Mössinger J
Institute of Tropical Medicine, University of Tübingen, Germany.
Parasitology. 1991 Aug;103 Pt 1:85-95. doi: 10.1017/s0031182000059321.
Adult Litomosoides carinii, recovered from cotton rats (Sigmodon hispidus) 4-5 months post-infection (p.i.), were cultivated in vitro with emphasis on investigations into the development of intra-uterine embryonic stages. Baseline values for the embryonic status of female worms were established immediately after recovery from the hosts. In such females, on average, 16% of the intra-uterine stages were fully formed microfilariae while the remainder belonged to the early embryonic classes that were characterized. For the evaluation of culture success, apart from survival of worms in vitro, the rate of microfilariae development (mf rate) served as a major parameter. Of the five basic culture media RPMI 1640, F12, L15, NCTC 135, and IMDM, tested without supplementation, RPMI 1640 yielded by far the best results (survival = 14 days; mf rate = 41%), and was therefore chosen as the routine medium. In comparison with 5% CO2 in nitrogen, a gas phase of 5% CO2 in air was superior, although the resulting oxygen tension of 138 mmHg in the medium was not physiological. Addition of 10% newborn or foetal bovine serum to the medium in some cases distinctly influenced results. Effects of different batches of sera varied from 'filaricidal' to 'very promoting'. Co-cultivation of worms and Sigmodon cells, or rhesus monkey LLCMK2 cells, only marginally improved results. Of the serum substitutes Ultroser G, BMS, and Clex, the latter had a moderately promoting effect. The protein supplements bovine serum albumin, transferrin and haemoglobin significantly improved results, and could replace certain batches of serum. Supplementation with the haemin moiety alone was less effective than with haemoglobin. The anti-oxidants glutathione plus ascorbic acid proved beneficial in combination with a serum supplement only. Results from other experiments with multiple supplementation also suggest that various supplements may act only in a synergistic manner. The longest average time that adult L. carinii survived in vitro was 3-4 weeks. The highest mf rate was 78%, which indicated that all normal embryonic stages present in the uteri of a female at the start of culture completed their development to microfilariae, however, oogenesis ceased in vitro. The parameters for embryonic development employed proved to be highly sensitive for the judgment of various culture conditions.
从感染后4 - 5个月的棉鼠（棉鼠属）体内获取的成年卡氏丝虫，在体外进行培养，重点研究子宫内胚胎阶段的发育情况。从宿主体内取出后立即确定雌虫胚胎状态的基线值。在这类雌虫中，平均有16%的子宫内阶段是完全发育的微丝蚴，其余的属于已鉴定的早期胚胎类别。为了评估培养的成功率，除了观察虫体在体外的存活情况外，微丝蚴发育率（mf率）作为一个主要参数。在未添加任何成分的情况下测试的五种基本培养基RPMI 1640、F12、L15、NCTC 135和IMDM中，RPMI 1640产生的结果最好（存活率 = 14天；mf率 = 41%），因此被选为常规培养基。与氮气中5%的二氧化碳相比，空气中5%的二氧化碳气相更优，尽管培养基中产生的138 mmHg的氧张力不符合生理状态。在某些情况下，向培养基中添加10%的新生牛血清或胎牛血清会明显影响结果。不同批次血清的效果从“杀丝虫”到“非常促进”不等。将虫体与棉鼠细胞或恒河猴LLCMK2细胞共同培养，结果仅略有改善。在血清替代品Ultroser G、BMS和Clex中，后者有适度的促进作用。蛋白质补充剂牛血清白蛋白、转铁蛋白和血红蛋白显著改善了结果，并且可以替代某些批次的血清。仅补充血红素部分的效果不如补充血红蛋白。抗氧化剂谷胱甘肽加抗坏血酸仅在与血清补充剂联合使用时被证明是有益的。其他多次补充实验的结果也表明，各种补充剂可能仅以协同方式起作用。成年卡氏丝虫在体外存活的最长平均时间为3 - 4周。最高的mf率为78%，这表明培养开始时雌虫子宫内存在的所有正常胚胎阶段都发育成了微丝蚴，然而，体外卵子发生停止。所采用的胚胎发育参数被证明对判断各种培养条件高度敏感。
