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Acinetobacter baumannii is an opportunistic multidrug-resistant pathogen that causes a significant mortality rate. The proteins containing Tetratrico Peptide Repeats (TPRs) are involved in the pathogenicity and virulence of bacteria and have different roles such as transfer of bacterial virulence factors to host cells, binding to the host cells and inhibition of phagolysosomal maturation. So, in this study, physicochemical properties of a new protein containing TPRs in A. baumannii which was named PcTPRs1 by this study were characterized and its 3D structure was predicted by in-silico tools. The protein B and T cell epitopes were mapped and its vaccine potential was in-silico and in-vivo investigated. Domain analysis indicated that the protein contains the Flp pilus assembly protein TadD domain which has three TPRs. The helix is dominant in the protein structure, and this protein is an outer membrane antigen which, is extremely conserved among A. baumannii strains; thus, has good properties to be applied as a recombinant vaccine. The best-predicted and refined model was applied in ligand-binding sites and conformational epitopes prediction. Based on epitope mapping results, several epitopes were characterized which could stimulate both immune systems. BLAST results showed the introduced epitopes are completely conserved among A. baumannii strains. The in-vivo analysis indicates that a 101 amino acid fragment of the protein which contains the best selected epitope, can produce a good protectivity against A. baumannii as well as the whole TPR protein and thus could be investigated as an effective subunit and potential vaccines.

Background Pseudomonas aeruginosa infection is the major cause of death in burn patients. Thus, in this study, a chimeric vaccine harboring the OprF185-350-OprI22-83-PcrV was designed and expressed in Escherichia coli. The immunogenicity of the recombinant chimer, OprI, OprF, and PcrV was studied in a burned mouse model. Methodology Recombinant proteins including the proposed chimer, OprF, OprI, and PcrV were expressed in the E.coli. Mice were immunized with the purified recombinant proteins, and the antibody titre was estimated in the sera obtained from immunized mice. Immunized and control mice were challenged with 2, 5, and 10xLD50 of the P. aeruginosa strains (PAO1, PAK, and R5), and microbial counts were measured in the skin, liver, spleen, and kidney of the studied mice. Results Results showed that the antibody titre (total IgG) was significantly increased by injection of 10 μg of chimeric protein in the experimental groups compared to the control groups. The antibody survival titre was high until 235 days after administration of the second booster. The survival rate of the mice infected with 10xLD50 was significantly increased and the number of bacteria was reduced, especially in the internal organs (kidney, spleen, and liver) compared to the mice immunized with any of the OprF, OprI, and PcrV proteins alone. Conclusion The findings of our study revealed that the chimeric protein is a promising vaccine candidate for control of the P. aeruginosa infection.

Background: Reduction in the quality and quantity of food can be due to the presence and growth of fungi in food. Responsible for contamination of feed and food, are some species of Aspergillus. Objectives: Objectives of this study were to investigate the essential oil of Satureja rechingeri as antifungal agent against strains of Aspergillus parasiticus NRRL2999 and Aspergillus flavus PICC-AF39 (productive aflatoxin B1). Methods: Antifungal activities of the oil to inhibit growth of aflatoxins of Aspergillus parasiticus NRRL2999 and Aspergillus flavus PICC-AF39 were studied. Antifungal properties of essential oil of S. rechingeri at a concentration of 107 and 105 spores/mL on aflatoxin fungus Aspergillus parasiticus NRRL2999 and Aspergillus flavus PICC-AF39 were studied. Antifungal assessment activity was performed using disk diffusion method. The minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) were determined. Results: Inhibition zone diameter for 20 µL oil 107 spores/mL of Aspergillus parasiticus NRRL2999 and Aspergillus flavus PICC-AF39, was reported 55, 70 mm as respectively and MIC and MFC, for the oil was respectively 150, 450 and 150, 300 ppm. Inhibition zone diameter for 20 µL oil 105 spores of Aspergillus parasiticus NRRL2999 and Aspergillus flavus PICC-AF39, respectively 65, 78 mm was reported and MIC and MFC, for the oil were 50, 200 and 50, 100 ppm respectively. Conclusions: Natural compounds such S. rechingeri for prevention of aflatoxinogen fungal growth are recommended because of strong antifungal activities.

BackgroundDue to the increasing resistance of pathogenic bacteria to common antibiotics, researchers are seeking alternative antimicrobial agents with plant origins.ObjectivesThe purpose of this study was to evaluate the activity of essential oil, extracted from the leaves of Eucalyptus camaldulensis, the major Eucalyptus species cultivated in Khuzestan, South of Iran, against the growth of drug-resistant bacteria.MethodsEssential oil was extracted from the leaves using the hydrodistillation method in a Clevenger apparatus. The constituents of the essential oil were determined by gas chromatography-mass spectrometry (GC-MS). Moreover, the antimicrobial activity of essential oil was assayed using the disk diffusion method. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were also determined using the macrodilution method.ResultsIsolation and identification of the main components of essential oil identified 1,8-cineole (55.2%) as the main component. The essential oil could control resistant pathogenic bacteria. The greatest effect of essential oil was reported against Klebsiella pneumoniae with an inhibition zone diameter of 35 mm and MIC and MBC of 500 and 1500 ppm, respectively. On the other hand, the lowest effect was reported against Salmonella infantis and Salmonella enteritidis with an inhibition zone diameter of 11 mm and MIC and MBC of 6,000 and 8,000 ppm, respectively.ConclusionsThe essential oil of E. camaldulensis (Myrtaceae family) grown in Iran exhibited significant activities against some Gram-positive and Gram-negative bacteria. Therefore, E. camaldulensis is an effective antibacterial and bactericidal agent in the treatment of infectious diseases

Cuminum cyminum and Rosmarinus officinalis were hydrodistilled and the essential oils obtained thereof were characterized by means of GC and GC–MS. C. cyminum and R. officinalis contained α-pinene (29.1%, 14.94%), 1,8-cineole (17.9%, 7.43%) and linalool (10.4%, 14.86%), respectively as the major compounds. C. cyminum oil exhibited stronger antimicrobial activity than did R. officinalis oil against E. coli, S. aureus and L. monocytogenes. Complete death time on exposure to C. cyminum and R. officinalis oils were 20 and 25 min, 180 and 240 min and 90 and 120 min for E. coli, S. aureus and L. monocytogenes, respectively. Radical-scavenging and antioxidant properties were tested by means of 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay and β-carotene bleaching test. These properties were compared to those of Thymus x-porlock essential oil, used as a reference ingredient. The radicalscavenging performance of the rosemary oil was better than that of C. cyminum. Results from antioxidant test were better than those provided by the radicalscavenging activity. C. cyminum and R. officinalis essential oils may be considered as potent agents in food preservation. INTRODUCTION The exploration of naturally occurring antimicrobials for food preservation receives increasing attention due to consumer awareness of natural food products and a growing concern of microbial resistance towards conventional preservatives (Gould, 1995). Researchers have been interested in biologically active compounds isolated from plant species for the elimination of pathogenic microorganisms because of the resistance that microorganisms have built against antibiotics (Essawi and Srour, 2000). To prolong the shelf-life of fruits and vegetables, the growth of microbial populations must be controlled and several post-harvest processes, such as washing and removal of damaged tissues, are employed to reduce initial high counts. It is well known that clean sanitation is essential in keeping the microbial population to a minimum, because storage life is shorter with high initial microbial loads (Bolin et al., 1977). Nychas (1995) reported antimicrobial activity of essential oils from oregano, thyme, sage, rosemary, clove, coriander, garlic and onion against both bacteria and fungi. Phenolic components, present in essential oils, have been known to possess antimicrobial activity and some are classified as “Generally Recognized as Safe” (GRAS) substances and therefore could be used to prevent post-harvest growth of native and contaminant bacteria (Kabara, 1991; Singh et al., 2001). Oxidation of lipids, which occurs during raw material storage, processing, heat treatment and further storage of final products, is one of the basic processes causing rancidity of food products, leading to their deterioration. Due to undesirable influences of oxidized lipids on the human organism, it seems to be essential to decrease contact with products of lipid oxidation in food (Karpinska et al., 2001). In order to prolong the storage stability of foods, synthetic antioxidants are used for industrial processing. But, according to toxicologists and nutritionists, the side effects of some synthetic antioxidants used in food processing such as, butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA), have already been documented. For 125 Proc. Int. WS on MAP Eds.: N. Chomchalow and V. Chantrasmi Acta Hort. 786, ISHS 2008 example, these substances can show carcinogenic effects in living organisms (Ames, 1983; Baardseth, 1989). From this point of view, governmental authorities and consumers are concerned about the safety of their food and about the potential effects of synthetic additives on health (Reische et al., 1998). The present study deals with the chemical composition, antibacterial, antioxidative and radical scavenging properties of the essential oils of Cuminum cyminum L. and Rosmarinus officinalis L. obtained by steam distillation. The rationale for this work was to test the possibility of creating a protective atmosphere by using natural compounds that could extend the shelf life of foodstuffs while minimizing organoleptic alterations. MATERIALS AND METHODS General The major equipment used were Clevenger, GC (9-A-Shimadzu), GC/MS (Varian3400), Microbial culture media (Merck), Shimadzu UV-2501PC spectrophotometer. Other chemicals were of analytical grade. Plant Materials The plants (Cuminum cyminum L. and Rosmarinus officinalis L.) were collected from National Botanical Garden of Iran during May-June 2005. Oil Extraction and Analysis The plant materials were steam distilled for 90 min in full glass apparatus. The oils were isolated using a Clevenger-type apparatus. The extraction was carried out for two hours after four hours maceration in 500 ml of water. The oils were stored in dark glass bottles in a freezer until they were used. Oil dilutions of 1/2, 1/4 and 1/8 were made with dimethylsulphoxide (DMSO). These dilutions were used in antibacterial analysis. Undiluted oil was taken as dilution 1. GC analysis was performed by GC (9-A-Shimadzu) gas chromatograph equipped with a flame ionization detector. Quantitation was carried out on Euro Chrom 2000 from KNAUER by area normalization method. The analysis was carried out using a DB-5 fused-silica column (30 m×0.25 mm, film thickness 0.25 μm) using a temperature program of 40–250C at a rate of 4C/min, injector temperature 250C, detector temperature 265C, carrier gas: helium (99.99%). The GC/MS unit consisted of Varian-3400 gas chromatograph coupled to a Saturn II ion trap detector. The column was same as of the GC under the same conditions stated above. The constituents were identified by comparison of their mass spectra with those in the computer library and with authentic compounds. The identifications were confirmed by comparison of their retention indices with those of authentic compounds or with literature data. Oil Dilution Solvent Bacterial strains were streaked on Mueller Hinton agar plates using sterile cotton swabs. Sterile blank disks loaded with 5μl of dimethylsulphoxide (DMSO) were placed on the agar plates and were incubated at 37C for 24 h. There was no antibacterial activity on the plates and hence DMSO was selected as a safe diluting agent for the oil. A volume of 5μl from each oil dilution followed by sterilization using a 0.45 μm membrane filter was added to sterile blank discs. The solvent also served as control. Microbial Strain and Growth Media E. coli (ATCC 25922), S. aureus (ATCC 25923) and L. monocytogenes (PTCC 1298) were employed in the study. Nutrient agar was used to maintain E. coli and S. aureus. Blood agar was used to maintain L. monocytogenes. Bacterial suspensions were made in Brain Heart Infusion (BHI) broth to a concentration of approximately 10 cfu/ml. Subsequent dilutions were made from the above suspension, which were then used in the tests.

Aflatoxin B1 (AFB1) is a highly toxic and carcinogenic metabolite produced by Aspergillus species on food and agricultural commodities. Natural products may regulate the cellular effects of aflatoxins and evidence suggests that aromatic organic compounds of spices can control the production of aflatoxins. With a view to controlling aflatoxin production, the essential oils from Rosmarinus officinalis and Trachyspermum copticum L. were obtained by hydrodistillation. Antifungal activities of the oils were studied with special reference to the inhibition of Aspergillus parasiticus growth and aflatoxin production. Minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oils were determined. T. copticum L. oil showed a stronger inhibitory effect than R. officinalis on the growth of A. parasiticus. Aflatoxin production was inhibited at 450 ppm of both oils with that of R. officinalis being stronger inhibitor. The oils were analyzed by GC and GC/MS. The major components of R. officinalis and T. copticum L. oils were Piperitone (23.65%), alpha-pinene (14.94%), Limonene (14.89%), 1,8-Cineole (7.43%) and Thymol (37.2%), P-Cymene (32.3%), gamma-Terpinene (27.3%) respectively. It is concluded that the essential oils could be safely used as preservative materials on some kinds of foods to protect them from toxigenic fungal infections.

Background: Aspergillus species produce the highly toxic and carcinogenic metabolite, Aflatoxin B1 (AFB1), on food and agricultural commodities. Some natural products are known to inhibit aflatoxin production. Objectives: With the aim of controlling aflatoxin production, the essential oils of Cuminum cyminum L. from the best known regions of Iran i.e. Alborz Mountain and Kerman region, were obtained by hydrodistillation. Materials and Methods: Antifungal activities of the oils to inhibit growth and aflatoxin productivity of A. flavus PICC-AF39, A. flavus PICC-AF24, and A. parasiticus NRRL-2999 were studied. Minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oil were determined. Sub-MIC was selected for the measurement of aflatoxins B and G concentration. Samples were analyzed either using a high performance liquid chromatography (HPLC) method with some minor modifications. Aflatoxins (AFs) were determined by reverse-phase HPLC and fluorescence detector with post column derivatization (PCD) involving bromination. Results: A significant reduction in Aflatoxin production was noted which was not due to the inhibitory effect but because of antifungal property of the oil. Interestingly, the oil promoted toxin production for the reasons yet to be investigated. The extent of aflatoxin production was dependent on the concentration of essential oil used. All toxin-producing fungi in this study produced higher amount of aflatoxin at low concentrations of the oil. 400 ppm concentration of C. cyminum L. from Alborz Mountain increased aflatoxin production to over fourfold. Aflatoxin productivity was declined at high concentration of the oil. Conclusions: Antimicrobial and antitoxigenic properties of natural products need a firmly established criterion before they could be offered to application.

Background: Aflatoxin B 1 (AFB 1 ) is a highly toxic and hepatocarcinogenic metabolite produced by Aspergillus species. Some natural products are known to kill fungi and destroy toxins and toxin-producing agents. Objectives: The purpose of this study is to provide experimental data on the antifungal activity of cumin oils and their components that could be considered suitable for application in foods and drugs. Materials and Methods: The essential oil (EO) of Cuminum cyminum L. collected from Alborz Mountain, Iran, was obtained by hydro-distillation. The oil was analyzed by gas chromatography (GC) and chromatography/mass spectrophotometry (GC/MS). The antifungal activity of the oil was studied with regard to the inhibition of the growth of Aspergillus flavus PICC-AF39 , Aspergillus flavus PICC-AF24, Aspergillus parasiticus NRRL-2999 and Aspergillus niger. The minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oil were determined. Results: α–Pinene (29.2%), limonene (21.7%), 1,8-cineole (18.1%), linalool (10.5%), linalyl acetate (4.8%), and α-terpineole (3.17%) were the major components of the essential oil from C. cyminum L., and the oil showed a strong inhibitory effect on fungal growth. Conclusions: Essential oils could be safely used as preservatives in pharmaceuticals as well as health and food products to protect them against toxigenic fungal infections.