Long-Read Sequencing and Hybrid Assembly for Genomic Analysis of Clinical Brucella melitensis Isolates.

Brucella melitensis is a key etiological agent of brucellosis and has been increasingly subject to characterization using sequencing methodologies. This study aimed to investigate and compare short-read, long-read, and hybrid assemblies of B. melitensis. Eighteen B. melitensis isolates from Southern Israel were sequenced using Illumina and the Oxford Nanopore (ONP) MinION, and hybrid assemblies were generated with ONP long reads scaffolded on Illumina short reads. Short reads were assembled with INNUca with SPADes, long reads and hybrid with dragonflye. Abricate with the virulence factor database (VFDB) and in silico PCR (for the genes BetB, BPE275, BSPB, manA, mviN, omp19, perA, PrpA, VceC, and ureI) were used for identifying virulence genes, and a total of 61 virulence genes were identified in short-read, long-read, and hybrid assemblies of all 18 isolates. The phylogenetic analysis using long-read assemblies revealed several inconsistencies in cluster assignment as compared to using hybrid and short-read assemblies. Overall, hybrid assembly provided the most comprehensive data, and stand-alone short-read sequencing provided comparable data to stand-alone long-read sequencing regarding virulence genes. For genomic epidemiology studies, stand-alone ONP sequencing may require further refinement in order to be useful in endemic settings.

Genomic Epidemiology of Clinical Brucella melitensis Isolates from Southern Israel.

Brucellosis, a zoonosis mainly transmitted by consumption of unpasteurized dairy products, is endemic in Southern Israel, mainly among the Bedouin Arab population. However, the genomic epidemiology of B. melitensis in this region has not yet been elucidated. A cohort of brucellosis cases (n = 118) diagnosed between 2017-2019 was studied using whole-genome sequencing (WGS). Phylogenetic analyses utilized core genome MLST (cgMLST) for all local isolates and core genome SNPs for 347 human-associated B. melitensis genomes, including Israeli and publicly available sequences. Israeli isolates formed two main clusters, presenting a notable diversity, with no clear dominance of a specific strain. On a global scale, the Israeli genomes clustered according to their geographical location, in proximity to genomes originating from the Middle East, and formed the largest cluster in the tree, suggesting relatively high conservation. Our study unveils the genomic epidemiology of B. melitensis in Southern Israel, implicating that rather than a common source, the transmission pattern of brucellosis among Bedouin communities is complex, predominantly local, and household-based. Further, genomic surveillance of B. melitensis is expected to inform future public health and veterinary interventions and clinical care.