Department of Medical Microbiology and Infectious Diseases, Erasmus University Medical Center Rotterdam, 3015 CN Rotterdam, The Netherlands.
Child Health Research Foundation, 23/2 SEL Huq Skypark, Block-B, Khilji Rd, Dhaka 1207, Bangladesh.
Genes (Basel). 2020 Nov 18;11(11):1365. doi: 10.3390/genes11111365.
Typhoid fever, caused by serovar Typhi ( Typhi), is a global health concern and its treatment is problematic due to the rise in antimicrobial resistance (AMR). Rapid detection of patients infected with AMR positive Typhi is, therefore, crucial to prevent further spreading. lustered egularly nterspaced hort alindromic epeats and CRISPR-associated genes (CRISPR-Cas), is an adaptive immune system that initially was used for typing purposes. Later, it was discovered to play a role in defense against phages and plasmids, including ones that carry AMR genes, and, at present, it is being explored for its usage in diagnostics. Despite the availability of whole-genome sequences (WGS), very few studied the CRISPR-Cas system of Typhi, let alone in typing purposes or relation to AMR. In the present study, we analyzed the CRISPR-Cas system of Typhi using WGS data of 1059 isolates obtained from Bangladesh, India, Nepal, and Pakistan in combination with demographic data and AMR status. Our results reveal that the Typhi CRISPR loci can be classified into two groups: A (evidence level >2) and B (evidence level ≤2), in which we identified a total of 47 unique spacers and 15 unique direct repeats. Further analysis of the identified spacers and repeats demonstrated specific patterns that harbored significant associations with genotype, demographic characteristics, and AMR status, thus raising the possibility of their usage as biomarkers. Potential spacer targets were identified and, interestingly, the phage-targeting spacers belonged to the group-A and plasmid-targeting spacers to the group-B CRISPR loci. Further analyses of the spacer targets led to the identification of an Typhi protospacer adjacent motif (PAM) sequence, TTTCA/T. New -genes known as , , and were also discovered in the Typhi genome. However, a specific variant of the gene was only identified in the extensively drug-resistant (XDR) lineage from Pakistan and ciprofloxacin-resistant lineage from Bangladesh. From this work, we conclude that there are strong correlations between variations identified in the Typhi CRISPR-Cas system and endemic AMR positive Typhi isolates.
伤寒由伤寒血清型（ Typhi ）引起，是一个全球性的健康问题，由于抗菌药物耐药性（AMR）的上升，其治疗变得成问题。因此，快速检测感染 AMR 阳性伤寒的患者对于防止进一步传播至关重要。规律成簇间隔短回文重复序列和 CRISPR 相关基因（CRISPR-Cas）是一种适应性免疫系统，最初用于分型目的。后来，人们发现它在防御噬菌体和质粒方面发挥作用，包括携带 AMR 基因的质粒，目前，它正在被探索用于诊断。尽管有全基因组序列（WGS）可用，但很少有研究针对伤寒的 CRISPR-Cas 系统，更不用说在分型目的或与 AMR 的关系上了。在本研究中，我们使用来自孟加拉国、印度、尼泊尔和巴基斯坦的 1059 个分离株的 WGS 数据，结合人口统计学数据和 AMR 状况，分析了伤寒的 CRISPR-Cas 系统。我们的结果表明，伤寒的 CRISPR 基因座可以分为两类：A（证据水平>2）和 B（证据水平≤2），其中我们总共鉴定了 47 个独特的间隔区和 15 个独特的直接重复区。对鉴定出的间隔区和重复区的进一步分析表明，存在与基因型、人口统计学特征和 AMR 状况显著相关的特定模式，从而提高了它们作为生物标志物的可能性。潜在的间隔区靶点被鉴定出来，有趣的是，靶向噬菌体的间隔区属于 A 组，而靶向质粒的间隔区属于 B 组 CRISPR 基因座。对间隔区靶点的进一步分析导致鉴定出一个伤寒原间隔基序（PAM）序列 TTTCA/T。在伤寒基因组中还发现了新的基因，分别称为、和。然而，只有在巴基斯坦的广泛耐药（XDR）谱系和孟加拉国的环丙沙星耐药谱系中才发现基因的特定变体。从这项工作中，我们得出结论，在伤寒的 CRISPR-Cas 系统中鉴定的变异与地方性 AMR 阳性伤寒分离株之间存在很强的相关性。
