Current situation of lead (Pb) exposure in raptors and waterfowl in Japan and difference in sensitivity to in vitro lead exposure among avian species.

Although lead (Pb) poisoning in wild birds has been considered a serious problem in Japan for over 30 years, there is little information about Pb exposure and its sources throughout Japan except for Hokkaido. Furthermore, to identify and effectively prioritize the conservation needs of highly vulnerable species, differences in sensitivity to Pb exposure among avian species need to be determined. Therefore, we investigated the current situation of Pb exposure in raptors (13 species, N = 82), waterfowl (eight species, N = 44) and crows (one species, N = 6) using concentration and isotope analysis. We employed blood or tissue samples collected in various Japanese facilities mainly in 2022 or 2023. We also carried out a comparative study of blood δ-ALAD sensitivity to in vitro Pb exposure using blood of nine avian species. Pb concentrations in the blood or tissues displayed increased levels (>0.1 µg/g blood) in two raptors (2.4%), ten waterfowl (23%) and one crow (17%). Among them, poisoning levels (>0.6 µg/g blood) were found in one black kite and one common teal. The sources of Pb isotope ratios in ten blood samples with high Pb levels were determined as deriving from shot pellets (N = 9) or rifle bullets (N = 1). In the δ-ALAD study, red-crowned crane showed the highest sensitivity among the nine tested avian species and was followed in order by five Accipitriformes species (including white-tailed and Steller's sea eagle), Blakiston's fish owl, Muscovy duck and chicken, suggesting a genetically driven variance in susceptibility. Further studies on contamination conditions and exposure sources are urgently needed to inform strict regulations on the usage of Pb ammunition. Furthermore, detailed examinations of δ-ALAD sensitivity, interspecific differences, and other factors involved in the variability in sensitivity to Pb are required to identify and prioritize highly sensitive species.

Evaluation of anticoagulant rodenticide sensitivity by examining in vivo and in vitro responses in avian species, focusing on raptors.

Anticoagulant rodenticides (ARs) are used to control pest rodent species but can result in secondary poisoning of non-target animals, especially raptors. In the present study, differences in AR sensitivity among avian species were evaluated by comparing in vivo warfarin pharmacokinetics and effects, measuring cytochrome P450s (CYPs) expression involved in AR metabolism, and conducting in vitro inhibition assays of the AR target enzyme Vitamin K 2,3-epoxide reductase (VKOR). Oral administration of warfarin at 4 mg/kg body weight did not prolong prothrombin time in chickens (Gallus gallus), rock pigeons (Columba livia), or Eastern buzzards (Buteo japonicus). Rock pigeons and buzzards exhibited shorter plasma half-life of warfarin compared to chickens. For the metabolite analysis, 4'-hydroxywarfarin was predominantly detected in all birds, while 10-hydroxywarfarin was only found in pigeons and raptors, indicating interspecific differences in AR metabolism among birds likely due to differential expression of CYP enzymes involved in the metabolism of ARs and variation of VKOR activities among these avian species. The present findings, and results of our earlier investigations, demonstrate pronounced differences in AR sensitivity and pharmacokinetics among bird species, and in particular raptors. While ecological risk assessment and mitigation efforts for ARs have been extensive, AR exposure and adverse effects in predatory and scavenging wildlife continues. Toxicokinetic and toxicodynamic data will assist in such risk assessments and mitigation efforts.

A Logistic Regression Model for Predicting the Risk of Subsequent Surgery among Patients with Newly Diagnosed Crohn's Disease Using a Brute Force Method.

Surgery avoidance is an important goal in Crohn's disease (CD) treatment and predicting the risk of subsequent surgery is important to determine adequate therapeutic strength for patients with newly diagnosed CD. Herein, we aimed to construct a prediction model for the risk of subsequent surgery based on disease characteristics at the patients' initial visit. We retrospectively collected disease characteristic data from 93 patients with newly diagnosed CD. A logistic regression model with a brute force method was used to maximize the area under the receiver operating characteristic curve (auROC) by employing a combination of potential predictors from 14 covariates (16,383). The auROC remained almost constant when one to 12 covariates were considered, reaching a peak of 0.89 at four covariates (small-bowel patency, extensive small-bowel lesions, main lesions, and the number of poor prognostic factors), and it decreased with increasing covariate size. The most significant predictors were small-bowel patency, extensive small-bowel lesions, and age or major lesions. Therefore, this prediction model using covariates may be helpful in determining the likelihood that a patient with newly diagnosed CD will require surgery, which can aid in appropriate treatment selection for high-risk patients.

Surveys of eleven species of wild and zoo birds and feeding experiments in white-tailed eagles reveal differences in the composition of the avian gut microbiome based on dietary habits between and within species.

The composition of the gut microbiome varies due to dietary habits. We investigated influences of diet on the composition of the gut microbiome using the feces of 11 avian species, which consumed grain-, fish- and meat-based diets. We analyzed gut microbiome diversity and composition by next-generation sequencing (NGS) of 16S ribosomal RNA. The grain-diet group had higher gut microbiome diversity than the meat- and fish-diet group. The ratio of Bacteroidetes and Firmicutes phyla was higher in the grain-diet group than in the meat- and fish-diet groups. The grain-diet group had a higher ratio of Veillonellaceae than the meat-diet group and a higher ratio of Eubacteriaceae than the fish-diet habit group. To clarify the influence of diet within the same species, white-tailed eagles (Haliaeetus albicilla, n=6) were divided into two groups, and given only deer meat or fish for approximately one month. The composition of the gut microbiome of individuals in both groups were analyzed by NGS. There were indications of fluctuation in the levels of some bacteria (Lactobacillus, Coriobacteriales, etc.) in each diet group. Moreover, one individual for each group which switched each diet in last week changed to each feature of composition of bacterial flora. The above results show that the composition of the gut microbiome differ depending on diet, even within the same species.

Vitamin A Promotes the Fusion of Autophagolysosomes and Prevents Excessive Inflammasome Activation in Dextran Sulfate Sodium-Induced Colitis.

Vitamin A ensures intestinal homeostasis, impacting acquired immunity and epithelial barrier function; however, its role in innate immunity is mostly unknown. Here, we studied the impact of vitamin A in different dextran sulfate sodium (DSS)-induced colitis animal models. Interestingly, more severe DSS-induced colitis was observed in vitamin A-deficient (VAD) mice than in vitamin A-sufficient (VAS) mice; the same was observed in VAD severe combined immunodeficient mice lacking T/B cells. Remarkably, IL-1ß production, LC3B-II expression, and inflammasome activity in the lamina propria were significantly elevated in VAD mice. Electron microscopy revealed numerous swollen mitochondria with severely disrupted cristae. In vitro, non-canonical inflammasome signaling-induced pyroptosis, LC3B-II and p62 expression, and mitochondrial superoxide levels were increased in murine macrophages (RAW 264.7) pretreated with retinoic acid receptor antagonist (Ro41-5253). These findings suggest that vitamin A plays a crucial role in the efficient fusion of autophagosomes with lysosomes in colitis.

Evaluation of Crohn's Disease Small-Bowel Mucosal Healing Using Capsule Endoscopy and Usefulness of Leucine-Rich α2-Glycoprotein.

Recently, the importance of achieving clinical and deep remissions with mucosal healing (MH) has been demonstrated as a therapeutic goal to avoid Crohn's disease (CD) surgical operations. Although ileocolonoscopy (CS) is considered the gold standard, there are increasing reports on the benefits of capsule endoscopy (CE) and serum leucine-rich α2-glycoprotein (LRG) for evaluating small-bowel lesions in CD. We evaluated the data of 20 patients with CD who underwent CE in our department between July 2020 and June 2021 and whose serum LRG level was measured within 2 months. Concerning the mean LRG value, there was no significant difference between the CS-MH and CS-non-MH groups. Conversely, the mean LRG level was 10.0 µg/mL in seven patients in the CE-MH group and 15.2 µg/mL in 11 patients in the CE-non-MH group with a significant difference between the two groups (p = 0.0025). This study's findings show that CE can sufficiently determine total MH in most cases, and LRG is useful for evaluating CD small-bowel MH because of its correlation with CE-MH. Furthermore, satisfying CS-MH criteria and a cut-off value of 13.4 µg/mL for LRG suggests its usefulness as a CD small-bowel MH marker, which could be incorporated into the treat-to-target strategy.

Experimental and natural infections of white-tailed sea eagles (Haliaeetus albicilla) with high pathogenicity avian influenza virus of H5 subtype.

White-tailed sea eagle (Haliaeetus albicilla), a regionally rare species of raptor, is threatened in several countries. To assess the risk of H5 high pathogenicity avian influenza (HPAI) viral infection in rare bird species, we performed experimental infections with a GS/GD96-lineage H5N6 HPAI virus of clade 2.3.4.4e in white-tailed sea eagles. Additionally, during the winter of 2020-2021 in Japan, we accidentally encountered a white-tailed sea eagle that had a fatal outcome due to natural infection with a GS/GD96-lineage H5N8 HPAI virus of clade 2.3.4.4b, allowing us to compare experimental and natural infections in the same rare raptor species. Our experiments demonstrated the susceptibility of white-tailed sea eagles to the GS/GD96-lineage H5 HPAI virus with efficient replication in systemic organs. The potential for the viruses to spread within the white-tailed sea eagle population through indirect transmission was also confirmed. Comprehensive comparisons of both viral distribution and histopathological observations between experimentally and naturally infected white-tailed sea eagles imply that viral replication in the brain is responsible for the disease severity and mortality in this species. These findings provide novel insights into the risk assessment of H5 HPAI viral infection in white-tailed sea eagles, proper diagnostic procedures, potential risks to artificially fed eagle populations and persons handling superficially healthy eagles, potential impact of intragastric infection on eagle outcomes, and possibility of severity of the disease being attributed to viral replication in the brain.

Behavior and toxic effects of Pb in a waterfowl model with oral exposure to Pb shots: Investigating Pb exposure in wild birds.

Among wild birds, lead (Pb) exposure caused by ingestion of ammunition is a worldwide problem. We aimed to reveal the behavior and toxic effect of Pb caused by ingesting Pb shots in waterfowl. Four male, eight-week old Muscovy ducks (Cairina moschata) were given three Pb shots (approximately 240 mg in total) orally and then fed for 29 days after exposure, simulating a low-dose Pb exposure in wild waterfowl. During the breeding period, blood samples were collected 10 times, and fecal samples every day. Additionally, 22 fresh tissue and 6 bone samples were obtained from each duck through the dissection. Although there were no gross abnormalities, the maximum blood Pb concentration of each duck ranged from 0.6 to 3.7 mg/L, reaching a threshold concentration indicative of clinical symptoms (>0.5 mg/L). δ-aminolevulinic acid dehydratase declined one day after exposure and remained low throughout the feeding period. Hematocrit also tended to decrease, indicating signs of anemia. The highest Pb accumulation was observed in the bones, followed by the kidneys, intestinal tracts, and liver. High Pb accumulation in the bones, which are known to have a long Pb half-life, suggested that Pb would remain in the body and possibly affect bird health beyond 28 days after exposure. Gene expression analysis showed a significant increase in the expression of the toll-like receptor-3 gene, which is involved in virus discrimination in the liver, suggesting a disruption of the immune system. Microbiota analyses showed a correlation between the blood Pb concentration and the abundances of Lachnospiraceae and Ruminococcaceae, suggesting that Pb affects lipid metabolism. These results provide fundamental data on Pb exposure in wild birds and a new perspective on the damage such exposure causes.

Automated evaluation of colon capsule endoscopic severity of ulcerative colitis using ResNet50.

Capsule endoscopy has been widely used as a non-invasive diagnostic tool for small or large intestinal lesions. In recent years, automated lesion detection systems using machine learning have been devised. This study aimed to develop an automated system for capsule endoscopic severity in patients with ulcerative colitis along the entire length of the colon using ResNet50. Capsule endoscopy videos from patients with ulcerative colitis were collected prospectively. Each single examination video file was partitioned into four segments: the cecum and ascending colon, transverse colon, descending and sigmoid colon, and rectum. Fifty still pictures (576 × 576 pixels) were extracted from each partitioned video. A patch (128 × 128 pixels) was trimmed from the still picture at every 32-pixel-strides. A total of 739,021 patch images were manually classified into six categories: 0) Mayo endoscopic subscore (MES) 0, 1) MES1, 2) MES2, 3) MES3, 4) inadequate quality for evaluation, and 5) ileal mucosa. ResNet50, a deep learning framework, was trained using 483,644 datasets and validated using 255,377 independent datasets. In total, 31 capsule endoscopy videos from 22 patients were collected. The accuracy rates of the training and validation datasets were 0.992 and 0.973, respectively. An automated evaluation system for the capsule endoscopic severity of ulcerative colitis was developed. This could be a useful tool for assessing topographic disease activity, thus decreasing the burden of image interpretation on endoscopists.

Analysis of lead distribution in avian organs by LA-ICP-MS: Study of experimentally lead-exposed ducks and kites.

Lead poisoning of wild birds by ingestion of lead ammunition occurs worldwide. Histopathological changes in organs of lead-intoxicated birds are widely known, and lead concentration of each organ is measurable using mass spectrometry. However, detailed lead localization at the suborgan level has remained elusive in lead-exposed birds. Here we investigated the detailed lead localization in organs of experimentally lead-exposed ducks and kites by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). In both the ducks and kites, lead accumulated diffusely in the liver, renal cortex, and brain. Lead accumulation was restricted to the red pulp in the spleen. With regard to species differences in lead distribution patterns, it is noteworthy that intensive lead accumulation was observed in the arterial walls only in the kites. In addition, the distribution of copper in the brain was altered in the lead-exposed ducks. Thus, the present study shows suborgan lead distribution in lead-exposed birds and its differences between avian species for the first time. These findings will provide fundamental information to understand the cellular processes of lead poisoning and the mechanisms of species differences in susceptibility to lead exposure.

Re-Invasion of H5N8 High Pathogenicity Avian Influenza Virus Clade 2.3.4.4b in Hokkaido, Japan, 2020.

Global dispersion of high pathogenicity avian influenza (HPAI), especially that caused by H5 clade 2.3.4.4, has threatened poultry industries and, potentially, human health. An HPAI virus, A/northern pintail/Hokkaido/M13/2020 (H5N8) (NP/Hok/20) belonging to clade 2.3.4.4b, was isolated from a fecal sample collected at a lake in Hokkaido, Japan where migratory birds rested, October 2020. In the phylogenetic trees of all eight gene segments, NP/Hok/20 fell into in the cluster of European isolates in 2020, but was distinct from the isolates in eastern Asia and Europe during the winter season of 2017-2018. The antigenic cartography indicates that the antigenicity of NP/Hok/20 was almost the same as that of previous isolates of H5 clade 2.3.4.4b, whereas the antigenic distances from NP/Hok/20 to the representative strains in clade 2.3.4.4e and to a strain in 2.3.4 were apparently distant. These data imply that HPAI virus clade 2.3.4.4b should have been delivered by bird migration despite the intercontinental distance, although it was not defined whether NP/Hok/20 was transported from Europe via Siberia where migratory birds nest in the summer season. Given the probability of perpetuation of transmission in the northern territory, periodic updates of intensive surveys on avian influenza at the global level are essential to prepare for future outbreaks of the HPAI virus.

Current situation regarding lead exposure in birds in Japan (2015-2018); lead exposure is still occurring.

Birds of a number of species have died as a result of lead (Pb) poisoning, including many Steller's sea eagles (Haliaeetus pelagicus) and white-tailed sea eagles (Haliaeetus albicilla) in Hokkaido, the northernmost island of Japan. To address this issue, the use of any type of Pb ammunition for hunting of large animals was prohibited in Hokkaido in 2004. However, Pb poisoning is still being reported in this area, and there are few regulations regarding the use of Pb ammunition in other parts of Japan, where it has been reported that eagles and water birds have been exposed to Pb. This study was performed to accurately determine the current level of Pb exposure of birds found dead in the field or dead in the wild bird centers in Japan (June 2015-May 2018) and to identify the sources of Pb. Pb exposure was found to still be occurring in raptors and water birds in various parts of Japan. Twenty-six point five % and 5.9% of the recorded deaths of Steller's sea eagles and white-tailed sea eagles, respectively, were found to have been poisoned by Pb. In addition, Pb isotope ratio analysis showed that both Pb rifle bullets and Pb shot pellets cause Pb exposure in birds, and these endangered eagles are also exposed to Pb in Hokkaido due to the illegal use of Pb ammunition. Changing to Pb-free ammunition, such as copper (Cu) rifle bullets, steel shot pellets, or bismuth shot pellets, will be essential for the conservation of avian species in Japan.

Prevalence of major depressive disorder among hemodialysis patients compared with healthy people in Japan using the Structured Clinical Interview for DSM-IV.

BACKGROUND: We investigated the prevalence of depression in hemodialysis (HD) patients using the Center for Epidemiologic Studies for Depression (CES-D) scale and the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (SCID) and compared the rates with those of community dwelling people in Japan. PATIENTS AND METHODS: A total of 99 patients undergoing HD were recruited. Blood sampling was performed no later than 2 weeks prior to assessment. As a reference group for SCID and CES-D evaluation, 404 age- and sex-matched healthy controls who had participated in the Iwaki Health Promotion Project were included in this study. The SCID and the CES-D scale were administered to all participants to diagnose their depression. Participants who met the criteria of a major depressive episode according to the SCID were classified as SCID depression and the participants whose CES-D score was 16 or higher were classified as CES-D depression. RESULTS: Ninety-nine HD patients completed the evaluation and data collection. There were no significant differences in age, sex, or CES-D scores between HD patients and controls. There were 12 cases of SCID depression in HD patients and four cases in controls. There was a significant difference between HD patients and controls in the prevalence of SCID depression. There were no significant differences between the two groups with regard to demographic or clinical data. There were 19 HD patients and 24 controls who showed CES-D depression. There was no significant difference between HD patients and controls in the prevalence of CES-D depression. There was a significant difference in potassium level between the two groups, but there were no significant differences in any of the other items. CONCLUSION: There were significantly more HD patients showing SCID depression than controls in the present study. In clinical settings, the SCID might be useful in surveying cases of depression detected by screening tools among HD patients.

Genetic and antigenic characterization of H5, H6 and H9 avian influenza viruses circulating in live bird markets with intervention in the center part of Vietnam.

A total of 3,045 environmental samples and oropharyngeal and cloacal swabs from apparently healthy poultry have been collected at three live bird markets (LBMs) at which practices were applied to reduce avian influenza (AI) virus transmission (intervention LBMs) and six conventional LBMs (non-intervention LBMs) in Thua Thien Hue province in 2014 to evaluate the efficacy of the intervention LBMs. The 178 AI viruses, including H3 (19 viruses), H4 (2), H5 (8), H6 (30), H9 (114), and H11 (5), were isolated from domestic ducks, muscovy ducks, chickens, and the environment. The prevalence of AI viruses in intervention LBMs (6.1%; 95% CI: 5.0-7.5) was similar to that in non-intervention LBMs (5.6%; 95% CI: 4.5-6.8; χ(2)=0.532; df=1; P=0.53) in the study area. Eight H5N6 highly pathogenic avian influenza (HPAI) viruses were isolated from apparently healthy ducks, muscovy ducks, and an environmental sample in an intervention LBM. The hemagglutinin genes of the H5N6 HPAI viruses belonged to the genetic clade 2.3.4.4, and the antigenicity of the H5N6 HPAI viruses differed from the H5N1 HPAI viruses previously circulating in Vietnam. Phylogenetic and antigenic analyses of the H6 and H9 viruses isolated in both types of LBMs revealed that they were closely related to the viruses isolated from domestic birds in China, Group II of H6 viruses and Y280 lineage of H9 viruses. These results indicate that the interventions currently applied in LBMs are insufficient to control AI. A risk analysis should be conducted to identify the key factors contributing to AI virus prevalence in intervention LBMs.

Experimental infection of highly and low pathogenic avian influenza viruses to chickens, ducks, tree sparrows, jungle crows, and black rats for the evaluation of their roles in virus transmission.

Highly pathogenic avian influenza viruses (HPAIVs) have spread in both poultry and wild birds. Determining transmission routes of these viruses during an outbreak is essential for the control of avian influenza. It has been widely postulated that migratory ducks play crucial roles in the widespread dissemination of HPAIVs in poultry by carrying viruses along with their migrations; however close contacts between wild migratory ducks and poultry are less likely in modern industrial poultry farming settings. Therefore, we conducted experimental infections of HPAIVs and low pathogenic avian influenza viruses (LPAIVs) to chickens, domestic ducks, tree sparrows, jungle crows, and black rats to evaluate their roles in virus transmission. The results showed that chickens, ducks, sparrows, and crows were highly susceptible to HPAIV infection. Significant titers of virus were recovered from the sparrows and crows infected with HPAIVs, which suggests that they potentially play roles of transmission of HPAIVs to poultry. In contrast, the growth of LPAIVs was limited in each of the animals tested compared with that of HPAIVs. The present results indicate that these common synanthropes play some roles in influenza virus transmission from wild birds to poultry.

Genetic and antigenic characterization of H5 and H7 influenza viruses isolated from migratory water birds in Hokkaido, Japan and Mongolia from 2010 to 2014.

Migratory water birds are the natural reservoir of influenza A viruses. H5 and H7 influenza viruses are isolated over the world and also circulate among poultry in Asia. In 2010, two H5N1 highly pathogenic avian influenza viruses (HPAIVs) were isolated from fecal samples of water birds on the flyway of migration from Siberia, Russia to the south in Hokkaido, Japan. H7N9 viruses are sporadically isolated from humans and circulate in poultry in China. To monitor whether these viruses have spread in the wild bird population, we conducted virological surveillance of avian influenza in migratory water birds in autumn from 2010 to 2014. A total of 8103 fecal samples from migratory water birds were collected in Japan and Mongolia, and 350 influenza viruses including 13 H5 and 19 H7 influenza viruses were isolated. A phylogenetic analysis revealed that all isolates are genetically closely related to viruses circulating among wild water birds. The results of the antigenic analysis indicated that the antigenicity of viruses in wild water birds is highly stable despite their nucleotide sequence diversity but is distinct from that of HPAIVs recently isolated in Asia. The present results suggest that HPAIVs and Chinese H7N9 viruses were not predominantly circulating in migratory water birds; however, continued monitoring of H5 and H7 influenza viruses both in domestic and wild birds is recommended for the control of avian influenza.