There has been a concerted effort to maintain an assurance population of cheetahs in North American zoos and conservation centers to combat dwindling wild populations. Wild cheetah ( Acinonyx jubatus) populations are considered “vulnerable” according to the International Union of Conservation Nature (IUCN) but are quickly approaching “endangered” status as populations continue to decline. Collectively, these data suggest that the MLV is capable of causing clinical disease and viral shedding in some cheetahs and represents evidence of interspecies transmission of virus between domestic and wild cats. The likely ancestral origin of these two isolates involves recombination events between Australian domestic cat and cheetah FHV-1 isolates. The remaining two cheetah FHV-1 isolates (unknown host vaccine status) were not associated with a clade. Eight cheetah FHV-1 isolates and the MLV were grouped in a clade along with FHV-1 isolates from domestic cats in the USA. The FHV-1 shed by vaccinated cheetahs were almost identical to the MLV, with few variants among viral genomes. Viral DNA was extracted for full-genome sequencing by Illumina MiSeq with viral genomes then used for phylogenomic and recombinational analyses. Ten FHV-1 isolates from ten captive cheetahs and one isolate from an MLV used to inoculate four of the host animals were analyzed. Modified live vaccines (MLV), intended for use in domestic cats, are used in some captive cheetah populations and have been anecdotally linked to disease in certain subpopulations. In addition, further analysis may include identification of virulence and antibiotic resistance genes for improved pathogen characterization.Feline herpesvirus type 1 (FHV-1) is endemic in captive cheetahs and sporadically causes devastating disease. The suggested protocol enables identification of outbreaks in early stages using a portable and low-cost device along with a streamlined downstream analysis, therefore having the potential to be incorporated in routine surveillance analysis workflows. aureus outbreak investigation, the protocol was able to identify two outbreaks in less than 31 h. All these estimated times were calculated considering the average time for six MRSA isolates per sequencing run. Overall, the developed protocol was able to at least discard an outbreak in 27 h (mean) after the bacterial identification and less than 33 h to confirm it. The phylogenetic analysis took 0.6 h to confirm an outbreak. Assemblies were achieved after 4 h (40 min per isolate) while the polishing was carried out in 7 min per isolate (42 min in total). After the sequencing run, it was possible to identify the ST in 2 h (20 min per isolate). The suggested protocol includes: (1) a 20 h sequencing run (2) identification of the sequence type (ST) (3) de novo genome assembly (4) polishing of the draft genomes and (5) phylogenetic analysis based on SNPs. aureus isolates was conducted to test the ONT-based protocol. Additionally, a real-time outbreak investigation of six clinical S. The validation of the protocol was performed using Illumina technology (MiSeq, Illumina). The protocol was developed using 42 methicillin-resistant Staphylococcus aureus (MRSA) isolates identified from former well-characterized outbreaks. This study developed and validated a complete analysis protocol for faster and more accurate surveillance and outbreak investigations of antibiotic-resistant microbes based on Oxford Nanopore Technologies (ONT) DNA whole-genome sequencing. Outbreak investigations are essential to control and prevent the dissemination of pathogens.
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