Link between Domoic Acid Production and Cell Physiology after Exchange of Bacterial Communities between Toxic Pseudo-nitzschia multiseries and Non-Toxic Pseudo-nitzschia delicatissima

Bacteria are known to influence domoic acid (DA) production by Pseudo-nitzschia spp., but the link between DA production and physiology of diatoms requires more investigation. We compared a toxic P. multiseries to a non-toxic P. delicatissima, investigating links between DA production, physiological parameters, and co-occurring bacteria. Bacterial communities in cultures of both species were reduced by antibiotic treatment, and each of the diatoms was inoculated with the bacterial community of the other species. The physiology of P. delicatissima was minimally affected by the absence of bacteria or the presence of alien bacteria, and no DA was detected. P. multiseries grew faster without bacteria, did not produce a significant amount of DA, and exhibited physiological characteristics of healthy cells. When grown with alien bacteria, P. multiseries did not grow and produced more DA; the physiology of these cells was affected, with decreases in chlorophyll content and photosynthetic efficiency, an increase in esterase activity, and almost 50% mortality of the cells. The alien bacterial community had morphological and cellular characteristics very different from the original bacteria, and the number of free-living bacteria per algal cell was much higher, suggesting the involvement of bacteria in DA production.     

Dangerous Relations in the Arctic Marine Food Web: Interactions between Toxin Producing Pseudo-nitzschia Diatoms and Calanus Copepodites

Diatoms of the genus Pseudo-nitzschia produce domoic acid (DA), a toxin that is vectored in the marine food web, thus causing serious problems for marine organisms and humans. In spite of this, knowledge of interactions between grazing zooplankton and diatoms is restricted. In this study, we examined the interactions between Calanus copepodites and toxin producing Pseudo-nitzschia. The copepodites were fed with different concentrations of toxic P. seriata and a strain of P. obtusa that previously was tested to be non-toxic. The ingestion rates did not differ among the diets (P. seriata, P. obtusa, a mixture of both species), and they accumulated 6%–16% of ingested DA (up to 420 µg per dry weight copepodite). When P. seriata was exposed to the copepodites, either through physical contact with the grazers or separated by a membrane, the toxicity of P. seriata increased (up to 3300%) suggesting the response to be chemically mediated. The induced response was also triggered when copepodites grazed on another diatom, supporting the hypothesis that the cues originate from the copepodite. Neither pH nor nutrient concentrations explained the induced DA production. Unexpectedly, P. obtusa also produced DA when exposed to grazing copepodites, thus representing the second reported toxic polar diatom.     

Domoic Acid Improves the Competitive Ability of Pseudo-nitzschia delicatissima against the Diatom Skeletonema marinoi

Because domoic acid, a neurotoxic secondary metabolite produced by marine diatoms in the genus Pseudo-nitzschia, is hypothesized to be part of a high affinity iron uptake system, we investigated whether domoic acid could improve the competitive ability of Pseudo-nitzschia delicatissima, and whether the availability of iron changed the outcome of competition experiments. We found that domoic acid had a slight negative effect on growth of the diatom Skeletonema marinoi when it was grown in monocultures. However, when S. marinoi was cultured with P. delicatissima the presence of domoic acid resulted in a reduction of S. marinoi cells by up to 38% and an increase in P. delicatissima cell numbers by up to 17% under iron replete conditions. Similar effects were not observed in low iron treatments. Domoic acid was not taken up by P. delicatissima cells. Overall, our results indicate that domoic acid can improve the competitive ability of Pseudo-nitzschia spp. and that iron is likely to be involved. This study provides an unusual example of indirect inhibition of competitor growth mediated by a secondary metabolite.     

Occurrence and characterization of fungi and oomycetes transmitted via potting mixtures and organic manures

A study was conducted to investigate the most common fungal and oomycete pathogens introduced into farms in Oman via potting mixtures and organic manures. A total of 37 commercial types of potting mixtures (2 local and 35 imported from overseas), 4 commercial types of organic manures and 11 non-commercial types of organic manures were included in the study. Identification of the isolated species was based on morphological characteristics, except for the most common species which were further identified using sequences of the internal transcribed spacer region of the ribosomal DNA (ITS rDNA). Fusarium spp. (14%), Pythium aphanidermatum (3%), Alternaria spp. (5%), Helminthosporium spp. (5%) and Cladosporium spp. (3%) were recovered at different frequencies from samples of potting mixtures. Fusarium solani (40%) and Fusarium equiseti (47%) were recovered at high frequencies from samples of organic manures. Isolations from organic manures also yielded Pythium periplocum (7%), Rhizoctonia solani (7%), Fusarium lichenicola (7%), Helminthosporium spp. (27%) and Alternaria spp. (27%). Trichoderma spp., Penicillium spp., Aspergillus spp. and Rhizopus spp. were found to be common in samples of potting mixtures and organic manures. Investigating sensitivity to hymexazol among 9 isolates of F. equiseti and 13 isolates of F. solani revealed variations among different isolates. The EC₅₀ values ranged from 1 to over 1200 (avg. 192 μg ml⁻¹) for F. equiseti isolates and from 135 to 789 (avg. 324 μg ml⁻¹) for F. solani isolates, indicating presence of resistance to this important fungicide among some Fusarium isolates. This appears to be the first report of contamination with R. solani, P. periplocum, F. solani, F. equiseti and F. lichenicola of organic manures. This study appears to report for the first time F. lichenicola in Oman and appears to be the first report of occurrence of resistance to hymexazol among F. equiseti and F. solani isolates.     

Combined Effects of Nitrogen Concentration and Seasonal Changes on the Production of Lipids in Nannochloropsis oculata

Instead of sole nutrient starvation to boost algal lipid production, we addressed nutrient limitation at two different seasons (autumn and spring) during outdoor cultivation in flat panel photobioreactors. Lipid accumulation, biomass and lipid productivity and changes in fatty acid composition of Nannochloropsis oculata were investigated under nitrogen (N) limitation (nitrate:phosphate N:P 5, N:P 2.5 molar ratio). N. oculata was able to maintain a high biomass productivity under N-limitation compared to N-sufficiency (N:P 20) at both seasons, which in spring resulted in nearly double lipid productivity under N-limited conditions (0.21 g L⁻¹ day⁻¹) compared to N-sufficiency (0.11 g L⁻¹ day⁻¹). Saturated and monounsaturated fatty acids increased from 76% to nearly 90% of total fatty acids in N-limited cultures. Higher biomass and lipid productivity in spring could, partly, be explained by higher irradiance, partly by greater harvesting rate (~30%). Our results indicate the potential for the production of algal high value products (i.e., polyunsaturated fatty acids) during both N-sufficiency and N-limitation. To meet the sustainability challenges of algal biomass production, we propose a dual-system process: Closed photobioreactors producing biomass for high value products and inoculum for larger raceway ponds recycling waste/exhaust streams to produce bulk chemicals for fuel, feed and industrial material.     

Shade delayed flowering and decreased photosynthesis, growth and yield of Sacha Inchi (Plukenetia volubilis) plants

We examined variation in the flowering phenology of Sacha Inchi (Plukenetia volubilis) plants monitored during an 8-month period with reference to a light gradient (20%, 52%, 75% and 100% of full sunlight), and related these results to photosynthetic features, growth and fruit yield. With strong phenotypically plastic response to light, relative growth rate and photosynthetic capability continually increased with increasing irradiance and their maximum values were much similar to those of the pioneer species. Shade did not increase the apparent quantum yield and reduce the light saturation point and respiration rate. Thus, it was suggested that P. volubilis is a high-light demanding species. Shade delayed initial flowering date and decreased flower and fruit parts. An earlier initiation of flowering was associated with higher flower biomass and plant total biomass, in turn; the latter was positively correlated with fruit biomass across light gradients. These results suggested that an optimized crop management practice should be adopted to ensure that P. volubilis be grown under conditions with minimal shading.     

Assessment of the Antimicrobial Activity of Algae Extracts on Bacteria Responsible of External Otitis

External otitis is a diffuse inflammation around the external auditory canal and auricle, which is often occurred by microbial infection. This disease is generally treated using antibiotics, but the frequent occurrence of antibiotic resistance requires the development of new antibiotic agents. In this context, unexplored bioactive natural candidates could be a chance for the production of targeted drugs provided with antimicrobial activity. In this paper, microbial pathogens were isolated from patients with external otitis using ear swabs for over one year, and the antimicrobial activity of the two methanol extracts from selected marine (Dunaliella salina) and freshwater (Pseudokirchneriella subcapitata) microalgae was tested on the isolated pathogens. Totally, 114 bacterial and 11 fungal strains were isolated, of which Staphylococcus spp. (28.8%) and Pseudomonas aeruginosa (P. aeruginosa) (24.8%) were the major pathogens. Only three Staphylococcus aureus (S. aureus) strains and 11 coagulase-negative Staphylococci showed resistance to methicillin. The two algal extracts showed interesting antimicrobial properties, which mostly inhibited the growth of isolated S. aureus, P. aeruginosa, Escherichia coli, and Klebsiella spp. with MICs range of 1.4 × 10⁹ to 2.2 × 10¹⁰ cells/mL. These results suggest that the two algae have potential as resources for the development of antimicrobial agents.     

Marine-Derived Quorum-Sensing Inhibitory Activities Enhance the Antibacterial Efficacy of Tobramycin against Pseudomonas aeruginosa

Bacterial epiphytes isolated from marine eukaryotes were screened for the production of quorum sensing inhibitory compounds (QSIs). Marine isolate KS8, identified as a Pseudoalteromonas sp., was found to display strong quorum sensing inhibitory (QSI) activity against acyl homoserine lactone (AHL)-based reporter strains Chromobacterium violaceum ATCC 12472 and CV026. KS8 supernatant significantly reduced biofilm biomass during biofilm formation (−63%) and in pre-established, mature P. aeruginosa PAO1 biofilms (−33%). KS8 supernatant also caused a 0.97-log reduction (−89%) and a 2-log reduction (−99%) in PAO1 biofilm viable counts in the biofilm formation assay and the biofilm eradication assay respectively. The crude organic extract of KS8 had a minimum inhibitory concentration (MIC) of 2 mg/mL against PAO1 but no minimum bactericidal concentration (MBC) was observed over the concentration range tested (MBC > 16 mg/mL). Sub-MIC concentrations (1 mg/mL) of KS8 crude organic extract significantly reduced the quorum sensing (QS)-dependent production of both pyoverdin and pyocyanin in P. aeruginosa PAO1 without affecting growth. A combinatorial approach using tobramycin and the crude organic extract at 1 mg/mL against planktonic P. aeruginosa PAO1 was found to increase the efficacy of tobramycin ten-fold, decreasing the MIC from 0.75 to 0.075 µg/mL. These data support the validity of approaches combining conventional antibiotic therapy with non-antibiotic compounds to improve the efficacy of current treatments.     

Role and effects of winter buds and rhizome morphology on the survival and growth of common reed (Phragmites australis)

To confirm the role of winter buds and rhizome morphology on the winter survival and subsequent growth of common reed (Phragmites australis), rhizome fragments of varying sizes with and without winter buds from three different P. australis populations were transplanted to small pots in a greenhouse and wintered by exposing them to a dry atmospheric condition. Survival ratios of the rhizome fragments with winter buds were much greater than those without winter buds regardless of population type, supporting that winter buds play a role as the insulator of rhizome fragments from dry atmosphere during winter season. Most growth characteristics of surviving P. australis from the rhizome fragments were similar to those in natural wetlands, indicating preserved growth characteristics through the rhizomes, which is an asexual organ. The average number of rhizome fragment nodes showed negative relationships with the number of blades per culm (P < .05) and basal culm diameter (P < .05) of surviving P. australis. In contrast, the diameter of planted rhizome fragments had significant positive relationships with the final shoot height (P < .05), above-ground dry weight (P < .05), root DW (P < .01), and total DW (P < .05). Rhizome volume also showed a similar correlation with the rhizome diameter. Thicker and larger rhizome fragments particularly with winter buds, indicating the young age of rhizome, seemed likely to guarantee greater survival and higher subsequent shoot growth and biomass production as well.     

LC-PUFA-Enriched Oil Production by Microalgae: Accumulation of Lipid and Triacylglycerols Containing n-3 LC-PUFA Is Triggered by Nitrogen Limitation and Inorganic Carbon Availability in the Marine Haptophyte Pavlova lutheri

In most microalgal species, triacyglycerols (TAG) contain mostly saturated and monounsaturated fatty acids, rather than PUFA, while PUFA-enriched oil is the form most desirable for dietary intake. The ability of some species to produce LC-PUFA-enriched oil is currently of specific interest. In this work, we investigated the role of sodium bicarbonate availability on lipid accumulation and n-3 LC-PUFA partitioning into TAG during batch cultivation of Pavlova lutheri. Maximum growth and nitrate uptake exhibit an optimum concentration and threshold tolerance to bicarbonate addition (~9 mM) above which both parameters decreased. Nonetheless, the transient highest cellular lipid and TAG contents were obtained at 18 mM bicarbonate, immediately after combined alkaline pH stress and nitrate depletion (day nine), while oil body and TAG accumulation were highly repressed with low carbon supply (2 mM). Despite decreases in the proportions of EPA and DHA, maximum volumetric and cellular EPA and DHA contents were obtained at this stage due to accumulation of TAG containing EPA/DHA. TAG accounted for 74% of the total fatty acid per cell, containing 55% and 67% of the overall cellular EPA and DHA contents, respectively. These results clearly demonstrate that inorganic carbon availability and elevated pH represent two limiting factors for lipid and TAG accumulation, as well as n-3 LC-PUFA partitioning into TAG, under nutrient-depleted P. lutheri cultures.     

Recent Advances in Exopolysaccharides from Paenibacillus spp.: Production,Isolation, Structure,and Bioactivities

This review provides a comprehensive summary of the most recent developments of various aspects (i.e., production, purification, structure, and bioactivity) of the exopolysaccharides (EPSs) from Paenibacillus spp. For the production, in particular, squid pen waste was first utilized successfully to produce a high yield of inexpensive EPSs from Paenibacillus sp. TKU023 and P. macerans TKU029. In addition, this technology for EPS production is prevailing because it is more environmentally friendly. The Paenibacillus spp. EPSs reported from various references constitute a structurally diverse class of biological macromolecules with different applications in the broad fields of pharmacy, cosmetics and bioremediation. The EPS produced by P. macerans TKU029 can increase in vivo skin hydration and may be a new source of natural moisturizers with potential value in cosmetics. However, the relationships between the structures and activities of these EPSs in many studies are not well established. The contents and data in this review will serve as useful references for further investigation, production, structure and application of Paenibacillus spp. EPSs in various fields.     

Extracts from Cladiella australis, Clavularia viridis and Klyxum simplex (Soft Corals) are Capable of Inhibiting the Growth of Human Oral Squamous Cell Carcinoma Cells

Many biomedical products have already been obtained from marine organisms. In order to search more therapeutic drugs against cancer, this study demonstrates the cytotoxicity effects of Cladiella australis, Clavularia viridis and Klyxum simplex extracts on human oral squamous cell carcinoma (SCC4, SCC9 and SCC25) cells using cell adhesion and cell viability assay. The morphological alterations in SCCs cells after treatment with three extracts, such as typical nuclear condensation, nuclear fragmentation and apoptotic bodies of cells were demonstrated by Hoechst stain. Flow cytometry indicated that three extracts sensitized SCC25 cells in the G₀/G₁ and S-G₂/M phases with a concomitant significantly increased sub-G₁ fraction, indicating cell death by apoptosis. This apoptosis process was accompanied by activation of caspase-3 expression after SCC25 cells were treated with three extracts. Thereby, it is possible that extracts of C. australis, C. viridis and K. simplex cause apoptosis of SCCs and warrant further research investigating the possible anti-oral cancer compounds in these soft corals.     

Relative abundance and damage by target and non-target insects on Bollgard and BollgardII cotton cultivars

Bollgard^® and BollgardII^® cotton cultivars were evaluated for their efficacy for control of bollworms and their effects on sucking insects and the abundance of natural enemies under bollworm insecticide protected and non-insecticide protected field conditions during the 2004 and 2005 cropping seasons. Bollgard cultivars are genetic transforms that produce Cry1Ac toxins from Bacillus thuringiensis (Bt) to control American bollworm, Helicoverpa armigera. Second generation BollgardII cultivars contain 2 toxins, Cry1Ac and Cry2Ab, and are more effective in controlling a broader range of caterpillar species. Bollgard cultivars were free of H. armigera damage until harvest under insecticide protected as well as non-protected conditions. Both BollgardII and Bollgard cultivars were infested with larvae of spotted bollworm, Earias vitella, and spiny bollworm, Earias insulana, at later crop growth stages. Neither BollgardII nor Bollgard cultivars were free of square (fruiting body), open boll and loculi damage. Bollworm damage did not reach economic threshold levels up to harvest. Densities of sucking insects (Amrasca biguttula biguttula, Bemisia tabaci, Aphis gossypi, and Thrips tabaci), of the foliage feeder Myllocerus undecimpustulatus and of predators (Chrysoperla spp., Orius spp., Coccinella spp., Brumus spp., Vespa spp., Lycosa spp., and Aranews spp.) were similar on Bollgard, BollgardII and conventional cultivars. The time of the first appearance of bollworms, sucking insects and predators on Bt cotton did not vary from conventional cotton varieties. Insecticidal protection based on economic threshold levels resulted in significant reductions in bollworm damage on conventional cultivars.     

Two Severe Cases of Tungiasis in Goat Kids in Uganda

Tungiasis ensues from the penetration and burrowing of female sand fleas (Tunga spp.; Siphonaptera: Tungidae) in the skin of mammals. There are few case reports of severe tungiasis in goats and in these cases the Tunga species were not in most cases clearly identified. Two cases of severe tungiasis caused by Tunga penetrans in goat kids from tungiasis-endemic rural Uganda are reported. These are the first severe cases of tungiasis in goats reported from outside South America.     

Comparative Studies on the Characteristic Fatty Acid Profiles of Four Different Chinese Medicinal Sargassum Seaweeds by GC-MS and Chemometrics

Sargassum seaweeds produce abundant biomass in China and have long been used as herbal medicine and food. Their characteristic fatty acid (FA) profiles and related potential function in promoting cardiovascular health (CVH) have not been systematically investigated. In this study, FA profiles of four medicinal Sargassum were characterized using GC-MS. Principal component analysis was used to discriminate the four medicinal Sargassum, and orthogonal projection to latent structures discriminant analysis was carried out between the two official species HAI ZAO and between the two folk medicine species HAI QIAN. In all of the algae investigated, the major SFA and MUFA were palmitic and stearic acid, respectively, while the major PUFAs were linoleic, arachidonic, and eicosapentaenoic acid. S. fusiforme and S. horneri had higher concentrations of PUFAs. With respect to CVH, all of the studied species, particularly S. fusiforme, exhibited satisfactory levels such as PUFA/SFA ratio and n-6/n-3 ratio. Each species possesses a unique FA profile and is discriminated clearly. Potential key FA markers (between the two Chinese official species, and between the two folk species) are assessed. The study provides characteristic fatty acid profiles of four Chinese medicinal Sargassum and their related potential function in promoting CVH.     

BMAA Inhibits Nitrogen Fixation in the Cyanobacterium Nostoc sp. PCC 7120

Cyanobacteria produce a range of secondary metabolites, one being the neurotoxic non-protein amino acid β-N-methylamino-L-alanine (BMAA), proposed to be a causative agent of human neurodegeneration. As for most cyanotoxins, the function of BMAA in cyanobacteria is unknown. Here, we examined the effects of BMAA on the physiology of the filamentous nitrogen-fixing cyanobacterium Nostoc sp. PCC 7120. Our data show that exogenously applied BMAA rapidly inhibits nitrogenase activity (acetylene reduction assay), even at micromolar concentrations, and that the inhibition was considerably more severe than that induced by combined nitrogen sources and most other amino acids. BMAA also caused growth arrest and massive cellular glycogen accumulation, as observed by electron microscopy. With nitrogen fixation being a process highly sensitive to oxygen species we propose that the BMAA effects found here may be related to the production of reactive oxygen species, as reported for other organisms.     

HPLC-analysis of the concentrations of free asparagine and glutamine in potato tubers grown with varying amounts of nitrogen

Summary The paper describes the rapid determination by HPLC of free asparagine and glutamine in potato tubers. The method has been applied to tubers from experiments in pots to which differing amounts of nitrogen had been added. Increasing levels of N-fertilizer, applied in inorganic or organic form, increased the concentrations of asparagine and glutamine in the tuber dry matter as well as their amounts per pot. amide concentrations are significantly correlated to those of crude protein (P<-0.001) and nitrate (P<-0.01).     

High-EPA Biomass from Nannochloropsis salina Cultivated in a Flat-Panel Photo-Bioreactor on a Process Water-Enriched Growth Medium

Nannochloropsis salina was grown on a mixture of standard growth media and pre-gasified industrial process water representing effluent from a local biogas plant. The study aimed to investigate the effects of enriched growth media and cultivation time on nutritional composition of Nannochloropsis salina biomass, with a focus on eicosapentaenoic acid (EPA). Variations in fatty acid composition, lipids, protein, amino acids, tocopherols and pigments were studied and results compared to algae cultivated on F/2 media as reference. Mixed growth media and process water enhanced the nutritional quality of Nannochloropsis salina in laboratory scale when compared to algae cultivated in standard F/2 medium. Data from laboratory scale translated to the large scale using a 4000 L flat panel photo-bioreactor system. The algae growth rate in winter conditions in Denmark was slow, but results revealed that large-scale cultivation of Nannochloropsis salina at these conditions could improve the nutritional properties such as EPA, tocopherol, protein and carotenoids compared to laboratory-scale cultivated microalgae. EPA reached 44.2% ± 2.30% of total fatty acids, and α-tocopherol reached 431 ± 28 µg/g of biomass dry weight after 21 days of cultivation. Variations in chemical compositions of Nannochloropsis salina were studied during the course of cultivation. Nannochloropsis salina can be presented as a good candidate for winter time cultivation in Denmark. The resulting biomass is a rich source of EPA and also a good source of protein (amino acids), tocopherols and carotenoids for potential use in aquaculture feed industry.     

Biological control of Macrophomina phaseolina by chemotactic fluorescent Pseudomonas aeruginosa PN1 and its plant growth promotory activity in chir-pine

Ten strains of Pseudomonas aeruginosa (PN1 ˜ PN10) isolated from rhizosphere of chir-pine were tested for their plant growth promontory properties and antagonistic activities against Macrophomina phaseolina in vitro and in vivo. P. aeruginosa PN1 produced siderophore, IAA, cyanogen and solubilized phosphorus, besides producing chitinase and β-1,3-glucanase. In dual culture, P. aeruginosa PN1 caused 69% colony growth inhibition. However, cell free culture filtrate also posed inhibitory effect but to a lesser extent. After 90 days, P. aeruginosa PN1 increased plant growth and biomass in pots trial containing M. phaseolina-infested soil. PN1 showed the strong chemotaxis toward root exudates resulting in effective root colonization. Moreover, increased population in rhizosphere of these bacteria was also recorded after 90 days of treatment. Thus, chemotactic fluorescent P. aeruginosa PN1 exhibited strong antagonistic property against M. phaseolina, suppressed the disease and improved plant growth of the seedlings of chir-pine proving potential biocontrol agent.