Background: We have previously reported that the addition of resistant maltodextrin (RMD), a ferm... more Background: We have previously reported that the addition of resistant maltodextrin (RMD), a fermentable functional fiber, to the diet increases fecal weight as well as the amount of fecal bifidobacteria. Here, we report on the targeted analysis of changes in potentially beneficial gut bacteria associated with the intervention. Objective: The primary objective of this study was to determine the effect of adding 0, 15 and 25 g RMD to the diets of healthy free-living adults on potentially beneficial gut bacteria. Methods: We expanded on our previously reported microbiota analysis in a double-blind, placebo-controlled feeding study (NCT02733263) by performing additional qPCR analyses targeting fecal lactic acid bacteria (LAB), Akkermansia muciniphila, Faecalibacterium prausnitzii and Fusicatenibacter saccharivorans in samples from 49 participants. Results: RMD resulted in an approximately two-fold increase in fecal Fusicatenibacter saccharivorans (p = 0.024 for 15 g/day RMD and p = 0.0...
In healthy adults resistant maltodextrin produces a greater change in fecal bifidobacteria counts... more In healthy adults resistant maltodextrin produces a greater change in fecal bifidobacteria counts and increases stool wet weight: a double-blind, randomized, controlled crossover study. Ntr (2018),
Studies have been made on the binding of 125I-glucagon by isolated chick hepatocytes. It was show... more Studies have been made on the binding of 125I-glucagon by isolated chick hepatocytes. It was shown that pH and temperature dependence of the binding does not differ from that in rat hepatocytes. Optimum binding was observed at pH 7.6, the rate of binding being higher at 37 degrees C as compared to that at 20 degrees C, although the binding capacity increased with the decrease in the temperature. Unlabeled glucagon was able to compete with 125I-glucagon at the binding sites. Scatchard plot was found to be curvilinear revealing two classes of the binding sites with Kd values 10(-9) and 10(-7) M at temperatures 20 and 37 degrees C correspondingly. Earlier studies revealed in rats the binding sites of a sole class with Kd value 10(-9) M. Preincubation of cells with native glucagon results in changes of labeled glucagon binding, the effect being proportional to the concentration of native glucagon. Preincubation effect was observed at 37 degrees C, being absent at 20 degrees C; the effec...
Background: Latent tuberculosis infection (LTBI) treatment is an effective strategy to eliminate ... more Background: Latent tuberculosis infection (LTBI) treatment is an effective strategy to eliminate TB in low-incidence settings. Shorter LTBI regimens incorporating the antimicrobial class of rifamycins are designed to improve treatment completion rates. Recent evidence suggests that the rifamycins could induce irreversible gut microbiota changes that impact future anti-TB immunity. Methods: To document the immediate effect of the rifamycins on the gut microbiota, we followed six patients with LTBI initiating four months of monotherapy with rifampin (4R; n=4) or three months of rifapentine in combination with isoniazid (3HP; n=2) and tracked recovery to baseline two months posttreatment completion. We collected stool samples parallel to the LTBI group from healthy volunteers (N=6) unexposed to the rifamycins. We used a questionnaire to collect diet, antibiotics, and lifestyle changes during follow-up. We profiled the gut microbiota using 16S rRNA amplicon sequencing (V1-V2 region). Re...
Tuberculosis (TB) preventive therapy (TPT) is an effective strategy to eliminate TB in low-incide... more Tuberculosis (TB) preventive therapy (TPT) is an effective strategy to eliminate TB in low-incidence settings. Shorter TPT regimens incorporating the antimicrobial class of rifamycins are designed to improve adherence and completion rates but carry the risk of modifications to the gut microbiota. We enrolled six subjects diagnosed with latent TB infection (LTBI) who accepted to initiate TPT. We also enrolled six healthy volunteers unexposed to the rifamycins. We profiled the gut microbiota using 16S rRNA amplicon sequencing (V1-V2 region) to document the immediate effect of rifamycin-based TPT on the gut microbiota composition and tracked recovery to baseline two months after TPT. Overall, TPT accounted for 17% of the variance in gut microbial community dissimilarity. This rifamycin-based TPT induced dysbiosis was characterized by a depletion of butyrate-producing taxa (Clostridium-XIVa and Roseburia) and expansion of potentially pathogenic taxa within the Firmicutes and Proteobacte...
Purpose: The purpose of this pilot study was to assess the differences in breast tissue microbiot... more Purpose: The purpose of this pilot study was to assess the differences in breast tissue microbiota composition by breast side (left versus right) within an individual woman and compare the microbiota of healthy and breast tumor tissue between women. We further aimed to determine whether certain bacterial taxa may be associated with breast tumors. Methods: Bilateral healthy breast tissue samples (n=36) were collected from ten women who received routine mammoplasty procedures at the University of Florida Department of Surgery. Archived breast tumor samples (n=10) were obtained from an established biorepository. Bacterial DNA was extracted from tissues, amplified via polymerase chain reaction (PCR), and sequenced on the Illumina MiSeq platform. Microbiota data were analyzed using QIIME and RStudio. Results: The most abundant phyla in both tumor and healthy tissues were Bacteroidetes, Firmicutes, Proteobacteria, and Actinobacteria. A total of 38 operational taxonomic units (OTUs) were found to be significantly different in terms of differential abundance between tumor and healthy tissues (absolute effect size range: 0.761-3.98). The OTUs with the largest absolute effect size associated with higher relative abundance in breast tumors were of Flavobacterium species (R2=3.98), Acinetobacter species (R2=3.64), [Mogibacteriaceae] family (R2=3.34), and Clostridiales order (R2=3.21). Alpha diversity (Shannon Diversity Index) was similar in healthy and tumor tissue (4.98 vs. 4.84; p=0.350). Based on unweighted UniFrac measures, breast tumor samples clustered distinctly from healthy samples (R2=0.10; p=0.001). Microbiota composition in healthy samples clustered within women (R2=0.20; p=0.012) and by breast side (left or right) within a woman (R2=0.36; p=0.001). Conclusion: We identified significant differences in microbiota composition between women and between breasts of the same woman. These results warrant further investigation to elucidate the potential relationship between breast tissue microbiota and breast cancer. Citation Format: Emily M. Klann, Jessica M. Williamson, Massimiliano S. Tagliamonte, Maria Ukhanova, Jaya Ruth Asirvatham, Harvey Chim, Lusine Yaghjyan, Volker Mai. Microbiota composition in bilateral healthy breast tissue and breast tumors [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr B31.
Peripheral immunity is thought to be dysregulated in Parkinson's disease (PD) and may provide an ... more Peripheral immunity is thought to be dysregulated in Parkinson's disease (PD) and may provide an avenue for novel immunotherapeutic interventions. Gut microbiota is a potential factor for modulating immunotherapy response. Considering the possibly complex role of the gut-brain axis in PD, we used a preclinical model to determine the effects of gut microbiota dynamics in mice receiving an immunotherapeutic intervention compared to controls. A total of 17 M83 heterozygous transgenic mice were used in this study. Mice in the treatment arm (N=10) received adoptive cellular therapy (ACT) by injection and control mice (N=7) were injected with saline at 8 weeks of age. All mice received peripheral α-syn fibrils to hasten parkinsonian symptoms via an intramuscular injection one week later (9 weeks of age; baseline). Fecal pellets were collected from all mice at three time points post injection (baseline, 6 weeks and 12 weeks). DNA from each stool sample was extracted, 16S rDNA amplified and sequenced, and analyzed using QIIME2 and RStudio. Differences in the relative abundance of bacterial taxa were observed over time between groups. No significant differences in alpha diversity were found between groups at any timepoint. UniFrac measures of phylogenetic distance between samples demonstrated distinct clustering between groups post-baseline (p=0.002). These differences suggest that the gut microbiome may be capable of influencing immunotherapy outcomes. Conclusively, we observed distinctly different microbiota dynamics in treated mice compared to those in the control group. These results suggest a correlation between the gut-brain axis, PD pathology, and immunotherapy.
Improvements in high-throughput sequencing technologies have spurred a large number of studies ai... more Improvements in high-throughput sequencing technologies have spurred a large number of studies aimed at obtaining a better understanding of the composition and the dynamics in gut microbiota and its associations with various human diseases, especially those in the intestinal tract. Here we briefly summarize results from three different such studies from our group, all of which used 454 based highthroughput 16S rRNA sequence analysis combined with other microbiota profiling methods to determine faecal microbiota composition. In the first study, a controlled feeding trial, we establish that energy gain from the consumption of up to 50 g/day of a resistant maltodextrin depends on the prevalent microbiota composition. Over time, resistant maltodextrin supplementation increased the proportion of total faecal bacteria as well as potentially beneficial bifidobacteria. Thus, energy gain from resistant maltodextrin in an individual appears to vary over time and depend on the adaptation of gut microbiota. We then illustrate the power of molecular tools for identifying (i) distortions in early microbiota development in pre-term infants and the presence of potentially novel pathogens contributing to necrotizing enterocolitis and (ii) a specific microbiota signature, based on discriminant analysis of the 16S rRNA sequences, that correlates with the prevalence of an early risk marker associated with colorectal carcinogenesis, intestinal adenoma, in elderly adults.
bioRxiv (Cold Spring Harbor Laboratory), Jan 25, 2020
The incidence of locally acquired dengue infections increased during the last decade in the Unite... more The incidence of locally acquired dengue infections increased during the last decade in the United States, compelling a sustained research effort on the dengue mosquito vector, Aedes aegypti, and its microbiome, which has been shown to influence virus transmission success. We examined the 'metavirome' of four populations of Ae. aegypti mosquitoes collected in 2016-2017 from Manatee County, Florida. Unexpectedly, we discovered that dengue virus serotype 4 (DENV4) was circulating in these mosquito populations, representing the first documented case of such a phenomenon in the absence of a local DENV4 human case in this county over a two-year period. We confirmed that all of the mosquito populations carried the same DENV4 strain, assembled its full genome, validated infection orthogonally by reverse transcriptase PCR, traced the virus origin, estimated the time period of its introduction to the Caribbean region, as well as explored the viral genetic signatures and mosquito-specific virome associations that potentially mediated DENV4 persistence in mosquitoes. We discuss
Next generation amplicon sequencing has created a plethora of data from human microbiomes. The ac... more Next generation amplicon sequencing has created a plethora of data from human microbiomes. The accessibility to this scientific data and its corresponding metadata is important for its reuse, to allow for new discoveries, verification of published results, and serving as path for reproducibility. Dietary fiber consumption has been associated with a variety of health benefits that are thought to be mediated by gut microbiota. To enable direct comparisons of the response of the gut microbiome to fiber, we obtained 16S rRNA sequencing data and its corresponding metadata from 11 fiber intervention studies for a total of 2,368 samples. We provide curated and pre-processed genetic data and common metadata for comparison across the different studies.
Introduction: Non-motor symptoms of Parkinson's disease (PD) such as gastrointestinal (GI) dy... more Introduction: Non-motor symptoms of Parkinson's disease (PD) such as gastrointestinal (GI) dysfunction are common, yet little is known about how modifying dietary intake impacts PD symptoms. The aim of this study in individuals with PD was to determine whether a Mediterranean diet intervention is feasible and affects GI function, intestinal permeability and fecal microbial communities.Methods: A single-arm, 5-week Mediterranean diet intervention study was conducted in eight people with PD. Daily and weekly questionnaires were administered to determine changes in GI symptoms. Urine and stool samples were collected at baseline and after 5 weeks to assess intestinal permeability and fecal microbial communities. Additionally, live-in partners of the participants with PD were matched as controls (n = 8) for baseline urine and stool samples.Results: Participants with PD increased intake of Mediterranean diet based on adherence scores from baseline to week 5 (4.4 ± 0.6 vs. 11.9 ± 0.7; ...
Probiotic supplements can contribute to maintaining health and ameliorating various disease sympt... more Probiotic supplements can contribute to maintaining health and ameliorating various disease symptoms. Probiotics can be delivered in many forms with crucial differences in their survival during gastrointestinal (GI) passage. Previously, a novel encapsulation, Probiotic Pearls™ Acidophilus, Integrative Therapeutics, LLC, USA (Pearls), was shown to increase survival in vitro after exposure to gastric conditions. Here, we compare fecal recovery in human volunteers consuming Pearls or a conventional hard-shelled gelatin capsule. We performed a randomized double-blinded, two-armed trial, with six healthy subjects in each 12-day study arm. In fecal samples collected at baseline, twice during the intervention period, and after washout, we compared colony counts between the two encapsulation methods. The identity of the colonies was confirmed by colony morphology, strainspecific PCR, and 16S rRNA gene sequencing. We further performed a comprehensive 16S rRNA gene sequencingbased analysis to identify differential effects on overall microbiota composition. We detected an average log increase in bifidobacteria of 0.152 cfu/g with gelatin and 0.651 cfu/g with Pearls capsules (p > 0.05). Total lactobacilli counts increased in both groups with no difference between the groups. However, the supplemented Lactobacillus acidophilus NCFM decreased to baseline levels within 7 days after end of supplementation with gelatin capsules while 3.11 log cfu/g higher counts compared to baseline (p = 0.05) remained for Pearls. Targeted qPCR largely confirmed the trends observed by viable plate counts. Protecting the probiotic strains by Pearls encapsulation results in higher recovery rates of the supplemented lactobacilli and bifidobacteria in fecal samples and increased persistence, suggesting an improved survival and viability that might increase efficacy towards achieving desired health benefits.
The aim of the studies was to determine the effects of calcium carbonate and calcium phosphate su... more The aim of the studies was to determine the effects of calcium carbonate and calcium phosphate supplementation on faecal Lactobacillus spp., with and without a probiotic supplement, in healthy adults. Study 1 comprised of a randomised, double-blind, crossover design; participants (n=15) received 2 capsules/d of 250 mg elemental calcium as calcium carbonate (Ca1) and calcium phosphate (Ca2) each for 2-week periods, with 2-week baseline and washout periods. Study 2 was a randomised, double-blind, crossover design; participants (n=17) received 2 capsules/d of Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011 (probiotic) alone, the probiotic with 2 capsules/d of Ca1, and probiotic with 2 capsules/d of Ca2 each for 2-week periods with 2-week baseline and washout periods. In both studies, stools were collected during the baseline, intervention and washout periods for Lactobacillus spp. quantification and qPCR analyses. Participants completed daily questionnaires of stool fr...
Angewandte Chemie (International ed. in English), Jan 20, 2015
Conventional antibiotics are ineffective against non-replicating bacteria (for example, bacteria ... more Conventional antibiotics are ineffective against non-replicating bacteria (for example, bacteria within biofilms). We report a series of halogenated phenazines (HP), inspired by marine antibiotic 1, that targets persistent bacteria. HP 14 demonstrated the most potent biofilm eradication activities to date against MRSA, MRSE, and VRE biofilms (MBEC=0.2-12.5 μM), as well as the effective killing of MRSA persister cells in non-biofilm cultures. Frontline MRSA treatments, vancomycin and daptomycin, were unable to eradicate MRSA biofilms or non-biofilm persisters alongside 14. HP 13 displayed potent antibacterial activity against slow-growing M. tuberculosis (MIC=3.13 μM), the leading cause of death by bacterial infection around the world. HP analogues effectively target persistent bacteria through a mechanism that is non-toxic to mammalian cells and could have a significant impact on treatments for chronic bacterial infections.
Purpose: To determine which dietary factors contribute to microbiota variation and identify corre... more Purpose: To determine which dietary factors contribute to microbiota variation and identify correlations with gut epithelial methylation pattern associated with the presence of high-risk polyps. Dietary exposures have long been suggested as important risk factors for cancers, especially those of the gastrointestinal tract. Diet is known to affect the commensal gut microbiota, which contributes important activities to the human host that are required for maintaining normal health. However, distortions in microbiota composition and associated microbial activities might contribute to disease processes including colorectal carcinogenesis. High-throughput sequencing based microbiota studies have revealed a large degree of intra- and inter-individual variation, the sources of which are currently not well understood. As part of this project we explored various ‘Big Data’ approaches that can facilitate an efficient mining of the large dataset generated by combining dietary intake data with ...
A probiotic formulation of Enterococcus faecium R0026 and Bacillus subtilis R0179 has been evalua... more A probiotic formulation of Enterococcus faecium R0026 and Bacillus subtilis R0179 has been evaluated in previous clinical trials. However, B. subtilis R0179 has not been evaluated as a single probiotic strain or in combination with other strains at doses higher than 0.1×109 cfu. To establish oral dose-response tolerance and gastrointestinal (GI) viability of B. subtilis R0179, a randomised, double-blind, placebo-controlled trial in healthy adults (n=81; 18-50 years old) was conducted. Participants received B. subtilis R0179 at 0.1, 1.0 or 10×109 cfu/capsule/day or placebo for four weeks. General wellness was assessed using a daily questionnaire evaluating GI, cephalic, ear-nose-throat, behavioural, emetic, and epidermal symptoms. GI symptoms were further evaluated using a weekly gastrointestinal symptom rating scale (GSRS). GI transit viability of B. subtilis R0179 was assessed by plating and microbiota analysis by 16S rRNA at baseline, week 4 of the intervention and washout. Genera...
Humans harbor distinct commensal microbiota at various anatomic sites. There has been renewed int... more Humans harbor distinct commensal microbiota at various anatomic sites. There has been renewed interest in the contributions of microbiota activities to human health and disease. The microbiota of the gut is the most complex of all anatomic sites in terms of total numbers of bacteria that interact closely with the mucosal immune system and contribute various functions to host physiology. Especially in the proximal large intestine a diverse microbiota ferments complex substrates such as dietary fiber and host mucins, but also metabolizes bile acids and phytoestrogens that reach the large intestine. It is now well established that microbiota composition differs between but over time also within individuals. However, a thorough understanding of the sources of variations in microbiota composition, which is an important requirement for large population based microbiota studies is lacking. Microbiota composition varies depending on what kind of sample is collected, most commonly stool samples, stool swabs or superficial rectal or intestinal biopsies, and the time of collection. Microbiota dynamics are affected by life style factors including diet and exercise that determine what nutrients reach the proximal colon and how fast these nutrients pass through (transit time). Here we review sample collection issues in gut microbiota studies and recent findings about dynamics in microbiota composition. We recommend standardizing human microbiota analysis methods to facilitate comparison and
Late onset sepsis (LOS) is a major contributor to neonatal morbidity and mortality, especially in... more Late onset sepsis (LOS) is a major contributor to neonatal morbidity and mortality, especially in premature infants. Distortions in the establishment of normal gut microbiota, commensal microbes that colonize the digestive tract, might increase the risk of LOS via disruption of the mucosal barrier with resultant translocation of luminal contents. Correlation of distortions of the intestinal microbiota with LOS is a necessary first step to design novel microbiota-based screening approaches that might lead to early interventions to prevent LOS in high risk infants. Using a case/control design nested in a cohort study of preterm infants, we analyzed stool samples that had been prospectively collected from ten preterm infants with LOS and from 18 matched controls. A 16S rRNA based approach was utilized to compare microbiota diversity and identify specific bacterial signatures that differed in their prevalence between cases and controls. Overall a-diversity (Chao1) was lower in cases two weeks before (p,0.05) but not one week before or at the time of diagnosis of LOS. Overall microbiota structure (Unifrac) appeared distinct in cases 2 weeks and 1 week before but not at diagnosis (p,0.05). Although we detected few operational taxonomic units (OTUs) unique or enriched in cases, we found many OTUs common in controls that were lacking in cases (p,0.01). Bifidobacteria counts were lower in cases at all time points. Our results support the hypothesis that a distortion in normal microbiota composition, and not an enrichment of potential pathogens, is associated with LOS in preterm infants.
Background: We have previously reported that the addition of resistant maltodextrin (RMD), a ferm... more Background: We have previously reported that the addition of resistant maltodextrin (RMD), a fermentable functional fiber, to the diet increases fecal weight as well as the amount of fecal bifidobacteria. Here, we report on the targeted analysis of changes in potentially beneficial gut bacteria associated with the intervention. Objective: The primary objective of this study was to determine the effect of adding 0, 15 and 25 g RMD to the diets of healthy free-living adults on potentially beneficial gut bacteria. Methods: We expanded on our previously reported microbiota analysis in a double-blind, placebo-controlled feeding study (NCT02733263) by performing additional qPCR analyses targeting fecal lactic acid bacteria (LAB), Akkermansia muciniphila, Faecalibacterium prausnitzii and Fusicatenibacter saccharivorans in samples from 49 participants. Results: RMD resulted in an approximately two-fold increase in fecal Fusicatenibacter saccharivorans (p = 0.024 for 15 g/day RMD and p = 0.0...
In healthy adults resistant maltodextrin produces a greater change in fecal bifidobacteria counts... more In healthy adults resistant maltodextrin produces a greater change in fecal bifidobacteria counts and increases stool wet weight: a double-blind, randomized, controlled crossover study. Ntr (2018),
Studies have been made on the binding of 125I-glucagon by isolated chick hepatocytes. It was show... more Studies have been made on the binding of 125I-glucagon by isolated chick hepatocytes. It was shown that pH and temperature dependence of the binding does not differ from that in rat hepatocytes. Optimum binding was observed at pH 7.6, the rate of binding being higher at 37 degrees C as compared to that at 20 degrees C, although the binding capacity increased with the decrease in the temperature. Unlabeled glucagon was able to compete with 125I-glucagon at the binding sites. Scatchard plot was found to be curvilinear revealing two classes of the binding sites with Kd values 10(-9) and 10(-7) M at temperatures 20 and 37 degrees C correspondingly. Earlier studies revealed in rats the binding sites of a sole class with Kd value 10(-9) M. Preincubation of cells with native glucagon results in changes of labeled glucagon binding, the effect being proportional to the concentration of native glucagon. Preincubation effect was observed at 37 degrees C, being absent at 20 degrees C; the effec...
Background: Latent tuberculosis infection (LTBI) treatment is an effective strategy to eliminate ... more Background: Latent tuberculosis infection (LTBI) treatment is an effective strategy to eliminate TB in low-incidence settings. Shorter LTBI regimens incorporating the antimicrobial class of rifamycins are designed to improve treatment completion rates. Recent evidence suggests that the rifamycins could induce irreversible gut microbiota changes that impact future anti-TB immunity. Methods: To document the immediate effect of the rifamycins on the gut microbiota, we followed six patients with LTBI initiating four months of monotherapy with rifampin (4R; n=4) or three months of rifapentine in combination with isoniazid (3HP; n=2) and tracked recovery to baseline two months posttreatment completion. We collected stool samples parallel to the LTBI group from healthy volunteers (N=6) unexposed to the rifamycins. We used a questionnaire to collect diet, antibiotics, and lifestyle changes during follow-up. We profiled the gut microbiota using 16S rRNA amplicon sequencing (V1-V2 region). Re...
Tuberculosis (TB) preventive therapy (TPT) is an effective strategy to eliminate TB in low-incide... more Tuberculosis (TB) preventive therapy (TPT) is an effective strategy to eliminate TB in low-incidence settings. Shorter TPT regimens incorporating the antimicrobial class of rifamycins are designed to improve adherence and completion rates but carry the risk of modifications to the gut microbiota. We enrolled six subjects diagnosed with latent TB infection (LTBI) who accepted to initiate TPT. We also enrolled six healthy volunteers unexposed to the rifamycins. We profiled the gut microbiota using 16S rRNA amplicon sequencing (V1-V2 region) to document the immediate effect of rifamycin-based TPT on the gut microbiota composition and tracked recovery to baseline two months after TPT. Overall, TPT accounted for 17% of the variance in gut microbial community dissimilarity. This rifamycin-based TPT induced dysbiosis was characterized by a depletion of butyrate-producing taxa (Clostridium-XIVa and Roseburia) and expansion of potentially pathogenic taxa within the Firmicutes and Proteobacte...
Purpose: The purpose of this pilot study was to assess the differences in breast tissue microbiot... more Purpose: The purpose of this pilot study was to assess the differences in breast tissue microbiota composition by breast side (left versus right) within an individual woman and compare the microbiota of healthy and breast tumor tissue between women. We further aimed to determine whether certain bacterial taxa may be associated with breast tumors. Methods: Bilateral healthy breast tissue samples (n=36) were collected from ten women who received routine mammoplasty procedures at the University of Florida Department of Surgery. Archived breast tumor samples (n=10) were obtained from an established biorepository. Bacterial DNA was extracted from tissues, amplified via polymerase chain reaction (PCR), and sequenced on the Illumina MiSeq platform. Microbiota data were analyzed using QIIME and RStudio. Results: The most abundant phyla in both tumor and healthy tissues were Bacteroidetes, Firmicutes, Proteobacteria, and Actinobacteria. A total of 38 operational taxonomic units (OTUs) were found to be significantly different in terms of differential abundance between tumor and healthy tissues (absolute effect size range: 0.761-3.98). The OTUs with the largest absolute effect size associated with higher relative abundance in breast tumors were of Flavobacterium species (R2=3.98), Acinetobacter species (R2=3.64), [Mogibacteriaceae] family (R2=3.34), and Clostridiales order (R2=3.21). Alpha diversity (Shannon Diversity Index) was similar in healthy and tumor tissue (4.98 vs. 4.84; p=0.350). Based on unweighted UniFrac measures, breast tumor samples clustered distinctly from healthy samples (R2=0.10; p=0.001). Microbiota composition in healthy samples clustered within women (R2=0.20; p=0.012) and by breast side (left or right) within a woman (R2=0.36; p=0.001). Conclusion: We identified significant differences in microbiota composition between women and between breasts of the same woman. These results warrant further investigation to elucidate the potential relationship between breast tissue microbiota and breast cancer. Citation Format: Emily M. Klann, Jessica M. Williamson, Massimiliano S. Tagliamonte, Maria Ukhanova, Jaya Ruth Asirvatham, Harvey Chim, Lusine Yaghjyan, Volker Mai. Microbiota composition in bilateral healthy breast tissue and breast tumors [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr B31.
Peripheral immunity is thought to be dysregulated in Parkinson's disease (PD) and may provide an ... more Peripheral immunity is thought to be dysregulated in Parkinson's disease (PD) and may provide an avenue for novel immunotherapeutic interventions. Gut microbiota is a potential factor for modulating immunotherapy response. Considering the possibly complex role of the gut-brain axis in PD, we used a preclinical model to determine the effects of gut microbiota dynamics in mice receiving an immunotherapeutic intervention compared to controls. A total of 17 M83 heterozygous transgenic mice were used in this study. Mice in the treatment arm (N=10) received adoptive cellular therapy (ACT) by injection and control mice (N=7) were injected with saline at 8 weeks of age. All mice received peripheral α-syn fibrils to hasten parkinsonian symptoms via an intramuscular injection one week later (9 weeks of age; baseline). Fecal pellets were collected from all mice at three time points post injection (baseline, 6 weeks and 12 weeks). DNA from each stool sample was extracted, 16S rDNA amplified and sequenced, and analyzed using QIIME2 and RStudio. Differences in the relative abundance of bacterial taxa were observed over time between groups. No significant differences in alpha diversity were found between groups at any timepoint. UniFrac measures of phylogenetic distance between samples demonstrated distinct clustering between groups post-baseline (p=0.002). These differences suggest that the gut microbiome may be capable of influencing immunotherapy outcomes. Conclusively, we observed distinctly different microbiota dynamics in treated mice compared to those in the control group. These results suggest a correlation between the gut-brain axis, PD pathology, and immunotherapy.
Improvements in high-throughput sequencing technologies have spurred a large number of studies ai... more Improvements in high-throughput sequencing technologies have spurred a large number of studies aimed at obtaining a better understanding of the composition and the dynamics in gut microbiota and its associations with various human diseases, especially those in the intestinal tract. Here we briefly summarize results from three different such studies from our group, all of which used 454 based highthroughput 16S rRNA sequence analysis combined with other microbiota profiling methods to determine faecal microbiota composition. In the first study, a controlled feeding trial, we establish that energy gain from the consumption of up to 50 g/day of a resistant maltodextrin depends on the prevalent microbiota composition. Over time, resistant maltodextrin supplementation increased the proportion of total faecal bacteria as well as potentially beneficial bifidobacteria. Thus, energy gain from resistant maltodextrin in an individual appears to vary over time and depend on the adaptation of gut microbiota. We then illustrate the power of molecular tools for identifying (i) distortions in early microbiota development in pre-term infants and the presence of potentially novel pathogens contributing to necrotizing enterocolitis and (ii) a specific microbiota signature, based on discriminant analysis of the 16S rRNA sequences, that correlates with the prevalence of an early risk marker associated with colorectal carcinogenesis, intestinal adenoma, in elderly adults.
bioRxiv (Cold Spring Harbor Laboratory), Jan 25, 2020
The incidence of locally acquired dengue infections increased during the last decade in the Unite... more The incidence of locally acquired dengue infections increased during the last decade in the United States, compelling a sustained research effort on the dengue mosquito vector, Aedes aegypti, and its microbiome, which has been shown to influence virus transmission success. We examined the 'metavirome' of four populations of Ae. aegypti mosquitoes collected in 2016-2017 from Manatee County, Florida. Unexpectedly, we discovered that dengue virus serotype 4 (DENV4) was circulating in these mosquito populations, representing the first documented case of such a phenomenon in the absence of a local DENV4 human case in this county over a two-year period. We confirmed that all of the mosquito populations carried the same DENV4 strain, assembled its full genome, validated infection orthogonally by reverse transcriptase PCR, traced the virus origin, estimated the time period of its introduction to the Caribbean region, as well as explored the viral genetic signatures and mosquito-specific virome associations that potentially mediated DENV4 persistence in mosquitoes. We discuss
Next generation amplicon sequencing has created a plethora of data from human microbiomes. The ac... more Next generation amplicon sequencing has created a plethora of data from human microbiomes. The accessibility to this scientific data and its corresponding metadata is important for its reuse, to allow for new discoveries, verification of published results, and serving as path for reproducibility. Dietary fiber consumption has been associated with a variety of health benefits that are thought to be mediated by gut microbiota. To enable direct comparisons of the response of the gut microbiome to fiber, we obtained 16S rRNA sequencing data and its corresponding metadata from 11 fiber intervention studies for a total of 2,368 samples. We provide curated and pre-processed genetic data and common metadata for comparison across the different studies.
Introduction: Non-motor symptoms of Parkinson's disease (PD) such as gastrointestinal (GI) dy... more Introduction: Non-motor symptoms of Parkinson's disease (PD) such as gastrointestinal (GI) dysfunction are common, yet little is known about how modifying dietary intake impacts PD symptoms. The aim of this study in individuals with PD was to determine whether a Mediterranean diet intervention is feasible and affects GI function, intestinal permeability and fecal microbial communities.Methods: A single-arm, 5-week Mediterranean diet intervention study was conducted in eight people with PD. Daily and weekly questionnaires were administered to determine changes in GI symptoms. Urine and stool samples were collected at baseline and after 5 weeks to assess intestinal permeability and fecal microbial communities. Additionally, live-in partners of the participants with PD were matched as controls (n = 8) for baseline urine and stool samples.Results: Participants with PD increased intake of Mediterranean diet based on adherence scores from baseline to week 5 (4.4 ± 0.6 vs. 11.9 ± 0.7; ...
Probiotic supplements can contribute to maintaining health and ameliorating various disease sympt... more Probiotic supplements can contribute to maintaining health and ameliorating various disease symptoms. Probiotics can be delivered in many forms with crucial differences in their survival during gastrointestinal (GI) passage. Previously, a novel encapsulation, Probiotic Pearls™ Acidophilus, Integrative Therapeutics, LLC, USA (Pearls), was shown to increase survival in vitro after exposure to gastric conditions. Here, we compare fecal recovery in human volunteers consuming Pearls or a conventional hard-shelled gelatin capsule. We performed a randomized double-blinded, two-armed trial, with six healthy subjects in each 12-day study arm. In fecal samples collected at baseline, twice during the intervention period, and after washout, we compared colony counts between the two encapsulation methods. The identity of the colonies was confirmed by colony morphology, strainspecific PCR, and 16S rRNA gene sequencing. We further performed a comprehensive 16S rRNA gene sequencingbased analysis to identify differential effects on overall microbiota composition. We detected an average log increase in bifidobacteria of 0.152 cfu/g with gelatin and 0.651 cfu/g with Pearls capsules (p > 0.05). Total lactobacilli counts increased in both groups with no difference between the groups. However, the supplemented Lactobacillus acidophilus NCFM decreased to baseline levels within 7 days after end of supplementation with gelatin capsules while 3.11 log cfu/g higher counts compared to baseline (p = 0.05) remained for Pearls. Targeted qPCR largely confirmed the trends observed by viable plate counts. Protecting the probiotic strains by Pearls encapsulation results in higher recovery rates of the supplemented lactobacilli and bifidobacteria in fecal samples and increased persistence, suggesting an improved survival and viability that might increase efficacy towards achieving desired health benefits.
The aim of the studies was to determine the effects of calcium carbonate and calcium phosphate su... more The aim of the studies was to determine the effects of calcium carbonate and calcium phosphate supplementation on faecal Lactobacillus spp., with and without a probiotic supplement, in healthy adults. Study 1 comprised of a randomised, double-blind, crossover design; participants (n=15) received 2 capsules/d of 250 mg elemental calcium as calcium carbonate (Ca1) and calcium phosphate (Ca2) each for 2-week periods, with 2-week baseline and washout periods. Study 2 was a randomised, double-blind, crossover design; participants (n=17) received 2 capsules/d of Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011 (probiotic) alone, the probiotic with 2 capsules/d of Ca1, and probiotic with 2 capsules/d of Ca2 each for 2-week periods with 2-week baseline and washout periods. In both studies, stools were collected during the baseline, intervention and washout periods for Lactobacillus spp. quantification and qPCR analyses. Participants completed daily questionnaires of stool fr...
Angewandte Chemie (International ed. in English), Jan 20, 2015
Conventional antibiotics are ineffective against non-replicating bacteria (for example, bacteria ... more Conventional antibiotics are ineffective against non-replicating bacteria (for example, bacteria within biofilms). We report a series of halogenated phenazines (HP), inspired by marine antibiotic 1, that targets persistent bacteria. HP 14 demonstrated the most potent biofilm eradication activities to date against MRSA, MRSE, and VRE biofilms (MBEC=0.2-12.5 μM), as well as the effective killing of MRSA persister cells in non-biofilm cultures. Frontline MRSA treatments, vancomycin and daptomycin, were unable to eradicate MRSA biofilms or non-biofilm persisters alongside 14. HP 13 displayed potent antibacterial activity against slow-growing M. tuberculosis (MIC=3.13 μM), the leading cause of death by bacterial infection around the world. HP analogues effectively target persistent bacteria through a mechanism that is non-toxic to mammalian cells and could have a significant impact on treatments for chronic bacterial infections.
Purpose: To determine which dietary factors contribute to microbiota variation and identify corre... more Purpose: To determine which dietary factors contribute to microbiota variation and identify correlations with gut epithelial methylation pattern associated with the presence of high-risk polyps. Dietary exposures have long been suggested as important risk factors for cancers, especially those of the gastrointestinal tract. Diet is known to affect the commensal gut microbiota, which contributes important activities to the human host that are required for maintaining normal health. However, distortions in microbiota composition and associated microbial activities might contribute to disease processes including colorectal carcinogenesis. High-throughput sequencing based microbiota studies have revealed a large degree of intra- and inter-individual variation, the sources of which are currently not well understood. As part of this project we explored various ‘Big Data’ approaches that can facilitate an efficient mining of the large dataset generated by combining dietary intake data with ...
A probiotic formulation of Enterococcus faecium R0026 and Bacillus subtilis R0179 has been evalua... more A probiotic formulation of Enterococcus faecium R0026 and Bacillus subtilis R0179 has been evaluated in previous clinical trials. However, B. subtilis R0179 has not been evaluated as a single probiotic strain or in combination with other strains at doses higher than 0.1×109 cfu. To establish oral dose-response tolerance and gastrointestinal (GI) viability of B. subtilis R0179, a randomised, double-blind, placebo-controlled trial in healthy adults (n=81; 18-50 years old) was conducted. Participants received B. subtilis R0179 at 0.1, 1.0 or 10×109 cfu/capsule/day or placebo for four weeks. General wellness was assessed using a daily questionnaire evaluating GI, cephalic, ear-nose-throat, behavioural, emetic, and epidermal symptoms. GI symptoms were further evaluated using a weekly gastrointestinal symptom rating scale (GSRS). GI transit viability of B. subtilis R0179 was assessed by plating and microbiota analysis by 16S rRNA at baseline, week 4 of the intervention and washout. Genera...
Humans harbor distinct commensal microbiota at various anatomic sites. There has been renewed int... more Humans harbor distinct commensal microbiota at various anatomic sites. There has been renewed interest in the contributions of microbiota activities to human health and disease. The microbiota of the gut is the most complex of all anatomic sites in terms of total numbers of bacteria that interact closely with the mucosal immune system and contribute various functions to host physiology. Especially in the proximal large intestine a diverse microbiota ferments complex substrates such as dietary fiber and host mucins, but also metabolizes bile acids and phytoestrogens that reach the large intestine. It is now well established that microbiota composition differs between but over time also within individuals. However, a thorough understanding of the sources of variations in microbiota composition, which is an important requirement for large population based microbiota studies is lacking. Microbiota composition varies depending on what kind of sample is collected, most commonly stool samples, stool swabs or superficial rectal or intestinal biopsies, and the time of collection. Microbiota dynamics are affected by life style factors including diet and exercise that determine what nutrients reach the proximal colon and how fast these nutrients pass through (transit time). Here we review sample collection issues in gut microbiota studies and recent findings about dynamics in microbiota composition. We recommend standardizing human microbiota analysis methods to facilitate comparison and
Late onset sepsis (LOS) is a major contributor to neonatal morbidity and mortality, especially in... more Late onset sepsis (LOS) is a major contributor to neonatal morbidity and mortality, especially in premature infants. Distortions in the establishment of normal gut microbiota, commensal microbes that colonize the digestive tract, might increase the risk of LOS via disruption of the mucosal barrier with resultant translocation of luminal contents. Correlation of distortions of the intestinal microbiota with LOS is a necessary first step to design novel microbiota-based screening approaches that might lead to early interventions to prevent LOS in high risk infants. Using a case/control design nested in a cohort study of preterm infants, we analyzed stool samples that had been prospectively collected from ten preterm infants with LOS and from 18 matched controls. A 16S rRNA based approach was utilized to compare microbiota diversity and identify specific bacterial signatures that differed in their prevalence between cases and controls. Overall a-diversity (Chao1) was lower in cases two weeks before (p,0.05) but not one week before or at the time of diagnosis of LOS. Overall microbiota structure (Unifrac) appeared distinct in cases 2 weeks and 1 week before but not at diagnosis (p,0.05). Although we detected few operational taxonomic units (OTUs) unique or enriched in cases, we found many OTUs common in controls that were lacking in cases (p,0.01). Bifidobacteria counts were lower in cases at all time points. Our results support the hypothesis that a distortion in normal microbiota composition, and not an enrichment of potential pathogens, is associated with LOS in preterm infants.
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Papers by MAria ukhanova