Test Bacterial Strains Research Articles

Antimicrobial properties and mechanism of volatile isoamyl acetate, a main flavour component of Japanese sake (Ginjo-shu).

To evaluate the antimicrobial properties of the main Ginjo-flavour components of sake, volatile isoamyl acetate and isoamyl alcohol. Volatile isoamyl acetate and isoamyl alcohol both inhibited growth of the five yeast and 10 bacterial test strains. The minimum inhibitory dose and minimum bactericidal (fungicidal) dose of isoamyl acetate were higher than those of isoamyl alcohol. Escherichia coli and Acetobacter aceti were markedly sensitive to isoamyl acetate and isoamyl alcohol. In E.coli exposed to isoamyl acetate for 5h, changes in expression were noted in proteins involved in sugar metabolism (MalE, MglB, TalB and PtsI), tricarboxylic acid cycle (AceA, Pfl and AcnB) and protein synthesis (EF-Tu, EF-G, and GlyS). Expression of acid and alcohol stress-response proteins was altered in E.coli exposed to isoamyl acetate. Esterase activity was detected in E.coli, suggesting that isoamyl acetate was hydrolyzed to acetic acid and isoamyl alcohol. Acetic acid and isoamyl alcohol damaged E.coli cell membranes and inactivated membrane proteins, impairing respiration. Volatile isoamyl acetate and isoamyl alcohol were effective in inactivating various micro-organisms, and antimicrobial mechanism of volatile isoamyl acetate against E.coli was clarified based on proteome analysis. To the best of our knowledge, this is the first report to examine the antimicrobial mechanism of volatile organic compound using proteome analysis combining two-dimensional difference gel electrophoresis with peptide mass fingerprinting.

Pulsed Light Decontamination of Endive Salad and Mung Bean Sprouts and Impact on Color and Respiration Activity

The objective of this study was the determination of the efficiency of pulsed light (PL) treatments for the decontamination of endive salad and mung bean sprouts, as well as the assessment of quality changes in relation to discoloration and alteration of respiration activity. Produce samples were artificially inoculated with two bacterial test strains Escherichia coli (DSM 498) and Listeria innocua (DSM 20649) and exposed to PL at different energy doses. The inactivation efficiency with regard to the naturally occurring microbiota was also investigated. Besides microbiological investigations, color changes were determined as well as the produce respiration during chilled storage. The results indicated that inactivation of more than 2 log was possible with one flash in the case of fresh-cut salad, while the reduction on mung bean sprouts was limited to approximately 1.6 log with one flash, irrespective if the natural flora or inoculated E. coli or L. innocua were considered. The UV part of the PL proved to be exclusively responsible for the inactivation of microorganisms. Significant lower levels of microbial counts of treated compared with untreated samples were maintained for up to 6 days. In the case of endive salad, a dose-dependent progressive discoloration and increase in respiration was diminished by applying optical bandpass filters, which only slightly affected the inactivation efficiency. In contrast, PL treatments showed a positive effect on color and general appearance of mung bean sprouts, while the respiration was almost unaffected. However, care must be taken with regard to efficiency-limiting matrix effects and impact on food quality. These aspects have to be assessed for each treated product. The integration of PL in industrial food processing plants could be an alternative way to improve the microbial quality of fresh produce, and therefore have a positive impact on public health by reducing the risk of contaminations with pathogenic bacteria.

Genotoxicity assessment of magnetic iron oxide nanoparticles with different particle sizes and surface coatings

Magnetic iron oxide nanoparticles (IONPs) have been widely used for various biomedical applications such as magnetic resonance imaging and drug delivery. However, their potential toxic effects, including genotoxicity, need to be thoroughly understood. In the present study, the genotoxicity of IONPs with different particle sizes (10, 30 nm) and surface coatings (PEG, PEI) were assessed using three standard genotoxicity assays, the Salmonella typhimurium reverse mutation assay (Ames test), the in vitro mammalian chromosome aberration test, and the in vivo micronucleus assay. In the Ames test, SMG-10 (PEG coating, 10 nm) showed a positive mutagenic response in all the five test bacterial strains with and without metabolic activation, whereas SEI-10 (PEI coating, 10 nm) showed no mutagenesis in all tester strains regardless of metabolic activation. SMG-30 (PEG coating, 30 nm) was not mutagenic in the absence of metabolic activation, and became mutagenic in the presence of metabolic activation. In the chromosomal aberration test, no increase in the incidence of chromosomal aberrations was observed for all three IONPs. In the in vivo micronucleus test, there was no evidence of increased micronuclei frequencies for all three IONPs, indicating that they were not clastogenic in vivo. Taken together, our results demonstrated that IONPs with PEG coating exhibited mutagenic activity without chromosomal and clastogenic abnormalities, and smaller IONPs (SMG-10) had stronger mutagenic potential than larger ones (SMG-30); whereas, IONPs with SEI coating (SEI-10) were not genotoxic in all three standard genotoxicity assays. This suggests that the mutagenicity of IONPs depends on their particle size and surface coating.

High Throughput Screening of Perfumery Raw Materials for Antimicrobial Properties

A microdilution protocol was developed and automated using a liquid handling station, allowing the determination of minimum inhibitory concentrations (MIC) of hydrophobic raw materials commonly used in the perfume industry (essential oils and synthetic chemicals). Tests were performed in 96-well microtiter plates against standard bacterial test strains and skin isolates involved in underarm malodor. The comparison with data previously reported in the literature indicated that the protocol was suitable, yielding MIC values that were in general agreement with those derived from manual test methods. For the majority of active test compounds, results showed a pronounced difference in susceptibility pattern between the Gram-positive and Gram-negative test strains used in this study. For a group of acyclic aliphatic aldehydes, a structure-activity relationship depending on the chain length was found.

Phytochemical constituents and antibacterial activity of some green leafy vegetables

To investigate the antibacterial activity and photochemicals of five green leafy vegetables against a panel of five bacteria strains. Disc diffusion method was used to determine the antibacterial activity, while kanamycin was used as a reference antibiotic. The phytochemical screening of the extracts was performed using standard methods. All methanol extracts were found active against all the test bacterial strains. Overall maximum extracts shows antibacterial activity which range from 6 to 15 mm. Proteins and carbohydrates was found in all the green leaves, whereas alkaloid, steroids, saponins, flavonoids, tannins were found in most of the test samples. The obtain result suggests that green leafy vegetables have moderate antibacterial activity and contain various pharmacologically active compounds and thus provide the scientific basis for the traditional uses of the studied vegetables in the treatment of bacterial infections.

Genotoxicity of styrene oligomers extracted from polystyrene intended for use in contact with food

Here, we conducted in vitro genotoxicity tests to evaluate the genotoxicity of styrene oligomers extracted from polystyrene intended for use in contact with food. Styrene oligomers were extracted with acetone and the extract was subjected to the Ames test (OECD test guideline No. 471) and the in vitro chromosomal aberration test (OECD test guideline No. 473) under good laboratory practice conditions. The concentrations of styrene dimers and trimers in the concentrated extract were 540 and 13,431 ppm, respectively. Extraction with acetone provided markedly higher concentrations of styrene oligomers compared with extraction with 50% ethanol aqueous solution, which is the food simulant currently recommended for use in safety assessments of polystyrene by both the United States Food and Drug Administration and the European Food Safety Authority. And these high concentrations of styrene dimers and trimers were utilized for the evaluation of genotoxicity in vitro. Ames tests using five bacterial tester strains were negative both in the presence or absence of metabolic activation. The in vitro chromosomal aberration test using Chinese hamster lung cells (CHL/IU) was also negative. Together, these results suggest that the risk of the genotoxicity of styrene oligomers that migrate from polystyrene food packaging into food is very low.

Inhibition of the mutagenic effects of N-methyl-N'-nitro-N-nitrosoguanidine and 9-Aminoacridine by indenopyridines in the Salmonella typhimurium tester strain 1537 and E. coli

The goal of the present research was to determine the protective potential of five newly synthesized indenopyridine derivatives against N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) and 9-aminoacridine (9-AA) induced mutagenesis. MNNG sensitive Escherichia coli WP2uvrA and 9-AA sensitive Salmonella typhimurium TA1537 were chosen as the bacterial tester strains. All of the test compounds showed significant antimutagenic activity at various tested concentrations. The inhibition rates ranged from 25.6% (Compound 2 - 1 mM/plate) to 68.2% (Compound 1 - 2.5 mM/plate) for MNNG and from 25.7% (Compound 4 - 1 mM/plate) to 76.1% (Compound 3 - 2.5 mM/plate) for 9-AA genotoxicity. Moreover, the mutagenicity of the test compounds was investigated by using the same strains. None of the test compounds has mutagenic properties on the bacterial strains at the highest concentration of 2.5 mM. Thus, the findings of the present study give valuable clues to develop new strategies for chemical prevention from MNNG and 9-AA genotoxicity by using synthetic indenopyridine derivatives.

Antioxidant and Antimicrobial Activity of Polyphenol Extracts from Wild Berry Fruits Grown in Southeast Serbia

Purpose: To assess the antioxidant and antimicrobial activities of polyphenolic extracts of three wild red wild berry fruit species from Southeast Serbia, viz, European cornel (Cornus mas), blackthorn (Prunus spinosa L.) and wild blackberry (Rubus fruticosus). Methods: Polyphenol content was determined using spectrophotometric and High performance liquid chromatography (HPLC) methods. Antioxidant activity was estimated by 2,2`- diphenyl - 1 - picrylhydrazyl (DPPH) test while reliminary antimicrobial tests were carried out by disc diffusion method in which antibacterial activity was evaluated by measuring the zone of inhibition against test bacterial strains. Broth microdilution method was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Results: The highest amount of total phenols was found in European cornel (8625.89 mg kg -1 FW). Galic acid, caffeic acid, p-coumaric acid, ferulic acid, (+)-catechin, procyanidin B2, (-)-epicatechin, quercetin, rutin and quercetin-3-glucoside were the main polyphenols in the fruit extracts. All extracts showed high scavenging effect on DPPH radical with IC 50 values ranging from 22.19 to 31.18 ml g -1 , as well as high antimicrobial activity on almost all the tested bacterial strains. Conclusion: Extracts of wild berry fruits may be suitable for the preparation of medicinal and nutritional products. Keywords: European cornel, Blackthorn, Wild blackberry, Polyphenols, Antioxidant, Antimicrobial

Evaluation of polymerized organo-selenium feed spacers to inhibit S. aureus and E. coli biofilm development in reverse osmosis systems

A feed spacer polymerized with organo-selenium was analyzed for biofouling inhibition to reverse osmosis membranes and feed spacers. The spacers polymerized with organo-selenium were tested for their ability to inhibit biomass formation and reduce biofilm thickness on the surface of RO membranes that the spacers were in contact with. The spacers and RO membranes were tested in a flow-cell system that exposed the sample membranes to normal operating conditions for RO membranes over a period of 24h. This system utilized a synthetic wastewater, selected as a high nutrient source to model a primary wastewater, and bacterial test strains were chosen to represent Gram-negative (Escherichia coli) and Gram-positive (Staphylococcus aureus) bacteria. Initial tests with spacers containing 0.55% organo-selenium in the spacer material showed inhibition levels of 2.0 logs for total biomass, and 2.7 logs for average biofilm thickness against S. aureus only. At a spacer concentration of 1% organo-selenium, inhibition levels of total specific biomass concentration averaged 2.9 logs for each bacterial strain, and averaged a reduction of biofilm thickness of 2.9 and 3.9 logs for S. aureus and E. coli.

Enhanced Production, Purification, Characterization and Mechanism of Action of Salivaricin 9 Lantibiotic Produced by Streptococcus salivarius NU10

BackgroundLantibiotics are small lanthionine-containing bacteriocins produced by lactic acid bacteria. Salivaricin 9 is a newly discovered lantibiotic produced by Streptococcus salivarius. In this study we present the mechanism of action of salivaricin 9 and some of its properties. Also we developed new methods to produce and purify the lantibiotic from strain NU10.Methodology / Principal FindingsSalivaricin 9 was found to be auto-regulated when an induction assay was applied and this finding was used to develop a successful salivaricin 9 production system in liquid medium. A combination of XAD-16 and cation exchange chromatography was used to purify the secondary metabolite which was shown to have a molecular weight of approximately 3000 Da by SDS-PAGE. MALDI-TOF MS analysis indicated the presence of salivaricin 9, a 2560 Da lantibiotic. Salivaricin 9 is a bactericidal molecule targeting the cytoplasmic membrane of sensitive cells. The membrane permeabilization assay showed that salivaricin 9 penetrated the cytoplasmic membrane and induced pore formation which resulted in cell death. The morphological changes of test bacterial strains incubated with salivaricin 9 were visualized using Scanning Electron Microscopy which confirmed a pore forming mechanism of inhibition. Salivaricin 9 retained biological stability when exposed to high temperature (90-100°C) and stayed bioactive at pH ranging 2 to 10. When treated with proteinase K or peptidase, salivaricin 9 lost all antimicrobial activity, while it remained active when treated with lyticase, catalase and certain detergents.ConclusionThe mechanism of antimicrobial action of a newly discovered lantibiotic salivaricin 9 was elucidated in this study. Salivaricin 9 penetrated the cytoplasmic membrane of its targeted cells and induced pore formation. This project has given new insights on lantibiotic peptides produced by S. salivarius isolated from the oral cavities of Malaysian subjects.

Validierung eines automatischen Testsystems für die Steriltestung von Amnionmembran

The use of cryopreserved amniotic membranes for the treatment of diseases and injuries of the surface of the eye is an established procedure in ophthalmological surgery. Before clinical use of cryopreserved amniotic membranes (AM) a careful testing for microbial contamination is essential to ensure a safe application. In this study the use of the BacT/Alert® test system was evaluated for screening of microbial growth in AMs. Minced fresh and cryopreserved AMs (approximately 5 × 5cm in size) were injected with 10ml of balanced salt solution in separate culture media test bottles and 10ml of cryopreservation medium bacterial and fungal test strains according to European Union (EU) regulations were applied to test the performance of the system. Approximately 10-100 colony forming units were applied on the samples prior to injection in the corresponding test bottles. Bottles were incubated at 37 °C for 7 days. Positive controls contained only balanced salt solution and the test strains while negative controls contained the test material without microbial test strains. Growth of the test strains was detected in all inoculated samples from non-processed and cryopreserved AM within the 7-day incubation period. In samples of the cryopreservation medium only growth of the fungus Candida albicans could be detected. The automated BacT/Alert test system is suitable for testing of microbial safety of amniotic membranes but not for testing the cryopreservation medium in clinical practice according to EU regulations.

New study of the essential oil, mineral composition and antibacterial activity of Pistacia lentiscus L. from Eastern Morocco

The present study describes the chemical composition of essential oils of Pistacia lentiscus L., collected from different regions of Eastern Morocco. The essential oils were obtained by hydro-distillation of the areal parts and analyzed by gas chromatography and gas chromatography coupled to a mass spectrometry. The study was conducted to determine the phytochemistry and antibacterial activities of oil from P. lentiscus leaves against both bacteria using the disc diffusion method. For Gram-negative, Salmonella sp., Klebsiella, Escherichia coli, and Pseudomonas, and for Gram-positive, Staphylococcus aureus, and Streptococcus were used as test bacterial strains. In the end, we completed the previous study by determining the minor and major mineral contents of leaves of P. lentiscus. The yield of P. lentiscus oil ranged between 0.13 and 0.23 %, with the chemical composition changing from one region to another. The variations are important between plant populations. The major oil components of PLL oil from Taforalt and Saidia (humid climate) were limonene and α-pinene, while myrcene, β-caryophyllene were found to be the major components of Laayoune and Jerada oil (semi-arid climate). For the mineral composition of PLL, the results show that trace element profiles of leaves from different regions differed significantly; the highest levels of K, Mg, Fe, and Ca were found in leaves of all locations, while the leaves collected from the Jerada region had the highest content of mineral substrates. This work also attempts to contribute to the knowledge of the nutritional properties of this plant; the results will be investigated for the evaluation of dietary information.

Substrate ambiguity among the nudix hydrolases: biologically significant, evolutionary remnant, or both?

Many members of the nudix hydrolase family exhibit considerable substrate multispecificity and ambiguity, which raises significant issues when assessing their functions in vivo and gives rise to errors in database annotation. Several display low antimutator activity when expressed in bacterial tester strains as well as some degree of activity in vitro towards mutagenic, oxidized nucleotides such as 8-oxo-dGTP. However, many of these show greater activity towards other nucleotides such as ADP-ribose or diadenosine tetraphosphate (Ap(4)A). The antimutator activities have tended to gain prominence in the literature, whereas they may in fact represent the residual activity of an ancestral antimutator enzyme that has become secondary to the more recently evolved major activity after gene duplication. Whether any meaningful antimutagenic function has also been retained in vivo requires very careful assessment. Then again, other examples of substrate ambiguity may indicate as yet unexplored regulatory systems. For example, bacterial Ap(4)A hydrolases also efficiently remove pyrophosphate from the 5' termini of mRNAs, suggesting a potential role for Ap(4)A in the control of bacterial mRNA turnover, while the ability of some eukaryotic mRNA decapping enzymes to degrade IDP and dIDP or diphosphoinositol polyphosphates (DIPs) may also be indicative of new regulatory networks in RNA metabolism. DIP phosphohydrolases also degrade diadenosine polyphosphates and inorganic polyphosphates, suggesting further avenues for investigation. This article uses these and other examples to highlight the need for a greater awareness of the possible significance of substrate ambiguity among the nudix hydrolases as well as the need to exert caution when interpreting incomplete analyses.

Antibacterial and antifungal activities of Actinobacteria isolated from Rathnagiri hills

Article history: Received on: 18/08/2012 Revised on: 29/08/2012 Accepted on: 05/09/2012 Available online: 29/10/2012 This study was performed to isolate actinomycete colonies having antibacterial and antifungal activity from soil samples. A total of 27 actinomycete colonies were isolated in pure culture from five soil samples using Starch casein agar medium. Entire isolates were screened for their antimicrobial activity by agar plug method against five each of human pathogenic bacteria and fungi. Of this, 7 strains inhibits B. substilis, 3 strains inhibits Klebseilla sp, 6 strains inhibits B. cerus, 5 strains inhibits S. aureus and only 2 strains inhibits E. coli. Incase of fungi all the actinobateria has moderate activity with less fungal strains, only 1 strain (RA 5) inhibits entire fungus except Penicillium sp. The metabolites from potent strain was produced by fermentation, separated by centrifugation, it was tested for their antimicrobial activity against the test bacterial and fungal strains by well diffusion and disc diffusion method. In this study, the metabolites from RA5 (identified as Streptomyces sp.) have showed good antibacterial and antifungal activity. Since many isolates showed inhibitory activity against indicator bacteria, it is suggestive that Rathnagiri hill’s soil could be an interesting source to explore for antibacterial secondary metabolites.

Chemoassay Screening of DNA-Reactive Mutagenicity with 4-(4-Nitrobenzyl)pyridine – Application to Epoxides, Oxetanes, and Sulfur Heterocycles

Organic electrophiles have the potential to covalently attack DNA bases, and thus initiate mutagenic and carcinogenic processes. In this context, aromatic nitrogen sites of the DNA bases are often particularly nucleophilic, with guanine N7 being one of the most favored sites of adduct formation with electrophilic xenobiotics. Employing 4-(4-nitrobenzyl)pyridine (NBP) as model nucleophile with a respective aromatic ═N- unit, a new kinetic variant of a photometric chemoassay for sensing the DNA reactivity of organic compounds is introduced and applied to 21 three- and four-membered oxygen and sulfur heterocycles (15 epoxides, two thiiranes, three oxetanes, and one thietane). Besides six unreactive compounds (oxetanes, thietane, and aliphatic epoxides with six or more side-chain carbons), second-order rate constants of the electrophile-NBP reaction, k(NBP), were obtained for 15 compounds, ranging from (1.16 ± 0.05)·10⁻³ to (36.5 ± 0.6)·10⁻³ L mol⁻¹ min⁻¹ in a methanol/tris-HCl buffer (16/84 v/v) reaction medium. Solvolysis as confounding factor was addressed by determining respective first-order rate constants k(solv). Analysis of the k(NBP) values resulted in structure-reactivity relationships, and comparison with literature data from the Ames test bacterial strains TA100, TA1535, and TA97 (Salmonella typhimurium) as well as from WP2 uvrA (Escherichia coli) revealed significant log-log relationships between the mutagenic potency of the heterocycles and their reactivity toward NBP. The latter demonstrates the potential of the NBP chemoassay as a nonanimal component of integrated testing strategies for REACH, enabling an efficient screening of organic electrophiles with respect to their DNA reactivity and associated mutagenicity and carcinogenicity.

Toxicologic Characterization of a Novel Explosive, Guanidinium 3,4-Dinitropyrazolate (GDNP), in Female Rats and Ames Mutagenicity Assay

Sustainable use of military training ranges requires the development of compounds that have a minimal impact to the environment when used in a weapon system. Guanidinium 3,4-dinitropyrazolate (GDNP) is a novel explosive compound of interest for application in some weapon systems. Little is known of its toxicologic properties. To ensure the health of potentially exposed personnel and the environment, initial toxicity investigations were conducted and the results were compared with another widely used energetic (hexahydro-1,3,5-trinitro-1,3,5-triazine [RDX]). In a microplate Ames assay, GDNP was not cytotoxic to bacterial tester strains at concentrations less than 100 μg/mL. However, GDNP was mutagenic to 4 of 5 bacterial strains with and without S9 metabolic incubation at concentrations as low as 0.7 μg/mL. Unlike RDX, GDNP did not have an affinity for the γ-aminobutyric acid(A) receptor convulsant site and was predicted to not induce seizure. After acute oral dosing in female rats, the median lethal dose in female rats of GDNP in tap water solution was determined to be 720 mg/kg. Daily oral exposure to 500 mg/kg per d of GDNP for 14 days caused weight loss, increased liver and spleen weights, and adverse histopathologic events in kidney and spleen. These adverse events were not observed in animals receiving lower doses of GDNP. In this study, the lowest-observed-adverse-effect-level from oral exposure to GDNP for 14 days was 500 mg/kg per d and the no-observable-adverse-effect-level was 152 mg/kg per d.

Evaluation of preservative efficacy in pharmaceutical products: the use of psychrotolerant, low-nutrient preferring microbes in challenge tests

Preservative efficacy in medicines is typically investigated using challenge tests. In such tests, the product is artificially contaminated with a high concentration of standard bacterial and fungal test strains such as Pseudomonas aeruginosa and Candida albicans. The rate and extent of reductions in inoculum viability over a specified period forms the basis for acceptance/rejection of preservative efficacy. None of the strains named for inclusion in the challenge test outlined in the European Pharmacopoeia are associated with the contamination of high-quality water used in pharmaceutical production. Alpha- and Betaproteobacteria are easily the most common microbes in waters intended for pharmaceutical production. In addition, none of the standard test strain panel prefer low-nutrient, dilute conditions or grow at or around refrigeration temperatures. This is important because the water activity and nutrient content of medicines can vary greatly and medicines are often stored cold. We investigate the relevance of these factors when testing preservative efficacy by including other strains in challenge tests. Psychrotolerant, low-nutrient preferring strains (Beta- and Alphaproteobacteria and a yeast) were isolated from pristine waters. These were compared in challenge tests with C. albicans and P. aeruginosa using different storage temperatures. Pharmaceutical products differing widely in water-content, pH and preservative systems were included in the study. Regardless of the type of medicine tested C. albicans always showed superior survival characteristics to the yeast isolate (Cryptococcus terricola). One of the three screened bacterial strains (a Sphingomonas sp.) survived significantly better than P. aeruginosa in all but one product tested. However, the results for all products taken together cannot easily be explained by reference to this strain's psychrotolerancy or its preference for dilute, low-nutrient environments. This study supports previous work indicating that the inclusion of wild-type test strains, in this instance strains that are suited to survival in high-quality waters, improves preservative efficacy tests. Use of a Sphingomonas sp. isolated from a pristine water as a challenge test strain, gave a more stringent indication of preservative efficacy in a wide range of pharmaceuticals than did P. aeruginosa.

Dual effect of macroalgal extracts on growth of bacteria in Western Baltic Sea

It is assumed that the biological characteristics of the bacterial strains used in bioactivity tests have strong influences on their susceptibility against antibacterial compounds. Therefore, the selection of bacterial test strains may rush conclusions on the effect of macroalgal extracts and metabolites on bacteria. To proof this assumption, we have analysed the biological activities of crude extracts of 16 macroalgae from the coastal waters of Kiel Fjord (Germany), and tested their effect against a panel of 10 microorganisms comprising 5 standard test strains of bacteria and 5 macroalga-associated bacteria. Fourteen macroalgae (88%) displayed antibacterial activity against at least one of the test strains. Despite the high proportion of extracts exhibiting antimicrobial activity, only 3 strains of the standard set were susceptible to macroalgal extracts and the overall activities were low (less than 80% of inhibition). Most of active extracts inhibited Bacillus subtilis, while no inhibition effects were found against Erwinia amylovora, Escherichia coli, and the macroalga-associated bacteria. In contrast, all extracts produced stimulatory growth effects of at least two of the tested bacteria. While growth stimulation of standard set of bacteria was rare (22.5% of total tests) with exception of plant pathogen Erwinia amylovora and two cases of Staphylococcus lentus, it was common among bacteria associated with macroalgae (77.5%), especially Bacillus algicola, Pseudomonas marincola and both algal-pathogenic bacteria. This study demonstrates that macroalgal extracts can display different effects, i.e., inhibition or stimulation of bacterial growth depending on the origin of the test strains, which are derived from a standard panel or from the marine environment, respectively.

A test for the “physiological phagemia” hypothesis—natural intestinal coliphages do not penetrate to the blood in horses

Soon after the discovery of bacteriophages in 1917, Felix d'Herelle demonstrated that they are present in humanand animal-associated microbiota, and speculated that these phages may play certain roles in the immunity against bacterial infections (d'Herelle 1926). That remains an intriguing but not yet proven possibility up to this day (reviewed in Gorski and Weber-Dabrowska 2005). It has been often reported that phages administrated orally or rectally in the course of phage therapy can penetrate through mucosa of gastrointestinal tract (GIT) to the blood (see Letarov et al. 2010; Gorski et al. 2006 for review) and it was suggested that natural intestinal bacteriophages might also translocate continuously from GIT to the blood. This state has been termed a “physiological phagemia” (Gorski et al. 2006). Coliphages, for example, were detected in a large fraction of commercial lots of animal sera used for eukaryotic cell cultures (reviewed in detail in Gorski et al. 2006). Since the bacteriophage communities present in different subjects are highly individual (Reyes et al. 2010; Golomidova et al. 2007; see also Letarov and Kulikov 2009), it is crucial to perform phage detection in the blood and in the intestinal contents simultaneously, using bacterial test strains able to detect the intestinal phages present in a particular subject to determine the extent to that intestinal phages in fact can routinely reach the blood. The aim of the work presented here is to quantify the presence of natural coliphages in the blood of the animals using a highly sensitive culturing approach. The choice of animal model was based on our data indicating that in contrast to humans and some other animal species (Reyes et al. 2010; see also refs in Letarov and Kulikov 2009), the intestinal viral community in horses is dominated by virulent phages that may occur in high titers in some healthy individuals (Golomidova et al. 2007; Kulikov et al. 2007; and our unpublished observations). Here, we present the analysis of bacteriophages in feces and in the blood of three horses. To select the animals for this study, we screened available population of horses for the titers of the fecal coliphages detectable on E.coli NM522 (Stratagene, USA) strain and selected three animals that yielded more than 10 plaque forming units (PFU)/g of coliphages. The indigenous coliform strains (ICSs) frequently detect different types of the coliphages in the same sample (Golomidova et al. 2007). In order to select a subset of these bacteria for subsequent phage detection, a series of 30 ICS isolates were obtained from each animal and subjected to whole-cell PCR fingerprinting using IS1tr (5′ATCAGTAAGTTGGA(G/A)(T/G)CATTACC-3′) and TR8834 (5′-ATCGGCGATGCGTTGACGAAT-3′) primers as described elsewhere (Isaeva et al. 2010; see also Golomidova et al. 2007). Briefly, the conventional PCR reactions (30 cycles consisting of 94°C for 15 s, 56°C for 30 s, 72°C for 45 s), using boiled cells from the plate as a template, were set up to generate strain-specific banding patterns. This analysis confirmed that all of the isolates were distinct, indicating M. Letarova and D. Strelkova contributed equally to this work. M. Letarova : S. Nevolina :A. Letarov (*) Winogradsky Institute of Microbiology RAS, Prospect 60-letiya Oktyabrya 7/2, 117312 Moscow, Russia e-mail: letarov@gmail.com

Pharmacological, genotoxic and phytochemical properties of selected South African medicinal plants used in treating stomach-related ailments

Ethnopharmacological relevanceThe evaluated medicinal plants are used in South African traditional medicine in treating stomach-related ailments. Aims of the studyThe study aimed at evaluating the pharmacological, genotoxic and phytochemical properties of the seven selected medicinal plants used for treating stomach-related ailments. Materials and methods: Ethyl acetate (EtOAc), ethanol (EtOH) 70% and water extracts of the selected plant parts were evaluated for their antimicrobial and anthelmintic activities using microdilution assays. Gram-positive bacteria (Enterococcus faecalis and Staphylococcus aureus), Gram-negative bacterium (Escherichia coli) and Candida albicans were used for antimicrobial assays. Caenorhabditis elegans was used for the anthelmintic assay. Plant extracts were also assayed for their cyclooxygenase-inhibitory activity against cyclooxygenase-1 and -2 enzymes. The Ames test was used to evaluate the genotoxicity of the plant extracts. A spectrophotometric method was used to determine the total phenolics, gallotannins, flavonoids and saponins. ResultsTwelve extracts exhibited minimum inhibitory concentration (MIC) <1mg/mL against the bacterial test strains, and five extracts exhibited MIC <1mg/mL against Candida albicans. The EtOAc extract of Tetradenia riparia had the best minimum lethal concentration (MLC) value (0.004mg/mL) against Caenorhabditis elegans. All the EtOAc extracts exhibited percentage inhibition in the range of 50.7–94.7% against COX-1 and -2 enzymes at 250μg/mL. All the plant extracts were non-mutagenic towards Salmonella typhimurium tester strains TA98, TA100 and TA1537 without metabolic activation. Phytochemical analysis revealed relatively high amounts of total phenolics, gallotannins and flavonoids in the evaluated plant extracts. ConclusionsThe general pharmacological activities exhibited by some of the plant extracts in this study support the traditional uses of the selected plants in treating stomach-related ailments. The Ames test showed that all the plant extracts were non-mutagenic but cytotoxicity tests are needed to ascertain the safety for long-term consumption.