Name: laboratory techniques used to maintain and differentiate biotypes of vibrio cholerae clinical and environmental isolates
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研究由新罕布什尔州INBRE通过机构发展奖（IDeA）支持，P20GM103506来自美国国家卫生研究院国家综合医学研究所。

注意：每个测定的时间考虑必须作为单个培养基制备需要不同的时间。例如，应将固体琼脂平板培养基充分冷却并干燥（1-2天）。额外的时间考虑（ 即单菌落和隔夜培养生长）在每个方案下规定， 见表2 。 媒体准备<ol><li> 1x磷酸缓冲盐水（PBS） <ol><li>称取4.0g NaCl，0.1g KCl，0.72g Na 2 HPO 4和0.12g KH 2 PO 4 。将组分?...

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Microbiol. 49 (11), 3739-3749 (2011).</li><li>Ghosh-Banerjee, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cholera+toxin+production+by+the+El+Tor+variant+of+Vibrio+cholerae+O1+compared+to+prototype+El+Tor+and+classical+biotypes.">Cholera toxin production by the El Tor variant of Vibrio cholerae O1 compared to prototype El Tor and classical biotypes.</a> J. Clin. Microbiol. 48 (11), 4283-4286 (2010).</li><li>Ansaruzzaman, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cholera+in+Mozambique,+variant+of+Vibrio+cholerae.">Cholera in Mozambique, variant of Vibrio cholerae.</a> Emerg. Infect. Dis. 10 (11), 2057-2059 (2004).</li><li>Ansaruzzaman, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+diversity+of+El+Tor+strains+of+Vibrio+cholerae+O1+with+hybrid+traits+isolated+from+Bangladesh+and+Mozambique.">Genetic diversity of El Tor strains of Vibrio cholerae O1 with hybrid traits isolated from Bangladesh and Mozambique.</a> Int. J. Med. Microbiol. 297 (6), 443-449 (2007).</li><li>Lan, R., Reeves, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pandemic+spread+of+cholera:+genetic+diversity+and+relationships+within+the+seventh+pandemic+clone+of+Vibrio+cholerae+determined+by+amplified+fragment+length+polymorphism.">Pandemic spread of cholera: genetic diversity and relationships within the seventh pandemic clone of Vibrio cholerae determined by amplified fragment length polymorphism.</a> J. Clin. Microbiol. 40 (1), 172-181 (2002).</li><li>Nair, G. B., Faruque, S. M., Bhuiyan, N. A., Kamruzzaman, M., Siddique, A. K., Sack, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+variants+of+Vibrio+cholerae+O1+biotype+El+Tor+with+attributes+of+the+classical+biotype+from+hospitalized+patients+with+acute+diarrhea+in+Bangladesh.">New variants of Vibrio cholerae O1 biotype El Tor with attributes of the classical biotype from hospitalized patients with acute diarrhea in Bangladesh.</a> J. Clin. Microbiol. 40 (9), 3296-3299 (2002).</li><li>Nair, G. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+of+Vibrio+cholerae+O1+strains+similar+to+pre-seventh+pandemic+El+Tor+strains+during+an+outbreak+of+gastrointestinal+disease+in+an+island+resort+in+Fiji.">Isolation of Vibrio cholerae O1 strains similar to pre-seventh pandemic El Tor strains during an outbreak of gastrointestinal disease in an island resort in Fiji.</a> J. Med. Microbiol. 55 (11), 1559-1562 (2006).</li><li>Nair, G. B., Mukhopadhyay, A. K., Safa, A., Takeda, Y., Faruque, S. M., Nair, G. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Emerging+hybrid+variants+of+Vibrio+cholerae+O1.">Emerging hybrid variants of Vibrio cholerae O1.</a> Vibrio cholerae—Genomics and Molecular Biology. , 179-190 (2008).</li><li>Safa, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+characteristics+of+Matlab+variants+of+Vibrio+cholerae+O1+that+are+hybrids+between+classical+and+El+Tor+biotypes.">Genetic characteristics of Matlab variants of Vibrio cholerae O1 that are hybrids between classical and El Tor biotypes.</a> J. Med. Microbiol. 55 (11), 1563-1569 (2006).</li><li>Nusrin, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diverse+CTX+phages+among+toxigenic+Vibrio+cholerae+O1+and+O139+strains+isolated+between+1994+and+2002+in+an+area+where+cholera+is+endemic.">Diverse CTX phages among toxigenic Vibrio cholerae O1 and O139 strains isolated between 1994 and 2002 in an area where cholera is endemic.</a> J. Clin. Microbiol. 42 (12), 5854-5856 (2004).</li><li>Carignan, B. M., Brumfield, K. D., Son, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single+nucleotide+polymorphisms+in+regulator-encoding+genes+have+an+additive+effect+on+virulence+gene+expression+in+a+Vibrio+cholerae+clinical+isolate.">Single nucleotide polymorphisms in regulator-encoding genes have an additive effect on virulence gene expression in a Vibrio cholerae clinical isolate.</a> mSphere. 1 (5), e00253 (2016).</li><li>Iwanaga, M., Yamamoto, K., Higa, N., Ichinose, Y., Nakasone, N., Tanabe, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Culture+conditions+for+stimulating+cholera+toxin+production+by+Vibrio+cholerae+O1+El+Tor.">Culture conditions for stimulating cholera toxin production by Vibrio cholerae O1 El Tor.</a> Microbiol. Immunol. 30 (11), 1075-1083 (1986).</li><li>DiRita, V. J., Claude, P., Georg, J., Mekalanos, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulatory+cascade+controls+virulence+in+Vibrio+cholerae.">Regulatory cascade controls virulence in Vibrio cholerae.</a> Proc. Natl. Acad. Sci. USA. 88 (12), 5403-5407 (1991).</li><li>Kovacikova, G., Lin, W., Skorupski, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Duel+regulation+of+genes+involved+in+acetoin+biosynthesis+and+motility/biofilm+formation+by+the+virulence+activator+AphA+and+the+acetate-responsive+Lys-R+type+regulator+AlsR+in+Vibrio+cholerae.">Duel regulation of genes involved in acetoin biosynthesis and motility/biofilm formation by the virulence activator AphA and the acetate-responsive Lys-R type regulator AlsR in Vibrio cholerae.</a> Mol. Microbiol. 57 (2), 420-433 (2005).</li><li>Vogel, H. J., Bonner, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acetylornithinase+of+Escherechia+coli:+partial+purification+and+some+properties.">Acetylornithinase of Escherechia coli: partial purification and some properties.</a> J. Biol. Chem. 218 (1), 97-106 (1956).</li><li>Son, M. S., Taylor, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+screens+and+biochemical+assays+to+characterize+Vibrio+cholerae+O1+biotypes:+classical+and+El+Tor.">Genetic screens and biochemical assays to characterize Vibrio cholerae O1 biotypes: classical and El Tor.</a> Curr. Protoc. 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在200多个确定的V。 霍乱血清群，只有O1和O139具有流行潜力。 O1血清群可以分为两种生物型：古典和El Tor。然而，已经出现了称为El Tor变体13,17的杂交菌株，具有El Tor生物型背景，并具有古典特征8,9,10,11,12,13,14,15,16,17。本手册中描述的方案旨在提供有兴趣表征和/或区分霍乱弧菌的各种临床和非临床分离株的研究者，具有可靠的多检测鉴定系统。使用可靠的多分析鉴定系统作为?...

霍乱是由食用含有水生革兰氏阴性菌霍乱弧菌的污染食物或水引起的远端小肠疾病。 霍乱的症状包括呕吐和不可控制的水样腹泻，导致严重脱水，如果不妥善治疗，将导致死亡。 V。基于细胞表面脂多糖O-抗原的结构， 霍乱弧菌可以分为200多个血清群。然而，只有2个血清群，O1和O139，已显示出流行或大流行的潜力1,2 。此外，血清群O139主要分离到东南亚3,4 ，而血清群O1在全球分布。此外，O1血清群可以分为两种主要生物型：古典和El Tor。经典生物型是造成头6头霍乱流行病1817年和1923年之间。正在进行的第七大流行病是El Tor生物型的结果，全球在环境5,6,7的环境中使古典生物型流行病学变异。最近，出现了具有古典和El Tor生物型8,​​9,10,11,12,13,14,15,16,17的区别特征的菌株，并且之后被称为El Tor变体13,17。一些El Tor变体已经显示出比以前观察到的更迅速和严重的疾病进展的升高的毒力能力，强调需要一个更全面的方法来代理身份识别和疾病预防和治疗8,9,18。虽然生物型鉴定并不立即决定治疗，但疫苗开发和未来治疗剂的进一步进步可能受益于生物型别的区别。 这里列出的第一系列协议将使调查人员能够正确维护V。 霍乱弧菌菌株在实验室设置。一致性和随后的分析需要分离物的储备准备和生长，其不是生物型依赖性的。然而，为了最佳地诱导毒力基因表达，需要独立的生物特异性培养技术。此外，本手稿中概述了各种遗传和生物化学测定的准备工作。 霍乱毒素（CT）和毒素共同作用培养的真菌（TCP）是主要调节剂ToxT在V的两种生物型中控制的两种主要的毒力因子。 霍乱弧菌 O1血清群20 。 CT是由五个CtxB组成的两部分毒素 亚单位围绕单个CtxA亚单位，并且负责与霍乱相关的快速电解质损失。 TCP是由tcp操纵子（ tcpABQCRDSTEF ）编码的IV型pilus ，并且涉及远端小肠的附着和定殖。 tcpA是tcp操纵子的第一个基因，其编码构建真菌8所必需的各个pilin亚单位。 ctxA的基因序列在经典和El Tor生物型之间是完全保守的，而ctxB和tcpA在两种生物型之间不同，但在每种生物型8中都是保守的。除了两个基本位置之外， ctxB在生物型之间是完全保守的（115和203）。在El Tor生物型中，胸腺嘧啶位于基因位置115和203，而经典生物型在这些碱基处含有胞嘧啶。 tcpA在每种生物型中都是完全保守的，但在生物型之间的多个基础上是不同的。这些遗传差异用作初级生物型鉴定标记，并且在测序包括这些位点的聚合酶链反应（PCR）扩增产物之后，可以将分离序列与野生型（WT）经典O395或WT E1 Tor N16961进行比较，以确定生物型背景的CT和TCP分别在给定的霍乱弧菌分离株中。 已经开发了许多方案来表征经典和El Tor生物型21,22,23之间的表型区别。多粘菌素B是一种肽抗生素，其在革兰氏阴性杆菌中损害外细胞膜的完整性通过多粘菌素B抗性测定可以观察到teria和多粘菌素B抗性21 。柠檬酸盐是克雷伯氏循环的主要底物，并且可以使用柠檬酸盐代谢测定法22来确定将柠檬酸盐代谢为唯一碳源的能力。 hapR编码霍乱弧菌 HapR中的全局调节子和主要群体感应调节因子，其结合各种启动子区域并调节基因和操纵子表达24 。 霍乱弧菌的一些致病菌株在hapR基因中具有天然存在的转录突变，导致毒力基因表达的密度依赖性调节丧失24,25 。使用牛奶琼脂培养基测量HapR调节的蛋白酶活性允许研究者鉴定特定分离物是否含有功能性HapR 23 。该溶血分析测试菌株分泌溶血红细胞的溶血酶的能力;溶血程度可以在血琼脂平板23上显现。运动性通常与霍乱弧菌中的毒力相关，可以使用运动性琼脂平板23进行分析。 Voges-Proskauer测定法测试菌株作为唯一碳源发酵葡萄糖并产生副产物乙偶姻21的能力 。随着El Tor变体的出现，很难预测任何给定的表型测定的结果，而没有广泛的基因型筛选，并且在推导V的生物型背景之前。 霍乱弧菌分离物，推荐进行这种测定23的装配，并将结果与表2中的参考菌株进行比较。 在这里，我们提出了一系列方案，集体利用上述方案基因型和表型检测用于更全面的表征V的方法 。 霍乱生物型此外，我们已经描述了已知V的基因型和表型区别。与常用的生物型参考菌株（WT经典O395，WT E1 Tor C6706和WT El Tor N16961; 表1 ）相比， 霍乱弧菌 El Tor变体（MQ1795和BAA-2163）。 El Tor变体的出现对先前使用的单一测定生物型表征方案的可靠性提出了挑战;然而，这种多重测定鉴定系统将允许更可靠地表征临床和环境V。 霍乱弧菌分离株

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			<td>Peqlab</td>
			<td>732-2789</td>
			<td>https://de.vwr.com/store/catalog/product.jsp?catalog_number=732-2789&#38;_DARGS=/store/cms/de.vwr.com/de_DE/header_2016111711383215.jsp_AF&#38;_dynSessConf=1766917479792147141&#38;targetURL=/store/catalog/product.jsp%3Fcatalog_number%3D732-2789&#38;lastLanguage=en&#38;/vwr/userprofiling/EditPersonalInfoFormHandler.updateLocale=&#38;_D%3AcurrentLanguage=+&#38;currentLanguage=en&#38;_D%3AlastLanguage=+&#38;_D%3A/vwr/userprofiling/EditPersonalInfoFormHandler.updateLocale=+</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Anhydrous K2HPO4</td>
			<td>Fisher Scientific</td>
			<td>P288-500</td>
			<td>https://www.fishersci.com/shop/products/potassium-phosphate-dibasic-anhydrous-crystalline-powder-certified-acs-fisher-chemical-5/p288500?searchHijack=true&#38;searchTerm=P288500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Blood Agar Plates</td>
			<td>Remel</td>
			<td>R01200</td>
			<td>Store at 4 &#176;C;&#160; http://www.remel.com/Catalog/Item.aspx?name=Blood+Agar</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Boric Acid</td>
			<td>Fisher Scientific</td>
			<td>A73-500</td>
			<td>https://www.fishersci.com/shop/products/boric-acid-crystalline-certified-acs-fisher-chemical-6/a73500?searchHijack=true&#38;searchTerm=A73500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bromothymol Blue</td>
			<td>Fisher Scientific</td>
			<td>B388-10</td>
			<td>https://www.fishersci.com/shop/products/bromothymol-blue-certified-acs-fisher-chemical/b38810?searchHijack=true&#38;searchTerm=B38810&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Cirtric acid &#183;H2O</td>
			<td>Fisher Scientific</td>
			<td>S72836-3</td>
			<td>https://www.fishersci.com/shop/products/citric-acid-monohydrate-4/s728363#?keyword=s728363</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Deoxynucleotide (dNTP) Solution Kit</td>
			<td>New England Biolabs</td>
			<td>N0446S</td>
			<td>Store at -20 &#176;C;&#160; https://www.neb.com/products/n0446-deoxynucleotide-solutionset</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Disodium EDTA</td>
			<td>Fisher Scientific</td>
			<td>S311-500</td>
			<td>https://www.fishersci.com/shop/products/ethylenediaminetetraacetic-acid-disodium-salt-dihydrate-crystalline-certified-acs-fisher-chemical-7/s311500?searchHijack=true&#38;searchTerm=S311500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">DNA Clean &#38; Concentrator&#8482; -25 Kit</td>
			<td>Zymo Research</td>
			<td>D4007</td>
			<td>http://www.zymoresearch.com/dna/dna-clean-up/zymoclean-gel-dna-recovery-kit</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">GelGreen Nucleic Acid Stain</td>
			<td>Biotium</td>
			<td>41005</td>
			<td>https://biotium.com/product/gelgreentm-nucleic-acid-gel-stain-10000x-in-water/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Genesys 10SUV-VIS Spectrophotometer</td>
			<td>Thermo Scientific</td>
			<td>840-208100</td>
			<td>https://www.thermofisher.com/order/catalog/product/840-208100?ICID=search-840-208100</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Gentra Puregene Yeast/Bact. Kit</td>
			<td>Qiagen</td>
			<td>158567</td>
			<td>https://www.qiagen.com/us/shop/sample-technologies/dna/dna-preparation/gentra-puregene-yeastbact-kit/#orderinginformation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HCl</td>
			<td>Fisher Scientific</td>
			<td>A144-212</td>
			<td>Corrosive;&#160; https://www.fishersci.com/shop/products/hydrochloric-acid-certified-acs-plus-fisher-chemical-10/a144212?searchHijack=true&#38;searchTerm=A144212&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">KCl</td>
			<td>Fisher Scientific</td>
			<td>P217-500</td>
			<td>https://www.fishersci.com/shop/products/potassium-chloride-crystalline-certified-acs-fisher-chemical-4/p217500?searchHijack=true&#38;searchTerm=P217500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">KH2PO4</td>
			<td>Fisher Scientific</td>
			<td>P285-500</td>
			<td>https://www.fishersci.com/shop/products/potassium-phosphate-monobasic-crystalline-certified-acs-fisher-chemical-5/p285500?searchHijack=true&#38;searchTerm=P285500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">KOH</td>
			<td>Fisher Scientific</td>
			<td>P250-500</td>
			<td>https://www.fishersci.com/shop/products/potassium-hydroxide-pellets-certified-acs-fisher-chemical-5/p250500?searchHijack=true&#38;searchTerm=P250500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Le Loop</td>
			<td>Decon Labs Inc.</td>
			<td>MP190-25</td>
			<td>http://deconlabs.com/products/leloop/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Le Stab</td>
			<td>Decon Labs Inc.</td>
			<td>MP186-5</td>
			<td>http://deconlabs.com/products/lestab/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MgSO4&#183;7H2O</td>
			<td>Fisher Scientific</td>
			<td>M63-500</td>
			<td>https://www.fishersci.com/shop/products/magnesium-sulfate-heptahydrate-crystalline-certified-acs-fisher-chemical-3/m63500?searchHijack=true&#38;searchTerm=M63500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mini-Sub Cell GT Horizontal Electrophoresis System</td>
			<td>Bio Rad Labs</td>
			<td>1704406</td>
			<td>http://www.bio-rad.com/en-us/product/mini-sub-cell-gt-cell?WT.srch=1&#38;WT.mc_id=aw-cbb-NA-sub_cell_systems_brand_gold&#38;WT.knsh_id=7eb1981f-a011-42a3-aece-1236ff453373</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MR-VP Broth</td>
			<td>Difco</td>
			<td>216300</td>
			<td>http://catalog.bd.com/nexus-ecat/getProductDetail?productId=216300&#38;parentCategory=&#38;parentCategoryName=&#38;categoryId=&#38;categoryName=&#38;searchUrl=%2FsearchResults%3Fkeyword%3Dmr-vp%2Bmedium%26typeOfSearch%3DproductSearch</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Na2HPO4</td>
			<td>Fisher Scientific</td>
			<td>S374-500</td>
			<td>https://www.fishersci.com/shop/products/sodium-phosphate-dibasic-anhydrous-granular-powder-certified-acs-fisher-chemical-5/s374500?searchHijack=true&#38;searchTerm=S374500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NaCl</td>
			<td>Fisher Scientific</td>
			<td>S271-10</td>
			<td>https://www.fishersci.com/shop/products/sodium-chloride-crystalline-certified-acs-fisher-chemical-6/s27110?searchHijack=true&#38;searchTerm=S27110&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NaHCO3</td>
			<td>Fisher Scientific</td>
			<td>S233-500</td>
			<td>https://www.fishersci.com/shop/products/sodium-bicarbonate-powder-certified-acs-fisher-chemical-5/s233500?searchHijack=true&#38;searchTerm=S233500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NaNH4HPO4&#183;4H2O</td>
			<td>Fisher Scientific</td>
			<td>S218-500</td>
			<td>https://www.fishersci.com/shop/products/sodium-ammonium-phosphate-tetrahydrate-crystalline-certified-fisher-chemical/s218500?searchHijack=true&#38;searchTerm=S218500&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NanoDrop Lite Spectrophtometer</td>
			<td>Thermo Scientific</td>
			<td>ND-LITE-PR</td>
			<td>https://www.thermofisher.com/order/catalog/product/ND-LITE-PR?ICID=search-ND-LITE-PR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Nonfat dry milk</td>
			<td>Nestle Carnation</td>
			<td>N/A</td>
			<td>N/A</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Peptone</td>
			<td>Becto, Dickinson and Co.</td>
			<td>211677</td>
			<td>http://catalog.bd.com/nexus-ecat/getProductDetail?productId=211677&#38;parentCategory=&#38;parentCategoryName=&#38;categoryId=&#38;categoryName=&#38;searchUrl=%2FsearchResults%3Fkeyword%3D211677%26typeOfSearch%3DproductSearch</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Petri Dishes (100 mm x 15 mm)</td>
			<td>Fisher Scientific</td>
			<td>FB0875712</td>
			<td>https://www.fishersci.com/shop/products/fisherbrand-petri-dishes-clear-lid-12/fb0875712#?keyword=FB0875712</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Petri Dishes (150 mm x 15 mm)</td>
			<td>Fisher Scientific</td>
			<td>FB0875714</td>
			<td>https://www.fishersci.com/shop/products/fisherbrand-petri-dishes-clear-lid-12/fb0875714?searchHijack=true&#38;searchTerm=FB0875714&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Polymyxin B sulfate salt</td>
			<td>Sigma-Aldrich</td>
			<td>P1004-10MU</td>
			<td>Store at 2-4 &#176;C; http://www.sigmaaldrich.com/catalog/product/sial/p1004?lang=en&#38;region=US</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Taq DNA Polymerase</td>
			<td>New England Biolabs</td>
			<td>M0273S</td>
			<td>Store at -20 &#176;C; https://www.neb.com/products/m0273-taq-dna-polymerase-with-standard-taq-buffer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Taq Reaction Buffer</td>
			<td>New England Biolabs</td>
			<td>M0273S</td>
			<td>Store at -20 &#176;C;&#160; https://www.neb.com/products/m0273-taq-dna-polymerase-with-standard-taq-buffer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Thermal Cycler Bio-Rad C1000 Touch&#8482;&#160;</td>
			<td>Bio Rad Labs</td>
			<td>1840148</td>
			<td>http://www.bio-rad.com/evportal/evolutionPortal.portal?_nfpb=true&#38;_pageLabel=search_page&#38;sfMode=search&#38;sfStartNumber=1&#38;clearQR=true&#38;js=1&#38;searchString=1840148&#38;database=productskus+productcategories+productdetails+abdProductDetails+msds+literatures+inserts+faqs+downloads+webpages+assays+genes+pathways+plates+promotions&#38;tabName=DIVISIONNAME</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Triphenyltetrazolium chloride</td>
			<td>Alfa Aesar</td>
			<td>A10870</td>
			<td>https://www.alfa.com/en/catalog/A10870/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tris Base</td>
			<td>Fisher Scientific</td>
			<td>BP152-1</td>
			<td>https://www.fishersci.com/shop/products/tris-base-white-crystals-crystalline-powder-molecular-biology-fisher-bioreagents-7/bp1521?searchHijack=true&#38;searchTerm=BP1521&#38;searchType=RAPID</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tris&#8729;HCl</td>
			<td>Calbiochem</td>
			<td>9310</td>
			<td>http://www.emdmillipore.com/US/en/product/OmniPur-TRIS-Hydrochloride---CAS-1185-53-1---Calbiochem,EMD_BIO-9310-OP</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tryptone</td>
			<td>Becto, Dickinson and Co.</td>
			<td>211705</td>
			<td>http://catalog.bd.com/nexus-ecat/getProductDetail?productId=211705&#38;parentCategory=&#38;parentCategoryName=&#38;categoryId=&#38;categoryName=&#38;searchUrl=%2FsearchResults%3Fkeyword%3D211705%26typeOfSearch%3DproductSearch</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Yeast Extract</td>
			<td>Becto, Dickinson and Co.</td>
			<td>212750</td>
			<td>http://catalog.bd.com/nexus-ecat/getProductDetail?productId=212750&#38;parentCategory=&#38;parentCategoryName=&#38;categoryId=&#38;categoryName=&#38;searchUrl=%2FsearchResults%3Fkeyword%3D212750%26typeOfSearch%3DproductSearch</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">&#945;-napthol</td>
			<td>MP Biomedicals</td>
			<td>204189</td>
			<td>http://www.mpbio.com/product.php?pid=05204189</td>
		</tr>
	</tbody>
</table>

laboratory techniques used to maintain and differentiate biotypes of vibrio cholerae clinical and environmental isolates

水生革兰氏阴性菌霍乱弧菌是感染性胃肠道霍乱的病原体。由于这种疾病的全球流行和严重程度， V 。 霍乱弧菌已经在环境和实验室环境中得到广泛的研究，需要适当的维护和培养技术。古典和El Tor是组成V的两个主要生物型。 霍乱弧菌 O1血清群，各自显示独特的基因型和表型特征，为生物型表征提供可靠的机制，并需要不同的毒力诱导培养条件。不管任何给定的感染或爆发的致病菌株的生物型是什么，该疾病的标准治疗涉及补充有抗生素方案的补液治疗。然而，生物型分类对于实验室研究可能是必要的，并且可能在生物医学领域具有更广泛的影响。在2000年初，鉴定出了经典和El Tor生物型的基因型和表型性状的临床分离株。新鉴定的杂种，称为El Tor变体，使得临床和环境分离生物型鉴定变得比以前的传统单试验鉴定方案更复杂。除了描述V。 霍乱弧菌一般维护和培养技术，该手稿描述了一系列基于特定基因（ ctxB和tcpA ）基于PCR的遗传筛选和表型测定（多粘菌素B抗性，柠檬酸盐代谢，蛋白水解活性，溶血活性，运动性和葡萄糖代谢通过Voges- Proskauer测定）共同用于表征和/或区分经典和El Tor生物型。一起，这些测定提供了一种有效的系统方法，作为替代使用，或另外用于代表性的昂贵的劳动密集型实验V。 霍乱临床（和环境）分离株。

为了妥善维护和使用任何细菌菌株，建议知道感兴趣的菌株的倍增时间。这里，常用的V的变化的生长速率。通过生长曲线证实霍乱弧菌菌株，并使用线性回归计算近似倍数。 WT El Tor N16961和El Tor变体MQ1795比WT经典O395（〜2 h）显示更短的倍增时间（分别为〜1 h和〜1 h）（ 图1 ; 表2 ）。 <p class="jove_content" fo:keep-together.within-pa...

Watch this Scientific Journal Video about Laboratory Techniques Used to Maintain and Differentiate Biotypes of Vibrio cholerae Clinical and Environmental Isolates at JoVE.com

Laboratory Techniques Used to Maintain and Differentiate Biotypes of Vibrio cholerae Clinical and Environmental Isolates (Video) | JoVE

该手稿除了进行一系列生物化学测定之外，还描述了适用于霍乱弧菌的维护技术，它们共同用于临床和环境V之间的快速可靠的分化。 霍乱弧菌生物型实验室设置。

用于维持和区分生物型的实验室技术<em&gt;霍乱弧菌</em&gt;临床和环境隔离

The aquatic Gram-negative bacterium Vibrio cholerae is the etiological agent of the infectious gastrointestinal disease cholera. Due to the global ...

Research

JoVE Journal

Immunology and Infection

Laboratory Techniques Used to Maintain and Differentiate Biotypes of Vibrio cholerae Clinical and Environmental Isolates

Using a Pan-Viral Microarray Assay (Virochip) to Screen Clinical Samples for Viral Pathogens

Using a Pan-Viral Microarray Assay Virochip to Screen Clinical Samples for Viral Pathogens (Video) | JoVE

使用泛病毒基因芯片检测（Virochip）屏幕病毒病原体的临床标本

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科研

免疫与感染

Multiplex Detection of Bacteria in Complex Clinical and Environmental Samples using Oligonucleotide-coupled Fluorescent Microspheres

Multiplex Detection of Bacteria in Complex Clinical and Environmental Samples using Oligonucleotide-coupled Fluorescent Microspheres (Video) | JoVE

在复杂的临床和环境样本使用寡核苷酸加上荧光微多元检测细菌

Bacterial vaginosis (BV) is a recurring polymicrobial syndrome that is characterized by a change in the "normal" microbiota from Lactobacillus-dominated ...

Colonization of Euprymna scolopes Squid by Vibrio fischeri

Colonization of Euprymna scolopes Squid by Vibrio fischeri (Video) | JoVE

定植 Euprymna scolopes鱿鱼由弧菌鲵</em

Specific bacteria are found in association with animal tissue1-5. Such host-bacterial associations (symbioses) can be detrimental (pathogenic), have no ...

TransFLP &#x2014; A Method to Genetically Modify Vibrio cholerae Based on Natural Transformation and FLP-recombination

TransFLP Ã¢â‚¬â€ A Method to Genetically Modify Vibrio cholerae Based on Natural Transformation and FLP-recombination (Video) | JoVE

TransFLP - A基因修改方法<em&gt;霍乱弧菌</em基于自然转化和FLP重组

Several methods are available to manipulate bacterial chromosomes1-3. Most of these protocols rely on the insertion of conditionally replicative plasmids ...

A Modified EPA Method 1623 that Uses Tangential Flow Hollow-fiber Ultrafiltration and Heat Dissociation Steps to Detect Waterborne Cryptosporidium and Giardia spp.

A Modified EPA Method 1623 that Uses Tangential Flow Hollow-fiber Ultrafiltration and Heat Dissociation Steps to Detect Waterborne Cryptosporidium and Giardia spp. (Video) | JoVE

一种改进的EPA方法1623使用切向流中空纤维超滤和热解离步骤检测水性<em&gt;隐</em&gt;和<em&gt;贾第虫属。</em

Cryptosporidium and Giardia species are two of the most prevalent protozoa that cause waterborne diarrheal disease outbreaks worldwide. To better ...

Safety Precautions and Operating Procedures in an (A)BSL-4 Laboratory: 1. Biosafety Level 4 Suit Laboratory Suite Entry and Exit Procedures

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安全注意事项和操作步骤在（A）BSL-4实验室：1.生物安全4级实验室西服套房出入境手续

Biosafety level 4 (BSL-4) suit laboratories are specifically designed to study high-consequence pathogens for which neither infection prophylaxes nor ...

A Rapid Strategy for the Isolation of New Faustoviruses from Environmental Samples Using Vermamoeba vermiformis

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一种快速战略新Faustoviruses从环境样品使用隔离<em&gt; Vermamoeba vermiformis</em

The isolation of giant viruses is of great interest in this new era of virology, especially since these giant viruses are related to protists. Giant ...

Developing a Salivary Antibody Multiplex Immunoassay to Measure Human Exposure to Environmental Pathogens

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制定唾液抗体多重免疫测定来测量人体暴露于环境中的病原体

The etiology and impacts of human exposure to environmental pathogens are of major concern worldwide and, thus, the ability to assess exposure and ...

Development of a More Sensitive and Specific Chromogenic Agar Medium for the Detection of Vibrio parahaemolyticus and Other Vibrio Species

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对于检测到的敏感性和特异性显色琼脂培养基发展<em&gt;副溶血性弧菌</em&gt;及其他<em&gt;弧菌</em&gt;物种

Foodborne infections in the US caused by Vibrio species have shown an upward trend. In the genus Vibrio, V. parahaemolyticus is responsible for the ...

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