利用非侵入式取樣方法分析BoLA-DRB3.2基因與乳房炎及泌乳性狀之關係

Abstract

乳產量與生乳組成分之提升常為泌乳牛之主要育種目標，然而長期選拔的過程中多半忽略如抗病性等重要之乳牛健康性狀。 舉例而言，乳房炎為泌乳牛最常見且重要之疾病，一旦牛隻感染乳房炎，不僅導致人力成本增加，且因於治療過程中之獸醫師費用、藥物，以及廢棄之生乳等因素導致增加額外之生產成本，更甚者可能造成乳牛提早淘汰。近年之研究發現某些疾病之抗性或可藉由免疫與遺傳標記篩選，針對此類基因進一步的研究或可解決因疾病而造成之乳量與組成分損失等課題。 主要組織相容複合體（major histocompatibility complex）與哺乳動物之免疫系統息息相關。牛隻主要組織相容複合體被稱為牛隻淋巴球抗原（bovine lymphocyte antigen, BoLA），由三個不同區域組成且與抗原呈獻有關；因此，BoLA基因已於不同疾病免疫與生產性狀之議題中廣泛地被研究。BoLA基因座上第三個DR區域（DRB3）已被發現具有相當高度之多態性，尤以第二個外顯子（exon 2）之多態性可能與牛隻免疫性狀相關。本試驗收集來自台灣大學試驗農場23頭荷蘭牛泌乳牛之生乳樣本取代傳統之血液與組織樣本，以探討利用非侵入式採樣於泌乳牛隻進行研究之可能性。試驗期間同時收集乳牛群性能改進計畫（Dairy Herd Improvement）資料以針對體細胞數（somatic cell counts）與生乳組成分與DRB3.2基因之關係進行討論。牛群依其於試驗期間體細胞數達臨床乳房炎等級之頻率分成三個群組，分別為易敏感性群組（100%）、一般性群組（40至80%），以及抗乳房炎群組（40%以下）。 本試驗結果顯示於23頭試驗牛隻中，有17頭個體可成功自乳品樣本中抽取DNA並進行PCR-SBT分析；其中抗乳房炎群組有6頭，一般性群組有7頭，易敏感性群組有4頭。於此17頭個體中共發現11個不同之DRB3.2交替基因型，*1201與*2703交替基因型僅見於抗乳房炎與一般性群組中，與加拿大東岸之荷蘭牛研究有相似之結果，顯示其為一良好之抗乳房炎交替基因型。*1501交替基因型之純合子僅見於易敏感性群組中，且帶有此純合子之個體其發炎程度均高於其他牛隻，顯示該交替基因型與乳房炎之敏感性相關，建議日後可列為育種選拔之參考。

Increasing productivity such as milk yields and milk components is always the main breeding goal for dairy cattle. However, some health traits such as disease resistance have been ignored for a long time. For example, mastitis is an important and world-wide disease in dairy herds and leads to not only add costs, including the expenditures of labors, vet service, drugs, but also the losses from lower production , discarding milk, and culling of cows. Nowadays, it is evident that disease resistance may be genetically determined. Selecting resistance to health problems can reduce the economical losses and increase milk quality, which is important to ranchman and consumers. Consequently, genetic and immunological markers have been widely studied expecting to be used in disease resistance selections. The major histocompatibility complex (MHC) groups are closely linked to mammalian immune system. For dairy cattle, MHC referred to as the bovine lymphocyte antigen (BoLA) complex, consisting of three classes and being responsible for antigens presentation. Hence, BoLA genes have been examined for various immune issues as well as production traits. The DRB3 locus of BoLA class II is very polymorphic, and exon 2 (DRB3.2) may potentially affect immunity in cattle. In this study, we collected milk samples instead of blood or tissue as the resource of DNA extractions from 23 dairy Holstein cows in NTU Experimental Farm, trying to evaluate the preliminary feasibility of constructing noninvasive sampling methods in dairy herds. Data from Dairy Herd Improvement (DHI) in recently five months were also used to investigate the relationship between somatic cell counts (SCC), milk components, and DRB3.2 alleles. According to the frequency of that SCC reached clinical mastitis we labeled the cows into three classifications, known as resistant group (under 40%), normal (40 to 80%), and sensitive one (100%). The results showed 17 individuals from 23 dairy cows could be sequenced successfully in PCR-SBT analysis, including six individuals in resistance group, seven in normal, and four in sensitive, respectively. Among these 17 individuals, 11 alleles were found. Alleles *1201 and *2703 were only found in resistance and normal group but not in sensitive one, and this result is in accordance with former study. Individual with allele *1501 homozygous expressed more serious mastitis level and only in sensitive group, which may tend toward inflammation easier. It may be the candidate allele of mastitis in breeding.