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Breast milk serves not only as a nutritional source, but also confers immune elements to the baby and modulates the microbiome. However, while researchers know it is a highly complex fluid, containing hundreds of bioactive molecules, what dictates and alters breast milk composition is not well understood. Kelsey Johnson, a geneticist at the University of Minnesota, studies how external factors impact breast milk. In her recent study, published in Nature Communications, she used a multiomics approach to characterize breast milk composition and discovered that maternal cytomegalovirus (CMV) infection can impact breast milk composition to the point of altering infant gut microbiome composition, thereby potentially disrupting growth during the first month of life.1
Human CMV infection is highly prevalent, as the virus can be found in approximately 85 percent of the world’s population.2 During lactation, CMV locally reactivates in the mammary gland. As such, while it can be passed from mother to infant in utero, CMV is commonly transmitted via breast milk.1 This is becoming a concern, Johnson noted, as there is a growing understanding that CMV transmission through breast milk can cause serious complications for pre-term infants, including sepsis, thrombocytopenia, and long-term neurodevelopmental impairment.3
Taking advantage of her background in genomics, Johnson set out to acquire a bigger picture of how CMV activation affected breast milk composition. “Historically, most studies of human milk have looked at one or a few components of interest based on a specific maternal trait,” Johnson explained. Instead of this approach, she and her colleagues opted to harness omics technology to examine CMV-associated transcriptomic and metabolomic changes in breast milk.
The researchers first sequenced milk samples from lactating mothers at one-month postpartum, looking for CMV DNA in order to establish the presence of the virus in the samples. They then performed transcriptomic analysis on these samples, revealing 34 genes that were upregulated in CMV-positive breast milk compared to CMV-negative. Many of these genes were associated with immune response-associated pathways. Johnson further found, using publicly available single-cell RNA-sequencing data, that these 34 upregulated genes tended to be more highly expressed in immune cells. Perhaps unsurprisingly, CMV-positive milk samples showed a higher estimated proportion of immune cells compared to CMV-negative milk.
Transcriptomic analysis also showed that IDO1, the gene encoding indoleamine 2,3-dioxygenase (IDO) was upregulated. IDO is the rate-limiting enzyme in the tryptophan-to-kynurenine metabolic pathway. Accordingly, metabolomic analysis surveying 458 different metabolites revealed two molecules showing increased abundance in CMV-positive breast milk samples: kynurenine and its metabolite kynurenic acid.
Johnson is not surprised that CMV presence in breast milk may be associated with the IDO1 tryptophan-to-kynurenine metabolic pathway, given that the immune response to CMV in human cells and primary tissues also features the activation of this pathway. She does find it fascinating though, since this pathway may modulate the infant gut. “Bifidobacteria utilization of tryptophan in the infant gut has positive effects on microbiome development and infant immune state. We were not able to interrogate those pathways in this study, but the presence of these differences is interesting.”
This study is a really good example of where the field is moving, as something that helps us determine what questions we should be asking next.
—Britt Goods, Dartmouth University
Finally, Johnson and her colleagues obtained fecal metagenomic data from full-term infants to investigate potential alterations to the gut microbiome stemming from CMV-containing breast milk. They found that infants exposed to CMV-positive milk had less abundance of Bifidobacterium species and higher abundances of Clostridium tertium. Other researchers have shown both of these features to be linked to adverse health outcomes in infants.4,5
Johnson cautions that further investigation is required to gauge the effect of CMV infection on breast milk over longer time periods, to look at variation between individuals, and differences across populations. “[These questions] definitely motivate my work and what I am thinking about in the future,” Johnson said. “There are things that vary a lot between individuals, such as fatty acids—which are impacted by genetics and diet and do not appear to be as tightly regulated by the mammary gland. Everybody also has a different immune profile depending on their historical exposure or vaccination record.”
More importantly, Johnson’s study highlights the presence of variation in human milk, the ability to characterize it through a multiomics approach, and how this variation impacts healthy infant development. According to Britt Goods, a biological engineer at Dartmouth College who was not associated with this study, “one of the most impressive things about this study…was all of the different data types they were able to generate and all of the different analyses they were able to do. It may sound really simple, but it is really challenging to pull that off effectively in a statistically powered manner.” Goods feels that breast milk research is still in its infancy, and that the scientific community needs to better understand it before trying to apply that knowledge to the clinic: “We do not know, for example, how changes in protein composition could be linked to alterations on the infant side. This study is a really good example of where the field is moving, as something that helps us determine what questions we should be asking next.”
For Johnson, those next steps include looking at a pre-term infant cohort to see if they replicate the results observed in this study and examining if differences in milk composition are linked to positive or negative clinical outcomes in pre-term infants. She is also interested in finding genetic variants associated with the milk metabolome that could hopefully help predict milk composition. “There are many cohorts where we do not have milk samples or milk metabolomics data, but we have genetic information,” she points out.
References
1. Johnson KE, et al. Human cytomegalovirus in breast milk is associated with milk composition and the infant gut microbiome and growth. Nat Commun. 2024;15:6216.
2. Zuhair M, et al. Estimation of the worldwide seroprevalence of cytomegalovirus: A systematic review and meta-analysis. Rev Med Virol. 2019;29(3):e2034.
3. Osterholm EA, Schleiss MR. Impact of breast milk-acquired cytomegalovirus infection in premature infants: Pathogenesis, prevention, and clinical consequences? Rev Med Virol. 2020;30(6):1-11.
4. Henrick BM, et al. Bifidobacteria-mediated immune system imprinting early in life. Cell, 2021;184(15):3884-3898.e11.
5. Kiu R, et al. Preterm infant-associated Clostridium tertium, Clostridium cadaveris, and Clostridium paraputrificum strains: Genomic and evolutionary insights. Genome Biol Evol. 2017;9(10):2707-2714.