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|標題:||生物殺菌劑Bacillus subtilis WG6-14作為產蛋雞益生菌及其排遺於植物病害防治應用之評估|
The use of biofungicide agent Bacillus subtilis WG6-14 as probiotics in chicken layer rearing and the evaluation of using faecess thereof produced on plant diseases management
γ-polyglutamic acid( PGA)
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|摘要:||Bacillus subtilis WG6-14 為台灣本土性可產生內生孢子之菌株，其原始分離自根圈土壤且具有廣效性抗真菌及細菌之功效。目前在生物殺菌劑之應用已相當成功，且商品化產品已成功應用在植物病害防治。此外，關於 Bacillus subtilis 亦已廣泛應用在動物飼養上，且展現益生菌之功效。本研究主要目的在於評估 Bacillus subtilis WG6-14 作為產蛋雞用益生菌之可能性。本研究將受試之 B. subtilis WG6-14 分別添加在飼料及飲水中，其中飼料試驗者係包括 30 隻 50 週齡之試驗雞隻，與 30 隻 50 週齡之對照雞隻；飲水試驗者係包括 600 隻 50 週齡之試驗雞隻，與 150 隻 50 週齡之對照雞隻。結果發現添加在飲水試驗中之試驗組，其飲水中多為典型 B. Subtilis WG6-14 菌落，而少有其他型態菌落出現；但對照組之飲水中則總菌量較高，且咸為枯草桿菌以外成員，待鑑定之微生物種類相當複雜。而將 WG6-14 添加於飲水中之試驗組，其糞便中之Salmonella 與 Staphylococcus 之微生物數量明顯較對照組低 1-2 個對數值，而 coliform 數量則明顯較對照組低約 1 個對數值。此外，在膽固醇部分，飲水中含有適量的 WG6-14 之試驗組，其蛋中膽固醇濃度亦較對照組降低約 24% 。本研究所見 WG 6-14 添加應用下，對於雞隻飼育上之諸多助益效果，顯示其於飼料添加上之應用性值得重視。 本研究進一步收集前述受試雞隻與對照雞隻之糞便排遺，以評估其應用在植物生長促進與植物病害防治之應用性。結果發現，未經堆肥化處理之雞糞，在應用上明顯可造成甘藍種子發芽不佳之影響，唯此一負面效果於 WG6-14 飼育下所收集到含有 WG6-14 之雞糞，在應用上較不明顯，且其處理植株之生長勢以葉寬受測結果看顯有比對照組為佳之現象。此外，本研究進一步發現，在雞糞混拌後，如先經數天堆置，則在小葉百喜草誘釣測試已可證實其可明顯降低土壤中存在的 R. solani AG4 之活力，此一對 R. Solani 之抑制效果在混拌含 WG6-14 雞糞 3 天即可見之而在利用未含有 WG6-14 之雞糞添加處理組，則於堆置 3 天後亦可見 AG4 活力降低的現象，然與含 WG6-14 孢子處理組相較，則有明顯差異，此一結果明白顯示堆置雞糞中含有適量 WG6-14 的存在，對於土中病原菌數量之降低確有正面效益。Bacillus 屬菌株已知可產生聚麩胺酸 (poly(γ-D-glutamic acid)(PGA))等多種聚氨基酸。針對聚麩胺酸所具有之優異保水功效一特質，本研究嘗試篩選可產生高量聚麩胺酸及兼具動物用益生菌應用性之菌株，以期能在供動物使用後，可進一步應用在植物生長及病害防治等方面。結果顯示，γ- PGA 之產生特性與對動物病原之抗生活性不見得有關，或可配合不同使用目的選取已知具不同功能之益生菌混合應用，其應可較單一菌株供應可獲較佳之成果。|
Bacillus subtilis WG6-14, a Taiwan-native endospore forming strain obtained from guava rhizosphere, has been shown to be antagonistic against wide spectrum important phytopathogenic fungi and bacteria. The development of its biofungicide application has been successful, its commercialized use in plant disease control is now in progress. As B. subtilis has been widely applied as direct-fed microbial (DFM) known with beneficial bioregulator function in animal science, the possible use of B. subtilis WG6-14 as probiotic in chicken layer rearing was explored. Both liquid and powder formulations of the tester strain (contains approximately 1010 endospores per ml/gm) produced by a fermentation pilot plant located at the campus were used for the conducted tests. The tester DFM was applied as supplements together with either the chicken feeds or water supply at final concentration approximately 108 endospores per ml/gm. A total of 30 heads (trial with powder formulation) and 750 heads (trial with liquid formulation) of chicken layers at 50 wks old were used for the tests. In liquid formulation applied trial, the fermentor produced broth culture of WG6-14 was administered to drinking water to 600 heads, the rest 150 heads without WG6-14 supplementation in drinking water were used as compared control. The daily egg yield and quality were monitored on a weekly basis. A paralleled analysis was shown by dilution plate count the micro flora within the drinking water and the layer little. The result obtained indicated the presence of solely WG6-14 in drinking water among the treated groups, the presence of contaminant microbes appeared to be non-detectable. In contrast to this, that among the non-treated group contained various kinds of contaminant microorganisms. Also worth noting was that the layer faeces from the control group consisted of bountiful amount of coliform bacteria and presence of substantial amount of Salmonella species and Staphylococcus species. Whereas the layer faeces collected from WG6-14 treated group contains approximately 104 cfu/gm of WG6-14, no Salmonella and a very much reduced numbers of coli-form and Staphylococcus spp. bacteria. Results obtained 5 weeks after treatment indicated a substantial decrease among WG6-14 treated layers the cholesterol content of egg yolks by approximately 24% as compared to the non-treated control. The observed beneficial effect of WG6-14 indicated its usefulness as probiotic feed additives to layer rearing.The chicken faecess collected from the experimental farm were evaluated further for plant growth promotion and the potential of plant disease control application. The amendment of collected chicken faeces obtained from the WG6-14 treaded birds resulted in certain growth promoting effect on cabbage foliage development, although the rate a test soil (artificially inoculated with Rhizoctonia solani AG4) with seedling emergence was affected. The compost chicken faeces collected from WG6-14 fed birds was found effective in reducing the inoculums potential of AG4. Bioassay by bahia-grass baiting indicated a significant reduction of R. solani AG4 inoculum potential by composting with 2% faeces amendment. It was worth noting that the decrease AG4 propagules was consistently greater in chicken faeces with WG6-14 comparing to that without WG6-14 amendment.Furthermore, Bacillus spp. are gram-positive bacteria, and are able to synthesize the polyamino acid (PAA), poly(γ-D-glutamic acid)(PGA) as a capsular substance or as a water-soluble slime. PGA is an excellent water-keeping material. The potential of Bacillus spp. for producing γ-PGA and the relationship of γ-PGA producing ability and the probiotic function were explored. Results obtained indicated the productivity of γ-PGA of a test strain did not correlate well the effectiveness of probiotic function. For the intended probiotics application, the use of mixed strains each with suitable desired function may be a more appropriated way than the application of a single strain.
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