Part II - THE COOL KID IN THE GAME! NGS FOR ANIMAL MICROBIOME
- Dr. Nihan
- Dec 11, 2025
- 8 min read
Last week, we talked about what NGS is, the three layers of data (Composition, Function, Activity), the toolbox from 16S to metatranscriptomics, and why seeing the whole microbial ecosystem beats chasing a single suspect. If you missed that one, I recommend starting there before diving into this.
Now the question is: where is this actually being used? We will also talk about where is it going next for those of you building products, running diagnostics, or treating patients.
Let me walk you through it.
Where Is Animal Microbiome NGS Actually Being Used?
1. Companion Animal Gut Microbiome
Back in 2011, a great work done by Handl et al. used massive parallel 16S rRNA gene pyrosequencing to show that healthy dog and cat feces contain far more diverse bacterial and fungal communities than anyone had documented before. That paper was a wake-up call for all of us: the gut is a special and unique ecosystem!
Since then, 16S rRNA sequencing has become the standard method for characterizing healthy versus diseased states, and responses to diet or therapy. Reviews like Suchodolski (2021) made this clear, while also warning us about the lack of standardization between studies and labs. I believe the strongest message for us was molecular tools are powerful, but validation really matters.
More recently, studies are moving toward:
Species-level baseline mapping in healthy dogs using full-length 16S (see Rojas et al., 2024 - 286 home-living dogs in North America, strict inclusion criteria, a real step forward from genus-level bar charts).
Linking microbiome patterns to chronic enteropathies, obesity, allergies, and therapeutic diets in more structured ways.
For clinicians, this is slowly translating into better reference ranges, more realistic dysbiosis indexes, and clearer expectations about what "normal" looks like across breeds, ages, and geographies.
Worth noting here: Suchodolski's validated qPCR-based Dysbiosis Index for dogs represents an important middle ground between single-pathogen PCR and full NGS - it measures seven key bacterial taxa and summarizes the results as one number. That kind of tool bridges the gap nicely for everyday clinical use. Still work in progress overall, but we are getting there.
2. Skin, Ear, and Urinary Microbiomes
NGS has also been applied beyond the gut. Skin, ear, and mucosal surfaces started to gain interest to look into more and more. Recent studies of canine skin have documented complex microbiota profiles in both healthy and atopic dogs, expanding earlier 16S-based descriptions and highlighting site-specific communities. A 2024 review by Santoro et al. provides a good update on our current understanding of skin barrier, cutaneous microbiome, and host defense peptides in canine atopic dermatitis.
Clinical case series (from MiDOG and collaborators, among others) show NGS helping identify bacterial and fungal drivers in:
Chronic otitis (including exotic species like rabbits
Chronic rhinosinusitis
Other difficult infections where culture was unrevealing or incomplete
This is particularly useful when the biofilm-associated infections keep recurring, previous antibiotics distort culture results and (my favorite) when fungi and bacteria are both in the picture.
3. Livestock and Aquaculture Microbiomes
NGS is now routine in research programs looking at feed efficiency, performance, and disease resilience. Companies like Porcinotec in Thailand use NGS to profile the gut microbiome of pigs and other livestock, aiming to adjust feed and management for better productivity and health.
Reviews highlight how farm animal gut microbiome studies increasingly rely on NGS to explore links between diet, microbiota, and antimicrobial resistance (AMR). In aquaculture, similar approaches are used to understand:
• Microbiome shifts under intensive farming
• Interactions between probiotics, pathogens, and water quality
This is where the One Health and AMR angles become very real since the farm animal gut is a reservoir, and NGS lets us see it.
4. Diagnostic Labs and One Health Programs
The shift from 'special research projects with NGS' to have it as a routine diagnostic menu item is one of the clearest signs that NGS is finding more place for itself in veterinary medicine. Here are couple examples from public sources I could find:
The Cornell Animal Health Diagnostic Center offers NGS services for viral diagnostics and exploratory metagenomics in a wide range of species and sample types. Their Virology Laboratory uses both MinION (Oxford Nanopore) and MiSeq (Illumina) platforms.
The University of Minnesota Veterinary Diagnostic Lab provides shotgun metagenomics for pathogen detection. They are refreshingly clear about limitations on their website - explicitly noting that failure to identify a pathogen does not guarantee absence, and listing specific reasons for potential false negatives including low viral titer, host DNA competition, and inadequate sequencing depth. That kind of transparency is exactly what we need.
Globally, One Health initiatives use NGS hubs for metagenomic surveillance of undiagnosed febrile and enteric diseases, integrating animal and human data.
Key Papers That Show What NGS Can Do
I got several requests from readers of the Part I for additional papers, so I am adding another bunch of good read pieces on the topic to read further on this topic:
Handl et al., 2011 - Massive parallel 16S rRNA gene pyrosequencing in healthy dogs and cats. This was one of the first large-scale molecular characterizations showing that normal fecal microbiota in dogs and cats are far more diverse than culture suggests. Set the stage for treating the gut as an ecosystem.
Suchodolski, 2021 - Analysis of the gut microbiome in dogs and cats. Comprehensive review of methods and clinical associations, with a clear warning about lack of standardization. Also introduces the Dysbiosis Index as a validated qPCR tool. Key message: molecular tools are powerful, but validation and consistent reporting are critical.
Shah et al., 2024 - Decoding the gut microbiome in companion animals. Summarizes evidence linking gut microbiome patterns to clinical conditions including inflammatory bowel disease, obesity, and neurological conditions. Emphasizes that most work still uses 16S, while deep shotgun metagenomics is needed for functional insights and resistance genes.
Rojas et al., 2024 - Species-level core fecal microbiome in healthy dogs. Uses full-length 16S (PacBio sequencing) to define a species-level core microbiome with strict inclusion criteria. Identified 23 bacterial species comprising the core microbiome. Provides a precise baseline crucial for interpreting dysbiosis scores, probiotics, or FMT approaches. Note: All authors are affiliated with AnimalBiome, a commercial microbiome testing company - doesn't invalidate the science but worth knowing.
Damerum et al., 2023 - NGS vs culture in exotic animals. Direct head-to-head comparison showing NGS detects many bacterial and most fungal pathogens missed by culture in exotic animal patients. Specifically, culture failed to grow 15% of bacterial and 81% of fungal pathogens identified by NGS. Strong practical signal that in certain contexts, NGS is not a luxury - it is the only way to see what is actually there.
You could easily add Afonso et al. (2023) on avian/poultry pathogen detection, or Domrazek et al. (2024) reviewing NGS applications in companion animal veterinary medicine more broadly. Together, these papers show a clear arc: description, baseline building, real diagnostic decisions, surveillance planning, and more.
The Fine Print: Limitations and Responsible Use
I love what NGS can do for sure but we always consider the hard points, as well, right?
No, a single microbiome report is not a diagnosis.
Without context (history, clinical exam, other tests), NGS is just a dense list of names and relative abundances. Over-interpretation is a real risk, especially in commercial "health score" reports. Before ordering a commercial microbiome test, ask about validation status, what reference databases are used, and how results are reported and interpreted.
Methodology matters.
Different labs use different extraction methods, primers, platforms, and bioinformatics pipelines. This means results are not always directly comparable across labs or over time. Sample collection and handling also matter - NGS results are highly sensitive to pre-analytical variables, so proper sample preservation and transport are essential.
False negatives can still happen.
Metagenomic tests can miss pathogens if the titer is low, host DNA overwhelms the sample, sequencing depth is insufficient, or nucleic acids are degraded. As Minnesota VDL puts it clearly on their website: failure to identify a pathogen is not a guarantee that the pathogen is absent.
Turnaround time and cost.
For some emergencies, you still need fast culture/PCR. Culture typically gives you results in 24-72 hours for many pathogens, while NGS usually takes 1-3 weeks. Cost-wise, expect roughly $150-400 per sample for 16S amplicon sequencing and $500-1500 or more for shotgun metagenomics, depending on the lab and depth of sequencing. NGS is currently best for complex, chronic, or high-value questions - rather than every otitis or diarrhea case.
Regulatory and validation landscape is evolving.
For animal health products, regulators will increasingly ask how NGS data were generated, controlled, and interpreted. This is an opportunity, but also a compliance challenge.
Where Is This Going Next?
Looking forward, I expect NGS in veterinary microbiome work to move in three directions:
Standardized panels and reports for specific indications. Chronic enteropathies, recurrent otitis/dermatitis, and complex exotics cases are likely to see structured NGS panels with clear interpretive frameworks rather than ad-hoc metagenomics for everything.
Integrated multi-omics decision support. Combining NGS with metabolomics, resistance gene profiling, and clinical data to support antibiotic deescalation, probiotic use, and diet changes in a traceable way.
Routine use in product development and post-market surveillance. Pet food, probiotic, and feed companies will use NGS not only to show "this product changes the microbiome," but to document safety, stability, and AMR impact over time.
Worth watching: long-read sequencing technologies like Oxford Nanopore and PacBio are gaining traction, especially for real-time and field applications. We are also seeing more work on equine microbiome, which I did not cover here but is an active area.
What This Means for Veterinarians and Animal Health Companies
For Clinicians:
-- I would still think of NGS as a tool for complex and recurrent cases instead of a replacement for basic diagnostics.
-- Ask for transparent methods and interpretation support when you use commercial microbiome tests.
-- Consider NGS when: culture has failed or is incomplete, you suspect biofilm or polymicrobial infections, fungi and bacteria might both be involved, or you are dealing with an exotic species where culture options are limited.
For Companies and Labs:
-- NGS can shorten development cycles, sharpen product positioning, and support more honest claims. But IF the studies are designed and analyzed properly for the method.
-- It is worth investing in study design, controls, and regulatory-aware reporting early before these datasets become central to your submissions and marketing.
At Sigma BioVet Sciences, we work in that middle space: helping translate complex NGS microbiome data into clear, defensible decisions for veterinary practices, diagnostic labs, and animal health companies.
If you are planning an animal microbiome study or considering adding NGS-based diagnostics to your offering, this is the time to ask the strategic questions (I just had to remind this to one of my clients recently - after their sequencing bills arrived!)
Until next time, stay curious and take care of your-micro-biome-self!
References
Afonso, C. L., & Afonso, A. M. (2023). Next-Generation Sequencing for the Detection of Microbial Agents in Avian Clinical Samples. Veterinary Sciences, 10(12), 690. https://doi.org/10.3390/vetsci10120690
Damerum, A., et al. (2023). Next-generation DNA sequencing offers diagnostic advantages over traditional culture testing. American Journal of Veterinary Research, 84(8). https://doi.org/10.2460/ajvr.23.03.0054
Domrazek, K., & Jurka, P. (2024). Application of Next-Generation Sequencing (NGS) Techniques for Selected Companion Animals. Animals, 14(11), 1578. https://doi.org/10.3390/ani14111578
Handl, S., et al. (2011). Massive parallel 16S rRNA gene pyrosequencing reveals highly diverse fecal bacterial and fungal communities in healthy dogs and cats. FEMS Microbiology Ecology, 76(2), 301-310. https://doi.org/10.1111/j.1574-6941.2011.01058.x
Rojas, C.A., et al. (2024). Species-level characterization of the core microbiome in healthy dogs using full-length 16S rRNA gene sequencing. Frontiers in Veterinary Science, 11, 1405470. https://doi.org/10.3389/fvets.2024.1405470
Santoro, D., et al. (2024). Update on the skin barrier, cutaneous microbiome and host defence peptides in canine atopic dermatitis. Veterinary Dermatology, 35(1), 5-14. https://doi.org/10.1111/vde.13215
Shah, H., et al. (2024). Decoding the gut microbiome in companion animals: impacts and innovations. Microorganisms, 12(9), 1831. https://doi.org/10.3390/microorganisms12091831
Suchodolski, J.S. (2021). Analysis of the gut microbiome in dogs and cats. Veterinary Clinical Pathology, 50(Suppl 1), 6-17. https://doi.org/10.1111/vcp.13031
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