Name: 作者聚焦：口腔 念珠菌 诊断推进 pSS 患者的临床治疗方案
Uploaded: 2024-03-01T14:25:00
Description: Watch this Scientific Journal Video about Author Spotlight: Oral Candida Diagnosis to Advance Clinical Treatment Regimen for pSS Patients at JoVE.com

本研究得到国家自然科学基金（82201084）、中国博士后科学基金（2022M722232）、北京市博士后科研基金（2023-ZZ-020）、首都医科大学附属北京天坛医院苗浚项目（2023MP10）和香港研究资助局优配研究金（27111820和17116521）资助。

下述所有程序均经首都医科大学附属北京天坛医院伦理委员会批准（编号：KY2023-177-01），参与本研究的所有患者均提供知情同意书。
1. 患者纳入和排除标准
<ol><li>纳入标准：如果患者满足 2016 年美国风湿病学会 （ACR）/欧洲抗风湿病联盟 （EULAR） 标准12，则将患者分类为 pSS 患者。具体来说，最终分类标准基于五个项目的加权总和：抗 SSA/Ro 抗...

pSS 相关口腔念珠菌的诊断和菌株鉴定

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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oral+complications+of+Sj&#246;gren's+syndrome.">Oral complications of Sj&#246;gren's syndrome.</a> Oral Maxillofac Surg Clin North Am. 26 (1), 55-62 (2014).</li><li>Mathews, S. A., Kurien, B. T., Scofield, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oral+manifestations+of+Sj&#246;gren's+syndrome.">Oral manifestations of Sj&#246;gren's syndrome.</a> J Dent Res. 87 (4), 308-318 (2008).</li><li>Leung, K. C. M., McMillan, A. S., Cheung, B. P. K., Leung, W. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sj&#246;gren's+syndrome+sufferers+have+increased+oral+yeast+levels+despite+regular+dental+care.">Sj&#246;gren's syndrome sufferers have increased oral yeast levels despite regular dental care.</a> Oral Dis. 14 (2), 163-173 (2008).</li><li>Lundstrom, I. M., Lindstrom, F. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subjective+and+clinical+oral+symptoms+in+patients+with+primary+Sj&#246;gren's+syndrome.">Subjective and clinical oral symptoms in patients with primary Sj&#246;gren's syndrome.</a> Clin Exp Rheumatol. 13 (6), 725-731 (1995).</li><li>Alam, M. Z., 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=Candida+identification:+a+journey+from+conventional+to+molecular+methods+in+medical+mycology.">Candida identification: a journey from conventional to molecular methods in medical mycology.</a> World J Microbiol Biotechnol. 30 (5), 1437-1451 (2014).</li><li>Bauters, T. G., Nelis, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+chromogenic+and+fluorogenic+membrane+filtration+methods+for+detection+of+four+Candida+species.">Comparison of chromogenic and fluorogenic membrane filtration methods for detection of four Candida species.</a> J Clin Microbiol. 40 (5), 1838-1839 (2002).</li><li>Soto-Rojas, A. E., Villa, A. R., Sifuentes-Osornio, J., Alarc&#243;n-Segovia, D., Kraus, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oral+candidiasis+and+Sj&#246;gren's+syndrome.">Oral candidiasis and Sj&#246;gren's syndrome.</a> J Rheumatol. 25 (5), 911-915 (1998).</li><li>Gilchrist, J. E., Campbell, J. E., Donnelly, C. B., Peeler, J. T., Delaney, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spiral+plate+method+for+bacterial+determination.">Spiral plate method for bacterial determination.</a> Appl Microbiol. 25 (2), 244-252 (1973).</li><li>Biomerieux. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ATB+FUNGUS+3+Sensitivity+test+for+yeast-like+fungi.+REF+14204.">ATB FUNGUS 3 Sensitivity test for yeast-like fungi. REF 14204.</a> Biomerieux. , (2014).</li><li>Zhang, L., 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=The+widely+used+ATB+FUNGUS+3+automated+readings+in+China+and+its+misleading+high+MICs+of+Candida+spp.+to+Azoles:+Challenges+for+developing+countries'+clinical+microbiology+labs.">The widely used ATB FUNGUS 3 automated readings in China and its misleading high MICs of Candida spp. to Azoles: Challenges for developing countries' clinical microbiology labs.</a> PLoS One. 9 (12), e114004 (2014).</li><li>Hu, L., Zhou, P., Zhao, W., Hua, H., Yan, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescence+staining+vs.+routine+KOH+smear+for+rapid+diagnosis+of+oral+candidiasis-A+diagnostic+test.">Fluorescence staining vs. routine KOH smear for rapid diagnosis of oral candidiasis-A diagnostic test.</a> Oral Dis. 26 (5), 941-947 (2020).</li><li>Samaranayake, L. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Candida+krusei+infections+and+fluconazole+therapy.">Candida krusei infections and fluconazole therapy.</a> Hong Kong Med J. 3 (3), 312-314 (1997).</li><li>Lee, Y., Puumala, E., Robbins, N., Cowen, L. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antifungal+drug+resistance:+Molecular+mechanisms+in+Candida+albicans+and+beyond.">Antifungal drug resistance: Molecular mechanisms in Candida albicans and beyond.</a> Chem Rev. 121 (6), 3390-3411 (2021).</li><li>Rihab, B., Lina, E. H., No&#233;mie, S. T., Jean, S., Marjolaine, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+experience+of+dry+mouth+and+screening+for+Sjogren's+syndrome+by+the+dentist:+patient-reported+experiences.">The experience of dry mouth and screening for Sjogren's syndrome by the dentist: patient-reported experiences.</a> BMC Oral Health. 23 (1), 1010 (2023).</li><li>Panpetch, W., 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=Oral+Candida+administration+in+a+Clostridium+difficile+mouse+model+worsens+disease+severity+but+is+attenuated+by+Bifidobacterium.">Oral Candida administration in a Clostridium difficile mouse model worsens disease severity but is attenuated by Bifidobacterium.</a> PLoS One. 14 (1), e210798 (2019).</li><li>Alwaily, E. R., Abood, M. S., Al Uobody, R. 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在目前的手稿中，我们提供了一系列系统、简单、可行的方法，用于检测口腔 念珠菌 感染、鉴定 念珠菌 菌株以及检测pSS患者常用药物的抗真菌敏感性。 
在实际环境中，并非本手稿中介绍的所有方法都是必需的。口腔 念珠菌 检测方法可以根据具体目的进行选择。KOH 涂片试验和 CFW 染色试验使用未经培养的样本，可以快速、初步地确定 pSS 患者是否存在?...

念珠菌病是由 念珠菌属引起的，念珠菌 属是机会性病原体。常见菌株包括 白色念珠菌、克鲁塞念珠菌、光滑念珠菌、副念珠 菌和 都柏林念珠菌1。 念珠菌 属引起的机会性感染包括浅表 念珠菌 感染和侵袭性念珠菌病。侵袭性念珠菌病主要发生在免疫功能低下的个体中;例如，在患有获得性免疫缺陷综合症 （AIDS） 的患者中，侵袭性念珠菌病会严重威胁生活质量甚至寿命2。浅表 念珠菌 感染，如皮肤黏膜念珠菌病，在患者中更常见3. 
口腔念珠菌病是一种浅表性 念珠菌 感染，是最常见的人类真菌感染。 白色念珠菌 是口腔的正常共生;据报道，在没有任何症状的一般人群中 ，念珠菌 属的携带率为 20%-75%4。 念珠菌 的过度生长会导致患者的局部不适，例如感觉味觉改变、口腔灼热感、营养不良导致的吞咽困难、恢复延迟和住院时间延长。长期口服念珠菌病可导致严重的侵袭性念珠菌病，导致严重的发病率和死亡率5.口腔念珠菌病也会增加患口腔癌的风险5.唾液腺功能受损是口腔念珠菌病的危险因素之一6.感染的开始是 念珠菌 粘附在上皮细胞壁上。其次是 念珠菌 的增殖和丝化以及生物膜7 的形成和成熟。生物膜极难根除，并且对常规抗真菌治疗具有抵抗力，这使得临床治疗成为生物膜相关口腔 念珠菌 感染的挑战7.
原发性干燥综合征 （pSS） 是一种自身免疫性疾病，其特征是口干、眼睛干涩和其他全身症状8.口干是pSS最常见的症状。唾液具有重要的生理抗真菌功能。一方面，它可以稀释和冲刷口腔，然后从粘膜中清除微生物。另一方面，唾液中含有抗菌蛋白，如乳铁蛋白、唾液过氧化物酶、溶菌酶、富含组氨酸的多肽和特异性抗念珠菌抗体，它们可以与口腔粘膜相互作用，防止 念珠菌6的过度生长。唾液流量减少可能使 pSS 患者易患口腔念珠菌病。一项对 61 例 pSS 患者进行的观察性横断面研究发现，13.1% 的 pSS 患者出现念珠菌病的口腔体征，发现 白色念珠 菌菌落形成单位 （CFU）/mL 与未刺激的全唾液 （UWS） 和刺激的全唾液 （SWS） 水平显着且呈负相关9。
本手稿中介绍的方法可以根据直接涂片或培养后的表征形态属性（例如菌丝和酵母细胞）识别口腔念珠菌感染10。使用念珠菌诊断琼脂对念珠菌进行菌株鉴定是基于具有不同形态的不同颜色菌落的形成，这些菌落是由物种特异性酶裂解显色底物引起的11。在本文中，介绍了检测口腔念珠菌感染的系统方法，包括传统的口腔念珠菌培养和氢氧化钾（KOH）涂片试验和Calcofluor White（CFW）染色试验的快速检查10。此外，还引入了抗真菌药敏试验，可指导临床口腔念珠菌感染治疗。在实际情况下，没有必要执行本手稿中介绍的所有方法;口腔念珠菌检测方法可根据目的选择。口腔念珠菌感染的检查和 pSS 患者念珠菌菌株的测定不仅有利于疾病治疗，而且有助于评估口腔念珠菌病的特征和物种特征。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.22 &#181;m Syringe Filter</td>
			<td>MERCK</td>
			<td>SLGV004SL</td>
			<td></td>
		</tr>
		<tr>
			<td>1.5 mL Centrifuge Tube</td>
			<td>axygen</td>
			<td>MCT-150-C-S</td>
			<td></td>
		</tr>
		<tr>
			<td>10 cm Microbiological Culture Dishes</td>
			<td>Jet Bio-Filtration Co. Ltd</td>
			<td>TCD000100</td>
			<td></td>
		</tr>
		<tr>
			<td>15 mL Centrifuge Tube</td>
			<td>Jet Bio-Filtration Co., Ltd</td>
			<td>CFT011150</td>
			<td></td>
		</tr>
		<tr>
			<td>50 mL Centrifuge Tube</td>
			<td>Corning</td>
			<td>430290</td>
			<td></td>
		</tr>
		<tr>
			<td>96-well Microplates</td>
			<td>Corning</td>
			<td>3599</td>
			<td></td>
		</tr>
		<tr>
			<td>API 0.85% NaCl ampule</td>
			<td>BIOMERIEUX</td>
			<td>20070</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcofluor White Stain</td>
			<td>Sigma-Aldrich</td>
			<td>18909-100ML-F</td>
			<td></td>
		</tr>
		<tr>
			<td>CHROMagar Candida</td>
			<td>CHROMagar</td>
			<td>P002860</td>
			<td>Candida diagnostic agar</td>
		</tr>
		<tr>
			<td>D(+)-Glucose Monohydrate</td>
			<td>Sinopharm Chemical Reagent Co.,Ltd</td>
			<td>10010518</td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum&#160;</td>
			<td>Gibco</td>
			<td>26170035</td>
			<td></td>
		</tr>
		<tr>
			<td>McFarland Standards Kit</td>
			<td>BIOMERIEUX</td>
			<td>70900</td>
			<td></td>
		</tr>
		<tr>
			<td>Nunc precision molded loops&#160;</td>
			<td>ThermoFisher Scientific</td>
			<td>254399</td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate Buffered Saline</td>
			<td>Gibco</td>
			<td>C10010500BT</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium Hydroxide</td>
			<td>Aladdin</td>
			<td>P112284-500g</td>
			<td></td>
		</tr>
		<tr>
			<td>PowerSoil DNA Isolation Kit&#160;</td>
			<td>MO BIO</td>
			<td>12888-50</td>
			<td></td>
		</tr>
		<tr>
			<td>RPMI 1640 Medium</td>
			<td>Gibco</td>
			<td>11875119</td>
			<td></td>
		</tr>
		<tr>
			<td>Sabourand&#39;s Agar Medium</td>
			<td>Solarbio</td>
			<td>S9710</td>
			<td></td>
		</tr>
		<tr>
			<td>Yeast Nitrogen Base Without Amino acids</td>
			<td>Solarbio</td>
			<td>Y8040</td>
			<td></td>
		</tr>
		<tr>
			<td>Yeast-like Fungal Susceptibility Kit (Microdilution)</td>
			<td>BIOMERIEUX</td>
			<td>14204</td>
			<td></td>
		</tr>
	</tbody>
</table>

examination of oral candida infection in primary sj&#246;gren&#39;s syndrome patients

原发性干燥综合征 （pSS） 是一种自身免疫性疾病，其特征是口干、眼睛干涩和其他全身症状等症状。由于pSS患者所经历的唾液分泌减退，口腔菌群失调经常发生。pSS 的一个常见并发症是口腔 念珠菌 感染。在本文中，作者描述了可以有效诊断口腔 念珠菌 感染并使用唾液、口腔粘膜拭子或 pSS 患者漱口水识别 念珠菌 菌株的系统方法。Sabouraud 葡萄糖琼脂 （SDA）、菌丝形成试验、氢氧化钾 （KOH） 涂片试验和钙荧光白 （CFW） 染色试验用于诊断口腔 念珠菌 感染。 念珠菌 诊断琼脂用于鉴定 念珠菌 菌株。最后，使用抗真菌药敏试验来确定适当的抗真菌药物治疗。这种标准化方法可以促进pSS相关口腔 念珠菌 感染的诊断、治疗和未来研究。使用这种方法进行早期诊断还可以防止由于延迟接受适当治疗而出现的任何并发症。

Candidiasis is caused by Candida spp., which are opportunistic pathogens. Common strains include Candida albicans, Candida krusei, Candida glabrata, Candida parapsilosis, and Candida dubliniensis1. Opportunistic infections caused by Candida spp. include superficial Candida infections and invasive candidiasis. Invasive candidiasis mainly happens in immunocompromised individuals; for example, in patients with acquired immunodeficiency syndrome (AIDS), invasive candidiasis can significantly threaten the quality of life or even the life span2. Superficial Candida infection, such as mucocutaneous candidiasis, is more common in patients3. 
Oral candidiasis, a superficial Candida infection, is the most common human fungal infection. Candida albicans is a normal commensal of the mouth; the carriage rate of Candida spp. has been reported to range from 20% to 75% in the general population without any symptoms4. Overgrowth of Candida can lead to local discomfort in patients, such as the altered taste of sensation, burning sensation in the mouth, dysphagia due to poor nutrition, delayed recovery, and prolonged hospital stay. Long-term oral candidiasis can lead to severe invasive candidiasis, resulting in significant morbidity and mortality5. Oral candidiasis can also increase the risk of oral cancer5. Impaired salivary gland function is among the risk factors for oral candidiasis6. The initiation of infection is the adhesion of Candida to epithelial cell walls. This is followed by the proliferation and filamentation of Candida and the formation and maturation of biofilm7. Biofilms are extremely difficult to eradicate and are resistant to conventional antifungal treatment, making clinical treatment a challenge for biofilm-associated oral Candida infection7.
Primary Sj&#246;gren's syndrome (pSS) is an autoimmune disease that is characterized by dry mouth, dry eyes, and other systematic symptoms8. Dry mouth is the most frequent symptom of pSS. Saliva has important physiological antifungal functions. On the one hand, it can dilute and scour the mouth and thereafter remove organisms from the mucosa. On the other hand, saliva contains antimicrobial proteins, such as lactoferrin, sialoperoxidase, lysozyme, histidine-rich polypeptides, and specific anti-candidal antibodies, which can interact with the oral mucosa and prevent the overgrowth of Candida6. The reduced saliva flow may predispose pSS patients to oral candidiasis. An observational cross-sectional study conducted in 61 pSS patients found that 13.1% of pSS patients presented oral signs of candidiasis and colony forming unit (CFU)/mL of Candida albicans was found significantly and negatively correlated with levels of unstimulated whole saliva (UWS) and stimulated whole saliva (SWS)9.
The methods introduced in this manuscript can identify oral Candida infection based on characterized morphological attributes (e.g., the hyphae and yeast cells) in direct smear or after culturing10. The strain identification of the Candida using a Candida diagnostic agar is based on the formation of different colored colonies with varied morphology, which result from the cleavage of chromogenic substrates by species-specific enzymes11. In this manuscript, systematic methods for detecting oral Candida infection are introduced, including traditional oral Candida culture and rapid examination by potassium hydroxide (KOH) smear test and calcofluor white (CFW) staining assay10. Additionally, the antifungal susceptibility test, which can guide clinical oral Candida infection treatment, has been introduced. In actual situations, it is not necessary to perform all of the introduced methods in this manuscript; the oral Candida detection method(s) can be selected according to the purpose. The examination of oral Candida infection and the determination of Candida strains in pSS patients not only benefit the disease treatment but also help assess the characterization of oral candidiasis and the species profiles.

作者聚焦：口腔 念珠菌 诊断推进 pSS 患者的临床治疗方案

原发性干燥综合征患者口腔 念珠菌 感染的检查

In this research, we present a protocol for the examination of oral candida infection in patients with primary Sjogren syndrome, which can be used for timely treatment to avoid infection-related complications. We provide a series of systematic, simple, and feasible methods for detecting oral candida infections, identifying candida strains, and testing the antifungal susceptibility of commonly-used drugs in patients with primary Sjogren's syndrome. Maybe our focus on the role of oral candida infection in the onset and the progression of primary Sjogren's syndrome pathologists in our future studies.

本研究选取12例pSS患者作为代表性患者，筛查口腔 念珠菌 感染（表1）。在这些患者中，一些患者表现出 念珠菌 感染的典型口腔病变，包括口角炎（以口角裂纹、脱屑和充血为特征， 图1A）、牙龈粘膜上鲜红色水肿、黄白色细状假膜（图1B）、舌萎缩和舌背粘膜鲜红色（图1C）。
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Watch this Scientific Journal Video about Author Spotlight: Oral Candida Diagnosis to Advance Clinical Treatment Regimen for pSS Patients at JoVE.com

在这里，我们提出了一种检查原发性干燥综合征患者口腔 念珠菌 感染的方案，该方案可用于及时治疗，此后避免相关并发症。

Primary Sj&#246;gren's syndrome (pSS) is an autoimmune disease characterized by symptoms such as dry mouth, dry eyes, and other systematic symptoms. Due ...

在这项研究中，我们提出了一种用于原发性干燥综合征患者口腔念珠菌感染检查的方案，该方案可用于及时治疗以避免与感染相关的并发症。我们为原发性干燥综合征患者口腔念珠菌感染的检测、念珠菌菌株的鉴定、常用药物的抗真菌敏感性等提供了一系列系统、简便、可行的方法。也许我们在未来的研究中关注口腔念珠菌感染在原发性干燥综合征病理学家的发病和进展中的作用。

examination-oral-candida-infection-primary-sjogrens-syndrome

Research

JoVE Journal

Medicine

Examination of Oral Candida Infection in Primary Sj&#246;gren&#39;s Syndrome Patients

Primary Sj&#246;gren's syndrome (pSS) is an autoimmune disease characterized by symptoms such as dry mouth, dry eyes, and other systematic symptoms. Due to the hyposalivation experienced by pSS patients, oral dysbacteriosis often occurs. A common complication of pSS is the oral Candida infection. In this article, the authors describe systematic methods that can effectively diagnose oral Candida infection and identify the Candida strains using saliva, oral mucosal swabs, or mouthwash from pSS patients. The Sabouraud's Dextrose Agar (SDA), hyphal formation assay, potassium hydroxide (KOH) smear test, and calcofluor white (CFW) staining assay are used for the diagnosis of oral Candida infection. A Candida diagnostic agar is used for the identification of Candida strains. Finally, antifungal susceptibility testing is used to determine appropriate antifungal drug treatment. This standardized method can enhance the diagnosis, treatment, and future research of pSS-related oral Candida infections. Early diagnosis, using this method, can also prevent any complications arising due to delay in receiving appropriate treatment.

Here, we present a protocol for the examination of oral Candida infection in patients with primary Sj&#246;gren's syndrome, which can be used for timely treatment and, thereafter, avoiding related complications.

作者聚焦：口腔 念珠菌 诊断推进 pSS 患者的临床治疗方案

作者聚焦：口腔念珠菌诊断推进 pSS 患者的临床治疗方案