Filtration extraction method using microfluidic channel for measuring environmental DNA

The environmental DNA (eDNA) method, which is widely applied for biomonitoring, is limited to laboratory analysis and processing. In this study, we developed a filtration/extraction component using a microfluidic channel, Biryu-Chip (BC), and a filtration/extraction method, BC method, to minimize the volume of the sample necessary for DNA extraction and subsequent PCR amplification. We tested the performance of the BC method and compared it with the Sterivex filtration/extraction method using aquarium and river water samples. We observed that using the BC method, the same concentration of the extracted DNA was obtained with 1/20–1/40 of the filtration volume of the Sterivex method, suggesting that the BC method can be widely used for eDNA measurement. In addition, we could perform on-site measurements of eDNA within 30 min using a mobile PCR device. Using the BC method, filtration and extraction could be performed easily and quickly. The PCR results obtained by the BC method were similar to those obtained by the Sterivex method. The BC method required fewer steps and therefore, the risk of DNA contamination could be reduced. When combined with a mobile PCR, the BC method can be applied to easily detect eDNA within 30 min from a few 10 mL of the water sample, even on-site.

Filtration extraction method using microfluidic channel for measuring environmental DNA

The environmental DNA (eDNA) method, which is widely applied for biomonitoring, is limited to laboratory analysis and processing. In this study, we developed a filtration/extraction component using a microfluidic channel, Biryu-Chip (BC), and a filtration/extraction method, BC method, to minimize the volume of the sample necessary for DNA extraction and subsequent PCR amplification. We tested the performance of the BC method and compared it with the Sterivex filtration/extraction method using aquarium and river water samples. We observed that using the BC method, the same concentration of the extracted DNA was obtained with 1/20-1/40 of the filtration volume of the Sterivex method, suggesting that the BC method can be widely used for eDNA measurement. In addition, we could perform on-site measurements of eDNA within 30 min using a mobile PCR device. Using the BC method, filtration and extraction could be performed easily and quickly. The PCR results obtained by the BC method were similar to those obtained by the Sterivex method. However, the BC method required fewer steps and therefore, the risk of DNA contamination could be reduced. When combined with a mobile PCR, the BC method can be applied to easily detect eDNA within 30 min from 10 mL of the water sample, even on-site.

Level and distribution of heavy metals in Miri River, Malaysia

Heavy metal pollution in water resources has become a serious and hazardous environmental problem all over the world because of its non-biodegradability, emanating from multiple sources, easy accumulation, and biological toxicity. This research was carried out to study the level and distribution of heavy metals at different sampling locations (upstream, midstream, and downstream), at different depths (0.5 m and 1.5 m from surface water level), and during low tide and high tide conditions in the Miri River of Miri City in Malaysia. The river water samples were collected and analyzed for Ca, Mg, Cu, Fe, Mn, Ni, Pb, and Zn by flame atomic absorption spectrophotometer. The concentration of Ca was found to be the highest in the Miri River, followed by Mg and Fe, and with traces of Cu, Mn, Ni, Pb, and Zn. An increase in the concentration of heavy metals, such as Cu, Mg, and Ni, was observed while flowing from upstream to downstream of the Miri River. Concentrations of heavy metals, such as Ca, Mg, Cu, and Zn, were clearly lower at 1.5 m depth than at 0.5 m depth. High tides in the river decrease the concentration of heavy metals, such as Ca, Cu, Mn, and Ni, compared to low tides. From this research, it gets clear that using the Miri River water for domestic and recreational purposes, washing, and fishing is detrimental to human health and the environment.

Correction to: Determination of organically bound fluorine sum parameters in river water samples—comparison of combustion ion chromatography (CIC) and high resolution-continuum source-graphite furnace molecular absorption spectrometry (HR-CS-GFMAS)

A Correction to this paper has been published: 10.1007/s00216-021-03675-z

Concentration and quantification of Tilapia tilapinevirus from water using a simple iron flocculation coupled with probe-based RT-qPCR

Tilapia tilapinevirus (also known as tilapia lake virus, TiLV) is an important virus responsible for die-off of farmed tilapia globally. Detection and quantification of the virus from environmental DNA/RNA (eDNA/eRNA) using pond water represents a potential, noninvasive routine approach for pathogen monitoring and early disease forecasting in aquaculture systems. Here, we report a simple iron flocculation method for viral concentration from water combined with a newly developed hydrolysis probe quantitative RT-qPCR method for detection and quantification of TiLV. The RT-qPCR method targeting a conserved region of TiLV genome segment 9 has a detection limit of 10 viral copies per uL of template. The method had a 100% analytical specificity and sensitivity for TiLV. The optimized iron flocculation method was able to recover 16.11 +/- 3.3% of virus from water samples spiked with viral cultures. During disease outbreak cases from an open-caged system and a closed hatchery system, both tilapia and water samples were collected for detection and quantification of TiLV. The results revealed that TiLV was detected from both clinically sick fish and asymptomatic fish. Most importantly, the virus was successfully detected from water samples collected from different locations in the affected farms e.g. river water samples from affected cages (8.50 x 102 to 2.79 x 104 copies/L) and fish-rearing water samples, sewage, and reservoir (4.29 x 102 to 3.53 x 103 copies/L) from affected and unaffected ponds of the hatchery. In summary, this study suggests that the eRNA detection system using iron flocculation coupled with probe based-RT-qPCR is feasible for concentration and quantification of TiLV from water. This approach might be useful for noninvasive monitoring of TiLV in tilapia aquaculture systems and facilitating appropriate decisions on biosecurity interventions needed.