Liquid-liquid phase inclusion bodies in acute and persistent parainfluenaza virus type 5 infections.

Cytoplasmic inclusion bodies (IBs) are a common feature of single-stranded, non-segmented, negative-strand RNA virus (Mononegavirales) infections and are thought to be regions of active virus transcription and replication. Here we followed the dynamics of IB formation and maintenance in cells infected with persistent and lytic/acute variants of the paramyxovirus, parainfluenza virus type 5 (PIV5). We show that there is a rapid increase in the number of small inclusions bodies up until approximately 12 h post-infection. Thereafter the number of inclusion bodies decreases but they increase in size, presumably due to the fusion of these liquid organelles that can be disrupted by osmotically shocking cells. No obvious differences were observed at these times between inclusion body formation in cells infected with lytic/acute and persistent viruses. IBs are also readily detected in cells persistently infected with PIV5, including in cells in which there is little or no ongoing virus transcription or replication. In situ hybridization shows that genomic RNA is primarily located in IBs, whilst viral mRNA is more diffusely distributed throughout the cytoplasm. Some, but not all, IBs show incorporation of 5-ethynyl-uridine (5EU), which is integrated into newly synthesized RNA, at early times post-infection. These results strongly suggest that, although genomic RNA is present in all IBs, IBs are not continuously active sites of virus transcription and replication. Disruption of IBs by osmotically shocking persistently infected cells does not increase virus protein synthesis, suggesting that in persistently infected cells most of the virus genomes are in a repressed state. The role of IBs in PIV5 replication and the establishment and maintenance of persistence is discussed.

Correction to: Incidental diagnosis of tuberous sclerosis complex by exome sequencing in three families with subclinical findings.

The published online version contain mistake in the author list. Instead of "A.M.Ilyas" it should have been "M.Ilyas ".

Incidental diagnosis of tuberous sclerosis complex by exome sequencing in three families with subclinical findings.

Tuberous sclerosis complex (TSC) is an autosomal-dominant neurocutaneous disorder characterized by lesions and benign tumors in multiple organ systems including the brain, skin, heart, eyes, kidneys, and lungs. The phenotype is highly variable, although penetrance is reportedly complete. We report the molecular diagnosis of TSC in individuals exhibiting extreme intra-familial variability, including the incidental diagnosis of asymptomatic family members. Exome sequencing was performed in three families, with probands referred for epilepsy, autism, and absent speech (Family 1); epileptic spasms (Family 2); and connective tissue disorders (Family 3.) Pathogenic variants in TSC1 or TSC2 were identified in nine individuals, including relatives with limited or no medical concerns at the time of testing. Of the nine individuals reported here, six had post-diagnosis examinations and three met clinical diagnostic criteria for TSC. One did not meet clinical criteria for a possible or definite diagnosis of TSC, and two had only a possible clinical diagnosis following post-diagnosis workup. These individuals as well as their mothers demonstrated limited features that would not raise concern for TSC in the absence of molecular results. In addition, three individuals exhibited epilepsy with normal brain MRIs, and two without seizures or intellectual disability had MRI findings fulfilling major criteria for TSC highlighting the difficulty providers face when relying on clinical criteria to guide genetic testing. Given the importance of a timely TSC diagnosis for clinical management, such cases demonstrate a potential benefit for clinical criteria to include seizures and an unbiased molecular approach to genetic testing.

Insights into the Fundamental Physiology of the Uncultured Fe-Oxidizing Bacterium Leptothrix ochracea.

Leptothrix ochracea is known for producing large volumes of iron oxyhydroxide sheaths that alter wetland biogeochemistry. For over a century, these delicate structures have fascinated microbiologists and geoscientists. Because L. ochracea still resists long-term in vitro culture, the debate regarding its metabolic classification dates back to 1885. We developed a novel culturing technique for L. ochracea using in situ natural waters and coupled this with single-cell genomics and nanoscale secondary-ion mass spectrophotometry (nanoSIMS) to probe L. ochracea's physiology. In microslide cultures L. ochracea doubled every 5.7 h and had an absolute growth requirement for ferrous iron, the genomic capacity for iron oxidation, and a branched electron transport chain with cytochromes putatively involved in lithotrophic iron oxidation. Additionally, its genome encoded several electron transport chain proteins, including a molybdopterin alternative complex III (ACIII), a cytochrome bd oxidase reductase, and several terminal oxidase genes. L. ochracea contained two key autotrophic proteins in the Calvin-Benson-Bassham cycle, a form II ribulose bisphosphate carboxylase, and a phosphoribulose kinase. L. ochracea also assimilated bicarbonate, although calculations suggest that bicarbonate assimilation is a small fraction of its total carbon assimilation. Finally, L. ochracea's fundamental physiology is a hybrid of those of the chemolithotrophic Gallionella-type iron-oxidizing bacteria and the sheathed, heterotrophic filamentous metal-oxidizing bacteria of the Leptothrix-Sphaerotilus genera. This allows L. ochracea to inhabit a unique niche within the neutrophilic iron seeps.IMPORTANCELeptothrix ochracea was one of three groups of organisms that Sergei Winogradsky used in the 1880s to develop his hypothesis on chemolithotrophy. L. ochracea continues to resist cultivation and appears to have an absolute requirement for organic-rich waters, suggesting that its true physiology remains unknown. Further, L. ochracea is an ecological engineer; a few L. ochracea cells can generate prodigious volumes of iron oxyhydroxides, changing the ecosystem's geochemistry and ecology. Therefore, to determine L. ochracea's basic physiology, we employed new single-cell techniques to demonstrate that L. ochracea oxidizes iron to generate energy and, despite having predicted genes for autotrophic growth, assimilates a fraction of the total CO2 that autotrophs do. Although not a true chemolithoautotroph, L. ochracea's physiological strategy allows it to be flexible and to extensively colonize iron-rich wetlands.

Scombrotoxic Fish Poisoning Secondary to Tuna Ingestion

Scombrotoxic poisoning results from the improper handling and refrigeration of fish containing naturally occurring histidine. Scombroid fish species such as tuna, mackerel and swordfish contain histidine, which is converted to histamine when inadequately chilled. European legislation states that scombroid fish species should be tested for the presence of histamine and mean values should be <100mg/kg1. The authors report an outbreak of scombrotoxic fish poisoning in 12 individuals following ingestion of tuna. Symptoms occurred rapidly and included flushing, headache, palpitations and diarrhoea. Fortunately, symptoms were short lived and self-limiting except in one individual, who required anti-histamine medication. Adequate refrigeration practices are crucial in preventing scombrotoxic food poisoning.

Equity, Diversity, and Inclusion Best Practices for Oral Health Researchers.

KNOWLEDGE TRANSFER STATEMENT: By following best practices related to equity, diversity, and inclusion, oral health researchers may be able to advance the study of health equity, grow a diverse research workforce, and better respond to pressing public health problems.

The Relations between Systems of Oppression and Oral Care Access in the United States.

We applied a structural intersectionality approach to cross-sectionally examine the relationships between macro-level systems of oppression, their intersections, and access to oral care in the United States. Whether and the extent to which the provision of government-funded dental services attenuates the emerging patterns of associations was also assessed in the study. To accomplish these objectives, individual-level information from over 300,000 respondents of the 2010 US Behavioral Risk Factor Surveillance System was linked with state-level data for 2000 and 2010 on structural racism, structural sexism, and income inequality, as provided by Homan et al. Using multilevel models, we investigated the relationships between systems of oppression and restricted access to oral health services among respondents at the intersections of race, gender, and poverty. The degree to which extended provision of government-funded dental services weakens the observed associations was determined in models stratified by state-level coverage of oral care. Our analyses bring to the fore intersectional groups (e.g., non-Hispanic Black women and men below the poverty line) with the highest odds of not seeing a dentist in the previous year. We also show that residing in states where high levels of structural sexism and income inequality intersect was associated with 1.3 greater odds (95% confidence interval, 1.1-1.5) of not accessing dental services in the 12 mo preceding the survey. Stratified analyses demonstrated that a more extensive provision of government-funded dental services attenuates associations between structural oppressions and restricted access to oral health care. On the basis of these and other findings, we urge researchers and health care planners to increase access to dental services in more effective and inclusive ways. Most important, we show that counteracting structural drivers of inequities in dental services access entails providing dental care for all.

Oral Health Equity Must Address Oppression: On the BSSOH Consensus Statement.

The Consensus Statement on Future Directions for the Behavioral and Social Sciences in Oral Health (Consensus Statement) is a transformational contribution to oral health research synthesizing a wide array of conceptual, methodological, and disciplinary concerns and offering strategies to improve scientific inquiry. The Consensus Statement explicates global oral health equity as a foundational concern of our field. Given this context, a key concern is missing from the statement: oppression!

NASA GEOS Composition Forecast Modeling System GEOS-CF v1.0: Stratospheric Composition.

The NASA Goddard Earth Observing System (GEOS) Composition Forecast (GEOS-CF) provides recent estimates and 5-day forecasts of atmospheric composition to the public in near-real time. To do this, the GEOS Earth system model is coupled with the GEOS-Chem tropospheric-stratospheric unified chemistry extension (UCX) to represent composition from the surface to the top of the GEOS atmosphere (0.01 hPa). The GEOS-CF system is described, including updates made to the GEOS-Chem UCX mechanism within GEOS-CF for improved representation of stratospheric chemistry. Comparisons are made against balloon, lidar, and satellite observations for stratospheric composition, including measurements of ozone (O3) and important nitrogen and chlorine species related to stratospheric O3 recovery. The GEOS-CF nudges the stratospheric O3 toward the GEOS Forward Processing (GEOS FP) assimilated O3 product; as a result the stratospheric O3 in the GEOS-CF historical estimate agrees well with observations. During abnormal dynamical and chemical environments such as the 2020 polar vortexes, the GEOS-CF O3 forecasts are more realistic than GEOS FP O3 forecasts because of the inclusion of the complex GEOS-Chem UCX stratospheric chemistry. Overall, the spatial patterns of the GEOS-CF simulated concentrations of stratospheric composition agree well with satellite observations. However, there are notable biases-such as low NO x and HNO3 in the polar regions and generally low HCl throughout the stratosphere-and future improvements to the chemistry mechanism and emissions are discussed. GEOS-CF is a new tool for the research community and instrument teams observing trace gases in the stratosphere and troposphere, providing near-real-time three-dimensional gridded information on atmospheric composition.

The cost of baby-led vs. parent-led approaches to introducing complementary foods in New Zealand.

Baby-led approaches to complementary feeding promote intake of family foods rather than infant specific foods, from the start of the complementary feeding period, which advocates suggest should be less expensive. However, this has never been formally examined. We recently completed a 2-year randomised controlled trial comparing baby-led (BLISS) and traditional spoon-feeding (Control) approaches to complementary feeding in 206 infants. Perceived expense was assessed at infant 7, 8, 9 and 12 months of age. The actual cost of intake (food offered, consumed and left over) was calculated from 3-day weighed diet records at 7 and 12 months of age. BLISS was perceived as less expensive than traditional feeding (P = 0.002), but comparisons of actual costs showed only small differences in total daily cost for food offered (NZ$0.20 and NZ$0.10 at 7 and 12 months, respectively), consumed (NZ$0.30, NZ$0.20) or left over (NZ$0.10, NZ$0.20). Baby-led approaches are not cheaper for families than traditional spoon-feeding.

RNA structure is a critical determinant of poly(A) site recognition by cleavage and polyadenylation specificity factor.

Sequence conservation among mammalian poly(A) sites is limited to the sequence AAUAAA, coupled with an amorphous downstream U- or GU-rich region. Since these sequences may also occur within the coding region of mRNAs, additional information must be required to define authentic poly(A) sites. Several poly(A) sites have been shown to contain sequences outside the core elements that enhance the efficiency of 3' processing in vivo and in vitro. The human immunodeficiency virus type 1, equine infectious anemia virus, and adenovirus L1 3' processing enhancers have been shown to promote the binding of cleavage and polyadenylation specificity factor (CPSF), the factor responsible for recognition of AAUAAA, to the pre-mRNA, thereby facilitating the assembly of a stable 3' processing complex. We have used in vitro selection to examine the mechanism by which the human immunodeficiency virus type 1 3' processing enhancer promotes the interaction of CPSF with the AAUAAA hexamer. Surprisingly, RNAs selected for efficient polyadenylation were related by structure rather than sequence. Therefore, in the absence of extensive sequence conservation, our results strongly suggest that RNA structure is a critical determinant of poly(A) site recognition by CPSF and may play a key role in poly(A) site definition.

Viral antibody titers. Comparison in patients with multiple sclerosis and rheumatoid arthritis.

Higher titers of antibodies to measles virus envelope antigens, hemolysin and hemagglutinin, Epstein-Barr virus (EBV) capsid antigen and nuclear antigen, and rubella virus hemagglutinin were demonstrated in serum samples of patients with multiple sclerosis and rheumatoid arthritis than in age- and sex-matched control subjects. A significant correlation was observed between antibodies to measles and rubella viruses both in patients with multiple sclerosis and rheumatoid arthritis, but such a correlation was not observed between antibodies to EBV and measles or rubella viruses. Whether elevated levels of antibodies to EBV are due to reactivation of the virus, or elevated levels of antibodies to all the enveloped viruses result from cross-reactions between viruses and host tissue, or perhaps reflect defects in immunoregulation, needs further investigation.

Cardiac troponin I for the diagnosis of acute myocardial infarction in the emergency department.

Cardiac troponin I (TnI) was tested in 316 consecutive patients with chest pain who were admitted to the emergency department, of whom 62 were discharged with a diagnosis of acute myocardial infarction (AMI). The TnI level was abnormal in 49 patients with AMI compared with 27 for creatine kinase (CK)-MB in the first specimen obtained at admission. All 62 patients with AMI were correctly diagnosed at admission with a combination of TnI and myoglobin testing. The overall peak performance of TnI testing in samples received within 24 hours of admission indicated high sensitivity (97%) and specificity (98%) for the diagnosis of AMI. The TnI was positive in elderly patients with myocardial injury and low CK and normal CK-MB values. These data suggest that testing for TnI could replace CK-MB and, in combination with myoglobin, could facilitate the rapid and effective triage of patients with chest pain in the emergency department.

Antibodies specific for measles virus envelope antigens and autoantibodies in patients with chronic active hepatitis.

Distinctly increased levels of antibodies to measles virus envelope antigens haemolysin and haemagglutinin were found in the sera of patients with chronic active hepatitis compared with a normal control group, using immunofluorescence and functional tests. Similarly, a higher incidence of smooth muscle antibody of both IgG and IgM classes was observed in the patients and an important correlation was found between haemolysin antibodies specific for measles virus and smooth muscle antibody of IgG and IgM classes. In contrast, there was no such correlation between the virus specific haemolysin antibodies and antinuclear antibodies. The increased levels of antibodies to measles virus envelope antigens and of autoantibodies may reflect defects in immunoregulation rather than persistent infection with measles virus.

Clinical, hematologic, and survival data from cats infected with feline immunodeficiency virus: 42 cases (1983-1988).

A retrospective study of stored feline serum samples was done to determine the infection rate of feline immunodeficiency virus in cats in central Missouri. Infected cats were compared with uninfected cats subjected to the same selection criteria on the basis of signalment, clinical signs, and CBC abnormalities. A significant incidence of virus infection was found in male cats. Neither age nor breed predilection could be identified. Infected cats were more likely to be anemic and leukopenic because of neutropenia. Cellulitis and neoplasia were more common in infected cats. A spectrum of disease severity was seen in infected cats ranging from no clinical signs to signs of severe chronic inflammatory disease. Infected cats were more likely to have clinical disease. Mean survival of infected cats was 24.4 months from the time of diagnosis.

Poster - Thurs Eve-33: Initial implementation of a novel, measurement-based IMRT QA method.

Current measurement-based QA for IMRT typically involves a composite dose delivery to a phantom. However, this approach does not allow a direct dosimetric evaluation of the delivered treatment with respect to the patient anatomy. In this work we implement a novel, measurement-based IMRT QA method which provides an accurate reconstruction of the 3D-dose distribution in the patient model. The RPC Head&Neck phantom and two clinical prostate cases have been examined to date. Step & shoot plans were developed satisfying required dose metrics. A 2D-array of dose chambers (MatriXX, IBA Dosimetry) was mounted on a linear accelerator to capture delivered fluence. The measurement data were read directly by the control software (COMPASS, IBA Dosimetry), which also provides the ability to import patient plan data from the TPS. The COMPASS software also includes a dose calculation engine and head fluence model and requires beam commissioning procedures analogous to those of a TPS. Reconstructed doses and DVHs were compared to those calculated by the TPS. The beam model in the COMPASS software was able to predict percentage depth dose and X and Y profiles for MLC-defined apertures ranging from 1×1-20×20 cm∧2 to within 1.5% (depth-dose), 2.0% (in-field profiles), and 2.5% (out-of-field profiles). Reconstructed doses in the test plans were mostly within 2% of those in the TPS. DVHs compared to <1.2%. Reconstructed doses were overlaid on CT data and contoured structures, to enable a clinically useful understanding of discrepancies as compared to the TPS plan. Research partially sponsored by IBA Dosimetry.

Properties of cerebral polyribosomes and membranes of the rough endoplasmic reticulum after ethanol dependence in rats.

Effects of physical dependence upon ethanol on the polyribosomal properties and the reconstitution of the rough endoplasmic reticulum (RER) of the brain has been examined. The purified free polyribosomes (polysomes), membrane-bound polysomes, and a fraction of RER membrane that has been stripped of polysomes were isolated from rat brain. RNA yields, amino acid incorporation activities, and electron micrographs established the purity of stripped membranes, but no differences were detected following the ethanol treatment. For the polyribosomal fractions, the stability of the mRNA/ribosomal complex was decreased after ethanol dependence as was the in vitro translation of endogenous mRNA. In the reconstitution reaction, the incubation of membranes from ethanol-treated animals with either source of purified bound polysomes resulted in higher yields of protein than when control membranes were used. The above results suggest that ethanol dependence affects the properties of both the RNA and membrane components of the RER of brain.