The inter-laboratory validation study of EpiSensA for predicting skin sensitization potential.

The Epidermal Sensitization Assay (EpiSensA) is a reconstructed human epidermis (RhE)-based gene expression assay for predicting the skin sensitization potential of chemicals. Since the RhE model is covered by a stratified stratum corneum, various kinds of test chemicals, including lipophilic ones and pre-/pro-haptens, can be tested with a route of exposure akin to an in vivo assay and human exposure. This article presents the results of a formally managed validation study of the EpiSensA that was carried out by three participating laboratories. The purpose of this validation study was to assess transferability of the EpiSensA to new laboratories along with its within- (WLR) and between-laboratory reproducibility (BLR). The validation study was organized into two independent stages. As demonstrated during the first stage, where three sensitizers and one non-sensitizer were correctly predicted by all participating laboratories, the EpiSensA was successfully transferred to all three participating laboratories. For Phase I of the second stage, each participating laboratory performed three experiments with an identical set of 15 coded test chemicals resulting in WLR of 93.3%, 93.3%, and 86.7%, respectively. Furthermore, when the results from the 15 test chemicals were combined with those of the additional 12 chemicals tested in Phase II of the second stage, the BLR for 27 test chemicals was 88.9%. Moreover, the predictive capacity among the three laboratories showed 92.6% sensitivity, 63.0% specificity, 82.7% accuracy, and 77.8% balanced accuracy based on murine local lymph node assay (LLNA) results. Overall, this validation study concluded that EpiSensA is easily transferable and sufficiently robust for assessing the skin sensitization potential of chemicals.

[Laboratory Performance Study of the Japanese Official Method to Detect Genetically Modified Papaya Line PRSV-YK].

Since the establishment of procedures for the safety assessment of food products that use recombinant DNA technology, the manufacture, import, and sale of genetically modified (GM) foods that have not undergone safety assessment are prohibited under the Food Sanitation Act. Therefore, a performance study to confirm the GM food testing operations of each laboratory is very important to ensure the reliability of the GM food monitoring system. In 2022, GM papaya line PRSV-YK-which has not yet been authorized in Japan-was selected for testing, and a papaya paste and a DNA solution were used as the test samples. With these samples, a laboratory performance study of the DNA extraction and real-time PCR operations was conducted. This confirmed that the 18 participating laboratories were generally performing the DNA extraction and real-time PCR operations correctly. However, some laboratories using certain DNA amplification reagent with some real-time PCR instruments were not able to determine the PRSV-YK detection test. This suggests that the PRSV-YK detection test may not be able to correctly detect samples containing GM papaya when performed with these combinations of instruments and reagent. In order to ensure the reliability of the PRSV-YK detection test, it is necessary to examine in detail how the combination of DNA polymerase reagents and real-time PCR instruments affects the detection limit, and to implement an appropriate solution.

Production of bacteriocin by Virgibacillus salexigens isolated from "terasi": a traditionally fermented shrimp paste in Indonesia.

A natural antibacterial-substance-producing gram-positive bacterium was isolated from terasi shrimp paste, a popular fermented product in Indonesia. This strain, a spore-forming and strictly aerobic bacterium, was identified as Virgibacillus salexigens by 16S rRNA gene sequence analysis. The antibacterial substance purified from the precipitated product in the culture supernatant of the strain using ammonium sulfate showed a broad inhibition spectrum against gram-positive bacteria, including a typical foodborne bacterium, namely, Listeria monocytogenes. The antibacterial activity of the substance was inactivated by treatments with various proteolytic enzymes. It was stable after heating or pH treatment, and approximately 60% of the initial activity remained even after heating at 121 °C for 15 min. In addition, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis indicated that its monoisotopic mass weight was 5318.4 Da (M+H)(+). On the basis of the results obtained by the automated Edman degradation technique and MALDI-TOF MS analysis, the substance can be classified as a member of Class IId bacteriocins, but it could not be identified as any of the previously purified substances except for the putative bacteriocin predicted from the draft genome sequence data of gram-positive bacteria such as Virgibacillus and Bacillus strains.

Isolation and characterization of halophilic lactic acid bacteria isolated from "terasi" shrimp paste: a traditional fermented seafood product in Indonesia.

Lactic acid bacteria from "terasi" shrimp paste, a highly popular fermented seafood in Indonesia were isolated and characterized. Viable cell counts were 10(4) to 10(6) cfu/g on MRS medium. All the isolates were catalase-negative, gram-positive cocci and were able to grow at 15% NaCl. Numerical phenotypic analysis showed that the isolates clustered into one group. However, they could be classified into two types: the Tetragenococcus halophilus group and the T. muriaticus group as revealed by a restriction fragment length polymorphism (RFLP) analysis and sequencing of the 16S rRNA gene. This study is the first to show that both species of Tetragenococcus are distributed in Indonesian fermented foods.