Smear layer removal ability and antibacterial activity of endodontic irrigants

INTRODUCTION : a variety of endodontic irrigants are available for endodontic irrigation. Irrigants must be effective in removing the smear layer created during endodontic therapy as well as eliminating bacteria. AIM : This in vitro study tested various alternating sequences of sodium hypochlorite, anolyte solution (electrochemically activated water), and EDTA for their ability to do this. METHOD : Forty-eight single canal teeth were randomly divided into six groups, prepared to working length, sterilised and inoculated with Enterococcus faecalis. Each group was assigned a different sequence of irrigants. Standard cultivation techniques were used to count the colonyforming units at each phase. Two SEM photomicrographs of each root’s coronal, middle and apical thirds were taken randomly and the number of patent dentinal tubules counted. Statistical analysis was completed using Oneway- ANOVA and multiple comparisons. RESULTS : Group 6 (n=10) protocol of 5ml anolyte followed by 3ml 18% EDTA showed the best smear layer removal results for all thirds of the canal. Chemical irrigation significantly decreased the intracanal E. faecalis CFUs. CONCLUSION : Within the limitations of the study anolyte solution followed by EDTA showed the best smear layer removal. The various sequences of NaOCl, anolyte solution, and EDTA all had similar antibacterial results.This study was partially funded with a grant from the Faculty Research Committee of the Faculty of Health Sciences, University of the Witwatersrand.http://www.sada.co.zaam2016Paraclinical Science

A comparison of the efficacy of various disinfection protocols in endodontic treatment : an in vitro study

INTRODUCTION : Disinfection is the main objective of root canal preparation and whilst irrigation is most commonly used, another method is Photo-activated Disinfection (PAD). AIMS AND OBJECTIVES : The aim of this in vitro study was to compare the efficacy of eradication of bacteria from root canals by six different disinfection protocols. METHODS : Root canals of 84 extracted human teeth were prepared to a standardised size and taper. The teeth were sterilised and then inoculated with E. faecalis. The teeth were randomly assigned to one of seven groups (n = 12), each of which underwent a different disinfection protocol. Dentine samples were plated onto BHI plates and incubated anaerobically. After five days, colony-forming units (cfu) were counted. The Pairwise Wilcoxon Rank Sum test and the Kruskal-Wallis test were used for statistical analysis of the data. RESULTS : The most effective disinfection protocol was: 3% NaOCl with 2% CHX, followed by 3% NaOCl with PAD, Chlor-XTRA and 2% CHX. The 3% NaOCl-protocol performed significantly better than PAD and distilled water. The PAD-protocol performed significantly better than distilled water. CONCLUSION : The most efficient protocol in eradicating E. faecalis from the root canals was 3% NaOCl followed by irrigation with 2% CHX.The Professor Cornelis H Pameijer Fellowshiphttp://www.sada.co.zaam201

Bacterial species from retailed poultry eggs in Tshwane, South Africa : implication for consumers

Food safety is an important public health issue and governments across the world are intensifying their efforts to improve the quantity, quality and the safety of national food supplies. Bacteria, especially Salmonella species, present in or on chicken meat and hens’ eggs in particular are the most common causes of food poisoning and the major sources of human salmonellosis. Literature reveals little information on the risk factors for salmonellae infection in Africa. The aim of this study was to determine which, if any, bacteria, especially Salmonella species, are present in and on hens’ eggs. Representative bacterial colonies were confirmed with Gram staining and then identified using the MALDI-TOF Biotyper assay. The genera identified were Escherichia coli (34%), Enterococcus faecalis (14%), Proteus mirabilis (9%), Klebshiella pneumoniae (7%), Salmonella Typhimurium (6%), Enterobacter cloacae (1%), Stenotrophomonas maltophilia (0.6%), Salmonella Dublin (0.6%) and Salmonella Braenderup (0.2%). Raw hens’ eggs and products containing raw hens’ eggs may contain pathogenic bacteria, thereby exposing a large number of consumers to the risk of contracting food poisoning when undercooked or uncooked hens’ eggs are consumed. SIGNIFICANCE : • Enterobacteriaceae counts are used as an indicator to evaluate the hygienic quality of food. • The presence of Salmonella species and other Enterobacteriaceae in raw hens’ eggs poses a health risk to consumers. • Any product in which raw eggs are used must be provided with a conspicuous label stating that it may contain pathogenic bacteria.Funding of the project was through the research funds of F.S.B. and E.M.B.http://www.sajs.co.zaam2018Food ScienceParaclinical Science

Sub-MICs of Carum copticum and Thymus vulgaris influence virulence factors and biofilm formation in Candida spp

BACKGROUND: Emergence of drug-resistant strains of Candida and inefficiency of conventional antifungal therapy has necessitated the search for alternative and new antifungal agents. Inhibition of virulence and biofilm are the potential drug targets. In this study, the oils of Carum copticum, Thymus vulgaris and their major active compound thymol as revealed by Gas chromatography and gas chromatography–mass spectrometry (GC-GC/MS) analysis were tested for their inhibitory activity against growth to determine sub-MIC values against 27 drug-resistant strains of Candida spp. METHODS: Brothmacrodilution method was used for determination of MIC of test oils against Candida strains. The spectrophotometric methods were used for detection and inhibition assays for virulence factors in Candida spp. Light and electron microscopy was performed to observe morphological effects of oils on biofilms. GC-GC/MS were used to evaluate the major active compounds of test oils. RESULTS: Virulence factors like proteinase and haemolysin were detected in 18 strains, both in solid and liquid media. A 70% of the test strains exhibited hydrophobicity and formed moderate to strong biofilms (OD280 0.5- > 1.0). Test oils exhibited MICs in the range of 45–360 μg.mL−1 against the majority of test strains. All the oils at 0.25× and 0.5× MICs induced >70% reduction in the cell surface hydrophobicity, proteinase and haemolysin production. At 0.5× MIC, thymol and T. vulgaris were most inhibitory against biofilm formation. At sub-MICs electron microscopic studies revealed the deformity of complex structures of biofilms formed and cell membranes appeared to be the target site of these agents. CONCLUSIONS: Therefore, our findings have highlighted the concentration dependent activity of oils of C. copticum and T. vulgaris against virulence factors and biofilms in proteinase and haemolysin producing drug-resistant strains of Candida spp. The above activities of test oils are supposed to be mainly contributed due to their major active compound thymol. Further mechanism involving anti-proteinase, anti-haemolysin and anti-biofilm activities of these oils and compounds are to be explored for possible exploitation in combating Candida infections.The Indian Council of Medical Research, New Delhi for financial support in the form of SRF to M S A Khan.http://www.biomedcentral.com/bmccomplementalternmedam201

The antibacterial activity of extracts of nine plant species with good activity against Escherichia coli against five other bacteria and cytotoxicity of extracts

Background: The development of antibiotic resistant bacteria stems from a number of factors, including inappropriate use of antibiotics in human and animal health and their prolonged use as growth promoters at sub-clinical doses in poultry and livestock production. We were interested in investigating plants that could be useful in protecting humans or animals against diarrhoea. We decided to work on extracts of nine plant species with good activity against Escherichia coli based on earlier work in the Phytomedicine Programme. Leaves of nine medicinal plant species with high antibacterial activity against Escherichia coli were extracted with acetone and their minimal inhibitory concentration (MIC) values determined using a microplate serial dilution technique against Gram-positive (Staphylococcus aureus, Enterococcus faecalis and Bacillus cereus) and Gram-negative (Escherichia coli, Salmonella Typhimurium and Pseudomonas aeruginosa) bacteria. Bioautography was used to determine the number of bioactive compounds in each extract. In vitro safety of the extracts was determined using the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide reduction assay on Vero cells. Results: The extracts were active against all the pathogens with average MICs ranging from 0.02 to 0.52 mg/ml. As expected E. coli was relatively sensitive, while E. faecalis and S. Typhimurium were more resistant to the extracts (average MICs of 0.28 mg/ml and 0.22 mg/ml respectively). Cremaspora triflora and Maesa lanceolata leaf extracts had higher activity than the other extracts against Gram-positive and Gram-negative pathogens with mean MICs of 0.07 mg/ml and 0.09 mg/ml respectively. Extracts of Maesa lanceolata and Hypericum roeperianum had the highest total antibacterial activity (TAA) at 1417 and 963 ml/g respectively. All extracts with the exception of that of Maesa lanceolata, Elaeodendron croceum and Calpurnia aurea had relatively low cytotoxicity with LC50 >20 μg/ml. Cremaspora triflora had the best selectivity index (SI) against S. aureus and E. coli of 2.87 and 1.15 respectively. Hypericum roeperianum had a SI of 1.10 against B. cereus. Bioautography revealed 1–6 visible antimicrobial compounds that were generally non-polar. Conclusions: There was a weak positive, but statistically non-significant correlation between the potency of the extracts and their cytotoxicity (R= 0.45, ρ > 0.05). The activity of the extracts on the test bacteria was in some cases not correlated with cytotoxicity, as shown by selectivity indices >1. This means that cellular toxicity was probably not due to compounds with antibacterial activity. Some of the extracts had a good potential for therapeutic use against the bacterial pathogens or for application in treating diarhoea. It does not appear that activity against E. coli is a good predictor of activity against Gram-negative rather than Gram-positive bacteria. Further investigation is in progress on C. triflora and H. roeperianum, both of which had promising activities and potential safety based on cytotoxicity.The National Research Foundation of South Africa (Eloff IPPR 953991), the Medical Research Council of South Africa and the University of Pretoria.https://bmccomplementalternmed.biomedcentral.com/am2017Paraclinical Science

Predicting the influence of multiple components on microbial inhibition using a logistic response model - a novel approach

BACKGROUND: There are several synergistic methods available. However, there is a vast discrepancy in the interpretation of the synergistic results. Also, these synergistic methods do not assess the influence the tested components (drugs, plant and natural extracts), have upon one another, when more than two components are combined. METHODS: A modified checkerboard method was used to evaluate the synergistic potential of Heteropyxis natalensis, Melaleuca alternifolia, Mentha piperita and the green tea extract known as TEAVIGO™. The synergistic combination was tested against the oral pathogens, Streptococcus mutans, Prevotella intermedia and Candida albicans. Inhibition data obtained from the checkerboard method, in the form of binary code, was used to compute a logistic response model with statistically significant results (p < 0.05). This information was used to construct a novel predictive inhibition model. RESULTS: Based on the predictive inhibition model for each microorganism, the oral pathogens tested were successfully inhibited (at 100% probability) with their respective synergistic combinations. The predictive inhibition model also provided information on the influence that different components have upon one another, and on the overall probability of inhibition. CONCLUSIONS: Using the logistic response model negates the need to ‘calculate’ synergism as the results are statistically significant. In successfully determining the influence multiple components have upon one another and their effect on microbial inhibition, a novel predictive model was established. This ability to screen multiple components may have far reaching effects in ethnopharmacology, agriculture and pharmaceuticals.The University of Pretoria holds a provisional South African patent (ZA2013/ 06534) relating to the content of the manuscript. No financial benefits have been received by the authors.The University of Pretoria and the Gen Foundation.http://www.biomedcentral.com/bmccomplementalternmedam201

The efficacy and safety of nine South African medicinal plants in controlling Bacillus anthracis Sterne vaccine strain

BACKGROUND : Anthrax is a zoonotic disease caused by Bacillus anthracis, a Gram-positive spore-forming bacterium. The presence of the bacteria and the toxins in the blood of infected hosts trigger a cascade of pathological events leading to death. Nine medicinal plants with good activities against other bacteria were selected to determine their in vitro antibacterial activity against Bacillus anthracis Sterne strain. The cytotoxicity of the extracts on Vero kidney cells was also determined. RESULTS : The minimum inhibitory concentration (MIC) values of the extracts against Bacillus anthracis Sterne strain ranged from 0.02 to 0.31 mg/ml. Excellent MIC values were observed for the following plant species: Maesa lanceolata (0.02 mg/ml), Bolusanthus speciosus, Hypericum roeperianum, Morus mesozygia (0.04 mg/ml) and Pittosporum viridiflorum (0.08 mg/ml). The total antibacterial activity of the extracts ranged from 92 to 5562 ml/g. Total activity presents the volume to which the extract from 1 g of plant material can be diluted and still inhibit microbial growth. Maesa lanceolata and Hypericum roeperianum had the highest total activity with values of 5562 and 2999 ml/g respectively. The extracts of Calpurnia aurea had the lowest total activity (92 ml/g). The cytotoxicity determined on Vero cells indicated that most of the extracts were relatively non-toxic compared to doxorubicin (LC50 8.3 ± 1.76 μg/ml), except for the extracts of Maesa lanceolata, Elaeodendron croceum and Calpurnia aurea with LC50 values at 2.38 ± 0.25, 5.20 ± 0.24 and 13 ± 2.26 μg/ml respectively. The selectivity index (SI) ranged from 0.02 to 1.66. Hypericum roeperianum had the best selectivity index, (SI = 1.66) and Elaeodendron croceum had lowest value (SI = 0.02). CONCLUSIONS : The crude acetone extracts of the selected plant species had promising antibacterial activity against Bacillus anthracis. Maesa lanceolata extracts could be useful as a disinfectant and Hypericum roeperianum could be useful to protect animals based on its high total activity and selectivity index. Further investigation of these plant extracts may lead to the development of new therapeutic agents to protect humans or animals against anthrax.The National Research Foundation of South Africa (Eloff 95991) and the University of Pretoria.http://www.biomedcentral.com/bmccom/plementalternmedam2016Paraclinical Science

Antimicrobial efficacy of nine different root canal irrigation solutions

Endodontic therapy is a debridement procedure that requires removal of the irritants from the canal and periapical tissues if the treatment is to be successful. It is well established that bacteria are the main etiological factors in the development of dentinal caries and its progression to pulpal and periapical disease. E. faecalis is the bacterial species most frequently recovered from root-filled teeth. Studies have shown that E. faecalis is able to withstand a high alkaline environment such as the one generated by calcium hydroxide. The resistance appears to be related to a cell proton-pump that is necessary for survival of the bacterium at high pH. Therefore, E. faecalis is able to form biofilms even in calcium-hydroxide-medicated root canals. In addition, under starved conditions, this resilient organism shows tolerance to sodium hypochlorite, heat, hydrogen peroxide, acid and ethanol. E. faecalis can also survive extended periods of starvation in water, within water-filled dentinal tubules, and in human serum.http://www.sada.co.zaam201

Anti-bacterial, free radical scavenging activity and cytotoxicity of acetone extracts of Grewia flava

BACKGROUND : Bacterial infections of the gastrointestinal tract (GIT) cause vomiting, diarrhoea and even systemic disease. There is a need for the development of natural products into alternative and safer medicines. OBJECTIVES : This study evaluated the anti-microbial activity of extracts prepared from berries, leaves,bark and roots of the edible plant Grewia flava. METHODS : The anti-bacterial activity was evaluated by the broth microdilution method. Anti-oxidant activity of the most active extracts was performed by 2, 2-diphenyl-1-picrylhydrazyl (DPPH) assay. The cytotoxicity of the extracts was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS : The acetone extracts of the leaves and roots showed the best activity with MIC values as low as 0.03 mg/mL against Staphylococcus aureus and Salmonella typhimurium and 0.07 mg/mL against Bacillus cereus, Escherichia coli and Staphylococcus aureus. Quantitative analysis of the scavenging ability showed that acetone extracts exhibited good free radical scavenging activity in a dose-dependent manner. The berries extract had the highest LC50 (lowest toxicity) of 551.68 68 μg/mL. CONCLUSION : Acetone extract of leaves and roots of Grewia flava contain anti-microbial and anti-oxidant compounds and could therefore be used as a natural product with little toxicity to host cells.https://www.ajol.info/index.php/ahshttp://www.bioline.org.br/hsam2017Community DentistryParaclinical Science

The anti-arthritic, anti-inflammatory, antioxidant activity and relationships with total phenolics and total flavonoids of nine South African plants used traditionally to treat arthritis

BACKGROUND : Oxidative stress predisposes the human and animal body to diseases like cancer, diabetes, arthritis, rheumatoid arthritis, atherosclerosis and chronic inflammatory disorders. Hence, this study seeks to determine the antioxidant, anti-inflammatory and anti-arthritic activities of acetone leaf extracts of nine South African medicinal plants that have been used traditionally to treat arthritis and inflammation. METHODS : The anti-inflammatory activity of the extracts was determined by investigating inhibition of nitric oxide production in lipopolysaccharide activated RAW 264.7 macrophages as well as 15-lipoxygenase enzyme inhibition. An anti-protein denaturation assay was used to determine the anti-arthritic properties of the extracts. The antioxidant activity was determined using the 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethyl-benzthiazoline-6-sulfonic acid) (ABTS) radical scavenging assays and ferric reducing antioxidant power (FRAP). The total phenolic and total flavonoid concentration of extracts were determined by using standard methods. RESULTS : All extracts inhibited nitric oxide production in a dose-dependent manner in the LPS-stimulated RAW 264.7 macrophages. Extracts of Maesa lanceolata and Heteromorpha arborescens inhibited NO production by 99.16 % and 89.48 % at a concentration of 30 μg/ml respectively. Elaeodendron croceum and Calpurnia aurea extracts had strong activity against 15-lipoxygenase activity with IC50 values of 26.23 and 34.70 μg/ml respectively. Morus mesozygia and Heteromorpha arborescens extracts had good in vitro anti-arthritic activity with IC50 values of 11.89 and 53.78 μg/ml, the positive control diclofenac sodium had IC50 value of 32.37 μg/ml. The free radical scavenging activity of the extracts in DPPH assays ranged between 7.72 and 154.77 μg/ml. Trolox equivalent antioxidant capacity (TEAC) and FRAP values ranged from 0.06 to 1.32 and 0.06 to 0.99 respectively. CONCLUSIONS : Results from this study support the traditional use of the selected medicinal plants in the management of arthritis and other inflammatory conditions. The free radical scavenging capacity of the extracts may be related to an immune boosting potential.ILE is a PhD student in the Phytomedicine Programme. JPD (PhD) is a post-doctoral fellow in the Phytomedicine Programme. FSB (PhD) is Senior Lecturer and mentor in the Phytomedicine Programme. LJM (PhD) is an Associate Professor and the current leader of the Phytomedicine Programme. JNE (DSc) is a Professor and the founder of the Phytomedicine Programme in the Department of Paraclinical Sciences of the Faculty of Veterinary Science, University of Pretoria.The National Research Foundation of South Africa (Eloff IPPR 953991) and the University of Pretoria.http://www.biomedcentral.com/bmccom/plementalternmedam2016Paraclinical Science