Discrimination of Mycobacterium leprae and Mycobacterium haemophilum in Clinical Isolates and Specimens by Multiplex PCR Assay and Prediction of Drug Susceptibility
Discrimination of Mycobacterium leprae and Mycobacterium haemophilum in Clinical Isolates and Specimens by Multiplex PCR Assay and Prediction of Drug Susceptibility
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多重PCR检测临床分离株和标本中麻风分枝杆菌和嗜血分枝杆菌的鉴别及药敏预测
DOI：
10.1128/jcm.01760-18
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发表时间：
2018
影响因子：
9.4
通讯作者：
Hoshino Yoshihiko
中科院分区：
文献类型：
--
作者：
Kitaoka Naoya;Fukano Hanako;Yoshida Mitsunori;Miyamoto Yuji;Mori Shuichi;Ishii Norihisa;Ato Manabu;Ohara Naoya;Hoshino Yoshihiko
下一代测序技术揭示了比可培养细菌(1)更多活的但不可培养的细菌(VBNC)的存在。分枝杆菌属中的一些物种不能培养，包括麻风分枝杆菌。同时，嗜血杆菌需要氯化血红素或柠檬酸铁铵才能在培养物中生长(2)，尽管在常规使用的培养液中缺乏这些成分使嗜血杆菌有效地进行VBNC(3)。麻风杆菌和嗜血杆菌都经常出现在皮损中。我们和其他人之前报告了嗜血杆菌感染患者的面部病变具有狮子相的外观和病理结果，可能误认为是麻风病(4，5)。麻风杆菌和嗜血杆菌在涂片上均为Ziehl-Neelsen染色阳性，但在培养上均为阴性(即VBNC)，这可能导致麻风杆菌的误诊。尽管针对麻风分枝杆菌36 kDa蛋白的常规聚合酶链式反应检测方法已经发展起来(7)，但这些检测方法对嗜血杆菌临床分离株可能会产生假阳性结果(8)。自2001年麻风杆菌全基因组测定以来，人们已经比较了几种检测麻风杆菌的分子生物学技术。用麻风杆菌特异性重复元件(RLEP)检测麻风杆菌是最敏感的检测方法(9，10)，但仍有40%的假阳性率(10，11)，这表明与麻风杆菌密切相关的不明、未培养或研究不足的分枝杆菌可能引起感染(11)。使用Mauve软件(12)比对麻风杆菌TN全基因组序列(GenBank登录号：NC_002677)和嗜血杆菌ATCC29548T(GenBank登录号：CP011883)参考菌株，我们鉴定了独特的插入/缺失(INDel)区域，并设计了能够根据扩增片段大小进行区分的特异PCR引物(见补充材料中的表S1)。
Next-generation sequencing technology revealed the existence of more viable but nonculturable bacteria (VBNC) than culturable bacteria (1). Several species in the genus Mycobacterium cannot be cultured, including Mycobacterium leprae. Meanwhile, Mycobacterium haemophilum requires hemin or ferric ammonium citrate for growth in culture (2), although the lack of these components in routinely used culture broths renders M. haemophilum effectively VBNC (3). Both M. leprae and M. haemophilum are frequently present in skin lesions. We and others previously reported that facial lesions in patients with M. haemophilum infection had a leonine facies appearance and pathological findings that could mistakenly suggest leprosy (4, 5). Both M. leprae and M. haemophilum yield positive Ziehl-Neelsen staining on smears but are negative in culture (ie, VBNC), which can result in misdiagnosis of M. leprae (6). Despite the development of regular PCR assays that target the 36-kDa protein of M. leprae (7), these assays can yield false-positive results for M. haemophilum clinical isolates (8). Since the determination of the complete M. leprae genome in 2001, several molecular biological techniques for M. leprae detection have been compared. M. leprae detection by M. leprae-specific repetitive element (RLEP) is the most sensitive assay (9, 10) but nonetheless has a 40% false-positive rate (10, 11), which suggests that infections caused by unidentified, unculturable, or understudied mycobacteria that are closely related to M. leprae could occur (11).Here, we developed a novel multiplex PCR assay of clinical isolates and specimens to differentiate M. haemophilum and M. leprae that cannot be isolated in in vitro culture and might be confused clinically. Using the Mauve software (12) to align wholegenome sequences of M. leprae TN (GenBank accession no. NC_002677) and M. haemophilum ATCC 29548T (GenBank accession no. CP011883) reference strains, we identified unique insertion/deletion (indel) regions and designed specific PCR primers that could allow differentiation according to amplicon size (see Table S1 in the supplemental material).
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免疫缺陷患者皮损中分离的嗜血分枝杆菌的细菌学特征
DOI：
10.11150/kansenshogakuzasshi1970.78.389
发表时间：
2004
期刊：
Kansenshogaku zasshi. The Journal of the Japanese Association for Infectious Diseases
影响因子：
--
作者：
H. Saito;K. Toda;I. Matsumoto;K. Matsuo;K. Nakanaga;N. Ishii
通讯作者：
N. Ishii
肾移植患者体内的嗜血分枝杆菌伪装成麻风病
影响因子：
--
作者：
Nathanial K. Copeland;N. Arora;T. Ferguson
通讯作者：
T. Ferguson
影响因子：
7
作者：
Darling, ACE;Mau, B;Perna, NT
通讯作者：
Perna, NT
嗜血分枝杆菌感染，具有类似麻风病的明显面部表现
DOI：
10.1111/1346-8138.12948
发表时间：
2015
期刊：
The Journal of Dermatology
影响因子：
--
作者：
K. Ishii;N. Ishii;K. Nakanaga;K. Nakano;I. Saito;A. Asahina
通讯作者：
A. Asahina
[由新微生物（嗜血分枝杆菌新种）引起的皮肤肉芽肿]。
DOI：
--
发表时间：
1978
期刊：
Harefuah
影响因子：
--
作者：
D. Naveh;T. Yankilewitz;A. Lagziel;D. Sompolinsky
通讯作者：
D. Sompolinsky