Mercury contamination disrupts ecosystems by causing toxicity and bioaccumulation in higher trophic levels. This study inspected the heavy metals content and the culturable diversity of mercury-resistant and detoxifying bacteria in samples from dumping site, thermal power plant and sewage treatment plant situated in/near Chandigarh, India. Most of the analyzed heavy metals were found above the permissible limit in these sites. Among 87 isolated bacteria, 45 could resist >200 µM ionic mercury (HgCl2). Markedly, 10 and 4 isolates harbored the key genes required for mercury detoxification, merA and merB gene, respectively. The merA gene-containing strain's phylogeny indicated the resistant strains belong to Alphaproteobacteria, Betaproteobacteria, Bacilli and Gammaproteobacteria. Nine isolates could volatilize HgCl2 efficiently, while two could volatilize organic mercury (CH3HgCl). The isolates Niallia circulans DCL_26, Serratia quinivorans DCS_A1, and Pseudomonas hibiscicola R_24 were found to resist and volatilize up to ≥300 μM HgCl2. Appreciably, S. quinivorans DCS_A1 could resist and remediate up to 6 μM of CH3HgCl. A 5-fold reduction in ionic mercury concentration was observed within 72 h of treatment for all three tested strains. Phytotoxicity assay performed using Arabidopsis thaliana and Brassica juncea seeds confirmed the strains DCL_26, DCS_A1, and R_24 have the ability to remove mercury from culture media as well as industrial effluent and reduce mercury toxicity significantly. The study provides a green technology based on microbial systems to remediate mercury-contaminated wastewater for its use in crop irrigation. However, further studies are required for the sustainable and safer application of these novel isolates to remediate mercury efficiently.

A novel mercury-resistant bacterium, designated strain DCL_24T, was isolated from the legacy waste at the Daddu Majra dumping site in Chandigarh, India. It showed resistance up to 300 mu M of inorganic mercury (mercuric chloride). The isolate was found to be a Gram-negative, facultative anaerobic, motile, and rod-shaped bacterium that can grow at 4 - 30 degrees C (optimum 25 degrees C), pH 6.0 - 12.0 (optimum 7.0), and 0 - 4.0 % (w/v) NaCl (optimum 0.5 - 2.0 %). The 16 S rRNA gene-based phylogenetic analysis showed that DCL_ 24 T shared a 97.53 % similarity with itsolosest type strain Rheinheimera muenzenbergensis E-49T. Insilico DNA-DNA hybridization and average nucleotide identity values were found to be 18.60 % and 73.77 %, respectively, between the genomes of DCL_24T and R. muenzenbergensis E-49T. The strain DCL_24T has 44.33 DNA G+C content (mol %). Based on the pheno-typic, chemotaxonomic, and genotypic data, the strain DCL_24T represents a novel species within the genus Rheinheimera, for which the name Rheinheimera metallidurans sp. nov is proposed. The type strain is DCL_24T (MTCC13203T = NBRC115780T = JCM 35551 T). The isolate was found to volatilize and remove mercury efficiently, as demonstrated by X-ray film and dithizone-based colorimetric methods. Around 92 % of mercury removal was observed within 48 h. The mercury-resistant determinant mer operon consisting of merA, encoding the mercuric reductase enzyme, and transport and regulatory genes (merT, merP, merD, and merR) were found in the isolate. Relative expression analysis of merA at increasing concentrations of HgCl2 was confirmed by quantitative real-time PCR. These data indicate the merA-mediated reduction of toxic Hg2+ into a non-toxic volatile Hg0. The phytotoxicity assay performed using Arabidopsis thaliana seeds further demonstrated the mercury toxicity reduction potential of DCL_24T. The study shows that this novel isolate, DCL_24T, is an inter-esting candidate for mercury bioremediation. However, further studies are required to assess the bioremediation efficacy of the strain under the harsh environmental conditions prevailing in polluted sites.

Abstract The unique environment of space is characterized by several stress factors, including intense radiation, microgravity, high vacuum and extreme temperatures, among others. These stress conditions individually or in-combination influence genetics and gene regulation and bring potential evolutionary changes in organisms that would not occur under the Earth's gravity regime (1 × g). Thus, space can be explored to support the emergence of new varieties of microbes and plants, that when selected for, can exhibit increased growth and yield, improved resistance to pathogens, enhanced tolerance to drought, low nutrient and disease, produce new metabolites and others. These properties may be more difficult to achieve using other approaches under 1 × g. This review provides an overview of the space microgravity and ionizing radiation conditions that significantly influence organisms. Changes in the genomics, physiology, phenotype, growth and metabolites of organisms in real and simulated microgravity and radiation conditions are illustrated. Results of space biological experiments show that the space environment has significant scientific, technological and commercial potential. Combined these potentials can help address the future of life on Earth, part of goal e of astrobiology.

This work aimed to study in-situ toxicity assessment of pulp and paper industry wastewater on Trigonella foenum-graecum L. Physico-chemical analysis of wastewater revealed high pH (8.1), total suspended solids, total dissolved solids, biochemical oxygen demand and chemical oxygen demand (89, 2160, 8670 and 25600 mg L−1 , respectively), electrical conductivity (2156 ms cm−1), and heavy metal (mg L−1) contents as Fe (79.61), Zn (49.06), Cu (6.07), Cd (6.07), Mg (19.66), and Ni (6.43), which were higher than compare to control The major pollutants detected via GC–MS analysis were hexadecanoic acid, octadecanoic acid, nonacosane, and β-sitosterol trimethylsilyl ether. UV–Vis spectrophotometry and Fourier transform infrared spectroscopy analyses of the wastewater confirmed the presence of residual organic and inorganic pollutants. The cytotoxicity and genotoxicity of the wastewater for plants were also assessed and revealed chromosomal and cytological aberrations in wastewater-treated T. foenum-graecum L. The accumulation of metals in and the effects of such accumulation on the physiology of T. foenum-graecum L. plants irrigated with wastewater were also studied, and it was concluded that antioxidant enzyme activity, chlorophyll, protein content, and dry weight were reduced in wastewater irrigated T. foenum-graecum L. plants compared to those in the control (fed with normal water). Scanning and transmission electron microscopy analyses of plant roots, root nodules, and stomata revealed the accumulation of heavy metals on the surfaces and in tissues. The present study highlights the urgent necessity of proper treatment of paper and pulp industry wastewater before their disposal into water bodies to control the damage to environment and aquatic life.

Living organisms adapt to changing environments using their amazing flexibility to remodel themselves by a process called evolution. Environmental stress causes selective pressure and is associated with genetic and phenotypic shifts for better modifications, maintenance, and functioning of organismal systems. The natural evolution process can be used in complement to rational strain engineering for the development of desired traits or phenotypes as well as for the production of novel biomaterials through the imposition of one or more selective pressures. Space provides a unique environment of stressors (e.g., weightlessness and high radiation) that organisms have never experienced on Earth. Cells in the outer space reorganize and develop or activate a range of molecular responses that lead to changes in cellular properties. Exposure of cells to the outer space will lead to the development of novel variants more efficiently than on Earth. For instance, natural crop varieties can be generated with higher nutrition value, yield, and improved features, such as resistance against high and low temperatures, salt stress, and microbial and pest attacks. The review summarizes the literature on the parameters of outer space that affect the growth and behavior of cells and organisms as well as complex colloidal systems. We illustrate an understanding of gravity-related basic biological mechanisms and enlighten the possibility to explore the outer space environment for application-oriented aspects. This will stimulate biological research in the pursuit of innovative approaches for the future of agriculture and health on Earth.

A mutated phytoene desaturase (pds) gene, pds-L504R, conferring resistance to the herbicide norflurazon has been reported as a dominant selectable marker for the genetic engineering of microalgae (Steinbrenner and Sandmann in Appl Environ Microbiol 72:7477–7484, 2006; Prasad et al. in Appl Microbiol Biotechnol 98(20):8629–8639, 2014). However, this mutated genomic clone harbors several introns and the entire expression cassette including its native promoter and terminator has a length > 5.6 kb, making it unsuitable as a standard selection marker. Therefore, we designed a synthetic, short pds gene (syn-pds-int) by removing introns and unwanted internal restriction sites, adding suitable restriction sites for cloning purposes, and introduced the first intron from the Chlamydomonas reinhardtii RbcS2 gene close to the 5′end without changing the amino acid sequence. The syn-pds-int gene (1872 bp) was cloned into pCAMBIA 1380 under the control of a short sequence (615 bp) of the promoter of pds (pCAMBIA 1380-syn-pds-int). This vector and the plasmid pCAMBIA1380-pds-L504R hosting the mutated genomic pds were used for transformation studies. To broaden the existing transformation portfolio, the rhodophyte Porphyridium purpureum was targeted. Agrobacterium-mediated transformation of P. purpureum with both the forms of pds gene, pds-L504R or syn-pds-int, yielded norflurazon-resistant (NR) cells. This is the first report of a successful nuclear transformation of P. purpureum. Transformation efficiency and lethal norflurazon dosage were determined to evaluate the usefulness of syn-pds-int gene and functionality of the short promoter of pds. PCR and Southern blot analysis confirmed transgene integration into the microalga. Both forms of pds gene expressed efficiently as evidenced by the stability, tolerance and the qRT-PCR analysis. The molecular toolkits and transformation method presented here could be used to genetically engineer P. purpureum for fundamental studies as well as for the production of high-value-added compounds.

The antibacterial peptide of Bacillus licheniformis MCC 2016 have potential biopreservative efficacy. Here, we report the purification process, properties, and mode of action of this antibacterial peptide for its potential application in the food industry. The antibacterial peptide from the cell-free supernatant was purified using a sequence of purification steps. The purified antibacterial peptide showed a specific activity of 68817 AU mg(-1) and 0.4% yield. Liquid chromatography-mass spectroscopy analysis showed an mz(-1) value of 279.28 for the active peptide. The SDS-PAGE analysis confirmed the antibacterial peptide is low-molecular weight and the size is between 3.0 and 3.5 kDa. Scanning electron microscopy, Fourier transform infrared spectroscopy, beta-gal induction assay and release of UV-absorbing materials indicated that the antibacterial peptide targets the cell wall of pathogens. Minimum inhibitory concentration of the antibacterial peptide against Listeria monocytogenes Scott A and others (Kocuria rhizophila ATCC 9341, Staphylococcus aureus FRI 722 and Salmonella typhimurium MTCC 1251) was found to be 1600 and 800 AU mL(-1), respectively. The antibacterial peptide is temperature and pH stable, proteolytic-enzyme-sensitive, low-molecular weight, cell wall active class I bacteriocin and exhibits remarkable antibacterial activity against pathogens, suggesting its application as a potential biopreservative in the food industry.

Background Photosynthetic euglenids are major contributors to fresh water ecosystems. Euglena gracilis in particular has noted metabolic flexibility, reflected by an ability to thrive in a range of harsh environments. E. gracilis has been a popular model organism and of considerable biotechnological interest, but the absence of a gene catalogue has hampered both basic research and translational efforts. Results We report a detailed transcriptome and partial genome for E. gracilis Z1. The nuclear genome is estimated to be around 500 Mb in size, and the transcriptome encodes over 36,000 proteins and the genome possesses less than 1% coding sequence. Annotation of coding sequences indicates a highly sophisticated endomembrane system, RNA processing mechanisms and nuclear genome contributions from several photosynthetic lineages. Multiple gene families, including likely signal transduction components, have been massively expanded. Alterations in protein abundance are controlled post-transcriptionally between light and dark conditions, surprisingly similar to trypanosomatids. Conclusions Our data provide evidence that a range of photosynthetic eukaryotes contributed to the Euglena nuclear genome, evidence in support of the ‘shopping bag’ hypothesis for plastid acquisition. We also suggest that euglenids possess unique regulatory mechanisms for achieving extreme adaptability, through mechanisms of paralog expansion and gene acquisition.

The marine microalga Pavlova lutheri has great nutritional value as it synthesizes and accumulates higher amounts of polyunsaturated fatty acids (PUFAs). It is commonly used in aquaculture. However, no transformation procedure has been realized for this industrially important microalga so far. Here, we report an efficient protocol for the nuclear transformation of P. lutheri. Agrobacterium-mediated transformation (AMT) of P. lutheri with a mutated genomic clone of phytoene desaturase (pds) gene, pds-L504R, from Haematococcus pluvialis yielded norflurazon-resistant P. lutheri cells. Ideal co-cultivation conditions for achieving higher numbers of transformants was found to be artificial seawater (ASW) medium, 100 μM acetosyringone, and a 24-h co-cultivation at 25 ± 1 °C. The integration of the introduced gene into the nuclear genome of P. lutheri was shown by PCR amplification of the T-DNA sequences from the genomic DNA of transformants and Southern blot analysis using T-DNA sequences as probes. The transgene expressed efficiently as evidenced by the results of stability and tolerance study, and the qRT-PCR analysis. Results clearly demonstrate the application of AMT approach and pds gene as a dominant selectable marker for the genetic engineering of P. lutheri for fundamental studies and biotechnological applications.

The bio-preservative efficacy of a partially purified antibacterial peptide (ppABP) produced by Bacillus licheniformis Me1 in an economical medium developed using agro-industry waste was evaluated by direct application in milk and milk-based food products. The addition of ppABP in milk samples stored at 4 ± 2 °C and 28 ± 2 °C resulted in the growth inhibition of pathogens Listeria monocytogenes Scott A, Micrococcus luteus ATCC 9341, and Staphylococcus aureus FRI 722. The shelf life of milk samples with added ppABP increased to 4 days at 28 ± 2 °C, whereas curdling and off-odor were noticed in samples without ppABP. Furthermore, the milk samples with ppABP were sensorily acceptable. Antilisterial effect was also observed in cheese and paneer samples treated with ppABP. These results clearly indicate that the ppABP of B. licheniformis Me1 can be utilized as a bio-preservative to control the growth of spoilage and pathogenic bacteria, thereby reducing the risk of food-borne diseases.

Photosynthetic euglenids are major components of aquatic ecosystems and relatives of trypanosomes. Euglena gracilis has considerable biotechnological potential and great adaptability, but exploitation remains hampered by the absence of a comprehensive gene catalogue. We address this by genome, RNA and protein sequencing: the E. gracilis genome is >2Gb, with 36,526 predicted proteins. Large lineage-specific paralog families are present, with evidence for flexibility in environmental monitoring, divergent mechanisms for metabolic control, and novel solutions for adaptation to extreme environments. Contributions from photosynthetic eukaryotes to the nuclear genome, consistent with the shopping bag model are found, together with transitions between kinetoplastid and canonical systems. Control of protein expression is almost exclusively post-transcriptional. These data are a major advance in understanding the nuclear genomes of euglenids and provide a platform for investigating the contributions of E. gracilis and its relatives to the biosphere.

Isochrysis galbana and Isochrysis sp. are economically important microalgae from the division of haptophytes. Here, we report Agrobacterium-mediated stable DNA transfer into their nuclear genomes. Initial studies were performed to standardize co-cultivation media and determine the sensitivity of the microalgae to selective agents. Up to 1 mg/ml of the antibiotic hygromycin did not inhibit growth, whereas both the haptophytes bleached in artificial seawater (ASW) medium containing micromolar concentrations of the herbicide norflurazon. Co-cultivation of Isochrysis sp. and I. galbana with Agrobacterium tumefaciens strain LBA 4404 harboring the binary vector pCAMBIA 1380-pds-L504R yielded norflurazon-resistant (NR) colonies visible on selective plates after 20–30 days. pCAMBIA 1380-pds-L540R was constructed by cloning a mutated genomic phytoene desaturase (pds) gene from Haematococcus pluvialis as a selectable marker gene into the binary vector system pCAMBIA 1380. Co-cultivation of Isochrysis sp. with A. tumefaciens in ASW medium containing 200 μM of acetosyringone for 72 h produced the highest number of NR cells. For I. galbana, 100 μM of acetosyringone, ASW medium, and 48 h co-cultivation period appeared to be optimum co-cultivation parameters. The NR colonies kept their resistance phenotype for at least 24 months, even in the absence of selective pressure. The transfer of the pds gene in NR cells was shown by PCR amplification of the T-DNA sequences from the genomic DNA of NR cells and Southern blot analysis using T-DNA sequences as probes. The genetic manipulation described here will allow metabolic engineering and a better understanding of several biochemical pathways in the future.

Saccharina (Laminaria) japonica, a safe, cheap, and readily available macroalga can be used as a substrate for various microbial fermentations. This work investigated the feasibility of S. japonica as a substrate for production of pigments by the fungus Talaromyces amestolkiae GT11 in solid-state fermentation without additional salt and/or nitrogen sources. Under optimized conditions, the pigment exhibited maximum absorption spectrum at 410 (yellow) and 510 nm (red), and the pigment yield of 1,153.5 (yellow) and 506.2 (red) OD units g−1 of dry fermented substrate were achieved with a particle size of 1.0 mm and pH 7, although visually the pigment was reddish in color. The optimum incubation period, pH, moisture, inoculum size, and temperature were observed to be at 192 h, pH 7.0, 80 % (w/w) moisture, 1.8 × 106 spores mL−1 of inoculum g-1 of dry substrate and 28 °C. Hence, this study indicates the suitability of utilization of S. japonica as a substrate for natural pigment production by T. amestolkiae GT11 which can be used in food, cosmetics and pharmaceutical industries for various applications.

In this study, the feasibility and applicability of marine algal biomass Saccharina (Laminaria) japonica as a sole substrate for the production of pigments by Talaromyces amestolkiae GT11 in submerged fermentation was evaluated. Results indicated that the fungus T. amestolkiae GT11 produced the highest amount of extracellular yellow (444.83 ± 22) and red (200.94 ± 12), and intracellular yellow (362.28 ± 34) and red (193.87 ± 10) pigments, utilizing 1% (w/v) of S. japonica powder at an initial pH of 5 and 30°C, as compared to other physiochemical parameters tested. The pH and thermostability analysis results demonstrated that even after 5 h of incubation the pigment was found to be highly stable at pH 6 and 40 ~ 60°C with 98% and 90.56 ~ 84.69% of residual absorbance, respectively. Apart from the application of pigment as a natural colorant instead of synthetic one in biotechnology industry, the fermented substrate itself can be exploited as food and feed with enhanced nutrient content, improved protein quality and fiber digestibility, etc. However, further studies concerning the safety and functional properties of the pigment and fermented substrate are required. Furthermore, this study provides the evidences about the biological method of making easily fermentable biomass for biorefiners or other metabolite production.

Aims An attempt was made to evaluate the effectiveness of partially purified antibacterial peptide (ppABP) produced by Bacillus licheniformis Me1 for food preservation by means of active packaging.Methods and Results: The active packaging films containing ppABP were developed using two different packing materials [low-density polyethylene (LDPE) and cellulose films] by two different methods: soaking and spread coating. The activated films showed antibacterial activity against pathogens. The release study of ppABP from coated film showed that the LDPE films liberated ppABP as soon as it comes in contact with water, while gradual release of coated ppABP was observed in case of cellulose films. The activated films showed residual activity in different simulating conditions, such as pH of food and storage temperatures. The activated films demonstrated its biopreservative efficacy in controlling the growth of pathogens in cheese and paneer.Conclusions: The ppABP-activated films were found to be effective for biopreservation. The ppABP from active films got diffused into the food matrix and reduced the growth rate and maximum growth population of the target micro-organism.Significance and impact of the Study: Both types of ppABP-activated films can be used as a packaging material to control spoilage and pathogenic organisms in food, thereby extending the shelf life of foods.

Probiotics have established their efficacy as dietary adjuncts providing benefits to consumers. However, selection of probiotics before incorporation into diet requires close scrutiny in the form of in vitro as well as in vivo tests. Three bacteriocinogenic Bacillus sp., namely, B. licheniformis Me1, B. flexus Hk1, and B. subtilis Bn1 previously isolated from milk, cheese and fermented beans, respectively, were characterized for typical in vitro probiotic criteria. When compared to probiotic Bacillus coagulans, all three cultures were found to possess better acid and bile tolerance. Cultures Me1 and Bn1, except Hk1, showed bile salt hydrolase activity. A marked difference in adhesion to hydrocarbons and auto-aggregation properties from 10–80 and 60–99%, respectively, were observed for the tested cultures. Highest antioxidant activity was measured for culture Hk1 (66.6%), whereas least activity of 53% was observed for culture Bn1. Cultures Me1 and Bn1 were sensitive to all the antibiotics tested, whereas Hk1 and B. coagulans showed resistance to the penicillin group of β-lactum antibiotics. All the tested cultures showed a broad spectrum of activity against food-borne pathogens. In co-cultivation studies, B. licheniformis Me1 completely inhibited the growth of the indicator pathogen Listeria monocytogenes ScottA. Overall, the test cultures exhibiting potential probiotic characteristics, particularly B. licheniformis Me1, can serve as probiotics of commercial interest.

Bacillus species are usually found in a number of food products, in which they cooperate with other microorganisms during fermentation and releases valuable compounds including enzymes and antimicrobial substances. Antibacterial substances produced by food-grade Bacillus spp. can find potential biopreservatives in food systems. In this study, Bacillus spp. were selectively isolated from various food sources based on their antibacterial activity against Micrococcus luteus ATCC 9341. Among 78 such isolates, 25 potent isolates were selected for further studies, and were characterized and differentiated by phenological and molecular approaches. From these 25 isolates, seven isolates were taken which showed considerable diversity in their characteristic features and were identified as B. subtilis Ec1, B. thuringiensis Ik15, B. licheniformis Me1, B. cereus Ik11, Bacillus flexus Hk1, B. megaterium Pk12 and B. amyloliquefaciens Bk1. These cultures have been deposited in Microbial Culture Collection, National Centre for Cell Science, India. Most of the isolates displayed wide range of inhibitory activity against both Gram-positive and Gramnegative food-borne pathogenic bacteria. The antibacterial substances produced by these isolates were partially characterised. The culture, B. licheniformis Me1 produces an antibacterial substance which is proteinaceous in nature, stable over a wide temperature and pH, and exhibits wide range of inhibitory spectrum. Based on these results, B. licheniformis Me1 was selected for further studies. The antibacterial peptide (ABP) produced by B. licheniformis Me1 was purified from culture supernatant using ammonium sulphate, butanol extraction and HPLC. A single active peak was obtained after RP-HPLC which showed an m/z value of 279.28, determined by mass-spectroscopy. The tricine SDS-PAGE analysis of the purified ABP revealed that it is low molecular weight (MW) peptide of 3 to 3.4 kDa. The effect of ABP on food-borne pathogens, such as Listeria monocytogenes Scott A, Staphylococcus aureus FRI 722, Salmonella typhimurium MTCC 1251 and B. cereus F 4433 revealed that it has bactericidal effect. The scanning electron microscopy images of ABP-treated cells revealed complete disruption of the cells. In addition, the mode of action studies using FTIR, release of UV absorbing materials, cell reporter assays showed that the target of action of the ABP was on the cell wall of the pathogens. An economical production media (consisting of corn steep liquor (a byproduct for corn milling), yeast extract and NaCl) was developed for maximum production of ABP. The optimum conditions for higher yield of ABP (51200 AU/ml) in this formulated media was found to be temperature 37°C, pH 8, agitation speed 150 cycles/min and an incubation time of 24 h. The technological properties and the mode of action of the ABP indicate that it may belong to the cationic- membrane-acting lantibiotic group. In order to verify the suitability of B. licheniformis Me1 for food application the safety of the culture was evaluated in vitro and in vivo. The isolate was nonhaemolytic and non-phospholytic. The in vivo toxicological safety analysis as determined using suitable animal models revealed that the culture is safe, non toxic, non-irritant, and non-mutagenic and can be used for food industry application. The ABP produced by this culture was analysed for the biopreservative effectiveness. Two different methodologies were tested, direct application and food packaging film application. The ABP was found to be efficient in controlling the growth of L. monocytogenes Scott A in milk and other dairy products, such as cheese and paneer. Furthermore, the ABP was used to develop active packaging films (LDPE and cellulose) and their efficacy in controlling growth of pathogens in packaged food was also evaluated. Both the active (LDPE and cellulose) films with ABP showed inhibitory activity against common food-borne pathogens tested. The absorption and release of ABP to and from the cellulose film was found to be better as compared to LDPE film, since the binding of ABP in the former case was evaluated to be more stable. The dairy products, such as cheese and paneer samples inoculated with L. monocytogenes Scott A, and packed with ABP activated cellulose and LDPE films showed a reduction in the viable count of the indicator organism. In addition, a comparative evaluation of the in vitro probiotic properties of the Bacillus cultures was carried out and the culture B. licheniformis Me1 exhibited potential probiotic properties, such as acid and bile tolerance, bile salt hydrolase activity, hydrophobicity, antioxidant activity, etc. With this work, a novel food-grade isolate, B. licheniformis Me1 was obtained which can find potential application in food industry as a probiotic or as biopreservative.

Reporter bacteria are beneficial for the rapid and sensitive screening of cultures producing peptide antibiotics, which can be an addition or alternative to the established antibiotics. This study was carried out to validate the usability of specific reporter strains for the target mediated identification of antibiotics produced by native Bacillus spp. isolated from different food sources. During preliminary classification, cell wall stress causing Bacillus isolates were screened by using reporter strain Bacillus subtilis BSF2470. The isolates which induced cell wall stress were further characterized for their specific mode of action by using other B. subtilis reporter strains (TMB 488, TMB 299 and TMB 279). The isolate B. licheniformis N12 was found to produce bacitracin confirmed by the response to reporter strain B. subtilis TMB 279 and by putative identification of bacitracin biosynthetic loci. The other isolate B. subtilis EC1 also induced B. subtilis TMB 279, but does not possess the bacitracin gene cluster indicating that it can be a novel, bacitracin like antibiotic. The different but related subsets of peptide antibiotics that bind the pyrophosphate moiety of the lipid carrier of cell wall biosynthesis can be identified using this whole cell based reporter strains.

In this study, an in vivo toxicological safety assessment of Bacillus licheniformis Me1, a native isolate from milk, was performed. An acute toxicity study in male albino Wistar rats demonstrated no treatment-related illness or mortality. A 90-day subchronic oral toxicity study using 2 doses (1.1 × 10(10) and 1.1 × 10(11) colony-forming unit [CFU]/kg body weight [BW], respectively) failed to show dose-dependent illness or mortality. Moreover, neither significant differences in serum biochemical and hematological analyses nor histopathological changes in organs or tissues were found when compared to the control groups. The no-observed-adverse-effect level (NOAEL) was found to be greater than 1.1 × 10(11) CFU/kg BW. The in vivo micronucleus assay in mice did not reveal any signs of genotoxic effect at any of the doses tested. Furthermore, dermal and acute eye irritation tests conducted in rabbits showed no edema or erythema and ocular lesions. These results suggest that B licheniformis Me1 can be considered safe for food industry applications.