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菲律宾奎松市耐多药结核病患者中使用多重聚合酶链反应鉴定非结核分枝杆菌

Identification of Nontuberculous Mycobacteria in Patients with Multidrug-resistant Tuberculosis in Quezon City, Philippines, Using Multiplex PCR.

作者信息

Cabanatan Michelle M, Bungay Alice Alma C, Villanueva Sharon Yvette Angelina M, Tobias-Altura Marohren C, Defensor Dario D, Lota Maria Margarita M

机构信息

Department of Medical Microbiology, College of Public Health, University of the Philippines Manila, Manila, Philippines.

Department of Pathology and Laboratory Medicine, Lung Center of the Philippines, Quezon City, Philippines.

出版信息

Acta Med Philipp. 2025 Mar 31;59(4):103-112. doi: 10.47895/amp.v59i4.9049. eCollection 2025.

Abstract

BACKGROUND AND OBJECTIVE

Nontuberculous mycobacteria (NTM) lung disease appears like tuberculosis infection but is resistant to primary anti-tuberculosis drugs. Hence, patients whose sputum sample tests positive for acid-fast bacilli (AFB) and bacterial culture for several times should be assessed for colonization or infection with NTM in a damaged lung secondary to TB. In such cases, though drug-resistant TB may be adequately treated, treatment may need to be directed towards the NTM as well. In NTM therapy, the duration and choice of treatment agent is based upon the specific organism and disease extent. This study used one-step multiplex PCR (mPCR) assay for rapid differentiation of solid cultures in Ogawa medium as (MTB) and/or NTM.

METHODS

A total of 80 stocked isolates obtained from the Lung Center of the Philippines from January to December 2018 were screened for NTM in terms of growth in Ogawa medium, acid fastness, and MPT64 TB antigen test result. These were from sputum specimens of multidrug-resistant tuberculosis (MDR-TB) patients. DNA was extracted from cultures (n=55) grown in Ogawa medium and one-step mPCR was performed to identify NTM to the species level.

RESULTS

Out of 80 samples screened, a total of 55 isolates were identified as NTM. One-step mPCR identified 12.73% (7/55) as , 34.55% (19/55) as , 1.82% (1/55) as , and 50.91% (28/55) were identified only up to genus spp. Neither complex nor was identified among the samples tested.

CONCLUSION

One-step mPCR was able to identify isolates as MTB or NTM coinciding with the initial screening using MPT64 TB antigen test. Multiplex PCR has given a more specific identification to the species level. The use of mPCR in identifying MTB and clinically significant NTM's is suitable for the adequate treatment of mycobacterial infection.

摘要

背景与目的

非结核分枝杆菌(NTM)肺病表现类似肺结核感染,但对一线抗结核药物耐药。因此,痰涂片抗酸杆菌(AFB)检测多次阳性且细菌培养阳性的患者,应评估是否存在继发于结核病的肺部受损后NTM定植或感染。在这种情况下,虽然耐多药结核病可能得到充分治疗,但可能也需要针对NTM进行治疗。在NTM治疗中,治疗疗程和治疗药物的选择基于特定病原体和疾病范围。本研究采用一步多重PCR(mPCR)检测法对小川培养基中的固体培养物进行快速鉴别,区分结核分枝杆菌(MTB)和/或NTM。

方法

对2018年1月至12月从菲律宾肺部中心获得的80株库存菌株,根据其在小川培养基中的生长情况、抗酸性和MPT64结核抗原检测结果进行NTM筛查。这些菌株来自耐多药结核病(MDR-TB)患者的痰标本。从在小川培养基中生长的培养物(n = 55)中提取DNA,并进行一步mPCR以将NTM鉴定到种水平。

结果

在80份筛查样本中,共鉴定出55株NTM。一步mPCR鉴定出12.73%(7/55)为堪萨斯分枝杆菌,34.55%(19/55)为胞内分枝杆菌,1.82%(1/55)为蟾蜍分枝杆菌,50.91%(28/55)仅鉴定到分枝杆菌属。在所检测的样本中未鉴定出鸟胞内分枝杆菌复合群和脓肿分枝杆菌。

结论

一步mPCR能够将菌株鉴定为MTB或NTM,与使用MPT64结核抗原检测的初步筛查结果一致。多重PCR能更特异性地鉴定到种水平。使用mPCR鉴定MTB和具有临床意义的NTM适用于分枝杆菌感染的充分治疗。

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