Probiotics in the intestinal tract of juvenile whiteleg shrimp Litopenaeus vannamei: modulation of the bacterial community

Molecular analysis of the 16S rDNA of the intestinal microbiota of whiteleg shrimp Litopenaeus vannamei was examined to investigate the effect of a Bacillus mix (Bacillus endophyticus YC3-b, Bacillus endophyticus C2-2, Bacillus tequilensisYC5-2) and the commercial probiotic (Alibio^®) on intestinal bacterial communities and resistance to Vibrio infection. PCR and single strain conformation polymorphism (SSCP) analyses were then performed on DNA extracted directly from guts. Injection of shrimp with V. parahaemolyticus at 2.5 × 10⁵ CFU g^?1 per shrimp followed 168 h after inoculation with Bacillus mix or the Alibio probiotic or the positive control. Diversity analyses showed that the bacterial community resulting from the Bacillus mix had the highest diversity and evenness and the bacterial community of the control had the lowest diversity. The bacterial community treated with probiotics mainly consisted of α- and γ-proteobacteria, fusobacteria, sphingobacteria, and flavobacteria, while the control mainly consisted of α-proteobacteria and flavobacteria. Differences were grouped using principal component analyses of PCR-SSCP of the microbiota, according to the time of inoculation. In Vibrio parahaemolyticus-infected shrimp, the Bacillus mix (~33 %) induced a significant increase in survival compared to Alibio (~21 %) and the control (~9 %). We conclude that administration of the Bacillus mix induced modulation of the intestinal microbiota of L. vannamei and increased its resistance to V. parahaemolyticus.     

Administration of Probiotics Improves the Brine Shrimp Production and Prevents Detrimental Effects of Pathogenic <Emphasis Type="Italic">Vibrio</Emphasis> Species

In this study, we evaluated a consortium of probiotic bacteria as an environmentally-friendly strategy for controlling pathogenic Vibrio species during the brine shrimp incubation period. Probiotic strains were initially selected on basis of (i) their ability to colonize the cyst surfaces, (ii) their absence of cross-inhibitory effects, and (iii) no detrimental effect on cyst hatching. The cysts and nauplius surfaces were immediately colonized after the application of selected probiotic strains, without detrimental effects on survival. Ten probiotic strains were mixed at similar proportions (probiotic consortium) and evaluated at different concentrations into brine shrimp cultures during incubation and early stages of development. Subsequently, these cultures were challenged with Vibrio parahaemolyticus and Vibrio harveyi. The probiotic consortium was effective to reduce the abundance of pathogenic Vibrio species and to prevent the mortality during Vibrio challenges; however, its effect was concentration-dependent and was successful at a starting concentration of 1.8?×?10⁶ CFU/ml. Our results suggest that this probiotic consortium offers an alternative to antimicrobial agents routinely used to reduce the incidence and prevalence of pathogenic Vibrio species in brine shrimp production.     

Microbial Flora of Pond-Reared Brown Shrimp (Penaeus aztecus)

Agar plate counts and microbial types are reported for brown shrimp reared in 2-acre natural marshland and in 0.5-acre artificial ponds during June to October 1970. Bacterial counts of pond-reared shrimp ranged from 5 × 10⁴ to 5.5 × 10⁶ per g. At final harvest in October, bacterial counts ranged from 2 × 10⁵ to 5.5 × 10⁶ per g. In marsh ponds, bacterial counts of shrimp and pond water were lowest in August when both water temperature and salinity were high. Coryneform bacteria and to a lesser extent Vibrio were the predominant isolates from fresh pond shrimp. Shrimp stored at 3 to 5 C for 7 days were acceptable as judged by appearance and odor. Between 7 and 14 days of refrigerated storage, bacterial counts increased sharply and about 50% of the samples became unacceptable. Refrigerated storage of pond shrimp caused increases in coryneform bacteria and micrococci and decreases in Vibrio, Flavobacterium, Moraxella, and Bacillus species. Pseudomonas species were not significant in fresh or stored pond shrimp. The microbial flora of pond water usually was dominated by coryneform bacteria, Flavobacterium, Moraxella, and Bacillus species.     

Acute Toxicity of Arsenic under Different Temperatures and Salinity Conditions on the White Shrimp Litopenaeus vannamei

The aim of this study was to determine acute toxicity in the post larvae of the white shrimp Litopenaeus vannamei after 96 h of exposure to dissolved arsenic under three different temperatures and salinity conditions. Recent reports have shown an increase in the presence of this metalloid in coastal waters, estuaries, and lagoons along the Mexican coast. The white shrimp stands out for its adaptability to temperature and salinity changes and for being the main product for many commercial fisheries; it has the highest volume of oceanic capture and production in Mexican shrimp farms. Lethal concentrations (LC₅₀–96 h) were obtained at nine different combinations (3?×?3 combinations in total) of temperature (20, 25, and 30 °C) and salinity (17, 25, and 33) showing mean LC₅₀–96 h values (±standard error) of 9.13?±?0.76, 9.17?±?0.56, and 6.23?±?0.57 mgAs?L^?1(at 20 °C and 17, 25, and 33 salinity); 12.29?±?2.09, 8.70?±?0.82, and 8.03?±?0.59 mgAs?L^?1 (at 25 °C and 17, 25, and 33 salinity); and 7.84?±?1.30, 8.49?±?1.40, and 7.54?±?0.51 mgAs?L^?1 (at 30 °C and 17, 25, and 33 salinity), respectively. No significant differences were observed for the optimal temperature and isosmotic point of maintenance (25 °C–S 25) for the species, with respect to the other experimental conditions tested, except for at 20 °C–S 33, which was the most toxic. Toxicity under 20 °C–S 33 conditions was also higher than 25 °C–S 17 and 20 °C (S 17 or 25). The least toxic condition was 25 °C–S 17. All this suggests that the toxic effect of arsenic is not affected by temperature changes; it depends on the osmoregulatory pattern developed by the shrimp, either hyperosmotic at low salinity or hiposmotic at high salinity, as observed at least on the extreme salinity conditions here tested (17 and 33). However, further studies testing salinities near the isosmotic point (between 20 and 30 salinities) are needed to clarify these mechanisms.     

Productivity and biochemical composition of <Emphasis Type="Italic">Tetradesmus obliquus</Emphasis> and <Emphasis Type="Italic">Phaeodactylum tricornutum</Emphasis>: effects of different cultivation approaches

The present work evaluated biomass productivity, carbon dioxide fixation rate, and biochemical composition of two microalgal species, Phaeodactylum tricornutum (Bacillariophyta) and Tetradesmus obliquus (Chlorophyta), cultivated indoors in high-technology photobioreactors (HT-PBR) and outdoors both in pilot ponds and low-technology photobioreactors in a greenhouse in southern Italy. Microalgae were grown in standard media, under nitrogen starvation, and in two liquid digestates obtained from anaerobic digestion of agro-zootechnical and vegetable biomass. P. tricornutum, cultivated in semi-continuous mode in indoor HT-PBRs with standard medium, showed a biomass productivity of 21.0?±?2.3 g m^?2 d^?1. Applying nitrogen starvation, the lipid productivity increased from 2.3 up to 4.5?±?0.5 g m^?2 d^?1, with a 24 % decrease of biomass productivity. For T. obliquus, a biomass productivity of 9.1?±?0.9 g m^?2 d^?1 in indoor HT-PBR was obtained using standard medium. Applying liquid digestates as fertilizers in open ponds, T. obliquus gave a biomass productivity (10.8?±?2.0 g m^?2 d^?1) not statistically different from complete medium such as P. tricornutum (6.5?±?2.2 g m^?2 d^?1). The biochemical data showed that the fatty acid composition of the microalgal biomass was affected by the different cultivation conditions for both microalgae. In conclusion, it was found that the microalgal productivity in standard medium was about doubled in HT-PBR compared to open ponds for P. tricornutum and was about 20 % higher for T. obliquus.     

Evaluation of commercial and natural food in experimental cultures of white shrimp based on stable carbon isotopes

The farming of shrimp is developing quickly worldwide, and recently, ingredients such as seaweeds in low proportion (25 to 4 %), incorporated into the commercial food, have been shown to improve the shrimp productive variables. The change of commercial foods to commercial feed with a proportion of natural food, and finally, to natural food has been little and simultaneously evaluated. The aim of our study was to determine the relative contribution of dietary carbon to the growth of Litopenaeus vannamei fed with a proportion of 4 % Sargassum (δ¹³C = ?20.9?±?0.05?‰), 4 % Ulva (δ¹³C = ?20.6?±?0.6?‰) meal, and a control diet (δ¹³C?=??22.6?±?0.2?‰) in 60-L tanks for 30 days, and finally, with the green seaweed Ulva spp. (δ¹³C = ?13.2?±?0.25?‰) and Ulva meal (δ¹³C = ?14.5?±?0.6?‰) in open-air ponds for 120 days, by measuring δ¹³C for each of the foods and in the muscle of shrimp. After 15 days, the rates of metabolic turnover (Δ¹³C = δ¹³C_shrimp ? δ¹³C_food) were constant until the end of the experiment in the tanks. The muscle of shrimp assimilated carbon from all diets, which demonstrated the potential use of combined diets and the optimization of their use in diets containing seaweed. Our data will be useful in future interpretations of field and laboratory isotopic values for this species.     

Biodegradation of Shrimp Biowaste by Marine Exiguobacterium sp. CFR26M and Concomitant Production of Extracellular Protease and Antioxidant Materials: Production and Process Optimization by Response Surface Methodology

Twelve marine bacterial cultures were screened for extracellular protease activity, and the bacterium CFR26M which exhibited the highest activity on caseinate agar plate was identified as an Exiguobacterium sp. Significant amount of extracellular protease (5.9?±?0.3 U/ml) and antioxidant materials, measured as 2,2′-diphenyl picrylhydrazyl (DPPH) radical scavenging activity (44.4?±?0.5 %), was produced by CFR26M in submerged fermentation using a shrimp biowaste medium. Response surface methodology (RSM) was employed to optimize the process variables for maximum production of protease and antioxidant materials by CFR26M. Among the seven variables screened by two-level 2**(7–2) fractional factorial design, the concentration of shrimp biowaste, sugar, and phosphate was found to be significant (p?≤?0.05). The optimum levels of these variables were determined by employing the central composite design (CCD) of RSM. The coefficient of determination (R ²) values of 0.9039 and 0.8924 for protease and antioxidant, respectively, indicates the accuracy of the CCD models. The optimum levels of shrimp biowaste, sugar, and phosphate were 21.2, 10.5, and 2.3 % (w/v) for production of protease and 28.8, 12, and 0.32 % (w/v) for production of antioxidant material, respectively. The concentration of shrimp biowaste, sugar, and phosphate had linear and quadratic effect on both protease and antioxidant productions. RSM optimization yielded 6.3-fold increases in protease activity and 1.6-fold in antioxidant material production. The crude protease of CFR26M had a maximum activity at 32?±?2 °C with pH 7.6. This is the first report on the use of marine Exiguobacterium sp. for concomitant production of protease and antioxidant materials from shrimp biowaste.     

Molecular Detection of Anaerobic Ammonium-Oxidizing (Anammox) Bacteria in High-Temperature Petroleum Reservoirs

Anaerobic ammonium-oxidizing (anammox) process plays an important role in the nitrogen cycle of the worldwide anoxic and mesophilic habitats. Recently, the existence and activity of anammox bacteria have been detected in some thermophilic environments, but their existence in the geothermal subterranean oil reservoirs is still not reported. This study investigated the abundance, distribution and functional diversity of anammox bacteria in nine out of 17 high-temperature oil reservoirs by molecular ecology analysis. High concentration (5.31–39.2 mg l^?1) of ammonium was detected in the production water from these oilfields with temperatures between 55°C and 75°C. Both 16S rRNA and hzo molecular biomarkers indicated the occurrence of anammox bacteria in nine out of 17 samples. Most of 16S rRNA gene phylotypes are closely related to the known anammox bacterial genera Candidatus Brocadia, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia, while hzo gene phylotypes are closely related to the genera Candidatus Anammoxoglobus, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia. The total bacterial and anammox bacterial densities were 6.4?±?0.5?×?10³ to 2.0?±?0.18?×?10⁶ cells ml^?1 and 6.6?±?0.51?×?10² to 4.9?±?0.36?×?10⁴ cell ml^?1, respectively. The cluster I of 16S rRNA gene sequences showed distant identity (<92%) to the known Candidatus Scalindua species, inferring this cluster of anammox bacteria to be a new species, and a tentative name Candidatus “Scalindua sinooilfield” was proposed. The results extended the existence of anammox bacteria to the high-temperature oil reservoirs.     

Genomic Analysis and Comparative Hexavalent Chromium Reduction Potential of Predominant Bacillus species Isolated from Chromite Mine Soil

Ten predominant bacteria (CSB 1-10), isolated from chromite mine soil of Sukinda, Odisha, were characterized by means of biochemical and 16S rRNA gene sequencing. All of the bacterial isolates were Gram-positive, spore-forming, and motile rods with diameter ranging from 1.57–2.79 μm and identified as Bacillus species. Based on 16S rRNA gene sequencing and phylogenetic tree construction, 10 Bacillus species were grouped into two clusters: Bacillus subtilis cluster with four species (two of B. amyloliquefaciens, and one each of B. tequilensis and B. mojavensis); and Bacillus cereus cluster containing six species of B. cereus. Secondary rRNA structure predicted for all 10 bacteria using 16S rRNA sequences revealed some degree of genetic variations among the species. Among the isolated bacteria, CSB-9 was found to be the most efficient chromate reducing strain (0.8 × 10^?4 mg mg^?1 h^?1) in comparison to the others. Chromate reduction of the bacteria was associated with the contribution of extracellular enzyme production, and highest enzyme activity (0.9 ± 0.09 U mL^?1) was observed in CSB-9 (B. amyloliquefaciens). The present study revealed the dominance of Bacillus species in the chromite mine soil and their potential for bioremediation of hexavalent chromium from the polluted environments.     

Litter dynamics and fine root production in Schizolobium parahyba var. amazonicum plantations and regrowth forest in Eastern Amazon

Forest plantations and agroforestry systems with Schizolobium parahyba var. amazonicum have greatly expanded in the Brazilian Amazon, generally as an alternative for reforesting degraded areas. To our knowledge there are no reports of above- and below-ground production in these forest systems. We quantified litter and fine root production in 6-yr old Schizolobium-based plantation forests (monospecific: MON, mixture: MIX, and agroforestry system: AFS) and in ~25-yr old regrowth forest (REG) over 8–12 months. We used litter traps and ingrowth cores to quantify litter and fine root production, respectively. Annual litter production was significantly lower in Schizolobium-based plantations (mean ± standard error, MON?=?5.92?±?0.15, MIX?=?6.08?±?0.13, AFS?=?6.63?±?0.13 Mg ha^?1 year^?1) than in regrowth forest (8.64?±?0.08 Mg ha^?1 year^?1). Schizolobium-based plantations showed significantly higher litter stock (MON?=?7.7?±?1.0, MIX?=?7.4?±?0.1 Mg ha^?1) than REG (5.9?±?1.3 Mg ha^?1). Total fine root production over an 8-month period was significantly higher in Schizolobium-based plantations (MON?=?3.8?±?0.2, MIX?=?3.4?±?0.2, AFS?=?2.7?±?0.1 Mg ha^?1) than in REG (1.1?±?0.03 Mg ha^?1). Six-yr old Schizolobium-based plantations and ~25-yr old regrowth forests showed comparable rates of litter + fine root production, suggesting that young forest plantations may be an interesting alternative to restore degraded areas due to early reestablishment of organic matter cycling under the studied conditions.     

Effect of co-substrate feeding on methane yield of anaerobic digestion of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis>

Microalgal production has many advantages over the use of terrestrial plants; therefore, increases in the use of microalgae for energy production can be expected. Algal biomass can be processed anaerobically to methane; however, the unfavorable C/N ratio of the substrate may have an inhibitory effect. The impact of the application of used cooking oil, maize silage, and mill residue on anaerobic co-digestion of the microalgal Chlorella vulgaris was studied in semi-continuous, laboratory-scale digestion. During the full period of the trial involving anaerobic digestion of algae in the case of mono-digestion and co-digestion with used cooking oil, maize silage, and mill residue, the volumetric methane yields were 0.38?±?0.07, 1.56?±?0.26, 1.19?±?0.18, and 1.16?±?0.13 L L^?1, respectively. Trials were carried out to determine the long-term effect of the total solid (TS) content of substrates (co-digestion of C. vulgaris and used cooking oil at 3.8 and 7.2 % of TS, respectively). Both designs could be increased to 5.5 g VS L^?1 d^?1, but a higher TS% resulted in increased methane production and a longer period of decline in the methane yield due to washout. The sharp decrease in methane content at the end of 90 days was accompanied by a reorganization of the methanogenic archaeal community.     

Microalgal cultivation in porous substrate bioreactor for extracellular polysaccharide production

Netrium digitus is a representative of the species-rich class Zygnematophyceae (Streptophyta). Its intensive extracellular polysaccharide (EPS) production makes this alga interesting for biotechnological applications with a focus on cosmetics and food additives. Quantitative data on growth and EPS production in suspension and, for the first time, in immobilized culture using lab-scale porous substrate bioreactors, so-called Twin-Layer (TL) systems, is presented. It is shown that the cell as well as the EPS dry weight content is increased at least sixfold in immobilized compared to suspension culture. Due to the high amount of EPS, the biofilms reach a thickness of more than 8 mm after 27 days at 70 μmol photons m^?2 s^?1 and with 1.5% CO₂ supply. Frequent exchange of the growth medium results in a linear cell biomass increase of 2.02?±?0.09 g m^?2 growth area day^?1 compared to 2.99?±?0.09 g m^?2 day^?1, when the medium is not exchanged. Under this mode of cultivation, the EPS production is lower and a final concentration of 12.18?±?1.25 g m^?2 compared to 20.76?±?0.85 g m^?2, when medium was exchanged, is reached. It is clearly demonstrated that the relatively slow growing, but excessively EPS producing, microalgal species N. digitus can be grown in porous substrate bioreactors and that this culturing technique is a promising alternative to suspension culture for the Zygnematophyceae.     

Effect of Seawater–Sewage Cross-Transplants on Bacterial Metabolism and Diversity

Bioassays experiments were conducted to determine the metabolic and community composition response of bacteria to transplants between relatively pristine coastal seawater and sewage-impacted seawater. There were four treatments: (1) pristine seawater bacteria?+?pristine seawater (Pb?+?Pw), (2) sewage-impacted bacteria?+?sewage-impacted water (Sb?+?Sw), (3) pristine seawater bacteria?+?sewage-impacted water (Pb?+?Sw), and (4) sewage-impacted bacteria?+?pristine seawater (Sb?+?Pw). Sewage-derived DOC was more labile and readily utilized by bacteria, which favored the growth of high nucleic acid (HNA) bacteria, resulting in high bacterial production (BP, 113?±?4.92 to 130?±?15.8 μg C l^?1?day^?1) and low respiration rate (BR, <67?±?11.3 μg C l^?1?day^?1), as well as high bacterial growth efficiency (BGE, 0.68?±?0.09 to 0.71?±?0.05). In contrast, at the relatively pristine site, bacteria utilized natural marine-derived dissolved organic matter (DOM) at the expense of lowering their growth efficiency (BGE, <0.32?±?0.02) with low BP (<62?±?6.3 μg C l^?1?day^?1) and high BR 133?±?14.2 μg C l^?1?day^?1). Sewage DOM input appeared to alter the partitioning of carbon between respiration and production of bacteria, resulting in a shift toward higher BGE, which would not enhance oxygen consumption. Taxonomic classification based on 454 pyrosequencing reads of the 16S rRNA gene amplicons revealed that changes in bacterial community structure occurred when seawater bacteria were transferred to the eutrophic sewage-impacted water. Sewage DOM fueled the growth of Gammma-proteobacteria and Epsilson-proteobacteria and reduced the bacterial richness, but the changes in the community were not apparent when sewage-impacted bacteria were transferred to pristine seawater.     

Effect of Mg, Si, and Sr on growth and antioxidant activity of the marine microalga Tetraselmis suecica

Efforts to increase the productivity of microalgal cultures have been focused on the improvement of photobioreactors, but little attention has been paid to the nutritional requirements of microalgae in order to improve culture media formulation. In this study, the main goal was obtaining a high productivity for Tetraselmis suecica (Chlorophyta) in semicontinuous culture by adding magnesium (Mg), silicon (Si), and strontium (Sr) at concentrations from 0.01 to 10 mM; at the time, the effect on steady-state cell density, biochemical composition, and antioxidant activity of T. suecica was evaluated. Because productivity is higher in high-density cultures, the work was focused many times to cell density. Mg (3 mM) and Sr (0.1 mM) added separately reached the highest steady-state cell density (7.0?×?10⁶?±?0.4 cells mL^?1) in comparison to control (4.2?±?0.1 cells mL^?1), but simultaneous addition had a synergic effect, achieving 8.7?×?10⁶?±?0.6 cells mL^?1. Silicon (3 mM) significantly affected the steady-state cell density, reaching 6.0?±?0.3 cells mL^?1 and increased the cell ash-free dry weight, reaching 127?±?7.9 pg cell^?1 in comparison to control (102.7?±?5.0 pg cell^?1), resulting in an ash-free dry weight productivity of 0.75?±?0.07 g?L^?1 day^?1. The highest fatty acids content and antioxidant activity, measured by 2, 2-diphenyl-1-picrylhydrazyl (DPPH) method were obtained with Sr 10 mM. Sr treatments showed a high correlation (R ²?=?0.98) between DPPH inhibition and polyphenolic content, explaining its high antioxidant activity. Therefore, the addition of Mg, Si, and Sr to culture medium of T. suecica is recommended to achieve high steady-state cell density in semicontinuous cultures.     

The membraneless bioelectrochemical reactor stimulates hydrogen fermentation by inhibiting methanogenic archaea

The membraneless bioelectrochemical reactor (Ml-BER) is useful for dark hydrogen fermentation. The effect of the electrochemical reaction on microorganisms in the Ml-BER was investigated using glucose as the substrate and compared with organisms in a membraneless non-bioelectrochemical reactor (Ml-NBER) and bioelectrochemical reactor (BER) with a proton exchange membrane. The potentials on the working electrode of the Ml-BER and BER with membrane were regulated to ?0.9 V (versus Ag/AgCl) to avoid water electrolysis with a carbon electrode. The Ml-BER showed suppressed methane production (19.8?±?9.1 mg-C·L^?1·day^?1) and increased hydrogen production (12.6?±?3.1 mg-H·L^?1·day^?1) at pH_out 6.2?±?0.1, and the major intermediate was butyrate (24.9?±?2.4 mM), suggesting efficient hydrogen fermentation. In contrast, the Ml-NBER showed high methane production (239.3?±?17.9 mg-C·L^?1·day^?1) and low hydrogen production (0.2?±?0.0 mg-H·L^?1·day^?1) at pH_out 6.3?±?0.1. In the cathodic chamber of the BER with membrane, methane production was high (276.3?±?20.4 mg-C·L^?1·day^?1) (pH_out, 7.2?±?0.1). In the anodic chamber of the BER with membrane (anode-BER), gas production was low because of high lactate production (43.6?±?1.7 mM) at pH_out 5.0?±?0.1. Methanogenic archaea were not detected in the Ml-BER and anode-BER. However, Methanosarcina sp. and Methanobacterium sp. were found in Ml-NBER. Prokaryotic copy numbers in the Ml-BER and Ml-NBER were similar, as were the bacterial community structures. Thus, the electrochemical reaction in the Ml-BER affected hydrogenotrophic and acetoclastic methanogens, but not the bacterial community.     

Evaluation of productions and economic returns from two brackishwater polyculture systems in tide‐fed ponds

To compare the production and economic performance of two polyculture systems with different species combinations in brackishwater tide‐fed ponds, a 180‐day trial was carried out. In the first combination (T1), mullets (Mugil cephalus, Liza tade and L. parsia at 3 : 1 : 1.3 ratio) and tiger shrimp (Penaeus monodon) and in the second (T2), milkfish (Chanos chanos) and tiger shrimp were stocked keeping the fishes and shrimp at 8000 and 20 000 numbers ha^?1, respectively, in both treatments with duplicate ponds. Since shrimp was an important component of these two systems with open water exchange, the dynamics of heterotrophic bacteria (THB) including Vibrio spp. (TVC) and the occurrence of viral infection agents were studied to understand the disease risks. Among the fishes in T1, M. cephalus attained the highest final weight of 92.29 ± 4.36 g followed by L. tade and L. parsia with 80.40 ± 4.02 and 54.02 ± 2.11 g, respectively. C. chanos in T2 had the highest net weight gain (127.85 g) and daily weight gain (0.71 g day^?1), while M. cephalus in T1 attained the highest specific growth rate (1.60% day^?1). Growth parameters of tiger shrimp were almost similar in both treatments, with no significant differences (P > 0.05). Though total production of fishes and shrimp was lower in T1 (689 kg ha^?1 180 day^?1), it was not significantly different (P > 0.05) from T2 (721 kg ha^?1 180 day^?1). Monthly THB and TVC were not alarming and the absence of viral infections in shrimp indicated no disease risk. Total income from T1 was significantly higher (P < 0.05) than that of T2, but net income and benefit‐cost ratio were insignificantly different between the treatments (P > 0.05). The present findings indicate that upon availability of stocking materials, both polyculture systems would be suitable farming options.     

Influence of silicate on enrichment of highly productive microalgae from a mixed culture

Microalgae have the potential to supply a biobased society with essential feedstocks like sugar and lipids. Besides being productive, strains used for this purpose should grow fast, be resistant to predators, and have good harvestability properties. Diatoms, a class of siliceous algae, have these and other preferred characteristics. In this paper, we describe the enrichment of microalgae in sequencing batch reactors with and without supply of silicate. Both reactors were operated with a light–dark cycle. To maximize storage compound production, carbon fixation and nitrogen uptake were uncoupled by limiting the availability of nitrate to the dark phase. After ten cycles, a stable culture was established in both reactors. The diatom Nitzschia sp. dominated the silicate-rich reactor, and the green algae Chlamydomonas sp. dominated the silicate-depleted reactor. Over the remaining 27 cycles of the experiment, the microalgal community structure did not change, indicating a highly stable system. Although the dominant microalga was highly dependent on the presence of silicate, the performance of both microalgal enrichments was similar. Polymers of glucose were stored during the nitrogen-limited light period. On organic matter dry weight basis, the sugar content of the biomass increased during the light period from 17?±?4 to 53?±?4 % for the silicate-limited culture, and from 14?±?4 to 43?±?4 % (w w ^?1) for the silicate excess culture. These results show that storage compound production can be achieved under various conditions, as long as a selective environment is maintained.     

Pigment production and isotopic fractionations in continuous culture: okenone producing purple sulfur bacteria Part II

Okenone is a carotenoid pigment unique to certain members of Chromatiaceae, the dominant family of purple sulfur bacteria (PSB) found in euxinic photic zones. Diagenetic alteration of okenone produces okenane, the only recognized molecular fossil unique to PSB. The in vivo concentrations of okenone and bacteriochlorophyll a (Bchl a) on a per cell basis were monitored and quantified as a function of light intensity in continuous cultures of the purple sulfur bacterium Marichromatium purpuratum (Mpurp1591). We show that okenone‐producing PSB have constant bacteriochlorophyll to carotenoid ratios in light‐harvesting antenna complexes. The in vivo concentrations of Bchl a, 0.151 ± 0.012 fmol cell^?1, and okenone, 0.103 ± 0.012 fmol cell^?1, were not dependent on average light intensity (10–225 Lux) at both steady and non‐steady states. This observation revealed that in autotrophic continuous cultures of Mpurp1591, there was a constant ratio for okenone to Bchl a of 1:1.5. Okenone was therefore constitutively produced in planktonic cultures of PSB, regardless of light intensity. This confirms the legitimacy of okenone as a signature for autotrophic planktonic PSB and by extrapolation water column euxinia. We measured the δ¹³C, δ¹⁵N, and δ³⁴S bulk biomass values from cells collected daily and determined the isotopic fractionations of Mpurp1591. There was no statistical relationship in the bulk isotope measurements or stable isotope fractionations to light intensity or cell density under steady and non‐steady‐state conditions. The carbon isotope fractionation between okenone and Bchl a with respect to overall bulk biomass (¹³ε_{pigment – biomass}) was 2.2 ± 0.4‰ and ?4.1 ± 0.9‰, respectively. The carbon isotopic fractionation () for the production of pigments in PSB is more variable than previously thought with our reported values for okenone at ?15.5 ± 1.2‰ and ?21.8 ± 1.7‰ for Bchl a.     

Characterization of the triacylglycerol profile in marine diatoms by ultra performance liquid chromatography coupled with electrospray ionization–quadrupole time-of-flight mass spectrometry

Diatoms are considered to have great potential as new biofuel sources because they can effectively accumulate triacylglycerols (TAGs). Detailed structure information of TAG in diatoms is much needed not only for the assessment of biofuel quality such as fatty acid chain length and unsaturation degree but also for the tracing of biosynthetic precursors because the biosynthesis of TAG is typically completed by utilizing the diacylglycerol acyltransferase in the cytoplasm. In this report, a comprehensive characterization of TAGs in marine diatoms was performed using ultra performance liquid chromatography–electrospray ionization–quadrupole time-of-flight mass spectrometry. Many types of major TAGs were identified for the first time in these diatoms: 12 TAGs in Chaetoceros debilis, 9 TAGs in Phaeodactylum tricornutum Bohlin, 16 TAGs in Nitzschia closterium f. minutissima, 16 TAGs in Thalassiosira weissflogii, 13 TAGs in Thalassiosira sp., 16 TAGs in Stephanodiscus asteaea and 7 TAGs in Skeletonema costatum. Semi-quantification of TAGs in these diatoms was also carried out, and it was found that the contents of individual TAGs ranged from 0.5?±?0.1 to 217.9?±?8.1 nmol mg^?1 total lipids. In addition, the total lipid contents in diatoms ranged from 143.6?±?16.3 to 201.1?±?16.3 mg g^?1 dry microalgae and the total TAG contents ranged from 36.8?±?9.5 to 793.2?±?54.4 nmol mg^?1 total lipids. By comparative analysis of the compositions and concentrations of major TAGs in the seven algal strains, N. closterium f. minutissima with high abundance of TAGs containing the most monounsaturated fatty acids (mainly palmitoleic acid) was considered as one of the most promising diatom strains for microalgal biofuel production. Additionally, based on the information of sn-2 fatty acid obtained (mainly C16 in the sn-2 position), we propose the hypothesis that TAGs in diatoms are mainly derived from lipids in chloroplasts through the prokaryotic biosynthesis pathway, including monogalactosyldiacylglycerol and digalactosyldiacylglycerol.     

Enhanced viability of Lactobacillus reuteri for probiotics production in mixed solid-state fermentation in the presence of Bacillus subtilis

In order to develop a multi-microbe probiotic preparation of Lactobacillus reuteri G8-5 and Bacillus subtilis MA139 in solid-state fermentation, a series of parameters were optimized sequentially in shake flask culture. The effect of supplementation of B. subtilis MA139 as starters on the viability of L. reuteri G8-5 was also explored. The results showed that the optimized process was as follows: water content, 50 %; initial pH of diluted molasses, 6.5; inocula volume, 2 %; flask dry contents, 30～35 g/250 g without sterilization; and fermentation time, 2 days. The multi-microbial preparations finally provided the maximum concentration of Lactobacillus of about 9.01?±?0.15 log CFU/g and spores of Bacillus of about 10.30?±?0.08 log CFU/g. Compared with pure fermentation of L. reuteri G8-5, significantly high viable cells, low value of pH, and reducing sugar in solid substrates were achieved in mixed fermentation in the presence of B. subtilis MA139 (P?<?0.05). Meanwhile, the mixed fermentation showed the significantly higher antimicrobial activity against E. coli K88 (P?<?0.05). Based on the overall results, the optimized process enhanced the production of multi-microbe probiotics in solid-state fermentation with low cost. Moreover, the viability of L. reuteri G8-5 could be significantly enhanced in the presence of B. subtilis MA139 in solid-state fermentation, which favored the production of probiotics for animal use.