Mutation studies of the gene encoding YuiC, a stationary phase survival protein in Bacillus subtilis

Aims@#YuiC is a stationary phase survival (Sps) protein from the Firmicute Bacillus subtilis that possesses muralytic activity to cleave bacterial cell-wall peptidoglycan. It has a small lytic transglycosylase (MltA) fold analogous to the resuscitation promoting factors (Rpfs) of Actinobacteria which have a hybrid of a mini lysozyme and soluble lytic transglycosylase (Slt35/70) fold. The present study aimed at identifying key residues of YuiC/Sps that are catalytically active and studying the effect of B. subtilis cell growth upon sps/yuiC deletion. @*Methodology and results@#Four forms of mutated yuiC were created through Site-directed, Ligase-Independent Mutagenesis Polymerase Chain Reaction (SLIM PCR) that include the substitutions of D129A, D151A, D162A and K102A. These individual mutated yuiC genes were cloned and expressed in the Escherichia coli BL21 (DE3) expression system and subsequently purified to homogeneity using affinity, cation exchange and size exclusion chromatography. The D129A variant was shown to be insoluble, indicating its role in maintaining the right protein folding of YuiC. The remaining three variants resulted in soluble proteins but were inactive on zymograms indicating that they may be responsible for catalysis. B. subtilis cells harbouring individual sps genes (yuiC, yabE, yocH and yorM) knocked out showed stationary phase defects and altered colony morphologies compared to the wild type. @*Conclusion, significance and impact of study@#This study has identified the key residues involved in catalysis of YuiC, which are the D151, D162 and K102. These are conserved in Sps domains. The catalytic mechanism of YuiC is similar to the mechanism reported for Neisseria gonorrhoeae MltA. sps/yuiC knock outs have implied that each sps/yuiC has a significant role on B. subtilis late growth stage. The B. subtilis YuiC/Sps model has given an insight into Sps functions in the final growth stage of the Firmicutes, which members include etiologic agents of anthrax, botulism and listeriosis. Inhibition of Sps protein may inactivate pathogen replication and facilitate entrance into a non-contagious dormant sporulation stage.

Blood glucose level, salivary PH and oral bacterial count in type 1 diabetic children

Laboratory tests and animal studies have shown a relationship between dental caries and high blood glucose level because of reduction in salivary pH level and the subsequent increase in salivary microbial count. If the acidic environment is left unregulated, it would eventually result in greatly disrupted oral microbial and mineral balance resulting in dental caries and subsequent tooth cavitations. Most bacteria have an optimum pH for growth in the range 6.5 - 7.5 with limits somewhere between 5 and 9. Acidophilic bacteria can grow at a low pH, and such organisms are very important in oral microbiology as the causative agents of caries: Lactobacilli and Streptococcus mutans produce acid as end products of metabolism of dietary sugars, are able to survive and grow in acidic conditions and have been found to have tendency toward higher count in diabetic patients. Subjects with juvenile diabetes are more likely to have higher oral bacterial count as compared to those suffering from adult-onset diabetes mellitus, which accounts for more risk in the progression of oral complications in these patients. No studies have so far been conducted in Pakistan that compares the prevalence of dental caries in diabetic children. This review therefore signifies the scope of such analysis especially in relation to the salivary pH level and salivary bacterial count. Hence, prophylaxis and treatment can be targeted more efficiently to the at risk subjects

Periodontal status of women during pregnancy

Objectives: Periodontal condition of women is considered to be debilitated during pregnancy. This is suggested to increase the chances of complications during pregnancy. This study aimed at obtaining information regarding periodontal status during pregnancy, which may be necessary for planning periodontal disease preventive programs for pregnant women in our population

Material and Methods: A cross-sectional study was conducted on a sample of 322 registered women at department of Gynecology and Obstetric. Periodontal condition was assessed by CPITN Index and recorded in a semi-structured performa. The data was coded and entered in SPSS version 11.0 and percentage frequencies with their mean and standard deviations were calculated. Exact Chi-sqaure test was used to analyze any significant changes observed in the CPITN scores amongst the study population, where level of significance was p < 0.05 with confidence interval 95%

Results: Only 17% of these women were observed to have healthy periodontal status and 83% of women had some signs involving a mean of 3 out of six sextants with different levels of signs of periodontal disease, with only less than half sextant with periodontal pockets. However, a moderate level of periodontal debilitation was observed in this population with a mean of 3 sextants having healthy periodontal status. The present study observed a constant level of periodontal status assessed by CPITN Index (p>0.05) in women during pregnancy

Conclusion: Although CPITN scores remained constant through out pregnancy and no significant change observed in this moderate level of diseased condition. However, shallow and deep periodontal pockets, although not very severe, are observed to be more prevalent. Therefore, special preventive and prophylaxis oral health care programs remain the need for these women during pregnancy

Oral health status in diabetic children

Objectives: diabetes causes numerous oral and salivary changes leading to cariogenic and gingival lesions. The present study was designed to elucidate the role of diabetes mellitus in oral health

Methods: a cross-sectional study including 60 diabetic children [case group] and 30 non diabetic children [control group] of age 3-14 years was conducted. HbA1c and blood glucose level was measured along with the oral health including gingival status and dental caries status was visually assessed. Gingival status was coded for healthy, marginal gingivitis and calculus. Dental caries status [decayed and filled] for both deciduous and permanent dentition was assessed. Data was recorded in a pre-coded oral health proforma, which was then entered and analyzed in SPSS version 10.0. Descriptive analysis such as percentage frequencies and means was performed. Exact Chi-square test was used to analyze any significant changes observed amongst the study population, where level of significance was p < 0.05 with confidence interval 95%

Results: the results show important difference between both groups of children. Dental caries level is significantly higher in diabetic children both in deciduous and permanent dentition than in non-diabetic children [p <0.05]. Gingival health was also observed to be debilitated in diabetic children than non- diabetic children [p < 0.05]

Conclusion: the study highlights that special preventive measures must be adopted to maintain a good oral health of the diabetic children

Periodontal infection: a potential risk factor for p re-term delivery of Low birth weight [PLBW] babies

Pre-term delivery of low-birth-weight [PLBW] babies is considered a major peri-natal problem in many countries and is contributing substantially to infant mortality and to childhood handicap. There is a reported incidence of pre-term delivery of low-birth-weight [PLBW] babies of 37% of all live births in Pakistan, which has a tremendous impact on health care system in this community. The prevalence of periodontal disease in Pakistan is also very high in all age groups and women of child bearing age [18-34 years] are no exception. Recent studies indicate periodontal infection as a potential independent risk factor for PLBW, and is considered to be 7 times more likely to be associated than any other risk factors. Several postulated mechanisms have been reviewed, including the virulence effects and role of asymptomatic bacteraemia, focusing on the bacterial load in periodontium facilitating its transmission from oral cavity to the uterus. The indication that periodontal disease is a potential risk factor for the delivery of PLBW; a high level of periodontal disease in women of child bearing age and similar high level of PLBW babies in country, calls for further longitudinal investigations that validate a causal relationship between periodontal infection and pre-term delivery of LBW babies in Pakistan. A review of literature and preliminary communication for a planned study is presented

Periodontal infection: a potential risk factor for pre-term delivery of low birth weight [PLBW] babies

Pre-term delivery of low-birth-weight [PLBW] babies is considered a major peri-natal problem in many countries and is contributing substantially to infant mortality and to childhood handicap. There is a reported incidence of pre-term delivery of low-birth-weight [PLBW] babies of 37% of all live births in Pakistan, which has a tremendous impact on health care system in this community. The prevalence of periodontal disease in Pakistan is also very high in all age groups and women of child bearing age [18-34 years] are no exception.Recent studies indicate periodontal infection as a potential independent risk factor for PLBW, and is considered to be 7 times more likely to be associated than any other risk factors. Several postulated mechanisms have been reviewed, including the virulence effects and role of asymptomatic bacteraemia, focusing on the bacterial load in periodontium facilitating its transmission from oral cavity to the uterus.The indication that periodontal disease is a potential risk factor for the delivery of PLBW; a high level of periodontal disease in women of child bearing age and similar high level of PLBW babies in country, calls for further longitudinal investigations that validate a causal relationship between periodontal infection and pre-term delivery of LBW babies in Pakistan. A review of literature and preliminary communication for a planned study is presented