Urethral Catheter Biofilms Reveal Plasticity in Bacterial Composition and Metabolism and Withstand Host Immune Defenses in Hypoxic Environment.

Biofilms composed of multiple microorganisms colonize the surfaces of indwelling urethral catheters that are used serially by neurogenic bladder patients and cause chronic infections. Well-adapted pathogens in this niche are Escherichia coli, Proteus, and Enterococcus spp., species that cycle through adhesion and multilayered cell growth, trigger host immune responses, are starved off nutrients, and then disperse. Viable microbial foci retained in the urinary tract recolonize catheter surfaces. The molecular adaptations of bacteria in catheter biofilms (CBs) are not well-understood, promising new insights into this pathology based on host and microbial meta-omics analyses from clinical specimens. We examined catheters from nine neurogenic bladder patients longitudinally over up to 6 months. Taxonomic analyses from 16S rRNA gene sequencing and liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics revealed that 95% of all catheter and corresponding urinary pellet (UP) samples contained bacteria. CB biomasses were dominated by Enterobacteriaceae spp. and often accompanied by lactic acid and anaerobic bacteria. Systemic antibiotic drug treatments of patients resulted in either transient or lasting microbial community perturbations. Neutrophil effector proteins were abundant not only in UP but also CB samples, indicating their penetration of biofilm surfaces. In the context of one patient who advanced to a kidney infection, Proteus mirabilis proteomic data suggested a combination of factors associated with this disease complication: CB biomasses were high; the bacteria produced urease alkalinizing the pH and triggering urinary salt deposition on luminal catheter surfaces; P. mirabilis utilized energy-producing respiratory systems more than in CBs from other patients. The NADH:quinone oxidoreductase II (Nqr), a Na+ translocating enzyme not operating as a proton pump, and the nitrate reductase A (Nar) equipped the pathogen with electron transport chains promoting growth under hypoxic conditions. Both P. mirabilis and E. coli featured repertoires of transition metal ion acquisition systems in response to human host-mediated iron and zinc sequestration. We discovered a new drug target, the Nqr respiratory system, whose deactivation may compromise P. mirabilis growth in a basic pH milieu. Animal models would not allow such molecular-level insights into polymicrobial biofilm metabolism and interactions because the complexity cannot be replicated.

Vertical Transmission of Gut Microbiome and Antimicrobial Resistance Genes in Infants Exposed to Antibiotics at Birth.

Vertical transmission of maternal microbes is a major route for establishing the gut microbiome in newborns. The impact of perinatal antibiotics on vertical transmission of microbes and antimicrobial resistance is not well understood. Using a metagenomic approach, we analyzed the fecal samples from mothers and vaginally delivered infants from a control group (10 pairs) and a treatment group (10 pairs) receiving perinatal antibiotics. Antibiotic-usage had a significant impact on the main source of inoculum in the gut microbiome of newborns. The control group had significantly more species transmitted from mothers to infants (P = .03) than the antibiotic-treated group. Approximately 72% of the gut microbial population of infants at 3-7 days after birth in the control group was transmitted from their mothers, versus only 25% in the antibiotic-treated group. In conclusion, perinatal antibiotics markedly disturbed vertical transmission and changed the source of gut colonization towards horizontal transfer from the environment to the infants.

Characterizing Microbial Signatures on Sculptures and Paintings of Similar Provenance.

The preservation of artwork challenges museums, collectors, and art enthusiasts. Currently, reducing moisture, adjusting the type of lighting, and preventing the formation of mold are primary methods to preserving and preventing deterioration. Other methods such as ones based in detailed knowledge of molecular biology such as microbial community characterization using polymerase chain reaction (PCR) and sequencing have yet to be explored. Such molecular biology approaches are essential to explore as some environmental bacteria are capable of oxidizing nonpolar chemical substances rich in hydrocarbons such as oil-based paints. Using 16S rDNA Illumina Sequencing, we demonstrate a novel finding that there are differing bacterial communities for artwork from roughly the same era when comparing paintings on wood, paintings on canvases, and sculptures made of stone and marble. We also demonstrate that there are specific genera such as Aeromonas known for having oxidase positive strains, present on paintings on wood and paintings on canvas that could potentially be responsible for deterioration and fading as such organisms produce water or hydrogen peroxide as a byproduct of cytochrome c oxidase activity. The advantages of these genomics-based approaches to characterizing the microbial population on deteriorating artwork provides immense potential by identifying potentially damaging species that may not be detected using conventional methods in addition to addressing challenges to identification, restoration, and preservation efforts.

Oral Microbial Species and Virulence Factors Associated with Oral Squamous Cell Carcinoma.

The human microbiome has been the focus of numerous research efforts to elucidate the pathogenesis of human diseases including cancer. Oral cancer mortality is high when compared with other cancers, as diagnosis often occurs during late stages. Its prevalence has increased in the USA over the past decade and accounts for over 40,000 new cancer patients each year. Additionally, oral cancer pathogenesis is not fully understood and is likely multifactorial. To unravel the relationships that are associated with the oral microbiome and their virulence factors, we used 16S rDNA and metagenomic sequencing to characterize the microbial composition and functional content in oral squamous cell carcinoma (OSCC) tumor tissue, non-tumor tissue, and saliva from 18 OSCC patients. Results indicate a higher number of bacteria belonging to the Fusobacteria, Bacteroidetes, and Firmicutes phyla associated with tumor tissue when compared with all other sample types. Additionally, saliva metaproteomics revealed a significant increase of Prevotella in five OSCC subjects, while Corynebacterium was mostly associated with ten healthy subjects. Lastly, we determined that there are adhesion and virulence factors associated with Streptococcus gordonii as well as from known oral pathogens belonging to the Fusobacterium genera found mostly in OSCC tissues. From these results, we propose that not only will the methods utilized in this study drastically improve OSCC diagnostics, but the organisms and specific virulence factors from the phyla detected in tumor tissue may be excellent biomarkers for characterizing disease progression.

Intestinal and hepatic microbiota changes associated with chronic ethanol administration in mice.

Alcohol-induced liver disease is closely related to translocation of bacterial products and bacteria from the intestine to the liver. However, it is not known whether bacterial translocation to the liver depends on certain intestinal microbiota changes that would predispose bacteria to translocate to the liver. In this study, we investigated the microbiota in the jejunum, ileum, cecum, feces and liver of mice subjected to chronic ethanol feeding using a Lieber DeCarli diet model of chronic ethanol feeding for 8 weeks. We demonstrate that chronic ethanol administration changes alpha diversity in the ileum and the liver and leads to compositional changes especially in the ileum. This is largely driven by an increase in gram-negative phyla - the source of endotoxins. Moreover, gram-negative Prevotella not only increased in the mucus layer of the ileum but also in liver samples. These results suggest that bacterial translocation to the liver might be associated with microbiota changes in the distal gastrointestinal tract.

Persistent Gut Microbial Dysbiosis in Children with Acute Lymphoblastic Leukemia (ALL) During Chemotherapy.

Prophylactic or therapeutic antibiotic use along with chemotherapy treatment potentially has a long-standing adverse effect on the resident gut microbiota. We have established a case-control cohort of 32 pediatric and adolescent acute lymphoblastic leukemia (ALL) patients and 25 healthy siblings (sibling controls) to assess the effect of chemotherapy as well as antibiotic prophylaxis on the gut microbiota. We observe that the microbiota diversity and richness of the ALL group is significantly lower than that of the control group at diagnosis and during chemotherapy. The microbiota diversity is even lower in antibiotics-exposed ALL patients. Although the gut microbial diversity tends to stabilize after 1-year post-chemotherapy, their abundances were altered because of chemotherapy and prophylactic antibiotic treatments. Specifically, the abundances of mucolytic gram-positive anaerobic bacteria, including Ruminococcus gnavus and Ruminococcus torques, tended to increase during the chemotherapy regimen and continued to be elevated 1 year beyond the initiation of chemotherapy. This dysbiosis may contribute to the development of gastrointestinal complications in ALL children following chemotherapy. These findings set the stage to further understand the role of the gut microbiome dynamics in ALL patients and their potential role in alleviating some of the adverse side effects of chemotherapy and antibiotics use in immunocompromised children.

Gastro-intestinal and oral microbiome signatures associated with healthy aging.

The human oral and gut microbiomes influence health via competition for a distinct niche in the body with pathogens, via metabolic capabilities that increase host digestive capacity and generate compounds engaged in signaling pathways and modulation of immune system functions. Old age alters our metabolic and regenerative capacity. Following recruitment of 65 human subjects in the age range of 70 to 82, we discerned healthy aging (HA) and non-healthy aging (NHA) cohorts discordant in the occurrence of one or more major diseases: (1) cancer, (2) acute or chronic cardiovascular diseases, (3) acute or chronic pulmonary diseases, (4) diabetes, and (5) stroke or neurodegenerative disorders. We analyzed these cohorts' oral microbiomes (saliva) and gut microbiomes (stool) to assess diversity and identify microbial biomarkers for HA. In contrast to the gut microbiome where no change was observed, we found that the saliva microbiome had higher α-diversity in the HA compared with the NHA group. We observed the genus Akkermansia to be significantly more abundant in the gut microbiota of the HA group. Akkermansia muciniphila is a colonic mucin-degrading bacterium believed to have beneficial effects on gastrointestinal health, particularly in the context of diabetes and obesity. Erysipelotrichaceae UCG-003 was a taxon increased in abundance in the HA cohort. Streptococcus was the only genus observed to be significantly decreased in abundance in both the gut and oral microbiomes of the HA cohort compared with the NHA cohort. Our data support the notion that these microbes are potential probiotics to decrease the risks of non-healthy aging.

Study of the impact of long-duration space missions at the International Space Station on the astronaut microbiome.

Over the course of a mission to the International Space Station (ISS) crew members are exposed to a number of stressors that can potentially alter the composition of their microbiomes and may have a negative impact on astronauts' health. Here we investigated the impact of long-term space exploration on the microbiome of nine astronauts that spent six to twelve months in the ISS. We present evidence showing that the microbial communities of the gastrointestinal tract, skin, nose and tongue change during the space mission. The composition of the intestinal microbiota became more similar across astronauts in space, mostly due to a drop in the abundance of a few bacterial taxa, some of which were also correlated with changes in the cytokine profile of crewmembers. Alterations in the skin microbiome that might contribute to the high frequency of skin rashes/hypersensitivity episodes experienced by astronauts in space were also observed. The results from this study demonstrate that the composition of the astronauts' microbiome is altered during space travel. The impact of those changes on crew health warrants further investigation before humans embark on long-duration voyages into outer space.

Microbial metagenome of urinary tract infection.

Urine culture and microscopy techniques are used to profile the bacterial species present in urinary tract infections. To gain insight into the urinary flora, we analyzed clinical laboratory features and the microbial metagenome of 121 clean-catch urine samples. 16S rDNA gene signatures were successfully obtained for 116 participants, while metagenome sequencing data was successfully generated for samples from 49 participants. Although 16S rDNA sequencing was more sensitive, metagenome sequencing allowed for a more comprehensive and unbiased representation of the microbial flora, including eukarya and viral pathogens, and of bacterial virulence factors. Urine samples positive by metagenome sequencing contained a plethora of bacterial (median 41 genera/sample), eukarya (median 2 species/sample) and viral sequences (median 3 viruses/sample). Genomic analyses suggested cases of infection with potential pathogens that are often missed during routine urine culture due to species specific growth requirements. While conventional microbiological methods are inadequate to identify a large diversity of microbial species that are present in urine, genomic approaches appear to more comprehensively and quantitatively describe the urinary microbiome.

Initial genome sequencing of the sugarcane CP 96-1252 complex hybrid.

The CP 96-1252 cultivar of sugarcane is a complex hybrid of commercial importance. DNA was extracted from lab-grown leaf tissue and sequenced. The raw Illumina DNA sequencing results provide 101 Gbp of genome sequence reads. The dataset is available from https://www.ncbi.nlm.nih.gov/bioproject/PRJNA345486/.

Intestinal fungi contribute to development of alcoholic liver disease.

Chronic liver disease with cirrhosis is the 12th leading cause of death in the United States, and alcoholic liver disease accounts for approximately half of all cirrhosis deaths. Chronic alcohol consumption is associated with intestinal bacterial dysbiosis, yet we understand little about the contribution of intestinal fungi, or mycobiota, to alcoholic liver disease. Here we have demonstrated that chronic alcohol administration increases mycobiota populations and translocation of fungal ß-glucan into systemic circulation in mice. Treating mice with antifungal agents reduced intestinal fungal overgrowth, decreased ß-glucan translocation, and ameliorated ethanol-induced liver disease. Using bone marrow chimeric mice, we found that ß-glucan induces liver inflammation via the C-type lectin-like receptor CLEC7A on Kupffer cells and possibly other bone marrow-derived cells. Subsequent increases in IL-1ß expression and secretion contributed to hepatocyte damage and promoted development of ethanol-induced liver disease. We observed that alcohol-dependent patients displayed reduced intestinal fungal diversity and Candida overgrowth. Compared with healthy individuals and patients with non-alcohol-related cirrhosis, alcoholic cirrhosis patients had increased systemic exposure and immune response to mycobiota. Moreover, the levels of extraintestinal exposure and immune response correlated with mortality. Thus, chronic alcohol consumption is associated with an altered mycobiota and translocation of fungal products. Manipulating the intestinal mycobiome might be an effective strategy for attenuating alcohol-related liver disease.

Gastrointestinal microbial populations can distinguish pediatric and adolescent Acute Lymphoblastic Leukemia (ALL) at the time of disease diagnosis.

BACKGROUND: An estimated 15,000 children and adolescents under the age of 19 years are diagnosed with leukemia, lymphoma and other tumors in the USA every year. All children and adolescent acute leukemia patients will undergo chemotherapy as part of their treatment regimen. Fortunately, survival rates for most pediatric cancers have improved at a remarkable pace over the past three decades, and the overall survival rate is greater than 90 % today. However, significant differences in survival rate have been found in different age groups (94 % in 1-9.99 years, 82 % in ≥10 years and 76 % in ≥15 years). ALL accounts for about three out of four cases of childhood leukemia. Intensive chemotherapy treatment coupled with prophylactic or therapeutic antibiotic use could potentially have a long-term effect on the resident gastrointestinal (GI) microbiome. The composition of GI microbiome and its changes upon chemotherapy in pediatric and adolescent leukemia patients is poorly understood. In this study, using 16S rRNA marker gene sequences we profile the GI microbial communities of pediatric and adolescent acute leukemia patients before and after chemotherapy treatment and compare with the microbiota of their healthy siblings. RESULTS: Our study cohort consisted of 51 participants, made up of matched pediatric and adolescent patients with ALL and a healthy sibling. We elucidated and compared the GI microbiota profiles of patients and their healthy sibling controls via analysis of 16S rRNA gene sequencing data. We assessed the GI microbiota composition in pediatric and adolescent patients with ALL during the course of chemotherapy by comparing stool samples taken before chemotherapy with stool samples collected at varying time points during the chemotherapeutic treatment. The microbiota profiles of both patients and control sibling groups are dominated by members of Bacteroides, Prevotella, and Faecalibacterium. At the genus level, both groups share many taxa in common, but the microbiota diversity of the patient group is significantly lower than that of the control group. It was possible to distinguish between the patient and control groups based on their microbiota profiles. The top taxa include Anaerostipes, Coprococcus, Roseburia, and Ruminococcus2 with relatively higher abundance in the control group. The observed microbiota changes are likely the result of several factors including a direct influence of therapeutic compounds on the gut flora and an indirect effect of chemotherapy on the immune system, which, in turn, affects the microbiota. CONCLUSIONS: This study provides significant information on GI microbiota populations in immunocompromised children and opens up the potential for developing novel diagnostics based on stool tests and therapies to improve the dysbiotic condition of the microbiota at the time of diagnosis and in the earliest stages of chemotherapy.

Intestinal REG3 Lectins Protect against Alcoholic Steatohepatitis by Reducing Mucosa-Associated Microbiota and Preventing Bacterial Translocation.

Approximately half of all deaths from liver cirrhosis, the tenth leading cause of mortality in the United States, are related to alcohol use. Chronic alcohol consumption is accompanied by intestinal dysbiosis and bacterial overgrowth, yet little is known about the factors that alter the microbial composition or their contribution to liver disease. We previously associated chronic alcohol consumption with lower intestinal levels of the antimicrobial-regenerating islet-derived (REG)-3 lectins. Here, we demonstrate that intestinal deficiency in REG3B or REG3G increases numbers of mucosa-associated bacteria and enhances bacterial translocation to the mesenteric lymph nodes and liver, promoting the progression of ethanol-induced fatty liver disease toward steatohepatitis. Overexpression of Reg3g in intestinal epithelial cells restricts bacterial colonization of mucosal surfaces, reduces bacterial translocation, and protects mice from alcohol-induced steatohepatitis. Thus, alcohol appears to impair control of the mucosa-associated microbiota, and subsequent breach of the mucosal barrier facilitates progression of alcoholic liver disease.