Biodegradation of γ-hexachlorocyclohexane by transgenic hairy root cultures of <Emphasis Type="Italic">Cucurbita moschata</Emphasis> that accumulate recombinant bacterial LinA

Key message

γ-HCH was successfully degraded using LinA-expressed transgenic hairy root cultures of Cucurbita moschata . Fusing an endoplasmic reticulum-targeting signal peptide to LinA was essential for stable accumulation in the hairy roots.

Abstract

The pesticide γ-hexachlorocyclohexane (γ-HCH) is a persistent organic pollutant (POP) that raises public health and environmental pollution concerns worldwide. Although several isolates of γ-HCH-degrading bacteria are available, inoculating them directly into γ-HCH-contaminated soil is ineffective because of the bacterial survival rate. Cucurbita species incorporate significant amounts of POPs from soils compared with other plant species. Here, we describe a novel bioremediation strategy that combines the bacterial degradation of γ-HCH and the efficient uptake of γ-HCH by Cucurbita species. We produced transgenic hairy root cultures of Cucurbita moschata that expressed recombinant bacterial linA, isolated from the bacterium Sphingobium japonicum UT26. The LinA protein was accumulated stably in the hairy root cultures by fusing an endoplasmic reticulum (ER)-targeting signal peptide to LinA. Then, we demonstrated that the cultures degraded more than 90 % of γ-HCH (1 ppm) overnight and produced the γ-HCH metabolite 1,2,4-trichlorobenzene, indicating that LinA degraded γ-HCH. These results indicate that the gene linA has high potential for phytoremediation of environmental γ-HCH.

     

Multi‐methods approach to characterizing the magnitude,impact, and spatial risk of Irrawaddy dolphin (Orcaella brevirostris) bycatch in small‐scale fisheries in Malampaya Sound,Philippines

Addressing the urgent conservation threat of marine mammal bycatch in small‐scale fisheries requires information on bycatch magnitude, gear types, population impacts, and risk. However, data on these proximate attributes are widely lacking in developing countries. This study used a multi‐methods approach integrating boat surveys and interviews with fishermen (n = 526) to assess proximate attributes of bycatch for the Critically Endangered subpopulation of Irrawaddy dolphins (Orcaella brevirostris) in Malampaya Sound, Philippines. Given an updated population estimate of 35 individuals (CV = 22.9%), the estimated rate of bycatch fatalities exceeds the potential biological removal rate. Spatial overlap scores were calculated to characterize overlap between dolphins and fisheries as a proxy of bycatch risk. These scores identified particularly high risk areas that could be prioritized for gear bans. However, completely reducing bycatch risk would require more widespread bans beyond these high risk areas. Implementing gear restrictions will be immensely challenging, given serious obstacles to fisheries management at this site. The multi‐methods approach describes the urgency of the bycatch problem and the changes required for mitigation, setting the stage for identifying and evaluating potential solutions. It can be readily applied to developing country sites to guide more efficient and complete data collection and conservation efforts.     

Oxidative stress in Perna perna and other bivalves as indicators of environmental stress in the Brazilian marine environment: antioxidants, lipid peroxidation and DNA damage

Oxidative stress can take place in marine bivalves under a series of environmental adverse conditions. The study of different systems related to oxidative stress in these organisms can give important information about their physiological status and also about environmental health. Bivalves have been proposed as good sentinel organisms in pollution monitoring studies through the analysis of biochemical biomarkers, and most of the biomarkers analyzed are those related to oxidative stress. However, it is very important to know how other environmental factors not associated to the presence of pollutants might affect these parameters. We have studied a series of mechanisms related to oxidative stress in mussels which inhabit the Brazilian coast, especially in Perna perna species, subjected to different stress conditions, such as the exposure to different contaminants in the laboratory and in the field, the exposure of mussels to air and re-submersion, simulating the tidal oscillations, and in mussels collected at different seasons. Both oxidative damage levels and antioxidant defense systems were strongly affected by the different environmental stress. This review summarizes the data obtained in some studies carried out in bivalves from the Brazilian coast.     

Two critical genes (HLA-DRB1 and ABCF1)in the HLA region are associated with the susceptibility to autoimmune pancreatitis

We have previously reported that autoimmune pancreatitis (AIP) is a bioclinical entity characterized by high serum immunoglobulin G4 concentrations and association with the HLA-DRB1*0405-DQB1*0401 haplotype. However, the precise identity of gene(s) within this haplotype directly responsible for AIP pathogenesis is yet to be established. To dissect the genetic contribution of the incriminated haplotype, we have now performed an association analysis within the human leukocyte antigen (HLA) region using various types of polymorphic markers. Genomic DNAs from 43 AIP patients and 213 unrelated Japanese controls were used in this analysis. In each DNA sample, we established the genotype of 25 microsatellite markers distributed throughout the HLA region, that of single nucleotide polymorphism within the 5'-flanking regions of the TNFA and IkBLI (also known as NFKBIL1) as well as HLA class I and II genes. The HLA-linked susceptibility regions for AIP were localized to two segments: HLA-DRB1 (*0405; OR = 3.20, P = 0.00063, Pc = 0.0016) -DQB1 (*0401; OR = 3.29, P = 0.00046, Pc = 0.0069) in the HLA class II and C3-2-11 microsatellite (allele 219; OR = 2.96, P = 0.0076, Pc = 0.099) in the HLA class I regions. Upon stratification analysis in search for a synergistic effect given the extensive linkage disequilibrium within the major histocompatibility complex, it was established that each segment contributed to disease pathogenesis. The two critical HLA regions for susceptibility to AIP are limited to the HLA-DRB1*0405-DQB1*0401 in the class II and the ABCF1 proximal to C3-2-11, telomeric of HLA-E, in the class I regions.     

The c‐MYC‐ABCB5 axis plays a pivotal role in 5‐fluorouracil resistance in human colon cancer cells

c‐MYC overexpression is frequently observed in various cancers including colon cancer and regulates many biological activities such as aberrant cell proliferation, apoptosis, genomic instability, immortalization and drug resistance. However, the mechanism by which c‐MYC confers drug resistance remains to be fully elucidated. In this study, we found that the c‐MYC expression level in primary colorectal cancer tissues correlated with the recurrence rate following 5‐fluorouracil (5‐FU)‐based adjuvant chemotherapy. Supporting this finding, overexpression of exogenous c‐MYC increased the survival rate following 5‐FU treatment in human colon cancer cells, and knockdown of endogenous c‐MYC decreased it. Furthermore, c‐MYC knockdown decreased the expression level of ABCB5, which is involved in 5‐FU resistance. Using a chromatin immunoprecipitation assay, we found that c‐MYC bound to the ABCB5 promoter region. c‐MYC inhibitor (10058‐F4) treatment inhibited c‐MYC binding to the ABCB5 promoter, leading to a decrease in ABCB5 expression level. ABCB5 knockdown decreased the survival rate following 5‐FU treatment as expected, and the ABCB5 expression level was increased in 5‐FU‐resistant human colon cancer cells. Finally, using a human colon cancer xenograft murine model, we found that the combined 5‐FU and 10058‐F4 treatment significantly decreased tumorigenicity in nude mice compared with 5‐FU or 10058‐F4 treatment alone. 10058‐F4 treatment decreased the ABCB5 expression level in the presence or absence of 5‐FU. In contrast, 5‐FU treatment alone increased the ABCB5 expression level. Taken together, these results suggest that c‐MYC confers resistance to 5‐FU through regulating ABCB5 expression in human colon cancer cells.     

The Neural Correlates of Shoulder Apprehension: A Functional MRI Study

Although shoulder apprehension is an established clinical finding and is important for the prevention of shoulder dislocation, how this subjective perception is evoked remains unclear. We elucidated the functional neuroplasticity associated with apprehension in patients with recurrent anterior shoulder instability (RSI) using functional magnetic resonance imaging (fMRI). Twelve healthy volunteers and 14 patients with right-sided RSI performed a motor imagery task and a passive shoulder motion task. Brain activity was compared between healthy participants and those with RSI and was correlated with the apprehension intensity reported by participants after each task. Compared to healthy volunteers, participants with RSI exhibited decreased brain activity in the motor network, but increased activity in the hippocampus and amygdala. During the passive motion task, participants with RSI exhibited decreased activity in the left premotor and primary motor/somatosensory areas. Furthermore, brain activity was correlated with apprehension intensity in the left amygdala and left thalamus during the motor imagery task (memory-induced), while a correlation between apprehension intensity and brain activity was found in the left prefrontal cortex during the passive motion task (instability-induced). Our findings provide insight into the pathophysiology of RSI by identifying its associated neural alterations. We elucidated that shoulder apprehension was induced by two different factors, namely instability and memory.     

Simple Resolution of Enantiomeric NMR Signals of α‐Amino Acids by Using Samarium(III) Nitrate With L‐Tartarate

Readily available L‐tartaric acid, which is a bidentate ligand with two chiral centers forming a seven‐membered chelate ring, was applied to the chiral ligand for the chiral nuclear magnetic resonance (NMR) shift reagent of samarium(III) formed in situ. This simple method does not cause serious signal broadening in the high magnetic field. Enantiomeric ¹³C and ¹H NMR signals and enantiotopic ¹H NMR signals of α‐amino acids were successfully resolved at pH 8.0 and the 1:3 molar ratio of Sm(NO₃)₃:L‐tartaric acid. It is elucidated that the enantiomeric signal resolution is attributed to the anisotropic magnetic environment for the enantiomers induced by the chiral L‐tartarato samarium(III) complex rather than differences in stability of the diastereomeric substrate adducts. The present ¹³C NMR signal resolution was also effective for the practical simultaneous analysis of plural kinds of DL‐amino acids. Chirality 27:353–357, 2015.© 2015 Wiley Periodicals, Inc.     

Human-type ABO blood groups as genetic markers for the management of a squirrel monkey (Saimiri sciureus) breeding colony

The human-type ABO blood groups were determined for 94 families of the squirrel monkey which included 151 animals. Four phenotypes of ABO blood groups (A, B, AB, and O) were detected. Family analysis revealed that the human-type ABO blood groups in this species were governed by three alleles, codominantA andB and silentO. There were intraspecific differences in the distribution of phenotypes and gene frequency among three populations imported by different routes at different times. The usefulness of ABO blood groups for defining the genetic variability of a squirrel monkey breeding colony through successive generations is discussed on the basis of the difference in distribution of ABO blood groups between wild-originated parental and its first colony-born populations.     

Full-term development of in vitro-matured, vitrified and fertilized bovine oocytes

In vitro-matured bovine oocytes were vitrified in a mixture of 2 M-dimethyl sulphoxide (DMSO), 1 M-acetamide and 3 M-propylene glycol dissolved in mTCM199. After vitrification and thawing, the oocytes were exposed to 2-0.1M-sucrose solution in 1 or 12 steps to remove the cryoprotectants. Then the oocytes were fertilized in vitro and co-cultured with a monolayer of cumulus cells for 7 days. Nine of 88 inseminated oocytes developed to the blastocyst stage. Three blastocysts were transferred to 3 recipients, resulting in 2 pregnancies.