Changes in Structure and Antigenicity of HIV-1 Env Trimers Resulting from Removal of a Conserved CD4 Binding Site-Proximal Glycan.

UNLABELLED: The envelope glycoprotein (Env) is the major target for HIV-1 broadly neutralizing antibodies (bNAbs). One of the mechanisms that HIV has evolved to escape the host's immune response is to mask conserved epitopes on Env with dense glycosylation. Previous studies have shown that the removal of a particular conserved glycan at N197 increases the neutralization sensitivity of the virus to antibodies targeting the CD4 binding site (CD4bs), making it a site of significant interest from the perspective of vaccine design. At present, the structural consequences that result from the removal of the N197 glycan have not been characterized. Using native-like SOSIP trimers, we examine the effects on antigenicity and local structural dynamics resulting from the removal of this glycan. A large increase in the binding of CD4bs and V3-targeting antibodies is observed for the N197Q mutant in trimeric Env, while no changes are observed with monomeric gp120. While the overall structure and thermostability are not altered, a subtle increase in the flexibility of the variable loops at the trimeric interface of adjacent protomers is evident in the N197Q mutant by hydrogen-deuterium exchange mass spectrometry. Structural modeling of the glycan chains suggests that the spatial occupancy of the N197 glycan leads to steric clashes with CD4bs antibodies in the Env trimer but not monomeric gp120. Our results indicate that the removal of the N197 glycan enhances the exposure of relevant bNAb epitopes on Env with a minimal impact on the overall trimeric structure. These findings present a simple modification for enhancing trimeric Env immunogens in vaccines. IMPORTANCE: The HIV-1 Env glycoprotein presents a dense patchwork of host cell-derived N-linked glycans. This so-called glycan shield is considered to be a major protective mechanism against immune recognition. While the positions of many N-linked glycans are isolate specific, some are highly conserved and are believed to play key functional roles. In this study, we examine the conserved, CD4 binding site-proximal N197 glycan and demonstrate that its removal both facilitates neutralizing antibody access to the CD4 binding site and modestly impacts the structural dynamics at the trimer crown without drastically altering global Env trimer stability. This indicates that surgical glycosylation site modification may be an effective way of sculpting epitope presentation in Env-based vaccines.

Building a Personalized Cancer Treatment System.

This paper reports the process by which a personalized cancer treatment system was built, following a user-centered approach. We give some background on personalized cancer treatment, the particular tumor chemosensitivity assay supported by the system, as well as some quality and legal issues related to such health systems. We describe how Contextual Design was applied when building the system. Contextual design is a user-centered design technique involving seven steps. We also provide some details about the system implementation. Finally, we explain how the Think-Aloud protocol and Heuristic Evaluation methods were used to evaluate the system and report its results. A qualitative assessment from the users perspective is also provided. Results from the heuristic evaluation indicate that only one of ten heuristics was missing from the system, while five were partially covered and four were fully covered.

Barriers to Healthcare Access for Immigrants in Costa Rica and Uruguay.

Access to public health has been, is, and will be a necessary right for any person in the world, motivating the proposal of universalist approaches as the best way to provide this service. However, we know that universalism is limited, at best, when it concerns immigrants. In this article, we focus on Costa Rica's and Uruguay's health systems, generally acknowledged as Latin America's most universal, to argue that there are important barriers that limit immigrants' access to public health insurance and health care. Applying a model based on the work by Niedzwiecki and Voorend (2019) that allows us to disaggregate the barriers to access into legal, institutional, de facto, and agency barriers, our analysis shows that migration and social policy interact to create barriers of different magnitudes, often conditioning healthcare access on migratory status, formal employment, and/or purchasing power. These limitations to universal social protection create important vulnerabilities, not only for the immigrants involved, but also for the health systems, and therefore for public health, highlighting the limitations of universalism.

Evaluation of hypereosinophilia in a case of FLT3-mutant acute myeloid leukemia treated with gilteritinib.

Acute myeloid leukemias (AMLs) frequently harbor activating mutations in Fms-like tyrosine kinase 3 (FLT3). The use of FLT3 inhibitors (FLT3i) is the standard of care for treatment of newly diagnosed and relapsed patients with AML. Differentiation responses including clinical differentiation syndrome have been previously reported with FLT3i when used as single agents in relapsed disease. We present a case of hypereosinophilia in a patient on FLT3i therapy with persistent FLT3 polymerase chain reaction (PCR) positivity in peripheral blood. We sorted mature leukocytes by lineage to determine if the eosinophils were leukemia-derived. FLT3 PCR and next-generation sequencing analysis demonstrated monocytic differentiation of the FLT3-ITD leukemic clone with reactive hypereosinophilia that was derived from a preleukemic SF3B1, FLT3 wild-type clone. Our case is the first to definitively demonstrate the emergence of clonal FLT3-ITD monocytes with FLT3i and the first to demonstrate a differentiation response following decitabine, venetoclax, and gilteritinib triplet therapy.

Using Metabolomics to Identify the Exposure and Functional Biomarkers of Ginger.

Liquid chromatography-mass spectrometry (LC-MS)-based metabolomics has become an important tool to increase our understanding of how diet affects human health. However, public and commercial mass spectral libraries of dietary metabolites are limited, resulting in the greatest challenge in converting mass spectrometry data into biological insights. In this study, we constructed an LC-MS/MS ginger library as an example to demonstrate the importance of dietary libraries for discovering food biomarkers. The functional and exposure biomarkers of ginger were investigated using plasma samples from mice treated with control and ginger extract diets. Our results showed clear discrimination between the metabolome of mice on normal and ginger extract diets. Using the in-house ginger library, we identified 20 ginger metabolites that can be used as exposure biomarkers of ginger. However, without the LC-MS/MS ginger library, none of the ginger metabolites could be accurately identified based on online mass databases. In addition, ginger treatment significantly impacts the endogenous metabolome, especially the purine metabolism and phenylalanine, tyrosine, and tryptophan biosynthesis. Overall, we demonstrated that the construction of LC-MS/MS spectra dietary libraries would enhance the ability to identify potential dietary biomarkers and correlate potential health benefits associated with food consumption.

Cranberry proanthocyanidins composite electrospun nanofibers as a potential alternative for bacterial entrapment applications.

The interaction between A-type interflavan bonds from cranberry proanthocyanidins (PAC) and surface virulence factors of extra-intestinal pathogenic Escherichia coli (ExPEC) was studied. Electrospun nanofibers (ESNF) were fabricated using PAC and polycaprolactone (PCL) solutions and their physical and chemical properties were characterized. The ability of PAC:PCL composite ESNF to interact with and entrap ExPEC strain 5011 (ExPEC-5011) was evaluated in vitro by plate culturing and when formulated as a biofilter and nanocoating. As a biofilter, the PAC:PCL ESNF exhibited a dose-dependent ability to entrap ExPEC-5011. Images from scanning electron and fluorescent microscopies revealed that ESNF sections with higher amounts of PAC led to higher bacterial entrapment. The effectiveness PAC:PCL ESNF to bind ExPEC when applied as a nanocoating was studied using ESNF-coated polyvinyl chloride intermittent catheter. Results indicate that ExPEC-5011 was entrapped well into the PAC:PCL ESNF coating on the catheter. Overall, our results suggest that incorporating the biomolecule PAC in ESNF is a potential means for applications requiring bacterial entrapment, such as biofunctionalization, biofiltration, and surface coating, among others.

Classification of proanthocyanidin profiles using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectra data combined with multivariate analysis.

Proanthocyanidin (PAC) profiles of apples (a-PAC), cranberries (c-PAC), and peanut skins (p-PAC) were determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Deconvolution of overlapping isotopic patterns indicated that in apples, only 5% of the PAC oligomers contain one or more A-type bonds, whereas in cranberries and peanut skins, 96% of the PAC oligomers contain one or more A-type bonds. MALDI-TOF MS data combined with multivariate analysis, such as principal component analysis (PCA) and linear discriminant analysis (LDA), were used to differentiate and discriminate a-PAC, c-PAC, and p-PAC from one another. Mixtures of c-PAC with either a-PAC or p-PAC at different w/w ratios were evaluated by LDA modeling. The LDA model classified the training, testing, and validation sets with 99.4%, 100%, and 94.2% accuracy. Results suggest that MALDI-TOF MS and multivariate analysis are useful in determining authenticity of PAC from different sources and mixtures of PAC sources.

Identification of A-Type Proanthocyanidins in Cranberry-Based Foods and Dietary Supplements by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry, First Action Method: 2019.05.

BACKGROUND: Cranberry proanthocyanidins (c-PAC) are oligomeric structures of flavan-3-ol units, which possess A-type interflavan bonds. c-PAC differs from other botanical sources because other PAC mostly have B-type interflavan bonds. Cranberry products used to alleviate and prevent urinary tract infections may suffer from adulteration, where c-PAC are replaced with less expensive botanical sources of PAC that contain B-type interflavan bonds. OBJECTIVE: Identifying the presence of A-type interflavan bonds in cranberry fruit and dietary supplements. METHODS: Thirty-five samples reported to contain A-type PAC (cranberry fruit and cranberry products) and 36 samples reported to contain B-type PAC (other botanical sources) were identified and differentiated using MALDI-TOF MS, deconvolution of overlapping isotope patterns, and principal component analysis (PCA). RESULTS: Our results show that both MALDI-TOF MS and deconvolution of overlapping isotope patterns were able to identify the presence of A-type interflavan bonds with a probability greater than 90% and a confidence of 95%. Deconvolution of MALDI-TOF MS spectra also determined the ratio of A-type to B-type interflavan bonds at each degree of polymerization in cranberry fruit and cranberry products, which is a distinguishing feature of c-PAC in comparison to other botanical sources of PAC. PCA shows clear differences based on the nature of the interflavan bonds. CONCLUSIONS: MALDI-TOF MS, deconvolution of overlapping isotope patterns of MALDI-TOF MS spectra, and PCA allow the identification, estimation, and differentiation of A-type interflavan bonds in cranberry-based foods and dietary supplements among other botanical sources containing mostly B-type interflavan bonds.

Inter-Laboratory Validation of 4-(Dimethylamino) Cinnamaldehyde (DMAC) Assay Using Cranberry Proanthocyanidin Standard for Quantification of Soluble Proanthocyanidins in Cranberry Foods and Dietary Supplements, First Action Official MethodSM: 2019.06.

BACKGROUND: Proanthocyanidins (PAC) are oligomers and polymers of flavan-3-ols with putative health benefits. PAC are prevalent in a wide variety of natural products and dietary supplements. OBJECTIVE: An inter-laboratory study was conducted to validate the 4-(dimethylamino)cinnamaldehyde (DMAC) colorimetric assay using a 96-well plate spectrophotometer for the accurate quantification of PAC in cranberry products and to evaluate the comparison of the procyanidin A2 (ProA2) dimer and cranberry PAC (c-PAC) reference standards. METHODS: Four test materials analyzed in this study included cranberry fiber powder, cranberry extract powder, concentrated cranberry juice, and a solution of cranberry PAC (30%, w/v). The samples were homogenized, extracted, sonicated, centrifuged, and analyzed using a 96-well plate spectrophotometer. RESULTS: Linearity for both the ProA2 and c-PAC standards was determined from 4.053 to 50.666 µg/mL and from 13.520 to 135.95 µg/mL, respectively. The relative standard deviation of repeatability (RSDr) values for the four materials analyzed, using both ProA2 and c-PAC standards, met the Standard Method Performance Requirements (SMPR®). Inter-laboratory precision using Horwitz ratio (HorRat) values for the four materials analyzed, using both ProA2 and c-PAC standards, satisfies the acceptance range in Appendix K of the Official Methods of Analysis (2003): Guidelines for Dietary Supplements and Botanicals. The limit of quantification (LOQ) was estimated to be 3.16 µg/mL. CONCLUSIONS: The results produced from this study demonstrate the utility of the c-PAC standard over the ProA2 standard and the advantages of using a 96-well plate spectrophotometer for the accurate quantification of PAC. HIGHLIGHTS: The use of a 96-well plate reader and c-PAC reference standard in the DMAC method improves accuracy and percision for quantification of soluble proanthocyanidins in cranberry foods and dietary supplements.

Synthesis of Fluorescent Proanthocyanidin-Cinnamaldehydes Pyrylium Products for Microscopic Detection of Interactions with Extra-Intestinal Pathogenic Escherichia coli.

Synthesis of proanthocyanidin-cinnamaldehydes pyrylium products (PCPP) was achieved by the condensation reaction of proanthocyanidins (PAC) with cinnamaldehyde and four cinnamaldehyde derivatives. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectra of PCPP show masses that correspond to (epi)catechin oligomers attached to single, double, or triple moieties of cinnamaldehydes. Synthesized PCPP exhibited fluorescence at higher excitation and emission wavelengths than PAC. Results indicate that PCPP were more bioactive for agglutinating extra-intestinal pathogenic Escherichia coli (ExPEC) compared to PAC. Scanning electron microscopy indicates that PCPP interact with ExPEC surface structures and suggests that PCPP have a higher affinity with the fimbriae-like structures of ExPEC than PAC. Fluorescent microscopy performed on in vitro and in vivo agglutination assays show that PCPP were entrapping ExPEC in a web-like network, thus demonstrating agglutination of ExPEC. This study demonstrated the potential of PCPP to improve our understanding of the temporal and dynamic interactions of PAC in in vitro and in vivo studies.

Characterization of proanthocyanidin-chitosan interactions in the formulation of composite nanoparticles using surface plasmon resonance.

Chitosan (CHT) interacts with proanthocyanidins (PAC) by a mechanism involving hydrogen bonding and ion-dipole interactions, allowing the spontaneous formation of PAC-CHT composite nanoparticles (PAC-CHT NPs). The interaction between PAC and CHT was characterized by ellipsometry, infrared spectroscopy, and surface plasmon resonance (SPR) to determine the effect of CHT molecular weight (MW), PAC to CHT ratios, and pH on the formulation of PAC-CHT NPs. These parameters also affect the size and morphology of PAC-CHT NPs. Results indicate that CHT MW and pH of the solution impact the interactions of PAC-CHT in two ways: (1) greater CHT MW increases the amount of PAC molecules that attach to the CHT chain, and (2) lower pH of the CHT solutions increases the amount PAC molecules that attach to the CHT chain. Results also show that higher CHT MW, CHT concentration, and pH of the CHT solutions increase the size of PAC-CHT NPs. In contrast, greater PAC concentrations decreases the size of PAC-CHT NPs. This study demonstrates that SPR is a useful technique for measuring the effect of changes in the interaction between PAC and CHT, which in turn affects the size and morphology of PAC-CHT NPs.

Isolation and Characterization of Blueberry Polyphenolic Components and Their Effects on Gut Barrier Dysfunction.

Highbush blueberries contain anthocyanins and proanthocyanidins that have antimicrobial and anti-inflammatory bioactivities. We isolated and characterized three polyphenolic fractions, a total polyphenol fraction (TPF), an anthocyanin-enriched fraction (AEF), and a proanthocyanidin-enriched fraction (PEF), from freeze-dried blueberry powder and evaluated their effects on an in vitro model of gut barrier dysfunction. High-performance liquid chromatography chromatograms illustrate successful fractionation of the blueberry powder into TPF, AEF, and PEF. AEF contained 21 anthocyanins, and PEF contained proanthocyanidin oligomers of (epi)catechin with primarily B-type interflavan bonds. The model uses a strain of Escherichia coli to disrupt a Caco-2 cell monolayer on Transwell inserts. Barrier function was measured by transepithelial electrical resistance (TEER), a marker of membrane permeability. All fractions were able to restore TEER values after an E. coli challenge when compared to the control, while AEF was able to attenuate the E. coli-induced decrease in TEER in a dose-dependent manner.

Composition of Anthocyanins and Proanthocyanidins in Three Tropical Vaccinium Species from Costa Rica.

Total polyphenol content (TPC), total flavonoid content (TFC), total anthocyanin content (TAC), and proanthocyanidin (PAC) content were determined in fruit from three tropical Vaccinium species (Vaccinium consanguineum, Vaccinium floribundum, and Vaccinium poasanum) from Costa Rica sampled at three stages of fruit development. Results show that TAC increased as the fruit developed, while TPC, TFC, and PAC content decreased. Anthocyanin profiles were evaluated using electrospray ionization tandem mass spectrometry. Cyanidin and delphinidin glycosides were the predominant anthocyanins for the three tropical Vaccinium species. Proanthocyanidins were characterized using attenuated total reflection Fourier transform infrared spectroscopy, nuclear magnetic resonance, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The presence of procyanidin structures with B-type interflavan bonds were observed, but deconvolution of mass spectrometry isotope patterns indicated that PACs with one or more A-type interflavan bonds accounted for more than 74% of the oligomers at each degree of polymerization.

Antimicrobial proanthocyanidin-chitosan composite nanoparticles loaded with gentamicin.

Cranberry proanthocyanidin-chitosan nanoparticles (PAC-CHT NPs) loaded with antibiotic gentamicin (GEN) (PAC-CHT-GEN NPs) were formulated and characterized according to size, polydispersity (PDI), surface charge, morphology, and encapsulation efficiency (EE). PAC-CHT-GEN NPs were evaluated for their ability to agglutinate E. coli, S. aureus, and P. aeruginosa and their bacteriostatic and bactericidal activity. Results indicate that the PAC-CHT-GEN NPs at 0.5:1.0, 1.0:1.0, and 2.0:1.0 weight ratios formed stable nanoparticles with sizes from 242.9 to 277.4 nm, a PDI from 0.344 to 0.391, and a zeta potential from 34.5 to 38.5 mV, and up to 94% EE. Results indicate that PAC-CHT-GEN NPs have the ability to agglutinate E. coli, S. aureus, and P. aeruginosa. Furthermore, PAC-CHT-GEN NPs exhibited greater bactericidal activity than GEN alone. Results suggested PAC-CHT-GEN NPs form stable, round-shaped, and bioactive nanoparticles with the potential to be use in the treatment of bacterial infections.

Confined benzene within InOF-1: contrasting CO2 and SO2 capture behaviours.

The confinement of small amounts of benzene in InOF-1 (Bz@InOF-1) shows a contradictory behavior in the capture of CO2 and SO2. While the capture of CO2 is increased 1.6 times, compared to the pristine material, the capture of SO2 shows a considerable decrease. To elucidate these behaviors, the interactions of CO2 and SO2 with Bz@InOF-1 were studied by DFT periodical calculations postulating a plausible explanation: (a) in the case of benzene and CO2, these molecules do not compete for the preferential adsorption sites within InOF-1, providing a cooperative CO2 capture enhancement and (b) benzene and SO2 strongly compete for these preferential adsorption sites inside the MOF material, reducing the total SO2 capture.

Exposure to methylphenidate in adolescence and adulthood modulates cross-sensitization to amphetamine in adulthood in three genetically variant female rat strains.

Attention Deficit Hyperactivity Disorder (ADHD) is a developmental, behavioral disorder that is characterized by patterns of impulsivity and limited attention. Stimulants, such as methylphenidate (MPD) and amphetamine (Amph), are utilized as first-line agents in the treatment of ADHD. While Amph is known to elicit dependence, MPD is not. Interdependence between MPD and Amph is a growing concern. Therefore, it is important to explore this interaction in animal models to gain insight on its mechanisms. In experimental studies, drugs that elicit behavioral sensitization and cross-sensitization in animals are considered to have the potential to elicit dependence. This study evaluated whether chronic repetitive MPD exposure in adolescence and/or adulthood in various genetic strains of female rats elicits behavioral sensitization as well as cross-sensitization with Amph. We used three strains: SHR (ADHD model), WKY, and SD rats. The three rat strains were exposed to chronic MPD in their adolescence and adulthood or exposed to chronic MPD only in adulthood. When the female rats were treated with MPD only in adulthood, the SHR strain exhibited cross-sensitization to Amph but the WKY and SD strains did not. Whereas, when the female rats were pretreated with MPD in adolescence and adulthood, the SHR and WKY strains exhibited cross-sensitization response to Amph, but the SD strain did not. This study showed that the genetic strain and age of exposure to MPD plays a crucial role in cross-sensitization to Amph in female rodents. Furthermore, we showed that genetics associated with ADHD pre-dispose animals to dependence between drugs even if the exposure starts in adulthood compared to control strains that did not show dependence with adult only MPD exposures. Genetic variability, age of initial drug exposure, and sex of the subject are key variables that should be accounted in studies that explore effects of psychostimulants.

Proanthocyanidin-chitosan composite nanoparticles prevent bacterial invasion and colonization of gut epithelial cells by extra-intestinal pathogenic Escherichia coli.

Cranberry proanthocyanidin-chitosan composite nanoparticles (PAC-CHT NPs) were formulated using 2:1, 5:1, 10:1, 15:1 20:1, 25:1, and 30:1 PAC to CHT weight ratio to form round shaped particles. The PAC-CHT NPs were characterized by size, polydispersity, surface charge, morphology, and PAC content. PAC-CHT NPs bioactivity was measured by agglutination of extra-intestinal pathogenic Escherichia coli (ExPEC) and inhibition of gut epithelial cell invasion by ExPEC. Results indicate that by increasing the PAC to CHT ratio 10:1 to 30:1 formed stable nanoparticles with diameters of 122.8 to 618.7 nm, a polydispersity index of approximated 0.4 to 0.5, and a zeta potential of 34.5 to 54.4 mV. PAC-CHT NPs ratio 30:1 agglutinated ExPEC and decreased the ability of ExPEC to invade epithelial cells in a dose-dependent manner. PAC-CHT NPs ratio 10:1 to 30:1 form stable, round-shaped, and bioactive nanoparticles for potential applications in the treatment of ExPEC bacterial infections.

Confined toluene within InOF-1: CO2 capture enhancement.

The toluene adsorption properties of InOF-1 are studied along with the confinement of small amounts of this non-polar molecule revealing a 1.38-fold increase in CO2 capture, from 5.26 wt% under anhydrous conditions to 7.28 wt% with a 1.5 wt% of pre-confined toluene at 298 K. The InOF-1 affinity towards toluene was experimentally quantified by ΔH ads (-46.81 kJ mol-1). InOF-1 is shown to be a promising material for CO2 capture under industrial conditions. Computational calculations (DFT and QTAIM) and DRIFTs in situ experiments provided a possible explanation for the experimental CO2 capture enhancement by showing how the toluene molecule is confined within InOF-1, which constructed a "bottleneck effect".

Electrospun plant mucilage nanofibers as biocompatible scaffolds for cell proliferation.

Electrospun nanofibers (ESNFs) were prepared from mucilage isolated from chan and linaza beans and mozote stem commercially available in Costa Rica. Poly(vinyl alcohol) (PVA) was used as an aiding agent. Mucilage/PVA mixed solutions of different volume ratios (100:0, 80:20, 60:40, 40:60, 20:80 and 0:100) were prepared and adjusted to be similar in viscosity and electrical conductivity suitable for electrospinning. Morphology of the ESNFs was examined using scanning electron microscopy (SEM). Fourier transform infrared spectrometer (FTIR) and differential scanning calorimetry (DSC) studies were used to characterize chemical composition and thermal characteristics of the nanofibers (NFs). The ability of the NFs to support fibroblast cell proliferation was investigated in vitro using the optimized mucilage/PVA solutions. Results show plant mucilage-based ESNFs are well-suited for fibroblast cell growth, significantly better than ESNFs of PVA; and the mucilage of chan beans is better than those of mozote and linaza for supporting cell proliferation.

Cranberry proanthocyanidin-chitosan hybrid nanoparticles as a potential inhibitor of extra-intestinal pathogenic Escherichia coli invasion of gut epithelial cells.

Chitosan interacts with proanthocyanidins through hydrogen-bonding, which allows encapsulation and development of stable nanoparticles via ionotropic gelation. Cranberry proanthocyanidins (PAC) are associated with the prevention of urinary tract infections and PAC inhibit invasion of gut epithelial cells by extra-intestinal pathogenic Escherichia coli (ExPEC). We determined the effect of cranberry proanthocyanidin-chitosan hybrid nanoparticles (PAC-CHTNp) on the ExPEC invasion of gut epithelial cells in vitro. PAC-CHTNp were characterized according to size, morphology, and bioactivity. Results showed a decrease in the size of the nanoparticles as the concentration of PAC was increased, indicating that PAC increases cross-linking by hydrogen-bonding on the surface of the chitosan nanoparticles. Nanoparticles were produced with diameters ranging from 367.3 nm to 293.2 nm. Additionally, PAC-CHTNp significantly inhibited the ability of ExPEC to invade the enterocytes by ~80% at 66 µg GAE/mL and by ~92% at 100 µg GAE/mL. Results also indicate that chitosan nanoparticles alone were not significantly different from controls in preventing ExPEC invasion of enterocytes (data not shown) and also there were not significant differences between PAC alone and PAC-CHTNp, suggesting that the new PAC-CHTNp could lead to an increase in the stability of encapsulated PAC, maintain the molecular adhesion of PAC to ExPEC.