Enrichment of polycyclic aromatic hydrocarbon metabolizing microorganisms on the oral mucosa of tobacco users.

Certain soil microbes resist and metabolize polycyclic aromatic hydrocarbons (PAHs). The same is true for a subset of skin microbes. In the human mouth, oral microbes have the potential to oxidize tobacco PAHs, thereby increasing these chemicals' ability to cause cancer of adjacent epithelium. We hypothesized that we could identify, in smokers, the oral mucosal microbes that can metabolize PAH. We isolated bacteria and fungi that survived long-term in minimal media with PAHs as the sole carbon source, under aerobic conditions, from the oral mucosa in 17 of 26 smokers and two of 14 nonsmokers. Of bacteria genera that survived harsh PAH exposure in vitro, most were found at trace levels, except for Staphylococcus, Actinomyces, and Kingella, which were more abundant. Two PAH-resistant strains of Candida albicans (C. albicans) were isolated from smokers. C. albicans was a prime candidate to contribute to carcinogenesis in tobacco users as it is found orally at high levels in tobacco users on the mucosa, and some Candida species can metabolize PAHs. However, when C. albicans isolates were tested for metabolism of two model PAH substrates, pyrene and phenanthrene, they were not capable, suggesting they cannot metabolize PAH under the conditions used. In conclusion, evidence for large scale microbial degradation of tobacco PAHs under aerobic conditions on the oral mucosa remains lacking, though nonabundant PAH metabolizers are certainly present.

Streptococcus endopeptidases promote HPV infection in vitro.

Both cervical and throat cancers are associated with human papillomavirus (HPV). HPV infection requires cleavage of the minor capsid protein L2 by furin. While furin is present in the vaginal epithelium, it is absent in oral epithelial basal cells where HPV infection occurs. The objective of this study was to investigate whether common oral bacteria express furin-like peptidases. By screening strains representing 12 oral Streptococcus and Enterococcus species, we identified that eight Streptococcus strains displayed high levels of furin-like peptidase activity, with S. gordonii V2016 the highest. We constructed null mutations for 14 genes encoding putative endopeptidases in S. gordonii V2016. Results showed that three endopeptidases, PepO, PulO, and SepM, had furin-like activities. All three mutants showed decreased natural transformation by chromosomal DNA, while the pepO mutant also showed reduced transformation by plasmid DNA, indicating involvement of these endopeptidases in competence development. The purified S. gordonii PepO protein promoted infection of epithelial 293TT cells in vitro by HPV16 pseudovirus. In conclusion, oral bacteria might promote HPV infection and contribute to HPV tissue tropism and subsequent carcinogenesis in the oral cavity and throat by providing furin-like endopeptidases.

Alcohol metabolism by oral streptococci and interaction with human papillomavirus leads to malignant transformation of oral keratinocytes.

Poor oral hygiene, ethanol consumption, and human papillomavirus (HPV) are associated with oral and esophageal cancers. However, the mechanism is not fully known. This study examines alcohol metabolism in Streptococcus and its interaction with HPV-16 in the malignant transformation of oral keratinocytes. The acetaldehyde-producing strain Streptococcus gordonii V2016 was analyzed for adh genes and activities of alcohol and aldehyde dehydrogenases. Streptococcus attachment to immortalized HPV-16 infected human oral keratinocytes, HOK (HPV/HOK-16B), human oral buccal keratinocytes, and foreskin keratinocytes was studied. Acetaldehyde, malondialdehyde, DNA damage, and abnormal proliferation among keratinocytes were also quantified. We found that S. gordonii V2016 expressed three primary alcohol dehydrogenases, AdhA, AdhB, and AdhE, which all oxidize ethanol to acetaldehyde, but their preferred substrates were 1-propanol, 1-butanol, and ethanol, respectively. S. gordonii V2016 did not show a detectable aldehyde dehydrogenase. AdhE is the major alcohol dehydrogenase in S. gordonii. Acetaldehyde and malondialdehyde production from permissible Streptococcus species significantly increased the bacterial attachment to keratinocytes, which was associated with an enhanced expression of furin to facilitate HPV infection and several malignant phenotypes including acetaldehyde adduct formation, abnormal proliferation, and enhanced migration through integrin-coated basement membrane by HPV-infected oral keratinocytes. Therefore, expression of multiple alcohol dehydrogenases with no functional aldehyde dehydrogenase contributes to excessive production of acetaldehyde from ethanol by oral streptococci. Oral Streptococcus species and HPV may cooperate to transform oral keratinocytes after ethanol exposure. These results suggest a significant clinical interaction, but further validation is warranted.

Multiple alcohol dehydrogenases but no functional acetaldehyde dehydrogenase causing excessive acetaldehyde production from ethanol by oral streptococci.

Ethanol consumption and poor oral hygiene are risk factors for oral and oesophageal cancers. Although oral streptococci have been found to produce excessive acetaldehyde from ethanol, little is known about the mechanism by which this carcinogen is produced. By screening 52 strains of diverse oral streptococcal species, we identified Streptococcus gordonii V2016 that produced the most acetaldehyde from ethanol. We then constructed gene deletion mutants in this strain and analysed them for alcohol and acetaldehyde dehydrogenases by zymograms. The results showed that S. gordonii V2016 expressed three primary alcohol dehydrogenases, AdhA, AdhB and AdhE, which all oxidize ethanol to acetaldehyde, but their preferred substrates were 1-propanol, 1-butanol and ethanol, respectively. Two additional dehydrogenases, S-AdhA and TdhA, were identified with specificities to the secondary alcohol 2-propanol and threonine, respectively, but not to ethanol. S. gordonii V2016 did not show a detectable acetaldehyde dehydrogenase even though its adhE gene encodes a putative bifunctional acetaldehyde/alcohol dehydrogenase. Mutants with adhE deletion showed greater tolerance to ethanol in comparison with the wild-type and mutant with adhA or adhB deletion, indicating that AdhE is the major alcohol dehydrogenase in S. gordonii. Analysis of 19 additional strains of S. gordonii, S. mitis, S. oralis, S. salivarius and S. sanguinis showed expressions of up to three alcohol dehydrogenases, but none showed detectable acetaldehyde dehydrogenase, except one strain that showed a novel ALDH. Therefore, expression of multiple alcohol dehydrogenases but no functional acetaldehyde dehydrogenase may contribute to excessive production of acetaldehyde from ethanol by certain oral streptococci.

Streptococci-human papilloma virus interaction with ethanol exposure leads to keratinocyte damage.

PURPOSE: Ethanol, human papilloma virus (HPV), and poor oral hygiene are risk factors that have been attributed to oral carcinogenesis. Streptococci sp and HPV infections are common in the head and neck, often associated with sexual activity. Although HPV is linked to head and neck squamous cell carcinoma, it is unclear whether there is a similar role for Streptococci sp. This cell study examines whether Streptococci sp and HPV-16 with exposure to ethyl alcohol (ETOH) can act as cofactors in the malignant transformation of oral keratinocytes. MATERIALS AND METHODS: ETOH (0.1%-20% vol/vol) was used to investigate Streptococci sp attachment with immortalized E6-expressing HPV/HOK-16B cells, human oral buccal keratinocytes, and foreskin keratinocytes. Streptococci sp (Streptococci mutans [LT11]) and various strains of acetaldehyde (AA) producer and nonproducer Streptococcus salivarius (110-1, 109-2, 101-7, and 107-1) and a lactic acid producer bacterium, Lactobacillus rhamnosus (24-1 and 25-2), were examined for interactions with keratinocytes by use of a green dye (percent of cells with colonies after 24 hours). Carcinogens, AA, malondialdehyde, DNA damage, and proliferation (5'-bromo-2-deoxyuridine) among keratinocytes were also quantified. RESULTS: AA and malondialdehyde production from permissible Streptococci sp significantly increased with attachment to keratinocytes, whereas L rhamnosus did not significantly attach to keratinocytes. This attachment was associated with enhanced levels of AA adduct formation, proliferation (5'-bromo-2-deoxyuridine incorporation), and enhanced migration through integrin-coated basement membrane by HPV oral keratinocytes, which are characteristics of a malignant phenotype. CONCLUSIONS: These cell studies suggest that oral Streptococci sp and HPV (HPV-16) cooperate to transform oral keratinocytes after low-level ETOH (1%) exposure. These results appear to suggest a significant clinical interaction, but further validation is warranted.

Gastroesophageal reflux disease (GERD): is there more to the story?

Gastroesophageal reflux disease (GERD) affects both men and women worldwide, with the most common symptom of GERD being frequent heartburn. If left untreated, more serious diseases including esophagitis and/or esophageal cancer may result. GERD has been commonly held to be the result of gastric acid refluxing into the esophagus. Recent work, however, has shown that there are acid-producing cells in the upper aerodigestive tract. In addition, acid-producing bacteria located within the upper gastrointestinal tract and oral cavity may also be a contributing factor in the onset of GERD. Proton pump inhibitors (PPIs) are commonly prescribed for treating GERD; these drugs are designed to stop the production of gastric acid by shutting down the H(+)/K(+)-ATPase enzyme located in parietal cells. PPI treatment is systemic and therefore significantly different than traditional antacids. Although a popular treatment choice, PPIs exhibit substantial interpatient variability and commonly fail to provide a complete cure to the disease. Recent studies have shown that H(+)/K(+)-ATPases are expressed in tissues outside the stomach, and the effects of PPIs in these nongastric tissues have not been fully explored. Likewise, acid-producing bacteria containing proton pumps are present in both the oral cavity and esophagus, and PPI use may also adversely affect these bacteria. The use of PPI therapy is further complicated by the two philosophical approaches to treating this disease: to treat only symptoms or to treat continuously. The latter approach frequently results in unwanted side effects which may be due to the PPIs acting on nongastric tissues or the microbes which colonize the upper aerodigestive tract.