How Breastfeeding Influences Viral Colonization of the Infant Gut

  • Viruses are known to colonize the human gut during early infancy, but the details of this colonization are still unclear.
  • A new study found that the viral colonization of the early infant gut occurs in a stepwise manner starting with viruses present in gut bacteria and later followed by viruses that replicate in human cells.
  • The study found that breastfeeding influences viral colonization of the infant gut, with viruses that grow in human cells more commonly found in exclusively formula-fed babies compared with those that were fully or partially breastfed.

 

The human gut microbiome is known to contain a large number of both bacteria and viruses. Viruses are absent from the infant gut at birth but colonize shortly after and can sometimes lead to gastrointestinal disorders (1–4). By one month of age, infants can have about a billion viruses per gram of stool, which is similar to the number of viruses present in older children and adults (5–7). But there is still a lot researchers don’t know about how viruses colonize the early infant gut to form the virus microbiome, known as the virome.

“So far most of the studies analyzing the gut microbiome have focused on bacteria and there are not that many studies on viruses,” says Dr. Guanxiang Liang of the University of Pennsylvania. “A 2015 study described the virome in infants and indicated that it is very dynamic, but there’s still a lot we don’t know,” he says (2). “Some people think that babies are born with viruses and are colonized before they were born, while others think babies are colonized after birth,” says Liang. “That’s one of the questions that prompted us to analyze the infant gut virome,” he says.

In a new study, Liang and his colleagues analyzed the colonization of the infant gut by viruses (8). They found that viral colonization occurs in a stepwise manner, starting with viruses present in bacteria—known as bacteriophages—that form the predominant viral population at one month of age. By four months of age, identifiable animal viruses that can replicate in human cells become more prominent. In addition, the researchers found that breastfeeding modulates viral colonization of the infant gut, with viruses that grow in human cells more commonly found in exclusively formula-fed babies compared with those that are fully or partially breastfed.

Studying the viruses in the infant gut was no easy task. Liang says that “99.9% of them cannot be cultured, so that’s the big challenge when you’re doing microbiome studies.” And even when researchers can sequence viruses, most of them can’t be matched to known sequences in virus databases. “They could be virus or maybe they’re not, and that’s another big challenge,” he says.

To investigate how viruses colonize the infant gut, the researchers analyzed stool samples from 20 healthy infants just after birth and at one and four months of age. “We first isolated the virus-like particles and we looked at them under the microscope,” says Liang. “We were the first group to do that in newborns, and we couldn’t see many viruses there,” he says. “But within one month the number of viruses can reach 1 billion per gram of feces, and that’s a very high number,” says Liang.

Liang and his colleagues then decided to investigate the source of these viruses. “We didn’t know where they were coming from,” says Liang. The researchers purified DNA and RNA from the virus-like particles and from the whole gut microbial communities, and used sequencing to characterize the early virome.

The researchers found that the infant gut viral community is assembled in distinct steps. Newborns were found to have few to no viruses, and the researchers found that most of the billion virus particles identified at one month of age were not animal viruses that could replicate in human cells. Instead, most of these viruses were those present in bacteria.

“There are two kinds of phages, temperate phages and lytic phages,” says Liang. “Lytic” phages get their name from their ability to “lyse” cells, i.e., to cause them to break apart by rupturing their cell membranes. “Lytic phages have ongoing replication and they infect the bacteria and lyse the bacterial cell to release themselves,” he says. “Temperate phages, on the other hand, infect the bacteria and integrate themselves into the bacterial genome,” says Liang. “They don’t lyse the bacterial cells immediately, but when they receive certain signals they can become lytic again and lyse the cells,” he says.

Liang decided to investigate which types of phages were more prevalent in the infant gut. He and his colleagues found that most of the virome community of one-month-old infants came from temperate phages rather than lytic phages. “When we checked later at four months, it seemed like more and more animal cell viruses came in, as well as some lytic phages,” says Liang.

The researchers then looked into the potential influence of breastfeeding on the infant virome. “A lot of studies have looked at how breast milk inhibits the infections of some viruses,” says Liang. Studies have shown that mixed feeding of formula and human milk may be protective against viruses compared with feeding only formula (9). Epidemiological studies have also shown protective effects of breastfeeding in reducing viral gastroenteritis and infant death (10,11).

Liang’s initial study of 20 infants appeared to indicate that breastfeeding had an effect, as it was associated with a lower accumulation of human viruses in stool samples. To validate these results, the researchers analyzed stool samples taken at 3–4 months of age from an additional 125 infants. This analysis also showed a protective effect of breastfeeding, with 30% of formula-fed babies positive for viruses that infect human cells compared with 9% of babies who were fed human milk or human milk together with formula.

Both the initial and follow-up analyses used samples from infants from an urban area in the United States. To check whether the results were more broadly applicable, the researchers analyzed samples from a different cohort of 4-month-old infants from Botswana. They again found that viruses that grow in human cells were more common in exclusively formula-fed babies compared with breastfed babies. “We tested the breastfeeding association again and we found similar results, which gave us confidence about these data,” says Liang. The study thus describes both the stepwise colonization of the early infant gut by viruses as well as the protective effects of breastfeeding on this viral colonization.

Researchers are still investigating the mechanisms by which breastfeeding may help protect against viruses. Several factors in human milk are known to inhibit viral colonization, including maternal antibodies, milk sugars known as human milk oligosaccharides, and human milk proteins such as lactoferrin (12–14). Breastfeeding may also increase the abundance of gut bacteria that serve as a source of the early bacteriophages that populate the virome. “Breastfeeding can increase the abundance of Bifidobacteria and Lactobacillus, and the phages could come from these bacteria, so it’s possible it’s an indirect effect of breast milk,” says Liang.

Liang is interested in performing longer-term follow-up studies of virome colonization. “One thing we haven’t figured out is the long-term outcome for the infant,” says Liang. “We only followed this cohort for four months, and so we could track them for one or two years or even longer to look at the dynamics of the virome and its influence on health outcomes,” he says. “Another thing we could look at is preterm babies and how [premature birth] influences the virome structure,” says Liang.

 

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10. Arifeen S, Black RE, Antelman G, Baqui A, Caulfield L, Becker S. Exclusive breastfeeding reduces acute respiratory infection and diarrhea deaths among infants in Dhaka slums. Pediatrics. 2001 Oct;108(4):E67.

11. Victora CG, Smith PG, Vaughan JP, Nobre LC, Lombardi C, Teixeira AM, Fuchs SC, Moreira LB, Gigante LP, Barros FC. Infant feeding and deaths due to diarrhea. A case-control study. Am J Epidemiol. 1989 May;129(5):1032-41.

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13. Shkoporov AN, Khokhlova EV, Fitzgerald CB, Stockdale SR, Draper LA, Ross RP, Hill C. ΦCrAss001 represents the most abundant bacteriophage family in the human gut and infects Bacteroides intestinalis. Nat Commun. 2018 Nov 14;9(1):4781.

14. Turin CG, Ochoa TJ. The role of maternal breast milk in preventing infantile diarrhea in the developing world. Curr Trop Med Rep. 2014 Jun 1;1(2):97-105.

 

Contributed by
Dr. Sandeep Ravindran
Freelance Science Writer
Sandeepr.com