University of Maryland Eastern Shore, Princess Anne, Maryland, USA
U.S. National Oceanic and Atmospheric Administration, National Ocean Service, National Centers for Coastal Ocean Science, Marine Spatial Ecology, Oxford, Maryland, USA.
Appl Environ Microbiol. 2020 Nov 10;86(23). doi: 10.1128/AEM.01581-20.
Oyster and seawater samples were collected from five sites in the Chesapeake Bay, MD, and three sites in the Delaware Bay, DE, from May to October 2016 and 2017. Abundances and detection frequencies for total and pathogenic and were compared using the standard most-probable-number-PCR (MPN-PCR) assay and a direct-plating (DP) method on CHROMagar Vibrio for total ( ) and pathogenic ( and ) genes and total () and pathogenic () genes. The colony overlay procedure for peptidases (COPP) assay was evaluated for total DP had high false-negative rates (14 to 77%) for most PCR targets and was deemed unsatisfactory. Logistic regression models of the COPP assay showed high concordances with MPN-PCR for and and in oysters (85.7 to 90.9%) and seawater (81.1 to 92.7%) when seawater temperature and salinity were factored into the model, suggesting that the COPP assay could potentially serve as a more rapid method to detect vibrios in oysters and seawater. Differences in total and pathogenic abundances between state sampling sites over different collection years were contrasted for oysters and seawater by MPN-PCR. Abundances of and were ∼8-fold higher in Delaware oysters than in Maryland oysters, whereas abundances of were nearly identical. For Delaware oysters, 93.5% were both and , compared to only 19.2% in Maryland. These results indicate that pathogenic was more prevalent in the Delaware Bay than in the Chesapeake Bay. While and cause shellfish-associated morbidity and mortality among shellfish consumers, current regulatory assays for vibrios are complex, time-consuming, labor-intensive, and relatively expensive. In this study, the rapid, simple, and inexpensive COPP assay was identified as a possible alternative to MPN-PCR for shellfish monitoring. This paper shows differences in total and pathogenic vibrios found in seawater and oysters from the commercially important Delaware and Chesapeake Bays. isolates from the Delaware Bay were more likely to contain commonly recognized pathogenicity genes than those from the Chesapeake Bay.
2016 年和 2017 年 5 月至 10 月,从马里兰州切萨皮克湾的五个地点和特拉华州特拉华湾的三个地点采集了牡蛎和海水样本。使用标准最可能数 PCR(MPN-PCR)检测法和直接平板(DP)法对总菌( )和致病菌( 和 )的 和 基因、总菌( )和致病菌( 和 )的 基因进行了比较。对蛋白酶菌落覆盖程序(COPP)检测法进行了评估,用于总菌的 DP 对大多数 PCR 靶标具有很高的假阴性率(14%至 77%),因此被认为不理想。COPP 检测法的逻辑回归模型显示,在将海水温度和盐度纳入模型后,与 MPN-PCR 对牡蛎(85.7%至 90.9%)和海水(81.1%至 92.7%)中的 和 以及 具有很高的一致性,表明 COPP 检测法有可能成为一种更快的方法来检测牡蛎和海水中的弧菌。通过 MPN-PCR 比较了不同年份在不同州采样点的牡蛎和海水中总菌 和致病菌 丰度的差异。与马里兰州的牡蛎相比,特拉华州的牡蛎中 和 的丰度高约 8 倍,而 的丰度几乎相同。对于特拉华州的牡蛎,93.5%的同时含有 和 ,而马里兰州只有 19.2%。这些结果表明,致病菌 在特拉华湾比在切萨皮克湾更为普遍。虽然 和 会导致贝类消费者贝类相关的发病率和死亡率,但目前用于弧菌的监管检测方法复杂、耗时、劳动密集且相对昂贵。在这项研究中,快速、简单且廉价的 COPP 检测法被确定为贝类监测中替代 MPN-PCR 的可能方法。本文显示了来自商业上重要的特拉华湾和切萨皮克湾的海水和牡蛎中总菌 和致病菌的差异。来自特拉华湾的 分离株比来自切萨皮克湾的分离株更有可能含有公认的致病性基因。