Comparison of traditional and environmental DNA survey methods for detecting rare and abundant freshwater fish

1. Detecting rare species is often a necessity for conservation and management, yet challenging for many field survey methods. Environmental DNA (eDNA) is a highly promising solution that has been shown to outperform many established survey methods.
2. Macquarie perch (Macquaria australasica) is an endangered native species that has declined significantly in range and abundance. Detection of M. australasica was compared with an abundant alien fish species (Oncorhynchus mykiss) using eDNA and three conventional survey methods: gill nets, electrofishing and fyke nets.
3. eDNA occupancy estimates for both fish species were compared using four different models to investigate what effect these differences have on false positives and false negatives for the rare and common fish species. These models used unadjusted eDNA detections in water samples, eDNA detections that have been screened using a limit of detection method to remove potential false positives, eDNA data supplemented with a second survey method, or eDNA data augmented with sequencing of positive polymerase chain reaction replicates.
4. eDNA surveying as a single detection method was found to be more efficient and sensitive compared with each capture method separately and combined. Occupancy estimates for the common and rare species did not vary significantly between the four site occupancy-detection models, suggesting that supplementary data may not have as much effect on occupancy estimates compared with other approaches such as temporal or spatial sampling.
5. We conclude that eDNA outperforms the three established survey methods for both a rare and common freshwater fish species. Although there was no significant effect of augmenting eDNA survey methods with other survey data, additional data may improve confidence in detection, and provide confirmatory evidence for unexpected or new detections of a species.

     

检测cDNA芯片中差异表达基因的非转换方法

cDNA芯片已经成为生物学试验中一种重要试验手段,与其相应的检测差异表达基因的统计方法也在快速地完善。目前通用的方法是将数据进行对数比转换,并相应进行标准化处理。鉴于对数转换的缺点,本研究提出一种非转换方法,即背景校正后直接进行数据标准化。研究表明:利用这种非转换方法,可以更有效地剔除试验中的“噪音,”提高检测的准确率。本研究利用Apo AI试验的芯片数据进行了效率验证,结果表明:与对数转换方法相比,非转换方法能检测出更多的差异表达基因,可以作为cDNA芯片数据分析的一种简便高效方法。     

Non‐indigenous fish in protected spaces: Trends in species distribution mediated by illegal stocking

1. Many freshwater non‐indigenous species (NISs) are stocked for recreational fishing, in some cases illegally in protected areas. In this study, fish communities were monitored using environmental DNA, electrofishing and anglers’ catches as the sources of samples in a mountainous Biosphere Reserve in Asturias (northern Spain), where stocking is forbidden.
2. Three NISs have been introduced illegally in the protected area and have shown increasing populations in the last two decades. Two species used as fishing bait, Squalius carolitertii (chub) and Phoxinus phoxinus (minnow), are expanding in running waters. Oncorhynchus mykiss (rainbow trout) was also detected and is likely to have been introduced for angling or from fish farm escapes.
3. The results suggest that sustained illegal stocking contributed to the increase of the three NISs. In contrast, Salmo trutta (brown trout) of northern European lineages, identified from *90 alleles at the LDH‐C1 locus, and formerly legally stocked for angling, is decreasing, most likely as a result of climate change. Climate change could also contribute to the expansion of the two non‐indigenous cyprinids to colder upstream areas.
4. Through the application of a social survey, it was found that unlike other population groups, anglers in the region significantly preferred stocking over environmental improvement for the management of fish populations. The results obtained suggest that raising the awareness of anglers about the importance of safeguarding native fish species could help to prevent the spread of NISs in protected areas.

     

Rhizosphere microbiota assemblage associated with wild and cultivated soybeans grown in three types of soil suspensions

Soil microbial community composition is determined by the soil type and the plant species. By sequencing the V3-V4 region of the bacterial 16S rRNA gene amplicons, the current study assessed the bacterial community assemblage in rhizosphere and bulks soils of wild (Glycine soja) and cultivated (Glycine max) soybeans grown in the suspensions of three important soil types in China, including black, red and soda-saline-alkali soils. The alpha-diversity of the bacterial community in the rhizosphere was significantly higher than that of the bulk soils suggesting that bulk soil lacks plant nurturing effect under the current study conditions. Black and red soils were enriched with nitrifying and nitrogen-fixing bacteria but the soda-saline-alkali soil suspension had more denitrifying bacteria, which may reflect agronomic unsuitability of the latter. We also observed a high abundance of Bradyrhizobium and Pseudomonas, enriched cellulolytic bacteria, as well as a highly connected molecular ecological network in the G. soja rhizosphere soil. Taken all, the current study suggest that wild soybeans may have evolved to recruit beneficial microbes in its rhizosphere that can promote nutrients requisition, biostasis and disease-resistance, therefore ecologically more resilient than cultivated soybeans.     

Environmental DNA as a monitoring tool for the endangered freshwater pearl mussel (Margaritifera margaritifera L.): a substitute for classical monitoring approaches?

1. Knowledge of species distribution is of utmost importance for conservation and management of endangered freshwater mussels. Conventional monitoring approaches are often time consuming and costly. The use of environmental DNA (eDNA) is a relatively new approach and considered as an effective tool to detect the presence of target species in aquatic environments. The aim of this study was to establish a nested PCR system to detect eDNA of Margaritifera margaritifera and to discuss the advantages and disadvantages of eDNA in mussel surveys compared with classical monitoring.
2. DNA of M. margaritifera was detected in 2 L water samples collected directly downstream (25 m) from pearl mussel populations (population size from 800–20 000), with an internal‐nested PCR approach greatly increasing the detection sensitivity (down to 10 fg target DNA). eDNA detection at greater distances downstream (500 and 1 000 m) of these populations failed, possibly due to DNA degradation or dilution processes. eDNA was also detected downstream of an extinct population, most likely resulting from overlooked mussels or the release of DNA from dead shells.
3. The eDNA approach proposed herein may be helpful in initial screening of streams that are otherwise difficult to monitor, or in the detection of buried juvenile mussels without disturbing their habitat. However, it cannot replace monitoring of population demography, and of other important information for conservation, and should thus only be seen as a supplementary tool.

Copyright © 2015 John Wiley & Sons, Ltd.     

Simultaneous analysis of the intestinal parasites and diet through eDNA metabarcoding

Agricultural expansion and intensification are having a huge impact on plant and arthropod diversity and abundance, affecting food availability for farmland birds. Difficult food access, in turn, can lead to immunosuppression and a higher incidence of parasites. In the studies designed to examine changes in the diet of birds and their parasites, metabarcoding is proving particularly useful. This technique requires mini-barcodes capable of amplifying the DNA of target organisms from fecal environmental DNA. To help to understand the impact of agricultural expansion on biodiversity, this study sought to design and identify mini-barcodes that might simultaneously assess diet and intestinal parasites from the feces of farmland birds. The capacity to identify diet and parasites of 2 existing and 3 newly developed mini-barcodes was tested “in silico” in relation to the behavior of a reference eukaryotic barcode. Among the newly designed mini-barcodes, MiniB18S_81 showed the higher taxonomic coverage of eukaryotic taxa and a greater amplification and identification capacity for diet and parasite taxa. Moreover, when it was tested on fecal samples from 5 different steppe bird species, MiniB18S_81 showed high taxonomic resolution of the most relevant diet and parasite phyla, Arthropoda, Nematoda, Platyhelminthes, and Apicomplexa at the order level. Thus, the mini-barcode developed emerges as an excellent tool to simultaneously provide detailed information regarding the diet and parasites of birds, essential for conservation and management.     

烤烟品种K326茎叶消减文库的构建及质量分析

利用抑制性消减杂交(SSH)技术,构建烤烟品种K326的茎叶抑制性消减cDNA文库.结果表明,文库的滴度为1.76×106cfu.mL-1,重组率为75%,随机扩增480个克隆,显示插入片段长度在200-1000 bp之间,符合建库的质量标准.     

Improving the detection of rare native fish species in environmental DNA metabarcoding surveys

1. The presence of threatened or endangered species often strongly influences management and conservation decisions. Within the Murray–Darling Basin (MDB), Australia, the presence of threatened native fish affects the management and allocation of water resources. In New South Wales, these decisions are currently based on traditional fisheries data and a predictive MaxEnt model. However, it is important to verify the model's predictive power given the implication it may have, but this requires methods with a high detection sensitivity for rare species.
2. Although the use of environmental DNA (eDNA) monitoring, in particular eDNA metabarcoding, achieves a higher detection sensitivity compared with traditional methods, earlier surveys in the MDB have shown that the highly abundant and invasive common carp (Cyprinus carpio) can reduce detection probabilities for rare species. Consequently, a polymerase chain reaction (PCR) blocking primer designed to block the amplification of carp eDNA could increase the detection probabilities for rare native species while simultaneously reducing the required sampling effort and survey costs. Although PCR blocking primers are often used in ancient DNA and dietary studies, no aquatic eDNA metabarcoding study to date has evaluated the potential benefits of using PCR blocking primers.
3. A laboratory and field-based pilot study was used to address this knowledge gap and assess the impact of a blocking primer, targeting cyprinid fishes (including carp), on the detection probabilities of native species and the minimum sampling effort required.
4. Results showed that the inclusion of the blocking primer increased the detection probabilities for native species by 10–20% and reduced the minimum required sampling effort by 25–50%. These findings provide important insights into possible methods for optimizing eDNA metabarcoding surveys for the detection of rare aquatic species.

     

Abstracts of Nippon Dojohiryogaku Zasshi

Abstract

The humus composition was analyzed and the humic acid characterized by UV and visible absorption spectroscopy in order to investigate the rotting and maturing process of city refuse compost according to the method of Kumada et al. During the composting process, the following findings were obtained: (1) the HT value was almost constant, but the HE/HT ratio varied somewhat, (2) HA increased with decrease in FA, and the PQ value so increased clearly, (3) the shoulder-like absorption at a wavelength near 270 nm weakened, and (4) the RF value of humic acid increased, whereas the Δ log K value seldom varied.

The IR spectrum of humic acid gradually changed as follows: (1) the absorption band in the 1700-1600 cm^-1 region and in the 1550-1500 cm^-1 region increased slightly, (2) the band in the 1100-1000 cm^-1 region decreased, and (3) the bands at 835 and 710 cm^-1 com pletely disappeared. On the whole, the shape of the IR spectrum of the city refuse compost became featureless. These changes were probably due to the oxidation which occurred in the composting process.