A South Dakota State University graduate student was recently honored for his research on pork by the American Society for Nutrition. Samitinjaya Dhakal received the Graduate Student Research Award — one of the highest possible honors for nutrition science graduate students — for his research “Effects of Lean Pork on Microbiota and Microbial-Metabolite Trimethylamine-N-Oxide: A Randomized Controlled Non-Inferiority Feeding Trial Based on Dietary Guidelines for Americans.”
Dhakal competed against 216 national and international graduate students including researchers from Cornell University, the University of Michigan and Purdue University, among others. Dhakal's research was also recognized through ASN's 2022 Emerging Leaders in Nutrition Science program designed for bright, upcoming doctoral and postdoctoral researchers.
Dhakal says his interest in nutritional health research began after completing his undergraduate degree.
"Pharmaceutical sciences is an important health science discipline that is designed to be treatment focused rather than prevention," he says. "Most of the modern medicines alleviate symptoms; very few medicines are made to cure diseases. When I explored more, after my undergrad, I found nutrition science to be an important field within the health science umbrella, where the focus is more on disease prevention. The field with such promise, yet has many ambiguities creating exciting prospects for future research."
In nutritional sciences, there are a lot of unsubstantiated concepts.
"Somedays, you hear fat is bad. Somedays, you hear carbohydrates are bad. Somedays, you hear butter is bad. Somedays, you hear margarine is bad," says Moul Dey, professor in the School of Health and Consumer Sciences and Dhakal's adviser. "It’s almost impossible for the consumer to discern what is real or what is good for them."
Dey says the challenge in nutritional concepts is that they are difficult to validate to the point where they are absolutely right or absolutely wrong about a food. Layers of evidence must accumulate from different research approaches. Some foods work for some people and not for others.
"Based on observational data alone we cannot know for sure what foods are health promoting and what foods are unhealthy," Dey says.
Evidence shows that things like ultra-processed foods, including processed red meat, are not health promoting. But what about fresh, lean, red meat? The notion has long been that red meat can lead to an increased risk for heart disease due to its higher saturated fat content. But red meat is also a good source of protein and micronutrients. So, was fresh, minimally processed, lean, red meat worse than poultry for omnivorous populations?
"That was the question we asked in this study, using a specific context," Dey says.
In the United States, over 95% of the population practice animal protein-based lifestyles. Americans overwhelmingly eat meat—red meat especially. Moving away from that protein, regardless of its perceived dangers, isn't likely to happen for most people. Further, as Dey explains, labeling something as harmful without sound evidence could potentially drive people to more harmful foods.
Searching for the truth behind red meat
Dey and Dhakal's research project began in 2018 through funding from the National Pork Board and the National Institute of Food and Agriculture. The study investigated Trimethylamine-N-Oxide (TMAO), a microbiota-dependent and primarily animal-protein-derived proatherogenic metabolite — a newer biomarker of atherosclerosis and a heart disease risk factor. According to Dey, the belief is that TMAO levels rise due to consuming red meat because of its precursors. However, any animal-sourced protein has TMAO precursors, Dey says.
"A study from Cornell University reported individuals' blood TMAO levels spiked the highest after consuming fish—much higher than that after consuming beef," Dey says. "However, we are told fish is healthy, but red meat has always had a negative connotation — especially considering its association with TMAO. You can see the ambiguity."
Gold standard of studies
The study's hypothesis was "pork minimally impacts the gut microbiota and microbiota-dependent TMAO response in omnivores," as outlined in their recently published research article in the Molecular Nutrition and Food Research journal, sharing co-authorship with Zahra Moazzami and Cydne Perry.
After screening almost 100 people for eligibility, the research team recruited 38 Brookings, South Dakota area participants, 50 years and older and at higher risk for atherosclerosis, to take part in a randomized, controlled, crossover feeding trial. Participants consumed their daily meals, snacks and drinks provided by the research team. Meals were prepared from scratch by dietetics and hospitality management students following the 2015–2020 Dietary Guidelines for Americans.
"Despite being around for over 40 years, our study is among a handful of research that have tested the U.S. Guidelines diet in a registered controlled feeding clinical trial," Dhakal says.
The researchers chose pork tenderloin — a red meat and the most popular animal protein worldwide — and chicken breast as the two primary proteins for the meals. After roasting and defatting, both have comparable calories and macronutrients, Dhakal notes. All other food was precisely portioned and matched for each participants to ensure consistency.
"Since poultry is widely recommended as heart healthier among animal proteins, chicken served as an active control for pork," per the study.
The researchers collected blood, buccal swabs, urine and stool samples four times during the trial to assess targeted and untargeted metabolites as well as metagenomic changes before and after the consumption of pork or chicken.
"It was a lot of work, but doing it this way helps minimize confounding effects, thus producing highest levels of evidence feasible in human nutrition research," Dey says. "We are indebted to our study participants who complied well with our strict protocols; many of them joined the study because they wanted to help academic research."
Pork not inferior to chicken
The feeding portion of this study took place in 2018-2019. Nutriomics assays and data analysis took another two years to complete.
At SDSU, Dhakal took courses in biostatistics, R programming and bioinformatics, which prepared him to analyze the massive data set of more than 350 biochemical changes in blood and more than 2,000 taxa of gut bacteria in hundreds of collected samples and replicates.
The data showed that substituting pork for chicken did not elicit a stronger TMAO response.
"What we found is that pork is statistically not inferior to chicken for TMAO," Dhakal says. "Its impact on TMAO precursor compounds, markers of lipid metabolism, oxidative stress and blood pressure were also not different."
"Instead of trying to say this is better and this is worse, we're trying to say that they're not really different using a non-inferiority trial design," Dey says. "That is another very new approach to look at dietary impacts on health; the concept however existed in pharmaceutical world for a long time to compare generics with brand name products."
As Dey and Dhakal noted in the study, a clear understanding of TMAO interaction is necessary in the context of popular dietary protein sources — something that had not widely been tested in a controlled feeding setting using the dietary guidelines.
More data from the trial remains to be analyzed and published, Dhakal says. Future avenues from this research could compare plant-based proteins with animal proteins.
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