{"title":"水蛭壶菌游动孢子活力测定方法的比较。","authors":"Adeline Chew, Matt West, Laura A Brannelly","doi":"10.3354/dao03749","DOIUrl":null,"url":null,"abstract":"<p><p>The emerging fungal pathogen Batrachochytrium dendrobatidis (Bd) threatens hundreds of amphibian species globally. During laboratory-based experiments it is often essential to quantify live Bd cells, but a comparison of the effectiveness of methods for counting and assessing the viability of the infectious zoospore life stage has not been done. A direct comparison of staining methods that assess viability will ensure that the most accurate and efficient method is used. Here, we compared the use of 2 relatively cheap common stains, trypan blue and methylene blue, and assessed their accuracy and precision for estimating the viability of Bd zoospores during both manual counting and colorimetric assays. We stained known proportions of killed Bd zoospores (0, 0.25, 0.50, 0.75, and 1.00) with each stain and estimated the proportion of stained (dead) and unstained (viable) cells in each sample using both manual counting and colorimetric assays. Trypan blue was found to be a much more effective stain than methylene blue for both microscopy and colorimetric assays. Additionally, counting zoospores via microscopy was both a more accurate and precise technique. We recommend using manual counts via microscopy using the trypan blue stain for assessing Bd zoospore viability.</p>","PeriodicalId":11252,"journal":{"name":"Diseases of aquatic organisms","volume":null,"pages":null},"PeriodicalIF":1.1000,"publicationDate":"2023-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Comparison of methods for determining Batrachochytrium dendrobatidis zoospore viability.\",\"authors\":\"Adeline Chew, Matt West, Laura A Brannelly\",\"doi\":\"10.3354/dao03749\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>The emerging fungal pathogen Batrachochytrium dendrobatidis (Bd) threatens hundreds of amphibian species globally. During laboratory-based experiments it is often essential to quantify live Bd cells, but a comparison of the effectiveness of methods for counting and assessing the viability of the infectious zoospore life stage has not been done. A direct comparison of staining methods that assess viability will ensure that the most accurate and efficient method is used. Here, we compared the use of 2 relatively cheap common stains, trypan blue and methylene blue, and assessed their accuracy and precision for estimating the viability of Bd zoospores during both manual counting and colorimetric assays. We stained known proportions of killed Bd zoospores (0, 0.25, 0.50, 0.75, and 1.00) with each stain and estimated the proportion of stained (dead) and unstained (viable) cells in each sample using both manual counting and colorimetric assays. Trypan blue was found to be a much more effective stain than methylene blue for both microscopy and colorimetric assays. Additionally, counting zoospores via microscopy was both a more accurate and precise technique. We recommend using manual counts via microscopy using the trypan blue stain for assessing Bd zoospore viability.</p>\",\"PeriodicalId\":11252,\"journal\":{\"name\":\"Diseases of aquatic organisms\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.1000,\"publicationDate\":\"2023-09-14\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Diseases of aquatic organisms\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://doi.org/10.3354/dao03749\",\"RegionNum\":4,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"FISHERIES\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Diseases of aquatic organisms","FirstCategoryId":"97","ListUrlMain":"https://doi.org/10.3354/dao03749","RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"FISHERIES","Score":null,"Total":0}
Comparison of methods for determining Batrachochytrium dendrobatidis zoospore viability.
The emerging fungal pathogen Batrachochytrium dendrobatidis (Bd) threatens hundreds of amphibian species globally. During laboratory-based experiments it is often essential to quantify live Bd cells, but a comparison of the effectiveness of methods for counting and assessing the viability of the infectious zoospore life stage has not been done. A direct comparison of staining methods that assess viability will ensure that the most accurate and efficient method is used. Here, we compared the use of 2 relatively cheap common stains, trypan blue and methylene blue, and assessed their accuracy and precision for estimating the viability of Bd zoospores during both manual counting and colorimetric assays. We stained known proportions of killed Bd zoospores (0, 0.25, 0.50, 0.75, and 1.00) with each stain and estimated the proportion of stained (dead) and unstained (viable) cells in each sample using both manual counting and colorimetric assays. Trypan blue was found to be a much more effective stain than methylene blue for both microscopy and colorimetric assays. Additionally, counting zoospores via microscopy was both a more accurate and precise technique. We recommend using manual counts via microscopy using the trypan blue stain for assessing Bd zoospore viability.
期刊介绍:
DAO publishes Research Articles, Reviews, and Notes, as well as Comments/Reply Comments (for details see DAO 48:161), Theme Sections and Opinion Pieces. For details consult the Guidelines for Authors. Papers may cover all forms of life - animals, plants and microorganisms - in marine, limnetic and brackish habitats. DAO''s scope includes any research focusing on diseases in aquatic organisms, specifically:
-Diseases caused by coexisting organisms, e.g. viruses, bacteria, fungi, protistans, metazoans; characterization of pathogens
-Diseases caused by abiotic factors (critical intensities of environmental properties, including pollution)-
Diseases due to internal circumstances (innate, idiopathic, genetic)-
Diseases due to proliferative disorders (neoplasms)-
Disease diagnosis, treatment and prevention-
Molecular aspects of diseases-
Nutritional disorders-
Stress and physical injuries-
Epidemiology/epizootiology-
Parasitology-
Toxicology-
Diseases of aquatic organisms affecting human health and well-being (with the focus on the aquatic organism)-
Diseases as indicators of humanity''s detrimental impact on nature-
Genomics, proteomics and metabolomics of disease-
Immunology and disease prevention-
Animal welfare-
Zoonosis