Differential detection of shrimp and crab for food labeling using polymerase chain reaction

Shrimp and crab are well-known as allergenic ingredients. According to Japanese food allergy labeling regulations, shrimp species (including prawns, crayfishes, and lobsters) and crab species must be differentially declared when ≥10 ppm (total protein) of an allergenic ingredient is present. However, the commercial ELISA tests for the detection of crustacean proteins cannot differentiate between shrimp and crab. Therefore, two methods were developed to discriminate shrimp and crab: a shrimp-PCR method with postamplification digestion and a crab-PCR method that specifically amplifies a fragment of the 16S rRNA gene. The sensitivity and specificity of both PCR methods were verified by experiments using DNA extracted from 15 shrimp species, 13 crab species, krill, mysid, mantis shrimp, other food samples (cephalopod, shellfish, and fish), incurred foods, and commercial food products. Both PCR methods could detect 5 pg of DNA extracted from target species and 50 ng of genomic DNA extracted from incurred foods containing 10 ppm (μg/g) total protein of shrimp or crab. The two PCR methods were considered to be specific enough to separately detect species belonging to shrimp and crab. Although false-positive and false-negative results were obtained from some nontarget crustacean species, the proposed PCR methods, when used in conjunction with ELISA tests, would be a useful tool for confirmation of the validity of food allergy labeling and management of processed food safety for allergic patients.     

Increase in soil carbon sequestration using rice husk charcoal without stimulating CH4 and N2O emissions in an Andosol paddy field in Japan

Abstract

Biochar application has been recognized as an effective option for promoting carbon (C) sequestration, but it may also affect the production and consumption of methane (CH₄) and nitrous oxide (N₂O) in soil. A 1-year field experiment was conducted to investigate the effects of rice husk charcoal application on rice (Oryza sativa L.) productivity and the balance of greenhouse gas exchanges in an Andosol paddy field. The experiment compared the treatments of rice husk charcoal applied at 10, 20 and 40 Mg ha^?1 (RC10, RC20 and RC40, respectively), rice husk applied at 20 Mg ha^?1 (RH20), and the control (CONT). Rice straw and grain yields did not significantly differ among the treatments. The seasonal cumulative CH₄ emissions were 38–47% higher from RC10, RC20 and RC40 than from the CONT. However, the increases were not in proportion to the application rates of rice husk charcoal, and their values did not significantly differ from the CONT. On the contrary, the RH20 treatment significantly increased the cumulative CH₄ emission by 227% compared to the CONT. The N₂O emissions during the measurement were not affected by the treatments. As a result, the combined global warming potential (GWP) of CH₄ and N₂O emissions was significantly higher in RH20 than in the other treatments. There was a positive linear correlation between C storage in the top 10 cm of soil and the application rate of rice husk charcoal. The increases in soil C contents compared to the CONT corresponded to 98–149% of the C amounts added as rice husk charcoal and 41% of the C added as rice husk. Carbon dioxide (CO₂) fluxes in the off season were not significantly different among RC10, RC20, RC40 and CONT, indicating that C added as rice husk charcoal remained in the soil during the fallow period. The CO₂ equivalent balance between soil C sequestration and the combined GWP indicates that the rice husk charcoal treatments stored more C in soil than the CONT, whereas the RH20 emitted more C than the CONT. These results suggest that rice husk charcoal application will contribute to mitigating global warming without sacrificing rice yields.     

Chemical Characteristics and Acid Buffering Capacity of Surface Soils in Japanese Forests

To determine the geological distribution of acid buffering capacity and exchangeable Al of forest soils in Japan, surface soils under forest vegetation were collected nationwide from a total of 1,034 sites. Generally, surface soils in Japanese forests are mostly acidic and low in both exchangeable cation content and exchangeable Al. The median of soil pH(H₂O), total exchangeable cations, and exchangeable Al are 5.1, 76 mmol₍₊₎Kg^?1, and 19.6 mmol₍₊₎kg^?1, respectively. Acid buffering capacities of selected soils were determined using a soil column, which was comparable to the capacity that resulted from cation exchanges with protons. Soils with high buffering capacity and low exchangeable Al are widely distributed in Japan, and overlap with the areas of Andisol distribution. Volcanogenic materials seem to mask soil characteristics derived from underlying geology even though they are not Andisols. However, central to western Honshu Island, Shikoku Island, and northern Kyushu Island showed weak acid buffering capacities with high exchangeable Al potential in surface soils.     

Infection and colonization of aseptically micropropagated sugarcane seedlings by nitrogen-fixing endophytic bacterium, Herbaspirillum sp. B501gfp1

In this study, we used Herbaspirillum sp. B501gfp1 (B501gfp1), an isolate from wild rice, to investigate the interaction between a non-host nitrogen-fixing endophytic bacterium and micropropagated sugarcane plants under aseptic condition. Two Japanese sugarcane plants (Saccharum sp.) cultivars (cvs) NiF8 and Ni15 were inoculated using B501gfp1 in two inoculum doses of 10⁸ and 10² bacterial-cells-per-milliliter suspension. The results showed that bacterial cells colonized both the root and stem tissues, and colonization was apparent in the intercellular spaces. Higher bacterial numbers were detected in plant tissues inoculated with the higher inoculum concentration treatment. Bacterial numbers also varied between the two cultivars, with the higher values determined in cv Ni15. This study provides evidence that Herbaspirillum sp. B501gfp1, a rice isolate, could colonize sugarcane tissues, suggesting non-specificity of host plant among endophytes.     

Variable-timing,fixed-rate application of cattle biogas effluent to rice using a leaf color chart: microcosm experiments in Vietnam

ABSTRACT

Livestock production plays a leading role in agricultural land-use change. Producing biogas from livestock waste and subsequently using the biogas effluent as fertilizer for crops is a promising option to solve environmental problems resulting from expanding livestock production. However, it is difficult to promptly and accurately measure the nitrogen (N) concentration of effluent for farmers in developing countries, making precise N management difficult. The objectives of the current study were (1) to evaluate the feasibility of variable-timing, fixed-rate application of cattle biogas effluent using a leaf color chart (LCC) for rice (Oryza sativa L.) and (2) to determine the optimum LCC threshold for grain yield. We conducted two microcosm experiments in the Mekong Delta of Vietnam in 2018 using eight treatments of N-fertilizer application. In the Zero treatment, we applied no N. In the Estd treatment, we split-applied N as effluent (E) at fixed rate and timing as the standard method. In E2.75, E3.00, E3.25, E3.50, and E3.75, we applied effluent whenever the LCC value went below 2.75, 3.00, 3.25, 3.50, and 3.75, respectively. In U3.25, we applied N as urea (U) whenever the LCC value fell below 3.25. The total effluent-N application rate ranged from 90 to 210 kg N ha^?1 season^?1. Rice growth was normal but there was a substantial yield gap between the two microcosm experiments due to the seasonal difference in solar radiation. Rice yield tended to increase with increasing LCC threshold. There was a positive linear relationship between LCC and chlorophyll content (SPAD) values (R ² = 0.73–0.79). Grain yield was well explained (R ² = 0.70–0.89) by the seasonal mean LCC or SPAD value. Plant total N uptake increased with increasing LCC threshold, but the three calculated indices of N use efficiency (NUE) – apparent N recovery, agronomic NUE, and internal NUE – were not always improved with a higher LCC threshold. Our results showed that the tested variable-timing, fixed-rate strategy for the application of cattle biogas effluent was feasible and the optimum LCC threshold for grain production was 3.75 under the current microcosm conditions.     

Promotion of the seed germination and seedling growth of cabbage by root-colonizing fungi isolated from oat and Eucalyptus roots

(Jpn. J. Soil Sci.Plant Nutr., 77, 257–263, 2006)
The symptom of blighted leaves was observed for the melon plants grown on an isolated soil bed after steam sterilization (SS) in a greenhouse in Kochi Prefecture, Japan. To clarify the causes of this symptom, soil and plant analyses were conducted in the first experiment. Manganese concentrations in the leaves with the symptom were higher than 1,000 mg kg−1and exchangeable Mn in the soil was higher than 30 mg kg−1, which indicated the observed symptom was due to Mn excess. In the second experiment, formation of Mn oxide-dissolving substances after SS was examined for several organic amendments, because large amounts of palm chips had been supplied to the soil before SS in the first experiment. As a treatment similar to SS, palm chips, bark compost, and cattle feces manure were autoclaved. The water extracts were collected before and after the autoclave treatment and the Mn oxide-dissolving capacities of the extracts were evaluated. The Mn oxide-dissolving capacity of the water extracts after SS was remarkably higher than before SS, and the value was highest in palm chips, and followed by bark compost, and cattle feces manure. Fractionation and HPLC analysis of the water extracts after SS revealed that the main substance behind Mn oxide reduction in palm tips was arabinose and that in bark manure was malic acid. Further, Mn oxide-dissolving capacities of the water extracts were almost completely explained by the amounts of arabinose or malic acid found in these materials. From these results, it was considered that reducing substances such as arabinose were released by SS from the palm chips which had been applied in large quantity before SS and these reducing substances dissolved the soil Mn oxides and increased the amounts of available Mn, which led to the occurrence of Mn toxicity to the plants.     

PCB 118 and Aryl Hydrocarbon Receptor Immunoassays for Screening Dioxins in Retail Fish

The efficacy of a combination of two enzyme-linked immunosorbent assay (ELISA) kits was examined for screening the toxic equivalent (TEQ) concentrations of dioxins in retail fish. The coplanar PCB-EIA system, which is a competitive immunoassay specific for polychlorinated biphenyl (PCB) 118, was tested as a screening method for mono- ortho PCBs. The Ah immunoassay (Ah-I), which is an ELISA-based aryl hydrocarbon receptor binding assay, was analyzed for its screening ability for non- ortho PCBs, polychlorinated dibenzo- p-dioxins (PCDDs), and dibenzofurans (PCDFs). Dilution and recovery tests using purified fish extracts revealed no major interference of the matrix in the PCB-EIA and suggested that the matrix effect was minimized in the Ah-I. Finally, the results for the fish samples ( n = 20) showed a strong correlation between this method and high-resolution gas chromatography coupled to high-resolution mass spectrometry for the determination of the TEQ concentrations of mono- ortho PCBs ( r = 0.99) and non- ortho PCBs and PCDD/Fs ( r = 0.97). These data indicate that our method is suitable for screening retail fish to determine the TEQ concentrations of dioxins.     

Epidemiological survey of Babesia gibsoni infection in dogs in eastern Japan

To determine the distribution of Babesia gibsoni infection in dogs in the eastern part of Japan, an epidemiological survey of dogs suspected of having B. gibsoni infection was attempted using the polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). Thirty-five of 115 such dogs (30.4%) were positive by PCR and/or ELISA. The 35 positive dogs consisted of 28 Tosa dogs, 4 American Pit Bull Terriers, and 3 mongrel dogs in Aomori, Fukushima, Ibaraki, Gunma, Chiba, Tokyo, Kanagawa, and Nagano Prefectures. The positive dogs had a significantly lower rate of tick exposure and a higher rate of bites by other dogs. Twenty-two of 35 B. gibsoni-positive dogs were infected with hemoplasma, and the rate of infection was significantly higher than that of B. gibsoni-negative dogs.