肉用鶏の腹水症に関する研究 : 形態学的観察による発生機序と発病要因の検討 Studies on ascites in broiler chickens : discussion of pathogenesis and induced factor by morphological observation
肉用鶏の腹水症に関する研究 : 形態学的観察による発生機序と発病要因の検討
Studies on ascites in broiler chickens : discussion of pathogenesis and induced factor by morphological observation
わが国における肉用鶏の腹水症の発生は1979年頃より多発の傾向を示している。現在までに報告された症例を整理すると，概略30日齢以降冬期の飼育群に発生し，その育成率低下は5%前後である。 原因については換気不良，フラゾリドンおよび他の中毒，寒冷感作を疑うものなどがあるが，さらにその発生機序に関しては不明な点が多い。 著者は野外発生例の疫学的調査から，飼育環境が換気不良状態であったことを指摘し，それをStressorとして2回の飼育実験を行ったところ野外例と同質の病変をみた。 野外例，飼育実験例を主として形態学的に整理し，本症の発生機序と発病要因について検討を試みた。I.検査材料および方法(野外例) 腹水症の発生のあったK，W，N農家の飼育環境調査および臨床観察を行い，53-66日齢の37羽を試験に供した。病理解剖・組織学的検査は全例，細菌学的検査は11例とK農家での給与飼料，寄生中検査は6例について実施した。(飼育実験1) 強度の換気不良環境(酸素濃度10～13%)で実験区3羽，対照区15羽をそれぞれ27日齢，34日齢まで飼育した。飼料はK農家事故発生時のものを用いた。全例の病理解剖・組織学的検査を実施した。(飼育実験2) 中～軽度の換気不良環境(酸素濃度15～20%)で飼育した。供試鶏は実験区，対照区それぞれ20，20羽で70日齢まで飼育した。病理学的検査は全例，細菌学的検査は実験区5例，対照区2例，ヘマトクリット値の測定は65日齢で両区それぞれ7例について実施した。II.成績(野外例)1. 発生状況…鶏舎の構造はK，Nで開放型，Wでドーム型で，給温方法はそれぞれ異なっていた。育成率は90.2～94.6%であった。K，Wでは各週1%，Nでは1～3週齢に集中して死亡，淘汰がみられ，腹水症はいずれも40日齢以降に確認している。飼育は冬期間で厳寒のため，いずれも1～2週齢時までは鶏舎を完全に密閉し，換気は行わなかった。以降日中のみわずかに行い，夜間は閉めきった。Nでは大型ストーブを用いていたが，吸気は舎外から取り入れているものの排気筒は鶏舎に開放されていた。 3農場5鶏舎についてガス測定を行ったところ，酸素濃度は1lotが有意に低く(16～20%)，炭酸ガスは全lotが0.1～0.15%，アンモニアガスは3/5lotで10ppmを示した。2. 臨床症状…腹水を伴った病鶏は腹囲膨満，翼下垂し，うずくまるものが多かった。歩かせるとペンギン様あるいは首を前に伸ばしアヒル様の歩行を示した。いずれの群でも軽い開口呼吸を伴っていた。3. 剖検所見…高率に腹水が認められ，50mlから300mlに及んでいた。腹水は麦桿色あるいはブドウ酒様の色調を帯び，フィブリン凝塊を容れていた。腹水の貯留部位は肝-腹膜腔であった。 心では心嚢液増量，右心室壁は菲薄，拡張，弛緩し，横径を増していた。腎静脈，肝静脈，後大静脈の著しいうっ滞が認められた。 肝では腫大と表面の不整が高率にみられ，辺縁は鈍となり，血量に富み暗赤色から赤橙色で脆いもの，褪色し硬度を増しているものなど各種段階がみられた。さらに黄白色斑や肝包膜下に漿液の貯留，表面にフィブリン被膜を載せているものもあった。 肺はうっ血水腫，腎は腫大し褪色しているもの，血量に富むものなどがみられた。 消化器では筋胃角質層の硬化，糜爛，潰瘍形成，腺胃の出血，十二脂腸のうっ血を認めた。 脾，胸腺，ファブリキュウス嚢の萎縮は高率であった。4. 組織所見…心;うっ血高度。心筋線維は萎縮，断裂し横紋消失，細胞質顆粒状，空胞形成，さらに凝固～融解壊死がみられた。一部の心筋では肥大がみられた。間質では水腫と間質線維の増殖，細胞浸潤がみられた。1例では中等度大の動脈の中膜が粗開し，肉芽の増生を伴っていた。 肝;肝静脈のうっ血高度で中心静脈，類洞は拡張し，小葉中心性に脂肪沈着，変性・壊死，類洞内皮の肥厚による硬変，変性部とグリッソン鞘の細胞浸潤，偽胆管増生，髄外造血像，包膜の肥厚が認められた。 肺;高度のうっ血と水腫を認めた。 腎;うっ血，尿細管上皮の変性，糸球体内皮核の増数による腫大がみられた。 脾，胸腺，ファブリキュウス嚢;著明なリンパ球の低形成が認められた。 骨髄;低形成を示した。 副腎;皮質に強い変化が認められた.多くの細胞は肥大し，核は大きく明るく，細胞質は淡染，あるものは細胞質内に空洞を容れていた。とくに表層に近い細胞群では肥大著しく，脱落しているものもみられた。一方細胞は容積を減じてきているものや，細胞境界不明，核はぼやけて構造不明となっている変性細胞集団も認め出血を伴っていた。変性細胞集団は表層に多くみられた。Sudan IIIによる染色では空洞は脂肪滴の沈着であり，変性細胞はリポイド顆粒を失っていた。5. 細菌検査所見…主として肺からブドウ球菌，シトロバクター，大腸菌が分離された。しかし共通性のないこと，菌数が少ないことなどから本症に有意なものとは考えられなかった。 飼料中の細菌培養所見は陰性であった。6. 寄生虫検査…陰性であった。(実験1) 実験区では強い呼吸困難がみられ，著しい発育の遅延が認められた。また27日齢に1例が死亡した。 剖検では心，肝，肺，腎はうっ血を主徴とし，それに伴い肝の表面不整，黄白色斑を伴っていた。 腺胃の潰瘍はそれぞれ1/3例に認めた。 脾，ファブリキュウス嚢は萎縮していた。しかしながら腹水の増量は認められなかった。 組織所見も野外例と同質の変化を示し，心筋の変性壊死，間質腺維の増生，動・静脈の変性，肝の小葉中心性の変性・壊死，脾，ファブリキュゥス嚢のリンパ組織の低形成，副腎の退行性変化が目立った。飼料による影響は認められなかった。(実験2) 実験区に呼吸困難がみられた。増体重は対照区に比し常に低く推移し，3週齢以降で有意差を認めた。52日齢で1例，58日齢で2例が死亡した。58日齢で死亡した1例に腹水の増量をみた。剖検では肝の脆弱化が目立ち，死亡2例では明瞭な変性壊死巣を伴っていた。組織所見は野外例，実験1と同質であった。なお骨髄は実験，対照区とも高率(94.1，88.9%)に低形成を示した。 細菌検査では実験区の肺，気管からブドウ球菌，大腸菌が分離されたが共通性，菌量から有意とは考えられなかった。70日齢時のヘマトクリット値は実験区33.6%，対照区30.9%で有意差を認めた。III.まとめおよび考察 野外で腹水症のあった3群，37羽の病鶏および換気不良環境による2回の飼育実験から病変を整理するとつぎのとおり要約できた。 1)副腎皮質細胞の肥大から変性壊死あるいは萎縮，2)心臓の障害およびそれに基づく循環障害からおこる肺，肝，腎の変化，3)消化器の出血，潰瘍の形成，4)リンパ組織系臓器の萎縮。 副腎皮質の障害は高度で，その機能亢進から疲憊までの過程がうかがえた。病変のおこり，分布，実験例で早期に出現してくること，赤血球増多症のみられることおよびその機能的な役割を鑑みるに，副腎皮質は本症での主座を占めると考えられた。 野外例の環境調査，飼育実験，病変の分布およびその起こりから腹水症の発生機序について以下のとおり考察した。 換気不良環境下では酸素欠乏により呼吸，脈拍数の増加に伴って肺動脈圧の上昇が起こり，心臓には機械的な負担が増大する。一部の心筋の肥大によって応答するが一定の限度があり疲労におちいる。このような心臓の機械的疲労に加え，酸素欠乏それ自身がStressorであることから，副腎皮質ホルモンの過剰分泌から失調までの間に引き起こされる心筋の変性・壊死，毛細血管の透過性充進による体液の平衡失調，血液の濃縮，赤血球増多症などにより心不全を招いたと考えられた。全身の循環障害のなかで，特に肝ではうっ血性肝硬変が生じており，腹水はその貯留部位から肝由来である。 腺胃，筋胃でみられた潰瘍と糜爛形成およびリンパ組織系臓器の低形成は副腎皮質ホルモンの過剰分泌期に生じた変化である。 以上のように副腎皮質は発生機序のうえで中枢的役割を演じており，本症は汎適応症候群として位置づけることができる。 本研究の飼育実験では，野外例の調査から指摘し得た換気不良環境をStressorとして与え野外例と同質の病変を得た。しかしながら副腎皮質は各種Stressorに非特異的に反応することを鑑みるに，野外でおこる本症の発病要因としては換気不良単独ではなく，寒冷，暑熱，過密，有害ガス，さらに肉用鶏のもつ特性すなわち急激な増体，過食，肥満などの各種Stressorが複雑に交錯し，それらが総和となって作用しているものと考えられた。
INTRODUCTION Ascites in broiler chickens has been getting become big problem from toward 1979 in Japan. From the many case reports, it has been ocurred from then on around 30 day old chickens during cold seasons reared. Incidence of the disease was estimated about 5 per cent. Insufficient ventilation, frazolidone poisening and coldness were suspected as the cause of disease. But, its pathogenesis was not clarified. Author pointed out that birds suffered from ascites were reared in environment of insufficient ventilation. Experiments given stress factor as insufficient ventilation were revealed as same changes as field cases. From results of environmental survey and morphological changes of field and experiment cases, in this study, pathogenesis and induced factor of ascites were discussed.MATERIALS AND METHODSField cases Clinical observations, environmental surveys and laboratry examinations were performed on flocks of K, W, N farm which had outbreaks of ascites. Gas concentration of 5 houses of 3 farms were measured by gas-detection-tube (Komyo Rikagaku Kogyo Co.). 37 chickens from the age of 53 to 66 day old were collected from 3 flocks of 3 farms for further examinations. 5 were dead and 32 were killed cases. All cases were examined macro and microscopicaly. For histological studies, specimens were fixed in 10% formalin, embedded in paraffin, and stained with hematoxylin and eosin. Some of specimens were stained with Sudan III. For bacteriological examination, materials from 11 cases were cultivated with sheep blood agar medium at the temperature of 37℃, 24 hours. Feed from K farm was cultivated sobouraud's glucose agar at the temperature of 37℃, 5 days. 6 cases from K farm were parasitologicaly examined for finding coccidium oocyst by flotation test. Experiment 1 18 chickens were prepared on this experiment. 3 birds were experimental lot, and 15 were control lot. Each lot was raised up to 27, 34 day old. Each lot was fed the by the feed given the birds suffered from ascites in K farm. Experimental lot was raised in sever hypoxic condision (O_2 concentration 10-13%). All cases were macro and microscopicaly examined as same procedure as field cases. Experiment 2 20 birds in experimental lot, and 19 birds in control lot were set. Each lot was raised up to 70 day old. Experimental lot was raised in the environment of mild hypoxia (O_2 concentration 15-20%). All cases were examined macro and microscopicaly. Materials from 5 cases of experimental lot and 2 cases of contol lot were cultivated with sheep blood agar medium for bacteriological examination. Hematocrits were estimated each 7 birds in experimental and contorl lot at the age of 65 day old.RESULTSField cases 1. Occurrences and environmental condisions Flocks were kept in open-system houses on K and N farm. W was windowless type. Heating system was a variety of floor heating -K, floor heating+gas brooder -W, gas brooder +stove -N. Incidence of the disease was estimated 9.8～5.4 %. Around 1% of birds were lost at the each stage of week old in flocks of K and W. Loss of birds was concentrated at the age of 1 to 3 week old in N. Ascitic birds were found out from above the age of 40 day old in each farm. Each flock occured of ascites was raised from November to March. Up to 2 week old, each flock was completely enclosed without ventilation because of sever cold in winter. On and after the 2 week old, doors of ventilation were slightly opened for regulation of temperature only on day time, then closed on night time. In case of N, flock was exposed exhausted gas from the stove, namely stove pipe was opend in the house. Oxygen in 1 out of 5 lots indicated significantly low concentration (16-20%). Carbon dioxide gas concentration in 5 lots were varied 0.5～0.15%. Ammonia gas concentration in 3 out of 5 lots indicated 10 ppm. 2. Clinical signs Disordered birds with ascites were manifested enlarged ventral part, dropness of wings and discordination. Sick birds showed duck or penguin like walk. Slight hardness of breath was observed on birds in each flock affected the disease. 3. Gross pathology Ascites was the most prominent feature, occuring in some 64% of cases. The amount of ascitic fluid varied form a few to 300 ml. Variably it was amber or wine-colored and it contained clots of fibrin. Ascites was in the cavity of liver-peritonium. The outstanding heart lesion was hydropericardium and dilatation of right atrium and ventricle. So that, wall of right-hand side ventricle was thin, flaccid and increased diameter of lumen. Venous congestion especially Vena cave caudalis was highly observed. Hepatic veins were severely engorged. Many cases of livers were enlarged with rough surfaces and rounded borders. Color of livers were varied from sanguish to paled. Many cases were hardened and some were fragile in palpation. Fibrin coat was covered on the surface of livers in cases of 30%. In some cases, yellowish necrotic mottling and vesicles on the surface could be seen. Lungs were edematous and congested. Many cases of kidneys were enlarged with paled color and others were normal size with congestion. In the gizzards, hardness of keratinoid membrane, erosion and ulceration were observed. Hemorrhage was seen in some cases of proventiculus. Congestion of duodenums were highly recognized. Striking atrophy of spleen, thymuse and bursa fabricius were highyly observed above 60% of the cases. 4. Micropathology Heart : Venous congestion was consistently observed in all cases. In many cases the myocardial fibers were shrinking and fargmentary, cytoplasm was devoid of striations, and granular or vacuolar degenerations were present. Additionaly, coagulation and liquefactive foci of necrosis were detected. In some cases the myocardiums were hypertrophic. In many cases of interstitium edema was prominent, and this was accompanied fibrosis and infiltration of leukocytes. In one case the medium sized artery in myocardium, tunica media was lost its original structur and it was organized with fibroblasts. Liver : In many cases there was prominent dilatation of hepatic veins and it affected thoughout central veins and sinusoids. In accordance with congestion, central fatty deposition, degeneration and necrosis of hepatic cells were occured in many cases. Thickning sinusoids and central fibrosis (cirrhosis) were observed in 30% of cases. These changes spread from central area of hepatic lobule to marginal side, in severe cases lesions formed network pattern with adjacent lobules. In some cases infiltration of leukocytes on necrotic lesions, and Glisson's capsuls, proliferation of bile-ducts and extramedullary hematopoiesis were recognized. Thickning of hepatic coat was highy observed. Lung : 64% of cases vessels were congested and edematous. Kidney : Venous congestion was highly observed. Degeneration of renal tubules and proliferation of glomerular endothelial cells were observed a frequency above 40%. Spleen, Thymus, Bursa fabricius : Lymph tissue atrophy was more generaly observed. In accordance with loss of lymph-cytes, network of reticular fibers were visible. Bone marrow : 41% of the cases were hypoplastic, both systems of erythrocyte and granulocyte were samely affected. Adrenal gland : The characteristic changes were observed on adrenal cortex. All the cases had at least some pathological changes. In 60% of the cases, many cells of cortex were swollen, nuclei were large and brighten, cytoplasms were week-staind with haematoxylin and eosin stain and contained various size of vacuoles. At the site of marginal area of adrenal gland, desquamation of hypertrophic cells were prominent. On the other hand, some of the lines of cortex cell were shrunk in spite of same specimen had many hypertrophic cells. Degenerative and necrotic foci were observed in many cases, but striking and extensive necrosis was located at marginal site of adrenal gland. From the results of Sudan III stain, vacuoles in cytoplasm were fatty deposition and necrotic cells lost lipoid granules. 5. Bacteriology Staphylococcus was isolated from trachea-1 case, lung-2 cases, air-sack-1 case, liver-1 case. Eschelishia coli was isolated from trachea-3 cases, lung-5 cases, heart-1 case, kidney-2 cases. Citrobactor was isolated from lung-1 case. Amount of bacteria was small and common bacteria was not isolated through cases and organs examined. The reasons above mentioned, bacterial infection could not be concerned with the diseas. Non of fungus was isolated from feed obtained K farm. 6. Parasitology Non of oocyst was detected. Experiment 1 Severe hardness of breath and stagnation of growth was observed in experimental lot. 1 case of experimental lot died at the age of 27 day old. In gross observation, all cases manifested venous congestions of heart, liver, lung and kidney. All cases of liver surface were rough and 2 of 3 cases yellowish necrotic mottles were visible. Hemorrhage of proventiculus and ulceration of gizzard were observed in each 1 of 3 cases. Spleens and bursa fabriciuses were generally small in size. Increasing of ascitic fluid was not found. Histopathological findings obtained experimental lot were as same qualities as field cases. Especially, agressive change of myocardium, fibrosis, degenerative change of artery and vein in heart, central degeneration and necrosis in liver, atrophy of lymph tissue in spleen and bursa fabricius, degenerative change in adreral cortex were prominent. Affection of feed had not been recognized. Experiment 2 Hardness of breath had been recognized in cases of experimental lot. Mean value of body weight of experiment lot had changed lower than control lot, from above 3 week old statistical significance was recognized. 1 case died at 52 day old. 2 died at 58 day old. Ascitic fluid was observed in a bird which was dead at 58 day old. Gross pathological findings were almost same as field cases. Livers of many cases were fragile and 2 cases dead at 58 day old were with prominent necrotic foci. Histological findings were as same characteristics as field cases. From bacteriological examination, Staphylococcus and Eschelishia coli were isolated from tracheas and lungs of experimental lot. These could not be concerned with bacterial infection because of a little amount of bacteria isolated. Results of the hematcrit estimation were as follows : Experimental birds (7) mean value 33.6% Control birds (7) mean value 30.9% The difference between these means was significant (p<0.05).CONCLUSION AND DISCUSSION Results of morphological studies throughout field and experimental cases, lesions were summerized as follows :(1) Progressive to agressive changes of cells of adrenal cortex.(2) Injury of heart, changes of lung, liver and kidney originated from heart failer.(3) Hemorrhage and ulceration in digestive tracts.(4) Atrophy of lymph tissues of lymph-system organs. From the results of morphological observation of adrenal cortex, its functional acceleration to exhaustion was infered. In this disease adrenal cortex performed main roles by the reasons as follows :(1) Injury of adrenal cortex was severe.(2) By the observation of experimental cases, changes of adrenal cortex were prior to all others.(3) Lesions and these distribution were characteristic.(4) Polycythemia was seen.(5) Lesions were in accordance with functional roles of adrenal cortex. From the results of field survey, experiment and morphorogical observation, pathogenesis of the ascites was considered as follow. Under the condition of insufficient ventilation (hypoxia), mechanical responsibility of heart was increased by raising of pulmonary arterial pressure with tachycardia and hyperpnoea. Though heart reacted with hypertrophy of myocardium, finally it was fatigued with work. On the other hand, hypoxia itself was a stresser, excretion of adrenocortical hormone changed from excessive to failer. During that process circulatory disturbances were encountered, those were degeneration or necrosis of myocardium, deranged metabolism of electrolytes causes an increase in the permeability of the capillaries, hemoconcentration and polycythemia. As the reason above mentioned, heart failer was induced. Among the general circulatry disturbanses, liver was severely affected and formed congestive cirrhosis. Ascitic fluid was originated from liver because of site of presence. Erosion and ulceration in digestive tracts and atrophy of lymph tissues were made during the stage of excessive excresion of adrenocortical hormone. In pathogenesis of the disease, adrenal cortex performed main role, so that "ascites in broiler chicken" was as same category as general adaptation syndrome. In this study, same lesions as field cases were obtained from experimental cases given hypoxic stress. However, adrenal cortex react nonspecifically to any stressors. Induced factors of the diseas in field, not only insufficient ventilation but also coldness, heat stroke, over population, poisonous gas, predisposition of broiler chicken ---- rapid growth, obesity and over feeding might be considered. Adrenal cortex could be affected by total stimulations combined many stressors.