THE INFECTING AGENT IN INFLUENZA: AN EXPERIMENTAL RESEARCH

BY Prof. T. Yamanouchi, Dr. K. Sakakami, Dr. S. Iwashima
Lancet: Volume 193, Issue 4997p971 、June 07, 1919

DURING the past winter influenza caused the loss of many lives 1n Japan. According to official statistics, up to the end of January, 1919, there were nearly 30,000,000 (of which 170,000 proved fatal. Inasmuch as 62 of our friends, doctors and nurses, offered themselves as subjects for experiment, we have been able to solve some important questions relating to this infectious disease. The experiments and results now recorded were made between Dec, 18, 1918, and the end of March, 1919.

Experiments.

• An emulsion of the sputa from 43 influenza patients was made in Ringer's solution. This emulsion was injected into the nose and throat of 12 healthy persons.
• A filtrate (by Berkefeld filter) of the same emulsion was injected into the nose and throat of 12 other healthy persons. The results of these experiments are very significant; among the subjects treated were six persons who had already bad influenza, and all six showed to no symptoms of illness. But all of the other 18 subjects, both those who had received the emulsion and those who had received only the filtrate were attacked by the disease, after so incubation of two or three days. Their fever was sometimes slight (37.5℃), sometimes very severe (39℃ or more). The subsequent symptoms were headache, sore-throat, lumbago, cough, and the like.
• A filtrate of blood of influenza patients was injected into the nose and throat of 6 more healthy persons. The results were precisely the same as in the previous experiments.
• We inoculated subcutaneously 4 healthy persons with the filtrate of the sputa emulsion and 4 others with a filtrate of the blood of influenza patients. They all, with the exception of one who had previously bad influenza, developed the disease after two- or three-days’ incubation.
• A pure culture of Pfeiffer’s bacillus and & mixed preparation of the pure Pfeiffer, along with pneumococci, streptococci, staphylococci, diplococci, and many other like microbes common in the sputa of influenza patients, were injected into the throat and nose- of 14 healthy people who had not bad influenza. There were no symptoms of illness following these injections.

Summary.

• The germ of influenza cannot be removed by filtering (filterable virus).
• The germs can infect through the mucous membrane and also by inoculation.
• The germs can be found in the sputum and the blood of influenza patients.
• The known bacilli, such as Pfeiffer's bacillus, pneumococci, and some diplococci are not the cause of influenza.
• We observed experimentally that all people who have previously had influenza or received the sputa emulsion or its filtrate are immune to the disease.

この冬、日本ではインフルエンザが多くの人命を奪った。公式統計によれば、1919年1月末までの罹患者数は約30,000,000人（うち死亡者数は17万人）であった。62名の友人、医師、看護師が本実験の被験者となってくれたので、いくつかの重要な問題を解決することができた。なお、この実験は1918年12月18日から1919年3月末までの間に行われたものである。

実験

• インフルエンザ患者43名の喀痰をリンゲル液で乳剤化した。この乳剤を健常者12名の鼻と喉に注入した。
• 同じ乳剤の濾液（バークフェルトフィルターで濾過した（筆者注：後述します））を、他の12人の健康な人の鼻と喉に注入した。
  この1.2.の実験結果は非常に重要である。その中にはすでにインフルエンザに罹患していた6人が含まれていたが、その6人全員、濾液を注入しても、インフルエンザの症状は見られなかった。しかし「乳剤を投与された者」も「濾液のみを投与された者」も、他の18名（訳注：つまりインフルエンザに罹患していない被験者18名）はすべて2～3日の潜伏期間を経て発病した。発熱は微熱（37.5℃）の時もあれば、非常に激しい（39℃以上）の時もあった。その後の症状は頭痛、咽頭痛、腰痛、咳などであった。
• インフルエンザ患者の血液の濾液をさらに健常者6名の鼻と喉に注射した。結果は前の実験と全く同じであった。
• 健常者4名に喀痰乳剤の濾液を皮下接種し、別 の4名にインフルエンザ患者の血液の濾液を皮下接種した。彼らはすべて、以前にインフルエンザに罹患していた1人を除いて、2、3日の潜伏後にインフルエンザを発症した。
• 肺炎球菌、連鎖球菌、ブドウ球菌、ディプロコッカス、およびインフルエンザ菌に罹患した患者の喀痰に共通する他の多くの類似の微生物とともに、純粋なファイファー菌の培養物と＆混合調製物を、14人の健康な人々の喉と鼻に注射したが、これらを注射してもインフルエンザは発症しなかった。

まとめ

• インフルエンザの病原はろ過しても除去できない（濾過性病原体だろう）。
  （筆者注：1884年にフランスの科学者シャルル・シャンベランは細菌を除去する磁器製の濾過器（シャンベラン濾過器（Chamberland filter）を考案。この濾過器は細菌を通さないので、特に浄水作成や実験での無菌操作に使われていました。この濾過器を細菌よりも小さい病原体（ビールス）は通過してしまうことが知られるようになり、ビールス学の黎明期にこの濾過器が活躍したのです。この論文の著者もシャンベラン濾過器を使っていたと思います。）
• この病原は粘膜を介して感染し、また接種によっても感染する。
• インフルエンザ患者の喀痰や血液中にもインフルエンザの病原体が存在する。
• ファイファー桿菌、肺炎球菌や、双球菌などの既知の菌は、インフルエンザの原因菌ではない。
• 以前にインフルエンザにかかったことがある人や、スプータ乳剤やその濾液をもらった人は全員免疫があることを実験的にできた。

5.3 Influenza Viruses

The history of the discovery of influenza virus is more complex than that of most other viruses: intermixed with the great pandemic of 1918, confounded by many contradictory claims, involving more than the usual number of players, and forced to overcome the insistence at the turn of the 20th century by the prominent bacteriologist, Richard Friedrich Johannes Pfeiffer (1858 – 1945), that the disease is caused by a bacterium, “Pfeiffer’s bacillus” (then Bacillus influenza, now Haemophilus influenzae)(Taubenberger et al., 2007). By 1922, the notion that influenza was caused by an ultrafilterable virus had gained much ground. At this time the great microbiologist Hans Zinsser (1878 – 1940) wrote in his Textbook of Bacteriology, “All evidence must be considered in connection with recent experiments upon the possibility of [influenza] causation by a filterable virus … ”(Zinsser, 1922). He cited several studies from 1918 – 19, including those of:

• (a) Charles Nicolle (1866 – 1936) and Charles Lebailly, of the Institute Pasteur Tunis, who ultrafiltered human blood and nasal secretions from uncomplicated influenza patients and instilled the filtrates into conjunctival sacs and nasal cavities of monkeys (sic “Macaccus Sinicus”) and human volunteers, obtaining flu-like clinical signs in most (Nicolle & Lebailly, 1918).
• (b) Ren e Dujarric de la Riviere (1885 – 1969), who ultrafiltered blood from four influenza patients and injected the combined filtrates into himself, clearly obtaining a flu-like disease (Dujarric de la Riviere, 1918).
• (c) Erich Leschke, who produced flu-like clinical signs in monkeys with ultrafiltered human nasal secretions (Leschke, 1919).
• (d) H. Selter, who ultrafiltered nasal secretions from cases and sprayed the filtrate into his own throat and that of an assistant, producing flu-like disease (Selter,1918).
• (e) T. Yamanouchi, K. Sakami, and S. Iwashima of the Government Institute for Infectious Diseases, Tokyo Imperial University (now the Institute of Medical Science, University of Tokyo), who carried out the most extensive experiments, processing nasopharyngeal secretions from 43 patients, ultrafiltering half, and instilling each into the nose of 24 volunteers (6 of those receiving the ultrafiltered material had just recovered from influenza). All of the 24, except those who had had influenza recently, came down with a flu-like disease after an incubation period of 2 and 3 days. They did other experiments as well, confirming their initial impressive findings (Yamanouchi et al., 1919).

• (e)東京帝国大学感染症研究所（現東京大学医科学研究所）の山内保、坂上弘蔵、岩島十三の3人は、最も大規模な実験を行った。43人の患者の鼻咽頭分泌液を処理し、半分を限外濾過し、それぞれを24人のボランティアの鼻に注入した（限外濾過を受けた人のうち6人はインフルエンザから回復したばかりであった）。 最近インフルエンザにかかった6人を除く24人全員が、2日と3日の潜伏期間の後、インフルエンザのような症状を呈した。 彼らは他の実験も行い、インフルエンザに関する世界で初めて重要な発見を確認した（山内ら、1919年）。