The Human Milk Glycome as a Defense Against Infectious Diseases

Viewpoint Cite This: ACS Infect. Dis. XXXX, XXX, XXX-XXX

pubs.acs.org/journal/aidcbc

The Human Milk Glycome as a Defense Against Infectious Diseases: Rationale, Challenges, and Opportunities Kelly M. Craft† and Steven D. Townsend*,†,‡ †

Department of Chemistry, Vanderbilt University, 7330 Stevenson Center, Nashville, Tennessee 37235, United States Institute of Chemical Biology, Vanderbilt University, 896 Preston Research Building, Nashville, Tennessee 37232, United States

‡

ABSTRACT: Each year over 3 million people die from infectious diseases with most of these deaths being poor and young children who live in low- and middle-income countries. Infectious diseases emerge for a multitude of reasons. On the social front, reasons include a breakdown of public health standards, international travel, and immigration (for financial, civil, and social reasons). At the molecular level, the modern rise of infectious diseases is tied to the juxtaposition of drug-resistant pathogens and a lack of new antimicrobials. The consequence is the possibility that humankind will return to the preantibiotic era wherein millions of people will perish from what should be trivial illnesses. Given the stakes, it is imperative that the chemistry community take leadership in delivering new antibiotic leads for clinical development. We believe this can happen through innovation in two areas. First is the development of novel chemical scaffolds to treat infections caused by multidrug-resistant pathogens. The second area, which is not exclusive to the first, is the generation of antibiotics that do not cause collateral damage to the host or the host’s microbiome. Both can be enabled through advances in chemical synthesis. It is with this general philosophy in mind that we hypothesized human milk oligosaccharides (HMOs) could serve as novel chemical scaffolds for antibacterial development. We provide herein a personal account of our laboratory’s progress toward the goal of using HMOs as a defense against infectious diseases.

■

A

BACKGROUND As their taglines often read, pharmaceutical companies are the greatest source of therapeutics in the world. Perhaps paradoxically, however, most have limited investments in the development of new antibiotics.6 Financially speaking, the return on investment for antibiotic development is poor as bacteria quickly develop resistance to small molecules. This means companies have a limited window to recoup the investment. A second issue is that new targets are often species, and even strain, specific. In the clinic, new antimicrobials are expected to feature broad-spectrum activity. This is unfortunate as highly effective antibiotics like fidaxomicin, a narrow spectrum macrolide antibiotic for Clostridium dif f icile that does not affect the host microbiome, are not competitive in a market which features broadly functioning warheads like vancomycin. Broadspectrum antimicrobials, however, predictably cause collateral damage to the host microbiome. Refreshingly, there are exceptions to this trend. Kaleido Biosciences is a new biotech company with the primary goal of developing orally available compounds that modulate the human microbiome. Excitingly, Kaleido has an interest in studying complex oligosaccharides and glycoconjugates in human gut microbiome assays that are designed to reflect healthy and diseased human bowels. At this stage, it is important to emphasize that, when it comes to clinical impact, quality is better than quantity. Going forward, it is imperative that new antimicrobials provide advances in treatment when compared to available therapies. Ideally, new compounds would function through new modes of

ntimicrobial resistance is among the most complex and concerning public health challenges facing humankind.1,2 Warnings that antibiotics are losing effectiveness due to clinical misuse and overuse have been largely ignored. Common illnesses, such as pneumonia, as well as the world’s most prevalent infectious diseases (human immunodeficiency virus (HIV), malaria, and tuberculosis) are becoming increasingly difficult to treat due to drug resistance. Infections caused specifically by antibiotic-resistant bacteria continue to challenge physicians (Table 1). The World Health Organization (WHO) has shown that rates of infection attributable to methicillinresistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecium (VRE), and fluoroquinolone-resistant Pseudomonas aeruginosa are increasing.3 In fact, more people in the United States die each year from MRSA infection than from HIV/acquired immunodeficiency syndrome (AIDS) and tuberculosis combined.4 Moreover, several antibiotic-resistant Gram-negative pathogens, including the Acinetobacter species, Escherichia coli, Klebsiella species, and multidrug-resistant (MDR) Pseudomonas aeruginosa, have emerged as significant players in human infection.5 Before initiating a discussion on what we view to be a frontier strategy in the treatment of infectious diseases, we believe it is beneficial to briefly inform the reader on the inspiration and risk parameters associated with the development of antimicrobial agents. To this end, we digress momentarily to recount various teachings from the fields of infectious disease and human milk glycobiology, both of which are relevant to our program mission. © XXXX American Chemical Society

Received: November 1, 2017

A

DOI: 10.1021/acsinfecdis.7b00209 ACS Infect. Dis. XXXX, XXX, XXX−XXX

ACS Infectious Diseases

Viewpoint

Table 1. WHO Priority Pathogens for New Antibiotic R&D7 priority level 1: critical

a

priority level 2: high

priority level 3: medium

pathogen

resistance

pathogen

resistance

pathogen

resistance

Acinetobacter baumannii Pseudomonas aeruginosa Enterobacteriaceaea

carbapenem carbapenem carbapenem, cephalosporin

Enterococcus faecium Staphylococcus aureus Helicobacter pylori Campylobacter Salmonella Neisseria gonorrhoeae

vancomycin methicillin, vancomycin clarithromycin fluoroquinolone fluoroquinolone cephalosporin fluoroquinolone

Streptococcus pneumoniae Haemophilus inf luenzae Shigella

penicillin ampicillin fluoroquinolone

Enterobacteriaceae include: Enterobacter spp., Escherichia coli, Klebsiella pneumonia, Morganella spp., Proteus spp., Providencia spp., and Serratia spp.

action.8−13 Upon analysis of the recent arsenal of drugs in the latest stages of development, it is evident that they do not advance our ability to treat infections of resistant pathogens, specifically Gram-negative species. Unfortunately, due to limited options, clinicians are repurposing once abandoned drugs.14 In our search for new scaffolds that may have new modes of action, we turned to human milk.

levels decrease. Over time, the gut microbiome achieves an adult-like composition. HMOs are believed to be a specific growth factor capable of enriching the gut flora. Breastfed infants have a microbiota rich in Bacteroides and Bifidobacteria. It has been demonstrated that some species of Bacteriodes consume long-chain HMOs via mucin-utilization pathways.33 Contrarily, a number of species of Bif idobacteria consume short-chain HMOs.34,35 One can infer from this data that long-chain HMOs serve as mimics to mucins and thus promote the growth of symbiotes, like Bacteroides, which can metabolize these molecules. Shorterchained HMOs, however, are structurally divergent from Olinked mucin-type glycans and glycoproteins. It is conceivable then that these molecules are used specifically by certain species of Bif idobacteria which do not metabolize mucins and thus would otherwise be outcompeted. In sum, HMOs select for the growth of both HMO-metabolizing Bif idobacterium species and mucin-metabolizing Bacteroides species (Table 2).36−38

■

HUMAN MILK OLIGOSACCHARIDES While breastfeeding has long had its critics, it is now widely accepted that nursing has a profoundly positive effect on the short- and long-term health of neonates.15 Under even the harshest of scenarios and even when the mother’s own nutrition is compromised, human milk provides all vitamins, nutrients, and macromolecules that are essential to the development of the child. Interestingly, the composition of macromolecules in human milk is dynamic and adapts itself to complement the neonate defense system.16−18 The host defense mechanisms of a newborn can be classified as nonspecific (innate) or specific (acquired).19−21 Nonspecific mechanisms are effective without prior exposure to a microorganism or its antigens. On the basis of the data, it appears as though human milk oligosaccharides (HMOs) are part of the nonspecific response.22−28 Historically, HMOs have been shown to inhibit the growth of a number of viral and bacterial pathogens.24−27,29 In the next two sections, we will provide brief overviews of the roles of HMOs as prebiotics and antimicrobial agents as both would be of primary interest to this community based on the WHO’s priority listing of bacterial pathogens.

Table 2. HMO-Promoted Growth of Symbiotic Bacteria symbiote

action

B. bifidum, B. longum

major strains found in breastfed infant feces can grow using HMOs as the sole carbon source metabolizes “small” oligosaccharides found in human milk major strains associated with adult gut flora do not grow efficiently on HMOs HMO use in B. f ragilis and B. thetaiotaomicron coupled to upregulation of mucin degradation pathways do not exhibit growth in the presence of HMOs do not digest complex HMOs

B. breve, B. adolescentis

B. fragilis, B. thetaiotaomicron

■

IMPACT OF THE HUMAN MILK GLYCOBIOME ON SYMBIOTIC BACTERIA Shortly after parturition, a newborn’s microbiome begins to develop as a succession of bacteria colonizes the neonatal gut. Additional factors, such as mode of delivery (cesarean vs vaginal birth) and antibiotic use, influence the gut flora.30,31 While little is known about the mechanisms that connect the variables described here to microbiota composition, it is well established that human milk and HMOs are a primary driver of healthy microbiome maturation. Human milk serves as a primary source of continued microbial inoculation as it can contain ca. 700 different species of bacteria.32 Moreover, because only ca. 1% of HMOs are absorbed into circulation, the majority reach the distal intestine where they can be metabolized by mutualistic symbiotic bacteria. Aerobic and facultative anaerobic species are favored early in life when the gut is oxygen replete. Anaerobic bacteria, such as Bif idobacteria and Bacteroides, are established as oxygen

B. ovatus, B. stercoris L. plantarum, L. acidophilus

L. reuteri, L. fermentum, S. thermophilus

metabolize neutral HMOs ferment lactose, glucose, Nacetylglucosamine, and fucose do not metabolize HMOs

reference 39

39

33 and 40 33 and 40 41 and 42

41 and 42

■

HMO-MEDIATED PATHOGEN PROTECTION It has been established that breastfed infants experience decreased instances of diarrhea, respiratory infection, urinary tract infection, ear infection, necrotizing enterocolitis (NEC), and sudden infant death syndrome (SIDS), compared to their formula-fed counterparts.15,43,44 In agreement with these findings, breastfed neonates tend to be colonized to a lesser B



Viewpoint

Table 3. HMO-Fostered Inhibition of Bacterial Pathogens bacterial species Acinetobacter baumannii Campylobacter jejuni

Candida albicans Clostridium dif f icile

Enterococcus faecium Escherichia coli

action

HMOs

inhibition of growth inhibition of adhesion to epithelial cells inhibition of inflammatory signaling decreased Campylobacter-attributable diarrhea inhibition of adhesion to epithelial cells interference with hyphal morphogenesis binding to exotoxins A (TcdA) and B (TcdB) (prevents interactions of toxin with cellular receptors)

faster vancomycin-resistant E. faecium (VRE) colonization reduction compared to non-HMO treatment interference with intracellular signals used by UPEC to cause cell damage inhibition of UPEC adhesion to epithelial cells

Streptococcus agalactiae

inhibition of EPEC adhesion to epithelial cells binding to heat-labile enterotoxin type 1 (HLT) inhibition of adhesion to epithelial cells inhibition of adhesion to epithelial cells inhibition of adhesion to epithelial cells reduction of adhesion to and internalization in pneumocytes bacteriostatic and antibiofilm (mechanism not yet determined)

Streptococcus pneumoniae

inhibition of adhesion to epithelial cells

Shigella dysenteriae

binding to Shiga toxins Stx2 and Stx1B5

Salmonella fyris

inhibition of adhesion to epithelial cells

Staphylococcus aureus

promotes growth without HMO metabolism; proposed to act as growth stimulant inhibition of biofilm inhibition of binding to HBGAs [Norovirus] inhibition of adhesion to epithelial cells [Rotavirus]

Haemophilus inf luenzae Helicobacter pylori Pseudomonas aeruginosa

Norovirus and Rotovirus

extent by infectious species such as E. coli, C. dif f, and C. jejuni. Importantly, many of these protective properties have been attributed to the HMO component of milk.45−50 For example, a study by the Donovan lab showed that HMO supplementation shorted the duration of rotavirus infection. Rotavirus is one of the leading causes of diarrhea in infants.51 Bovine milk, from which most formula is based, however, contains a negligible oligosaccharide component. Additionally, bovine milk oligosaccharides (BMOs) lack the structural complexity and diversity of HMOs.17 Consequently, formula-fed infants unfortunately do not obtain comparable oligosaccharidefostered protections as those that are breastfed. Broadly speaking, HMO-fostered protection can be broken down into two categories. The first is protection resulting from the selective metabolism of HMOs by symbiotic bacteria. Selective utilization by symbiotes, such as Bifidobacteria, affords these species a competitive edge over pathogens which cannot metabolize HMOs. In a study by the Miller lab, it was found that 0 of 10 Enterobacteriaceae strains tested, including several E. coli strains and one Shigella dysenteriae strain, were incapable of growing on the HMOs 2′-fucosyllactose (2′-FL), 6′sialyllactose (6′-SL), and lacto-N-netotetraose (LNnT). Several of these strains were, however, able to grow on galactooligosaccharides (GOS) and mono- and disaccharide HMO components.52 As a result of this selective metabolism,

reference

pooled HMOs 2′-FL other 2-linked fucosylated oligosaccharides

unpublished 55, 56, and 59

pooled HMOs

60

fucosylated single-entity HMOs (e.g., LNFP I, LNFP III) acidic single-entity HMOs (e.g., LST b and c) LNT, LNnH mixtures of fucosylated HMOs

61 and 62

acidic and neutral HMO mixtures

64−68

63

neutral and acidic single-entity HMOs (e.g., 2′-FL, 6′-SL, LNFP I and II)

high molecular weight fraction of milk acidic HMOs (e.g., 3′-SL and 6′-SL) 2′-FL and 3-FL 3′-SL and 6′-SL neutral HMOs mixtures single-entity HMOs (e.g., LNT and LNFP I) pooled HMOs low and high molecular weight milk fractions single-entity HMOs (e.g., LNT) acidic and neutral single-entity HMOs (e.g., 2′-FL, 6′-SL, LNDFH I, LNFP III) acidic and neutral low molecular weight HMOs (e.g., 3-FL and 6′-FL) pooled HMOs

69 70 71 and 72

sialylated HMOs (e.g., 3′-SL and 6′-FL)

53, 54, and 74

57 and 58

69 68 65 73

symbiotes can grow and outcompete harmful pathogens. Moreover, metabolism of HMOs results in the production of short-chain fatty acids (SCFAs). SCFAs lower the pH of the gut which further stunts the growth of many pathogenic species.46 The second protective mechanism arises from more direct interaction with pathogens. Namely, HMOs can act as antiadhesive antimicrobials by serving as soluble decoy receptors for pathogens or pathogenic virulence agents such as toxins. This ability is made possible by the resemblance of HMOs to various cell surface glycan receptors. As a result, pathogens bind to HMOs rather than to cell surface glycans thereby prohibiting the binding of pathogenic species to epithelial cells which is often the first step of infection. For instance, the potential for HMOs to protect against norovirus infection is hypothesized by Hansman and coworkers to be due to commonalities between HMO and histo-blood group antigen (HBGA) fucosylation patterns. These structural similarities would allow HMOs to act as natural decoys for the HBGA binding pocket of noroviruses.53,54 Similar findings by the Newburg lab have been seen for C. jejuni wherein α-1,2 fucosylated HMOs were able to inhibit adherence to host cell receptors.55,56 A summary of HMO-fostered protections against numerous bacterial pathogens is provided in Table 3. C



Viewpoint

nevertheless often cite this method as a shortcoming of their studies. This proclamation of limitation is more often than not followed by the authors’ lament about the lack of access to single-entity compounds and/or the prohibitively expensive nature of the few commercially available HMOs. In general, our team is against pooling milk samples for evaluation as it removes the ability to observe differences in HMO activity based on maternal phenotype. Additionally, this approach can make identifying specific HMO structure activity relationships akin to finding a needle in a very large haystack. A hallmark example of the limitations associated with using pooled milk samples can be seen in a recent study by the Bode lab which investigated the effects of HMOs on NEC in a neonatal rat model.75 Initial studies found pooled HMOs to reduce NEC incidence. Further investigation, however, revealed that this protection was unique to a single HMO, disialyllactoN-tetraose (DSLNT). Furthermore, it was found that both sialic acid residues were required to protect against NEC. In addition to issues related to procuring HMOs, an obscure issue is that the composition of human milk changes on a regular basis. While this is certainly evolutionary genius, perhaps to defend against resistance, the scientific consequence is that, once a sample has been exhausted, one will never be able to replicate the biological results observed from the initial screening! Moreover, based on the average milk donation, approximately 4−5 bacterial strains can be assayed from a single milk sample. Thus, innovative solutions are necessary to determine which HMOs, or which combinations of HMOs, are most important for anti-infective activity. These results would in turn drive innovation in both chemical and chemoenyzmatic synthesis. As carbohydrate synthesis continues to advance, more HMOs are becoming available.76−78 These HMOs are, however, generally restricted to small (tri- and tetrasaccharides) and structurally simple compounds. More specifically, these structures are generally restricted to compounds featuring minimal decorations or elongations of lactose, which is the disaccharide found at the reducing end of all HMOs. Examples include monofucosylated or sialylated lactose and lactose elongated in a linear fashion with one lacto-N-biose or Nacetyllactosamine residue. Unfortunately, these HMOs represent only a small portion of the human milk glycome which is estimated to contain well-over 200 unique structures.79 It is our hope that the high-level nature of the problem will continue to inspire novel strategies and tactics to produce complex HMOs.77,78 There exist exciting opportunities for multidisciplinary teams to learn exactly how HMOs confer beneficial effects.80 For example, the NIH has established a glycoscience common fund program to develop new methodologies and resources to study glycans. The goal of the program is to make carbohydrates, including HMOs, accessible to the broader biomedical research community. Recently, UC San Diego has initiated an amazing effort to study this topic. Prof. Lars Bode, an expert in HMO research, is leading an initiative which will endow a faculty chair in human milk research and support seed grants for collaborative projects. Hopefully, as time progresses, more initiatives will be established at UCSD and other universities to assist multidisciplinary teams in performing research on this frontier topic.

While inhibition of binding to cell surface glycans is a common mode of HMO-attributable protection, there nevertheless remain cases where the mechanism of inhibition has yet to be established. Work from our laboratory as well as work from the Bode laboratory has shown that HMOs have both bacteriostatic and antibiofilm activities against Streptococcus agalactiae (Group B Step, GBS). While the Bode laboratory has evidence which suggests that HMOs may serve as alternative substrates capable of impairing growth kinetics, a definitive mechanism of inhibition has yet to be determined.57,58 The relationship between HMOs and S. aureus remains ambiguous. An early study by the McGuire lab showed that HMOs actually stimulated the growth of S. aureus.73 The increased growth was not, however, attributable to the metabolism of HMOs. As a result, it was suggested that HMOs perhaps serve as growth stimulants. Recent work in our laboratory showed HMOs to act as antibiolfilm agents against S. aureus, although again, a mechanism for this activity has yet to be determined.

■

CHALLENGES AND FUTURE OPPORTUNITIES It was first observed in 1900 that human milk governs the composition of the infant gut flora. A century later, the community still has a poor understanding of the mode of action for antimicrobial activity. Specifically, questions remain about exactly how HMOs select for the growth of symbiotic species while also providing the means to defeat a number of pathogens (Table 4). Table 4. Key Roadblocks to Leveraging the Human Milk Glycome To Fight Infectious Diseases objective

stage

identify which pathogens are susceptible to HMOs characterize and purify anti-infective HMOs determine mechanism of antimicrobial or antivirulence activity chemically/chemoenzymatically synthesize single-entity HMOs use HMOs as dietary supplements

late early early early late

We believe the future of HMO research ultimately will involve orally administering synthetic HMOs as a new generation of antimicrobial agents or dietary supplements. In fact, a number of for-profit and nonprofit companies are pursuing this concept. For example, Sugarlogix is a biotech start-up looking to synthesize HMOs as a supplement for infant formula. Our personal view is that, depending on the pathogen, HMOs could serve as either therapeutics to treat current infections or prophylactics to prevent infection. For instance, HMO antimicrobial cocktails could be delivered to children at risk for infectious diseases. The greatest barrier to research in the field of HMO glycobiology is the limited availability of HMOs. At a basic level, there is a limited supply of donor milk accessible in the United States. Rightfully so, this product is prioritized for sick neonates who are most likely to benefit from exclusive consumption of human milk. Given the need to prioritize milk distribution, the amount of breast milk available to researchers for preliminary studies remains small. Reflective of this reality, a common theme in HMO glycobiology research remains that researchers resort to pooling together small donor milk samples in order to generate enough material for evaluation. While the use of pooled milk has helped elucidate several actions and benefits of HMOs, researchers D



■

Viewpoint

(9) Minbiole, K. P. C., Jennings, M. C., Ator, L. E., Black, J. W., Grenier, M. C., LaDow, J. E., Caran, K. L., Seifert, K., and Wuest, W. M. (2016) From antimicrobial activity to mechanism of resistance: the multifaceted role of simple quaternary ammonium compounds in bacterial eradication. Tetrahedron 72 (25), 3559−3566. (10) Jennings, M. C., Minbiole, K. P. C., and Wuest, W. M. (2015) Quaternary Ammonium Compounds: An Antimicrobial Mainstay and Platform for Innovation to Address Bacterial Resistance. ACS Infect. Dis. 1 (7), 288−303. (11) Nguyen, T. V., Blackledge, M. S., Lindsey, E. A., Minrovic, B. M., Ackart, D. F., Jeon, A. B., Obregon-Henao, A., Melander, R. J., Basaraba, R. J., and Melander, C. (2017) The Discovery of 2Aminobenzimidazoles That Sensitize Mycobacterium smegmatis and M. tuberculosis to beta-Lactam Antibiotics in a Pattern Distinct from beta-Lactamase Inhibitors. Angew. Chem., Int. Ed. 56 (14), 3940−3944. (12) Shapiro, J. A., and Wencewicz, T. A. (2017) Structure-function studies of acinetobactin analogs. Metallomics 9 (5), 463−470. (13) Wencewicz, T. A. (2016) New antibiotics from Nature’s chemical inventory. Bioorg. Med. Chem. 24 (24), 6227−6252. (14) Corona, A., and Cattaneo, D. (2017) Dosing Colistin Properly: Let’s Save ″Our Last Resort Old Drug!″. Clin. Infect. Dis. 65 (5), 870− 870. (15) Kamudoni, P., Maleta, K., Shi, Z., and Holmboe-Ottesen, G. (2015) Exclusive breastfeeding duration during the first 6 months of life is positively associated with length-for-age among infants 6−12 months old, in Mangochi district, Malawi. Eur. J. Clin. Nutr. 69 (1), 96−101. (16) Newburg, D. S. (2013) Glycobiology of human milk. Biochemistry (Moscow) 78 (7), 771−85. (17) Bode, L. (2012) Human milk oligosaccharides: every baby needs a sugar mama. Glycobiology 22 (9), 1147−62. (18) Kunz, C., Rudloff, S., Baier, W., Klein, N., and Strobel, S. (2000) Oligosaccharides in human milk: structural, functional, and metabolic aspects. Annu. Rev. Nutr. 20, 699−722. (19) Hettinga, K., van Valenberg, H., de Vries, S., Boeren, S., van Hooijdonk, T., van Arendonk, J., and Vervoort, J. (2011) The host defense proteome of human and bovine milk. PLoS One 6 (4), e19433. (20) Newburg, D. S. (1996) Oligosaccharides and glycoconjugates in human milk: their role in host defense. J. Mammary Gland Biol. Neoplasia 1 (3), 271−83. (21) Xanthou, M., Bines, J., and Walker, W. A. (1995) Human milk and intestinal host defense in newborns: an update. Adv. Pediatr. 42, 171−208. (22) Newburg, D. S., and Morelli, L. (2015) Human milk and infant intestinal mucosal glycans guide succession of the neonatal intestinal microbiota. Pediatr. Res. 77 (1−2), 115−20. (23) Yu, H., Lau, K., Thon, V., Autran, C. A., Jantscher-Krenn, E., Xue, M., Li, Y., Sugiarto, G., Qu, J., Mu, S., Ding, L., Bode, L., and Chen, X. (2014) Synthetic disialyl hexasaccharides protect neonatal rats from necrotizing enterocolitis. Angew. Chem., Int. Ed. 53 (26), 6687−91. (24) Musilova, S., Rada, V., Vlkova, E., and Bunesova, V. (2014) Beneficial effects of human milk oligosaccharides on gut microbiota. Benefic. Microbes 5 (3), 273−83. (25) Liu, B., and Newburg, D. S. (2013) Human milk glycoproteins protect infants against human pathogens. Breastfeed Med. 8 (4), 354− 62. (26) Newburg, D. S. (2009) Neonatal protection by an innate immune system of human milk consisting of oligosaccharides and glycans. J. Anim. Sci. 87 (13 Suppl), 26−34. (27) Morrow, A. L., Ruiz-Palacios, G. M., Jiang, X., and Newburg, D. S. (2005) Human-milk glycans that inhibit pathogen binding protect breast-feeding infants against infectious diarrhea. J. Nutr. 135 (5), 1304−1307. (28) Klein, N., Schwertmann, A., Peters, M., Kunz, C., and Strobel, S. (2000) Immunomodulatory effects of breast milk oligosaccharides. Adv. Exp. Med. Biol. 478, 251−259.

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Kelly M. Craft: 0000-0001-9753-2801 Steven D. Townsend: 0000-0001-5362-7235 Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS S.D.T. would like to acknowledge Vanderbilt University and the Institute of Chemical Biology for financial support. K.M.C. acknowledges support from the Vanderbilt Chemical Biology Interface (CBI) training program (T32 GM065086), the Vanderbilt Pre3 Initiative for a travel grant, and a Mitchum E. Warren, Jr. Graduate Research Fellowship.

■

ABBREVIATIONS HMO, human milk oligosaccharide; WHO, World Health Organization; MRSA, methicillin-resistant S. aureus; VRE, vancomycin-resistant E. faecium; HIV, human immunodeficiency virus; AIDS, acquired immunodeficiency syndrome; MDR, multidrug-resistant; spp, multiple species; NEC, necrotizing enterocolitis; SIDS, sudden infant death syndrome; SCFA, short-chain fatty acids; GBS, Group B Step; 2′-FL, 2′fucosyllactose; 3-FL, 3-fucosyllactose; 6′-SL, 6′-sialyllactose; LNnT, lacto-N-netotetraose; GOS, galactooligosaccharides; LNFP I, lacto-N-fucopentaose I; LNFP III, lacto-N-fucopentaose III; LST b, sialyl-lacto-N-tetraose b; LST c, sialyl-lacto-Ntetraose c; LNT, lacto-N-tetraose; LNnH, lacto-N-neohexaose; UPEC, uropathogenic E. coli; EPEC, enteropathogenic E. coli; LNFP II, lacto-N-fucopentaose II; 3′-SL, 3′-sialyllactose; LNDFH I, lacto-N-difucohexaose I; HBGA, histo-blood group antigens; DSLNT, disialyllacto-N-tetraose

■

REFERENCES

(1) Melander, R. J., and Melander, C. (2017) The Challenge of Overcoming Antibiotic Resistance: An Adjuvant Approach? ACS Infect. Dis. 3 (8), 559−563. (2) Rossiter, S. E., Fletcher, M. H., and Wuest, W. M. (2017) Natural Products as Platforms To Overcome Antibiotic Resistance. Chem. Rev. 117 (19), 12415−12474. (3) Loomba, P. S., Taneja, J., and Mishra, B. (2010) Methicillin and Vancomycin Resistant S. aureus in Hospitalized Patients. J. Glob. Infect. Dis. 2 (3), 275−83. (4) Klein, E., Smith, D. L., and Laxminarayan, R. (2007) Hospitalizations and deaths caused by methicillin-resistant Staphylococcus aureus, United States, 1999−2005. Emerging Infect. Dis. 13 (12), 1840−6. (5) Sudan, S., Cherian, C., Goldstein, E., Hines, D. W., Glenn, G., and Chopra, T. (2015) Trends in Carbapenem Susceptibility of Acinetobacter baumannii, Pseudomonas aeruginosa and Klebsiella pneumoniae Isolates From Long-Term Acute Care Hospitals Across the United States-An Analysis of Antibiograms. Open Forum Infect. Dis. 2 (Suppl 1), 177. (6) Fernandes, P., and Martens, E. (2017) Antibiotics in late clinical development. Biochem. Pharmacol. 133, 152−163. (7) WHO. (February 27, 2017) Global Priority List of AntibioticResistant Bacteria to Guide Research, Discovery, and Development of New Antibiotics, WHO, Geneva. (8) Jennings, M. C., Forman, M. E., Duggan, S. M., Minbiole, K. P. C., and Wuest, W. M. (2017) Efflux Pumps Might Not Be the Major Drivers of QAC Resistance in Methicillin-Resistant Staphylococcus aureus. ChemBioChem 18 (16), 1573−1577. E



Viewpoint

(29) Newburg, D. S., and Walker, W. A. (2007) Protection of the neonate by the innate immune system of developing gut and of human milk. Pediatr. Res. 61 (1), 2−8. (30) Gasparrini, A. J., Crofts, T. S., Gibson, M. K., Tarr, P. I., Warner, B. B., and Dantas, G. (2016) Antibiotic perturbation of the preterm infant gut microbiome and resistome. Gut Microbes 7, 443−449. (31) Praveen, P., Jordan, F., Priami, C., and Morine, M. J. (2015) The role of breast-feeding in infant immune system: a systems perspective on the intestinal microbiome. Microbiome 3, 41. (32) Cabrera-Rubio, R., Collado, M. C., Laitinen, K., Salminen, S., Isolauri, E., and Mira, A. (2012) The human milk microbiome changes over lactation and is shaped by maternal weight and mode of delivery. Am. J. Clin. Nutr. 96 (3), 544−551. (33) Marcobal, A., Barboza, M., Sonnenburg, E. D., Pudlo, N., Martens, E. C., Desai, P., Lebrilla, C. B., Weimer, B. C., Mills, D. A., German, J. B., and Sonnenburg, J. L. (2011) Bacteroides in the infant gut consume milk oligosaccharides via mucus-utilization pathways. Cell Host Microbe 10 (5), 507−14. (34) LoCascio, R. G., Ninonuevo, M. R., Freeman, S. L., Sela, D. A., Grimm, R., Lebrilla, C. B., Mills, D. A., and German, J. B. (2007) Glycoprofiling of bifidobacterial consumption of human milk oligosaccharides demonstrates strain specific, preferential consumption of small chain glycans secreted in early human lactation. J. Agric. Food Chem. 55 (22), 8914−9. (35) Sela, D. A., Chapman, J., Adeuya, A., Kim, J. H., Chen, F., Whitehead, T. R., Lapidus, A., Rokhsar, D. S., Lebrilla, C. B., German, J. B., Price, N. P., Richardson, P. M., and Mills, D. A. (2008) The genome sequence of Bifidobacterium longum subsp. infantis reveals adaptations for milk utilization within the infant microbiome. Proc. Natl. Acad. Sci. U. S. A. 105 (48), 18964−9. (36) Bashiardes, S., Thaiss, C. A., and Elinav, E. (2016) It’s in the Milk: Feeding the Microbiome to Promote Infant Growth. Cell Metab. 23 (3), 393−4. (37) DiBartolomeo, M. E., and Claud, E. C. (2016) The Developing Microbiome of the Preterm Infant. Clin. Ther. 38 (4), 733−9. (38) Lim, E. S., Wang, D., and Holtz, L. R. (2016) The Bacterial Microbiome and Virome Milestones of Infant Development. Trends Microbiol. 24, 801. (39) Yu, Z. T., Chen, C., and Newburg, D. S. (2013) Utilization of major fucosylated and sialylated human milk oligosaccharides by isolated human gut microbes. Glycobiology 23 (11), 1281−92. (40) Marcobal, A., Barboza, M., Froehlich, J. W., Block, D. E., German, J. B., Lebrilla, C. B., and Mills, D. A. (2010) Consumption of human milk oligosaccharides by gut-related microbes. J. Agric. Food Chem. 58 (9), 5334−40. (41) Schwab, C., and Ganzle, M. (2011) Lactic acid bacteria fermentation of human milk oligosaccharide components, human milk oligosaccharides and galactooligosaccharides. FEMS Microbiol. Lett. 315 (2), 141−8. (42) Ganzle, M. G., and Follador, R. (2012) Metabolism of oligosaccharides and starch in lactobacilli: a review. Front. Microbiol. 3, 340. (43) Division of Family Health World Health Organization. (1982) The prevalence and duration of breast-feeding: a critical review of available information. World Health Stat. Q. 35 (2), 92−116. (44) Bartick, M., Stuebe, A., Shealy, K. R., Walker, M., and GrummerStrawn, L. M. (2009) Closing the quality gap: promoting evidencebased breastfeeding care in the hospital. Pediatrics 124 (4), e793−802. (45) Blaymore Bier, J. A., Oliver, T., Ferguson, A., and Vohr, B. R. (2002) Human milk reduces outpatient upper respiratory symptoms in premature infants during their first year of life. J. Perinatol. 22 (5), 354−9. (46) Ackerman, D. L., Craft, K. M., and Townsend, S. D. (2017) Infant food applications of complex carbohydrates: Structure, synthesis, and function. Carbohydr. Res. 437, 16−27. (47) Stuebe, A. (2009) The Risks of Not Breastfeeding for Mothers and Infants. Rev. Obstet. Gynecol. 2 (4), 222−231. (48) Eglash, A., Montgomery, A., and Wood, J. (2008) Breastfeeding. DM, Dis.-Mon. 54 (6), 343−411.

(49) Newburg, D. S., Ruiz-Palacios, G. M., and Morrow, A. L. (2005) Human milk glycans protect infants against enteric pathogens. Annu. Rev. Nutr. 25, 37−58. (50) Penders, J., Thijs, C., Vink, C., Stelma, F. F., Snijders, B., Kummeling, I., van den Brandt, P. A., and Stobberingh, E. E. (2006) Factors influencing the composition of the intestinal microbiota in early infancy. Pediatrics 118 (2), 511−21. (51) Li, M., Monaco, M. H., Wang, M., Comstock, S. S., Kuhlenschmidt, T. B., Fahey, G. C., Jr., Miller, M. J., Kuhlenschmidt, M. S., and Donovan, S. M. (2014) Human milk oligosaccharides shorten rotavirus-induced diarrhea and modulate piglet mucosal immunity and colonic microbiota. ISME J. 8 (8), 1609− 1620. (52) Hoeflinger, J. L., Davis, S. R., Chow, J., and Miller, M. J. (2015) In vitro impact of human milk oligosaccharides on Enterobacteriaceae growth. J. Agric. Food Chem. 63 (12), 3295−302. (53) Schroten, H., Hanisch, F. G., and Hansman, G. S. (2016) Human Norovirus Interactions with Histo-Blood Group Antigens and Human Milk Oligosaccharides. J. Virol. 90 (13), 5855−9. (54) Koromyslova, A., Tripathi, S., Morozov, V., Schroten, H., and Hansman, G. S. (2017) Human norovirus inhibition by a human milk oligosaccharide. Virology 508, 81−89. (55) Ruiz-Palacios, G. M., Cervantes, L. E., Ramos, P., ChavezMunguia, B., and Newburg, D. S. (2003) Campylobacter jejuni binds intestinal H(O) antigen (Fuc alpha 1, 2Gal beta 1, 4GlcNAc), and fucosyloligosaccharides of human milk inhibit its binding and infection. J. Biol. Chem. 278 (16), 14112−20. (56) Morrow, A. L., Ruiz-Palacios, G. M., Altaye, M., Jiang, X., Guerrero, M. L., Meinzen-Derr, J. K., Farkas, T., Chaturvedi, P., Pickering, L. K., and Newburg, D. S. (2004) Human milk oligosaccharides are associated with protection against diarrhea in breast-fed infants. J. Pediatr. 145 (3), 297−303. (57) Ackerman, D. L., Doster, R. S., Weitkamp, J. H., Aronoff, D. M., Gaddy, J. A., and Townsend, S. D. (2017) Human Milk Oligosaccharides Exhibit Antimicrobial and Antibiofilm Properties against Group B Streptococcus. ACS Infect. Dis. 3, 595. (58) Lin, A. E., Autran, C. A., Szyszka, A., Escajadillo, T., Huang, M., Godula, K., Prudden, A. R., Boons, G. J., Lewis, A. L., Doran, K. S., Nizet, V., and Bode, L. (2017) Human milk oligosaccharides inhibit growth of group B Streptococcus. J. Biol. Chem. 292, 11243. (59) Yu, Z. T., Nanthakumar, N. N., and Newburg, D. S. (2016) The Human Milk Oligosaccharide 2′-Fucosyllactose Quenches Campylobacter jejuni-Induced Inflammation in Human Epithelial Cells HEp-2 and HT-29 and in Mouse Intestinal Mucosa. J. Nutr. 146 (10), 1980− 1990. (60) Gonia, S., Tuepker, M., Heisel, T., Autran, C., Bode, L., and Gale, C. A. (2015) Human Milk Oligosaccharides Inhibit Candida albicans Invasion of Human Premature Intestinal Epithelial Cells. J. Nutr. 145 (9), 1992−8. (61) El-Hawiet, A., Kitova, E. N., Kitov, P. I., Eugenio, L., Ng, K. K., Mulvey, G. L., Dingle, T. C., Szpacenko, A., Armstrong, G. D., and Klassen, J. S. (2011) Binding of Clostridium difficile toxins to human milk oligosaccharides. Glycobiology 21 (9), 1217−27. (62) Nguyen, T. T., Kim, J. W., Park, J. S., Hwang, K. H., Jang, T. S., Kim, C. H., and Kim, D. (2016) Identification of Oligosaccharides in Human Milk Bound onto the Toxin A Carbohydrate Binding Site of Clostridium difficile. J. Microbiol. Biotechnol. 26 (4), 659−65. (63) Champion, E., McConnell, B., and Dekany, G. (2016) Mixtures of human milk oligosaccharides for treatment of bacterial infections, Patent WO2016063262A1. (64) Lin, A. E., Autran, C. A., Espanola, S. D., Bode, L., and Nizet, V. (2014) Human milk oligosaccharides protect bladder epithelial cells against uropathogenic Escherichia coli invasion and cytotoxicity. J. Infect. Dis. 209 (3), 389−98. (65) Coppa, G. V., Zampini, L., Galeazzi, T., Facinelli, B., Ferrante, L., Capretti, R., and Orazio, G. (2006) Human milk oligosaccharides inhibit the adhesion to Caco-2 cells of diarrheal pathogens: Escherichia coli, Vibrio cholerae, and Salmonella fyris. Pediatr. Res. 59 (3), 377−82. F



Viewpoint

(66) Cravioto, A., Tello, A., Villafan, H., Ruiz, J., del Vedovo, S., and Neeser, J. R. (1991) Inhibition of localized adhesion of enteropathogenic Escherichia coli to HEp-2 cells by immunoglobulin and oligosaccharide fractions of human colostrum and breast milk. J. Infect. Dis. 163 (6), 1247−1255. (67) Coppa, G. V., Gabrielli, O., Giorgi, P., Catassi, C., Montanari, M. P., Varaldo, P. E., and Nichols, B. L. (1990) Preliminary study of breastfeeding and bacterial adhesion to uroepithelial cells. Lancet 335 (8689), 569−71. (68) El-Hawiet, A., Kitova, E. N., and Klassen, J. S. (2015) Recognition of human milk oligosaccharides by bacterial exotoxins. Glycobiology 25 (8), 845−54. (69) Andersson, B., Porras, O., Hanson, L. A., Lagergard, T., and Svanborg-Eden, C. (1986) Inhibition of attachment of Streptococcus pneumoniae and Haemophilus influenzae by human milk and receptor oligosaccharides. J. Infect. Dis. 153 (2), 232−7. (70) Simon, P. M., Goode, P. L., Mobasseri, A., and Zopf, D. (1997) Inhibition of Helicobacter pylori binding to gastrointestinal epithelial cells by sialic acid-containing oligosaccharides. Infect. Immun. 65 (2), 750−757. (71) Weichert, S., Jennewein, S., Hufner, E., Weiss, C., Borkowski, J., Putze, J., and Schroten, H. (2013) Bioengineered 2′-fucosyllactose and 3-fucosyllactose inhibit the adhesion of Pseudomonas aeruginosa and enteric pathogens to human intestinal and respiratory cell lines. Nutr. Res. (N. Y., NY, U. S.) 33 (10), 831−8. (72) Marotta, M., Ryan, J. T., and Hickey, R. M. (2014) The predominant milk oligosaccharide 6′-sialyllactose reduces the internalisation of Pseudomonas aeruginosa in human pneumocytes. J. Funct. Foods 6, 367−373. (73) Hunt, K. M., Preuss, J., Nissan, C., Davlin, C. A., Williams, J. E., Shafii, B., Richardson, A. D., McGuire, M. K., Bode, L., and McGuire, M. A. (2012) Human milk oligosaccharides promote the growth of staphylococci. Appl. Environ. Microbiol. 78 (14), 4763−70. (74) Hester, S. N., Chen, X., Li, M., Monaco, M. H., Comstock, S. S., Kuhlenschmidt, T. B., Kuhlenschmidt, M. S., and Donovan, S. M. (2013) Human milk oligosaccharides inhibit rotavirus infectivity in vitro and in acutely infected piglets. Br. J. Nutr. 110 (7), 1233−42. (75) Jantscher-Krenn, E., Zherebtsov, M., Nissan, C., Goth, K., Guner, Y. S., Naidu, N., Choudhury, B., Grishin, A. V., Ford, H. R., and Bode, L. (2012) The human milk oligosaccharide disialyllacto-Ntetraose prevents necrotising enterocolitis in neonatal rats. Gut 61 (10), 1417−25. (76) Craft, K. M., and Townsend, S. D. (2017) Synthesis of lacto-Ntetraose. Carbohydr. Res. 440−441, 43−50. (77) Xiao, Z., Guo, Y., Liu, Y., Li, L., Zhang, Q., Wen, L., Wang, X., Kondengaden, S. M., Wu, Z., Zhou, J., Cao, X., Li, X., Ma, C., and Wang, P. G. (2016) Chemoenzymatic Synthesis of a Library of Human Milk Oligosaccharides. J. Org. Chem. 81 (14), 5851−65. (78) Prudden, A. R., Liu, L., Capicciotti, C. J., Wolfert, M. A., Wang, S., Gao, Z. W., Meng, L., Moremen, K. W., and Boons, G. J. (2017) Synthesis of asymmetrical multiantennary human milk oligosaccharides. Proc. Natl. Acad. Sci. U. S. A. 114 (27), 6954−6959. (79) Wu, S., Grimm, R., German, J. B., and Lebrilla, C. B. (2011) Annotation and structural analysis of sialylated human milk oligosaccharides. J. Proteome Res. 10 (2), 856−68. (80) Boltje, T. J., Buskas, T., and Boons, G. J. (2009) Opportunities and challenges in synthetic oligosaccharide and glycoconjugate research. Nat. Chem. 1 (8), 611−22.

G


The Human Milk Glycome as a Defense Against Infectious Diseases

Recommend Documents