence of herpes simplex virus type 1 in latently infected rats after trauma to oral tissues

Tooth extraction in rats was used to trigger a latent HSV-1 infection. HSV-1 was inoculated unilaterally in the rat palates, tight weeks later two molars were removed bilaterally. The trigeminal ganglia were co-cultivated and HSV-1 was isolated from 63% of the ganglia on the infected sides hut from only 11% on control sides. The immune response pattern was analysed by immunoblotting of rat serum, and strong reactivity to HSV-1 specific cell polypeptides and glycoprotems (ICP6, gC, pgC, gD) was seen after reactivation. The extraction sockets were histopathologically evaluated and showed healing on the infected side in 26%, compared to 63% in contralateral control sockets. The effect of acyclovir (ACV) treatment was elucidated and was found to influence the subsequent development of antibodies and to promote healing of the sockets. Vesiculation in intra- and subepithelial tissue was present on the infected side in 58% but in only 12% of ACV-treated animals. The present study in rats has shown that a latent HSV-1 infection can be established and reactivated by tooth extraction. Reactivation resulted in delayed healing of sockets on the latently infected side but not on the Contralateral control side. HSV-1 reactivation was demonstrated serologically by immunoblotting. Healing was significantly promoted by administration of ACV. which also supports the contention that HSV-1 interferes with the healing process.     

单纯疱疹病毒Ⅰ型与白塞病关系的探讨

为了探讨单纯疱疹病毒(HSV-1)与白塞病(Behcet's disease,BD)的关系,本研究应用聚合酶链反应(P('R))技术检测38例BD患者口腔溃疡愈合前后的HSV-1 DNA,免疫组化技术观察溃疡粘膜HSV-1抗原表达。结果,溃疡刮片标本HSV-1 DNA检出率为34.2％(13/38),对照组为6.7％(2/30);原刮片部位之愈后粘膜HSV-1 DNA检出7例,占愈合粘膜标本的53.8％(7/13),HSV-1抗原表达阳性率为61.3％(8/13);愈后粘膜HSV-1 DNA阳性者,其HSV-1抗原表达亦全部阳性。这些结果提示HSV-1可能参与BD病损形成,病损愈后的粘膜仍有HSV-1潜伏及其基因表达。     

单纯疱疹病毒Ⅰ型体外感染人口腔上皮细胞的实验研究

目的研究单纯疱疹病毒Ⅰ型(HSV-1)体外感染人口腔上皮细胞的途径和方式。方法在体外利用非洲绿猴肾细胞大量扩增获得HSV-1,将此病毒体外感染人口腔上皮细胞,建立HSV-1体外感染人口腔上皮细胞模型,并收集HSV-1感染人口腔上皮细胞后的上清液转移感染Vero细胞。利用倒置显微镜对病毒感染进行形态学鉴定。应用聚合酶链式反应技术(PCR)对病毒核酸进行检测。结果倒置显微镜下人口腔上皮细胞未观察到典型的细胞病变,上清液转移感染后的Vero细胞发生典型细胞病变。利用PCR法可在单纯疱疹病毒Ⅰ型感染后的人口腔上皮细胞内检测到病毒核酸。结论HSV-1可直接感染人口腔上皮细胞。     

Effect of essential oils containing and alcohol-free chlorhexidine mouthrinses on cariogenic micro-organisms in human saliva

Abstract

Objective. The aim of this study was to evaluate the effect on mutans streptococci and lactobacilli in saliva of mouthrinsing with essential oils and an alcohol-free chlorhexidine. Materials and method. Twenty healthy volunteers (mean age 59 years) participated in the double-blind randomized cross-over study. Three mouthrinses were used in 16 days rinsing periods in addition to their regular mechanical oral hygiene: a solution with essential oils (EO; Listerine), a solution with alcohol-free chlorhexidine (CHX; Paroex) and water (negative control). The mouthrinse periods were separated by 3-month washout periods. At days 0 (baseline) and 17 (end) of each mouthrinse period, paraffin stimulated whole saliva was collected in order to analyse CFU/ml saliva of mutans streptococci and lactobacilli. Results. Only the CHX rinse showed a significant difference for CFU mutans streptococci between baseline and end (p = 0.004). The CFU mutans streptococci at the end of the rinse periods showed statistically significant differences between CHX vs EO (p = 0.039) and CHX vs water (p = 0.022). The difference in CFU lactobacilli between baseline and end was significant for CHX (p = 0.031), but not for the other rinses. No statistically significant differences for lactobacilli were found at the end of the rinse periods between the mouthrinses. Conclusion. A significant reduction in amount of cariogenic bacteria in saliva was observed after 16 days of alcohol-free chlorhexidine mouthrinse but not after the essential oils rinse. The high number of participant's not changing to a bacterial class with a reduced number of micro-organisms showed that both rinses had little clinical significance as a caries preventing treatment method, which can decrease the number of CFU cariogenic micro-organisms.     

Detection of herpes simplex virus type 1 shedding in the oral cavity by polymerase chain reaction and enzyme-linked immunosorbent assay at the prodromal stage of recrudescent herpes labialis

Recrudescent herpes labialis (RHL) is a disease caused by herpes simplex virus (HSV). predominantly type 1 (HSV-1). We have monitored HSV-1 shedding in the oral cavity by polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISA) using digoxigenin-labeled primers designed to amplify a 278 bp segment of the HSV-1 UL 42 region. Prodromal RHL was confirmed by thermographic imaging in 22 patients. Infectious virus was not detected using tissue culture for virus isolation (0/22). Using PCR and agarose gel electrophoresis. We could detect HSV-1 DNA in 8/22 patients. Using a biotinylated-probe internal to the predicted sequence of the PCR product. HSV-1 DNA was detected in 10/22 of the patients by ELISA. We conclude that HSV-1 DNA is shed into the oral cavity of patients presenting with sub-clinical RHL and that the PCR-EL1SA technique represents a more sensitive method to monitor HSV-1 shedding than conventional tissue culturing or PCR-electrophoresis alone.     

The controversial natural history of oral herpes simplex virus type 1 infection

The natural history of oral herpes simplex virus type 1 (HSV‐1) infection in the immunocompetent host is complex and rich in controversial phenomena, namely the role of unapparent transmission in primary infection acquisition, the high frequency of asymptomatic primary and recurrent infections, the lack of immunogenicity of HSV‐1 internalized in the soma (cell body) of the sensory neurons of the trigeminal ganglion, the lytic activity of HSV‐1 in the soma of neurons that is inhibited in the sensory neurons of the trigeminal ganglion and often uncontrolled in the other neurons, the role of keratin in promoting the development of recurrence episodes in immunocompetent hosts, the virus–host Nash equilibrium, the paradoxical HSV‐1‐seronegative individuals who shed HSV‐1 through saliva, the limited efficacy of anti‐HSV vaccines, and why the oral route of infection is the least likely to produce severe complications. The natural history of oral HSV‐1 infection is also a history of symbiosis between humans and virus that may switch from mutualism to parasitism and vice versa. This balance is typical of microorganisms that are highly coevolved with humans, and its knowledge is essential to oral healthcare providers to perform adequate diagnosis and provide proper individual‐based HSV‐1 infection therapy.     

单纯疱疹病毒1型体外感染人口腔上皮细胞的初步探讨

目的 观察单纯疱疹病毒1型(herpes simplex virus type 1,HSV-1)体外感染人口腔上皮细胞的途径和方式,为研究HSV-1导致牙周病的机制提供依据.方法 在体外扩增获得HSV-1,将其感染人口腔上皮细胞,再转移感染Vero细胞.倒置显微镜及透射电镜下观察.应用聚合酶链反应(PCR)技术对病毒核酸进行检测.结果 倒置显微镜下人口腔上皮细胞未见细胞病变,转移感染后的Vero细胞发生典型细胞病变.透射电镜下在人口腔上皮细胞内未观察到病毒颗粒,在转移感染后的Vero细胞里可见典型的病毒颗粒.HSV-1感染后的人口腔上皮细胞内可检测到病毒核酸.结论 HSV-1可直接感染人口腔上皮细胞,为进一步研究HSV-1导致牙周病的机制提供了较好的体外模型.     

The effect of herbal,essential oil and chlorhexidine mouthrinse on de novo plaque formation

To cite this article:
Int J Dent Hygiene DOI: 10.1111/j.1601‐5037.2012.00556.x
Singh A, Daing A, Dixit J. The effect of herbal, essential oil and chlorhexidine mouthrinse on de novo plaque formation. Abstract: Background: Brushing and flossing are the most widely accepted procedures, the ‘gold standard’, for controlling bacterial plaque, but these mechanical methods have limitations. Based on results derived from several clinical trials, essential oil (EO) mouthrinse (Listerine^®) and a chlorhexidine mouthrinse have been accepted by ADA to be used as an adjunct to routine mechanical oral hygiene measures however, both of them are associated with side effects, therefore, the present study was undertaken to evaluate the antiplaque efficacy of a new herbal formulation as compared to an EO and chlorhexidine rinse. Materials and method: The study was a single blind parallel randomized controlled trial involving four groups. 48 volunteers refrained from all oral hygiene measures for 4 days, but rinsed instead twice daily with 10 ml of a herbal (HM), EO, chlorhexidine (CHX) or a placebo (PL) solution. Plaque index and plaque area (PA) was assessed on Day 4. Results: The HM and EO showed a significant inhibition of plaque regrowth compared to PL (P < 0.001), but the lowest values of PI and PA were obtained with CHX. Statistically significant difference in plaque parameters was observed when CHX was compared to HM and EO, and HM to EO rinse. Conclusion: The new herbal mouthrinse had a promising plaque inhibitory potential but it not as efficacious as chlorhexidine in preventing plaque regrowth.     

1型单纯疱疹病毒三叉神经节潜伏感染再激活对大鼠三叉神经痛阈的影响

目的 建立SD大鼠1型单纯疱疹病毒（HSV-1）三叉神经节潜伏感染和再激活的动物模型,利用痛阈检测观察HSV-1神经节感染和再激活对三叉神经痛阈的影响。方法 SD大鼠随机分为实验组和对照组,实验组角膜划痕滴加 HSV-1接种病毒,建立神经节潜伏感染模型;对照组仅滴加生理盐水。8周后,再将实验组大鼠分为紫外照射组和非紫外照射组,实施紫外线照射,干预诱导潜伏感染再激活。利用Von Frey纤维丝测痛仪检测各组大鼠触须垫机械痛阈。提取大鼠三叉神经节组织,RT-PCR检测HSV-GD、LAT、ICP27的表达水平,验证HSV-1神经节潜伏感染和再激活是否成功。结果 大鼠HSV-1神经节潜伏感染和再激活成功;实验组大鼠在HSV-1潜伏感染状态下和再激活状态下,发生疼痛刺激样反应,痛阈有降低趋势,尤其在再激活后更加明显。结论 三叉神经节HSV-1潜伏感染再激活可以诱发动物三叉神经痛阈降低,提示HSV-1可能是三叉神经痛的病因。     

Subclinical reactivation of herpes simplex virus type 1 in the oral cavity

Reactivation in the oral cavity either symptomatically (recrudescence) or without symptoms (recurrence) may contribute to the transmission of herpes simplex virus type 1 (HSV-1), especially in critical areas of exposure such as dentistry. In order to measure the frequency of HSV-1 reactivation, nested polymerase chain reaction (PCR) was performed on oral swabs collected from 30 healthy people over a period of 58-161 days. In total 19 of 25 (76%) seropositive people were PCR-positive at least once, 6 of these 19 (32%) had recrudescence and 13 (68%) had only asymptomatic reactivation. Frequencies of additional recurrences were higher in people showing symptomatic reactivation than in those who had only recurrences. Recrudescence is a risk factor for elevated levels of asymptomatic HSV-shedding. In most cases HSV-1 was detected only by nested PCR investigated by early onset of therapy or time span before sampling.     

Oral pseudomembranous candidiasis, herpes simplex virus-1 infection, and oral mucositis in head and neck cancer patients receiving radiotherapy and granulocyte-macrophage colony-stimulating factor (GM-CSF) mouthwash

Oral pseudomembranous candidiasis (OPC) was evaluated in 61 patients receiving head and neck radiotherapy (RT). Herpes simplex virus-1 (HSV-1) reactivation was also investigated in 14 patients. According to the agreed protocol, granulocyte-macrophage colony-stimulating factor (GM-CSF) mouthwash was administered in 46 patients with radiation-induced ulcers. Candidiasis was diagnosed in 31 patients. Candida albicans was the most frequent isolate. Multiple Candida species were isolated from the lesions of four patients. Concurrent candidiasis and radiation-induced ulcers were observed in 17 patients. Viral culture and the polymerase chain reaction disclosed the presence of HSV-1 in five patients. Twenty of the 46 patients, with initial mucositis grade II and grade III, completed RT with mucositis grade I, indicating a beneficial effect of GMCSF mouthwash, although further controlled studies are necessary to verify that. In conclusion, OPC was an important infection in patients undergoing radiotherapy. The role of HSV-1 in oral mucositis during head and neck radiotherapy needs additional study.     

The effect of polyvinyl pyrrolidone on the clinical activity of 0.09% and 0.2% chlorhexidine mouthrinses

BACKGROUND: Previous studies have shown that polyvinyl pyrrolidone (PVP) added to a chlorhexidine rinse reduced extrinsic dental stain but at the expense of a reduction in plaque inhibitory activity. This effect appeared due to a reduction in the effective chlorhexidine dose to levels where dose response studies show plaque inhibition falls off rapidly. The aim of these 2 clinical studies was to determine if PVP could be added to chlorhexidine rinses to maintain efficacy and reduce staining. METHOD: Study 1 involved 42 healthy dentate volunteers and was a blind, randomised, 7 treatment, crossover design balanced for residual effects. The rinses were: 1. 0.09% chlorhexidine to which was added, 2. 1% PVP, 3. 3% PVP, 4. 5% PVP, 5. 7% PVP, 6. Placebo, 7. Essential oil product. Rinses were used 2x on day one of each period after a prophylaxis. Subjects suspended tooth cleaning for 24 h and were then scored for plaque area. Study 2 used the experimental gingivitis model, involved 24 healthy dentate subjects and was a blind, randomised, 3 treatment, crossover design balanced for residual effects. The rinses were 1. 0.2% chlorhexidine, 2. 0.2% chlorhexidine/10% PVP, 3. Placebo. At baseline and the end of each study period subjects were rendered plaque, stain and calculus free, suspended oral hygiene and rinsed 2x per day. Plaque, gingivitis and stain were scored at baseline, 1, 2, and 3 weeks. Calculus was scored at baseline and 3 weeks. RESULTS: Study 1: Buccal plaque scores were significantly lower with all rinses compared to placebo. Also all buccal plaque scores were significantly lower with chlorhexidine and chlorhexidine/PVP rinses compared to the essential oil/phenolic rinse. There were no significant differences between the chlorhexidine rinse and the chlorhexidine/PVP rinses. Analyses for buccal and lingual plaque combined produced, with one exception, the same results for rinse comparisons as for buccal plaque alone. Thus the essential oil/phenolic rinse just failed to reach significance compared to placebo. Study 2: Plaque and gingivitis scores were significantly lower with positive control and test rinses compared to placebo but with no difference between these rinses. Tooth and tongue stain was significantly higher with the positive control and test rinses compared to placebo but not significantly different between these 2 rinses. Calculus scores were not significantly different between the three study rinses. CONCLUSION: Taken with previous data, the balance of evidence does not support PVP as an inhibitor of staining associated with chlorhexidine. These data are further evidence that chlorhexidine oral hygiene products, which, do not or claim not to cause staining, are most probably lacking efficacy.     

Herpes simplex virus type 1 and type 2 infection in human oral mucosa in culture

To examine the sensitivity of human oral mucosa to herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) infection, human gingival mucosa explants were infected with either HSV-1 or HSV-2 in vitro and the expression of virus specific antigen was examined by the immunofluorescent antibody technique. HSV-2 antigen was found in the basement membrane, basal cell layer and lower prickle cell layer. This finding was consistent with the HSV-1 infection. Electron microscopic study revealed the presence of nucleocapsids and enveloped virus particles in the basal cells of HSV-2-infected organ cultures. These findings indicate that human gingival mucosa is sensitive to infection with HSV-2, as well as HSV-1, and that the virus may replicate in the undifferentiated epithelial cells of mucosal epithelium.     

Striated muscle involvement in experimental oral infection by herpes simplex virus type 1

Herpes simplex virus type 1 is one of the most frequent causes of oral infection in humans, especially during early childhood. Several experimental models have been developed to study the pathogenesis of this virus but all of them employed adult animals. In this work, we developed an experimental model that uses mice younger than 4 days old, to more closely resemble human infection. Mice were infected subcutaneously with the prototype strain McIntyre of Herpes simplex‐1, and the progression of infection was studied by immunoperoxidase. All animals died within 24–72 h post‐infection, while viral antigens were found in the oral epithelium, nerves and brain. The most striking result was the finding of viral antigens in the nucleus and cytoplasm of cells belonging to striated muscles. Organotypic cultures of striated muscles were performed, and viral replication was observed in them by immunocytochemistry, electron microscopy and viral isolation. We conclude that the infection of striated muscles is present from the onset of oral infection and, eventually, could explain some clinical observations in humans.     

Modulation of herpes simplex virus type 1 replication by human salivary secretions

Saliva functions to protect the oral cavity from pathogenic invasion by modulating the ability of microbes to colonize the oral surfaces or limiting their growth and/or viability. Although the role of salivary secretions in the modulation of the oral bacteria flora has received considerable attention, little is known concerning its role in viral pathogenesis. Accordingly, the purpose of this study was to assess the effect of salivary secretions on herpes simplex virus type 1 (HSV-1) replication. Initially, HSV-1 plaque and liter reduction assays were performed to determine the ability of human submandibular/sublingual (HSMSL) and parotid (HPS) salivas to inhibit the early stages of HSV-1 infection (adsorption and penetration). Our results suggested that both HSMSL and HPS possess cell-protective and virus neutralization activities, with HSMSL being more active than HPS. Additional experiments were performed to determine the effect of saliva on the yield of virus progeny. Again, HSMSL caused a greater reduction of HSV-1 replication than did HPS. A similar effect could not be obtained using vaccinia, suggesting that this inhibitory activity of human saliva is selective. Collectively, these results suggest that human salivary secretions can modulate the replication of HSV-I in vitro.     

Oral shedding of herpes simplex virus type 1 in immunocompetent persons

BACKGROUND: Reactivation of herpes simplex virus-1 (HSV-1) can result in recurrent herpes labialis lesions (RHL) and in oral shedding of virus. This study utilized polymerase chain reaction (PCR) to document the frequency and quantity of such shedding. METHODS: Thirty adults with greater than three RHL episodes per year were followed through one recurrence. They collected swabs for PCR every 12 h starting at prodrome and for 10 days thereafter. Shedding was analyzed with regard to frequency, timing and quantity. RESULTS: HSV-1 was detectable in 87% of participants for a mean of 4 days. Shedding occurred most frequently during the vesicle/ulcer stage (91% of subjects), but was common in both clinical and subclinical stages (50% vs. 23%, average log DNA copy number/ml(2) 2.6 vs. 1.4). CONCLUSION: The majority of RHL patients shed viral DNA. Shedding occurred before and after the appearance of clinical lesions. Such findings may be useful in designing methods to reduce viral shedding and prevent transmission.     

Enhanced replication of herpes simplex virus type 1 in human cells.

The effects of DNA-damaging agents on the replication of herpes simplex virus type 1 (HSV-1) were assessed in vitro. Monolayers of human lung fibroblast cell lines were exposed to DNA-damaging agents (methyl methanesulfonate [MMS], methyl methanethiosulfonate [MMTS], ultraviolet light [UV], or gamma radiation [GR]) at specific intervals, before or after inoculation with low levels of HSV-1. The ability of cell monolayers to support HSV-1 replication was measured by direct plaque assay and was compared with that of untreated control samples. In this system, monolayers of different cell lines infected with identical HSV-1 strains demonstrated dissimilar levels of recovery of the infectious virus. Exposure of DNA-repair-competent cell cultures to DNA-damaging agents produced time-dependent enhanced virus replication. Treatment with agent before virus inoculation significantly (p less than 0.025) increased the number of plaques by 10 to 68%, compared with untreated control cultures, while treatment with agent after virus adsorption significantly increased (p less than 0.025) the number of plaques by 7 to 15%. In a parallel series of experiments, cells deficient in DNA repair (xeroderma pigmentosum) failed to support enhanced virus replication. These results suggest that after exposure to DNA-damaging agents, fibroblasts competent in DNA repair amplify the replication of HSV-1, and that DNA-repair mechanisms that act on a variety of chromosomal lesions may be involved in the repair and biological activation of HSV-1 genomes.     

Rapid detection of human herpes simplex virus type 1 in saliva.

Herpes simplex virus type 1 (HSV1) can be rapidly identified in saliva from patients with acute herpetic gingivostomatitis, by in vitro amplification using the polymerase chain reaction and specific primers. Amplification of DNA results in a product of 110 bp length corresponding to the region 1381-1490 bp of the HSV1 thymidine kinase gene. The specificity of the reaction was demonstrated in three ways: (i) the presence of a Sma 1 restriction enzyme site in the amplified product sequence; (ii) Southern blot using a biotinylated HSV1-specific oligonucleotide probe and (iii) direct sequencing of amplified product. At high titres of virus (> 5 x 10(5) virions/ml saliva), saliva may be added directly to the amplification assay for detection purposes. However, at lower titres of HSV1 viral DNA must be purified from saliva before in vitro amplification. HSV was identified in the saliva from symptomatic patients with acute herpetic gingivostomatitis and was absent in saliva collected from controls.     

The effect of a polyhexamethylene biguanide mouthrinse compared to an essential oil rinse and a chlorhexidine rinse on bacterial counts and 4-day plaque regrowth

OBJECTIVES: For various clinical applications, polyhexamethylene biguanide hydrochloride (PHMB) has been used for many years as an antiseptic in medicine. Recently, a 0.04% PHMB mouthwash was shown to inhibit plaque regrowth and to reduce oral bacterial counts. In this study, a 0.12% PHMB mouthrinse (A) was compared with a negative control placebo rinse (10% ethanol, flavour) (B), a positive control 0.12% chlorhexidine rinse (C), and a commercially available mouthrinse containing essential oils (Listerine) (D). MATERIALS AND METHODS: The study was a double-blind, randomised 4-replicate 4 x 4 Latin square cross-over design in which plaque regrowth was measured. The in vivo antibacterial effect was assessed by taking bacterial counts from the tooth surface and mucosa 4 h after the first rinse with the preparations on day 1 and prior to the clinical examination on day 5. 16 volunteers participated and, on day 1 of each study period, were rendered plaque-free, ceased toothcleaning, and rinsed 2x daily with the allocated mouthrinse. On day 5, plaque was scored and smears were collected according to the protocol. Washout periods were 9 days. Data were analysed using ANOVA with Bonferroni HSD adjustment for multiple comparisons (significance level alpha=0.05). RESULTS: The 0.12% PHMB mouthrinse (A) was significantly more effective in inhibiting plaque than the placebo (B) but no significant differences could be observed between A and 0.12% chlorhexidine (C), or between A and Listerine (D). Bacterial count reductions on the tooth surface with PHMB (A) were significantly greater compared to the placebo (B) after 4 h and significantly greater compared to B and D after 5 days. Chlorhexidine (C) was more effective than A after 5 days. On the mucosa, chlorhexidine (C) was significantly more effective in reducing bacterial counts than the other 3 treatments at both time points investigated. PHMB (A) was significantly more effective in reducing bacterial counts than the placebo (B) after 4 h and after 5 days, and than D after 4 h. CONCLUSION: Consistent with a previous study, a PHMB mouthrinse was shown to inhibit plaque recolonisation and to reduce oral bacterial counts, indicating that PHMB may find applications in the prevention of plaque-associated diseases.