Effects of three-dimensional spheroid culture on equine mesenchymal stem cell plasticity

Mesenchymal stem cells (MSCs) are useful candidates for tissue engineering and cell therapy fields. We optimize culture conditions of equine adipose tissue-derived MSCs (eAD-MSCs) for treatment of horse fractures. To investigate enhancing properties of three-dimensional (3D) culture system in eAD-MSCs, we performed various sized spheroid formation and determined changes in gene expression levels to obtain different sized spheroid for cell therapy. eAD-MSCs were successfully isolated from horse tailhead. Using hanging drop method, spheroid formation was generated for three days. Quantitative real-time PCR was performed to analyze gene expression. As results, expression levels of pluripotent markers were increased depending on spheroid size and the production of PGE₂ was increased in spheroid formation compared to that in monolayer. Ki-67 showed a remarkable increase in the spheroid formed with 2.0?×?10⁵ cells/drop as compared to that in the monolayer. Expression levels of angiogenesis-inducing factors such as VEGF, IL-6, IL-8, and IL-18 were significantly increased in spheroid formation compared to those in the monolayer. Expression levels of bone morphogenesis-inducing factors such as Cox-2 and TGF-β1 were also significantly increased in spheroid formation compared to those in the monolayer. Expression levels of osteocyte-specific markers such as RUNX2, osteocalcin, and differentiation potential were also significantly increased in spheroid formation compared to those in the monolayer. Therefore, spheroid formation of eAD-MSCs through the hanging drop method can increases the expression of angiogenesis-inducing and bone morphogenesis-inducing factors under optimal culture conditions.     

Positive nitrogen balance of <Emphasis Type="Italic">Acacia mangium</Emphasis> woodlots as fallows in the Philippines based on <Superscript>15</Superscript>N natural abundance data of N<Subscript>2</Subscript> fixation

Nitrogen inputs from biological nitrogen fixation contribute to productivity and sustainability of agroforestry systems but they need to be able to offset export of N when trees are harvested. This study assessed magnitudes of biological nitrogen fixation (natural ¹⁵N abundance) and N balance of Acacia mangium woodlots grown in farmer’s fields, and determined if N₂ fixation capacity was affected by tree age. Tree biomass, standing litter, understory vegetation and soil samplings were conducted in 15 farmer’s fields growing A. mangium as a form of sequential agroforestry in Claveria, Misamis Oriental, Philippines. The trees corresponded to ages of 4, 6, 8, 10 and 12 years, and were replicated three times. Samples from different plant parts and soils (0–100 cm) were collected and analyzed for δ¹⁵N and nutrients. The B-value, needed as a reference of isotopic discrimination when fully reliant on atmospheric N, was generated by growing A. mangium in an N₂-free sand culture in the glasshouse. Isotopic discrimination occurring during N₂ fixation and metabolic processes indicated variation of δ¹⁵N values in the order of nodules > old leaves > young leaves > stems > litterfall and roots of the trees grown in the field, with values ranging from −0.8 to 3.5‰ except nodules which were enriched and significantly different from other plant parts (P < 0.0001). Isotopic discrimination was not affected by tree age (P > 0.05). Plants grown in N free sand culture exhibited the same pattern of isotopic discrimination as plants grown in the field. The estimated B-value for the whole plant of A. mangium was −0.86‰. Mature tree stands of 12 years accumulated up to 1994 kg N ha⁻¹ in aboveground biomass. Average proportion of N derived from N₂ fixation of A. mangium was 54% (±22) and was not affected by age (P > 0.05). Average yearly quantities of N₂ fixed were 128 kg N ha⁻¹ in above-ground biomass amounting to 1208 kg N fixed ha⁻¹ over 12 years. Harvest of 12-year old trees removed approximately 91% of standing aboveground biomass from the site as timber and fuel wood. The resulting net N balance was +151 kg N ha⁻¹ derived from remaining leaves, twigs, standing litter, and +562 kg N ha⁻¹ when tree roots were included in the calculation. The fast growing A. mangium appears to be a viable fallow option for managing N in these systems. However, other nutrients have to be replaced by using part of the timber and fuel wood sales to compensate for large amounts of nutrient removed in order for the system to be sustainable.     

Newly recorded species of the genus Isohypsibius （Tardigrada; Hypsibiidae） from China

This paper reports two newly recorded species, lsohypsibius lunulatus Iharos, 1966 and lsohypsibiusprosostomus Thulin, 1928, of the genus lsohypsibius （Tardigrada; Hypsibiidae） from China. The specimens of lsohysibius lunulatus were collected from Taibai Mt （34°18′N, 107°42′E） at 2,500 m a.s.1, and those oflsohypsibius prosostomus from Taibai Mt （34°10′N, 107°35′E） at 2,000 m above sea level. All specimens are deposited at the College of Life Sciences, Shaanxi Normal University, China. A key to the Chinese species of lsohypsibius was also given.     

Immune responses elicited in mice with recombinant <Emphasis Type="Italic">Lactococcus lactis</Emphasis> expressing F4 fimbrial adhesin FaeG by oral immunization

Enterotoxigenic Escherichia coli (ETEC) is a major pathogenic agent causing piglet diarrhea. The major subunit and adhesin FaeG of F4⁺ ETEC is an important virulence factor with strong immunogenicity. To determine whether Lactococcus lactis (L. lactis) could effectively deliver FaeG to the mucosal immune system, recombinant L. lactis expressing FaeG was constructed, and immune responses in mice following oral route delivery of recombinant L. lactis were explored. The production of FaeG expressed in L. lactis was up to approximately 10% of soluble whole-cell proteins, and recombinant FaeG (rFaeG) possessed good immunoreactivity by Western blot analysis. Oral immunization with recombinant L. lactis expressing FaeG induced F4-specific mucosal and systemic immune responses in the mice. In addition, high dose recombinant L. lactis or co-administration of high dose recombinant L. lactis with CTB enhanced the immune responses. These results suggested that L. lactis expressing FaeG was a promising candidate vaccine against ETEC.     

Tall fescue forage mass in a grass-legume mixture: predicted efficiency of indirect selection

High fertilizer prices and improved environmental stewardship have increased interest in grass-legume mixed pastures. It has been hypothesized, but not validated, that the ecological combining ability between grasses and legumes can be improved by breeding specifically for mixture performance. This experiment examined the predicted efficiency of selection in a grass monoculture environment to indirectly improve tall fescue (Lolium arundinaceum (Schreb.) Darbysh.) forage mass in a grass-legume mixture. Heritability, genetic and rank correlations, and selection efficiencies were estimated for forage mass in a tall fescue half-sib population grown as spaced-plants overseeded with either turf-type tall fescue (monoculture) or alfalfa (mixture). Heritability for tall fescue forage mass in monoculture ranged from 0.32 to 0.70 and were always similar or greater than those in mixture (range 0.27–0.55) for four successive harvests and annual total. Genetic correlations between monoculture and mixture tall fescue forage mass varied with values of 0.48, 0.92, ?0.31, 0.70, and 0.25 in June, July, August, October, and annual total, respectively. Indirect selection efficiencies exceeded or approached direct selection for mixtures only in July and October (1.29, and 0.73, respectively). Whereas, indirect selection efficiencies were low in June, August, and annual forage mass (0.58, ?0.31, and 0.28, respectively). Moreover, low Spearman’s rank correlations (?0.03 to 0.35) indicated differing half-sib family performance between the monoculture and mixture environments. Results indicate that direct selection should be used to improve tall fescue forage mass in a grass-legume mixture, and support the hypothesis of increasing ecological combining ability by breeding for mixtures per se.     

Assessing the Impacts of Time-to-Detection Distribution Assumptions on Detection Probability Estimation

Abundance estimates from animal point-count surveys require accurate estimates of detection probabilities. The standard model for estimating detection from removal-sampled point-count surveys assumes that organisms at a survey site are detected at a constant rate; however, this assumption can often lead to biased estimates. We consider a class of N-mixture models that allows for detection heterogeneity over time through a flexibly defined time-to-detection distribution (TTDD) and allows for fixed and random effects for both abundance and detection. Our model is thus a combination of survival time-to-event analysis with unknown-N, unknown-p abundance estimation. We specifically explore two-parameter families of TTDDs, e.g., gamma, that can additionally include a mixture component to model increased probability of detection in the initial observation period. Based on simulation analyses, we find that modeling a TTDD by using a two-parameter family is necessary when data have a chance of arising from a distribution of this nature. In addition, models with a mixture component can outperform non-mixture models even when the truth is non-mixture. Finally, we analyze an Ovenbird data set from the Chippewa National Forest using mixed effect models for both abundance and detection. We demonstrate that the effects of explanatory variables on abundance and detection are consistent across mixture TTDDs but that flexible TTDDs result in lower estimated probabilities of detection and therefore higher estimates of abundance.Supplementary materials accompanying this paper appear on-line.     

Optimisation of larval culture of the mussel <Emphasis Type="Italic">Mytilus</Emphasis><Emphasis Type="Italic">edulis</Emphasis> (L.)

The blue mussel Mytilus edulis is a commercially important species whose fishery and culture generally relies on natural spat collection. Hatchery-production could provide an alternative source of seed, enabling reliable expansion of the industry. Mussel spawning and larval rearing trials were carried out to optimise elements of hatchery production. Culturing fertilised eggs at low density (20–200 eggs cm⁻²) rather than high density (400–720 eggs cm⁻²) significantly improved the quality of first veliger larvae and differences in this improvement were evident between the eggs from different females (maternal effects). Veliger larval growth at 17 or 21°C was significantly faster than growth at 14°C. Feeding veliger larvae an identical total ration either daily or at 2–3 day intervals did not significantly affect their growth. Different microalgal diets (1: Isochrysis sp. (clone T-ISO), 2: Chaetoceros calcitrans forma pumilus, 3: C. muelleri, 4: mixed Isochrysis sp. (clone T-ISO) and C. calcitrans f. pumilus, and 5: mixed Isochrysis sp. (clone T-ISO) and C. muelleri) were tested on veliger larval growth and mixed diets outperformed single-species diets.     

Analysis of rice blast resistance gene <Emphasis Type="Italic">Pi</Emphasis>-<Emphasis Type="Italic">z</Emphasis> in rice germplasm using pathogenicity assays and DNA markers

The Pi-z gene in rice confers resistance to a wide range of races of the rice blast fungus, Magnaporthe oryzae. The objective of this study was to characterize Pi-z in 111 rice germplasm accessions using DNA markers and pathogenicity assays. The existence of Pi-z in rice germplasm was detected by using four simple sequence repeat (SSR) markers (RM527, AP4791, AP5659-1, AP5659-5) closely linked to Pi-z, and was verified using pathogenicity assays with an avirulent strain (IE1k) and two virulent races (IB33 and IB49). Among 111 germplasm accessions evaluated, 73 were found to contain the Pi-z gene using both SSR markers and pathogenicity assays. The remaining 38 germplasm accessions were found to be inconsistent in their responses to the blast races IB33, IEIk and IB49 with expected SSR marker alleles, suggesting the presence of unexpected SSR alleles and additional R gene(s). These characterized germplasm can be used for genetic studies and marker-assisted breeding for improving blast resistance in rice.     

Laboratory characterization of firebrands involved in spot fires

• Introduction  

Wildfires are considered the most important disturbance in the Mediterranean Basin, and some are propagated over long distances due to lift-off and ignition of firebrands.     

Evaluation of two Indian native chicken breeds for reproduction traits and heritability of juvenile growth traits

The present study was conducted to evaluate two Indian native chicken breeds, namely, Aseel and Kadaknath for fertility, hatchability, genetic parameters of juvenile growth traits, and semen quality traits at the onset of sexual maturity. The fertility was similar in Aseel (86.96%) and Kadaknath (85.15%); however, a relatively higher hatchability was observed in Kadaknath (77.94%) than Aseel (70.74%). Heritability estimates of body weights at 4 weeks of age were almost similar in Aseel (0.37) and Kadaknath (0.39), while the estimate of body weight at 6 weeks of age was higher in Aseel (0.42) than Kadaknath (0.31). The heritability estimate of shank length at 6 weeks of age was lower in Aseel (0.16) compared to Kadaknath (0.35). The age at first egg in the flock was comparable in Aseel (148 days) and Kadaknath (150 days). Aseel breed with significantly (P ≤ 0.001) higher body weight, absolute and relative testes weights had significantly higher semen volume (P ≤ 0.05) and sperm motility (P ≤ 0.01) but had lower seminal plasma cholesterol level (P ≤ 0.05) as compared to Kadaknath. It can be concluded that there is a scope for genetic improvement of these two native breeds for juvenile growth traits since heritability estimates of these traits were relatively high.