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ABSTRACT Urinary tract infections (UTIs) caused by _Escherichia coli_ create a large burden on healthcare and frequently lead to recurrent infections. Part of the success of _E. coli_ as an

uropathogenic bacterium can be attributed to its ability to form quiescent intracellular reservoirs in bladder cells and its persistence after antibiotic treatment. Cranberry juice and

related products have been used for the prevention of UTIs with varying degrees of success. In this study, a group of cranberry pectic oligosaccharides (cPOS) were found to both inhibit

quiescence and reduce the population of persister cells formed by the uropathogenic strain, CFT073. This is the first report detailing constituents of cranberry with the ability to modulate

these important physiological aspects of uropathogenic _E. coli_. Further studies investigating cranberry should be keen to include oligosaccharides as part of the ‘active’ cocktail of

chemical compounds. SIMILAR CONTENT BEING VIEWED BY OTHERS LYOPHILIZED CELL-FREE SUPERNATANTS OF _LIMOSILACTOBACILLUS FERMENTUM_ T0701 EXHIBITED ANTIBACTERIAL ACTIVITY AGAINST _HELICOBACTER

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access 23 November 2022 INTRODUCTION Bacterial urinary tract infections (UTIs) affect up to 150 million people each year and are among the most common infections worldwide1. It is estimated

that the total societal cost burden of UTIs is $3.5 billion each year in the US alone2. Over 50% of women will experience a UTI, with an estimated recurrence rate of about 25%3. While UTIs

are typically curable, recurrent infections can require continuous antibiotic prophylaxis for prevention. Frequent use of antibiotics, however, can lead to adverse events such as colitis,

the development of antibiotic resistant UTIs, or the development of complicated UTIs2,3,4. Uropathogenic _Escherichia coli_ (UPEC) is responsible for 65–90% of UTI cases2,4,5,6. For

recurrent UTIs, the original infecting UPEC strain is responsible 77% of the time7. While the mechanism of recurrence is not fully understood, it is thought that intracellular, biofilm-like

communities (IBCs) and quiescent intracellular reservoirs (QIRs) are contributing factors8,9,10. IBCs form when UPEC from urine invade and replicate inside of superficial bladder cells6,10.

As the infection progresses, subsequent UPEC invasion of urothelial transitional cells can lead to establishment of QIRs. Cells residing in QIRs can remain viable for months6,8,9. Upon

exfoliation of bladder epithelial cells, UPEC from QIRs would be able to re-colonize urine and cause a recurrent infection6. Since antibiotics generally target actively dividing bacteria,

quiescent UPEC cells are unlikely to be eradicated by antibiotics, thus contributing to treatment failures and recurrence8,10. Persistence is a physiologic state similar to quiescence for

UPEC11,12,13,14. Persister cells are a small subpopulation of bacteria that exist in a dormant, non-dividing state characterized by high antibiotic tolerance11,12,14. Persister cells can be

isolated and regrown following initial antibiotic treatment, and these populations retain sensitivity to the initial antibiotic and can consistently form another persister cell fraction15.

Similar to QIR cells, persister cells are strongly implicated in the pathogenesis of chronic infections, including recurrent UTIs16,17,18,19,20. While proven to be metabolically distinct,

the quiescent and persistent physiological states of UPEC are both likely to cause recurrent UTIs and treatment failures16. Therapies aimed at preventing or reversing quiescence and

persistence could significantly reduce the burden of UPEC recurrent UTIs. Cranberry (_Vaccinium macrocarpon_) products have been extensively researched for their ability to prevent UTIs,

especially those caused by UPEC21,22,23,24,25,26. Past studies have focused on cranberry proanthocyanidins (PACs) with A-type linkages24,27 and fructose28 since these have been shown to

interfere with UPEC adherence to bladder cells. Organic acids present in cranberry, including quinic, shikimic, malic and citric acids, were recently demonstrated to help reduce _E. coli_

populations in an experimental mouse model of UTI29. Other recent studies have suggested that cranberry complex carbohydrates might also contribute to the prevention of UTIs. For example,

xyloglucan oligosaccharides from cranberry have been shown to interfere with cell adhesion and biofilm formation by uropathogenic _E. coli_30,31,32. We hypothesized that certain cranberry

constituents might interfere with quiescent and persister cells since these are implicated in UTI recurrence. Herein, we report the isolation and chemical characterization of cranberry

pectic oligosaccharides (cPOS) that prevent quiescence in the prototypical UPEC strain CFT073 and significantly reduce the population of UPEC persister cells in the presence of ampicillin.

RESULTS THE UPEC STRAIN CFT073 DEMONSTRATES _IN VITRO_ QUIESCENCE ON GLUCOSE MINIMAL MEDIA AGAR PLATES AND RESUMES GROWTH IN RESPONSE TO CRANBERRY CONSTITUENTS We previously reported that

the UPEC strain CFT073 grows normally in liquid glucose minimal media but exhibits _in vitro_ quiescence on glucose minimal media agar plates when plated at low density (≤106 CFU)16. It was

found that cells exhibiting quiescence under these conditions could be stimulated to grow by transferring a colony of _E. coli_ MG1655 onto the plate or by applying an amino acid cocktail

comprising various combinations of lysine, methionine, and tyrosine. Following addition of the stimulus to the agar surface and 24 hour incubation at 37 °C, growing CFT073 cells appear as

visible colonies radiating out from the site of the stimulus (see Fig. 1 as an example). The diameter of the zone of growing CFT073 continues to expand with additional incubation time. This

experiment is a qualitative test for stimuli that reverse _in vitro_ quiescence of UPEC strains. Because cranberry has been traditionally used for preventing UTI, we hypothesized that

certain cranberry constituents might reverse UPEC quiescence. The above quiescence assay was used to test cranberry phytochemicals and guide the separation of the active components (Fig. 

S1). A pectinase degraded cranberry hull extract was separated using C18 reversed phase chromatography to afford an oligosaccharide-enriched fraction, Cranf1b, that induced bacteria colony

formation when added (100 µg in 10 µL water) to the surface of an agar plate containing quiescent CFT073 (data not shown). Cranf1b was further separated using porous graphitized carbon (PGC)

cartridges. Only a sub-fraction that eluted with 10% ACN/H2O/0.1%TFA showed reversal of _E. coli_ CFT073 quiescence (Fig. 1). This sub-fraction (cPOS) was further separated using HPLC

chromatography and characterized using a combination of NMR, HR-MS and LC-MS/MS. CHEMICAL CHARACTERIZATION OF CRANBERRY CONSTITUENTS THAT REVERSE QUIESCENCE The 1H and HSQC NMR spectra of

the active cranberry fraction revealed proton and carbon resonances with chemical shifts characteristic for poly-uronic acids. Additional structural features included a methoxy group at δ

3.70/52.8 and a vinylic methine at δ 6.00/111.8 (Figs. S2 and S3). Monosaccharide composition analysis of cPOS determined that the oligosaccharides were comprised of galacturonic acid units

(Fig. S4). The active fraction was further analyzed by hydrophilic interaction liquid chromatography (HILIC) HPLC (Fig. 2). HILIC LC-MS data showed that the predominate ions were consistent

with poly-galacturonic acid methyl esters with various degrees of polymerization (DP) (Fig. 3). Each oligosaccharide was present as a pair of α- and β- anomers in equilibrium. These results

were in agreement with HR-ESI-MS and HR-ESI-MS/MS data obtained for collected fractions (Figs. S5 and S6). The most abundant ions indicated the presence of fragments with one or two free

carboxylic acids and one additional degree of unsaturation. UV absorption maxima at 235 nm was consistent with the presence of an α, β-unsaturated carboxylic acid, which is in agreement of

the vinylic methine observed in NMR, and accounting for the additional unsaturation. The major components were therefore determined to be unsaturated pectic oligosaccharides, referred to as

uGXmY (uG: unsaturated poly-Galacturonic acid; X: degree of polymerization; Y: number of methyl esters). Saturated poly-galacturonic acid methyl esters were unexpectedly found to be present

only trace amounts in cPOS (data not shown). We next isolated two unsaturated poly-galacturonic acid methyl esters, uG3m2 (HRMS (_m/z_): [M−H]−, C20H27O18, 555.1205, calcd. 555.1197) and

uG4m3 (1, HRMS (_m/z_): [M−H]−, C27H37O24, 745.1692, calcd.745.1674) from the cPOS mixture. Compounds eluting after 1 were pooled into a single fraction containing cPOS with higher DPs

(HDP-cPOS). Purified 1 was characterized using 1D and 2D NMR spectra (Figs. S7–S12) and compared to previously reported poly-galacturonic acids33,34,35,36. Superimposition of HSQC spectra

for both 1 and cPOS showed that both contained similar correlation signals (Fig. S3), confirming 1 as a representative constituent of cPOS. The 1H and 13C NMR chemical shifts of 1 are listed

in Table S1. Briefly, five spin systems were identified and were assigned to the four subunits (A-α/A-β, B, C, D) of 1 (Fig. 4). Key HMBC and NOE correlations are shown in Fig. 4. HMBC

correlations between the methoxy protons and the C-6 carbonyls were observed for subunits A, B and D, indicating that the free carboxylic acid was located on subunit C. The C-6 of subunit C

also has a higher NMR chemical shift (δ 175.0 ppm), consistent with a carboxylic acid at this position. The MS/MS fragmentation patterns indicated sequential neutral losses of 190 Da for 1

(Fig. 5), further supporting the structure. Three previously reported iridoid glucosides were isolated from the cPOS fraction as minor impurities and identified as 6,7-dihydromonotropein,

deacetylasperulsidic acid and monotropein37,38,39,40. These iridoid glucosides were inactive in the _in vitro_ quiescence inhibition assay (data not shown). We found that these could be

removed from the cPOS fraction by exploiting their differential solubility in ethanol. Cranf1b was triturated with 95% aqueous ethanol prior to PGC separation, producing a cPOS fraction

(cPOS-t) devoid of iridoid glycosides as determined by HPLC analysis (Fig. 2) and LC-MS ion extraction (data not shown). PURIFIED OLIGOSACCHARIDES UG3M2 AND UG4M3 INHIBIT _E. COLI_ CFT073

QUIESCENCE The isolated oligosaccharides uG4m3 and uG3m2 were tested alongside cPOS and HDP-cPOS for reversal of _E. coli_ CFT073 quiescence (Fig. 1). All four samples, applied as 100 

μg/10μL aqueous solutions to the agar surface, stimulated the growth of quiescent bacteria, indicating an inhibition of quiescence. The quiescence assay was repeated using glucose-free

plates to test if bacteria growth was simply due to bacteria using the added oligosaccharides as a carbon source. The addition of uG3m2 (100 μg/10μL) onto glucose-free plates containing

quiescent CFT073 did not result in visible growth of bacteria colonies (Fig. 6). However, co-spotting 100 μg uG3m2 and 2 mg glucose onto agar containing quiescent CFT073 stimulated bacteria

growth. This result is consistent with the cranberry oligosaccharides stimulating the quiescent UPEC to grow on glucose. _E. COLI_ CFT073 GENERATES A LOW LEVEL OF PERSISTER CELLS IN LIQUID

GLUCOSE M9 MINIMAL MEDIUM CONTAINING CRANBERRY PECTIC OLIGOSACCHARIDES Normal microbial populations contain persister cells as small subpopulations (10−3 to 10−4%) of dormant cells that are

tolerant to antibiotics. Upon regrowth in the absence of the drug, persister cells produce cultures that regain full sensitivity to the antibiotic. Persister cells likely play a role in

chronic infections13. We previously found that _E. coli_ CFT073 generates high levels of persister cells in liquid glucose M9 minimal medium in the presence of ampicillin (100 µg/mL), and

the addition of a mixture of the 20 standard L-amino acids (100 µg/mL each) reduced persister cell populations by 100-fold during ampicillin treatment16. Since cPOS reversed _E. coli_ CFT073

quiescence on glucose plates, we were interested in determining whether cPOS addition to cultures of _E. coli_ CFT073 grown in glucose M9 minimal medium in the presence of ampicillin would

generate fewer persister cells. Overnight cultures of _E. coli_ CFT073 grown on 0.4% glucose M9 minimal medium were diluted 20-fold into fresh 0.2% glucose M9 minimal medium (_A_600 of 0.1,

~108 CFU/mL) containing or lacking ampicillin (100 µg/mL), and viable counts were measured after 4 h and 24 h at 37 °C (Fig. 7). As previously observed16, the viable cell counts in the _E.

coli_ CFT073 cultures decreased slightly during the first 4 h of incubation in both the presence and absence of ampicillin. By 24 h, _E. coli_ CFT073 had grown to just over 109 CFU/mL in the

absence of ampicillin (Fig. 7). In contrast, the presence of ampicillin reduced the viable counts an additional ~1000-fold over the next 20 h, to about 106 CFU/mL. As shown by the results

in Fig. 7, in the presence of 1 mg/mL cPOS and absence of ampicillin, _E. coli_ CFT073 viable counts increased rapidly, reaching stationary phase within 4 h. Importantly, in the presence of

both the cPOS and ampicillin, _E. coli_ CFT073 viable counts decreased rapidly within 4 h, and reached a level of about 3 × 102 CFU/mL after 24 h (Fig. 7). Therefore, treating CFT073 with

both cPOS and ampicillin resulted in >1000-fold fewer persister cells versus treatment with ampicillin alone. Similar reduction of persisters was obtained when conducting the assay with

cPOS-t (Fig. S13). DISCUSSION The oligosaccharides discussed in this manuscript suggest previously undiscovered benefits of cranberry constituents for mitigating UPEC infections. Other than

specific amino acid cocktails, cPOS represents the first molecules shown to reverse a quiescent phenotype and reduce a persister cell fraction in the UPEC strain CFT073. Inhibition of UPEC

quiescence is intriguing since this phenotype has been linked with recurrent UTIs6. Persistence and quiescence are both phenotypes involving dormant bacteria cells; however, it remains to be

determined if the physiologic change(s) induced in UPEC by cPOS is shared between the two cell types. Prevention of UPEC dormancy may allow host defense mechanisms and common treatment

approaches to be more effective. CFT073 is a prototypic UPEC strain belonging to the phylogenetic group B2 multilocus sequence type 73 strains (ST73). ST73 is a major UPEC lineage,

accounting for 11% and 16.6% of UPEC isolated from patients in recent studies41,42. It was previously shown that CFT073 enters a quiescent state when plated onto M9 minimal medium agar

plates containing glucose at concentrations ≤106 CFU16. A recent study found that 30 of 38 ST73 strains (78.9%) entered the quiescent state on M9 glucose plates, demonstrating that

quiescence is a common phenotype for this UPEC lineage. UPEC strains belonging to other sequence types (ST141, ST104, ST394, and ST998) have also demonstrated the same _in vitro_ quiescent

phenotype16. Additional studies are underway to determine if cPOS reverses quiescence in UPEC strains other than CFT073. We have shown that cranberry pectic oligosaccharides reduce a

persister cell fraction of the UPEC strain CFT073 formed in the presence of ampicillin. Ampicillin belongs to the beta-lactam class of antibiotics and inhibits enzymes critical for cell wall

biosynthesis. Further studies are needed to determine if cPOS similarly reduces persister cell fractions when used with other antibacterial agents, especially those that inhibit bacteria

via other mechanisms of action such as the fluoroquinolones. The dehydration on subunit D is likely due to the enzymatic treatment used to break down insoluble pectic polysaccharides during

cranberry juice manufacturing33. Some pectinases can selectively hydrolyze glycosidic bonds by eliminative cleavage and cause oxygen-aglycone bond breakage43,44,45. For example, a

mechanistic study on a pectin lyase A (PLA) derived from _Aspergillus niger_, of the same origin as the Klerzyme 150 in this study, showed specific cleavage of a glycosidic bond adjacent to

a methyl-esterified galacturonic acid residue, leaving a 4,5-unsaturated galacturonic acid methyl ester at the non-reducing end46. Interestingly, it was also pointed out that free carboxylic

acids generally occurred second from the non-reducing end, which is in agreement with our assignment of 146. Previous studies of cranberry oligosaccharides have focused on xyloglucans and

their antibiofilm and antiadhesion properties30,31,32. A recent report showed that xyloglucans can be found in pig urine following oral administration31. In the study presented here,

xyloglucans were separated from cPOS by PGC chromatography and were inactive when tested in the quiescence and persister cells assays. Hence, there may be distinct health benefits from the

structurally different oligosaccharide fractions found in cranberry. The results presented here encourage further investigation of cPOS as agents to combat recurrent _E. coli_ UTIs and

support current drug treatment options. Our findings complement those concerning other cranberry constituents (e.g. PACs, organic acids and xyloglucans) and their possible benefits in

preventing UTI infections. It is rational to speculate that the full benefit of cranberry may not be realized with one chemical class, but rather that multiple classes act in concert.

Reviews of the published clinical data show inconsistencies in chemical characterization and preparation of cranberry products tested for UTI prevention22. The optimal formulation and use of

cranberry products for UTI prevention will require further knowledge of the active constituents and mechanisms of action. Until then, cranberry juice or whole berry products might be best

used until more is known regarding how and why cranberry may aid in preventing UTIs. METHODS GENERAL EXPERIMENTAL PROCEDURES NMR experiments were conducted using an Agilent Inova NMR 500 MHz

spectrometer in D2O (99.99%, Sigma-Aldrich, St. Louis, MO, USA) at 25 °C. LC-MS analysis was performed on a Prominence UFLC system (Shimadzu, Kyoto, Japan) coupled to a QTRAP 4500 mass

spectrometer (AB Sciex, Framingham, MA, USA) with electrospray ionization source on negative ion mode with scans from _m/z_ 200 to _m/z_ 2000. High Resolution Electrospray Ionization mass

spectra and tandem mass spectra (HR-ESI-MS and HR-ESI-MS/MS) were acquired using a TripleTOF 4600 spectrometer (AB Sciex) operating in negative ion mode. Flash chromatography was completed

using an Agilent 971-FP flash purification system (Agilent Technologies, Santa Clara, CA, USA) with Biotage SNAP KP-C18-HS 120 g cartridges (Biotage, Charlotte, NC, USA). HPLC separations

were performed on a Shimadzu Prominence i-series HPLC (Shimadzu). SEPARATION OF CRANBERRY PECTIC OLIGOSACCHARIDES Cranberry hulls were degraded with pectinase (Klerzyme 150, DSM Food

Specialties, Heerlen, Netherlands) and fractionated as previously described with modifications30. Briefly, cranberry pectinase treated hull extract was fractionated using a C18 flash column

sequentially eluted with 500 mL 100% deionized (d.i.) water, 500 mL 15% methanol/H2O and 500 mL 100% methanol. Fractions from each step-gradient were individually pooled into three major

fractions, Cranf1W (38.1%, w/w, dry), Cranf1b (23.8%, w/w, dry) and Cranf1M (28.1%, w/w, dry). Cranf1b was then subjected to separation using a PGC HyperSep Hypercarb SPE cartridge (1 gram,

Thermofisher Scientific, Waltham, MA, USA). The PGC cartridge was firstly conditioned by 50% ACN/water and then equilibrated with 100% d.i. water. Two mL of 20 mg/mL Cranf1b aqueous solution

was then loaded onto the PGC cartridge and eluted sequentially with d.i. water, 10% ACN/H2O/0.1% TFA and 30% ACN/H2O/0.1% TFA. The 10% ACN/H2O/0.1% TFA PGC eluent (cPOS, 15% w/w, dry) was

then separated using a TSKgel Amide-80 HR HILIC column (4.6 × 250 mm, 5 μm, TOSOH Bioscience, Tokyo, Japan) at 1 mL/min and 35 °C. The column was eluted with a linear gradient from 80%

ACN/H2O/0.1% formic acid to 40% ACN/H2O/0.1% formic acid over 30 min. uG3m2 and 1 were collected at 10 min and 12.5 min, respectively (Fig. 2). Trituration of Cranf1b to remove iridoid

terpenes was accomplished by sonicating 200 mg Cranf1b in 30 mL 95% ethanol/water and then centrifuging the solids at 17,000x g for 8 min. The collected solids were then dissolved in d.i.

H2O and lyophilized. The trituration step was repeated twice prior to further separation by PGC chromatography to provide sample cPOS-t (91.7%, w/w dry). URONIC ACID IDENTIFICATION OF CPOS

The composition of the poly-galacturonic acid backbone of cPOS was determined by acid hydrolysis of cPOS followed by derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP)47. Briefly, 10 

mg cPOS was hydrolyzed in 1 mL 2 M TFA at 90 °C for 5 h. After cooling to room temperature, the reaction mixture was centrifuged at 3000 rpm for 5 min. The supernatant was evaporated _in

vacuo_ to remove residual TFA. The hydrolysate mixture was dissolved in 1 mL distilled water and treated with 30 µL NaOH (0.3 M) and 20 µL PMP solution (0.5 M in methanol) for

derivatization. The mixture was incubated at 70 °C for 60 min, cooled to room temperature and neutralized with 30 µL of HCl (0.3 M). The mixture was then extracted with 1 mL chloroform. The

aqueous layer was filtered by passing through a 0.45 µm syringe filter. Glucuronic acid (2.0 mM), galacturonic acid (2.0 mM) and a distilled water blank were derivatized the same as

described above. HPLC analyses of PMP-labeled samples were conducted on a Prominence i-series system (Shimadzu) using a C18 column (4.6 mm × 250 mm, 5 𝜇m, J.T. Baker Inc., Phillipsburg, NJ,

USA). The injection volume was 20 µL with a flow rate of 1.0 mL/min at 35 °C. Mobile phase A was 10% ACN/water with 0.045% KH2 PO4–0.05% triethylamine buffer (pH 7.5) and mobile phase B was

100% ACN. The column was eluted with 6% B for 4 min, 6–12% for 1 min and then 12% B for 20 min. The UV detection wavelength was 254 nm. BACTERIAL STRAINS AND CULTIVATION _E. coli_ CFT073 was

obtained from a cryogenically frozen stock at the laboratory of P.S.C. and stored at −80 °C in a 1:1 mixture of LB broth and 50% glycerol by volume. LB broth and LB agar were used for

routine cultivation. Liquid M9 minimal medium was prepared as described previously16, and M9 minimal medium agar plates were prepared with 1.5% noble agar to avoid impurities present in

bacteriological agar. _IN VITRO_ QUIESCENCE INHIBITION ASSAY The procedure for this assay closely follows a published protocol16. In general, overnight cultures of _E. coli_ CFT073 were

prepared in 0.4% glucose M9 minimal medium as aforementioned. Bacteria from this culture were diluted to a final concentration of 105 CFU in 4 mL of liquid overlay media (0.2% glucose M9

minimal medium with 0.9% noble agar at 45 °C). Each 4 mL overlay inoculum was poured over a pre-warmed (37 °C) 0.2% glucose M9 minimal media agar plate immediately after inoculation. These

plates were allowed to solidify at room temperature with lids slightly ajar. Once solidified, test spots were added to the overlay media and allowed to dry before incubating the plate upside

down at 37 °C for 24 h. Test spots were 10 μL of each test solution at 10 mg/mL in M9 minimal media unless indicated otherwise. Non-growing _E. coli_ in the overlay were considered to be

quiescent if growth could be induced with the positive control (3 co-spots; 5 μL each of tyrosine, lysine, and methionine solutions at 0.1 mg/mL; co-spots added sequentially to the identical

location on the agar). To test the effects in glucose-free conditions, glucose was not added to any of the media components prior to completing the assay. When adding glucose back to the

glucose-free plates, a 10 μL spot of 20% glucose in water (2 mg; sufficient to support bacteria growth) was added directly following and to the identical location as the dried test spot. For

all quiescence assays, a positive result was defined as observable bacteria growth at the site of a test sample following 24 h incubation. A negative result was defined as no observable

bacteria growth at the site of a test sample. Images of agar plates were made after 24 h using a Molecular imager Gel Doc XR+ (Bio-rad, Hercules, CA, USA) system with Image Lab Software.

PERSISTER CELL VIABILITY ASSAY The procedure for this assay closely followed a published protocol16. _E. coli_ CFT073 was streaked from cryogenically frozen stocks onto LB agar plates and

incubated overnight at 37 °C. A loopful of cells from the plate was added to 10 mL 0.4% glucose M9 minimal media in a 125 mL culture flask and incubated overnight at 37 °C and 200 rpm.

Cultures were diluted in fresh 0.2% glucose M9 minimal media supplemented with test samples (1 mg/mL) to an optical density (OD600) of 0.1 (~108 CFU/mL) and grown in the presence of

ampicillin sodium at 10x MIC (0.1 mg/mL) to generate persister fractions. Cultures were grown in the absence of ampicillin and/or cranberry constituents as controls. The cultures were

incubated shaking (200 rpm) at 37 °C and viable counts were measured at 0, 4, and 24 hours by plating on LB media. The persister cell viability assays were completed in triplicate and

analyzed for statistical significance. STATISTICS Persister cell viability assays were compared using a two-tailed student’s _t_ test. _P_-values ≤ 0.05 were considered statistically

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Ocean Spray Cranberries, Inc. (Lakeville-Middleboro, MA, USA). RD was financially supported by the Omar Magnate Foundation Fellowship. Research reported in this publication was made possible

by the use of equipment and services available through the RI-INBRE Centralized Research Core Facility, which is supported by the Institutional Development Award (IDeA) Network for

Biomedical Research Excellence from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM103430, and by the National Science

Foundation EPSCoR Cooperative Agreement #EPS-1004057. AUTHOR INFORMATION Author notes * These authors contributed equally: Jiadong Sun and Robert W. Deering. AUTHORS AND AFFILIATIONS *

Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, Kingston, RI, 02881, USA Jiadong Sun, Robert W. Deering, Zhiyuan Peng, Laila Najia, 

Navindra P. Seeram & David C. Rowley * Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD,

20814, USA Jiadong Sun * Ocean Spray Cranberries, Inc., One Ocean Spray Drive, Lakeville-Middleboro, MA, 02349, USA Christina Khoo * Department of Cell and Molecular Biology, University of

Rhode Island, Kingston, RI, 02881, USA Paul S. Cohen Authors * Jiadong Sun View author publications You can also search for this author inPubMed Google Scholar * Robert W. Deering View

author publications You can also search for this author inPubMed Google Scholar * Zhiyuan Peng View author publications You can also search for this author inPubMed Google Scholar * Laila

Najia View author publications You can also search for this author inPubMed Google Scholar * Christina Khoo View author publications You can also search for this author inPubMed Google

Scholar * Paul S. Cohen View author publications You can also search for this author inPubMed Google Scholar * Navindra P. Seeram View author publications You can also search for this author

inPubMed Google Scholar * David C. Rowley View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS D.C.R., J.S., R.W.D. and P.S.C. conceived and

designed the experiments. J.S. and R.W.D. prepared the manuscript and contributed equally to this work. J.S. and R.W.D. performed chemical purification and characterization of cranberry

materials. R.W.D and L.N. performed _in vitro_ quiescence and persister cell assays. Z.P. contributed to Figures 2 and S3. L.N. contributed to Figure 8. C.K., P.S.C., N.P.S and D.C.R edited

and approved the final version of the manuscript. CORRESPONDING AUTHOR Correspondence to David C. Rowley. ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing interests.

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CITE THIS ARTICLE Sun, J., Deering, R.W., Peng, Z. _et al._ Pectic Oligosaccharides from Cranberry Prevent Quiescence and Persistence in the Uropathogenic _Escherichia coli_ CFT073. _Sci

Rep_ 9, 19590 (2019). https://doi.org/10.1038/s41598-019-56005-w Download citation * Received: 06 March 2019 * Accepted: 04 December 2019 * Published: 20 December 2019 * DOI:

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