Molt-dependent transcriptomic analysis of cement proteins in the barnacle Amphibalanus amphitrite

Abstract Background A complete understanding of barnacle adhesion remains elusive as the process occurs within and beneath the confines of a rigid calcified shell. Barnacle cement is mainly proteinaceous and several individual proteins have been identified in the hardened cement at the barnacle-substrate interface. Little is known about the molt- and tissue-specific expression of cement protein genes but could offer valuable insight into the complex multi-step processes of barnacle growth and adhesion. Methods The main body and sub-mantle tissue of the barnacle Amphibalanus amphitrite (basionym Balanus amphitrite) were collected in pre- and post-molt stages. RNA-seq technology was used to analyze the transcriptome for differential gene expression at these two stages and liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) was used to analyze the protein content of barnacle secretions. Results We report on the transcriptomic analysis of barnacle cement gland tissue in pre- and post-molt growth stages and proteomic investigation of barnacle secretions. While no significant difference was found in the expression of cement proteins genes at pre- and post-molting stages, expression levels were highly elevated in the sub-mantle tissue (where the cement glands are located) compared to the main barnacle body. We report the discovery of a novel 114kD cement protein, which is identified in material secreted onto various surfaces by adult barnacles and with the encoding gene highly expressed in the sub-mantle tissue. Further differential gene expression analysis of the sub-mantle tissue samples reveals a limited number of genes highly expressed in pre-molt samples with a range of functions including cuticular development, biominerialization, and proteolytic activity. Conclusions The expression of cement protein genes appears to remain constant through the molt cycle and is largely confined to the sub-mantle tissue. Our results reveal a novel and potentially prominent protein to the mix of cement-related components in A. amphitrite. Despite the lack of a complete genome, sample collection allowed for extended transcriptomic analysis of pre- and post-molt barnacle samples and identified a number of highly-expressed genes. Our results highlight the complexities of this sessile marine organism as it grows via molt cycles and increases the area over which it exhibits robust adhesion to its substrate.http://deepblue.lib.umich.edu/bitstream/2027.42/115487/1/12864_2015_Article_2076.pd

A retrospective epidemiologic study to define risk factors, microbiology, and clinical outcomes of infective endocarditis in a large tertiary-care teaching hospital

Objective: This study aims to define risk factors as well as their association with microbiology and clinical outcomes in a large US infective endocarditis population. Methods: Hospital records were searched for appropriate infective endocarditis–related ICD codes from 16 July 2007 to 13 August 2015. A total of 363 cases were retrospectively identified that met definite Modified Duke Criteria for infective endocarditis and were analyzed by age group, causative organism, and associated risk factors for use of valvular surgical intervention, 30/90/180-day mortality after admission, and embolic phenomena. Results: Chronic hemodialysis was the most common risk factor (26.7% of cases). Of all age groups, those aged 78+ years had the lowest 30-day mortality but those aged 58–77 years had the highest mortality (p = 0.039). Staphylococcus aureus was the most prevalent causative organism. Those aged 78–97 years were more likely to have enterococcal infective endocarditis than those aged 18–27 years (p = 0.0144). Chronic hemodialysis associated infective endocarditis was more likely to be caused by coagulase-negative staphylococcus (p = 0.0121) and have a higher 30-day mortality (p = 0.141) than intravenous drug use associated infective endocarditis. Intravenous drug use and chronic hemodialysis were similarly likely to be caused by S. aureus . Intravenous drug use associated infective endocarditis was more likely to be caused by viridans group streptococci (p = 0.0001). Fungal infective endocarditis was most likely to embolize. Chronic hemodialysis patients were less likely to undergo valvular surgery (p = 0.001) and those with chronic hemodialysis who did had lower mortality than those only managed medically that did not reach statistical significance (p = 0.2991). Infective endocarditis caused by coagulase-negative staphylococci had the greatest 30-day mortality at 31.3% but did not reach statistical significance over all other causative organisms (p = 0.060). Conclusion: In our infective endocarditis population, S. aureus is the predominant causative organism. Chronic hemodialysis is the most common risk factor present in infective endocarditis populations and has greater association with coagulase-negative staphylococci and 30-day mortality. Intravenous drug use had the lowest mortality among risk factors with a similar proportion of S. aureus infective endocarditis compared to chronic hemodialysis but a higher proportion of viridans group streptococci infective endocarditis cases. Further study will need to be performed on prevention and treatment of infective endocarditis in chronic hemodialysis patients

Clinical outcomes and inflammatory marker levels in patients with Covid-19 and obesity at an inner-city safety net hospital.

ObjectivesPatients with Covid-19 and obesity have worse clinical outcomes which may be driven by increased inflammation. This study aimed to characterize the association between clinical outcomes in patients with obesity and inflammatory markers.MethodsWe analyzed data for patients aged ≥18 years admitted with a positive SARS-CoV-2 PCR test. We used multivariate logistic regression to determine the association between BMI and intensive care unit (ICU) transfer and all-cause mortality. Inflammatory markers (C-reactive protein [CRP], lactate dehydrogenase [LDH], ferritin, and D-dimer) were compared between patients with and without obesity (body mass index [BMI] ≥30 kg/m2).ResultsOf 791 patients with Covid-19, 361 (45.6%) had obesity. In multivariate analyses, BMI ≥35 was associated with a higher odds of ICU transfer (adjusted odds ratio [aOR] 2.388 (95% confidence interval [CI]: 1.074-5.310) and hospital mortality (aOR = 4.3, 95% CI: 1.69-10.82). Compared to those with BMIConclusionsPatients with obesity were more likely to have poor outcomes even without increased inflammation

Vibrio campbellii hmgA-mediated pyomelanization impairs quorum sensing, virulence, and cellular fitness

Melanization due to the inactivation of the homogentisate-1,2-dioxygenase gene (hmgA) has been demonstrated to increase stress resistance, persistence, and virulence in some bacterial species but such pigmented mutants have not been observed in pathogenic members of the Vibrio Harveyi clade. In this study, we used Vibrio campbellii ATCC BAA-1116 as model organism to understand how melanization affected cellular phenotype, metabolism, and virulence. An in-frame deletion of the hmgA gene resulted in the overproduction of a pigment in cell culture supernatants and cellular membranes that was identified as pyomelanin. Unlike previous demonstrations in Vibrio cholerae, Burkholderia cepacia, and Pseudomonas aeruginosa, the pigmented V. campbellii mutant did not show increased UV resistance and was found to be ~2.7 times less virulent than the wild type strain in Penaeus monodon shrimp virulence assays. However, the extracted pyomelanin pigment did confer a higher resistance to oxidative stress when incubated with wild type cells. Microarray-based transcriptomic analyses revealed that the hmgA gene deletion and subsequent pyomelanin production negatively effected the expression of 129 genes primarily involved in energy production, amino acid, and lipid metabolism, and protein translation and turnover. This transcriptional response was mediated in part by an impairment of the quorum sensing regulon as transcripts of the quorum sensing high cell density master regulator LuxR and other operonic members of this regulon were significantly less abundant in the hmgA mutant. Taken together, the results suggest that the pyomelanization of V. campbellii sufficiently impairs the metabolic activities of this organism and renders it less fit and virulent than its isogenic wild type strain.United States. Office of Naval Researc

Adaptation of the Black Yeast <em>Wangiella dermatitidis</em> to Ionizing Radiation: Molecular and Cellular Mechanisms

<div><p>Observations of enhanced growth of melanized fungi under low-dose ionizing radiation in the laboratory and in the damaged Chernobyl nuclear reactor suggest they have adapted the ability to survive or even benefit from exposure to ionizing radiation. However, the cellular and molecular mechanism of fungal responses to such radiation remains poorly understood. Using the black yeast <em>Wangiella dermatitidis</em> as a model, we confirmed that ionizing radiation enhanced cell growth by increasing cell division and cell size. Using RNA-seq technology, we compared the transcriptomic profiles of the wild type and the melanin-deficient <em>wdpks1</em> mutant under irradiation and non-irradiation conditions. It was found that more than 3000 genes were differentially expressed when these two strains were constantly exposed to a low dose of ionizing radiation and that half were regulated at least two fold in either direction. Functional analysis indicated that many genes for amino acid and carbohydrate metabolism and cell cycle progression were down-regulated and that a number of antioxidant genes and genes affecting membrane fluidity were up-regulated in both irradiated strains. However, the expression of ribosomal biogenesis genes was significantly up-regulated in the irradiated wild-type strain but not in the irradiated <em>wdpks1</em> mutant, implying that melanin might help to contribute radiation energy for protein translation. Furthermore, we demonstrated that long-term exposure to low doses of radiation significantly increased survivability of both the wild-type and the <em>wdpks1</em> mutant, which was correlated with reduced levels of reactive oxygen species (ROS), increased production of carotenoid and induced expression of genes encoding translesion DNA synthesis. Our results represent the first functional genomic study of how melanized fungal cells respond to low dose ionizing radiation and provide clues for the identification of biological processes, molecular pathways and individual genes regulated by radiation.</p> </div

Up-regulated (2 fold) Ribi/RP genes.

<p>Up-regulated (2 fold) Ribi/RP genes.</p

Expression profiles of RP/Ribi genes regulated by ionizing radiation in the wild type and <i>wdpks1</i> strains.

<p>Each column represents a strain. Each row represents log2 ratio of transcripts between irradiated and non-irradiated cells for a single gene. Genes were ordered based on level of expression changes in the wild type strain.</p

Antioxidant gene regulation.

<p>Antioxidant gene regulation.</p

Verification of RNA-seq transcript changes with RT-PCR.

<p>a. Transcript ratio between irradiated and non-irradiated in the wild type strain from RNA-seq data.</p><p>b. Transcript ratio between irradiated and non-irradiated in the wild type strain from qRT-PCR data. qRT-PCR Ct values were derived from averages of duplicates from two independent biological samples.</p><p>c. Undivisible duo to no transcript was detected from non-irradiated samples.</p