First extraction of eDNA from tree hole water to detect tree frogs: a simple field method piloted in Madagascar

AbstractEnvironmental DNA (eDNA) is becoming an increasingly used tool for monitoring cryptic species within terrestrial and aquatic systems. We present the first method for extracting water from tree holes for eDNA studies of tree-dwelling frogs, and the first use of eDNA for amphibian monitoring in Madagascar. This pilot study expands on a previously developed method and aims to provide a simple field protocol for DNA extraction from very small water samples, using a relatively inexpensive kit compared to other collection methods. We collected 20 ml of water from tree holes in Ambohitantely Special Reserve in Madagascar, with the aim to survey for the Critically Endangered tree frog Anodonthyla vallani, and we developed species specific cytochrome c oxidase 1 primers for this species. While our two samples did not detect A. vallani, we successfully extracted up to 16.6 ng/µl of eDNA from the samples and using 16S rRNA primers barcoded the tree frog Plethodontohyla mihanika in one of the samples. Despite just two samples being collected, we highlight the future potential of eDNA from tree holes for investigating cryptic habitat specialist amphibians given we extracted frog eDNA from just 20 ml of water. The method provides a rapid, simple, and cost-effective method which can assist cryptic species monitoring in challenging and time-consuming field conditions and should be developed further for frog surveying in Madagascar and beyond. The newly developed primers can be used for further work using this eDNA method to survey threatened Anodonthyla frog species.

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Towards environmental DNA-based bioassessment of freshwater reservoirs with small volumes of water: robust molecular protocols

10.1101/2021.11.21.469426 ◽

2021 ◽

Author(s):

Rebecca Ker Loh ◽

Sujatha Narayanan Kutty ◽

Darren Chong Jinn Yeo ◽

Rudolf Meier

Keyword(s):

Water Sample ◽

Environmental Dna ◽

Cost Effective ◽

Sampling Time ◽

Small Water ◽

Tropical Reservoirs ◽

Frequent Monitoring ◽

Planktonic Crustaceans ◽

Water Volumes ◽

Dna Template

Bioassessment of freshwater quality via eDNA is rapidly developing into a powerful alternative to traditional methods involving collecting, sorting, and identifying macroinvertebrates based on morphology. Particularly attractive would be methods that can use remote-controlled boats for sampling because it would allow for cost-effective, and frequent monitoring at multiple sites. The latter will be particularly important for tropical reservoirs that require year-around surveillance. We here optimize molecular protocols for capturing reservoir-specific differences in metazoan communities based on small water volumes (15 mL). The optimization is based on samples from two freshwater reservoirs with very different water qualities ("reservoir signal"). Each reservoir was sampled at three sites ("biological replicates"). For each water sample, the DNA was extracted twice ("technical replicates"). We then tested how much DNA template (0.1 ng to 15 ng) and how many PCR cycles (25 or 35) minimized variance between technical replicates. We find that 15 mL is sufficient for capturing the reservoir signal regardless of sampling time, template amounts, or PCR cycle numbers. Indeed, extrapolation from our results suggests that <1 mL would be sufficient because only 17 of 59 metazoan mOTUs (mainly planktonic crustaceans and rotifers) detected with a 313bp COI minibarcode were shared. We find that the use of 35 PCR cycles significantly lowered the number of detected species and that template amounts <0.5 ng yielded somewhat higher variance between technical replicates. Despite extensive trials, the variance between technical replicates remained high (Bray-Curtis: 5-20%; Jaccard: 10-40%) and we predict that it will be difficult to reduce this variance further. However, the overall reservoir differences are so strong that all biological and technical replicates can be correctly assigned.

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The effect of silica desiccation under different storage conditions on filter-immobilized environmental DNA

BMC Research Notes ◽

10.1186/s13104-021-05530-x ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Michael J. Allison ◽

Jessica M. Round ◽

Lauren C. Bergman ◽

Ali Mirabzadeh ◽

Heather Allen ◽

...

Keyword(s):

Silica Gel ◽

Storage Temperature ◽

Environmental Dna ◽

Cost Effective ◽

Storage Conditions ◽

Systematic Evaluation ◽

Silica Bead ◽

Gel Beads ◽

Polymerase Chain

Abstract Objective Silica gel beads have promise as a non-toxic, cost-effective, portable method for storing environmental DNA (eDNA) immobilized on filter membranes. Consequently, many ecological surveys are turning to silica bead filter desiccation rather than ethanol preservation. However, no systematic evaluation of silica bead storage conditions or duration past 1 week has been published. The present study evaluates the quality of filter-immobilized eDNA desiccated with silica gel under different storage conditions for over a year using targeted quantitative real-time polymerase chain reaction (qPCR)-based assays. Results While the detection of relatively abundant eDNA target was stable over 15 months from either ethanol- or silica gel-preserved filters at − 20 and 4 °C, silica gel out-performed ethanol preservation at 23 °C by preventing a progressive decrease in eDNA sample quality. Silica gel filter desiccation preserved low abundance eDNA equally well up to 1 month regardless of storage temperature (18, 4, or − 20 °C). However only storage at − 20 °C prevented a noticeable decrease in detectability at 5 and 12 months. The results indicate that brief storage of eDNA filters with silica gel beads up to 1 month can be successfully accomplished at a range of temperatures. However, longer-term storage should be at − 20 °C to maximize sample integrity.

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Chemical Composition and Thermal Behavior of Kraft Lignins

Forests ◽

10.3390/f10060483 ◽

2019 ◽

Vol 10 (6) ◽

pp. 483 ◽

Cited By ~ 3

Author(s):

Aleš Ház ◽

Michal Jablonský ◽

Igor Šurina ◽

František Kačík ◽

Tatiana Bubeníková ◽

...

Keyword(s):

Molecular Weight ◽

Molecular Weight Distribution ◽

Weight Distribution ◽

Cost Effective ◽

Activation Energies ◽

Industrial Applications ◽

Raw Material ◽

Molecular Weight Determination ◽

Two Samples ◽

Process Measurements

Lignin has great potential for utilization as a green raw material or as an additive in various industrial applications, such as energy, valuable chemicals, or cost-effective materials. In this study, we assessed a commercial form of lignin isolated using LignoBoost technology (LB lignin) as well as three other types of lignin (two samples of non-wood lignins and one hardwood kraft lignin) isolated from the waste liquors produced during the pulping process. Measurements were taken for elemental analysis, methoxyl and ash content, higher heating values, thermogravimetric analysis, and molecular weight determination. We found that the elemental composition of the isolated lignins affected their thermal stability, activation energies, and higher heating values. The lignin samples examined showed varying amounts of functional groups, inorganic component compositions, and molecular weight distributions. Mean activation energies ranged from 93 to 281 kJ/mol. Lignins with bimodal molecular weight distribution were thermally decomposed in two stages, whereas the LB lignin showing a unimodal molecular weight distribution was decomposed in a single thermal stage. Based on its thermal properties, the LB lignin may find direct applications in biocomposites where a higher thermal resistance is required.

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Challenges for assessing vertebrate diversity in turbid Saharan water-bodies using environmental DNA

Genome ◽

10.1139/gen-2018-0071 ◽

2018 ◽

Vol 61 (11) ◽

pp. 807-814 ◽

Cited By ~ 8

Author(s):

Bastian Egeter ◽

Sara Peixoto ◽

José C. Brito ◽

Simon Jarman ◽

Pamela Puppo ◽

...

Keyword(s):

Environmental Dna ◽

Water Bodies ◽

Sampling Point ◽

Sahara Desert ◽

Turbid Water ◽

Vertebrate Species ◽

Sampling Protocol ◽

Filtering Method ◽

Small Water ◽

Human Dna

The Sahara desert is the largest warm desert in the world and a poorly explored area. Small water-bodies occur across the desert and are crucial habitats for vertebrate biodiversity. Environmental DNA (eDNA) is a powerful tool for species detection and is being increasingly used to conduct biodiversity assessments. However, there are a number of difficulties with sampling eDNA from such turbid water-bodies and it is often not feasible to rely on electrical tools in remote desert environments. We trialled a manually powered filtering method in Mauritania, using pre-filtration to circumvent problems posed by turbid water in remote arid areas. From nine vertebrate species expected in the water-bodies, four were detected visually, two via metabarcoding, and one via both methods. Difficulties filtering turbid water led to severe constraints, limiting the sampling protocol to only one sampling point per study site, which alone may largely explain why many of the expected vertebrate species were not detected. The amplification of human DNA using general vertebrate primers is also likely to have contributed to the low number of taxa identified. Here we highlight a number of challenges that need to be overcome to successfully conduct metabarcoding eDNA studies for vertebrates in desert environments in Africa.

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Solar Photovoltaic Water Pumping to Alleviate Drought in Remote Locations

ASME 2006 Power Conference ◽

10.1115/power2006-88215 ◽

2006 ◽

Cited By ~ 1

Author(s):

Kala Meah ◽

Steven Fletcher ◽

Yu Wan ◽

Sadrul Ula

Keyword(s):

Performance Monitoring ◽

Cost Effective ◽

Small Scale ◽

Solar Water ◽

Small Water ◽

Water Pumping ◽

Western Us ◽

Water Right ◽

And Performance ◽

The University

Many parts of the western US is rural in nature and consequently do not have electrical distribution lines in many parts of farms and ranches. Distribution line extension costs can run from $15,000 to $25,000 per mile, thereby making availability of electricity to small water pumping projects economically unattractive. Solar photo-voltaic (PV) powered water pumping is more cost effective in these small scale applications. Many western states including Wyoming are passing through fifth year of drought with the consequent shortages of water for many applications. Wyoming State Climatologist is predicting a possible 5–10 years of drought. Drought impacts the surface water right away, while it takes much longer to impact the underground aquifers. To mitigate the effect on the livestock and wildlife, Wyoming Governor Dave Freudenthal initiated a solar water pumping initiative in cooperation with the University of Wyoming, County Conservation Districts, Rural Electric Cooperatives, and ranching organizations. Solar water pumping has several advantages over traditional systems; for example, diesel or propane engines require not only expensive fuels, they also create noise and air pollution in many remote pristine areas. Solar systems are environment friendly, low maintenance and have no fuel cost. In this paper the design, installation and performance monitoring of the solar system for small scale remote water pumping will be presented.

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The Bionomics of some Tree-hole Mosquitos

Bulletin of Entomological Research ◽

10.1017/s0007485300020927 ◽

1929 ◽

Vol 20 (1) ◽

pp. 45-58 ◽

Cited By ~ 3

Author(s):

Mary V. F. Beattie ◽

Lucy J. Howland

Keyword(s):

Algal Species ◽

Animal Food ◽

Tree Hole ◽

Tree Holes

1. 138 larvae of Anopheles, Finlaya and Orthopodomyia, collected from three tree-holes described in Part I, were dissected and their contents examined.2. Small algae, a few animals, bacteria and considerable quantities of rotting vegetation were found in them.3. The algal and animal food seems to be insufficient for the anabolic processes of the larvae.4. It is possible that bacteria may play some part in their nutrition, especially of Orthopodomyia.5. It is also suggested that the larvae are able to utilise any food constituents present in the vegetable debris.I wish to thank Dr. P. A. Buxton for the interest he has shown during the progress of this work, and Professor F. E. Fritsch for help in identification of the algal species.

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Larval breeding habitats and ecological factors influence the species composition of mosquito (Diptera : Culicidae) in the parks of Dhaka city, Bangladesh

Bangladesh Journal of Zoology ◽

10.3329/bjz.v45i2.35706 ◽

2018 ◽

Vol 45 (2) ◽

pp. 111-122 ◽

Cited By ~ 2

Author(s):

Afroza Sultana ◽

Sharmin Hasan ◽

Mosharraf Hossain ◽

Abdul Alim ◽

Mohammad Al Mamun ◽

...

Keyword(s):

Mosquito Control ◽

Mosquito Species ◽

Ecological Factors ◽

Control Program ◽

Breeding Habitat ◽

Coconut Shell ◽

Larval Abundance ◽

Tree Hole ◽

Breeding Habitats ◽

Tree Holes

Mosquito larval ecology is prerequisite for determining the larval abundance and species assemblage in mosquito control program. The study explored the association of five mosquito species with their breeding habitat diversity and species distribution in three selected parks from May to October, 2015. A total of 3217 mosquito larvae were reported from six breeding habitats, namely tree hole, leaf axils, water bottle, tire, drain and coconut shell. The frequency of the three species (Ae. aegypti, Ae. albopictus and Ar. subalbatus) coexistence was higher in tree holes (82.4%) than that of the other coexisting species. Pearson Chi-square result revealed that the association of species was significantly dependent on the breeding habitats. ANOVA further suggested that mosquito density varied across habitats where among the highest density of Cx. quinquefasciatus (3.87 ± 0.22) found in drain, followed by both Ae. albopictus (2.02 ± 0.17) and Ar. subalbatus (0.50 ± 0.09) in tree holes and Ae. aegypti (1.25 ± 0.23) in coconut shell. Cx. tritaeniorhynchus occurred in drain with the least observed density (0.03 ± 0.01). CCA results suggested that Aedes species were likely preferred to oviposit in shaded habitats where pH was associated with Ae. albopictus and dissolved oxygen was with Ae. aegypti and Ar. subalbatus. Culex species were positively associated with the habitats characterizing muddy bottom and emerging vegetation but not with any of the physico-chemical parameters. These findings concluded that ecological factors influence mosquito species to favor their breeding habitats can be helpful in controlling targeted vector species as well as the mosquito borne diseases.Bangladesh J. Zool. 45(2): 111-122, 2017

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Limited Microcystin, Anatoxin and Cylindrospermopsin Production by Cyanobacteria from Microbial Mats in Cold Deserts

Toxins ◽

10.3390/toxins12040244 ◽

2020 ◽

Vol 12 (4) ◽

pp. 244 ◽

Cited By ~ 1

Author(s):

Nataliia Khomutovska ◽

Małgorzata Sandzewicz ◽

Łukasz Łach ◽

Małgorzata Suska-Malawska ◽

Monika Chmielewska ◽

...

Keyword(s):

Microbial Mats ◽

Sequence Similarity ◽

Extreme Environment ◽

Toxin Production ◽

High Mountain ◽

Rrna Gene ◽

High Sequence Similarity ◽

Small Water ◽

Two Samples ◽

Liquid Chromatography Tandem Mass

Toxic metabolites are produced by many cyanobacterial species. There are limited data on toxigenic benthic, mat-forming cyanobacteria, and information on toxic cyanobacteria from Central Asia is even more scarce. In the present study, we examined cyanobacterial diversity and community structure, the presence of genes involved in toxin production and the occurrence of cyanotoxins in cyanobacterial mats from small water bodies in a cold high-mountain desert of Eastern Pamir. Diversity was explored using amplicon-based sequencing targeting the V3-V4 region of the 16S rRNA gene, toxin potential using PCR-based methods (mcy, nda, ana, sxt), and toxins by enzyme-linked immunosorbent assays (ELISAs) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Molecular identification of cyanobacteria showed a high similarity of abundant taxa to Nostoc PCC-73102, Nostoc PCC-7524, Nodularia PCC-935 and Leptolyngbya CYN68. The PCRs revealed the presence of mcyE and/or ndaF genes in 11 samples and mcyD in six. The partial sequences of the mcyE gene showed high sequence similarity to Nostoc, Planktothrix and uncultured cyanobacteria. LC-MS/MS analysis identified six microcystin congeners in two samples and unknown peptides in one. These results suggest that, in this extreme environment, cyanobacteria do not commonly produce microcystins, anatoxins and cylindrospermopsins, despite the high diversity and widespread occurrence of potentially toxic taxa.

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The tadpole of Ecnomiohyla sukia Savage & Kubicki, 2010 (Amphibia: Hylidae)

Zootaxa ◽

10.11646/zootaxa.2793.1.6 ◽

2011 ◽

Vol 2793 (1) ◽

pp. 63

Author(s):

HEINZ HOFFMANN ◽

BRIAN KUBICKI

Keyword(s):

Costa Rica ◽

Larval Development ◽

Tree Frog ◽

Adult Frog ◽

Tree Hole ◽

Adult Tree ◽

The Village

Near the village of Guayacán de Siquirres, Limón province, Costa Rica (N 10°02”, W 83°31’) the call of a frog revealed a breeding hole about 15 m high in a Guarea tree. An adult was taken from this water-filled cavity along with some tadpoles. The adult tree frog was identified by B. Kubicki as Ecnomiohyla sukia Savage and Kubicki, 2010. Its sex was not determined. Ten tadpoles were extracted from the water-filled tree hole and it appeared they had only a few days of larval development, but the exact age was undeterminable. The conspecificity of the tadpoles found with the adult frog E. sukia was established by rearing them to metamorphic stages and froglets. The following description is based on tadpoles from stages 25, 36–37, and 41 (Gosner 1960).

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Transparency is in the eye of the beholder: the effects of identity and negative perceptions on ratings of transparency via surveys

International Review of Administrative Sciences ◽

10.1177/0020852315615197 ◽

2016 ◽

Vol 83 (1_suppl) ◽

pp. 177-194

Author(s):

Heungsik Park ◽

John Blenkinsopp

Keyword(s):

Cost Effective ◽

Negative Influence ◽

Public Employees ◽

Survey Period ◽

Factor Solution ◽

Public Servants ◽

Cost Effective Approach ◽

Two Samples ◽

Negative Perceptions ◽

Survey Responses

Surveys are a commonly used means of measuring transparency levels, but they are potentially vulnerable to perceptual biases. This study sought to examine perceptual differences by the respondents’ identities as general citizens or public employees, and the possible negative perceptions that one group may have of the other concerning responses to a survey-based measure of transparency. The survey was designed on the basis of existing literature, suggesting that transparency has up to six facets. Two samples were taken: from citizens who visited district offices to file civil applications during the survey period; and from public employees involved in processing these applications. A total of 472 surveys were used for analysis: 233 citizens and 239 public employees. The results indicated that the two groups had different understandings of transparency. Data from public employees produced a three-factor solution, which was labeled as Efficiency, Reliability, and Access. For citizens, a two-factor solution was a better fit, with the factors being described as Accessibility (a wider notion than Access) and Utility. The findings suggest that public employees adopt a somewhat technical view of transparency, whereas citizens have more practical concerns about it. Only citizens’ unfavorable perception of public employees had a negative influence on the level of transparency. This study contributes to the understanding of how public employees and citizens have qualitatively different perceptions of transparency. Points for practitioners To assess progress in governmental transparency, we must measure it, and surveys offer an accessible and potentially cost-effective approach. However, the survey responses of citizens and public employees show that they understand transparency in qualitatively different ways, with citizens’ perceptions of transparency also influenced by their perceptions of public employees. If governments are to increase public trust in policymaking and administration, they must focus on improving transparency as it is understood by the public rather than how it is understood by public servants.

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