Differences in pathogenicity of three animal isolates of Mycobacterium species in a mouse model

Animal mycobacterioses are among the most important zoonoses worldwide. These are generally caused by either Mycobacterium tuberculosis (MTB), M. bovis (MBO) or M. avium (MAV). To test the hypothesis that different species of pathogenic mycobacteria isolated from varied anatomic locations or animal species differ in virulence and pathogenicity, we performed experiments with three mycobacteria strains (NTSE-3(MTB), NTSE-4(MBO) and NTSE-5 (MAV)) obtained from animal species. Spoligotyping analysis was used to confirm both MTB and MBO strains while the MAV strain was confirmed by 16s rDNA sequencing. BALB/c mice were intranasally infected with the three strains at low and high CFU doses to evaluate variations in pathogenicity. Clinical and pathological parameters were assessed. Infected mice were euthanized at 80 days post-inoculation (dpi). Measures of lung and body weights indicated that the MBO infected group had higher mortality, more weight loss, higher bacterial burden and more severe lesions in lungs than the other two groups. Cytokine profiles showed higher levels of TNF-α for MBO versus MTB, while MAV had the highest amounts of IFN-β in vitro and in vivo. In vitro levels of other cytokines such as IL-1β, IL-10, IL-12, IL-17, and IFN-β showed that Th1 cells had the strongest response in MBO infected mice and that Th2 cells were inhibited. We found that the level of virulence among the three isolates decreased in the following order MBO>MTB>MAV.

动物分枝杆菌病是全球范围内最为重要的人畜共患病之一。此类疾病通常由结核分枝杆菌（Mycobacterium tuberculosis, MTB）、牛分枝杆菌（Mycobacterium bovis, MBO）或鸟分枝杆菌（Mycobacterium avium, MAV）引发。为验证「从不同解剖部位或动物物种中分离的致病性分枝杆菌菌种，其毒力与致病性存在差异」这一假说，我们采用3株从动物体内分离得到的分枝杆菌菌株开展实验：NTSE-3（MTB）、NTSE-4（MBO）及NTSE-5（MAV）。采用间隔区寡核苷酸分型（Spoligotyping）技术对MTB与MBO菌株进行鉴定，MAV菌株则通过16S rDNA测序完成验证。以低、高两种菌落形成单位（Colony-Forming Units, CFU）剂量，通过鼻腔接种方式将3株菌株感染BALB/c小鼠，以评估其致病性差异。对感染小鼠的临床及病理参数进行检测，并于接种后80天（days post-inoculation, dpi）对其实施安乐死。肺脏重量与体重检测结果显示，相较于另外两组，MBO感染组小鼠的死亡率更高、体重丢失量更大、细菌载量更高，且肺部病变更为严重。细胞因子谱分析结果显示，MBO感染组的肿瘤坏死因子-α（Tumor Necrosis Factor-α, TNF-α）水平高于MTB感染组；而MAV感染组在体外与体内环境中均呈现最高的干扰素-β（Interferon-β, IFN-β）表达量。其余细胞因子（如白细胞介素-1β、白细胞介素-10、白细胞介素-12、白细胞介素-17及干扰素-β）的体外检测结果表明，MBO感染小鼠体内的辅助性T细胞1（Th1）应答最强，辅助性T细胞2（Th2）的活性则受到抑制。本研究发现，3株分离菌株的毒力水平由高到低依次为：MBO＞MTB＞MAV。