Purification and Characterization of a Novel Anti-<Emphasis Type="Italic">Campylobacter</Emphasis> Bacteriocin Produced by <Emphasis Type="Italic">Lactobacillus curvatus</Emphasis> DN317

The lactic acid bacteria (LAB) microbiota of Saudi chicken ceca was determined. From 60 samples, 204 isolates of lactic acid bacteria were obtained. Three isolates produced antimicrobial activities against Campylobacter jejuni, Listeria monocytogenes, and Bacillus subtilis. The isolate DN317, which had the highest activity against Campylobacter jejuni ATCC 33560, was identified as Lactobacillus curvatus (GenBank accession numbers: KX353849 and KX353850). Full inhibitory activity was observed after a 2-h incubation with the supernatant at pH values between 4 and 8. Only 16% of the activity was conserved after a treatment at 121 °C for 15 min. The use of proteinase K, pepsin, chymotrypsin, trypsin, papain, and lysozyme drastically reduced the antimicrobial activity. However, lipase, catalase, and lysozyme had no effect on this activity. The active peptide produced by Lactobacillus curvatus DN317 was purified by precipitation with an 80% saturated ammonium sulfate solution, and two steps of reversed phase HPLC on a C18 column. The molecular weight of this peptide was 4448 Da as determined by MALDI-ToF. N-terminal sequence analysis using Edman degradation revealed 47 amino acid residues (UniProt Knowledgebase accession number C0HK82) revealing homology with the amino acid sequences of sakacin P and curvaticin L442. The antimicrobial activity of the bacteriocin, namely curvaticin DN317, was found to be bacteriostatic against Campylobacter jejuni ATCC 33560. The use of microbial antagonism by LAB is one of the best ways to control microorganisms safely in foods. This result constitutes a reasonable advance in the antimicrobial field because of its potential applications in food technology.     

5-Nitroimidazole-derived Schiff bases and their copper(II) complexes exhibit potent antimicrobial activity against pathogenic anaerobic bacteria

In the present work a family of novel secnidazole-derived Schiff base compounds and their copper(II) complexes were synthesized. The antimicrobial activities of the compounds were evaluated against clinically important anaerobic bacterial strains. The compounds exhibited in vitro antibacterial activity against Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides vulgatus, Bacteroides ovatus, Parabacteroides distasonis and Fusubacterium nucleatum pathogenic anaerobic bacteria. Upon coordination to copper(II) the antibacterial activity significantly increased in several cases. Some derivatives were even more active than the antimicrobial drugs secnidazole and metronidazole. Therefore, the compounds under study are suitable for in vivo evaluation and the microorganisms should be classified as susceptible to them. Electrochemical studies on the reduction of the nitro group revealed that the compounds show comparable reduction potentials, which are in the same range of the bio-reducible drugs secnidazole and benznidazole. The nitro group reduction potential is more favorable for the copper(II) complexes than for the starting ligands. Hence, the antimicrobial activities of the compounds under study might in part be related to intracellular bio-reduction activation. Considering the increasing resistance rates of anaerobic bacteria against a wide range of antimicrobial drugs, the present work constitutes an important contribution to the development of new antibacterial drug candidates.     

Comparison of Antimicrobial Properties of Silver Nanoparticles Synthesized from Selected Bacteria

Green silver nanoparticle (AgNP) biosynthesis is facilitated by the enzyme mediated reduction of Ag ions by plants, fungi and bacteria. The antimicrobial activity of green AgNPs is useful to overcome the challenge of antimicrobial resistance. Antimicrobial properties of biosynthesized AgNPs depend on multiple factors including culture conditions and the microbial source. The antimicrobial activity of AgNPs biosynthesized by Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923 and Acinetobacter baumannii (confirmed clinical isolate) were investigated in this study. Biosynthesis conditions (AgNO₃ concentration, pH, incubation temperature and incubation time) were optimized to obtain the maximum AgNP yield. Presence of AgNPs was confirmed by observing a characteristic UV–Visible absorbance peak in 420–435 nm range. AgNP biosynthesis was optimal at 0.4 g/L AgNO₃ concentration under alkaline conditions at 60–70 °C. The biosynthesized AgNPs showed higher stability compared to chemogenized AgNPs in the presence of electrolytes. AgNPs synthesized by P. aeruginosa were the most stable while NPs of S. aureus were the least stable. AgNPs synthesized by P. aeruginosa and S. aureus showed good antimicrobial potential against E. coli, P. aeruginosa, S. aureus, MRSA and Candida albicans. AgNPs synthesized by S. aureus had greater antimicrobial activity. The antimicrobial activity of NPs may vary depending on the size and the morphology of NPs.     

Bioprospecting of <Emphasis Type="Italic">Diaporthe terebinthifolii</Emphasis> LGMF907 for antimicrobial compounds

Antibiotic-resistant bacteria have been observed with increasing frequency over the past decades, driving the search for new drugs and stimulating the interest in natural products sources. Endophytic fungi from medicinal plants represent a great source of novel bioactive compounds useful to pharmaceutical and agronomical purposes. Diaporthe terebinthifolii is an endophytic species isolated from Schinus terebinthifolius, a plant used in popular medicine for several health problems. The strain D. terebinthifolii LGMF907 was previously reported by our group to produce secondary metabolites with biological activity against phytopathogens. Based on these data, strain LGMF907 was chosen for bioprospecting against microorganisms of clinical importance and for characterization of major secondary metabolites. In this study, different culture conditions were evaluated and the biological activity of this strain was expanded. The crude extracts demonstrated high antibacterial activity against Escherichia coli, Micrococcus luteus, Saccharomyces cerevisiae, methicillin-sensitive Staphylococcus aureus, and methicillin-resistant S. aureus. The compounds diaporthin and orthosporin were characterized and also showed activity against the clinical microorganisms evaluated. This study discloses the first isolation of diaporthin and orthosporin from D. terebinthifolii, and revealed the potential of this endophytic fungus to produce secondary metabolites with antimicrobial activity.     

Possible Probiotic Lactic Acid Bacteria Isolated from Oysters (<Emphasis Type="Italic">Crassostrea gigas</Emphasis>)

We attempted to isolate lactic acid bacteria (LAB) from the marine oyster (Crassostrea gigas) and selected several environmental stress-resistant isolates for the development of a future probiotic adjuvant for marine aquaculture. Twenty-six presumptive LAB isolates were extracted from oysters and screened (by an agar diffusion assay) for antimicrobial activity toward various pathogens: Vibrio parahaemolyticus, Streptococcus iniae, and Edwardsiella tarda. Eight isolates had an antibacterial activity toward V. parahaemolyticus; in particular, 6 isolates showed a growth-inhibitory activity, with inhibition zone diameters > 15 mm. Of these, 5 isolates (JL17, JL18, JL28, HL7, and HL32) were also active against S. iniae and E. tarda. Enterococcus faecium HL7 was selected as the isolate most resistant to environmental stressors: the minimum NaCl, ethanol, and hydrogen peroxide concentrations at which HL7 cells lost their viability were 1.9 M, 11%, and 0.013%, respectively. When an antibiotic sensitivity test was performed on E. faecium HL7, this isolate was found to be resistant to trimethoprim/sulfamethoxazole, cephalothin, ampicillin, rifampin, gentamicin, cefotaxime, cefepime, cefotetan, nalidixic acid, and kanamycin. While the oyster model studies provided indication that E. faecium HL7 could be a good candidate as biocontrol agent against V. vulnificus, further optimization is needed in the actual animal rearing situation.     

Genomic characterisation,detection of genes encoding virulence factors and evaluation of antibiotic resistance of <Emphasis Type="Italic">Trueperella pyogenes</Emphasis> isolated from cattle with clinical metritis

Trueperella pyogenes is one of the most important microorganisms causing metritis in post-partum cattle. Co-infection with other bacterial species such as Escherichia coli or Fusobacterium necrofurom increases the severity of the disease and the persistence of bacteria in utero. The aim of this study was to investigate the frequency of T. pyogenes strains, and their virulence and antimicrobial resistant profiles in metritis cases. The study was carried out on 200 samples obtained from metritis discharges of postpartum cattle on 18 farms around Tehran, Iran. Sixty-five T. pyogenes isolates (32.5%) were identified, of which 16 isolates were detected as pure cultures and the other 49 isolates from cultures most commonly mixed with E. coli or F. necrofurom. In terms of diversity in biochemical characteristic of T. pyogenes strains, 8 different biotypes were identified among the isolates. Single or multi antimicrobial resistance was observed in 48 isolates (73.9%), which was mostly against trimethoprim sulfamethoxazole, azithromycin, erythromycin and streptomycin. The tetracycline resistance gene tetW and macrolide resistance genes ermB and ermX were detected in 30, 18 and 25 isolates, respectively. In the screening of genes encoding virulence factors, fimA and plo genes were identified in all tested isolates. Genes encoding nanP, nanH, fimC, fimG, fimE and cbpA were detected in 50, 54, 45, 40, 50 and 37 of isolates, respectively. Thirteen different genotypes were observed in these T. pyogenes isolates. A significant association between clonal types and virulence factor genes, biochemical profile, CAMP test result, severity of the disease and sampling time was detected.     

The Phenolic Contents and Antimicrobial Activities of Some Marine Algae from the Mediterranean Sea (Algeria)

In this study, three marine algae collected from western coast of algerian mediteranean sea (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) were tested using the agar-well diffusion method for their production of antibacterial and antifungal agents on various organisms that cause diseases of humans and plants (Eschirichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Salmonella sp, Candida albicans, and Penicillium sp.). The total phenol content and antimicrobial activity were determined using different crude seaweeds extracts (methanol, diethylether, and chloroform). The results show that the chloroform extracts of (Ulva lactuca and Corallina elongata) had the highest activity against E. coli and Salmonella sp. The methanol extract obtained from (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) showed antifungal activity for Candida albicans. The results of the study revealed that the seaweeds from Algeria appear to have immense potential as a source of antibacterial and antifungal compounds; they can be used in treating diseases caused by these organisms.     

Biological activity of new flavonoid from <Emphasis Type="Italic">Hieracium pilosella</Emphasis> L.

Hieracium pilosella L. (Asteraceae) is a well-known plant used in ethno-medicine as its inflorescences are particularly rich in beneficial polyphenolics. This research aimed to elucidate the structure of a new flavone glycoside isolated from the inflorescences of Hieracium pilosella and evaluate its antioxidant, antimicrobial and antiproliferative activities. The chromatographic methods were successfully applied to isolate the new flavonoid. Its structure was determined by subsequent UV, NMR and MS experiments and identified as isoetin 4′-O-β-D-glucopyranoside. Free radical scavenging capacity was examined by measuring the scavenging activity of the new isoetin derivative on 2,2-diphenyl-1-picrylhydrazyl (DPPH). The compound was also screened for spectrum of antimicrobial activity using the agar well diffusion method. Minimum inhibitory concentration (MIC) for Pseudomonas aeruginosa ATCC 9027 was performed by the micro-dilution broth method. The antiproliferative effect of tested glycoside was assessed in two human tumor cell lines derived from lung (A549) and colon (HT-29) carcinoma and cell proliferation was determined by means of MTT method. The tested compound showed high antiradical activity, reducing the DPPH? with EC₅₀ 7.9 μM (3.7 µg/ml) and exhibited narrow antimicrobial spectrum among tested microorganisms. The compound was active against Pseudomonas aeruginosa ATCC 9027 (MIC 125 μg/ml) which is prone to causing infections that are difficult to treat due to it developing extremely rapid antibiotic resistance. In the antiproliferative studies, cell proliferation of the colon (HT-29) carcinoma cell line was significantly decreased after exposure to the compound. The results indicate that isoetin 4′-O-β-D-glucopyranoside possesses antioxidant capacity and very promising antibacterial activity and could have uses as an effective antipseudomonal agent as well a antiproliferative agent.     

Effect of salt stress on the antimicrobial activity of <Emphasis Type="Italic">Ruta chalepensis</Emphasis> essential oils

In the current investigation, the biological activities of essential oils obtained from organs of Ruta chalepensis plants grown under salt stress (0, 50 and 100 mM NaCl) were analyzed. Their chemical composition was often investigated by GC/FID and GC–MS and the antimicrobial activities towards eight bacteria (Salmonella All, Salmonella K, Escherichia coli 45AG, Escherichia coli 45AI, Staphylococcus aureus 9402, Staphylococcus aureus 02B145, Listeria 477 and Pseudomonas aeruginosa ATCC 10145) and five fungi strains (Aspergillus, Saccharomycee crvisiale, Streptomyces griseus, Fusarium solani and Penicillium thomii) were studied. Results revealed that salt increased essential oil production in leaves at 50 and 100 mM NaCl. A total of 20 compounds were identified in leaves, undecan-2-one, nonan-2-one and geijerene being the dominant ones. In stems, 21 compounds were found; they were dominated by decan-2-one, geijerene, nonan-2-one and undecan-2-one. In contrast, roots exhibited a large variation with 25 volatile compounds and octyl acetate, methyl decanoate, phytyl acetate were the major ones. Salt stress induced significant antibacterial activity changes, mainly in leaves and stems. In leaves, the minimum inhibitory and bactericidal concentration decreased at 100 mM NaCl against Listeria 477, the two strains of E. coli (45AG and 45AI) and P. aeruginosa but it increased versus other bacteria. In stems, salt increased oil antibacterial activity against all strains except P. aeruginosa ATCC 10145. Root oil showed the least antibacterial activity under saline conditions versus Listeria 477 and P. aeruginosa ATCC 10145. As regards antifungal activity, NaCl reduced the antifungal activity of essential oils against the majority of fungi strains.     

Study of ectoparasitism of ultramicrobacteria of the genus <Emphasis Type="Italic">Kaistia</Emphasis>, strains NF1 and NF3 by electron and fluorescence microscopy

Transmission electron and fluorescence microscopy was used to study the character of the interaction of free-living ultramicrobacterial (UMB) strains NF1 and NF3, affiliated with the genus Kaistia, and seven species of gram-positive and gram-negative heterotrophic bacteria. Strains NF1 and NF3 were found to exhibit parasitic activity against gram-positive Bacillus subtilis and gram-negative Acidovorax delafildii. UMB cells are tightly attached to the envelopes of the victim cells and induce their lysis, thus demonstrating the features of typical ectoparasitism. The selectivity of parasitism of the studied UMB to the victim bacteria has been shown: only two soil microorganisms of the seven test objects, B. subtilis ATCC 6633 and an aerobic gramnegative bacterium A. delafildii 39, were found to be sensitive to UMB attack. Other bacteria (Micrococcus luteus VKM Ac-2230, Staphylococcus aureus 209-P, Pseudomonas putida BS394, Escherichia coli C 600, and Pantoea agglomerans ATCC 27155) were not attacked by UMB. It was established for the first time that free-living UMB may be facultative parasites not only of phototrophic bacteria, as we have previously demonstrated [1], but of heterotrophic bacteria as well. The UMB under study seem to play an important role in the regulation of the quantity of microorganisms and in the functioning of microbial communities in some natural ecotopes.     

Analysis of antimicrobial and immunomodulatory substances produced by heterofermentative <Emphasis Type="Italic">Lactobacillus reuteri</Emphasis>

Antimicrobial and immunomodulatory potential of various Lactobacillus reuteri strains is closely connected to their metabolite production profile under given cultivation conditions. We determined the in vitro production of antimicrobial substances such as organic acids, ethanol, and reuterin by four strains of L. reuteri (L. reuteri E, L. reuteri KO5, L. reuteri CCM 3625, and L. reuteri ATCC 55730). All studied L. reuteri strains showed the ability to produce lactic acid, acetic acid, and ethanol with concominant consumption of glucose and together with phenyllactic acid—a potent antifungal compound—with concominant consumption of phenylalanine. The reuterin production from glycerol was confirmed for all analyzed lactobacilli strains except L. reuteri CCM 3625. Production of organic acids, ethanol, and reuterin is significantly involved in antimicrobial activity of lactobacilli which was determined using the dual-culture overlay diffusion method against six indicator bacteria and five indicator moulds. In comparison to the referential L. reuteri ATCC 55730, the highest inhibition potential was observed against Escherichia coli CCM 3988 and Pseudomonas aeruginosa CCM 3955. Among analyzed indicators of moulds, the growth of Alternaria alternata CCM F-128 was the most inhibited by all four analyzed L. reuteri strains. Finally, the immunomodulatory potential of analyzed lactobacilli were proven by the determination of the in vitro production of biogenic amines histamine and tyramine. L. reuteri CCM 3625 was able to produce tyramine, and L. reuteri E and L. reuteri KO5 were able to produce histamine under given cultivation conditions.     

<Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> as a Probiotic Potential from Kouzeh Cheese (Traditional Iranian Cheese) and Its Antimicrobial Activity

The aim of this study is to isolate and identify Lactobacillus plantarum isolates from traditional cheese, Kouzeh, and evaluate their antimicrobial activity against some food pathogens. In total, 56 lactic acid bacteria were isolated by morphological and biochemical methods, 12 of which were identified as Lactobacillus plantarum by biochemical method and 11 were confirmed by molecular method. For analyzing the antimicrobial activity of these isolates properly, diffusion method was performed. The isolates were identified by 318 bp band dedicated for L. plantarum. The isolated L. plantarum represented an inhibitory activity against four of the pathogenic bacteria and showed different inhibition halos against each other. The larger halos were observed against Staphylococcus aureus and Staphylococcus epidermidis (15 ± 0.3 and 14.8 ± 0.7 mm, respectively). The inhibition halo of Escherichia coli was smaller than that of other pathogen and some L. plantarum did not show any inhibitory activity against E. coli, which were resistant to antimicrobial compounds produced by L. plantarum. The isolated L. plantarum isolates with the antimicrobial activity in this study had strong probiotic properties. These results indicated the nutritional value of Kouzeh cheese and usage of the isolated isolates as probiotic strains.     

A New Broad-Spectrum Peptide Antibiotic Produced by <Emphasis Type="Italic">Bacillus brevis</Emphasis> Strain MH9 Isolated from Margalla Hills of Islamabad,Pakistan

The antimicrobial peptide from a bacterial strain is isolated from soil sample of Margalla Hills of Islamabad, Pakistan. The peptide is found to significantly inhibit the growth of both Gram-positive (Staphylococcus aureus ATCC 6538 and Micrococcus luteus ATCC 10240) and Gram-negative (Escherichia coli ATCC 25922 and Salmonella typhi ATCC 14028) bacteria as compared to gramicidin as standard. The bacterium is identified as Bacillus brevis strain MH9 based on phenotype and phylogenetic analysis. The antibacterial polypeptide was produced optimally at 35 °C after 48 h of growth, precipitated by 50 % ammonium sulphate, and further purified using HPLC. The sequential steps of purification decrease the peptide contents with prominent antibacterial activity. The peptide composed of 11 amino acid was further characterized by FT-IR and NMR. Results suggested that the peptide molecule is a novel antibacterial agent that is effective against both Gram-positive and Gram-negative bacteria. This study may have important implications for new peptide antibiotic that could be a new addition to treat infections.     

Controlled biosynthesis of AgCl nanoparticles by a thermotolerant <Emphasis Type="Italic">Aspergillus terreus</Emphasis> in the L-Tryptophan supplemented media: Characterization and antimicrobial activity

A simple and green method was developed for the extracellular biosynthesis of silver chloride nanoparticles, free from silver nanoparticles, using cell-free filtrate of a thermotolerant fungal strain Aspergillus terreus 8. The synthesized silver chloride nanoparticles exhibited characteristic absorption maximum at 275 nm. As-fabricated AgCl-NPs were characterized by UV-vis spectroscopy, XRD, SEM-EDX, and FT-IR. The biosynthesized silver chloride nanoparticles exhibited strong antimicrobial activity towards pathogenic microorganisms such as Fusarium oxysporum f. sp. vasinfectum and Verticillium dahliae. The synthesized silver chloride nanoparticles can be exploited as a promising new biocide bionanocomposite against pathogenic microorganisms.     

Phylogeny and bioactivity of epiphytic Gram-positive bacteria isolated from three co-occurring antarctic macroalgae

Marine macroalgae are emerging as an untapped source of novel microbial diversity and, therefore, of new bioactive secondary metabolites. This study was aimed at assessing the diversity and antimicrobial activity of the culturable Gram-positive bacteria associated with the surface of three co-occurring Antarctic macroalgae. Specimens of Adenocystis utricularis (brown alga), Iridaea cordata (red alga) and Monostroma hariotii (green alga) were collected from the intertidal zone of King George Island, Antarctica. Gram-positive bacteria were investigated by cultivation-based methods and 16S rRNA gene sequencing, and screened for antimicrobial activity against a panel of pathogenic microorganisms. Isolates were found to belong to 12 families, with a dominance of Microbacteriaceae and Micrococcaceae. Seventeen genera of Actinobacteria and 2 of Firmicutes were cultured from the three macroalgae, containing 29 phylotypes. Three phylotypes within Actinobacteria were regarded as potentially novel species. Sixteen isolates belonging to the genera Agrococcus, Arthrobacter, Micrococcus, Pseudarthrobacter, Pseudonocardia, Sanguibacter, Staphylococcus, Streptomyces and Tessaracoccus exhibited antibiotic activity against at least one of the indicator strains. The bacterial phylotype composition was distinct among the three macroalgae species, suggesting that these macroalgae host species-specific Gram-positive associates. The results highlight the importance of Antarctic macroalgae as a rich source of Gram-positive bacterial diversity and potentially novel species, and a reservoir of bacteria producing biologically active compounds with pharmacological potential.     

Biodegradation of sulfonamides by <Emphasis Type="Italic">Shewanella oneidensis</Emphasis> MR-1 and <Emphasis Type="Italic">Shewanella</Emphasis> sp. strain MR-4

Because of extensive sulfonamides application in aquaculture and animal husbandry and the consequent increase in sulfonamides discharged into the environment, strategies to remediate sulfonamide-contaminated environments are essential. In this study, the resistance of Shewanella oneidensis MR-1 and Shewanella sp. strain MR-4 to the sulfonamides sulfapyridine (SPY) and sulfamethoxazole (SMX) were determined, and sulfonamides degradation by these strains was assessed. Shewanella oneidensis MR-1 and Shewanella sp. strain MR-4 were resistant to SPY and SMX concentrations as high as 60 mg/L. After incubation for 5 days, 23.91 ± 1.80 and 23.43 ± 2.98% of SPY and 59.88 ± 1.23 and 63.89 ± 3.09% of SMX contained in the medium were degraded by S. oneidensis MR-1 and Shewanella sp. strain MR-4, respectively. The effects of the initial concentration of the sulfonamides and initial pH of the medium on biodegradation, and the degradation of different sulfonamides were assessed. The products were measured by LC–MS; with SPY as a substrate, 2-AP (2-aminopyridine) was the main stable metabolite, and with SMX as a substrate, 3A5MI (3-amino-5-methyl-isoxazole) was the main stable metabolite. The co-occurrence of 2-AP or 3A5MI and 4-aminobenzenesulfonic acid suggests that the initial step in the biodegradation of the two sulfonamides is S–N bond cleavage. These results suggest that S. oneidensis MR-1 and Shewanella sp. strain MR-4 are potential bacterial resources for biodegrading sulfonamides and therefore bioremediation of sulfonamide-polluted environments.     

Zinc oxide and titanium dioxide nanoparticles induce oxidative stress,inhibit growth,and attenuate biofilm formation activity of <Emphasis Type="Italic">Streptococcus mitis</Emphasis>

Streptococcus mitis from the oral cavity causes endocarditis and other systemic infections. Rising resistance against traditional antibiotics amongst oral bacteria further aggravates the problem. Therefore, antimicrobial and antibiofilm activities of zinc oxide and titanium dioxide nanoparticles (NPs) synthesized and characterized during this study against S. mitis ATCC 6249 and Ora-20 were evaluated in search of alternative antimicrobial agents. ZnO and TiO₂-NPs exhibited an average size of 35 and 13 nm, respectively. The IC₅₀ values of ZnO and TiO₂-NPs against S. mitis ATCC 6249 were 37 and 77 µg ml^?1, respectively, while the IC₅₀ values against S. mitis Ora-20 isolate were 31 and 53 µg ml^?1, respectively. Live and dead staining, biofilm formation on the surface of polystyrene plates, and extracellular polysaccharide production show the same pattern. Exposure to these nanoparticles also shows an increase (26–83 %) in super oxide dismutase (SOD) activity. Three genes, namely bapA1, sodA, and gtfB like genes from these bacteria were identified and sequenced for quantitative real-time PCR analysis. An increase in sodA gene (1.4- to 2.4-folds) levels and a decrease in gtfB gene (0.5- to 0.9-folds) levels in both bacteria following exposure to ZnO and TiO₂-NPs were observed. Results presented in this study verify that ZnO-NPs and TiO₂-NPs can control the growth and biofilm formation activities of these strains at very low concentration and hence can be used as alternative antimicrobial agents for oral hygiene.     

Synthesis of functionalized benzo[<Emphasis Type="Italic">f</Emphasis>]2<Emphasis Type="Italic">H</Emphasis>-chromenes and evaluation of their antimicrobial activities

Knoevenagel cyclocondensations of α-hydroxy naphthaldehyde with β-oxodithioesters and ketene dithioacetals yielded 2H-benzo[f]chromene-2-thiones and 2H-benzo[f]chromen-2-ones, respectively, in high yields. The newly synthesized compounds were evaluated for antifungal and antibacterial activities. Among them, compounds (2-furyl)(3-thioxo-3H-benzo[f]chromen-2-yl)methanone and phenyl(3-oxo-3H-benzo[f]chromen-2-yl)methanone exhibited excellent antifungal activity against tested fungi Curvularia lunata and Fusarium moniliforme. The highest antibacterial activity against the tested bacteria Escherichia coli and Staphylococcus aureus was observed for (4-chlorophenyl)(3-oxo-3H-benzo[f]chromen-2-yl)methanone. The results of antimicrobial screening demonstrate that (2-furyl)(3-thioxo-3H-benzo[f]chromen-2-yl)methanone, phenyl(3-oxo-3H-benzo[f]chromen-2-yl)methanone, and (4-chlorophenyl)(3-oxo-3H-benzo[f]chromen-2-yl)methanone are promising as antimicrobial drugs.     

Antifungal activity of 3-acetylbenzamide produced by actinomycete WA23-4-4 from the intestinal tract of <Emphasis Type="Italic">Periplaneta americana</Emphasis>

Actinomycetes are well-known for producing numerous bioactive secondary metabolites. In this study, primary screening by antifungal activity assay found one actinomycete strain WA23-4-4 isolated from the intestinal tract of Periplaneta americana that exhibited broad spectrum antifungal activity. 16S rDNA gene analysis of strain WA23-4-4 revealed close similarity to Streptomyces nogalater (AB045886) with 86.6% sequence similarity. Strain WA23-4-4 was considered as a novel Streptomyces and the 16s rDNA sequence has been submitted to GenBank (accession no. KX291006). The maximum antifungal activity of WA23-4-4 was achieved when culture conditions were optimized to pH 8.0, with 12% inoculum concentration and 210 ml ISP2 medium, which remained stable between the 5^th and the 9^th day. 3-Acetyl benzoyl amide was isolated by ethyl acetate extraction of WA23-4-4 fermentation broth, and its molecular formula was determined as C₉H₉NO₂ based on MS, IR, ¹H, and ¹³C NMR analyses. The compound showed significant antifungal activity against Candida albicans ATCC 10231 (MIC: 31.25 μg/ml) and Aspergillus niger ATCC 16404 (MIC: 31.25 μg/ml). However, the compound had higher MIC values against Trichophyton rubrum ATCC 60836 (MIC: 500 μg/ml) and Aspergillus fumigatus ATCC 96918 (MIC: 1,000 μg/ml). SEM analysis showed damage to the cell membrane of Candida albicans ATCC 10231 and to the mycelium of Aspergillus niger ATCC 16404 after being treatment with 3-acetyl benzoyl amide. In conclusion, this is the first time that 3-acetyl benzoyl amide has been identified from an actinomycete and this compound exhibited antifungal activity against Candida albicans ATCC 10231 and Aspergillus niger ATCC 16404.     

LyeTx I,a potent antimicrobial peptide from the venom of the spider <Emphasis Type="Italic">Lycosa erythrognatha</Emphasis>

LyeTx I, an antimicrobial peptide isolated from the venom of Lycosa erythrognatha, known as wolf spider, has been synthesised and its structural profile studied by using the CD and NMR techniques. LyeTx I has shown to be active against bacteria (Escherichia coli and Staphylococcus aureus) and fungi (Candida krusei and Cryptococcus neoformans) and able to alter the permeabilisation of l-α-phosphatidylcholine-liposomes (POPC) in a dose-dependent manner. In POPC containing cholesterol or ergosterol, permeabilisation has either decreased about five times or remained unchanged, respectively. These results, along with the observed low haemolytic activity, indicated that antimicrobial membranes, rather than vertebrate membranes seem to be the preferential targets. However, the complexity of biological membranes compared to liposomes must be taken in account. Besides, other membrane components, such as proteins and even specific lipids, cannot be discarded to be important to the preferential action of the LyeTx I to the tested microorganisms. The secondary structure of LyeTx I shows a small random-coil region at the N-terminus followed by an α-helix that reached the amidated C-terminus, which might favour the peptide-membrane interaction. The high activity against bacteria together with the moderate activity against fungi and the low haemolytic activity have indicated LyeTx I as a good prototype for developing new antibiotic peptides.