Publications by authors named "Steven R Gill"

71 Publications

Microbiomics of irrigation with xylitol or in chronic rhinosinusitis.

Laryngoscope Investig Otolaryngol 2021 Feb 21;6(1):64-70. Epub 2021 Jan 21.

Department of Otolaryngology Head and Neck Surgery University of Rochester Medical Center Rochester New York USA.

Objective: Topical sinonasal rinse therapies may alter the local microbiome and improve disease control in chronic rhinosinusitis (CRS). The objective of this study was to examine microbiome changes in post-surgical CRS patients when rinsing with commercially available products containing xylitol or .

Methods: A crossover-type protocol with a washout period was designed. Swab samples from anterior ethmoid cavities of CRS patients were collected prospectively at baseline. Subjects were provided packets containing either W136 or xylitol in non-blinded fashion and instructed to add it to their rinse bottles daily for 28 days, after which another swab was taken. A saline wash-out period was completed and a third swab taken. A final 28-day regimen of the opposite product was followed by a final swab. DNA extraction and sequencing of the 16S rRNA gene allowed for global microbiome analysis.

Results: We enrolled 25 subjects with CRS and 10 controls resulting in 70 adequate samples. Increased detection of was observed after use of . No significant trends in alpha or beta diversity as a result of treatment were observed. SNOT-22 score did not change significantly following treatment with xylitol, , or saline.

Conclusion: We did not detect any major clinical or microbiome-level effect due to treatment with two topical rinse products. Further research is needed to elucidate their clinical utility and possible probiotic effect.

Level Of Evidence: 3.
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http://dx.doi.org/10.1002/lio2.524DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7883620PMC
February 2021

Staphylococcus aureus Tolerance and Genomic Response to Photodynamic Inactivation.

mSphere 2021 01 6;6(1). Epub 2021 Jan 6.

Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, New York, USA

is an opportunistic pathogen with a clinical spectrum ranging from asymptomatic skin colonization to invasive infections. While traditional antibiotic therapies can be effective against , the increasing prevalence of antibiotic-resistant strains results in treatment failures and high mortality rates. Photodynamic inactivation (PDI) is an innovative and promising alternative to antibiotics. While progress has been made in our understanding of the bacterial response to PDI, major gaps remain in our knowledge of PDI tolerance, the global cellular response, and adaptive genomic mutations acquired as a result of PDI. To address these gaps, HG003 and isogenic mutants with mutations in , , , and exposed to single or multiple doses of PDI were assessed for survival and tolerance and examined by global transcriptome and genome analyses to identify regulatory and genetic adaptations that contribute to tolerance. Pathways in inorganic ion transport, oxidative response, DNA replication recombination and repair, and cell wall and membrane biogenesis were identified in a global cellular response to PDI. Tolerance to PDI was associated with superoxide dismutase and the global methylhydroquinone (MHQ)-quinone transcriptome network. Genome analysis of PDI-tolerant HG003 identified a nonsynonymous mutation in the quinone binding domain of the transcriptional repressor QsrR, which mediates quinone sensing and oxidant response. Acquisition of a heritable QsrR mutation through repeated PDI treatment demonstrates selective adaption of to PDI. PDI tolerance of a gene deletion in HG003 confirmed that QsrR regulates the response to PDI. can cause disease at most body sites, with illness ranging from asymptomatic infection to death. The increasing prevalence of antibiotic-resistant strains results in treatment failures and high mortality rates. acquires resistance to antibiotics through multiple mechanisms, often by genetic variation that alters antimicrobial targets. Photodynamic inactivation (PDI), which employs a combination of a nontoxic dye and low-intensity visible light, is a promising alternative to antibiotics that effectively eradicates in human infections when antibiotics are no longer effective. In this study, we demonstrate that repeated exposure to PDI results in resistance of to further PDI treatment and identify the underlying bacterial mechanisms that contribute to resistance. This work supports further analysis of these mechanisms and refinement of this novel technology as an adjunctive treatment for infections.
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http://dx.doi.org/10.1128/mSphere.00762-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7845598PMC
January 2021

The Interplay Between Respiratory Microbiota and Innate Immunity in Flavor E-Cigarette Vaping Induced Lung Dysfunction.

Front Microbiol 2020 16;11:589501. Epub 2020 Dec 16.

Department of Environmental Medicine, University of Rochester Medical Center, Rochester, NY, United States.

Global usage of electronic nicotine delivery systems (ENDS) has been increasing in the last decade. ENDS are non-combustible tobacco products that heat and aerosolize a liquid containing humectants, with added flavorings and often nicotine. Though ENDS are promoted as a less harmful alternative to smoking, current evidence links their use to a wide range of deleterious health effects including acute and chronic lung damage. ENDS can elicit an inflammatory response and impair the innate immune response in the lungs. Exposure to ENDS flavorings results in abnormal activation of the lung epithelial cells and β-defensins, dysfunction of the macrophage phagocytic activity, increased levels of mucin (MUC5AC) and abnormal activation of the neutrophilic response (NETosis). ENDS menthol flavorings disrupt innate immunity and might be associated with allergies and asthma through activation of transient receptor potential ankyrin 1 (TRAP1). Recent studies have expanded our understanding of the relationship between the homeostasis of lung innate immunity and the immunomodulatory effect of the host-microbiota interaction. Alterations of the normal respiratory microbiota have been associated with chronic obstructive pulmonary disease (COPD), asthma, atopy and cystic fibrosis complications which are strongly associated with smoking and potentially with ENDS use. Little is known about the short-and long-term effects of ENDS on the respiratory microbiota, their impact on the innate immune response and their link to pulmonary health and disease. Here we review the interaction between the innate immune system and the respiratory microbiota in the pathogenesis of ENDS-induced pulmonary dysfunction and identify future areas of research.
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http://dx.doi.org/10.3389/fmicb.2020.589501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7772214PMC
December 2020

Identification of Penicillin Binding Protein 4 (PBP4) as a critical factor for Staphylococcus aureus bone invasion during osteomyelitis in mice.

PLoS Pathog 2020 10 22;16(10):e1008988. Epub 2020 Oct 22.

Center for Musculoskeletal Research, University of Rochester Medical Center, Rochester, NY, United States of America.

Staphylococcus aureus infection of bone is challenging to treat because it colonizes the osteocyte lacuno-canalicular network (OLCN) of cortical bone. To elucidate factors involved in OLCN invasion and identify novel drug targets, we completed a hypothesis-driven screen of 24 S. aureus transposon insertion mutant strains for their ability to propagate through 0.5 μm-sized pores in the Microfluidic Silicon Membrane Canalicular Arrays (μSiM-CA), developed to model S. aureus invasion of the OLCN. This screen identified the uncanonical S. aureus transpeptidase, penicillin binding protein 4 (PBP4), as a necessary gene for S. aureus deformation and propagation through nanopores. In vivo studies revealed that Δpbp4 infected tibiae treated with vancomycin showed a significant 12-fold reduction in bacterial load compared to WT infected tibiae treated with vancomycin (p<0.05). Additionally, Δpbp4 infected tibiae displayed a remarkable decrease in pathogenic bone-loss at the implant site with and without vancomycin therapy. Most importantly, Δpbp4 S. aureus failed to invade and colonize the OLCN despite high bacterial loads on the implant and in adjacent tissues. Together, these results demonstrate that PBP4 is required for S. aureus colonization of the OLCN and suggest that inhibitors may be synergistic with standard of care antibiotics ineffective against bacteria within the OLCN.
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http://dx.doi.org/10.1371/journal.ppat.1008988DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608983PMC
October 2020

Airway gene expression correlates of RSV disease severity and microbiome composition in infants.

J Infect Dis 2020 Sep 14. Epub 2020 Sep 14.

Division of Neonatology and Pediatric Molecular and Personalized Medicine Program, University of Rochester Medical Center, Rochester NY, USA.

Rationale: Respiratory syncytial virus (RSV) is the leading cause of severe respiratory disease in infants. The causes and correlates of severe illness in the majority of infants are poorly defined.

Objectives: We identified molecular correlates of illness severity from the airways of infants infected with RSV.

Methods: We recruited a cohort of RSV-infected infants and simultaneously assayed the molecular status of their airways and the presence of airway microbiota. Rigorous statistical approaches identified gene expression patterns associated with disease severity and microbiota composition, separately and in combination.

Measurements And Main Results: We measured comprehensive airway gene expression patterns in 106 infants with primary RSV infection. We identified an airway gene expression signature of severe illness dominated by excessive chemokine expression. We also found an association between H. influenzae, disease severity and airway lymphocyte accumulation. Exploring the time of onset of clinical symptoms revealed acute activation of interferon (IFN) signaling following RSV infection in infants with mild or moderate illness, which was absent in subjects with severe illness.

Conclusion: Our data reveal that airway gene expression patterns distinguish mild/moderate from severe illness severity. Furthermore, our data identify biomarkers that may be therapeutic targets or useful for measuring efficacy of intervention responses.
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http://dx.doi.org/10.1093/infdis/jiaa576DOI Listing
September 2020

Temporal Dysbiosis of Infant Nasal Microbiota Relative to Respiratory Syncytial Virus Infection.

J Infect Dis 2020 Sep 14. Epub 2020 Sep 14.

Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA.

Rationale: Respiratory Syncytial Virus (RSV) is a leading cause of infant respiratory disease. Infant airway microbiota has been associated with respiratory disease risk and severity. The extent to which interactions between RSV and microbiota occur in the airway, and their impact on respiratory disease susceptibility and severity, are unknown.

Objectives: Characterize temporal associations between microbiota and RSV infection before, during, and after infants' first respiratory illness.

Methods: 16S rRNA microbiota profiling of two infant cohorts in the first year of life: 1) a cross-sectional cohort of 89 RSV infected infants sampled during illness and 102 matched healthy controls, and 2) a matched longitudinal cohort of 12 infants who developed RSV infection and 12 who did not, sampled before, during, and after infection.

Results: We identified 12 taxa significantly associated with RSV infection. All 12 taxa were differentially abundant during infection, with 8 associated with disease severity. Nasal microbiota composition was more discriminative of healthy vs. infected than of disease severity.

Conclusions: Our findings elucidate the chronology of nasal microbiota dysbiosis and suggest an altered developmental trajectory associated with RSV infection. Microbial temporal dynamics reveal indicators of disease risk, correlates of illness and severity, and impact of RSV infection on microbiota composition.
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http://dx.doi.org/10.1093/infdis/jiaa577DOI Listing
September 2020

Best Practices for Successfully Writing and Publishing a Genome Announcement in .

Microbiol Resour Announc 2020 Sep 3;9(36). Epub 2020 Sep 3.

Department of Biological Sciences, University of Southern California, Los Angeles, California, USA.

(MRA) provides peer-reviewed announcements of scientific resources for the microbial research community. We describe the best practices for writing an announcement that ensures that these publications are truly useful resources. Adhering to these best practices can lead to successful publication without the need for extensive revisions.
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http://dx.doi.org/10.1128/MRA.00763-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7471381PMC
September 2020

Oral microbiome: possible harbinger for children's health.

Int J Oral Sci 2020 04 30;12(1):12. Epub 2020 Apr 30.

Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, NY, USA.

The human microbiome functions as an intricate and coordinated microbial network, residing throughout the mucosal surfaces of the skin, oral cavity, gastrointestinal tract, respiratory tract, and reproductive system. The oral microbiome encompasses a highly diverse microbiota, consisting of over 700 microorganisms, including bacteria, fungi, and viruses. As our understanding of the relationship between the oral microbiome and human health has evolved, we have identified a diverse array of oral and systemic diseases associated with this microbial community, including but not limited to caries, periodontal diseases, oral cancer, colorectal cancer, pancreatic cancer, and inflammatory bowel syndrome. The potential predictive relationship between the oral microbiota and these human diseases suggests that the oral cavity is an ideal site for disease diagnosis and development of rapid point-of-care tests. The oral cavity is easily accessible with a non-invasive collection of biological samples. We can envision a future where early life salivary diagnostic tools will be used to predict and prevent future disease via analyzing and shaping the infant's oral microbiome. In this review, we present evidence for the establishment of the oral microbiome during early childhood, the capability of using childhood oral microbiome to predict future oral and systemic diseases, and the limitations of the current evidence.
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http://dx.doi.org/10.1038/s41368-020-0082-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7190716PMC
April 2020

The gut microbiome-joint connection: implications in osteoarthritis.

Curr Opin Rheumatol 2020 01;32(1):92-101

Department of Orthopedics, Anschutz Medical Campus.

Purpose Of Review: Osteoarthritis is a debilitating disease leading to joint degeneration, inflammation, pain, and disability. Despite efforts to develop a disease modifying treatment, the only accepted and available clinical approaches involve palliation. Although many factors contribute to the development of osteoarthritis, the gut microbiome has recently emerged as an important pathogenic factor in osteoarthritis initiation and progression. This review examines the literature to date regarding the link between the gut microbiome and osteoarthritis.

Recent Findings: Studies showing correlations between serum levels of bacterial metabolites and joint degeneration were the first links connecting a dysbiosis of the gut microbiome with osteoarthritis. Further investigations have demonstrated that microbial community shifts induced by antibiotics, a germ-free environment or high-fat are important underlying factors in joint homeostasis and osteoarthritis. It follows that strategies to manipulate the microbiome have demonstrated efficacy in mitigating joint degeneration in osteoarthritis. Moreover, we have observed that dietary supplementation with nutraceuticals that are joint protective may exert their influence via shifts in the gut microbiome.

Summary: Although role of the microbiome in osteoarthritis is an area of intense study, no clear mechanism of action has been determined. Increased understanding of how the two factors interact may provide mechanistic insight into osteoarthritis and lead to disease modifying treatments.
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http://dx.doi.org/10.1097/BOR.0000000000000681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6903327PMC
January 2020

Evolving concepts in bone infection: redefining "biofilm", "acute vs. chronic osteomyelitis", "the immune proteome" and "local antibiotic therapy".

Bone Res 2019 15;7:20. Epub 2019 Jul 15.

1Center for Musculoskeletal Research, University of Rochester Medical Center, Rochester, NY USA.

Osteomyelitis is a devastating disease caused by microbial infection of bone. While the frequency of infection following elective orthopedic surgery is low, rates of reinfection are disturbingly high. is responsible for the majority of chronic osteomyelitis cases and is often considered to be incurable due to bacterial persistence deep within bone. Unfortunately, there is no consensus on clinical classifications of osteomyelitis and the ensuing treatment algorithm. Given the high patient morbidity, mortality, and economic burden caused by osteomyelitis, it is important to elucidate mechanisms of bone infection to inform novel strategies for prevention and curative treatment. Recent discoveries in this field have identified three distinct reservoirs of bacterial biofilm including: abscess communities in the local soft tissue and bone marrow, glycocalyx formation on implant hardware and necrotic tissue, and colonization of the osteocyte-lacuno canalicular network (OLCN) of cortical bone. In contrast, intracellular persistence in bone cells has not been substantiated in vivo, which challenges this mode of chronic osteomyelitis. There have also been major advances in our understanding of the immune proteome against , from clinical studies of serum antibodies and media enriched for newly synthesized antibodies (MENSA), which may provide new opportunities for osteomyelitis diagnosis, prognosis, and vaccine development. Finally, novel therapies such as antimicrobial implant coatings and antibiotic impregnated 3D-printed scaffolds represent promising strategies for preventing and managing this devastating disease. Here, we review these recent advances and highlight translational opportunities towards a cure.
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http://dx.doi.org/10.1038/s41413-019-0061-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6804538PMC
July 2019

Microbiome-Transcriptome Interactions Related to Severity of Respiratory Syncytial Virus Infection.

Sci Rep 2019 09 25;9(1):13824. Epub 2019 Sep 25.

Channing Division of Network Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, 02115, USA.

Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections and hospital visits during infancy and childhood. Although risk factors for RSV infection have been identified, the role of microbial species in the respiratory tract is only partially known. We aimed to understand the impact of interactions between the nasal microbiome and host transcriptome on the severity and clinical outcomes of RSV infection. We used 16 S rRNA sequencing to characterize the nasal microbiome of infants with RSV infection. We used RNA sequencing to interrogate the transcriptome of CD4 T cells obtained from the same set of infants. After dimension reduction through principal component (PC) analysis, we performed an integrative analysis to identify significant co-variation between microbial clade and gene expression PCs. We then employed LIONESS (Linear Interpolation to Obtain Network Estimates for Single Samples) to estimate the clade-gene association patterns for each infant. Our network-based integrative analysis identified several clade-gene associations significantly related to the severity of RSV infection. The microbial taxa with the highest loadings in the implicated clade PCs included Moraxella, Corynebacterium, Streptococcus, Haemophilus influenzae, and Staphylococcus. Interestingly, many of the genes with the highest loadings in the implicated gene PCs are encoded in mitochondrial DNA, while others are involved in the host immune response. This study on microbiome-transcriptome interactions provides insights into how the host immune system mounts a response against RSV and specific infectious agents in nasal microbiota.
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http://dx.doi.org/10.1038/s41598-019-50217-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6761288PMC
September 2019

Measuring the Severity of Respiratory Illness in the First 2 Years of Life in Preterm and Term Infants.

J Pediatr 2019 11 31;214:12-19.e3. Epub 2019 Jul 31.

Department of Pediatrics, University of Rochester Medical Center, Rochester, NY; Department of Environmental Medicine, University of Rochester Medical Center, Rochester, NY.

Objective: To develop a valid research tool to measure infant respiratory illness severity using parent-reported symptoms.

Study Design: Nose and throat swabs were collected monthly for 1 year and during respiratory illnesses for 2 years in a prospective study of term and preterm infants in the Prematurity, Respiratory Outcomes, Immune System and Microbiome study. Viral pathogens were detected using Taqman Array Cards. Parents recorded symptoms during respiratory illnesses using a Childhood Origins of Asthma (COAST) scorecard. The COAST score was validated using linear mixed effects regression modeling to evaluate associations with hospitalization and specific infections. A data-driven method was also used to compute symptom weights and derive a new score, the Infant Research Respiratory Infection Severity Score (IRRISS). Linear mixed effects regression modeling was repeated with the IRRISS illness data.

Results: From April 2013 to April 2017, 50 term, 40 late preterm, and 28 extremely low gestational age (<29 weeks of gestation) infants had 303 respiratory illness visits with viral testing and parent-reported symptoms. A range of illness severity was described with 39% of illness scores suggestive of severe disease. Both the COAST score and IRRISS were associated with respiratory syncytial virus infection and hospitalization. Gestational age and human rhinovirus infection were inversely associated with both scoring systems. The IRRISS and COAST scores were highly correlated (r = 0.93; P < .0001).

Conclusions: Using parent-reported symptoms, we validated the COAST score as a measure of respiratory illness severity in infants. The new IRRISS score performed as well as the COAST score.
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http://dx.doi.org/10.1016/j.jpeds.2019.06.061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6815715PMC
November 2019

An in vitro platform for elucidating the molecular genetics of S. aureus invasion of the osteocyte lacuno-canalicular network during chronic osteomyelitis.

Nanomedicine 2019 10 24;21:102039. Epub 2019 Jun 24.

Department of Biomedical Engineering, University of Rochester Medical Center, Rochester, NY. Electronic address:

Staphylococcus aureus osteomyelitis is a devasting disease that often leads to amputation. Recent findings have shown that S. aureus is capable of invading the osteocyte lacuno-canalicular network (OLCN) of cortical bone during chronic osteomyelitis. Normally a 1 μm non-motile cocci, S. aureus deforms smaller than 0.5 μm in the sub-micron channels of the OLCN. Here we present the μSiM-CA (Microfluidic - Silicon Membrane - Canalicular Array) as an in vitro screening platform for the genetic mechanisms of S. aureus invasion. The μSiM-CA platform features an ultrathin silicon membrane with defined pores that mimic the openings of canaliculi. While we anticipated that S. aureus lacking the accessory gene regulator (agr) quorum-sensing system would not be capable of invading the OLCN, we found no differences in propagation compared to wild type in the μSiM-CA. However the μSiM-CA proved predictive as we also found that the agr mutant strain invaded the OLCN of murine tibiae.
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http://dx.doi.org/10.1016/j.nano.2019.102039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6814543PMC
October 2019

Aims, Study Design, and Enrollment Results From the Assessing Predictors of Infant Respiratory Syncytial Virus Effects and Severity Study.

JMIR Res Protoc 2019 Jun 6;8(6):e12907. Epub 2019 Jun 6.

University of Rochester School of Medicine and Dentistry, Rochester, NY, United States.

Background: The majority of infants hospitalized with primary respiratory syncytial virus (RSV) infection have no obvious risk factors for severe disease.

Objective: The aim of this study (Assessing Predictors of Infant RSV Effects and Severity, AsPIRES) was to identify factors associated with severe disease in full-term healthy infants younger than 10 months with primary RSV infection.

Methods: RSV infected infants were enrolled from 3 cohorts during consecutive winters from August 2012 to April 2016 in Rochester, New York. A birth cohort was prospectively enrolled and followed through their first winter for development of RSV infection. An outpatient supplemental cohort was enrolled in the emergency department or pediatric offices, and a hospital cohort was enrolled on admission with RSV infection. RSV was diagnosed by reverse transcriptase-polymerase chain reaction. Demographic and clinical data were recorded and samples collected for assays: buccal swab (cytomegalovirus polymerase chain reaction, PCR), nasal swab (RSV qualitative PCR, complete viral gene sequence, 16S ribosomal ribonucleic acid [RNA] amplicon microbiota analysis), nasal wash (chemokine and cytokine assays), nasal brush (nasal respiratory epithelial cell gene expression using RNA sequencing [RNAseq]), and 2 to 3 ml of heparinized blood (flow cytometry, RNAseq analysis of purified cluster of differentiation [CD]4+, CD8+, B cells and natural killer cells, and RSV-specific antibody). Cord blood (RSV-specific antibody) was also collected for the birth cohort. Univariate and multivariate logistic regression will be used for analysis of data using a continuous Global Respiratory Severity Score (GRSS) as the outcome variable. Novel statistical methods will be developed for integration of the large complex datasets.

Results: A total of 453 infants were enrolled into the 3 cohorts; 226 in the birth cohort, 60 in the supplemental cohort, and 78 in the hospital cohort. A total of 126 birth cohort infants remained in the study and were evaluated for 150 respiratory illnesses. Of the 60 RSV positive infants in the supplemental cohort, 42 completed the study, whereas all 78 of the RSV positive hospital cohort infants completed the study. A GRSS was calculated for each RSV-infected infant and is being used to analyze each of the complex datasets by correlation with disease severity in univariate and multivariate methods.

Conclusions: The AsPIRES study will provide insights into the complex pathogenesis of RSV infection in healthy full-term infants with primary RSV infection. The analysis will allow assessment of multiple factors potentially influencing the severity of RSV infection including the level of RSV specific antibodies, the innate immune response of nasal epithelial cells, the adaptive response by various lymphocyte subsets, the resident airway microbiota, and viral factors. Results of this study will inform disease interventions such as vaccines and antiviral therapies.
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http://dx.doi.org/10.2196/12907DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6595944PMC
June 2019

The Response of and Contributes to Virulence and Metabolism.

J Bacteriol 2019 05 9;201(9). Epub 2019 Apr 9.

Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, Rochester, New York, USA

causes a wide spectrum of disease, with the site and severity of infection dependent on virulence traits encoded within genetically distinct clonal complexes (CCs) and bacterial responses to host innate immunity. The production of nitric oxide (NO) by activated phagocytes is a major host response to which metabolically adapts through multiple strategies that are conserved in all CCs, including an nitric oxide synthase (Nos). Previous genome analysis of CC30, a lineage associated with chronic endocardial and osteoarticular infections, revealed a putative NO reductase (Nor) not found in other CCs that potentially contributes to NO resistance and clinical outcome. Here, we demonstrate that Nor has true nitric oxide reductase activity, with expression enhanced by NO stress and anaerobic growth. Furthermore, we demonstrate that is regulated by MgrA and SrrAB, which modulate virulence and hypoxic response. Transcriptome analysis of the UAMS-1, UAMS-1 Δ, and UAMS-1 Δ strains under NO stress and anaerobic growth demonstrates that Nor contributes to nucleotide metabolism and Nos to glycolysis. We demonstrate that Nor and Nos contribute to enhanced survival in the presence of human human polymorphonuclear cells and have organ-specific seeding in a tail vein infection model. Nor contributes to abscess formation in an osteological implant model. We also demonstrate that Nor has a role in metabolism and virulence. The regulation overlap between Nor and Nos points to an intriguing link between regulation of intracellular NO, metabolic adaptation, and persistence in the CC30 lineage. can cause disease at most body sites, and illness spans asymptomatic infection to death. The variety of clinical presentations is due to the diversity of strains, which are grouped into distinct clonal complexes (CCs) based on genetic differences. The ability of CC30 to cause chronic infections relies on its ability to evade the oxidative/nitrosative defenses of the immune system and survive under different environmental conditions, including differences in oxygen and nitric oxide concentrations. The significance of this work is the exploration of unique genes involved in resisting NO stress and anoxia. A better understanding of the functions that control the response of CC30 to NO and oxygen will guide the treatment of severe disease presentations.
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http://dx.doi.org/10.1128/JB.00107-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6456865PMC
May 2019

Neonatal gut and respiratory microbiota: coordinated development through time and space.

Microbiome 2018 10 26;6(1):193. Epub 2018 Oct 26.

Genomics Research Center, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA.

Background: Postnatal development of early life microbiota influences immunity, metabolism, neurodevelopment, and infant health. Microbiome development occurs at multiple body sites, with distinct community compositions and functions. Associations between microbiota at multiple sites represent an unexplored influence on the infant microbiome. Here, we examined co-occurrence patterns of gut and respiratory microbiota in pre- and full-term infants over the first year of life, a period critical to neonatal development.

Results: Gut and respiratory microbiota collected as longitudinal rectal, throat, and nasal samples from 38 pre-term and 44 full-term infants were first clustered into community state types (CSTs) on the basis of their compositional profiles. Multiple methods were used to relate the occurrence of CSTs to temporal microbiota development and measures of infant maturity, including gestational age (GA) at birth, week of life (WOL), and post-menstrual age (PMA). Manifestation of CSTs followed one of three patterns with respect to infant maturity: (1) chronological, with CST occurrence frequency solely a function of post-natal age (WOL), (2) idiosyncratic to maturity at birth, with the interval of CST occurrence dependent on infant post-natal age but the frequency of occurrence dependent on GA at birth, and (3) convergent, in which CSTs appear first in infants of greater maturity at birth, with occurrence frequency in pre-terms converging after a post-natal interval proportional to pre-maturity. The composition of CSTs was highly dissimilar between different body sites, but the CST of any one body site was highly predictive of the CSTs at other body sites. There were significant associations between the abundance of individual taxa at each body site and the CSTs of the other body sites, which persisted after stringent control for the non-linear effects of infant maturity. Canonical correlations exist between the microbiota composition at each pair of body sites, with the strongest correlations between proximal locations.

Conclusion: These findings suggest that early microbiota is shaped by neonatal innate and adaptive developmental responses. Temporal progression of CST occurrence is influenced by infant maturity at birth and post-natal age. Significant associations of microbiota across body sites reveal distal connections and coordinated development of the infant microbial ecosystem.
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http://dx.doi.org/10.1186/s40168-018-0566-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6204011PMC
October 2018

Tracking Anti- Antibodies Produced and during Foot Salvage Therapy for Diabetic Foot Infections Reveals Prognostic Insights and Evidence of Diversified Humoral Immunity.

Infect Immun 2018 12 20;86(12). Epub 2018 Nov 20.

Center for Musculoskeletal Research and Department of Orthopaedics, University of Rochester Medical Center, Rochester, New York, USA

Management of foot salvage therapy (FST) for diabetic foot infections (DFI) is challenging due to the absence of reliable diagnostics to identify the etiologic agent and prognostics to justify aggressive treatments. As is the most common pathogen associated with DFI, we aimed to develop a multiplex immunoassay of IgG in serum and medium enriched for newly synthesized anti- antibodies (MENSA) generated from cultured peripheral blood mononuclear cells of DFI patients undergoing FST. Wound samples were collected from 26 DFI patients to identify the infecting bacterial species via 16S rRNA sequencing. Blood was obtained over 12 weeks of FST to assess anti- IgG levels in sera and MENSA. The results showed that 17 out of 26 infections were polymicrobial and 12 were positive for While antibody titers in serum and MENSA displayed similar diagnostic potentials to detect infection, MENSA showed a 2-fold-greater signal-to-background ratio. Multivariate analyses revealed increases in predictive power of diagnosing infections (area under the receiver operating characteristic curve [AUC] > 0.85) only when combining titers against different classes of antigens, suggesting cross-functional antigenic diversity. Anti- IgG levels in MENSA decreased with successful FST and rose with reinfection. In contrast, IgG levels in serum remained unchanged throughout the 12-week FST. Collectively, these results demonstrate the applicability of serum and MENSA for diagnosis of DFI with increased power by combining functionally distinct titers. We also found that tracking MENSA has prognostic potential to guide clinical decisions during FST.
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http://dx.doi.org/10.1128/IAI.00629-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6246899PMC
December 2018

Exacerbated Foot Infections in Obese/Diabetic Mice Are Associated with Impaired Germinal Center Reactions, Ig Class Switching, and Humoral Immunity.

J Immunol 2018 07 1;201(2):560-572. Epub 2018 Jun 1.

Department of Pathology and Laboratory Medicine, University of Rochester, Rochester, NY 14642;

Obese patients with type 2 diabetes (T2D) are at an increased risk of foot infection, with impaired immune function believed to be a critical factor in the infectious process. In this study, we test the hypothesis that humoral immune defects contribute to exacerbated foot infection in a murine model of obesity/T2D. C57BL/6J mice were rendered obese and T2D by a high-fat diet for 3 mo and were compared with controls receiving a low-fat diet. Following injection of into the footpad, obese/T2D mice had greater foot swelling and reduced clearance than controls. Obese/T2D mice also had impaired humoral immune responses as indicated by lower total IgG levels and lower anti- Ab production. Within the draining popliteal lymph nodes of obese/T2D mice, germinal center formation was reduced, and the percentage of germinal center T and B cells was decreased by 40-50%. Activation of both T and B lymphocytes was similarly suppressed in obese/T2D mice. Impaired humoral immunity in obesity/T2D was independent of active infection, as a similarly impaired humoral immune response was demonstrated when mice were administered an digest. Isolated splenic B cells from obese/T2D mice activated normally but had markedly suppressed expression of , with diminished IgG and IgE responses. These results demonstrate impaired humoral immune responses in obesity/T2D, including B cell-specific defects in Ab production and class-switch recombination. Together, the defects in humoral immunity may contribute to the increased risk of foot infection in obese/T2D patients.
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http://dx.doi.org/10.4049/jimmunol.1800253DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6039240PMC
July 2018

Targeting the gut microbiome to treat the osteoarthritis of obesity.

JCI Insight 2018 04 19;3(8). Epub 2018 Apr 19.

Center for Musculoskeletal Research.

Obesity is a risk factor for osteoarthritis (OA), the greatest cause of disability in the US. The impact of obesity on OA is driven by systemic inflammation, and increased systemic inflammation is now understood to be caused by gut microbiome dysbiosis. Oligofructose, a nondigestible prebiotic fiber, can restore a lean gut microbial community profile in the context of obesity, suggesting a potentially novel approach to treat the OA of obesity. Here, we report that - compared with the lean murine gut - obesity is associated with loss of beneficial Bifidobacteria, while key proinflammatory species gain in abundance. A downstream systemic inflammatory signature culminates with macrophage migration to the synovium and accelerated knee OA. Oligofructose supplementation restores the lean gut microbiome in obese mice, in part, by supporting key commensal microflora, particularly Bifidobacterium pseudolongum. This is associated with reduced inflammation in the colon, circulation, and knee and protection from OA. This observation of a gut microbiome-OA connection sets the stage for discovery of potentially new OA therapeutics involving strategic manipulation of specific microbial species inhabiting the intestinal space.
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http://dx.doi.org/10.1172/jci.insight.95997DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5931133PMC
April 2018

Prevalence of High-Risk Human Papillomavirus in Tonsil Tissue in Healthy Adults and Colocalization in Biofilm of Tonsillar Crypts.

JAMA Otolaryngol Head Neck Surg 2018 Mar;144(3):231-237

Department of Otolaryngology, University of Rochester Medical Center, Rochester, New York.

Importance: The pathogenesis of human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma is currently an important topic of elucidation. The presence of latent HPV infection in tonsil tissue of healthy adults may provide an explanation for a component of this process and contribute to the understanding of HPV-associated squamous cell carcinoma oncogenesis of the oropharynx.

Objective: To determine the prevalence of oropharyngeal HPV and to determine the spatial relationship between the virus and crypt biofilm in tonsil tissue.

Design, Setting, And Participants: A retrospective, cross-sectional study was carried out using samples obtained from tonsils that were archived at a university hospital following elective nononcologic tonsillectomy from 2012 to 2015. Samples consisted of formalin-fixed paraffin embedded samples of tumor-free tonsil tissue from 102 adults between the ages of 20 and 39 years.

Exposures: Human papillomavirus status was assessed by polymerase chain reaction, and high-risk subtypes 16 and 18 were assessed with quantitative polymerase chain reaction assay. Samples that demonstrated presence of HPV were then analyzed by in situ hybridization to localize the viral capsid protein. These samples were then stained with concanavalin A to establish biofilm presence and morphology. These samples were also stained with diamidino-phenylindole (DAPI) to visualize location of the virus in relation to cell nuclei. These data were then assembled for aggregate analysis to colocalize HPV in the biofilm of the tonsillar crypts.

Main Outcomes And Measures: Outcome measurements were determined prior to data collection and include prevalence of high-risk HPV types 16 and 18 in tonsil tissue of otherwise healthy adults, as well as demonstration with immunohistochemistry of HPV in tonsillar crypt biofilm.

Results: In 102 otherwise healthy adults (55 [53.9%] female; age range, 20-39 years), the overall prevalence of HPV in tonsils was 4.9% (n = 5); and high-risk type 16 or 18, 3.9% (n = 4). In this sample population, in situ hybridization colocalized HPV virus to the biofilm of the tonsillar crypts.

Conclusions And Relevance: Biofilm is present in the tonsillar crypts in a considerable proportion of tonsil tissues and may be reproducibly identified. Human papillomavirus is demonstrated to colocalize to the crypt biofilm. This has important implications with respect to the determination of HPV prevalence rates in the oropharynx. It may also play a role in the pathogenesis of HPV-related oropharyngeal carcinoma.
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http://dx.doi.org/10.1001/jamaoto.2017.2916DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5885877PMC
March 2018

Impact of prematurity and nutrition on the developing gut microbiome and preterm infant growth.

Microbiome 2017 Dec 11;5(1):158. Epub 2017 Dec 11.

Genomics Research Center, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA.

Background: Identification of factors that influence the neonatal gut microbiome is urgently needed to guide clinical practices that support growth of healthy preterm infants. Here, we examined the influence of nutrition and common practices on the gut microbiota and growth in a cohort of preterm infants.

Results: With weekly gut microbiota samples spanning postmenstrual age (PMA) 24 to 46 weeks, we developed two models to test associations between the microbiota, nutrition and growth: a categorical model with three successive microbiota phases (P1, P2, and P3) and a model with two periods (early and late PMA) defined by microbiota composition and PMA, respectively. The more significant associations with phase led us to use a phase-based framework for the majority of our analyses. Phase transitions were characterized by rapid shifts in the microbiota, with transition out of P1 occurring nearly simultaneously with the change from meconium to normal stool. The rate of phase progression was positively associated with gestational age at birth, and delayed transition to a P3 microbiota was associated with growth failure. We found distinct bacterial metabolic functions in P1-3 and significant associations between nutrition, microbiota phase, and infant growth.

Conclusion: The phase-dependent impact of nutrition on infant growth along with phase-specific metabolic functions suggests a pioneering potential for improving growth outcomes by tailoring nutrient intake to microbiota phase.
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http://dx.doi.org/10.1186/s40168-017-0377-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5725645PMC
December 2017

Obesity/type 2 diabetes increases inflammation, periosteal reactive bone formation, and osteolysis during Staphylococcus aureus implant-associated bone infection.

J Orthop Res 2018 06 3;36(6):1614-1623. Epub 2018 Jan 3.

Department of Pathology and Laboratory Medicine, University of Rochester, Box 626, 601 Elmwood Ave., Rochester, New York, 14642.

Obese and type 2 diabetic (T2D) patients have a fivefold increased rate of infection following placement of an indwelling orthopaedic device. Though implant infections are associated with inflammation, periosteal reactive bone formation, and osteolysis, the effect of obesity/T2D on these complicating factors has not been studied. To address this question, C57BL/6J mice were fed a high fat diet (60% Kcal from fat) to induce obesity/T2D, or a control diet (10% Kcal from fat) for 3 months, and challenged with a transtibial pin coated with a bioluminescent USA300 strain of S. aureus. In the resulting infected bone, obesity/T2D was associated with increased S. aureus proliferation and colony forming units. RNA sequencing of the infected tibiae on days 7 and 14 revealed an increase in 635 genes in obese/T2D mice relative to controls. Pathways associated with ossification, angiogenesis, and immunity were enriched. MicroCT and histology on days 21 and 35 demonstrated significant increased periosteal reactive bone formation in infected obese/T2D mice versus infected controls (p < 0.05). The enhanced periosteal bone formation was associated with increased osteoblastic activity and robust endochondral ossification, with persistant cartilage on day 21 that was only observed in infected obesity/T2D. Osteolysis and osteoclast numbers in obesity/T2D were also significantly increased versus infected controls (p < 0.05). Consistent with an up-regulated immune transcriptome, macrophages were more abundant within both the periosteum and the new reactive bone of obese/T2D mice. In conclusion, we find that implant-associated S. aureus osteomyelitis in obesity/T2D is associated with increased inflammation, reactive bone formation, and osteolysis. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1614-1623, 2018.
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http://dx.doi.org/10.1002/jor.23831DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995608PMC
June 2018

Editor's Highlight: Variation in Methylmercury Metabolism and Elimination Status in Humans Following Fish Consumption.

Toxicol Sci 2018 02;161(2):443-453

Department of Environmental Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642.

Evaluating the potential for methylmercury (MeHg) toxicity relies on accurately predicting the mercury (Hg) body burden that results from eating fish. Hg body burden is directly determined by the slow elimination kinetics of MeHg in the human body (kel = 0.014 days-1 or t1/2 =50 days). Existing studies on MeHg half-life in humans demonstrate a wide range values (t1/2 = 30 to >150 days) and has lead to uncertainty in the derivation of a regulatory standard for acceptable daily oral intake. The causes of variation in MeHg toxicokinetics in humans remain little explored. Here we characterize variation in human MeHg metabolism and elimination rate (kel) in 37 adult volunteers who consumed 3 fish meals. We determined MeHg elimination rates via longitudinal Hg analysis in single hairs using laser ablation inductively coupled plasma mass spectrometry. We also measured MeHg metabolism (biotransformation) via speciation of fecal Hg. We find an average kel = 0.0157 days-1 (t1/2 = 44 days) amongst a more than 2-fold variation in kel across the cohort (0.0248-0.0112 days-1; t1/2 = 28-62 days). Although MeHg biotransformation varied widely between individuals, it showed a positive association with elimination rates across the cohort. A more than 2-fold change in kel over a period of 2 years was seen in some individuals. In 2 individuals, who received antibiotic for unrelated health issues, elimination rate was seen to slow significantly. Associations of kel with age, body mass index, gender, and fish eating habits were not observed. We establish that a measure of methylmercury metabolism and eliminaiton status (MerMES) can reduce uncertainty in determining an individual's MeHg toxicokinetics subsequent to eating fish.
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http://dx.doi.org/10.1093/toxsci/kfx226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5837402PMC
February 2018

Transcriptomic Biomarkers to Discriminate Bacterial from Nonbacterial Infection in Adults Hospitalized with Respiratory Illness.

Sci Rep 2017 07 26;7(1):6548. Epub 2017 Jul 26.

Division of Infectious Diseases, Department of Medicine, University of Rochester School Medicine and Rochester General Hospital, Rochester, NY, USA.

Lower respiratory tract infection (LRTI) commonly causes hospitalization in adults. Because bacterial diagnostic tests are not accurate, antibiotics are frequently prescribed. Peripheral blood gene expression to identify subjects with bacterial infection is a promising strategy. We evaluated whole blood profiling using RNASeq to discriminate infectious agents in adults with microbiologically defined LRTI. Hospitalized adults with LRTI symptoms were recruited. Clinical data and blood was collected, and comprehensive microbiologic testing performed. Gene expression was measured using RNASeq and qPCR. Genes discriminatory for bacterial infection were identified using the Bonferroni-corrected Wilcoxon test. Constrained logistic models to predict bacterial infection were fit using screened LASSO. We enrolled 94 subjects who were microbiologically classified; 53 as "non-bacterial" and 41 as "bacterial". RNAseq and qPCR confirmed significant differences in mean expression for 10 genes previously identified as discriminatory for bacterial LRTI. A novel dimension reduction strategy selected three pathways (lymphocyte, α-linoleic acid metabolism, IGF regulation) including eleven genes as optimal markers for discriminating bacterial infection (naïve AUC = 0.94; nested CV-AUC = 0.86). Using these genes, we constructed a classifier for bacterial LRTI with 90% (79% CV) sensitivity and 83% (76% CV) specificity. This novel, pathway-based gene set displays promise as a method to distinguish bacterial from nonbacterial LRTI.
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http://dx.doi.org/10.1038/s41598-017-06738-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5529430PMC
July 2017

Adaptive Upregulation of Clumping Factor A (ClfA) by Staphylococcus aureus in the Obese, Type 2 Diabetic Host Mediates Increased Virulence.

Infect Immun 2017 06 23;85(6). Epub 2017 May 23.

Department of Pathology & Laboratory Medicine, University of Rochester, Rochester, New York, USA

Obesity and associated type 2 diabetes (T2D) are important risk factors for infection following orthopedic implant surgery. , the most common pathogen in bone infections, adapts to multiple environments to survive and evade host immune responses. Whether adaptation of to the unique environment of the obese/T2D host accounts for its increased virulence and persistence in this population is unknown. Thus, we assessed implant-associated osteomyelitis in normal versus high-fat-diet obese/T2D mice and found that infection was more severe, including increases in bone abscesses relative to nondiabetic controls. isolated from bone of obese/T2D mice displayed marked upregulation of four adhesion genes (, , , and ), all with binding affinity for fibrin(ogen). Immunostaining of infected bone revealed increased fibrin deposition surrounding bacterial abscesses in obese/T2D mice. coagulation assays demonstrated a hypercoagulable state in obese/T2D mice that was comparable to that of diabetic patients. with an inactivating mutation in clumping factor A () showed a reduction in bone infection severity that eliminated the effect of obesity/T2D, while infections in control mice were unchanged. In infected mice that overexpress plasminogen activator inhibitor-1 (PAI-1), expression and fibrin-encapsulated abscess communities in bone were also increased, further linking fibrin deposition to expression of and infection severity. Together, these results demonstrate an adaptation by to obesity/T2D with increased expression of that is associated with the hypercoagulable state of the host and increased virulence of .
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http://dx.doi.org/10.1128/IAI.01005-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5442639PMC
June 2017

Antimicrobials from human skin commensal bacteria protect against and are deficient in atopic dermatitis.

Sci Transl Med 2017 02;9(378)

Department of Dermatology, University of California, San Diego, La Jolla, CA 92092, USA.

The microbiome can promote or disrupt human health by influencing both adaptive and innate immune functions. We tested whether bacteria that normally reside on human skin participate in host defense by killing , a pathogen commonly found in patients with atopic dermatitis (AD) and an important factor that exacerbates this disease. High-throughput screening for antimicrobial activity against was performed on isolates of coagulase-negative (CoNS) collected from the skin of healthy and AD subjects. CoNS strains with antimicrobial activity were common on the normal population but rare on AD subjects. A low frequency of strains with antimicrobial activity correlated with colonization by The antimicrobial activity was identified as previously unknown antimicrobial peptides (AMPs) produced by CoNS species including and These AMPs were strain-specific, highly potent, selectively killed , and synergized with the human AMP LL-37. Application of these CoNS strains to mice confirmed their defense function in vivo relative to application of nonactive strains. Strikingly, reintroduction of antimicrobial CoNS strains to human subjects with AD decreased colonization by These findings show how commensal skin bacteria protect against pathogens and demonstrate how dysbiosis of the skin microbiome can lead to disease.
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http://dx.doi.org/10.1126/scitranslmed.aah4680DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5600545PMC
February 2017

Viral Respiratory Infections in Preterm Infants during and after Hospitalization.

J Pediatr 2017 03 30;182:53-58.e3. Epub 2016 Dec 30.

Department of Pediatrics, University of Rochester Medical Center, Rochester, NY; Department of Environmental Medicine, University of Rochester Medical Center, Rochester, NY.

Objective: To determine the burden of viral respiratory infections in preterm infants both during and subsequent to neonatal intensive care unit (NICU) hospitalization and to compare this with term infants living in the community.

Study Design: From March 2013 through March 2015, we enrolled 189 newborns (96 term and 93 preterm) into a prospective, longitudinal study obtaining nose/throat swabs within 7 days of birth, weekly while hospitalized and then monthly to 4 months after hospital discharge. Taqman array cards were used to identify 16 viral respiratory pathogens by real-time polymerase chain reaction. Demographic, clinical, and laboratory data were gathered from electronic medical records, and parent interview while hospitalized with interval histories collected at monthly visits. The hospital course of all preterm infants who underwent late-onset sepsis evaluations was reviewed.

Results: Over 119 weeks, we collected 618 nose/throat swabs from at risk preterm infants in our level IV regional NICU. Only 4 infants had viral respiratory infections, all less than 28 weeks gestation at birth. Two infants were symptomatic with the infections recognized by the clinical team. The daily risk of acquiring a respiratory viral infection in preterm infants in the NICU was significantly lower than in the full term cohort living in the community. Once discharged from the hospital, viral respiratory infections were common in all infants.

Conclusions: Viral respiratory infections are infrequent in a NICU with strict infection prevention strategies and do not appear to cause unrecognized illness. Both preterm and term infants living in the community quickly acquire respiratory viral infections.
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http://dx.doi.org/10.1016/j.jpeds.2016.11.077DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5328856PMC
March 2017

Candida albicans Carriage in Children with Severe Early Childhood Caries (S-ECC) and Maternal Relatedness.

PLoS One 2016 14;11(10):e0164242. Epub 2016 Oct 14.

Eastman Institute for Oral Health, University of Rochester Medical Center, Rochester, NY, United States of America.

Introduction: Candida albicans has been detected together with Streptococcus mutans in high numbers in plaque-biofilm from children with early childhood caries (ECC). The goal of this study was to examine the C. albicans carriage in children with severe early childhood caries (S-ECC) and the maternal relatedness.

Methods: Subjects in this pilot cross-sectional study were recruited based on a convenient sample. DMFT(S)/dmft(s) caries and plaque scores were assessed during a comprehensive oral exam. Social-demographic and related background information was collected through a questionnaire. Saliva and plaque sample from all children and mother subjects were collected. C. albicans were isolated by BBL™ CHROMagar™ and also identified using germ tube test. S. mutans was isolated using Mitis Salivarius with Bacitracin selective medium and identified by colony morphology. Genetic relatedness was examined using restriction endonuclease analysis of the C. albicans genome using BssHII (REAG-B). Multilocus sequence typing was used to examine the clustering information of isolated C. albicans. Spot assay was performed to examine the C. albicans Caspofungin susceptibility between S-ECC children and their mothers. All statistical analyses (power analysis for sample size, Spearman's correlation coefficient and multiple regression analyses) were implemented with SAS 9.4.

Results: A total of 18 S-ECC child-mother pairs and 17 caries free child-mother pairs were enrolled in the study. Results indicated high C. albicans carriage rate in the oral cavity (saliva and plaque) of both S-ECC children and their mothers (>80%). Spearman's correlation coefficient also indicated a significant correlation between salivary and plaque C. albicans and S. mutans carriage (p<0.01) and caries severity (p<0.05). The levels of C. albicans in the prepared saliva and plaque sample (1ml resuspension) of S-ECC children were 1.3 ± 4.5 x104 cfu/ml and 1.2 ± 3.5 x104 cfu/ml (~3-log higher vs. caries-free children). Among 18 child-mother pairs, >60% of them demonstrated identical C. albicans REAG-B pattern. C. albicans isolated from >65% of child-mother pairs demonstrated similar susceptibility to caspofungin in spot assay, while no caspofungin resistant strains were seen when compared with C. albicans wild-type strain SC5314. Interestingly, the regression analysis showed that factors such as antibiotic usage, birth weight, inhaler use, brushing frequency, and daycare attendance had no significant effect on the oral carriage of C. albicans in the S-ECC children.

Conclusions: Our results reveal that both the child with S-ECC and the mother were highly infected with C. albicans, while most of the strains were genetically related, suggesting that the mother might be a source for C. albicans acquisition in the oral cavity of children affected by the disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0164242PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5065202PMC
May 2017

The Healthy Infant Nasal Transcriptome: A Benchmark Study.

Sci Rep 2016 Sep 23;6:33994. Epub 2016 Sep 23.

Division of Neonatology and Pediatric Molecular and Personalized Medicine Program, University of Rochester Medical Center, Rochester NY, USA.

Responses by resident cells are likely to play a key role in determining the severity of respiratory disease. However, sampling of the airways poses a significant challenge, particularly in infants and children. Here, we report a reliable method for obtaining nasal epithelial cell RNA from infants for genome-wide transcriptomic analysis, and describe baseline expression characteristics in an asymptomatic cohort. Nasal epithelial cells were collected by brushing of the inferior turbinates, and gene expression was interrogated by RNA-seq analysis. Reliable recovery of RNA occurred in the absence of adverse events. We observed high expression of epithelial cell markers and similarity to the transcriptome for intrapulmonary airway epithelial cells. We identified genes displaying low and high expression variability, both inherently, and in response to environmental exposures. The greatest gene expression differences in this asymptomatic cohort were associated with the presence of known pathogenic viruses and/or bacteria. Robust bacteria-associated gene expression patterns were significantly associated with the presence of Moraxella. In summary, we have developed a reliable method for interrogating the infant airway transcriptome by sampling the nasal epithelium. Our data demonstrates both the fidelity and feasibility of our methodology, and describes normal gene expression and variation within a healthy infant cohort.
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http://dx.doi.org/10.1038/srep33994DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034274PMC
September 2016

Sequential Evolution of Vancomycin-Intermediate Resistance Alters Virulence in Staphylococcus aureus: Pharmacokinetic/Pharmacodynamic Targets for Vancomycin Exposure.

Antimicrob Agents Chemother 2015 Dec 28;60(3):1584-91. Epub 2015 Dec 28.

Laboratory for Antimicrobial Pharmacodynamics, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, Buffalo, New York, USA New York State Center of Excellence in Life Sciences and Bioinformatics, University at Buffalo, Buffalo, New York, USA

Staphylococcus aureus possesses exceptional virulence and a remarkable ability to adapt in the face of antibiotic therapy. We examined the in vitro evolution of S. aureus in response to escalating vancomycin exposure by evaluating bacterial killing and the progression of resistance. A hollow-fiber infection model was utilized to simulate human doses of vancomycin increasing from 0.5 to 4 g every 12 h (q12h) versus a high inoculum (10(8) CFU/ml) of methicillin-resistant S. aureus (MRSA) USA300 and USA400. Host-pathogen interactions using Galleria mellonella and accessory gene regulator (agr) expression were studied in serially obtained isolates. In both USA300 and USA400 MRSA isolates, vancomycin exposure up to 2 g q12h resulted in persistence and regrowth, whereas 4 g administered q12h achieved sustained killing against both strains. As vancomycin exposure increased from 0.5 to 2 g q12h, the bacterial population shifted toward vancomycin-intermediate resistance, and collateral increases in the MICs of daptomycin and televancin were observed over 10 days. Guideline-recommended exposure of a ratio of the area under the concentration-time curve for the free, unbound fraction of the drug to the MIC (fAUC/MIC ratio) of 200 displayed a 0.344-log bacterial reduction in area, whereas fAUC/MICs of 371 and 554 were needed to achieve 1.00- and 2.00-log reductions in area, respectively. The stepwise increase in resistance paralleled a decrease in G. mellonella mortality (P = 0.021) and a gradual decline of RNAIII expression over 10 days. Currently recommended doses of vancomycin resulted in amplification of resistance and collateral damage to other antibiotics. Decreases in agr expression and virulence during therapy may be an adaptive mechanism of S. aureus persistence.
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http://dx.doi.org/10.1128/AAC.02657-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4776009PMC
December 2015