Identification and validation of reference genes for reliable analysis of differential gene expression during antibiotic induced persister formation in Klebsiella pneumoniae using qPCR.

The study of differential gene expression in persister cells is compounded by ceasure of conventional cellular metabolic pathways during persistence. There is, hence, a requirement to identify and validate suitable reference genes whose expression remains stable during persistence. We evaluated the suitability of five genes viz. dnaJ, groEL, rpoB, kp751, kp4432 as references to study gene expression using real-time polymerase chain reaction (qPCR) during persister cell formation in Klebsiella pneumoniae. Results obtained showed that while dnaJ and groEL suffered from unstable expression; rpoB, kp751 and kp4432 showed stable expression. Further, it was observed that data normalization using either of the stable genes viz. rpoB, kp751, kp4432 alone, resulted in either too low expression levels or too high variation among replicates. Our study indicates the concurrent use of kp4432 and rpoB as reference genes to be the most suitable for reliable analysis of differential gene expression during antibiotic induced persister formation in K. pneumoniae. kp4432 and rpoB encode NAD-dependant phenylacetaldehyde dehydrogenase and DNA-directed RNA polymerase beta subunit respectively. The outcome of this study will increase the utility of qPCR in studying the temporal changes in gene expression during persistence. The study will also aid in understanding mechanisms underlying persister cell formation particularly in K. pneumoniae.

Tooth loss and dementia: An oro-neural connection. A cross-sectional study.

BACKGROUND: Recent cross-sectional studies have suggested that chronic inflammation resulting due to gingival and periodontal diseases may have potential reversible risk factors for dementia. Recently, cognition, memory deficit, and along with the behavioral defect have been considered as characteristics of dementia. Age is one of the main factors which is also known to be associated with dementia. The aim of the present study was to establish the correlation between tooth loss and diagnosis of cognitive impairment/dementia in elderly patients. MATERIALS AND METHODS: A cross-sectional study, comprising of 300 patients (Age: 50-80 years), was conducted. For the assessment of cognitive impairment/dementia, Mini-Mental Status Examination (MMSE) was conducted. All the results were analyzed using the Chi-square and Kruskal-Wallis ANOVA test. RESULTS: Number of teeth was found to be directly associated with MMSE. Less number of teeth was associated with lower MMSE score, irrespective of gender. The level of statistical significance was found to be P < 0.001. Factors, such as socioeconomic status, school education, and marital status, were found to have a statistically significant impact on the dementia scale. CONCLUSION: The current study found a significant association between tooth loss and cognitive impairment. Along with age factors such as socioeconomic status, education, and marital status were also found to have a direct or indirect impact on dementia. More prospective studies and clinical trials are required to be conducted to confirm the observations of this study.