Section for Arctic Veterinary Medicine, Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, Tromsø, Norway.
PLoS One. 2013 Jul 25;8(7):e70186. doi: 10.1371/journal.pone.0070186. Print 2013.
A high prevalence of Brucellapinnipedialis serology and bacteriology positive animals has been found in the Northeast Atlantic stock of hooded seal (Cystophoracristata); however no associated gross pathological changes have been identified. Marine mammal brucellae have previously displayed different infection patterns in human and murine macrophages. To investigate if marine mammal Brucella spp. are able to invade and multiply in cells originating from a presumed host species, we infected alveolar macrophages from hooded seal with a B. pinnipedialis hooded seal isolate. Hooded seal alveolar macrophages were also challenged with B. pinnipedialis reference strain (NCTC 12890) from harbor seal (Phocavitulina), B. ceti reference strain (NCTC 12891) from harbor porpoise (Phocoenaphocoena) and a B. ceti Atlantic white-sided dolphin (Lagenorhynchusacutus) isolate (M83/07/1), to evaluate possible species-specific differences. Brucella suis 1330 was included as a positive control. Alveolar macrophages were obtained by post mortem bronchoalveolar lavage of euthanized hooded seals. Phenotyping of cells in the lavage fluid was executed by flow cytometry using the surface markers CD14 and CD18. Cultured lavage cells were identified as alveolar macrophages based on morphology, expression of surface markers and phagocytic ability. Alveolar macrophages were challenged with Brucella spp. in a gentamicin protection assay. Following infection, cell lysates from different time points were plated and evaluated quantitatively for colony forming units. Intracellular presence of B. pinnipedialis hooded seal isolate was verified by immunocytochemistry. Our results show that the marine mammal brucellae were able to enter hooded seal alveolar macrophages; however, they did not multiply intracellularly and were eliminated within 48 hours, to the contrary of B. suis that showed the classical pattern of a pathogenic strain. In conclusion, none of the four marine mammal strains tested were able to establish a persistent infection in primary alveolar macrophages from hooded seal.
在北大西洋冠海豹（Cystophoracristata）种群中，已发现大量的 Brucellapinnipedialis 血清学和细菌学阳性动物；然而，尚未确定与之相关的明显的病理变化。海洋哺乳动物布鲁氏菌在人类和鼠类巨噬细胞中的感染模式先前已有报道。为了研究海洋哺乳动物布鲁氏菌是否能够入侵并在来源于假定宿主物种的细胞中繁殖，我们用一株冠海豹分离的 B. pinnipedialis 感染了冠海豹的肺泡巨噬细胞。我们还用来自港海豹（Phocavitulina）的 B. pinnipedialis 参考株（NCTC 12890）、来自港湾海豚（Phocoenaphocoena）的 B. ceti 参考株（NCTC 12891）和一株来自大西洋白海豚（Lagenorhynchusacutus）的 B. ceti 分离株（M83/07/1）对冠海豹肺泡巨噬细胞进行了挑战，以评估可能存在的种间差异。Brucella suis 1330 被用作阳性对照。肺泡巨噬细胞是通过安乐死的冠海豹死后支气管肺泡灌洗获得的。灌洗液中的细胞表型通过使用表面标记物 CD14 和 CD18 的流式细胞术进行分析。根据形态、表面标记物的表达和吞噬能力，培养的灌洗细胞被鉴定为肺泡巨噬细胞。在庆大霉素保护试验中，用布鲁氏菌属对肺泡巨噬细胞进行了攻击。感染后，从不同时间点的细胞裂解物中进行平板培养，并进行定量的菌落形成单位评估。通过免疫细胞化学验证了冠海豹分离株 B. pinnipedialis 的细胞内存在。我们的结果表明，海洋哺乳动物布鲁氏菌能够进入冠海豹的肺泡巨噬细胞；然而，它们并没有在细胞内繁殖，而是在 48 小时内被清除，这与 B. suis 形成了鲜明的对比，后者显示出典型的致病菌模式。总之，在所测试的四种海洋哺乳动物菌株中，没有一种能够在冠海豹的原代肺泡巨噬细胞中建立持续感染。
