2017 Fiscal Year Annual Research Report
ネパールのヒトー家畜ー野生動物境界領域における結核の伝播動態調査
| Project/Area Number |
17F17408
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| Research Institution | Hokkaido University |
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Principal Investigator |
鈴木 定彦 北海道大学, 人獣共通感染症リサーチセンター, 教授 (90206540)
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| Co-Investigator(Kenkyū-buntansha) |
THAPA JEEWAN 北海道大学, 学内共同利用施設等, 外国人特別研究員
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| Project Period (FY) |
2017-11-10 – 2020-03-31
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| Keywords | M. tuberculosis complex / disease dynamics / Nepal |
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| Outline of Annual Research Achievements |
The main purpose of the research is to better understand the disease dynamics of Mycobacterium tuberculosis complex (M. tuberculosis, M. orygis, M. bovis) and potentially others across the wildlife/human/livestock interface in the landscape of Chitwan National Park (CNP) in Nepal.
A detailed protocol for tuberculosis (TB) suspected sample collection and bacteria isolation from wild animals and livestock was developed. We developed a fecal-TB assay to perform TB surveillance work for monitoring TB. From February 3 to March 4, 2018, a field sample collection trip to Nepal was conducted. Forty TB suspected samples comprising of 10 elephant trunk discharges and 30 fecal samples from different TB suspected wild animals were collected. The samples were processed for culture and DNA extraction. However, TB was not detected from those samples. Beside this, we are performing postmortem on any wild animals especially rhinoceros and elephants for TB surveillance work. During this time, we collected TB suspected samples from the lungs from a dead elephant. The samples are now placed for bacteria culture.
We have discovered methyl green as a suitable non-fluorescent dye to detect positive loop mediated isothermal amplification (LAMP) results. We are testing and validating our LAMP system by using human and animal clinical samples. The LAMP method was used to diagnose TB on 142 clinical human samples and was evaluated with 69 new TB suspected samples. The sensitivity in culture positive samples was 92.8% (13/14) and specificity in culture negative samples was 96.3% (53/55).
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
The research plan is developed and have progressed well. We are further modifying fecal-TB assay by spiking different concentration of M. bovis BCG in feces and processing it for culture and DNA extraction. Our collaborators are performing postmortem of wild animals in CNP landscape and will be collecting more TB suspected samples. The TB suspected elephant samples, that is now in culture will be evaluated. We are preparing a manuscript on our methyl green based TB-LAMP study.
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| Strategy for Future Research Activity |
We will validate fecal-TB assay and transfer this technology to our collaborators for long term surveillance of TB in animals of Nepal. We will also transfer methyl-green based TB-LAMP and PCR technology to our collaborators. We will continue to collect TB suspected samples form dead animals and process them for bacteria isolation. We will be collaborating with our collaborators for zoonotic TB sample collection. We will perform whole genome analysis of previously isolated M. orygis isolates, elephant M. tuberculosis isolates and all the potential MTBC isolates and perform their comprehensive analysis. Finally, we will combine our field observation and genetic findings to understand disease dynamics of TB across the wildlife/human/livestock interface in Nepal.
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