Poster 020: GFP Transduction Using Adenoviral Vector in Normal Oral Epithelium
1
females) who visited the Department of Oral Medicine of Kagoshima University Hospital were diagnosed to have candida glostitis base on culture methods. We utilized the USB-microscope M2 (Scalar) to thus examine can- dida glostitis which were suspected and at the same time diagnosed by conventional culture methods to detect Candida species. We applied gentian violet (0.1% meth- ylrosaniline chloride solution) on the surface of the tongue and then washed it off using physiological saline and then observed the papillae of the tongue by the USB-microscope M2. Method of Data Analysis: We confirmed the presence of Candida species where the stained area by culture method. Results: Eighteen cases were thus detected to have Candida species by based on the findings obtained using the USB-microscope M2 with gentian violet staining. In all cases some parts of the lingual papillae were stained blue by gentian violet. Candida species were cultured from this part which was stained by gentian violet and pseudohyphae were observed Conclusion: The detection of oral Candida species using microscopic evaluations may therefore be a useful new method. References Tam M.T., et al. Gram stain method shows better sensitivity than clinical criteria for detection of bacterial vaginosis in survellance of pregnant, low-income women in a clinical setting. Infection Disease in obstetric Gynecology; 6:204-208. 1998 Chaiban G., et al. A rapid method of impregnating endtrocheal tubes and urinary catheters with gendine: a novel antiseptic agent. J. of Antimicrobial Chemoteraphy; 55: 51-56. 2005 POSTER 020 GFP Transduction Using Adenoviral Vector in Normal Oral Epithelium Seong-Gon Kim, DDS, PhD, Dept. of OMFS, Sacred Heart Hospital, Hallym University, #896 Pyungchon- Dong, Dongan-Gu, Anyang, 431-070, Korea (Kim SG; Song KH; Lee SK; Nah HD) Statement of the Problem: The aim of this study was to determine the conditions suitable for viral infections in the normal oral mucosa and the possibility of topical gene therapy to the oral mucosa using a viral vector. Materials and Methods: Freshly taken fragments of the palate and tongue from a mouse were used for the organ culture. The specimens were exposed to the green flu- orescent protein (GFP)-adenoviral vector for 1 hour. The control was not exposed to the virus. The initial viral titer was 6.3x1011 pfu/ml and the virus was used after dilution. The dilution ratio was 1: 1,000, 1:10,000, and 1: 100,000. The samples were cultured on a stainless steel wire mesh in an organ culture dish. A stereoscopic examination was performed every 24 hours for 6 days. The specimens were then fixed and examined immuno- histochemically. Results: There was no visible expression in the con- trol, and the cells exposed to 6.3x106 pfu/ml, and 6.3x107pfu/ml. The initial expression was observed at 24 hours after infection in both the palate and tongue in the cells exposed to 6.3x108pfu/ml, and the expression level increased until 3 days after the infection. In both cell types, the expression was mainly observed at the resection margin. The immunohistochemical study re- vealed that the epithelial cells were positive to GFP. Conclusion: A topically applied adenovirus containing specific genetic information of GFP can transduce the GFP into the normal oral epithelial cells of a resection margin in an organ culture provided a sufficient concen- tration and exposure time are used. POSTER 021 Expression of CfxA2 Beta-Lactamase Gene of Obligate Anaerobic Gram- Negative Rods in Oral Cavity Rie Iwai, DDS, Japan (Kinoshita S; Matsumoto K; Tabushi M; Motita S) Statement of the Problem: Obligate anaerobic Gram- negative rods were dominantly isolated from various of odontogenic infectious diseases. Some antimictobial agents were usually used for treatment of the disease, but be-ta-lactams-resistant bacteria were found during that treatment. Be-ta-lactam antibiotics resistance im- pedes the result of medication for the disease. The pur- pose of this study is to elucidate the transformation and expression of the gene extracted from beta-lactamase producing obligate anaerobic Gram-negative bacteria in the oral cavity by inducing the gene into Escherichia coli. Materials and Methods: Six strains of obligate anaero- bic Gram-negative rods were isolated from the oral cav- ity. The DNA of each strain was extracted and examined with PCR using 9 primers (EcToho, SfTEM, KpCAZ, MmTEM, CfxA2, KoAmpCA, KoAmpCB, EcGc, AbOXA21). The PCR products were introduced into Escherichia coli by TA cloning technique, and cultured at selected plate contained ampicillin. Beta-lactamase productivity of transconjugants were investigated nitro- cefin. Method of Data Analysis: PCR products differented a few sequence respectively between 922 – 944 bp. After incubation of 48 hours, 3 transconjugants grew. So the beta-lactam resistance was expressioned in 3 strains (us- ing CfxA2 primer). But another strains did not grow. Results: The above was an obvious fact that the trans- formation gene was beta-lactamase gene. Scientific Poster Session AAOMS • 2007 43.e11
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Transcript of Poster 020: GFP Transduction Using Adenoviral Vector in Normal Oral Epithelium
females) who visited the Department of Oral Medicine ofKagoshima University Hospital were diagnosed to havecandida glostitis base on culture methods. We utilizedthe USB-microscope M2 (Scalar) to thus examine can-dida glostitis which were suspected and at the same timediagnosed by conventional culture methods to detectCandida species. We applied gentian violet (0.1% meth-ylrosaniline chloride solution) on the surface of thetongue and then washed it off using physiological salineand then observed the papillae of the tongue by theUSB-microscope M2.
Method of Data Analysis: We confirmed the presenceof Candida species where the stained area by culturemethod.
Results: Eighteen cases were thus detected to haveCandida species by based on the findings obtained usingthe USB-microscope M2 with gentian violet staining. Inall cases some parts of the lingual papillae were stainedblue by gentian violet. Candida species were culturedfrom this part which was stained by gentian violet andpseudohyphae were observed
Conclusion: The detection of oral Candida speciesusing microscopic evaluations may therefore be a usefulnew method.
References
Tam M.T., et al. Gram stain method shows better sensitivity thanclinical criteria for detection of bacterial vaginosis in survellance ofpregnant, low-income women in a clinical setting. Infection Disease inobstetric Gynecology; 6:204-208. 1998
Chaiban G., et al. A rapid method of impregnating endtrocheal tubesand urinary catheters with gendine: a novel antiseptic agent. J. ofAntimicrobial Chemoteraphy; 55: 51-56. 2005
POSTER 020GFP Transduction Using AdenoviralVector in Normal Oral EpitheliumSeong-Gon Kim, DDS, PhD, Dept. of OMFS, SacredHeart Hospital, Hallym University, #896 Pyungchon-Dong, Dongan-Gu, Anyang, 431-070, Korea (Kim SG;Song KH; Lee SK; Nah HD)
Statement of the Problem: The aim of this study was todetermine the conditions suitable for viral infections inthe normal oral mucosa and the possibility of topicalgene therapy to the oral mucosa using a viral vector.
Materials and Methods: Freshly taken fragments of thepalate and tongue from a mouse were used for the organculture. The specimens were exposed to the green flu-orescent protein (GFP)-adenoviral vector for 1 hour. Thecontrol was not exposed to the virus. The initial viraltiter was 6.3x1011 pfu/ml and the virus was used afterdilution. The dilution ratio was 1: 1,000, 1:10,000, and 1:100,000. The samples were cultured on a stainless steelwire mesh in an organ culture dish. A stereoscopic
examination was performed every 24 hours for 6 days.The specimens were then fixed and examined immuno-histochemically.
Results: There was no visible expression in the con-trol, and the cells exposed to 6.3x106 pfu/ml, and6.3x107pfu/ml. The initial expression was observed at24 hours after infection in both the palate and tongue inthe cells exposed to 6.3x108pfu/ml, and the expressionlevel increased until 3 days after the infection. In bothcell types, the expression was mainly observed at theresection margin. The immunohistochemical study re-vealed that the epithelial cells were positive to GFP.
Conclusion: A topically applied adenovirus containingspecific genetic information of GFP can transduce theGFP into the normal oral epithelial cells of a resectionmargin in an organ culture provided a sufficient concen-tration and exposure time are used.
POSTER 021Expression of CfxA2 Beta-LactamaseGene of Obligate Anaerobic Gram-Negative Rods in Oral CavityRie Iwai, DDS, Japan (Kinoshita S; Matsumoto K;Tabushi M; Motita S)
Statement of the Problem: Obligate anaerobic Gram-negative rods were dominantly isolated from various ofodontogenic infectious diseases. Some antimictobialagents were usually used for treatment of the disease,but be-ta-lactams-resistant bacteria were found duringthat treatment. Be-ta-lactam antibiotics resistance im-pedes the result of medication for the disease. The pur-pose of this study is to elucidate the transformation andexpression of the gene extracted from beta-lactamaseproducing obligate anaerobic Gram-negative bacteria inthe oral cavity by inducing the gene into Escherichiacoli.
Materials and Methods: Six strains of obligate anaero-bic Gram-negative rods were isolated from the oral cav-ity. The DNA of each strain was extracted and examinedwith PCR using 9 primers (EcToho, SfTEM, KpCAZ,MmTEM, CfxA2, KoAmpCA, KoAmpCB, EcGc,AbOXA21). The PCR products were introduced intoEscherichia coli by TA cloning technique, and culturedat selected plate contained ampicillin. Beta-lactamaseproductivity of transconjugants were investigated nitro-cefin.
Method of Data Analysis: PCR products differented afew sequence respectively between 922 – 944 bp. Afterincubation of 48 hours, 3 transconjugants grew. So thebeta-lactam resistance was expressioned in 3 strains (us-ing CfxA2 primer). But another strains did not grow.
Results: The above was an obvious fact that the trans-formation gene was beta-lactamase gene.
Scientific Poster Session
AAOMS • 2007 43.e11
