Rapid degradation of longer DNA fragments enables the improved estimation of distribution and biomass using environmental DNA

The advent of environmental DNA (eDNA) analysis methods has enabled rapid and wide‐range ecological monitoring in aquatic ecosystems, but there is a dearth of information on eDNA degradation. The results of previous studies suggest that the decay rate of eDNA varies depending on the length of DNA fragments. To examine this hypothesis, we compared temporal change in copy number of long eDNA fragments (719 bp) with that of short eDNA fragments (127 bp). First, we isolated rearing water from a target fish species, Japanese Jack Mackerel (Trachurus japonicus), and then quantified the copy number of the long and short eDNA fragments in 1 L water samples after isolating the water from the fish. Long DNA fragments showed a higher decay rate than short fragments. Next, we measured the eDNA copy numbers of long and short DNA fragments using field samples, and compared them with fish biomass as measured by echo intensity. Although a previous study suggested that short eDNA fragments could be overestimated because of nontarget eDNA from a nearby fish market and carcasses, the eDNA concentrations of long fragments were correlated with echo intensity. This suggests that the concentration of longer eDNA fragments reflects fish biomass more accurately than the previous study by removing the effects of the fish market and carcasses. The length‐related differences in eDNA have a substantial potential to improve estimation of species biomass.     

Effect of oxygen and temperature on the dynamic of the dominant bacterial populations of pig manure and on the persistence of pig-associated genetic markers, assessed in river water microcosms

Aims: The aim is to evaluate the dynamic of Bacteroides–Prevotella and Bacillus–Streptococcus–Lactobacillus populations originating from pig manure and the persistence of pig‐associated markers belonging to these groups according to temperature and oxygen. Methods and Results:  River water was inoculated with pig manure and incubated under microaerophilic and aerobic conditions, at 4 and 20°C over 43 days. The diversity of bacterial populations was analysed by capillary electrophoresis‐single‐strand conformation polymorphism. The persistence of the pig‐associated markers was measured by real‐time PCR and compared with the survival of Escherichia coli and enterococci. Decay was characterized by the estimation of the time needed to produce a 1‐log reduction (T90). The greatest changes were observed at 20°C under aerobic conditions, leading to a reduction in the diversity of the bacterial populations and in the concentrations of the Pig‐1‐Bac, Pig‐2‐Bac and Lactobacillus amylovorus markers with a T90 of 10·5, 8·1 and 17·2 days, respectively. Conclusions: Oxygen and temperature were found to have a combined effect on the persistence of the pig‐associated markers in river waters. Significance and Impact of the Study: The persistence profiles of the Pig‐1‐Bac, Pig‐2‐Bac and Lact. amylovorus markers in addition to their high specificity and sensitivity support their use as relevant markers to identify pig faecal contamination in river waters.     

环境温度对白头鹎代谢产热和蒸发失水的影响

基础代谢率(BMR)是恒温动物维持正常生理机能的最小产热速率,是动物在清醒时维持身体各项基本功能所需的最小能量值。"能量需求"假设认为,鸟类短期BMR的调整可以通过改变内部器官的大小来适应能量需求。以白头鹎(Pycnonotus sinensis)为研究对象,分别在10℃和30℃的不同环境中适应4周后,测定了其BMR、蒸发失水(EWL)和内部器官(心、肝、肾、胃、小肠和整体消化道)的重量,同时分析了白头鹎内部器官的变化及与BMR的关系。驯化4周后,白头鹎暖温组(30℃)的体重有明显降低的趋势;经协方差校正体重后,低温组(10℃)单位体重BMR与整体BMR明显高于暖温组,同时两组间EWL表现出显著差异。协方差分析表明,低温组白头鹎的肝脏、肾脏、小肠及总消化道重量显著高于暖温组。低温引起白头鹎能量需求增加,内部器官发生相应改变,要求摄入更多的氧气以维持代谢平衡,导致散失更多的蒸发水。     

Effect of thermo-velocity barriers on fish: influence of water temperature,flow velocity and body size on the volitional swimming capacity of northern straight-mouth nase (Pseudochondrostoma duriense)

Water temperature and flow velocity directly affect the fish swimming capacity, and thus, both variables influence the fish passage through river barriers. Nonetheless, their effects are usually disregarded in fishway engineering and management. This study aims to evaluate the volitional swimming capacity of the northern straight-mouth nase (Pseudochondrostoma duriense), considering the possible effects of water temperature, flow velocity and body size. For this, the maximum distance, swim speed and fatigue time (FT) were studied in an outdoor open-channel flume in the Duero River (Burgos, Spain) against three nominal velocities (1.5, 2.5 and 3 m s⁻¹) and temperatures (5.5, 13.5 and 18.5°C), also including the changes between swimming modes (prolonged and sprint). Results showed that a nase of 20.8 cm mean fork length can develop a median swim speed that exceeds 20.7 BL s⁻¹ (4.31 m s⁻¹) during a median time of 3.4 s in sprint mode, or 12.2 BL s⁻¹ (2.55 m s⁻¹) for 23.7 s in prolonged mode under the warmest scenario. During prolonged swimming mode, fish were able to reach further distances in warmer water conditions for all situations, due to a greater swimming speed and FT, whereas during sprint mode, warmer conditions increased the swim speed maintaining the FT. In conclusion, the studied temperature range and flow velocity range influence fish swimming performance, endurance and distance travelled, although with some differences depending on the swimming mode. The provided information goes a step forward in the definition of real fish swimming capacities, and in turn, will contribute to establish clear passage criteria for thermo-velocity barriers, allowing the calculation of the proportion of fish able to pass a barrier under different working scenarios, as well designing of the optimized solutions to improve the fish passage through river barriers.     

Amphibious adaptations in a newly recognized amphibious fish: Terrestrial locomotion and the influences of body size and temperature

Amphibious animals are adapted for both aquatic and terrestrial habitats. The conflicting requirements for dual habitats are perhaps most pronounced in the air‐breathing fishes, which represent an intermediate stage between the totally aquatic habitat and terrestrial colonization. A key requirement for amphibious fishes is terrestrial locomotion. The different densities and compositions of air and water impose constraints for efficient terrestrial locomotion that differ from those required for aquatic locomotion. I investigated terrestrial locomotion in a small South African fish, Galaxias ‘nebula’, by exposing 60 individual fish to air in specially designed raceways and quantifying movement type and occurrence as a function of availability of water, fish size and environmental temperature. Nebula showed a sustained undulating form of terrestrial locomotion characteristic of amphibious fishes and also a transient ballistic locomotion (jumps) typical of fully aquatic species. Terrestrial movement was influenced by fish size, with medium‐sized fish undertaking more jumps towards water, and fewer jumps away from water, than their smaller or larger conspecifics. In contrast, axial undulation was mainly influenced by temperature. However, there was no consistent pattern in temperature effects presumably because temperature is just one of a suit of environmental factors that may affect terrestrial locomotion. Nebula's amphibious adaptations allow it to cope with the unpredictability inherent in its natural environment.     

Residual DNA analysis in biologics development: review of measurement and quantitation technologies and future directions

Residual DNA (rDNA) is comprised of deoxyribonucleic acid (DNA) fragments and longer length molecules originating from the host organism that may be present in samples from recombinant biological processes. Although similar in basic structural base pair units, rDNA may exist in different sizes and physical forms. Interest in measuring rDNA in recombinant products is based primarily on demonstration of effective purification during manufacturing, but also on some hypothetical concerns that, in rare cases, depending on the host expression system, some DNA sequences may be potentially infectious or oncogenic (e.g., HIV virus and the Ras oncogene, respectively). Recent studies suggest that a sequence known as long interspersed nucleotide element-1 (LINE-1), widely distributed in the mammalian genome, is active as a retrotransposon that can be transcribed to RNA, reverse-transcribed into DNA and inserts into a new site in genome. This integration process could potentially disrupt critical gene functions or induce tumorigenesis in mammals. Genomic DNA from microbial sources, on the other hand, could add to risk of immunogenicity to the target recombinant protein being expressed, due to the high CpG content and unmethylated DNA sequence. For these and other reasons, it is necessary for manufacturers to show clearance of DNA throughout production processes and to confirm low levels in the final drug substance using an appropriately specific and quantitative analytical method. The heterogeneity of potential rDNA sequences that might be makes the testing of all potential analytes challenging. The most common methodology for rDNA quantitation used currently is real-time polymerase chain reaction (RT-PCR), a robust and proven technology. Like most rDNA quantitation methods, the specificity of RT-PCR is limited by the sequences to which the primers are directed. To address this, primase-based whole genome amplification is introduced herein. This paper will review the recent advancement in rDNA quantitation and recent findings regarding potential risks of immunogenicity, infectivity, and oncogenicity of rDNA.     

The influence of temperature and habitat on the distribution of chiselmouth, Acrocheilus alutaceus, in British Columbia

We intensively sampled fish in rivers and streams within a single major drainage basin (the Blackwater River) and across major drainages in British Columbia to assess the factors influencing distribution of chiselmouth, Acrocheilus alutaceus, and to develop models for predicting chislemouth presence. Chiselmouth were typically absent from sites with maximum temperatures below 20°C or 2100 annual degree days, both within a single drainage and between larger drainages. Indices of stream size (bankfull channel width and basin area) were the most significant predictors of chiselmouth presence within the Blackwater drainage (p=0.016 and p=0.032, respectively), and inclusion of thermal variables only marginally increased classification success. In contrast, bankfull channel width and basin area were poor predictors of chiselmouth presence in mainstem habitat within larger drainage basins throughout British Columbia. Inclusion of thermal variables (particularly degree days > 12°C) doubled correct classification rates of chiselmouth presence across larger drainage basins. These habitat associations suggest that water temperature is the primary constraint on presence of chiselmouth populations in larger drainages across a landscape, while selection of different habitat types (mainstem habitat over smaller tributaries) determines distribution within any given basin.     

The effects of temperature, pH and water hardness on winter starvation of young-of-the-year smallmouth bass, Micropterus dolomieui Lacepede

Year-class strength in northern populations of smallmouth bass is strongly influenced by winter starvation of young-of-the-year. We examined starvation among young bass under both winter and summer light and temperature conditions. During starvation, body condition declines to a specific level and then the fish dies. Body condition at death is a well defined function of body size that remains relatively constant over a wide range of environmental conditions. Starvation rate varies systematically with body size, temperature, pH and water hardness. Available stored energy increases more rapidly with body size than starvation rate. Therefore, lifetime under starvation conditions tends to increase with increasing body size. The Q₁₀ for starvation rate over the temperature range 2.5-8° C is 2.2. Starvation rate increases as pH declines from 7.0-4.9: the rate at pH 4.9 is ∼ 1.25 times the rate at pH 7 Starvation rate decreases as Ca concentration increases from 1 mgl⁻¹ to 80mgl⁻¹: the rate at 80 mg Ca 1⁻¹ is ∼0.80 times the rate at 1 mgl⁻¹.     

Modelling the effect of temperature, water activity and pH on the growth of Serpula lacrymans

Aims: To predict the risk factors for building infestation by Serpula lacrymans, which is one of the most destructive fungi causing timber decay in buildings. Methods and Results: The growth rate was assessed on malt extract agar media at temperatures between 1·5 and 45°C, at water activity (a_w) over the range of 0·800–0·993 and at pH ranges from 1·5 to 11·0. The radial growth rate (μ) and the lag phase (λ) were estimated from the radial growth kinetics via the plots radius vs time. These parameters were then modelled as a function of the environmental factors tested. Models derived from the cardinal model (CM) were used to fit the experimental data and allowed an estimation of the optimal and limit values for fungal growth. Optimal growth rate occurred at 20°C, at high a_w level (0·993) and at a pH range between 4·0 and 6·0. The strain effect on the temperature parameters was further evaluated using 14 strains of S. lacrymans. The robustness of the temperature model was validated on data sets measured in two different wood‐based media (Quercus robur L. and Picea abies). Conclusions: The two‐step procedure of exponential model with latency followed by the CM with inflection gives reliable predictions for the growth conditions of a filamentous fungus in our study. The procedure was validated for the study of abiotic factors on the growth rate of S. lacrymans. Significance and Impact of the Study: This work describes the usefulness of evaluating the effect of physico‐chemical factors on fungal growth in predictive building mycology. Consequently, the developed mathematical models for predicting fungal growth on a macroscopic scale can be used as a tool for risk assessment of timber decay in buildings.     

Effects of particle size distribution,compositional and color properties of ground corn on quality of distillers dried grains with solubles (DDGS)

Oftentimes, corn processors believe that ground corn (raw material) and distillers dried grains with solubles (DDGS) are interrelated in certain quality parameters. Yet, previous studies, although rather limited, have not established this relationship. In this study, six ground corn samples and their resulting DDGS were analyzed for particle size distribution (PSD), using a series of six selected US standard sieves: Nos. 8, 12, 18, 35, 60, and 100, and a pan. The original sample and sieve sized fractions were measured for contents of moisture, protein, oil, ash and starch, and surface color. Total carbohydrate (CHO) and total non-starch CHO were also calculated. Results show that the geometric mean diameter (d_gw) of particles varied with individual corn and DDGS samples, and that d_gw of DDGS was larger than that of corn (0.696 vs. 0.479 mm, average values), indicating that during conversion of corn to DDGS, certain particles became enlarged. For d_gw and mass frequency of individual particle size classes, the relationship between ground corn and DDGS varied, but PSD of the whole sample was well correlated between them (r = 0.807). Upon conversion from corn to DDGS, on an average, protein was concentrated 3.59 times; oil, 3.40 times; ash, 3.32 times; and total non-starch CHO, 2.89 times. There were some positive correlations in contents of protein and non-starch CHO and in L value between corn and DDGS. Yet, variations in nutrients and color attributes were larger in DDGS than in corn. For either corn or DDGS, these variations were larger in sieved fractions than in the whole fraction. Raw material, processing method and addition of yeasts are among major factors considered for causing larger variations in these attributes among DDGS. The study partially supports the common belief by processors that quality attributes of corn affect those of DDGS.     

Effect of water activity and temperature on germination and growth of Penicillium digitatum,P. italicum and Geotrichum candidum

AIMS: This study compares the effect of temperature (4-37 degrees C) and water activity (aw: 0.99-0.87) and their interactions on the germination rates, lag times prior to germination and mycelial growth 'in vitro' of Penicillium digitatum, P. italicum and Geotrichum candidum, the main postharvest pathogens affecting citrus fruits. METHODS AND RESULTS: Germination and growth were markedly influenced by temperature and aw. Generally, lag times were longer and germination and growth rates were slower when conditions of temperature and aw were far from optimum. All the studied species were able to germinate over a range of 4-30 degrees C at 0.995 aw, although in non-optimal conditions P. digitatum only reached 40-60% of germinated conidia. At low temperatures, P. italicum germinated and grew faster than P. digitatum and G. candidum, particularly at 0.95 aw. Penicillium italicum was also able to germinate and grow in the driest studied conditions (0.87 aw), while G. candidum did not germinate under 0.95 aw. CONCLUSIONS: Knowledge of the ecological requirements of these fungi is important in order to understand their behaviour in natural situations and to predict fungal spoilage on citrus fruits.     

Effect of springtime water temperature on the time of emergence and size of Pteronarcys californica in the Henry's Fork catchment, Idaho, U.S.A.

1. The emergence time of Pteronarcys californica in streams in the Henry's Fork catchment, Idaho, U.S.A. was negatively correlated with mean April water temperature. Emergence was in mid- to late May at sites influenced by groundwater, where April water temperature averaged 7.9 °C. Adults emerged in mid-June in streams receiving run-off from snowmelt (mean April water temperature 5.4 °C). Intermediate emergence times were observed in a regulated section of river where water temperature was influenced, on one bank, by dam release (mean April water temperature 4.5 °C) and, on the other, by a spring-fed tributary stream (mean April water temperature 6.3 °C).
2. During each of the three study years, emergence was earlier on the bank of the regulated section that was warmer during April and May. The mean body length of P. californica exuviae, collected from the warm side of the river, averaged 1.2 mm longer than those collected from the cold side.
3. We tested the effect on emergence of altering springtime water temperature by translocating P. californica in cages from one location to another during April. Individuals moved to sites with higher April water temperature emerged earlier than individuals that remained at the site from which they were collected.     

浙江南麂列岛大檑山屿水仙自然居群的物种多样性、环境解释及空间分布格局分析

在野外调查基础上,对浙江南麂列岛大檑山屿水仙(Narcissus tazetta var.chinensis M.Roem.)自然居群5个典型生境(乔木林地、林缘、弃耕地、荒坡和灌木林地)的物种组成和α多样性指数进行分析,并采用典范对应分析(CCA)、最小生成树(MST)分析和空间点格局分析(SPPA)分别研究水仙自然居群物种分布与主要环境因子的关系、水仙与其伴生种的种间关系及空间分布格局.结果表明:浙江南麂列岛大檑山屿水仙自然居群5个生境共有植物28种,其中,乔木林地、林缘、弃耕地、荒坡和灌木林地分别有16、13、9、8和8种.5个生境中水仙的重要值均最高.弃耕地和乔木林地的Shannon-Wiener多样性指数较高,灌木林地和林缘其次,荒坡最低.5个生境的聚类分析结果显示:弃耕地和乔木林地聚为一组,其他3个生境聚为另一组.CCA排序结果显示:水仙的分布与坡度呈负相关,与土壤的电导率、含水量和温度以及坡向和海拔呈正相关,说明水仙适宜生长于坡度平缓以及土壤的含水量、电导率和温度适中的生境.MST分析结果显示:水仙与其伴生种羊蹄(Rumex japonicus Houtt.)、鬼针草(Bidens pilosa Linn.)、野艾蒿(Artemisia lavandulifolia DC.)的种间关系最近,这3个伴生种应作为水仙自然居群恢复生物治理的重点防控对象.SPPA分析结果显示:荒坡中水仙群丛分布数量相对较多,集中分布于东北角,西部和南部较少;在0.0~0.1 m尺度时,水仙群丛表现为随机分布;在大于0.1 m尺度时,表现为集中分布.研究结果显示:浙江南麂列岛大檑山屿水仙自然居群主要集中分布在东南坡的弃耕地,海岛生境异质性与自身繁殖特性是影响南麂列岛水仙自然居群恢复的关键因素,建议对水仙群落动态进行长期监测和相关研究,并结合岛屿生态系统进行综合管理.     

Genomic structure, polymorphism and expression analysis of the growth hormone (GH) gene in female and male Half-smooth tongue sole (Cynoglossus semilaevis)

Growth hormone (GH) is a polypeptide which is an important regulator of development and somatic growth in teleosts, and may be associated with the mechanisms which drive sexual growth dimorphism in the Half-smooth tongue sole (Cynoglossus semilaevis). In this study, the full length gh cDNA was cloned from C. semilaevis by homology cloning and the rapid amplification of cDNA ends polymerase chain reaction (RACE-PCR). The full-length gh cDNA is 826 bp and contains an open reading frame (ORF) of 603 bp encoding a protein of 200 amino acids (AA). The precursor of gh consists of a 17 amino-acid signal peptide followed by a 183 amino-acid mature polypeptide. GH gene sequences obtained from female and male adults consist of 3428 bp and 3371 bp, respectively, each of which includes six exons and five introns, and the difference in the GH gene size was mainly caused by the microsatellites. When 14 tissues from females, normal males and extra-large male adults were analyzed for sex-specific tissue expression, the gh mRNA was found to be predominantly expressed in the pituitary, and the expression levels in females were 3.6 times as much as those in normal males, while the mRNA expression in extra-large males was 1.7 times as much as those in normal males. Sex differences in gh mRNA expression during development were also examined by using a full-sib family of C. semilaevis, and the gh mRNA was detected at all of the 12 time points sampled from 10 to 380 days-old. A significant increase in gh mRNA was detected starting in 80 day old fish and was then followed by a drop to very low levels starting at 230 day old fish. Differential expression indicated that the gh expression level in females was significantly higher than males (P < 0.01) at all of the stages except for 10 days-old. Two microsatellite loci were identified in the second intron of the GH gene. Using these two polymorphic markers to genotype 224 individuals, there was no significant difference between the females and males in the Bohai Sea, the Yellow Sea and the hatchery samples.