Cultures ofP Research Articles

Green synthesis, characterization, and evaluation of antibacterial activities of cobalt nanoparticles produced by marine fungal speciesPericonia prolifica

AbstractBio-nanotechnology provided an ecofriendly synthesis route for various metal nanoparticles by utilizing different biological systems, especially microorganisms, which act as an alternative to the physical and chemical methods. Cobalt nanoparticles (CoNPs) were synthesized byPericonia prolifica(Anast.) from intertidal decayed wood samples from the mangrove treeAvicennia marina(Forsk.) of Tarout Island, located in the Arabian Gulf Sea of Saudi Arabia. CoNPs were characterized by X-ray diffraction, transmission electron microscopy, Fourier-transform infrared spectroscopy, and atomic force microscope. The extract of the culture ofP. prolificawas used as a bio-reductant agent, during which the culturing process proved to have great potential to be applied on an industrial scale, as it was a time-saving, inexpensive, and adequate amount of biomass being produced at the end of the process. A preliminary antibacterial test against one Gram-positive resistant bacteria (i.e., Methicillin-resistantStaphylococcus aureus) and other Gram-negative resistant bacteria was performed using a disk diffusion assay. The antibacterial results witnessed the key role that metal size plays in causing higher activity and also in causing severe damage to the bacterial cells by inactivating its membrane permeability, leading to bacterial cell death.

Cryopreservation OfPyramimonas Mucifera

BACKGROUND:It is important to appreciate microalgal diversity, better understand their ecosystem functioning and therefore implement conservation measures. The National Biodiversity Act of South Africa has a marine and coastal component which promotes such investigations.OBJECTIVE:To develop a cryostorage method for the marine unicellular algal speciesPyramimonas mucifera.MATERIALS AND METHODS:Cell viability, measured by propidium iodide, was used to determine both optimal exposure time to 10 % DMSO and survival following thawing of cryopreserved cells. Cryopreservation was achieved by a two-step cooling method.RESULTS & DISCUSSION:A 30-min DMSO exposure was selected forP. mucifera, as cells following such treatment retained cell shape and integrity. Although density was significantly reduced after cryopreservation, the surviving cells were capable of returning to viability levels equal to those of the untreated control (>90%).CONCLUSION:Cultures ofP. muciferacan be successfully cryopreserved and propidium iodide provides a useful indication of culture vitality.

Biofilm inhibition and pathogenicity attenuation in bacteria by Proteus mirabilis.

Biofilms play an important role in the antibiotic resistance of encased bacteria, and biofilm formation is regulated by quorum sensing (QS). Inhibiting the QS system may, therefore, degrade the integrity of a biofilm and expose the bacterial pathogens within it to the deleterious effects of molecules such as antibiotics. Moreover, the use of QS inhibitors (QSIs) may provide a novel approach for treating bacterial infections of aquacultures. In the present study, the bacterium Proteus mirabilis was identified as a potential producer of QSIs. Varying concentrations (0.1–1.1%) of filtrates prepared from the culture of P. mirabilis inhibited biofilm formation by the pathogens Pseudomonas aeruginosa, Vibrio harveyi and Staphylococcus aureus by as much as 58.9%, 41.5% and 41.9%, respectively. These filtrates as well as the crude aqueous extracts prepared from them increased the sensitivities of pathogens to the inhibitory effects of kanamycin. The filtrates also showed pathogenicity attenuation potential in P. aeruginosa by decreasing the production of virulence factors. Moreover, the filtrates did not influence the planktonic growth of these pathogens. The results indicate that P. mirabilis may act as a non-specific (or broad-spectrum) inhibitor of biofilm formation that will help control infectious diseases that adversely affect the aquaculture industry.

Antibacterial Activity ofMiswak(SalvadorapersicaL.)Stem Aqueous and Ethanolic Extracts in vitro

The aim was to evaluate the in vitro antibacterial activity of three concentrations (50, 75 and 100 μg/ml) of aqueous and ethanoliccrudeextractsofmiswak(Salvadorapersica) on threepathogenic bacteria (Staphylococcus aureus, Streptococcus pyogenesand Pseudomonas aeruginosa)using agar well-diffusion method.Phytochemical analysis revealed that the extracts contained alkaloids, flavonoids, phenols, tannins and steroids. Antibacterial evaluations demonstrated that both extracts inhibited the growth of the investigated bacteria in a concentration-dependent manner, but the aqueous extract was more efficient than the ethanolicextract.Thehighest growth-inhibition (GI) zone(22.1± 2.3mm)was observed in the culture ofP. aeruginosaat the concentration 100 μg/mlof the aqueous extract, while the lowest (12.3± 1.2 mm) was inthe culture of S.aureus using the ethanolic extract at the concentration 50 μg/ml. These findings suggest the antibacterial potential of Salvadorapersica stem, which might be related to its chemical constituents..

An Antipersister Strategy for Treatment of Chronic Pseudomonas aeruginosa Infections.

Bacterial persisters are a quasidormant subpopulation of cells that are tolerant to antibiotic treatment. The combination of the aminoglycoside tobramycin with fumarate as an antibacterial potentiator utilizes an antipersister strategy that is aimed at reducing recurrent Pseudomonas aeruginosa infections by enhancing the killing of P. aeruginosa persisters. Stationary-phase cultures of P. aeruginosa were used to generate persister cells. A range of tobramycin concentrations was tested with a range of metabolite concentrations to determine the potentiation effect of the metabolite under a variety of conditions, including a range of pH values and in the presence of azithromycin or cystic fibrosis (CF) patient sputum. In addition, 96-well dish biofilm and colony biofilm assays were performed, and the cytotoxicity of the tobramycin-fumarate combination was determined utilizing a lactate dehydrogenase (LDH) assay. Enhanced killing of up to 6 orders of magnitude of P. aeruginosa persisters over a range of CF isolates, including mucoid and nonmucoid strains, was observed for the tobramycin-fumarate combination compared to killing with tobramycin alone. Furthermore, significant fumarate-mediated potentiation was seen in the presence of azithromycin or CF patient sputum. Fumarate also reduced the cytotoxicity of tobramycin-treated P. aeruginosa to human epithelial airway cells. Finally, in mucoid and nonmucoid CF isolates, complete eradication of P. aeruginosa biofilm was observed in the colony biofilm assay due to fumarate potentiation. These data suggest that a combination of tobramycin with fumarate as an antibacterial potentiator may be an attractive therapeutic for eliminating recurrent P. aeruginosa infections in CF patients through the eradication of bacterial persisters.

Propionibacterium acnes Recovered from Atherosclerotic Human Carotid Arteries Undergoes Biofilm Dispersion and Releases Lipolytic and Proteolytic Enzymes in Response to Norepinephrine Challenge In Vitro.

In the present study, human atherosclerotic carotid arteries were examined following endarterectomy for the presence of the Gram-positive bacterium Propionibacterium acnes and its potential association with biofilm structures within the arterial wall. The P. acnes 16S rRNA gene was detectable in 4 of 15 carotid artery samples, and viable P. acnes was one among 10 different bacterial species recoverable in culture. Fluorescence in situ hybridization analysis of 5 additional atherosclerotic carotid arteries demonstrated biofilm bacteria within all samples, with P. acnes detectable in 4 samples. We also demonstrated that laboratory-grown cultures of P. acnes biofilms were susceptible to induction of a biofilm dispersion response when challenged with physiologically relevant levels of norepinephrine in the presence of iron-bound transferrin or with free iron. The production and release of lipolytic and proteolytic extracellular enzymes by P. acnes were shown to increase in iron-induced dispersed biofilms, and these dispersion-induced P. acnes VP1 biofilms showed increased expression of mRNAs for the triacylglycerol lipases PPA2105 and PPA1796 and the hyaluronate lyase PPA380 compared to that in untreated biofilms. These results demonstrate that P. acnes can infect the carotid arteries of humans with atherosclerosis as a component of multispecies biofilms and that dispersion is inducible for this organism, at least in vitro, with physiologically relevant levels of norepinephrine resulting in the production and release of degradative enzymes.

In vivo antimalarial activity and mechanisms of action of 4-nerolidylcatechol derivatives.

4-Nerolidylcatechol (1) is an abundant antiplasmodial metabolite that is isolated from Piper peltatum roots. O-Acylation or O-alkylation of compound 1 provides derivatives exhibiting improved stability and significant in vitro antiplasmodial activity. The aim of this work was to study the in vitro inhibition of hemozoin formation, inhibition of isoprenoid biosynthesis in Plasmodium falciparum cultures, and in vivo antimalarial activity of several 4-nerolidylcatechol derivatives. 1,2-O,O-Diacetyl-4-nerolidylcatechol (2) inhibited in vitro hemozoin formation by up to 50%. In metabolic labeling studies using [1-(n)-(3)H]geranylgeranyl pyrophosphate, diester 2: significantly inhibited the biosynthesis of isoprenoid metabolites ubiquinone 8, menaquinone 4, and dolichol 12 in cultures of P. falciparum 3D7. Similarly, 2-O-benzyl-4-nerolidylcatechol (3) significantly inhibited the biosynthesis of dolichol 12. P. falciparum in vitro protein synthesis was not affected by compounds 2 or 3. At oral doses of 50 mg per kg of body weight per day, compound 2 suppressed Plasmodium berghei NK65 in infected BALB/c mice by 44%. This in vivo result for derivative 2 represents marked improvement over that obtained previously for natural product 1. Compound 2 was not detected in mouse blood 1 h after oral ingestion or in mixtures with mouse blood/blood plasma in vitro. However, it was detected after in vitro contact with human blood or blood plasma. Derivatives of 4-nerolidylcatechol exhibit parasite-specific modes of action, such as inhibition of isoprenoid biosynthesis and inhibition of hemozoin formation, and they therefore merit further investigation for their antimalarial potential.

Studies on Molecular Characterizations of the Outer Membrane Proteins, Lipids Profile, and Exopolysaccharides of Antibiotic Resistant StrainPseudomonas aeruginosa

Susceptibility of the tested Pseudomonas aeruginosa strain to two different antibiotics, tetracycline (TE) and ciprofloxacin (CIP), was carried out using liquid dilution method. Minimum inhibitory concentrations of TE and CIP were 9.0 and 6.0 mg/100 mL, respectively. Some metabolic changes due to both, the mode of action of TE and CIP on P. aeruginosa and its resistance to high concentrations of antibiotics (sub-MIC) were detected. The total cellular protein contents decreased after antibiotic treatment, while outer membrane protein (OMP) contents were approximately constant for both treated and untreated cells. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the OMPs for untreated and TE and CIP treated cells indicated that the molecular changes were achieved as; lost in, induction and stability of some protein bands as a result of antibiotics treatment. Five bands (with mol. wt. 71.75, 54.8, 31.72, 28.63, and 20.33 KDa) were stable in both treated and untreated tested strains, while two bands (with mol. wt. 194.8 and 118.3 KDa) were induced and the lost of only one band (with mol. wt. 142.5 KDa) after antibiotics treatment. On the other hand, total lipids and phospholipids increased in antibiotic treated cells, while neutral lipids decreased. Also, there was observable stability in the number of fatty acids in untreated and treated cells (11 fatty acids). The unsaturation index was decreased to 56% and 17.6% in both TE and CIP treatments, respectively. The produced amount of EPSs in untreated cultures of P. aeruginosa was relatively higher than in treated cultures with sub-MICs of TE and CIP antibiotics. It was also observed that the amounts of exopolysaccharides (EPSs) increased by increasing the incubation period up to five days of incubation in case of untreated and antibiotic treated cultures.

Evaluation of different methods to preventPenicillium nordicum growth on and ochratoxin A production in country-style sausages

Penicillium nordicum is the main ochratoxin A (OTA)-producing fungal species isolated from artisanal sausages of different areas of Italy and other European countries.P. nordicum grows on and produces OTA in the sausage casing, thereby posing a health risk to Italian consumers, who commonly eat both the meat and casing of artisanal sausages. Different methods have been proposed to eliminate OTA andP. nordicum on the casings, including brushing, brushing and washing, the use of fungal starter cultures (Penicillium nalgiovense) and gaseous ozone. The aim of this study was to compare these methods with regard to eliminating the risk of OTA contamination in sausages. Three concentrations of gaseous ozone were assessed, i.e. 0.5, 1.0 and 1.5 mg/l. Only 1.5 mg/l preventedP. nordicum growth and production of OTA in sausages without influencing the physicochemical parameters and the sensory characteristic of the products. Application of this gaseous ozone concentration was then compared to the other methods, i.e. brushing, brushing and washing, a one-step starter culture ofP. nalgiovense (inoculation after casing) and a two-step starter culture (inoculation after both casing and drying). The results obtained showed that only the use of gaseous ozone and a two-step starter culture ofP. nalgiovense achieved complete inhibition ofP. nordicum growth and OTA production. However, sensory analysis of the treated sausages showed that the use of starter cultures was the best method.

The Putative Enoyl-Coenzyme A Hydratase DspI Is Required for Production of the Pseudomonas aeruginosa Biofilm Dispersion Autoinducer cis -2-Decenoic Acid

In the present study, we report the identification of a putative enoyl-coenzyme A (CoA) hydratase/isomerase that is required for synthesis of the biofilm dispersion autoinducer cis-2-decenoic acid in the human pathogen Pseudomonas aeruginosa. The protein is encoded by PA14_54640 (PA0745), named dspI for dispersion inducer. The gene sequence for this protein shows significant homology to RpfF in Xanthomonas campestris. Inactivation of dspI was shown to abolish biofilm dispersion autoinduction in continuous cultures of P. aeruginosa and resulted in biofilms that were significantly greater in thickness and biomass than those of the parental wild-type strain. Dispersion was shown to be inducible in dspI mutants by the exogenous addition of synthetic cis-2-decenoic acid or by complementation of ΔdspI in trans under the control of an arabinose-inducible promoter. Mutation of dspI was also shown to abolish cis-2-decenoic acid production, as revealed by gas chromatography-mass spectrometry (GC-MS) analysis of cell-free spent culture medium. The transcript abundance of dspI correlated with cell density, as determined by quantitative reverse transcriptase (RT) PCR. This regulation is consistent with the characterization of cis-2-decenoic acid as a cell-to-cell communication molecule that regulates biofilm dispersion in a cell density-dependent manner.

Nematodes from the Victoria Land coast, Antarctica and comparisons with culturedPanagrolaimus davidi

AbstractMorphological and genetic information was used to determine the species identity and relationships of four species of nematodes from the Victoria Land coast, Antarctica; and between laboratory and field populations ofPanagrolaimus davidiTimm. The 18S and D3 expansion segments of 28S rRNA were successfully amplified from the four species:Scottnema lindsayaeTimm,Plectus murrayiYeates,Eudorylaimusspp. andPanagrolaimus davidi. The four species from field populations mapped to three different nematode clades, indicating that if they are survivors in, rather than colonizers of, Antarctica they originate from several Gondwanan ancestors. The laboratory culture ofP. davidiis shown to be a different species to that of the field samples ofP. davidi, which were genetically identical to each other. There are several possible explanations for this observation, which require further investigation.

Camphor Pathway Redux: Functional Recombinant Expression of 2,5- and 3,6-Diketocamphane Monooxygenases of Pseudomonas putida ATCC 17453 with Their Cognate Flavin Reductase Catalyzing Baeyer-Villiger Reactions

Whereas the biochemical properties of the monooxygenase components that catalyze the oxidation of 2,5-diketocamphane and 3,6-diketocamphane (2,5-DKCMO and 3,6-DKCMO, respectively) in the initial catabolic steps of (+) and (-) isomeric forms of camphor (CAM) metabolism in Pseudomonas putida ATCC 17453 are relatively well characterized, the actual identity of the flavin reductase (Fred) component that provides the reduced flavin to the oxygenases has hitherto been ill defined. In this study, a 37-kDa Fred was purified from a camphor-induced culture of P. putida ATCC 17453 and this facilitated cloning and characterization of the requisite protein. The active Fred is a homodimer with a subunit molecular weight of 18,000 that uses NADH as an electron donor (Km = 32 μM), and it catalyzes the reduction of flavin mononucleotide (FMN) (Km = 3.6 μM; kcat = 283 s(-1)) in preference to flavin adenine dinucleotide (FAD) (Km = 19 μM; kcat = 128 s(-1)). Sequence determination of ∼40 kb of the CAM degradation plasmid revealed the locations of two isofunctional 2,5-DKCMO genes (camE25-1 for 2,5-DKCMO-1 and camE25-2 for 2,5-DKCMO-2) as well as that of a 3,6-DKCMO-encoding gene (camE36). In addition, by pulsed-field gel electrophoresis, the CAM plasmid was established to be linear and ∼533 kb in length. To enable functional assessment of the two-component monooxygenase system in Baeyer-Villiger oxidations, recombinant plasmids expressing Fred in tandem with the respective 2,5-DKCMO- and 3,6-DKCMO-encoding genes in Escherichia coli were constructed. Comparative substrate profiling of the isofunctional 2,5-DCKMOs did not yield obvious differences in Baeyer-Villiger biooxidations, but they are distinct from 3,6-DKCMO in the stereoselective oxygenations with various mono- and bicyclic ketone substrates.

Bioremediation of Direct Blue 14 and Extracellular Ligninolytic Enzyme Production by White Rot Fungi:PleurotusSpp.

In the present investigation, four species of white rot fungi (Pleurotus), that is, P. flabellatus, P. florida, P. ostreatus and P. sajor-caju were used for decolorization of direct blue 14 (DB14). Among all four species of Pleurotus, P. flabellatus showed the fastest decolorization in petri plates on different concentration, that is, 200 mg/L, 400 mg/L, and 600 mg/L. All these four species were also evaluated for extracellular ligninolytic enzymes (laccase and manganese peroxidase) production and it was observed that the twelve days old culture of P. flabellatus showed the maximum enzymatic activity, that is, 915.7 U/mL and 769.2 U/mL of laccase and manganese peroxidase, respectively. Other three Pleurotus species took more time for dye decolorization and exhibited less enzymatic activities. The rate of decolorization of DB14 dye solution (20 mg/L) by crude enzymes isolated from P. flabellatus was very fast, and it was observed that up to 90.39% dye solution was decolorized in 6 hrs of incubation.

Bioremediation of Parboiled Rice Effluent Supplemented with Biodiesel-Derived Glycerol UsingPichia pastorisX-33

This paper describes the use of Pichia pastoris X-33 as a bioremediator to reduce the chemical oxygen demand (COD), total Kjeldahl nitrogen (TKN), and phosphorus (P-PO4 3−), after culture in parboiled rice effluent supplemented with p.a. glycerol or a glycerol by-product of the biodiesel industry. The greatest reduction in the COD (55%), TKN (45%), and P-PO4 3− (52%) of the effluent was observed in cultures of P. pastoris X-33 supplemented with 15 g ·L−1 of biodiesel-derived glycerol. Furthermore, the overall biomass yield was 2.1 g ·L−1. These data suggest that biodiesel-derived glycerol is an efficient carbon source for the bioremediation of parboiled rice effluent and biomass production.

Life history and morphological studies ofPunctaria tenuissima(Chordariaceae, Phaeophyceae), a new record for the Azores

AbstractPunctaria tenuissima(Chordariaceae, Phaeophyceae) is reported for the first time from the Azores. Erect thalli were collected on the Island of São Miguel; they were up to 3 cm long, flattened and often twisted. The plurilocular sporangia were formed from surface cells that were quadrate or rectangular in surface view. Unilocular sporangia were not observed in the field. In culture, the plurispores ofP. tenuissimadeveloped intoHecatonema-like, tufted, prostrate thalli that formed plurilocular sporangia. The plurispores of the prostrate thalli cultured at 15°C with a long day (LD) photoperiod developed into new prostrate thalli, which in turn formed plurilocular sporangia. This cycle was repeated 5 times, resulting in several generations of reproductive prostrate thalli. When these cultures were cooled to 10°C with a short day (SD) photoperiod, new erect thalli developed from the prostrate thalli resembling the thalli collected in the field. These erect blades produced unilocular and plurilocular sporangia on the same or on different thalli. The plurispores and unispores produced by the erect thalli at 10°C under SD conditions once again developed into new prostrate thalli. This is the first report of unilocular sporangia formed in cultures ofP. tenuissima. Sexual reproduction was not observed. The culture conditions, particularly temperature and daylength, appeared to influence the formation of erect blades. ThePunctaria-like thalli were produced in 10°C/SD conditions, whilst theHecatonema-like thalli were produced under 10°–15°C/LD and SD conditions. These results are similar to those reported forP. tenuissimafrom other locations and suggest the occurrence of both direct and heteromorphic life histories in the Azorean algae.

Potential of Oyster Mushrooms for the Biocontrol of Sugar Beet Nematode (Heterodera Schachtii)

Potential of Oyster Mushrooms for the Biocontrol of Sugar Beet Nematode (Heterodera Schachtii)The potential of oyster mushrooms to attack and kill sugar beet cyst nematode (Heterodera schachtii) was studied. The ability ofPleurotus ostreatus, P. sajor-caju, P. florida, P. flabellatus, P. ostreatus(sporeless),P. eryngiiandHypsizygus ulmariusto prey on the cyst nematode was confirmed. Nematodes were inoculated on water agar plates on which a single sparse fungal colony of one of the above mentioned fungi was grown. Nematodes were quickly immobilized after inoculation the plates with eachPleurotusspecies, with mortality of about 96%, 72%, 55%, 52%, 20%, 23% and 62%, respectively. Nematicidal activities of culture filtrates ofPleurotusspecies andH. ulmariusagainstH. schachtiiwere also studiedin vitro.Extracts ofP. ostreatuscould paralyse 90% of nematodes whereasP. eryngiicould paralyse only 50% of nematodes. The efficacy of old mushroom compost from cultures ofP. ostreatusandP. sajor-cajuin controlling cysts on sugar beet (Beta vulgarisICI) were studied under greenhouse conditions. The results showed that 100 and 200 grams of mushroom compost per 3 kilogram/pot could significantly control cysts, reducing more than 85% cysts in soil. Presence of cysts on the roots and larvae inside roots were compared with the control treatment.

Quantification of 2,4-Diacetylphloroglucinol-Producing Pseudomonas fluorescens Strains in the Plant Rhizosphere by Real-Time PCR

A real-time PCR SYBR green assay was developed to quantify populations of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing (phlD+) strains of Pseudomonas fluorescens in soil and the rhizosphere. Primers were designed and PCR conditions were optimized to specifically amplify the phlD gene from four different genotypes of phlD+ P. fluorescens. Using purified genomic DNA and genomic DNA extracted from washes of wheat roots spiked with bacteria, standard curves relating the threshold cycles (C(T)s) and copies of the phlD gene were generated for P. fluorescens strains belonging to genotypes A (Pf-5), B (Q2-87), D (Q8r1-96 and FTAD1R34), and I (FTAD1R36). The detection limits of the optimized real-time PCR assay were 60 to 600 fg (8 to 80 CFU) for genomic DNA isolated from pure cultures of P. fluorescens and 600 fg to 6.0 pg (80 to 800 CFU, corresponding to log 4 to 5 phlD+ strain CFU/rhizosphere) for bacterial DNA extracted from plant root washes. The real-time PCR assay was utilized to quantify phlD+ pseudomonads in the wheat rhizosphere. Regression analysis of population densities detected by real-time PCR and by a previously described phlD-specific PCR-based dilution endpoint assay indicated a significant linear relationship (P = 0.0016, r2 = 0.2). Validation of real-time PCR assays with environmental samples was performed with two different soils and demonstrated the detection of more than one genotype in Quincy take-all decline soil. The greatest advantage of the developed real-time PCR is culture independence, which allows determination of population densities and the genotype composition of 2,4-DAPG producers directly from the plant rhizospheres and soil.

Antiplasmodial Activity of Lauryl-Lysine Oligomers

The ever evolving resistance of the most virulent malaria parasite, Plasmodium falciparum, to antimalarials necessitates the continuous development of new drugs. Our previous analysis of the antimalarial activities of the hemolytic antimicrobial peptides dermaseptins and their acylated derivatives implicated the importance of hydrophobicity and charge for drug action. Following these findings, an oligoacyllysine (OAK) tetramer designed to mimic the characteristics of dermaseptin was synthesized and assessed for its antimalarial activity in cultures of P. falciparum. The tetramer inhibited the growth of different plasmodial strains at low micromolar concentrations (mean 50% inhibitory concentration [IC(50)], 1.8 microM). A structure-activity relationship study involving eight derivatives unraveled smaller, more potent OAK analogs (IC(50)s, 0.08 to 0.14 microM). The most potent analogs were the most selective, with selectivity ratios of 3 orders of magnitude. Selectivity was strongly influenced by the self-assembly properties resulting from interactions between hydrophobic OAKs, as has been observed with conventional antimicrobial peptides. Further investigations performed with a representative OAK revealed that the ring and trophozoite stages of the parasite developmental cycle were equally sensitive to the compound. A shortcoming of the tested compound was the need for long incubation times in order for it to exert its full effect. Nevertheless, the encouraging results obtained in this study regarding the efficiency and selectivity of some compounds establish them as leads for further development.

A Novel ChimericPlasmodium vivaxCircumsporozoite Protein Induces Biologically Functional Antibodies That Recognize both VK210 and VK247 Sporozoites

A successful vaccine against Plasmodium vivax malaria would significantly improve the health and quality of the lives of more than 1 billion people around the world. A subunit vaccine is the only option in the absence of long-term culture of P. vivax parasites. The circumsporozoite protein that covers the surface of Plasmodium sporozoites is one of the best-studied malarial antigens and the most promising vaccine in clinical trials. We report here the development of a novel "immunologically optimal" recombinant vaccine expressed in Escherichia coli that encodes a chimeric CS protein encompassing repeats from the two major alleles, VK210 and VK247. This molecule is widely recognized by sera from patients naturally exposed to P. vivax infection and induces a highly potent immune response in genetically disparate strains of mice. Antibodies from immunized animals recognize both VK210 and VK247 sporozoites. Furthermore, these antibodies appear to be protective in nature since they cause the agglutination of live sporozoites, an in vitro surrogate of sporozoite infectivity. These results strongly suggest that recombinant CS is biologically active and highly immunogenic across major histocompatibility complex strains and raises the prospect that in humans this vaccine may induce protective immune responses.

Enhancement of Antimicrobial Activity against Pseudomonas aeruginosa by Coadministration of G10KHc and Tobramycin

Pseudomonas aeruginosa is a common opportunistic human pathogen that is associated with life-threatening acute infections and chronic airway colonization during cystic fibrosis. Previously, we converted the wide-spectrum antimicrobial peptide novispirin G10 into a selectively-targeted antimicrobial peptide (STAMP), G10KHc. Compared to novispirin G10, the STAMP had an enhanced ability to kill Pseudomonas mendocina. In this study, we explored the activity of G10KHc against P. aeruginosa. G10KHc was found to be highly active (as active as tobramycin) against P. aeruginosa clinical isolates. Most interestingly, we observed a synergistic-like enhancement in killing activity when biofilms and planktonic cultures of P. aeruginosa were cotreated with G10KHc and tobramycin. The data indicate that the mechanism of enhanced activity may involve increased tobramycin uptake due to G10KHc-mediated cell membrane disruption. These results suggest that G10KHc may be useful against P. aeruginosa during acute and chronic infection states, especially when it is coadministered with tobramycin.