- 學位論文
台灣豬鏈球菌血清型別、毒力相關基因譜型及藥物敏感性之綜合分析
The comprehensive analysis between serotype, virulence-associated genes profile, and antimicrobial susceptibility of Streptococcus suis in Taiwan
摘要
豬鏈球菌（Streptococcus suis, S. suis）為重要人畜共通傳染病病原之一，在豬隻可造成腦膜炎、胸膜炎、關節炎或肺炎病變，導致豬隻生長遲滯及死亡。S. suis共區分成29種血清型別（serotype），特定型別具有較強致病能力，例如第2血清型別。此外，其致病機轉牽涉多種毒力相關因子，例如：溶血素（suilysin）、胞壁酸酶釋放蛋白質（muramidase-released protein）及細胞外蛋白質因子（extracellular protein factor）等，可用以評估菌株侵害能力。因此，在S. suis感染症之臨床病例診斷流程中，檢測血清型別及毒力相關基因（virulence-associated genes, VAGs）皆為重要指標。本疾病尚無商業化疫苗可供有效防護，臨床上多使用抗生素進行預防性投藥或病豬治療，故應例行檢測菌株藥物敏感性並追蹤抗藥性變化，以提供謹慎用藥的選擇依據。為了解台灣病豬分離之S. suis菌株血清型別、毒力相關基因譜型與藥物敏感性，本研究自2015至2019年針對S. suis相關病灶之檢體進行菌株分離，以分子生物學方法檢測血清型別和毒力相關基因（sly、mrp、epf），並將相同牧場來源的菌株以17種抗生素進行最小抑制濃度試驗（minimum inhibitory concentration, MIC）。本研究共收集406株S. suis樣本，檢出的22種血清型別中，主要盛行型別為第3型（20.0%）、第8型（11.3%）、第2型（10.3%）及第4型（10.1%），與亞洲主流型別類似，其中第8型比例較其他國家研究多，為台灣菌株的特點之一；sly－/mrp－/epf－（40.9%）、sly－/mrp＋/epf－（29.6%）及sly＋/mrp＋/epf－（18.7%）譜型較盛行。將血清型別對應毒力相關基因譜型，serotype 3 - sly－/mrp＋/epf－（17.4%）和serotype 8 - sly－/mrp－/epf－（10.6%）的組合模式最具代表性（p < 0.01、p < 0.05）。比較菌株於不同感染部位之分離率，血清型別第1型及epf基因與全身性感染病灶較有關聯，血清型別第2、3、4、8型及sly、mrp基因則與呼吸系統病灶較有關聯。46株菌株完成MIC試驗，對amoxicillin、cefotaxime、ceftiofur及florfenicol較具敏感性，而oxytetracycline、doxycycline、tilmicosin、tylosin及lincospectin則不適用於治療感染S. suis之病豬，與各國研究結論相符。本篇為全球首先使用常態化抗藥性解釋模型（normalized resistance interpretation, NRI）分析S. suis菌株的研究，部份菌株對β-內醯胺類藥物具敏感性但為非野型（non-wild type）分類，可能已具有潛在抗藥性能力，須特別留意未來田間盛行率變化。總結而言，本研究提供台灣S. suis菌株血清型別、毒力相關基因譜型及藥物敏感性等豐富資料，並進行多項綜合分析，有助於未來釐清S. suis致病機轉及相關疫苗開發等研究，並提供臨床治療藥物選擇之依據。
並列摘要
Streptococcus suis (S. suis) is an important zoonotic pathogen in porcine industry over worldwide, causing meningitis, septicemia, arthritis, pneumonia, and even acute death. It has been classified into 29 serotypes based on the differentiation of capsule polysaccharide structures, and some specific serotypes have higher pathogenicity, for instance, serotype 2. Multiple virulence-associated factors are involved in S. suis pathogenesis mechanism such as suilysin (SLY), muramidase-released protein (MRP), and extracellular protein factor (EF). These factors could be key indicators to pathogenicity of the isolates. Thus, serotyping and virulence-associated genes (VAGs) screening are meaningful in diagnosis routine for clinical cases. Without available and efficient commercial vaccine, antibiotic treatment to prevent or control the outbreak of S. suis is still the primary choice in field. Therefore, it is necessary to perform the antimicrobial susceptibility tests and track the resistance dynamic for prudent use references. To investigate serotypes, VAGs profiles, and antimicrobial susceptibility of S. suis from diseased pigs in Taiwan, totally 406 clinical isolates were obtained from S. suis-associated lesions from 2015 to 2019. Serotypes and VAGs (sly, mrp, epf) were both detected by multiplex PCR assays. The isolates from the same farms were subsequently tested for antimicrobial susceptibility against 17 antibiotics by determining minimum inhibitory concentration (MIC) assay. Amount 22 detected serotypes, the predominant serotypes in Taiwan were serotype 3 (20.0%), 8 (11.3%), 2 (10.3%), and 4 (10.1%). This result is similar to those of other Asian countries. In addition, the prevalence of serotype 8 is higher that is one of features in serotype distribution in Taiwan. Main VAGs profiles showed as: sly－/mrp－/epf－ (40.9%), sly－/mrp＋/epf－ (29.6%), and sly＋/mrp＋/epf－ (18.7%). Combined together, serotype 3 - sly－/mrp＋/epf－ (17.4%, p < 0.01) and serotype 8 - sly－/mrp－/epf－ (10.6%, p < 0.05) were dominant patterns as compared to others. Comparing the isolation rates of strains in different lesions, serotype 1 and epf gene were more related to systemic lesions; by contrast, serotype 2, 3, 4, 8 and sly, mrp genes were related to respiratory lesions. Forty-six isolates were performed MIC tests. The results show that isolates are susceptible for amoxicillin, cefotaxime, ceftiofur, and florfenicol; however, resistant to oxytetracycline, doxycycline, tilmicosin, tylosin, and lincospectin, which are not recommended to cure ill pigs affected by S. suis. The tendency is also similar to the previous studies. The present study is the first one to apply normalized resistance interpretation (NRI) to analyze MIC distribution of clinical S. suis isolates over the world. It is necessary to follow up the prevalence of those non-wild type isolates with susceptibility to β-lactam antibiotics, which may have equipped potential resistant abilities. In general, this study provides multiple information on serotype, VAGs profile, and also antimicrobial susceptibility of S. suis isolates in Taiwan. It is valuable for further pathogenesis investigation, vaccine developing, and medication reference establishment.
參考文獻
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