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Novel immunoassay for diagnosis of ongoing Clostridioides difficile infections using serum and medium enriched for newly synthesized antibodies (MENSA)
Journal of Immunological Methods ( IF 1.6 ) Pub Date : 2020-11-19 , DOI: 10.1016/j.jim.2020.112932
Natalie S Haddad 1 , Sophia Nozick 1 , Geena Kim 1 , Shant Ohanian 1 , Colleen Kraft 2 , Paulina A Rebolledo 3 , Yun Wang 4 , Hao Wu 5 , Adam Bressler 6 , Sang Nguyet Thi Le 7 , Merin Kuruvilla 7 , L Edward Cannon 1 , F Eun-Hyung Lee 8 , John L Daiss 9
Affiliation  

Background

Clostridioides difficile infections (CDI) have been a challenging and increasingly serious concern in recent years. While early and accurate diagnosis is crucial, available assays have frustrating limitations.

Objective

Develop a simple, blood-based immunoassay to accurately diagnose patients suffering from active CDI.

Materials and methods

Uninfected controls (N = 95) and CDI patients (N = 167) were recruited from Atlanta area hospitals. Blood samples were collected from patients within twelve days of a positive CDI test and processed to yield serum and PBMCs cultured to yield medium enriched for newly synthesized antibodies (MENSA). Multiplex immunoassays measured Ig responses to ten recombinant C. difficile antigens.

Results

Sixty-six percent of CDI patients produced measurable responses to C. difficile antigens in their serum or MENSA within twelve days of a positive CDI test. Fifty-two of the 167 CDI patients (31%) were detectable in both serum and MENSA, but 32/167 (19%) were detectable only in MENSA, and 27/167 (16%) were detectable only in serum.

Discussion

We describe the results of a multiplex immunoassay for the diagnosis of ongoing CDI in hospitalized patients. Our assay resolved patients into four categories: MENSA-positive only, serum-positive only, MENSA- and serum-positive, and MENSA- and serum-negative. The 30% of patients who were MENSA-positive only may be accounted for by nascent antibody secretion prior to seroconversion. Conversely, the serum-positive only subset may have been more advanced in their disease course. Immunocompromise and misdiagnosis may have contributed to the 34% of CDI patients who were not identified using MENSA or serum immunoassays.

Importance

While there was considerable overlap between patients identified through MENSA and serum, each method detected a distinctive patient group. The combined use of both MENSA and serum to detect CDI patients resulted in the greatest identification of CDI patients. Together, longitudinal analysis of MENSA and serum will provide a more accurate evaluation of successful host humoral immune responses in CDI patients.



中文翻译:


使用富含新合成抗体的血清和培养基诊断正在进行的艰难梭菌感染的新型免疫测定法 (MENSA)


 背景


近年来,艰难梭菌感染(CDI)一直是一个具有挑战性且日益严重的问题。虽然早期准确的诊断至关重要,但现有的检测方法存在令人沮丧的局限性。

 客观的


开发一种简单的基于血液的免疫测定法来准确诊断患有活动性 CDI 的患者。

 材料和方法


从亚特兰大地区医院招募未感染对照者 ( N = 95) 和 CDI 患者 ( N = 167)。在 CDI 检测呈阳性后 12 天内从患者身上采集血样,并进行处理以产生血清和 PBMC,培养后产生富含新合成抗体 (MENSA) 的培养基。多重免疫测定测量了对十种重组艰难梭菌抗原的 Ig 反应。

 结果


66% 的 CDI 患者在 CDI 检测呈阳性后 12 天内对其血清或 MENSA 中的艰难梭菌抗原产生了可测量的反应。 167 名 CDI 患者中,52 名 (31%) 在血清和 MENSA 中均可检测到,但 32/167 (19%) 仅在 MENSA 中可检测到,27/167 (16%) 仅在血清中可检测到。

 讨论


我们描述了用于诊断住院患者持续性 CDI 的多重免疫测定结果。我们的检测将患者分为四类:仅 MENSA 阳性、仅血清阳性、MENSA 和血清阳性、以及 MENSA 和血清阴性。 30% 的 MENSA 阳性患者可能是由于血清转化前新生抗体分泌所致。相反,仅血清阳性的亚群可能在其病程中处于更晚期。 34% 的 CDI 患者未能通过 MENSA 或血清免疫测定法确诊,可能是免疫功能低下和误诊造成的。

 重要性


虽然通过 MENSA 和血清鉴定的患者之间存在相当大的重叠,但每种方法都检测到了独特的患者群体。结合使用 MENSA 和血清来检测 CDI 患者,可以最大程度地识别 CDI 患者。总之,MENSA 和血清的纵向分析将为 CDI 患者成功的宿主体液免疫反应提供更准确的评估。

更新日期:2020-11-19
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