Advancing Diagnostics Evidence Feed as a Fomite

Animal gut microbial populations can be dynamic, and Salmonella populations are diverse, with multiple serovars co-existing. Anitox collaborated with Dr. Nikki Shariat and the University of Georgia’s Poultry Research and Diagnostic Center on the study, Animal feed containing diverse populations of Salmonella. CRISPR technology was used in this study to determine if multiple serovars can co-exist in Salmonella-positive animal feed.

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What can Advancing Diagnostics tell us about Feed as a Fomite?

CRISPR technology has been used previously to identify serovars isolated through traditional methodology in samples from commercial feed mills. Traditional Salmonella recovery from feed takes many steps and multiple days and is highly dependent on the human selection of a single colony to determine a serovar.

Advancing diagnostics such as CRISPR prompt us to wonder:

• Could we shorten the method or garner more information from the analysis?
• What if, instead of getting a single serotype from a Salmonella-positive result, you got three?

More information is crucial in an industry that is feeling more pressure to implement Salmonella reduction strategies pre-harvest. Knowing what serotypes are present in feed will illuminate how Salmonella populations survive throughout the production chain.

How Do You Typically Find Salmonella in Feed?

Conventional Salmonella isolation from feed is a time-consuming multistep process that typically indicates the presence of a single serotype.

• Pre-enrichment – Feed is added to a buffered non-selective media and incubated for 24 hours, creating the perfect environment for all of the microbes within the feed to grow.
• Enrichment – Samples from the incubated pre-enriched feed samples are added to selective broths and incubated for another 24 hours. Selective broths are tailored to provide for the specific nutrient profiles required by Salmonella. This allows for Salmonella to grow uninhibited by background microorganisms.
• Culture Growth on Selective agar – Sterile loops are used to collect a very small liquid sample from the incubated enrichment broths. These agars cater to the nutrient needs and capitalize on the biochemical properties of Salmonella to confirm its presence within the enriched sample.
• Culture Growth on Non-selective Agar – One Salmonella colony is removed from a positive selective agar plate and streaked onto a nonselective agar plate. The goal here is to grow an entire healthy culture plate from one single colony. 
• Serotyping and Molecular Identification – Colonies are removed from the nonselective agar Salmonella culture plate and exposed to different serotype antigens. A reaction of the culture with any of these antigens allows us to understand what group of Salmonella exists within the feed sample. Molecular detection methods can also be performed, giving more detailed serotype information.

How were CRISPR diagnostics applied in the study?

Dr. Shariat and Anitox researchers used CRISPR analysis to evaluate Salmonella found in feed samples and throughout each step of the conventional methodology.

These experiments found that Salmonella populations within feed are more diverse than initially thought and demonstrated that populations also diversify within each step of the conventional methods. Selective media is used to allow Salmonella to outcompete other bacteria found in feed. CRISPR technology illuminated that different Salmonella serotypes also compete within samples. Some serotypes are better adapted to different types of selective media and are, therefore, more likely to be found through conventional methods.

Advantages of CRISPR Diagnostics

So, we know that feed is a fomite for Salmonella. But does it matter if we know exactly what populations are present in Salmonella-positive samples?

Not all serotypes of Salmonella have similar impacts on human or animal health. A better understanding of Salmonella populations in feed and food animal production may allow for better pre-harvest interventions. For example, if we know one serotype of Salmonella is prevalent in feed and feed mills but doesn’t typically get found until samples are collected and tested from the farms - we are able to understand that acting in the feed and feed mills better prevent the introduction of that serotype onto the farm.

Another benefit to diagnostics such as CRISPR is their ability to provide detailed information on Salmonella populations from an earlier step in the Salmonella recovery process. Getting results faster allows producers to implement action earlier and control pathogens faster.

Lastly, understanding Salmonella populations at any given time allows us to evaluate mitigation efforts better. For example, many feed producers use heat to sanitize their feed. This appears effective because samples taken at the mill are tested and found to be Salmonella free. However, a sample of feed taken from the truck or feed bin is tested and found to host a few different serotypes of Salmonella, meaning contamination occurred somewhere between sampling at the mill, truck and feed bin.

Furthermore, the population comparisons between the truck and feed bin samples show a single shared serotype. From this, we can see that feed contamination likely occurred on the truck and somewhere between the truck and the feed bin. The potential to get this detailed understanding of pathogen behavior within the feed and food production chain allows for improved corrective action. It helps us understand what kinds of interventions are most impactful.

Pathogen control interventions are expensive, and it is essential that only genuinely effective tools are used.

If you’re ready to implement feed pathogen control or if you want more information on recovering Salmonella in feed contact us. 

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