Abstract

Marine mammals strand along the California coastline for a variety of ailments, including primary or secondary bacterial and fungal infections.^1-3 Diagnostic assessment of these microbial infections is indicated to guide therapeutic plans during rehabilitation. Historically, conventional culture and sensitivity testing (CS) has been the standard diagnostic tools to identify microbial agents and provide guidance for antimicrobial selection. Next-generation DNA sequencing (NGS) is a more recently developed technology utilized to diagnose bacterial and fungal infections, as well as detect acquired and intrinsic antibiotic resistance genes.^4-9 The goal of this study was to compare next-generation DNA sequencing to conventional culture and sensitivity as a diagnostic tool for infections in marine mammals.

Study subjects consisted of California sea lions (Zalophus californianus) undergoing treatment in a rehabilitation setting at the Pacific Marine Mammal Center, Laguna Beach, California. Samples (n=30) were collected in duplicate for conventional culture and sensitivity testing and next-generation DNA sequencing. Sample types included swabs of open wounds or closed abscesses of the skin, oral cavity, cornea, as well as urine for urinary tract infections. Conventional culture and sensitivity testing included aerobic incubation for all samples, with anaerobic incubation (73%, 22/30) and fungal (13%, 4/30) culture for a subset of the samples.

In 90.0% (27/30) and 75.0% (3/4) of the cases, conventional culture and sensitivity reported bacterial or fungal growth, respectively, while next-generation DNA sequencing testing reported 100% (30/30) of bacterial positive and 60.0% (18/30) of fungal positive results. Specifically, next-generation sequencing had a higher sensitivity in detecting clinically relevant bacterial and fungal pathogens, including but not limited to: Campylobacter pinnipediorum, Clostridium spp., Corynebacterium spp., Fusobacterium spp., Morganella morganii, Mycoplasma spp., Neisseria spp., and Aspergillus tonophilus, Candida albicans, Malassezia spp., Trichosporon spp. Substantial numbers of both bacterial and fungal pathogens were missed by conventional culture testing, but identified using next-generation DNA sequencing.

Next-generation DNA sequencing may be more beneficial in a clinical setting when compared to conventional culture and sensitivity, particularly when considering the addition of anaerobic and fungal cultures to aerobic incubation. Fungal cultures are not routinely submitted during initial clinical evaluation due to cost prohibition. Next-generation DNA sequencing provides results consisting of aerobic, anaerobic, and fungal agents with antibiotic-resistant genetic information for less than half the cost of standard conventional culture and sensitivity submitted to a reference laboratory with a faster turnaround time (48–72 hours) compared to the 7–16 days or more required for conventional anaerobic and fungal cultures. Next-generation DNA sequencing allows for more cost-effective diagnostic abilities and facilitates faster clinically relevant antibiotic choices for therapeutic plans. This study showcases the utility of next-generation DNA sequencing-based diagnostics in marine mammal medicine.

Acknowledgements

The authors wish to thank the staff and volunteers at Pacific Marine Mammal Center. The authors also wish to thank the Pacific Life Foundation and the Doris Day Animal Foundation for their generous donations supporting our patient care efforts. Rescue and rehabilitation activities were conducted under the NOAA stranding agreement.

*Presenting author

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