Combining phylogeny and coevolution improves the inference of interaction partners among paralogous proteins.

Predicting protein-protein interactions from sequences is an important goal of computational biology. Various sources of information can be used to this end. Starting from the sequences of two interacting protein families, one can use phylogeny or residue coevolution to infer which paralogs are specific interaction partners within each species. We show that these two signals can be combined to improve the performance of the inference of interaction partners among paralogs. For this, we first align the sequence-similarity graphs of the two families through simulated annealing, yielding a robust partial pairing. We next use this partial pairing to seed a coevolution-based iterative pairing algorithm. This combined method improves performance over either separate method. The improvement obtained is striking in the difficult cases where the average number of paralogs per species is large or where the total number of sequences is modest.

Social determinants of health and disparate disability accumulation in a cohort of Black, Hispanic, and White patients with multiple sclerosis.

BACKGROUND: Black and Hispanic patients with multiple sclerosis (MS) have been shown to accumulate greater multiple sclerosis-associated disability (MSAD) than White patients. Disparities in social determinants of health (SDOH) among these groups have also been reported. OBJECTIVE: To determine the extent to which associations of race and ethnicity with MSAD may be attributable to differences in SDOH. METHODS: Retrospective chart analysis of patients at an academic MS center grouped by self-identified Black (n = 95), Hispanic (n = 93), and White (n = 98) race/ethnicity. Individual patient addresses were geocoded and matched with neighborhood-level area deprivation index (ADI) and social vulnerability index (SVI). RESULTS: Average Expanded Disability Status Scale (EDSS) scores at last-recorded evaluations of White patients (1.7 ± 2.0) were significantly lower than Black (2.8 ± 2.4, p = 0.001) and Hispanic (2.6 ± 2.6, p = 0.020) patients. Neither Black race nor Hispanic ethnicity was significantly associated with EDSS in multivariable linear regression models that included individual-level SDOH indicators and either ADI or SVI. CONCLUSION: Black race and Hispanic ethnicity are not significantly associated with EDSS in models that include individual and neighborhood-level SDOH indicators. Further research should elucidate mechanisms by which structural inequities affect MS disease course.

MicrobeAnnotator: a user-friendly, comprehensive functional annotation pipeline for microbial genomes.

BACKGROUND: High-throughput sequencing has increased the number of available microbial genomes recovered from isolates, single cells, and metagenomes. Accordingly, fast and comprehensive functional gene annotation pipelines are needed to analyze and compare these genomes. Although several approaches exist for genome annotation, these are typically not designed for easy incorporation into analysis pipelines, do not combine results from different annotation databases or offer easy-to-use summaries of metabolic reconstructions, and typically require large amounts of computing power for high-throughput analysis not available to the average user. RESULTS: Here, we introduce MicrobeAnnotator, a fully automated, easy-to-use pipeline for the comprehensive functional annotation of microbial genomes that combines results from several reference protein databases and returns the matching annotations together with key metadata such as the interlinked identifiers of matching reference proteins from multiple databases [KEGG Orthology (KO), Enzyme Commission (E.C.), Gene Ontology (GO), Pfam, and InterPro]. Further, the functional annotations are summarized into Kyoto Encyclopedia of Genes and Genomes (KEGG) modules as part of a graphical output (heatmap) that allows the user to quickly detect differences among (multiple) query genomes and cluster the genomes based on their metabolic similarity. MicrobeAnnotator is implemented in Python 3 and is freely available under an open-source Artistic License 2.0 from https://github.com/cruizperez/MicrobeAnnotator . CONCLUSIONS: We demonstrated the capabilities of MicrobeAnnotator by annotating 100 Escherichia coli and 78 environmental Candidate Phyla Radiation (CPR) bacterial genomes and comparing the results to those of other popular tools. We showed that the use of multiple annotation databases allows MicrobeAnnotator to recover more annotations per genome compared to faster tools that use reduced databases and is computationally efficient for use in personal computers. The output of MicrobeAnnotator can be easily incorporated into other analysis pipelines while the results of other annotation tools can be seemingly incorporated into MicrobeAnnotator to generate summary plots.

Volume Matters: Longitudinal Retrospective Cohort Study of Outcomes Following Consultation and Standardization of Adrenal Surgery.

PURPOSE: Subspecialization of adrenal surgery through regionalization has not been adequately evaluated. We assessed implementation of subspecialization and the association of regionalization with adrenalectomy outcomes in a community-based setting. METHODS: In this longitudinal retrospective cohort study, we used an interrupted time series analysis on consecutive adrenal surgeries at Kaiser Permanente Northern California, 2010-2019. The intervention was regionalization of surgery in 2016. Main outcomes include surgical volumes, operative time, length of stay, 30-day return-to-care, and 30-day complications obtained from the electronic medical record. t-Tests and multivariable models were used to analyze time trends in outcomes after accounting for changes in patient and disease characteristics. RESULTS: In total, 850 adrenal surgery cases were eligible. Between 2010 and 2019, the annual incidence of surgery (per 100,000 persons) increased from 2.4 (95% CI 1.9-3.1) to 4.1 (95% CI 3.5-4.8). Average annual surgeon volume increased from 2.4 (95% CI 1.6-3.1) to 9.9 (95% CI 4.9-14.9), while hospital volume increased from 3.5 (95% CI 2.3-4.6) to 15.4 (95% CI 6.9-24.0). Operative time was 34 (23-45) min faster in 2018-2019 compared with 2010-2011. After regionalization, same-day discharges increased to 64% in 2019 (p < 0.0001). The frequency of return-to-care (p = 0.69) and the overall complication rate (p = 0.31) did not change. CONCLUSIONS: Regionalizing adrenal surgery through surgical subspecialization and standardized care pathways was feasible and decreased operative time, and hospital stay, while increasing the frequency of same-day discharges without increasing return-to-care or complications.

Management of primary skin cancer during a pandemic: Multidisciplinary recommendations.

During the coronavirus disease 2019 (COVID-19) pandemic, providers and patients must engage in shared decision making regarding the pros and cons of early versus delayed interventions for localized skin cancer. Patients at highest risk of COVID-19 complications are older; are immunosuppressed; and have diabetes, cancer, or cardiopulmonary disease, with multiple comorbidities associated with worse outcomes. Physicians must weigh the patient's risk of COVID-19 complications in the event of exposure against the risk of worse oncologic outcomes from delaying cancer therapy. Herein, the authors have summarized current data regarding the risk of COVID-19 complications and mortality based on age and comorbidities and have reviewed the literature assessing how treatment delays affect oncologic outcomes. They also have provided multidisciplinary recommendations regarding the timing of local therapy for early-stage skin cancers during this pandemic with input from experts at 11 different institutions. For patients with Merkel cell carcinoma, the authors recommend prioritizing treatment, but a short delay can be considered for patients with favorable T1 disease who are at higher risk of COVID-19 complications. For patients with melanoma, the authors recommend delaying the treatment of patients with T0 to T1 disease for 3 months if there is no macroscopic residual disease at the time of biopsy. Treatment of tumors ≥T2 can be delayed for 3 months if the biopsy margins are negative. For patients with cutaneous squamous cell carcinoma, those with Brigham and Women's Hospital T1 to T2a disease can have their treatment delayed for 2 to 3 months unless there is rapid growth, symptomatic lesions, or the patient is immunocompromised. The treatment of tumors ≥T2b should be prioritized, but a 1-month to 2-month delay is unlikely to worsen disease-specific mortality. For patients with squamous cell carcinoma in situ and basal cell carcinoma, treatment can be deferred for 3 months unless the individual is highly symptomatic.

Concurrent chemo-radiotherapy with proton therapy: reduced toxicity with comparable oncological outcomes vs photon chemo-radiotherapy.

Concurrent chemo-radiotherapy is a commonly employed curative treatment approach for locally advanced cancers but is associated with considerable morbidity. Chemo-radiotherapy using proton therapy may be able to reduce side effects of treatment and improve efficacy, but this remains an area of controversy and data are relatively limited. We comment on recently published studies and discuss future directions for proton therapy.

Computational Drug Repositioning for Chagas Disease Using Protein-Ligand Interaction Profiling.

Chagas disease, caused by Trypanosoma cruzi (T. cruzi), affects nearly eight million people worldwide. There are currently only limited treatment options, which cause several side effects and have drug resistance. Thus, there is a great need for a novel, improved Chagas treatment. Bifunctional enzyme dihydrofolate reductase-thymidylate synthase (DHFR-TS) has emerged as a promising pharmacological target. Moreover, some human dihydrofolate reductase (HsDHFR) inhibitors such as trimetrexate also inhibit T. cruzi DHFR-TS (TcDHFR-TS). These compounds serve as a starting point and a reference in a screening campaign to search for new TcDHFR-TS inhibitors. In this paper, a novel virtual screening approach was developed that combines classical docking with protein-ligand interaction profiling to identify drug repositioning opportunities against T. cruzi infection. In this approach, some food and drug administration (FDA)-approved drugs that were predicted to bind with high affinity to TcDHFR-TS and whose predicted molecular interactions are conserved among known inhibitors were selected. Overall, ten putative TcDHFR-TS inhibitors were identified. These exhibited a similar interaction profile and a higher computed binding affinity, compared to trimetrexate. Nilotinib, glipizide, glyburide and gliquidone were tested on T. cruzi epimastigotes and showed growth inhibitory activity in the micromolar range. Therefore, these compounds could lead to the development of new treatment options for Chagas disease.

Proteome-wide analysis of human motif-domain interactions mapped on influenza a virus.

BACKGROUND: The influenza A virus (IAV) is a constant threat for humans worldwide. The understanding of motif-domain protein participation is essential to combat the pathogen. RESULTS: In this study, a data mining approach was employed to extract influenza-human Protein-Protein interactions (PPI) from VirusMentha,Virus MINT, IntAct, and Pfam databases, to mine motif-domain interactions (MDIs) stored as Regular Expressions (RegExp) in 3DID database. A total of 107 RegExp related to human MDIs were searched on 51,242 protein fragments from H1N1, H1N2, H2N2, H3N2 and H5N1 strains obtained from Virus Variation database. A total 46 MDIs were frequently mapped on the IAV proteins and shared between the different strains. IAV kept host-like MDIs that were associated with the virus survival, which could be related to essential biological process such as microtubule-based processes, regulation of cell cycle check point, regulation of replication and transcription of DNA, etc. in human cells. The amino acid motifs were searched for matches in the immune epitope database and it was found that some motifs are part of experimentally determined epitopes on IAV, implying that such interactions exist. CONCLUSION: The directed data-mining method employed could be used to identify functional motifs in other viruses for envisioning new therapies.

Probing the global kinome and phosphoproteome in Chlamydomonas reinhardtii via sequential enrichment and quantitative proteomics.

The identification of dynamic protein phosphorylation events is critical for understanding kinase/phosphatase-regulated signaling pathways. To date, protein phosphorylation and kinase expression have been examined independently in photosynthetic organisms. Here we present a method to study the global kinome and phosphoproteome in tandem in a model photosynthetic organism, the alga Chlamydomonas reinhardtii (Chlamydomonas), using mass spectrometry-based label-free proteomics. A dual enrichment strategy targets intact protein kinases via capture on immobilized multiplexed inhibitor beads with subsequent proteolytic digestion of unbound proteins and peptide-based phosphorylation enrichment. To increase depth of coverage, both data-dependent and data-independent (via SWATH, Sequential Windowed Acquisition of All Theoretical Fragment Ion Mass Spectra) mass spectrometric acquisitions were performed to obtain a more than 50% increase in coverage of the enriched Chlamydomonas kinome over coverage found with no enrichment. The quantitative phosphoproteomic dataset yielded 2250 phosphopeptides and 1314 localized phosphosites with excellent reproducibility across biological replicates (90% of quantified sites with coefficient of variation below 11%). This approach enables simultaneous investigation of kinases and phosphorylation events at the global level to facilitate understanding of kinase networks and their influence in cell signaling events.

Energy-Dispersive Total-Reflection Resonant Inelastic X-ray Scattering as a Tool for Elemental Speciation in Contaminated Water.

This work presents a state-of the-art analytical methodology, by which chemical state information on metallic elements is obtained for liquid samples in a fast and simple manner. This method overcomes limitations of conventional X-ray techniques, such as X-ray absorption spectroscopy, by applying resonant inelastic X-ray scattering under total reflection geometry (TRIXS). TRIXS is particularly applicable for the analysis of small quantity of liquid samples deposited on polished reflectors. This feature is relevant for the chemical speciation of metallic trace elements contained in water samples, since the degree of their toxicity depends crucially on the concentration of specific chemical species included. The analytical merits of the proposed methodology were studied at Elettra Sincrotrone Trieste and at the Brazilian Synchrotron Light Laboratory. Contaminated water samples with low concentration of different chromium and manganese compounds were measured. Results prove the analytical potential of the TRIXS technique in characterizing different chemical species of metallic elements in water samples.

Quantifying the Importance of the Rare Biosphere for Microbial Community Response to Organic Pollutants in a Freshwater Ecosystem.

A single liter of water contains hundreds, if not thousands, of bacterial and archaeal species, each of which typically makes up a very small fraction of the total microbial community (<0.1%), the so-called "rare biosphere." How often, and via what mechanisms, e.g., clonal amplification versus horizontal gene transfer, the rare taxa and genes contribute to microbial community response to environmental perturbations represent important unanswered questions toward better understanding the value and modeling of microbial diversity. We tested whether rare species frequently responded to changing environmental conditions by establishing 20-liter planktonic mesocosms with water from Lake Lanier (Georgia, USA) and perturbing them with organic compounds that are rarely detected in the lake, including 2,4-dichlorophenoxyacetic acid (2,4-D), 4-nitrophenol (4-NP), and caffeine. The populations of the degraders of these compounds were initially below the detection limit of quantitative PCR (qPCR) or metagenomic sequencing methods, but they increased substantially in abundance after perturbation. Sequencing of several degraders (isolates) and time-series metagenomic data sets revealed distinct cooccurring alleles of degradation genes, frequently carried on transmissible plasmids, especially for the 2,4-D mesocosms, and distinct species dominating the post-enrichment microbial communities from each replicated mesocosm. This diversity of species and genes also underlies distinct degradation profiles among replicated mesocosms. Collectively, these results supported the hypothesis that the rare biosphere can serve as a genetic reservoir, which can be frequently missed by metagenomics but enables community response to changing environmental conditions caused by organic pollutants, and they provided insights into the size of the pool of rare genes and species.IMPORTANCE A single liter of water or gram of soil contains hundreds of low-abundance bacterial and archaeal species, the so called rare biosphere. The value of this astonishing biodiversity for ecosystem functioning remains poorly understood, primarily due to the fact that microbial community analysis frequently focuses on abundant organisms. Using a combination of culture-dependent and culture-independent (metagenomics) techniques, we showed that rare taxa and genes commonly contribute to the microbial community response to organic pollutants. Our findings should have implications for future studies that aim to study the role of rare species in environmental processes, including environmental bioremediation efforts of oil spills or other contaminants.

Novel fMRI working memory paradigm accurately detects cognitive impairment in multiple sclerosis.

BACKGROUND: Cognitive impairment (CI) cannot be diagnosed by magnetic resonance imaging (MRI). Functional magnetic resonance imaging (fMRI) paradigms, such as the immediate/delayed memory task (I/DMT), detect varying degrees of working memory (WM). Preliminary findings using I/DMT showed differences in blood oxygenation level dependent (BOLD) activation between impaired (MSCI, n = 12) and non-impaired (MSNI, n = 9) multiple sclerosis (MS) patients. OBJECTIVES: The aim of the study was to confirm CI detection based on I/DMT BOLD activation in a larger cohort of MS patients. The role of T2 lesion volume (LV) and Expanded Disability Status Scale (EDSS) in magnitude of BOLD signal was also sought. METHODS: A total of 50 patients (EDSS mean ( m) = 3.2, disease duration (DD) m = 12 years, and age m = 40 years) underwent the Minimal Assessment of Cognitive Function in Multiple Sclerosis (MACFIMS) and I/DMT. Working memory activation (WMa) represents BOLD signal during DMT minus signal during IMT. CI was based on MACFIMS. RESULTS: A total of 10 MSNI, 30 MSCI, and 4 borderline patients were included in the analyses. Analysis of variance (ANOVA) showed MSNI had significantly greater WMa than MSCI, in the left prefrontal cortex and left supplementary motor area ( p = 0.032). Regression analysis showed significant inverse correlations between WMa and T2 LV/EDSS in similar areas ( p = 0.005, 0.004, respectively). CONCLUSION: I/DMT-based BOLD activation detects CI in MS. Larger studies are needed to confirm these findings.

Unlocking the electronic genome of halogenobenzenes.

The drive to develop new organic materials for use in optoelectronic devices has created the need to understand the fundamental role functionalization plays concerning the electronic properties of conjugated molecules. Here density functional theory (DFT) is used to investigate how the HOMO-LUMO gaps of halogenobenzenes are affected as a function of substituent size, position, electronegativity, ionization potential, and polarizability. A detailed molecular orbital analysis is also provided. It is shown that the molecular static polarizability and ionization potential of the bound halogens are the primary physical descriptors governing the HOMO-LUMO gap within halogenobenzenes. Two secondary descriptors controlling the HOMO-LUMO gap in these materials are the aromaticity of the halogen substituted benzene rings (as monitored via the harmonic oscillator method of aromaticity index [HOMA]) and the reduced population of the halogen atomic orbitals in the frontier MOs (%XHOMO or %XLUMO). The molecular polarizability and aromaticity, as well as %XHOMO and %XLUMO, are shown to be a function of halogen electronegativity and size, as well as number and position on the ring. It is ultimately demonstrated that halogenobenzenes which are most polarizable and are either least aromatic and/or exhibit the smallest %XLUMO (or largest %XHOMO) values, have the smallest HOMO-LUMO gaps.

Microbial and Functional Diversity within the Phyllosphere of Espeletia Species in an Andean High-Mountain Ecosystem.

Microbial populations residing in close contact with plants can be found in the rhizosphere, in the phyllosphere as epiphytes on the surface, or inside plants as endophytes. Here, we analyzed the microbiota associated with Espeletia plants, endemic to the Páramo environment of the Andes Mountains and a unique model for studying microbial populations and their adaptations to the adverse conditions of high-mountain neotropical ecosystems. Communities were analyzed using samples from the rhizosphere, necromass, and young and mature leaves, the last two analyzed separately as endophytes and epiphytes. The taxonomic composition determined by performing sequencing of the V5-V6 region of the 16S rRNA gene indicated differences among populations of the leaf phyllosphere, the necromass, and the rhizosphere, with predominance of some phyla but only few shared operational taxonomic units (OTUs). Functional profiles predicted on the basis of taxonomic affiliations differed from those obtained by GeoChip microarray analysis, which separated community functional capacities based on plant microenvironment. The identified metabolic pathways provided insight regarding microbial strategies for colonization and survival in these ecosystems. This study of novel plant phyllosphere microbiomes and their putative functional ecology is also the first step for future bioprospecting studies in search of enzymes, compounds, or microorganisms relevant to industry or for remediation efforts.

Exploratory methodology for retrieving oxidation state information from X-ray resonant Raman scattering spectrometry.

It has been observed recently that the resonant Raman scattering (RRS) peak of an X-ray spectrum contains information about the chemical environment of the irradiated matter. This information is extracted with complex processing of the spectrum data. Principal component analysis (PCA) is a statistical multivariate technique that allows exploring the variance-covariance structure of a set of data, through a few linear combinations of the original variables. This methodology can be applied to obtain information from RRS spectra. To analyze its potentiality, several measurements of different oxides in surface nanolayers were measured in total reflection conditions using synchrotron radiation. Multivariate analysis techniques, in particular, PCA, were used to obtain the information encrypted in the RRS peak, and to establish a new methodology, simpler and more accurate. The results show that multivariate analysis techniques are suitable for the analysis of this kind of spectra, foreseeing its application in future research.

Multi-environment multi-QTL association mapping identifies disease resistance QTL in barley germplasm from Latin America.

KEY MESSAGE: Multi-environment multi-QTL mixed models were used in a GWAS context to identify QTL for disease resistance. The use of mega-environments aided the interpretation of environment-specific and general QTL. Diseases represent a major constraint for barley (Hordeum vulgare L.) production in Latin America. Spot blotch (caused by Cochliobolus sativus), stripe rust (caused by Puccinia striiformis f.sp. hordei) and leaf rust (caused by Puccinia hordei) are three of the most important diseases that affect the crop in the region. Since fungicide application is not an economically or environmentally sound solution, the development of durably resistant varieties is a priority for breeding programs. Therefore, new resistance sources are needed. The objective of this work was to detect genomic regions associated with field level plant resistance to spot blotch, stripe rust, and leaf rust in Latin American germplasm. Disease severities measured in multi-environment trials across the Americas and 1,096 SNPs in a population of 360 genotypes were used to identify genomic regions associated with disease resistance. Optimized experimental design and spatial modeling were used in each trial to estimate genotypic means. Genome-Wide Association Mapping (GWAS) in each environment was used to detect Quantitative Trait Loci (QTL). All significant environment-specific QTL were subsequently included in a multi-environment-multi-QTL (MEMQ) model. Geographical origin and inflorescence type were the main determinants of population structure. Spot blotch severity was low to intermediate while leaf and stripe rust severity was high in all environments. Mega-environments were defined by locations for spot blotch and leaf rust. Significant marker-trait associations for spot blotch (9 QTL), leaf (6 QTL) and stripe rust (7 QTL) and both global and environment-specific QTL were detected that will be useful for future breeding efforts.

Anti-Trypanosoma cruzi and anti-leishmanial activity by quinoxaline-7-carboxylate 1,4-di-N-oxide derivatives.

In this work, a novel series of ethyl and methyl quinoxaline-7-carboxylate 1,4-di-N-oxide derivatives were evaluated in vitro on Trypanosoma cruzi trypomastigotes and Leishmania mexicana promastigotes, and cytotoxicity activity in murine macrophages was tested. In silico molecular docking simulations of trypanothione reductase were also done. Three compounds of 33 quinoxaline-7-carboxylate 1,4-di-N-oxide derivatives showed better anti-T. cruzi activity than nifurtimox and beznidazole; two compounds had better anti-leishmanial activity that amphotericin-B, and two compounds showed better activity against both parasites than reference drugs. Compounds M2, M7, M8 and E5, showed low cytotoxic activity on the host cell. The in silico studies suggest that compound M2 is a potential trypanothione reductase inhibitor.

Pearls & Oy-sters: KLHL11 IgG Paraneoplastic-Associated Hearing Loss and Rhombencephalitis in a Woman With Metastatic Müllerian Tumor.

Kelch-like protein-11 (KLHL11) immunoglobulin G (IgG) is a recently reported paraneoplastic autoantibody associated with rhombencephalitis, which commonly presents with ataxia, diplopia, vertigo, hearing loss, tinnitus, and gaze palsies. The association of this high-risk paraneoplastic autoantibody with testicular germ cell tumors is widely accepted, but it has not been associated with Müllerian tumors. In this study, we report a woman without a known germ cell tumor presenting with signs and symptoms suggesting autoimmune encephalitis. She was found to have metastatic ovarian serous carcinoma with KLHL11 immunoreactivity on histopathology. This case demonstrates a rare cancer association of KLHL11 IgG-seropositive rhombencephalitis with Müllerian tumor and highlights that this autoantibody can also be detected in female patients. Thus, this case expands on the current knowledge of KLHL11-related autoimmune encephalitis including the paraneoplastic presentation, associated tumor types, and management of this syndrome in women.

Serum cholesterol levels are inversely associated with the risk of malignancy in subjects with Bethesda category IV thyroid nodules.

INTRODUCTION: Some epidemiological data suggest that there may be an inverse relationship between cholesterol levels and the risk of thyroid cancer in the overall population. The present study was aimed to evaluate the lipid profile specifically in subjects with Bethesda category IV thyroid nodules, and compare whether there were any differences between those with benign and malignant nodules. METHODS: Single-centre, retrospective study on 204 subjects treated by partial or total thyroidectomy for excision of a Bethesda category IV thyroid nodule, who had undergone a blood lipid profile test in the 12 months prior to surgery. In addition to lipid measures, other demographic, clinical, biochemical and ultrasound data were collected. RESULTS: Seventy-five subjects (36.8%) were diagnosed with thyroid carcinoma in the definitive histopathological examination. Patients with thyroid cancer had lower levels of total cholesterol, LDL-cholesterol and non-HDL-cholesterol than subjects with benign thyroid diseases. There were no differences in HDL-cholesterol, triglycerides or total cholesterol/HDL-cholesterol ratio. There were no differences either between groups in other clinical, biochemical and ultrasound variables, including the use of lipid-lowering drugs. In multivariate analysis, only LDL-cholesterol was independently associated with malignancy. Subjects with follicular carcinoma showed the lowest cholesterol levels, while those with papillary carcinoma had intermediate values between the group with follicular carcinoma and the group with benign thyroid diseases. CONCLUSIONS: In subjects with cytologically indeterminate Bethesda category IV thyroid nodules, levels of total cholesterol, non-HDL-cholesterol and, particularly, LDL-cholesterol are lower among those with malignant nodules.