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Low-fat, reduced-fat, whole-fat—we talk about milk fat as if it were a singular ingredient, when milk fat is actually made up of several thousand different fats. Mammalian milk fat is, in fact, the most complex lipid in nature [1]. A new research field, called lipidomics, allows researchers to quantify this complexity, by identifying and measuring all the thousands of fats at once.

For the first time, this systemic approach was applied to milk fat from multiple mammalian species, including humans, pigs, and cows [2]. Just as genomic comparisons across species can identify conserved genetic traits and those that are unique to a species, comparing milk lipidomes can uncover fats shared across mammalian lineages as well as species-specific patterns in milk lipid composition.

The study focused on triglycerides (TGs), which make up 98% of milk lipids. TGs are comprised of three fatty acids attached to a glycerol backbone. Their diversity comes from the combinations of fatty acids, of which there are hundreds [1]. From seven mammals—humans, rhesus macaque, crab-eating macaque, pig, cow, domestic yak, and goat—the researchers detected 472 distinct types of fat [2]. Eighty-five percent of these fats demonstrated intensity differences among species. For example, cow milk had more TGs made up of long-chain monounsaturated fatty acids, human milk had more TGs with medium- and long-chain polyunsaturated fatty acids (LCPUFA), and pig milk, which was the most distinct, had more TGs made from LCPUFA and very-long-chain PUFA (VLCPUFA) [2].

The researchers suggested that the higher LCPUFA and VLCPUFA in pig milk could reflect the need to pass on essential nutrients in a relatively shorter lactation period (pigs usually nurse for just over a month, whereas cows nurse for up to 10 months, and humans can nurse for several years) [2]. In humans, however, higher quantities of LCPUFA and a distinct lipidome profile from the two other primates included in the study were hypothesized to represent an adaptation to increased requirements for neonatal brain growth. The authors go on to argue that their results “show that after the human-monkey species’ divergence approximately 30 million years ago, human ancestors started to produce milk with a higher abundance of [TGs] containing long-chain fatty acids with high levels of unsaturation” [2].

This proposition fits nicely with our understanding of fatty acids and brain growth. In mammals, brain growth is associated with increased incorporation of LCPUFA, particularly omega-3 LCPUFA such as docosahexaenoic acid (DHA), in neural tissues. The faster the brain is growing, the more LCPUFA it is incorporating [3]. Humans, relative to other mammals (including other non-human primates), have a unique pattern of postnatal brain growth; they continue to grow their brain at the fetal rate of growth for the first 18 months of life [4]. All of this suggests that human infants, relative to other primates and mammals, would have an increased need for milk LCPUFA, which would be reflected in increased production of milk LCPUFA by human mothers.

Unfortunately, the lipidome study lacks sufficient data to support this evolutionary hypothesis. Fatty acids are the most variable milk nutrient, particularly LCPUFA. The mammary gland is unable to make LCPUFA; milk LCPUFA represent LCPUFA from the mother’s current diet, LCPUFA from maternal fat stores (representing the mother’s past diet), or synthesis from precursor fatty acids that come from the maternal diet [3]. The lipidome study included only 19 human milk samples from two human populations (Russian and Chinese) and did not control for or discuss their results in the context of diet.

The same can be said for their non-human primate data, represented by only two milk samples from each of the two monkey species. It is not specified, but one can assume that these are captive-living monkeys and could be consuming diets that differ from their wild counterparts in the types and quantities of fatty acids. As such, they are unlikely to represent “monkey milk fatty acid patterns” in a way that would permit discussions of evolutionary divergence in milk composition.

My colleague Richard Bazinet and I previously demonstrated [5] that captive and wild-living primates have distinct milk fatty acid profiles due to differences in preformed sources of LCPUFA in their diet, the same phenomenon seen in cross-cultural studies of human milk fatty acids [6]. Captive-living primates are often fed Monkey Chow, which can contain soy, corn, or vegetable oils as well as dried fish meal, all of which are sources of LCPUFA that would be lacking in a wild diet. As a result, many of the non-human primate milks we studied exceeded human milk in the proportion of LCPUFA, even DHA. Contrary to the lipidome study, we found that human milk fatty acid profiles fit well within a larger anthropoid (i.e., monkeys and apes) pattern rather than demonstrating a distinct composition [5].

In addition to diet, there are several known genetic variants that can influence the concentration of milk LCPUFA, such as the FADS1 and FADS2 genes, which code for enzymes that influence PUFA metabolism [7]. It was surprising that both maternal diet and genetic variants were mentioned in the lipidome paper’s introduction as factors that could influence milk fat composition but were not discussed in the context of the results.

Milk fat is the most complex lipid in nature. And lipids, because of their interaction with maternal factors like diet, might be the most complex nutrient to study in milk. Lipidomics, when integrated with maternal dietary and genetic data, could be the ideal way to capture the complexity of milk fat composition and investigate shared and unique features across mammalian milks.

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Dentists have plenty to do these days. During the pandemic, for weeks and months at a time, countries have put in place policies that have postponed many a dental check-up. Probably millions. Meanwhile, forced to stay home, people’s diets have shifted. One analysis of the Brisighella Heart Study cohort found that participants ate more yogurt and drank more milk than usual during Italy’s February–April 2020 lockdown. They also guzzled more sugars and sweets [1]. While no dentist expects the extra sugar and sweets to make their job any easier, the elevated yogurt and milk intake just might, depending that is, on whether individuals consumed dairy milk products or plant-based alternatives.

There is growing evidence that the distinction matters for tooth health. Milk is thought to protect teeth against the formation of caries. However, the emerging data on various forms of plant-based milk alternatives is not so positive. Some findings and patient case studies have prompted dentists to issue warnings to one another in the correspondence pages of professional journals [2].

These concerns emerge in the context of a long history of research into the causes of dental caries. Since 1890, micro-organisms able to ferment sugar have been suspected of playing a role in the formation of caries [3]. Even though milk contains moderate amounts of lactose, it is not thought to damage teeth because its other components have a protective effect. Caries develop when enamel “demineralizes” (or dissolves, such that calcium and phosphate leak out). Thus, conditions are primed for demineralization when the mouth’s pH dips below a threshold—unless a lot of calcium and phosphate are already present in solution [3]. (When immersed in these ions, enamel in fact has been shown to not start dissolving even when the pH drops to 2.5 [4].) The calcium and phosphate content of milk and other dairy products thus gets in the way of potential harm caused by a reduction in pH from the fermentation of lactose. Moreover, milk proteins have an additional teeth-protecting effect. They form a layer over enamel that interferes with the growth of troublesome biofilms.

Nearly 20 years ago, a World Health Organization report gathered the available evidence into a report on the effects of dairy on dental health, and concluded a possible decreased risk of dental caries associated with milk consumption [5]. In the years since that WHO report, writes one recent review of the subject, “observational epidemiological studies have adjusted for potential confounders and have reported that milk consumption is associated with lower caries experience or incidence” [6].

Studies to this effect are numerous. For example, one paper published this year reports an assessment of caries in children as young as three years old. The children in question were attending kindergarten across Poland’s 16 provinces. The study found clear differences in the number and severity of caries among the children who were only given milk or water to drink before bed, and those who were allowed other (sugary) beverages [7].

Yet, research findings such of this are rare for plant-based milk alternatives. More often, warnings emerge out of them. From oats to almonds, milk alternatives are becoming increasingly popular. The constituents of these non-dairy options tend to naturally lack many of the vitamins and minerals in plain milk. Hence, calcium and vitamin D are usually added by manufacturers, though the same cannot be said for iodine—present in real milk but not in most plant-based alternatives—as one compositional analysis recently reported [8]. Iodine is needed for the body to make thyroid hormones, and is important in neurological function.

To be sure, adding calcium to plant-based versions of milk could help to quench the effect of fermented “free” sugars on tooth enamel. However, dentists’ main concern with these products is less obvious, as it does not taste sweet at all. Some of these plant-based alternatives contain starchy carbohydrates that have been associated with the formation of caries. The oats in oat milk, for example, may be broken down to form maltose, a comfortable staple of the plaque bacteria that contributes to caries. This means that even when consumers opt for apparently healthy “unsweetened” versions of some plant-based milk alternatives, they are often likely to be selecting products that are less good for their teeth than plain old milk.

The question now is what to do with dentists’ concerns. Where government advice communicates milk as being promoting of oral health, some kind of specification that this refers to dairy milks would be helpful. There has been some discussion of the dental risks of starchy foods in New Zealand [9] but little elsewhere.

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No one likes having a sneezing fit due to seasonal allergies or struggling to breathe during an asthma attack. It turns out our propensity to such allergic and autoimmune reactions may come down to what’s in our gut—or rather, what was there when we were infants. A new study finds that whether a particular bacterium, Bifidobacterium longum subspecies infantis (B. infantis), is present in infant guts influences early immune development and could thus reduce the risk of allergic and autoimmune conditions later in life. The infant gut microbiome has been shown to play a particularly crucial role in the development of the immune system. An abnormal early gut microbiome is associated with immune dysregulation, which can lead to several disorders including colic, asthma, allergies, type-1 diabetes, and Crohn’s disease [1-6].

Even though the gut microbiome as a whole has important effects on the immune system, identifying the specific microbes involved has been a challenge. One way to identify beneficial gut bacteria may be to see which ones can utilize sugars in human milk, known as human milk oligosaccharides (HMOs). Breastfeeding is known to help develop better immune-microbe relationships, and HMOs appear to play an important role in promoting the growth of beneficial gut microbes.

When researchers looked for bacteria adapted to metabolizing HMOs, they identified B. infantis, and the EVC001 strain of B. infantis harbors all fully functional HMO-utilization genes [7,8]. Studies have shown that some B. infantis strains, such as EVC001, are able to stably and persistently colonize and dominate the intestinal microbiome of breastfed infants, leading to reductions in a fecal marker of intestinal inflammation [9,10]. Previous work has also shown that B. infantis is commonly found in breastfed infants in countries such as Bangladesh and Malawi where incidence of immune-mediated disorders is low [11,12]. In contrast, it’s rarely found in Europe and North America [13,14].

“It’s very compelling that the countries that still have this strain of bacteria don’t have the levels of autoimmune and allergic diseases that we do in first world nations,” says Dr. Bethany Henrick, Director of Immunology and Diagnostics at Evolve BioSystems. “I think the reason we lost it is unclear, but putting it back does not seem to do any harm,” says Henrick.

Along with Dr. Petter Brodin of the Karolinska Institute and other colleagues, Henrick led a new study investigating the effects of B. infantis on infant immune development [15]. They found that B. infantis was associated with immune changes that help regulate inflammation and reduce the risk of allergic and autoimmune conditions. “We’re showing in this paper that one of the consequences of getting it back into babies is proper immune programming and moving away from autoimmune and allergic phenotypes,” she says.

Henrick previously found that infants fed B. infantis EVC001 had significantly less enteric inflammation or inflammation in the gut [9]. “That was pretty profound, and as an immunologist I felt like that could impact the development of or programming of the immune system,” says Henrick. “Research in the past five to ten years has shown that the adaptive immune system in particular is programmed during the first 100 days of life,” she says.

In the new study, Henrick and her colleagues examined immune development in 208 infants, evaluating bifidobacterial species and other microbes expressing HMO-utilization genes. They also assessed the beneficial effects of microbes expressing HMO-utilization genes in a second cohort of 40 breastfed infants in California, with half receiving B. infantis EVC001 and the other half given no supplementation.

“There was this ‘aha!’ moment where we found stark differences between the kids that were colonized with B. infantis EVC001 compared with the controls,” says Henrick. A lack of Bifidobacterium and depletion of genes required for HMO utilization were associated with increased markers of both systemic and intestinal inflammation and immune dysregulation during the first months of life. In contrast, feeding infants B. infantis EVC001 increased levels of regulatory molecules known as cytokines and decreased levels of pro-inflammatory cytokines. B. infantis EVC001 metabolites also appeared to shift human immune cells in a way that favored immune and inflammatory regulation and away from the development of autoimmune and allergic diseases. “Putting B. infantis back into a baby creates a bimodal shift in the cytokines and in the immune cell imprinting, and we have never seen that before with other bacteria,” says Henrick.

Henrick is planning follow-up studies with collaborators to further investigate the impact of putting B. infantis back in infant guts, including looking specifically at its potential effects on preventing or modulating eczema or type 1 diabetes. She is also planning to test the effects of reintroducing B. infantis in larger groups of infants followed up over longer time periods.

These results show how a specific bacterium, B. infantis EVC001, can influence the programming of immune cells early in a breastfed infant’s life away from pro-inflammatory responses associated with immune-related conditions while producing regulatory cells that improve the body’s ability to control inflammation. “Our data do provide a very strong mechanism of action that we can skew the immune system away from cell subtypes that are indicative or strongly associated with the development of autoimmune and allergic diseases in kids, so that we’re putting these kids on the proper immune trajectory,” says Henrick.

References

The thought of maggots, fungus, and mites infesting your cheese might make you feel queasy, but researchers are looking into how these unconventional cheese-making methods might actually release peptides, or amino acid sequences, that could be beneficial for your health. In a new study, scientists at the University of California, Davis, profiled the array of peptides found in four particularly pungent cheeses and discovered a huge diversity of peptides—between 2900 and 4700 per cheese [1].

“We selected a few unique cheeses from Europe that had different microorganisms living on them that can contribute to protein breakdown,” says Randall Robinson, a Postdoctoral Fellow who authored the new paper. “We looked at what peptides were formed and their potential bioactivities.”

Milk contains a suite of complex proteins, and during cheese processing and ripening, acids and enzymes break down these large proteins into smaller peptides, or short amino acid sequences similar to proteins but with different functions. Some of these peptides have been found to be bioactive, meaning they can have antihypertensive, antibacterial, or anti-inflammatory properties that can be beneficial for your health [2-6].

“When cheese is made, the milk proteins get coagulated—there’s some protein breakdown,” says Robinson. “Then, during the ripening process, the proteins can be digested a little bit more and potentially form more bioactive peptides. Its something that people haven’t looked at in a lot of types of cheeses.”

Previous studies have looked at the peptide profiles of more commonplace cheeses such as Emmental, Cheddar, and Manchego [7-9], but this time the research team wanted to study a few cheeses—Casu Marzu, Mimolette, Stilton, and Taleggio—with unconventional preparation methods.

“Casu Marzu is the famous one because it’s illegal in a lot of countries,” explains Robinson. “It has fly eggs on it. They hatch and the fly maggots start eating the cheese and digesting it.” The Italian sheep milk cheese is made from Pecorino that’s been exposed to Piophila casei maggots, which after two to three months, make the cheese soft and creamy.

Mimolette, a bovine cheese from France, was another polarizing cheese. The hard cheese has mites added to its surface, and the cheese is banned in the United States. Stilton is a bovine cheese from the United Kingdom, and its interior is inoculated with the mold Penicillium roqueforti, giving it a sharp, distinctive blue cheese taste. Taleggio is an Italian cheese ripened in humid conditions and rind-washed with saltwater, so a unique set of yeasts and molds flourish on the cheese. “It’s really soft so it almost melts at room temperature,” says Robinson. “The taste wasn’t too bad if you like cheese. Stilton was super strong. I didn’t like that one.”

In addition to figuring out what peptides could be found in the cheeses, the research team was also interested in comparing the peptide profiles from both the center and the rind of the cheeses—something that previous studies hadn’t yet looked at. Because the interior and exterior of the cheeses are exposed to very different environments and enzymatic activity, they would have differences in the peptides released.

For the study, Robinson took samples from both the inside and outside of the cheeses and separated the peptides from the proteins and fats. The peptides were separated using liquid chromatography, and the amino acid sequences were analyzed using mass spectrometry [1].

What the research team found was a huge diversity of peptides: 2933 for Casu Marzu, 3520 for Taleggio, 3408 for Stilton, and 4701 for Mimolette. Previous research that used older technology isolated fewer than 200 peptides in their respective studies [7-12], demonstrating that the peptide ensembles found in cheese are much more diverse and complex than previously thought. Among the four cheeses, the team found 111 bioactive peptide sequences, with Mimolette containing the highest number at 88 [1]. The majority of these bioactive peptides were found to be antimicrobial or potentially antihypertensive.

The study also quantified the differences of peptidomes on the rinds versus the insides of the cheeses and found drastic differences. In Stilton, for example, only 4.3% of the total peptides were found on both the inside and outside of the cheese [1]. “In the rinds of the cheese we could see digestion of some proteins that weren’t digested very well on the interior,” says Robinson, who gave the example of β-lactoglobulin, a notoriously hard-to-digest protein. Casu Marzu, Mimolette, and Taleggio all showed more fragments of the broken down protein on the rind than the center.

“Lactoferrin was another one that was in high abundance, which is kind of interesting because it’s an antibacterial protein, and when it gets digested, some of the other peptides are also antibacterial,” says Robinson. Mimolette had a surprisingly large number of lactoferrin-derived peptides at 117, and all but one were found on the rind.

Robinson said that a major direction for future research would be to analyze the bioactivities of more dairy peptides. Out of the thousands of peptides identified in his study, less than 5% had been studied for potential bioactivity. “There’s this huge collection of peptides where we don’t really know what they do yet, but they could be influencing our health in ways that we don’t know about,” he says. For peptides that have known health benefits, more research needs to be done to determine how and in what quantities they could be consumed in order to confer a possible health benefit. Studying the diversity of dairy peptides is still in its infancy, and the field has exciting potential when it comes discovering possible health benefits.

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