Journal of fish diseases最新文献

{"title":"First Report of Dual Infection of Hepatopancreatic Parvovirus and Enterocytozoon hepatopenaei in Wild and Farmed Macrobrachium rosenbergii.","authors":"Amala Jiji, Nandiath Karayi Sanil, Nalini Poojary, Jeena Kezhedath, Sreedharan Krishnan, Megha K Bedekar, K V Rajendran","doi":"10.1111/jfd.14165","DOIUrl":"https://doi.org/10.1111/jfd.14165","url":null,"abstract":"<p><p>The freshwater prawn Macrobrachium rosenbergii is widely farmed in India, and except for Macrobrachium nodavirus (MrNV), no significant pathogens infecting this species have been reported. In this perspective, a histology- and PCR-based screening of M. rosenbergii was carried out for pathogens such as hepatopancreatic parvovirus (HPV), Enterocytozoon hepatopenaei (EHP) and infectious hypodermal and haematopoietic necrosis virus (IHHNV). For this, 335 samples were collected from five locations in Kerala, Maharashtra and West Bengal, India. Fifty animals, consisting of post-larvae, sub-adults and adults, were subjected to histological analysis, and the majority of the post-larvae (from Kerala) and a few sub-adult samples (from West Bengal) showed eosinophilic to basophilic, intranuclear inclusions, typical of HPV infection. The prevalence of HPV and EHP was found to be 61.3% and 95.4%, respectively, and 56.8% of the samples displayed multiple infections. None of the samples showed the presence of IHHNV. The sequence analysis of PCR amplicons indicated 90%-93% and 100% similarity to the capsid protein gene of HPV and the spore wall protein gene of EHP, respectively. This report formed the first documentation of HPV and EHP in M. rosenbergii from India and the first report of dual infections of these pathogens in freshwater prawn.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14165"},"PeriodicalIF":2.2,"publicationDate":"2025-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144266412","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Pilot Study on the Histological and Ultrastructural Characteristics of Major Immune Organs (Spleen, Head Kidney, and Trunk Kidney) and Analysis of Pathological Features in Sichuan Taimen (Hucho bleekeri).","authors":"Qinyao Wei, Hua Li, Yeyu Chen, Zhao Liu, Yanling Chen, Yi Yu, Quanyu Tu, Guiliang Liu, Zhaobin Song, Jun Du, Huanchao Yang","doi":"10.1111/jfd.14168","DOIUrl":"https://doi.org/10.1111/jfd.14168","url":null,"abstract":"<p><p>Sichuan taimen (Hucho bleekeri) is a glacial relict and critically endangered fish species restricted to the Yangtze River drainage in China. The lack of knowledge regarding the morphology of immune organs may limit the timely detection of health threats, thereby hindering effective conservation efforts. To address this gap, the pilot study provides the first histological and ultrastructural characterisation of the main immune organs (spleen, head kidney, and trunk kidney) of H. bleekeri. The spleen comprises red and white pulp; the red pulp is abundant in erythrocytes, while the white pulp contains leukocytes within a reticular network. Melano-macrophage centers, involved in immune and phagocytic activities, are prominent in the spleen and kidneys. The head kidney functions as a primary haematopoietic organ, exhibiting distinct lymphoid and non-lymphoid zones supported by a reticular framework. The trunk kidney serves an excretory role, and its tissue contains dispersed renal corpuscles. Ultrastructural analyses revealed typical cellular features, including well-developed rough endoplasmic reticulum and melano-macrophage-associated lysosomes. Notably, structural abnormalities-including mitochondrial swelling, cristae fragmentation, and mild dilation of the rough endoplasmic reticulum-were observed in the head kidney; these may potentially reflect early immune activation or sublethal environmental stress. These baseline findings would provide critical morphological references and lay a foundation for future studies on immune function, disease pathology, and conservation strategies in H. bleekeri.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14168"},"PeriodicalIF":2.2,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144248241","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Co-Infection of Fungi and Bacteria in the Gills of Kuruma Shrimp (Penaeus japonicus) Afflicted by Fusarium Disease.","authors":"Yohei Sakamoto, Kentaro Imaizumi, Satoshi Kawato, Miho Furukawa, Kayo Konishi, Reiko Nozaki, Keiichiro Koiwai, Hidehiro Kondo, Ikuo Hirono","doi":"10.1111/jfd.14167","DOIUrl":"https://doi.org/10.1111/jfd.14167","url":null,"abstract":"<p><p>Fusarium disease is an infectious condition that occurs in the cultivation of kuruma shrimp (Penaeus japonicus), primarily presenting as lesions in the gills. These lesions contain proteolytic enzymes of likely bacterial origin. This is the first study to demonstrate fungal-bacterial mixed infection in gill lesions of Fusarium-infected kuruma shrimp. Tenacibaculum spp. showing proteolytic activity were isolated from the lesions, whereas Fusarium spp. showed no such activity. Two Tenacibaculum strains were identified as Tenacibaculum mesophilum based on whole-genome sequencing (WGS) using MiSeq and GridION. Comparative analysis with the type strain DSM 13764 revealed > 95% nucleotide identity and 82.7% DNA-DNA hybridization similarity, with average genome coverage of 250×. Genomic analysis predicted that the genomes encode multiple protease and antibiotic resistance genes. Histopathology and SEM revealed that Fusarium hyphae had penetrated gill tissue, accompanied by infiltration of blood cells. Both fungal hyphae and T. mesophilum co-localised around gill cells. Bacterial diversity, assessed by 16S rRNA gene sequencing (V4 region), declined with tissue disintegration. Immune-related host genes such as anti-lipopolysaccharide factor (ALF) and lactate dehydrogenase (LDH) were upregulated, suggesting innate immune activation. These findings provide novel insights into the disease pathogenesis and contribute to the establishment of early detection systems and biosecurity-based control strategies.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14167"},"PeriodicalIF":2.2,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144258146","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluating Type III Secretion System Genes (escE, esaE and eseJ) of Edwardsiella piscicida for Virulence in Japanese Flounder (Paralichthys olivaceus).","authors":"Yasuhiko Kawato, Takamitsu Sakai, Yukihiro Nakagawa, Chigusa Suezawa, Jun Okuda","doi":"10.1111/jfd.14163","DOIUrl":"https://doi.org/10.1111/jfd.14163","url":null,"abstract":"<p><p>Edwardsiella piscicida, an etiological agent of edwardsiellosis in finfish, causes severe damage to the aquaculture industry in the world. The virulence of E. piscicida is known to be associated with the type III secretion system (T3SS) which is a protein transport nanomachine that injects effector proteins directly into host cells to be resistant to phagocyte killing. The functions and importance of the T3SS genes for the virulence of E. piscicida have been identified using murine macrophages or model fish. In this study, we evaluated the contribution of the T3SS genes consisting of orf13 (escE), orf19 (esaE) and orf29/30 (eseJ) for the virulence in a natural host using Japanese flounder (Paralichthys olivaceus). Intracellular replication ability in macrophage-like cells isolated from Japanese flounder was significantly decreased in all knockout mutants compared to the wild-type strain. In the challenge test using Japanese flounder, the virulence of Δorf13 and Δorf19 mutants was significantly decreased, but not for Δorf29 and Δorf30 mutants. Thus, escE and esaE genes are considered to highly affect the virulence in Japanese flounder. However, the eseJ gene may not significantly contribute to the virulence, although it could regulate intracellular survival in phagocytes of Japanese flounder.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14163"},"PeriodicalIF":2.2,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144248240","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"First Report of Cutaneous Leiomyoma in Plotosus canius (Hamilton, 1822) From the Coastal Waters of Parangipettai, India.","authors":"Yosuva Mariasingarayan, Jeyapragash Danaraj, Singaravel Vijayapoopathi, Nguyen Dinh-Hung","doi":"10.1111/jfd.14164","DOIUrl":"https://doi.org/10.1111/jfd.14164","url":null,"abstract":"<p><p>A tumour-bearing Plotosus canius was collected from the Parangipettai fish landing centre on the southeast coast of India. The neoplastic growth measured 3.5 × 2.8 × 1.5 cm and exhibited a dark red coloration interspersed with greyish-white regions. The lesion showed overlying epidermal ulceration and hyperchromatic, hyperaemic features, contrasting with the surrounding tissue that retained the species normal pigmentation. The excised tumour was vascularised and originated from the underlying musculature. Histopathological examination of the ulcerated peripheral region revealed spindle-shaped cells arranged in interlacing fascicles, possessing elongated nuclei, eosinophilic cytoplasm, and lacking significant pleomorphism or mitotic activity. The central region contained spindle-shaped cells arranged in intersecting fascicles and whorled patterns, with uniform elongated nuclei and moderately eosinophilic cytoplasm. The tumour architecture appeared well-organised, exhibiting variable cellular densities and a collagenous stroma with moderate-grade infiltration, but no signs of necrosis. This study presents the first documented case of cutaneous leiomyoma in Plotosus canius from the coastal waters of Parangipettai, India, and provides a detailed histopathological characterisation of the tumour in this species. Therefore, the potential role of environmental pollutants is speculative and warrants further investigation.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14164"},"PeriodicalIF":2.2,"publicationDate":"2025-06-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144225677","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Spleen as an Alternative Tissue for Estimating Tetracapsuloides bryosalmonae Load, Prevalence and Its Relationship With Proliferative Kidney Disease in Brown Trout.","authors":"Magnus Lauringson, Lilian Pukk, Siim Kahar, Oksana Burimski, Riho Gross, Veljo Kisand, Anti Vasemägi","doi":"10.1111/jfd.14148","DOIUrl":"https://doi.org/10.1111/jfd.14148","url":null,"abstract":"<p><p>Accurate pathogen detection is essential in fish health management and disease prevention. Pathogens often target different host tissues, and monitoring alternative target organs can provide important insights into disease progression. We evaluated the spleen as an alternative to the kidney for estimating the load and prevalence of the myxozoan parasite Tetracapsuloides bryosalmonae (Tb) and its relationships with proliferative kidney disease (PKD) in juvenile brown trout. We sampled 238 brown trout across nine rivers and quantified parasite load in both tissues using qPCR. Parasite load showed a strong positive correlation between the two tissues for both raw and log10-transformed data (Pearson's r = 0.60-0.71, Spearman's ρ = 0.78), with the spleen exhibiting, on average, 4.1-fold lower parasite load compared to the kidney. Tb was found in 191 specimens, consisting of 167 spleen and 190 kidney detections. The relationships between parasite load and PKD symptoms (renal hyperplasia and anaemia) were comparable for both tissues, and segmented regression line analysis indicated that health parameters deteriorate faster after exceeding a certain parasite load threshold. In conclusion, these results suggest that the spleen may serve as a viable alternative to the kidney for Tb monitoring, providing useful insights into Tb presence, load, and PKD progression in salmonids.</p>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":" ","pages":"e14148"},"PeriodicalIF":2.2,"publicationDate":"2025-06-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144225678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Establishment and Characterisation of Skin Cell Line of Schizothorax prenanti and Its Application Into Pathogen Infection","authors":"Yuhong Shi,&nbsp;Qi Zhang,&nbsp;Na Wang,&nbsp;Hua Tian","doi":"10.1111/jfd.14145","DOIUrl":"10.1111/jfd.14145","url":null,"abstract":"<div>\u0000 \u0000 <p><i>Schizothorax prenanti</i> is an important economic Cyprinidae fish endemic to the upper reaches of the Yangtze River in China. The wild population of <i>S. prenanti</i> continues to decline and has been listed as an endangered fish because of environmental pollution and overfishing. Herein, the skin cell line (SPSK) of <i>S. prenanti</i> was established using the tissue block method to aid in protecting <i>S. prenanti</i> at the cellular level and provide a skin cell line that can be applied in functional genomics and disease aetiology of the spring viraemia of carp virus (SVCV), which is highly infectious in Cyprinidae fish. The SPSK cell line was sub-cultured to more than 30 generations at 24°C in L-15 medium supplemented with 15% fetal bovine serum (FBS). Karyotype analysis further revealed that the chromosome number of SPSK ranged between 140 and 149, with 146 accounting for the highest proportion. Significant fluorescent signals were observed after transfection of SPSK with pEGFP-N1 and Cy3-siRNA, with a 30% and 90% transfection efficiency, respectively. Severe cytopathic effects (CPE) were also observed when SPSK was infected with SVCV, and the SVCV glycoprotein gene was detected by RT-PCR, indicating that SPSK was susceptible to SVCV. To further explore the mechanism of bacterial infection, transcriptome analysis was conducted for LPS treated SPSK cells and 9099 differentially expressed genes were identified. These genes significantly enriched into pathways including the Haematopoietic Cell Lineage and Primary immunodeficiency. Furthermore, seven predominantly expressed epidermal maker genes were identified by transcriptomic data, suggesting that SPSK cells were mainly derived from skin epidermis, composed of epidermal stem cell, Merkel cell, and immune cell. The establishment and characterisation of SPSK revealed its application in functional genomics and aetiology studies, making it a favourable tool for exploring disease control in <i>S. prenanti</i> and recovering fish resources.</p>\u0000 </div>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":"48 11","pages":""},"PeriodicalIF":2.2,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144208619","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Reassessment of Myxobolus toyamai Morphology in Koi Carp Using an Enhanced Giemsa Warming Technique: A First Report From Vietnam","authors":"Truong Dinh Hoai,&nbsp;Dang Thi Hoa,&nbsp;Doan Thi Nhinh,&nbsp;Nguyen Thi Huong Giang,&nbsp;Andrew P. Shinn","doi":"10.1111/jfd.14152","DOIUrl":"10.1111/jfd.14152","url":null,"abstract":"<div>\u0000 \u0000 <p><i>Myxobolus toyamai</i>, a well-established gill parasite of common carp, <i>Cyprinus carpio</i>, has been implicated in infections across multiple continents. Despite numerous attempts to classify this parasite based on the structural features of its polar capsules, existing staining techniques have proven inadequate, resulting in unclear structural definitions. This study represents the first report of <i>M. toyamai</i> infection in koi fish in Vietnam, along with the optimisation of a staining technique using Giemsa warming to accurately define its morphological characteristics. In addition, the study includes the analysis of the 18S rRNA gene and histopathological evaluations. The results show that the Giemsa warming staining method effectively clarifies the structure of the polar capsules, revealing distinct measurements that differ from those reported in previous studies. Furthermore, sequence analysis of the 18S rDNA gene demonstrates a high degree of similarity to <i>M. toyamai</i>, confirming its identification in the examined specimens.</p>\u0000 </div>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":"48 11","pages":""},"PeriodicalIF":2.2,"publicationDate":"2025-06-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144199348","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Molecular Diversity of Aquatic Pseudomonas spp. With Inhibitory Activity Against Saprolegnia Water Mould (Oomycota)","authors":"Edit Eszterbauer,&nbsp;Noémi Erdei,&nbsp;Patrícia Kóczán,&nbsp;Réka Vámosi,&nbsp;Milán Farkas,&nbsp;Győző L. Kaján,&nbsp;Dóra Sipos,&nbsp;Domonkos Sváb,&nbsp;Edit Kaszab","doi":"10.1111/jfd.14156","DOIUrl":"10.1111/jfd.14156","url":null,"abstract":"<div>\u0000 \u0000 <p>Several <i>Pseudomonas</i> bacterial species demonstrate potential for biological control. We examined 24 aquatic <i>Pseudomonas</i> isolates mainly from fish farms and natural waters in Hungary, with siderophore production and inhibitory effects on <i>Saprolegnia</i> water moulds. We identified six <i>Pseudomonas</i> species (<i>P. paracarnis</i>, <i>P. shahriarae</i>, <i>P. salmasensis</i>, <i>P. canadensis</i>, <i>P. chlororaphis</i> subsp. <i>aurantiaca</i> and <i>P. danubii</i>) distributed across five distinct phylogenetic clusters based on 16S ribosomal RNA and RNA polymerase sigma factor, <i>rpoD</i> gene sequences. The MALDI-TOF MS protein profiles showed species distribution patterns and clustering of the most inhibitory strains. In vitro experiments revealed that three strains of <i>P. danubii</i>, one <i>P. chlororaphis</i> subsp. <i>aurantiaca</i> and one unidentified <i>Pseudomonas</i> sp. B379-HU demonstrated the highest inhibitory activity in both bacterial culture and supernatants. Partial inhibition caused hyphae branching and nodule formation, whereas complete inhibition led to fragmentation and death of hyphae. This study is the first to document inhibitory activity in these <i>Pseudomonas</i> species, suggesting potential for water mould control. For fish health safety and to mitigate the risk of antibiotic resistance, utilising the extracellular inhibitory substances offers the most promising approach for future application.</p>\u0000 </div>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":"48 11","pages":""},"PeriodicalIF":2.2,"publicationDate":"2025-06-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144199347","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of a Visual Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Morganella morganii","authors":"Kek Heng Chua,&nbsp;Tien-Tien Vicky Lau,&nbsp;Suat Moi Puah","doi":"10.1111/jfd.14159","DOIUrl":"10.1111/jfd.14159","url":null,"abstract":"<div>\u0000 \u0000 <p><i>Morganella morganii</i> is a Gram-negative bacterium associated with foodborne diseases and histamine toxicity in fish, posing significant public health concerns. Traditional detection methods, such as polymerase chain reaction (PCR), often require costly equipment, restricting their applicability in resource-limited environments. In this study, we present the development of a cost-effective and user-friendly colorimetric loop-mediated isothermal amplification (LAMP) assay. We designed six primers targeting the urease gene and optimized the assay to produce visual results using a pH-sensitive phenol red dye. The LAMP assay delivers results in 25 min and can detect <i>M. morganii</i> at concentrations as low as 5 pg (1.16 × 10⁶ copies/reaction) and 2.6 × 10⁵ CFU/mL (2.5 × 10⁶ copies/reaction) in pure cultures, demonstrating high specificity, with no cross-reactivity against 10 other bacterial strains. A comparison with PCR methods using 30 samples revealed a 100% concordance rate, indicating robust diagnostic agreement. Additionally, the assay successfully identified <i>M. morganii</i> in spiked aquaculture water samples at concentrations up to 10⁷ CFU/mL. In summary, this LAMP assay operates at a constant temperature and provides clear visual results, serving as a potential rapid detection method for applications in aquaculture settings.</p>\u0000 </div>","PeriodicalId":15849,"journal":{"name":"Journal of fish diseases","volume":"48 11","pages":""},"PeriodicalIF":2.2,"publicationDate":"2025-05-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144191881","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}