Gomaa Basant, Lu Jingjun, Abdelhamed Hossam, Banes Michelle, Pechanova Olga, Pechan Tibor, Arick Mark A, Karsi Attila, Lawrence Mark L
Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS 39762, USA.
Institute for Genomics, Biocomputing, and Biotechnology, Mississippi State University, Mississippi State, MS 39762, USA.
Foods. 2024 Apr 24;13(9):1302. doi: 10.3390/foods13091302.
is the causative agent of listeriosis, a severe foodborne illness characterized by septicemia, meningitis, encephalitis, abortions, and occasional death in infants and immunocompromised individuals. is composed of four genetic lineages (I, II, III, and IV) and fourteen serotypes. The aim of the current study was to identify proteins that can serve as biomarkers for detection of genetic lineage III strains based on simple antibody-based methods. Liquid chromatography (LC) with electrospray ionization tandem mass spectrometry (ESI MS/MS) followed by bioinformatics and computational analysis were performed on three strains (NRRL B-33007, NRRL B-33014, and NRRL B-33077), which were used as reference strains for lineages I, II, and III, respectively. Results from ESI MS/MS revealed 42 unique proteins present in NRRL B-33077 and absent in NRRL B-33007 and NRRL B-33014 strains. BLAST analysis of the 42 proteins against a broader panel of >80 sequenced strains from lineages I and II revealed four proteins [TM2 domain-containing protein (NRRL B-33077_2770), DUF3916 domain-containing protein (NRRL B-33077_1897), DNA adenine methylase (NRRL B-33077_1926), and protein RhsA (NRRL B-33077_1129)] that have no homology with any sequenced strains in lineages I and II. The four genes that encode these proteins were expressed in strain DE3 and purified. Polyclonal antibodies were prepared against purified recombinant proteins. ELISA using the polyclonal antibodies against 12 lineage I, II, and III isolates indicated that TM2 protein and DNA adenine methylase (Dam) detected all lineage III strains with no reaction to lineage I and II strains. In conclusion, two proteins including TM2 protein and Dam are potentially useful biomarkers for detection and differentiation of lineage III strains in clinical, environmental, and food processing facilities. Furthermore, these results validate the approach of using a combination of proteomics and bioinformatics to identify useful protein biomarkers.
是李斯特菌病的病原体，李斯特菌病是一种严重的食源性疾病，其特征为败血症、脑膜炎、脑炎、流产，以及婴儿和免疫功能低下个体偶尔死亡。由四个遗传谱系（I、II、III和IV）和十四种血清型组成。本研究的目的是基于简单的基于抗体的方法，鉴定可作为检测遗传谱系III菌株生物标志物的蛋白质。对三株菌株（NRRL B - 33007、NRRL B - 33014和NRRL B - 33077）进行了液相色谱（LC）与电喷雾电离串联质谱（ESI MS/MS）分析，随后进行生物信息学和计算分析，这三株菌株分别用作谱系I、II和III的参考菌株。ESI MS/MS的结果显示，NRRL B - 33077中存在42种独特蛋白质，而NRRL B - 33007和NRRL B - 33014菌株中不存在。对这42种蛋白质针对来自谱系I和II的80多种测序菌株进行的BLAST分析显示，有四种蛋白质[含TM2结构域的蛋白质（NRRL B - 33077_2770）、含DUF3916结构域的蛋白质（NRRL B - 33077_1897）、DNA腺嘌呤甲基化酶（NRRL B - 33077_1926）和蛋白质RhsA（NRRL B - 33077_1129）]与谱系I和II中的任何测序菌株均无同源性。编码这些蛋白质的四个基因在菌株DE3中表达并纯化。制备了针对纯化重组蛋白的多克隆抗体。使用针对12株谱系I、II和III分离株的多克隆抗体进行的ELISA表明，TM2蛋白和DNA腺嘌呤甲基化酶（Dam）可检测到所有谱系III菌株，而与谱系I和II菌株无反应。总之，包括TM2蛋白和Dam在内的两种蛋白质可能是临床、环境和食品加工设施中检测和区分谱系III菌株的有用生物标志物。此外，这些结果验证了使用蛋白质组学和生物信息学相结合的方法来鉴定有用蛋白质生物标志物的有效性。
