与巴基羔羊细菌性腹泻易感性相关的核苷酸序列变异、基因表达及血清免疫和抗氧化标记物概况

IF 2.3 2区 农林科学 Q1 VETERINARY SCIENCES BMC Veterinary Research Pub Date : 2024-10-11 DOI:10.1186/s12917-024-04288-1
Asmaa Darwish, Eman Ebissy, Amani Hafez, Ahmed Ateya, Ahmed El-Sayed
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引用次数: 0

摘要

背景:尽管将腹泻描述为一种临床症状比描述为一种疾病更为准确。腹泻是绵羊医学中最重要的问题之一,尤其是对羔羊而言,由于其发病率和死亡率高、生长迟缓、兽医成本高,被认为是经济损失的主要来源。沙门氏菌和肠毒性大肠杆菌是导致腹泻的最常见、最具商业价值的病原体:本研究旨在监测腹泻羔羊的核苷酸序列变异、基因表达、血清炎症和氧化应激生物标志物。另一个目的是确定引起腹泻的沙门氏菌和大肠杆菌的不同病理类型和毒力基因:从 50 只腹泻羔羊和 50 只健康羔羊身上采集血液样本,然后分成 3 份,第一份加入 EDTA 进行全血细胞计数、DNA 和 RNA 提取。第二份样本中加入 5000 I.U. 的肝素钙,第三份样本则使用干净的普通试管。第二和第三部分离心提取血清和血浆,直至完成生化和免疫分析。收集粪便样本进行细菌学检查,并通过 PCR 分析鉴定细菌。对免疫(SELL、JAK2、SLC11A1、IL10、FEZF1、NCF4、LITAF、SBD2、NFKB、TNF-α、IL1B、IL6、LGALS 和 CATH1)进行 PCR-DNA 测序、健康羔羊和腹泻羔羊的抗氧化基因(SOD1、CAT、GPX1、GST、Nrf2、Keap1、HMOX1 和 NQO1)以及胃肠道健康基因(CALB1、GT 和 MUC2)。结果在所有腹泻羔羊中都检测到了大肠杆菌(astA、Vt2e (Stx2e)、CFA/I、groES 和 luxS)和沙门氏菌(invA、SopB、bcfC 和 avrA)的致病性病毒基因标记。免疫、抗氧化和肠道健康基因的 PCR-DNA 测序发现 11 个单核苷酸多态性(SNPs)与巴基羔羊的腹泻抵抗力或腹泻易感性有关。免疫、抗氧化和肠道健康(CALB1、GT和MUC2)基因的转录水平在健康羔羊和腹泻羔羊之间存在差异。所调查基因的核苷酸序列在 GenBank 中的参考序列与所调查动物的参考序列之间的变异验证了所有已确定的 SNPs。腹泻羔羊出现了明显的红细胞增多症、中性粒细胞增多症和淋巴细胞减少症(P = 0.001)。血清 IL-1α、IL-1β、IL-6、TNF-α(分别为 90.5 ± 1.7、101.8 ± 1.7、72.3 ± 6.6、71.26 ± 4.89 Pg/ml)、血清 Fb、Cp、Hp、SAA(分别为 230.7 ± 12.4 mg/dl、6.5 ± 0.07 mg/dl、2.5 ± 0.09 g/dl、7.4 ± 0.4 mg/L)、自由基(MDA、NO)、皮质醇(6.91 ± 0.18 μg/dl)和生长激素,腹泻羔羊血清IL-10(81.71 ± 1.05 Pg/ml)、抗氧化剂(CAT、GPx)、胰岛素、三碘甲状腺原氨酸(T3)和甲状腺素(T4)显著下降(P = 0.001):研究结果证实了以下理论:根据与炎症、抗氧化剂和肠道健康有关的基因 SNPs 选择羔羊,标记辅助选择(MAS)可用于预测和预防巴尔基绵羊腹泻。为了制定有效的管理方案并确定疾病发生的最易感风险期,还可以利用所研究基因的基因表达谱、促炎细胞因子和急性期蛋白作为羔羊肠炎的替代生物标记物。
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Nucleotide sequence variants, gene expression and serum profile of immune and antioxidant markers associated with bacterial diarrhea susceptibility in Barki lambs.

Background: Despite the fact that diarrhea is more accurately described as a clinical symptom than a disease. Diarrhea is one of the most important issues in ovine medicine, particularly in lambs, and because of high morbidity and mortality rate, sluggish growth performance, and veterinary costs, it is believed to be a major source of economic loss. Salmonella and enterotoxigenic Escherichia coli are the most common and commercially significant agents responsible for diarrhea.

Objective: The objective of this study was to monitor the nucleotide sequence variations, gene expression, serum inflammatory and oxidative stress biomarkers in diarrheic lambs. Another aim was to identify different pathotypes and virulence genes of Salmonella and E. coli causing diarrhea.

Methodology: Blood samples were taken from 50 Barki who were diarrheal and 50 who appeared to be healthy, and then divided in 3 portions, with EDTA added to the first part for CBC, DNA and RNA extraction. The second sample received 5000 I.U. of heparin calcium, and a clean plain tube was used for the third component. The second and third sections were centrifuged to extract serum and plasma until the biochemical and immunological analysis was completed. Fecal samples were collected for bacteriological examination, and the bacteria were identified by PCR analysis. PCR-DNA sequencing was conducted for immune (SELL, JAK2, SLC11A1, IL10, FEZF1, NCF4, LITAF, SBD2, NFKB, TNF-α, IL1B, IL6, LGALS, and CATH1), antioxidant (SOD1, CAT, GPX1, GST, Nrf2, Keap1, HMOX1, and NQO1), and GIT health (CALB1, GT, and MUC2) genes in healthy and diarrheic lambs.

Results: Virulent genetic markers of pathogenic characteristics of E. coli (astA, Vt2e (Stx2e), CFA/I, groES and luxS) and Salmonella (invA, SopB, bcfC and avrA) were detected in all diarrheic lambs. PCR-DNA sequencing of immune, antioxidant and intestinal health genes found eleven single nucleotide polymorphisms (SNPs) linked to either diarrhea resistance or susceptibility in Barki lambs. Transcript levels of immune, antioxidant, and GIT health (CALB1, GT, and MUC2) genes varied between healthy and diarrheic lambs. Nucleotide sequence variation of the genes under inquiry between reference sequences in GenBank and those of the animals under investigation verified all identified SNPs. Significant (P = 0.001) erythrocytosis, neutrophilic leukocytosis, with lymphocytopenia were observed in diarrheic lambs. Significant (P = 0.001) increases in serum IL-1α, IL-1β, IL-6, TNF-α (90.5 ± 1.7, 101.8 ± 1.7, 72.3 ± 6.6, 71.26 ± 4.89 Pg/ml, respectively), serum Fb, Cp, Hp, SAA (230.7 ± 12.4 mg/dl, 6.5 ± 0.07 mg/dl, 2.5 ± 0.09 g/dl, 7.4 ± 0.4 mg/L, respectively), free radicals (MDA, NO), cortisol (6.91 ± 0.18 μg/dl) and growth hormone, with significant (P = 0.001) decreases in serum IL-10 (81.71 ± 1.05 Pg/ml), antioxidants (CAT, GPx), insulin, triiodothyronine (T3) and thyroxine (T4) in diarrheic lambs.

Conclusions: The study's findings provided credence to the theory that marker-assisted selection (MAS) could be used to predict and prevent diarrhea in Barki sheep by selecting lambs based on SNPs in genes linked to inflammation, antioxidants, and intestinal health. In order to establish an efficient management protocol and determine the most susceptible risk period for disease occurrence, gene expression profiles of the genes under investigation, pro-inflammatory cytokines and acute phase proteins may also be utilized as proxy biomarkers for lamb enteritis.

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来源期刊
BMC Veterinary Research
BMC Veterinary Research VETERINARY SCIENCES-
CiteScore
4.80
自引率
3.80%
发文量
420
审稿时长
3-6 weeks
期刊介绍: BMC Veterinary Research is an open access, peer-reviewed journal that considers articles on all aspects of veterinary science and medicine, including the epidemiology, diagnosis, prevention and treatment of medical conditions of domestic, companion, farm and wild animals, as well as the biomedical processes that underlie their health.
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