Relationship between Subclinical Hypothyroidism and Uremic Pruritis in Hemodialysis Patients.

BACKGROUND AND AIM: Uremic pruritus (UP) is one of the most distressing symptoms in hemodialysis (HD) patients. Subclinical hypothyroidism (SCH) is a biochemical condition with high prevalence in HD patients. The present multicentric study aimed to assess the relationship between UP and SCH in HD patients. METHODS: The present cross-sectional study included 328 HD patients. All patients were submitted to careful history through clinical examination and standard laboratory assessment. Pruritis was evaluated using the pruritis visual analog scale (VAS). Patients were diagnosed with SCH if they had TSH levels above the upper limit of the normal reference range in association with normal free thyroxine (FT4) levels. RESULTS: Among the studied patients, there were 196 patients (59.8 %) with UP. Comparison between patients with UP and patients without revealed that patients in the former group had significantly longer HD duration (median (IQR): 47.5 (27.0-72.5) versus 36.0 (23.0-50.5) months, p < 0.001) and lower Kt/v (median (IQR): 1.4 (1.09-1.7) versus 1.54 (1.12-1.91), p = 0.009). Moreover, they had significantly higher ferritin (median (IQR): 653.0 (526.0-800.0) versus 628.0 (470.8- 716.0) ng/mL), hsCRP (median (IQR): 12.0 (8.0-14.0) versus 8.0 (6.0-9.0) mg/dL, p < 0.001) and TSH levels (median (IQR): 4.34 (1.98-5.2) versus 3.34 (1.9-4.85) µIU/ml) with a significantly higher frequency of SCH (45.9 % versus 28.8 %, p = 0.002). Logistic regression analysis identified hemodialysis duration (OR (95%) CI): 1.02 (1.009-1.028), p < 0.001), ferritin levels (OR (95% CI): 1.002 (1.001-1.003), p < 0.001), and SCH (OR (95% CI): 0.54 (0.32-0.89), p = 0.016) as significant predictors of UP. CONCLUSION: The present study suggested a possible link between SCH and the development of UP in HD patients.

Effect of Different Levels of Metabolizing Energy on Productive Performance of Two Grower Quail Strains.

Two strains of Japanese quail (Coturnix coturnix japonica), including 250 birds of desert and 250 birds of white color, all one day old and unsexed, were divided into five treatment groups with 50 replicates in each group. These treatments included five levels of metabolism energy (ME) levels, including 2700, 2800, 2900, 3000, and 3100 Kcal/Kg diet. The study included one stage from day 1 to day 42 of birds' age. The results confirmed a statistically significant difference (P≤0.05) caused by ME levels in the body weight, weight gain, feed conversion ratio (feed [g] weight gain [g]), water consumption, water conversion ratio (ml weight gain [g]), protein conversion ratio (protein [g] weight gain [g]), energy conversion ratio (Kcal weight gain [g]), carcass weight, in addition to albumin and triglyceride. Therefore, the results showed significant effects (P≤0.05) of ME levels and the interaction on feed consumption, protein consumption, edible giblet percentage, tenderness, and juiciness. Significant differences (P≤0.05) were also caused by ME levels in the total cholesterol. In addition, significant differences (P≤0.05) have been found in the interaction on mortality percentage. Net return (Iraqi Dinar/live weight [Kg]) for desert quail was better than that for the white quail (2900 Kcal/Kg diet), and the interaction effect was stronger on the desert strain with 2900 Kcal than the white strain.

Relation between erythropoietin resistance and metabolic syndrome in hemodialysis patients: A multicentric propensity score matched analysis.

Erythropoietin (EPO) resistance is frequently reported in hemodialysis (HD) patients. Metabolic syndrome (MetS) is a common biochemical condition that comprises central obesity, dyslipidemia, hypertension, and hyperglycemia. The present study aimed to assess the relation between MetS and EPO resistance in HD patients. The present multicentric study included 150 patients with EPO resistance and 150 patients without EPO resistance. Short-acting EPO resistance was diagnosed if the erythropoietin resistance index is ≥1.0 IU/kg/gHb. Comparison between patients with EPO resistance and patients without resistance revealed that the former group had significantly higher body mass index, lower hemoglobin levels, lower albumin levels, higher ferritin levels, and higher high-sensitivity C-reactive protein (hsCRP) levels. In addition, patients in the EPO resistance group had significantly higher frequency of MetS (75.3% vs 38.0%, p < 0.001) and higher number of MetS components (2.7 ± 1.3 vs 1.8 ± 1.6, p < 0.001). Multivariate logistic regression analysis identified lower albumin levels (OR (95% CI): 0.072 (0.016-0.313), p < 0.001), higher ferritin levels (OR (95% CI): 1.05 (1.033-1.066), p< 0.001), higher hsCRP levels (OR (95% CI): 1.041 (1.007-1.077), p = 0.018), and MetS (OR (95% CI): 36.68 (2.893-465.05), p = 0.005) as predictors of EPO resistance in the studied patients. The present study identified MetS as a predictor of EPO resistance in HD patients. Other predictors include serum ferritin, hsCRP, and albumin levels.

Incidence of pediatric rigid esophagoscopy for foreign body removal before and after coin currency implementation in Saudi Arabia in 2017.

BACKGROUND: Foreign body (FB) ingestion is a common problem in children, which can lead to severe complications. Coins are the most common FB ingested. Since coin currency was recently implemented in Saudi Arabia, we decided to assess whether any changes have occurred in the incidence of FB removal by esophagoscopy. OBJECTIVES: Incidence of rigid esophagoscopy for ingested FB removal before and after coin currency implementation. DESIGN: Medical record review SETTINGS: Main referral hospital in Jazan region. PATIENTS AND METHODS: Our study included pediatric patients who underwent rigid esophagoscopy for removal of FB between February 2015 and July 2020 in the otorhinolaryngology department. We reported the incidence, age, gender, and type of FB. As the coin currency implementation started on December 2, 2017, the data were analyzed before and after this date. MAIN OUTCOME MEASURES: Annual incidence of pediatric rigid esophagoscopy for removal of ingested FB and type of the FB. SAMPLE SIZE: 124 patients RESULTS: The median age and interquartile range was 6.0 (5.0) years. After implementation of coins in 2017, 104 cases were reported over 32 months; before that date, 20 cases were reported over 34 months. Coins were the FB in 2 cases (10%) before implementation of coins and in 83 cases (79.8%) after implementation (P=.0001). CONCLUSIONS: The annual incidence of pediatric esophagoscopy for FB removal has increased more than five times since implementation of coin currency. This increase is exclusively related to the increase in coins as a FB. LIMITATIONS: Retrospective study. CONFLICT OF INTEREST: None.

Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method.

Sequencing of the 16S rRNA gene (16S) has long been a go-to method for microbiome characterization due to its accessibility and lower cost compared to shotgun metagenomic sequencing (SMS). However, 16S sequencing rarely provides species-level resolution and cannot provide direct assessment of other taxa (e.g., viruses and fungi) or functional gene content. Shallow shotgun metagenomic sequencing (SSMS) has emerged as an approach to bridge the gap between 16S sequencing and deep metagenomic sequencing. SSMS is cost-competitive with 16S sequencing, while also providing species-level resolution and functional gene content insights. In the present study, we evaluated the effects of sequencing depth on marker gene-mapping- and alignment-based annotation of bacteria in healthy human stool samples. The number of identified taxa decreased with lower sequencing depths, particularly with the marker gene-mapping-based approach. Other annotations, including viruses and pathways, also showed a depth-dependent effect on feature recovery. These results refine the understanding of the suitability and shortcomings of SSMS, as well as annotation tools for metagenomic analyses in human stool samples. Results may also translate to other sample types and may open the opportunity to explore the effect of sequencing depth and annotation method.

Integrated analysis of population genomics, transcriptomics and virulence provides novel insights into Streptococcus pyogenes pathogenesis.

Streptococcus pyogenes causes 700 million human infections annually worldwide, yet, despite a century of intensive effort, there is no licensed vaccine against this bacterium. Although a number of large-scale genomic studies of bacterial pathogens have been published, the relationships among the genome, transcriptome, and virulence in large bacterial populations remain poorly understood. We sequenced the genomes of 2,101 emm28 S. pyogenes invasive strains, from which we selected 492 phylogenetically diverse strains for transcriptome analysis and 50 strains for virulence assessment. Data integration provided a novel understanding of the virulence mechanisms of this model organism. Genome-wide association study, expression quantitative trait loci analysis, machine learning, and isogenic mutant strains identified and confirmed a one-nucleotide indel in an intergenic region that significantly alters global transcript profiles and ultimately virulence. The integrative strategy that we used is generally applicable to any microbe and may lead to new therapeutics for many human pathogens.

Diagnostic accuracy of tablet-based software for the detection of concussion.

Despite the high prevalence of traumatic brain injuries (TBI), there are few rapid and straightforward tests to improve its assessment. To this end, we developed a tablet-based software battery ("BrainCheck") for concussion detection that is well suited to sports, emergency department, and clinical settings. This article is a study of the diagnostic accuracy of BrainCheck. We administered BrainCheck to 30 TBI patients and 30 pain-matched controls at a hospital Emergency Department (ED), and 538 healthy individuals at 10 control test sites. We compared the results of the tablet-based assessment against physician diagnoses derived from brain scans, clinical examination, and the SCAT3 test, a traditional measure of TBI. We found consistent distributions of normative data and high test-retest reliability. Based on these assessments, we defined a composite score that distinguishes TBI from non-TBI individuals with high sensitivity (83%) and specificity (87%). We conclude that our testing application provides a rapid, portable testing method for TBI.

Genomic Characteristics Behind the Spread of Bacteremic Group A Streptococcus Type emm89 in Finland, 2004-2014.

BACKGROUND: Many countries worldwide have reported increasing numbers of emm89 group A Streptococcus (GAS) infections during last decade. Pathogen genetic factors linked to this increase need assessment. METHODS: We investigated epidemiological characteristics of emm89 GAS bacteremic infections, including 7-day and 30-day case-fatality rates, in Finland during 2004-2014 and linked them to whole-genome sequencing data obtained from corresponding strains. The Fisher exact test and exact logistic regression were used to compare differences between bacteremic infections due to emm89 GAS belonging to different genetic clades and subclades. RESULTS: Out of 1928 cases of GAS bacteremic infection, 278 were caused by emm89 GAS. We identified 2 genetically distinct clades, arbitrarily designated clade 2 and clade 3. Both clades were present during 2004-2008, but clade 3 increased rapidly from 2009 onward. Six subclades (designated subclades A-F) were identified within clade 3, based on phylogenetic core genome analysis. The case-fatality rate differed significantly between subclades (P < .05), with subclade D having the highest 30-day estimated case-fatality rate (19% vs 3%-14%). CONCLUSIONS: A new emm89 clone, clade 3, emerged in 2009 and spread rapidly in Finland. Patients infected with certain subclades of clade 3 were significantly more likely to die. A specific polymerase chain reaction assay was developed to follow the spread of subclade D in 2015.

Genomic Landscape of Intrahost Variation in Group A Streptococcus: Repeated and Abundant Mutational Inactivation of the fabT Gene Encoding a Regulator of Fatty Acid Synthesis.

To obtain new information about Streptococcus pyogenes intrahost genetic variation during invasive infection, we sequenced the genomes of 2,954 serotype M1 strains recovered from a nonhuman primate experimental model of necrotizing fasciitis. A total of 644 strains (21.8%) acquired polymorphisms relative to the input parental strain. The fabT gene, encoding a transcriptional regulator of fatty acid biosynthesis genes, contained 54.5% of these changes. The great majority of polymorphisms were predicted to deleteriously alter FabT function. Transcriptome-sequencing (RNA-seq) analysis of a wild-type strain and an isogenic fabT deletion mutant strain found that between 3.7 and 28.5% of the S. pyogenes transcripts were differentially expressed, depending on the growth temperature (35°C or 40°C) and growth phase (mid-exponential or stationary phase). Genes implicated in fatty acid synthesis and lipid metabolism were significantly upregulated in the fabT deletion mutant strain. FabT also directly or indirectly regulated central carbon metabolism genes, including pyruvate hub enzymes and fermentation pathways and virulence genes. Deletion of fabT decreased virulence in a nonhuman primate model of necrotizing fasciitis. In addition, the fabT deletion strain had significantly decreased survival in human whole blood and during phagocytic interaction with polymorphonuclear leukocytes ex vivo We conclude that FabT mutant progeny arise during infection, constitute a metabolically distinct subpopulation, and are less virulent in the experimental models used here.

Transcriptome Remodeling Contributes to Epidemic Disease Caused by the Human Pathogen Streptococcus pyogenes.

UNLABELLED: For over a century, a fundamental objective in infection biology research has been to understand the molecular processes contributing to the origin and perpetuation of epidemics. Divergent hypotheses have emerged concerning the extent to which environmental events or pathogen evolution dominates in these processes. Remarkably few studies bear on this important issue. Based on population pathogenomic analysis of 1,200 Streptococcus pyogenes type emm89 infection isolates, we report that a series of horizontal gene transfer events produced a new pathogenic genotype with increased ability to cause infection, leading to an epidemic wave of disease on at least two continents. In the aggregate, these and other genetic changes substantially remodeled the transcriptomes of the evolved progeny, causing extensive differential expression of virulence genes and altered pathogen-host interaction, including enhanced immune evasion. Our findings delineate the precise molecular genetic changes that occurred and enhance our understanding of the evolutionary processes that contribute to the emergence and persistence of epidemically successful pathogen clones. The data have significant implications for understanding bacterial epidemics and for translational research efforts to blunt their detrimental effects. IMPORTANCE: The confluence of studies of molecular events underlying pathogen strain emergence, evolutionary genetic processes mediating altered virulence, and epidemics is in its infancy. Although understanding these events is necessary to develop new or improved strategies to protect health, surprisingly few studies have addressed this issue, in particular, at the comprehensive population genomic level. Herein we establish that substantial remodeling of the transcriptome of the human-specific pathogen Streptococcus pyogenes by horizontal gene flow and other evolutionary genetic changes is a central factor in precipitating and perpetuating epidemic disease. The data unambiguously show that the key outcome of these molecular events is evolution of a new, more virulent pathogenic genotype. Our findings provide new understanding of epidemic disease.

Trading Capsule for Increased Cytotoxin Production: Contribution to Virulence of a Newly Emerged Clade of emm89 Streptococcus pyogenes.

UNLABELLED: Strains of emm89 Streptococcus pyogenes have become one of the major causes of invasive infections worldwide in the last 10 years. We recently sequenced the genome of 1,125 emm89 strains and identified three major phylogenetic groups, designated clade 1, clade 2, and the epidemic clade 3. Epidemic clade 3 strains, which now cause the great majority of infections, have two distinct genetic features compared to clade 1 and clade 2 strains. First, all clade 3 organisms have a variant 3 nga promoter region pattern, which is associated with increased production of secreted cytolytic toxins SPN (S. pyogenes NADase) and SLO (streptolysin O). Second, all clade 3 strains lack the hasABC locus mediating hyaluronic acid capsule synthesis, whereas this locus is intact in clade 1 and clade 2 strains. We constructed isogenic mutant strains that produce different levels of SPN and SLO toxins and capsule (none, low, or high). Here we report that emm89 strains with elevated toxin production are significantly more virulent than low-toxin producers. Importantly, we also show that capsule production is dispensable for virulence in strains that already produce high levels of SPN and SLO. Our results provide new understanding about the molecular mechanisms contributing to the rapid emergence and molecular pathogenesis of epidemic clade 3 emm89 S. pyogenes. IMPORTANCE: S. pyogenes (group A streptococcus [GAS]) causes pharyngitis ("strep throat"), necrotizing fasciitis, and other human infections. Serious infections caused by emm89 S. pyogenes strains have recently increased in frequency in many countries. Based on whole-genome sequence analysis of 1,125 strains recovered from patients on two continents, we discovered that a new emm89 clone, termed clade 3, has two distinct genetic features compared to its predecessors: (i) absence of the genes encoding antiphagocytic hyaluronic acid capsule virulence factor and (ii) increased production of the secreted cytolytic toxins SPN and SLO. emm89 S. pyogenes strains with the clade 3 phenotype (absence of capsule and high expression of SPN and SLO) are highly virulent in mice. These findings provide new understanding of how new virulent clones emerge and cause severe infections worldwide. This newfound knowledge of S. pyogenes virulence can be used to help understand future epidemics and conduct new translational research.

The majority of 9,729 group A streptococcus strains causing disease secrete SpeB cysteine protease: pathogenesis implications.

Group A streptococcus (GAS), the causative agent of pharyngitis and necrotizing fasciitis, secretes the potent cysteine protease SpeB. Several lines of evidence suggest that SpeB is an important virulence factor. SpeB is expressed in human infections, protects mice from lethal challenge when used as a vaccine, and contributes significantly to tissue destruction and dissemination in animal models. However, recent descriptions of mutations in genes implicated in SpeB production have led to the idea that GAS may be under selective pressure to decrease secreted SpeB protease activity during infection. Thus, two divergent hypotheses have been proposed. One postulates that SpeB is a key contributor to pathogenesis; the other, that GAS is under selection to decrease SpeB during infection. In order to distinguish between these alternative hypotheses, we performed casein hydrolysis assays to measure the SpeB protease activity secreted by 6,775 GAS strains recovered from infected humans. The results demonstrated that 84.3% of the strains have a wild-type SpeB protease phenotype. The availability of whole-genome sequence data allowed us to determine the relative frequencies of mutations in genes implicated in SpeB production. The most abundantly mutated genes were direct transcription regulators. We also sequenced the genomes of 2,954 GAS isolates recovered from nonhuman primates with experimental necrotizing fasciitis. No mutations that would result in a SpeB-deficient phenotype were identified. Taken together, these data unambiguously demonstrate that the great majority of GAS strains recovered from infected humans secrete wild-type levels of SpeB protease activity. Our data confirm the important role of SpeB in GAS pathogenesis and help end a long-standing controversy.

A molecular trigger for intercontinental epidemics of group A Streptococcus.

The identification of the molecular events responsible for strain emergence, enhanced virulence, and epidemicity has been a long-pursued goal in infectious diseases research. A recent analysis of 3,615 genomes of serotype M1 group A Streptococcus strains (the so-called "flesh-eating" bacterium) identified a recombination event that coincides with the global M1 pandemic beginning in the early 1980s. Here, we have shown that the allelic variation that results from this recombination event, which replaces the chromosomal region encoding secreted NADase and streptolysin O, is the key driver of increased toxin production and enhanced infection severity of the M1 pandemic strains. Using isoallelic mutant strains, we found that 3 polymorphisms in this toxin gene region increase resistance to killing by human polymorphonuclear leukocytes, increase bacterial proliferation, and increase virulence in animal models of pharyngitis and necrotizing fasciitis. Genome sequencing of an additional 1,125 streptococcal strains and virulence studies revealed that a highly similar recombinational replacement event underlies an ongoing intercontinental epidemic of serotype M89 group A Streptococcus infections. By identifying the molecular changes that enhance upper respiratory tract fitness, increased resistance to innate immunity, and increased tissue destruction, we describe a mechanism that underpins epidemic streptococcal infections, which have affected many millions of people.

Evolutionary pathway to increased virulence and epidemic group A Streptococcus disease derived from 3,615 genome sequences.

We sequenced the genomes of 3,615 strains of serotype Emm protein 1 (M1) group A Streptococcus to unravel the nature and timing of molecular events contributing to the emergence, dissemination, and genetic diversification of an unusually virulent clone that now causes epidemic human infections worldwide. We discovered that the contemporary epidemic clone emerged in stepwise fashion from a precursor cell that first contained the phage encoding an extracellular DNase virulence factor (streptococcal DNase D2, SdaD2) and subsequently acquired the phage encoding the SpeA1 variant of the streptococcal pyrogenic exotoxin A superantigen. The SpeA2 toxin variant evolved from SpeA1 by a single-nucleotide change in the M1 progenitor strain before acquisition by horizontal gene transfer of a large chromosomal region encoding secreted toxins NAD(+)-glycohydrolase and streptolysin O. Acquisition of this 36-kb region in the early 1980s into just one cell containing the phage-encoded sdaD2 and speA2 genes was the final major molecular event preceding the emergence and rapid intercontinental spread of the contemporary epidemic clone. Thus, we resolve a decades-old controversy about the type and sequence of genomic alterations that produced this explosive epidemic. Analysis of comprehensive, population-based contemporary invasive strains from seven countries identified strong patterns of temporal population structure. Compared with a preepidemic reference strain, the contemporary clone is significantly more virulent in nonhuman primate models of pharyngitis and necrotizing fasciitis. A key finding is that the molecular evolutionary events transpiring in just one bacterial cell ultimately have produced millions of human infections worldwide.

Bacterial discrimination by dictyostelid amoebae reveals the complexity of ancient interspecies interactions.

BACKGROUND: Amoebae and bacteria interact within predator-prey and host-pathogen relationships, but the general response of amoeba to bacteria is not well understood. The amoeba Dictyostelium discoideum feeds on, and is colonized by, diverse bacterial species, including Gram-positive [Gram(+)] and Gram-negative [Gram(-)] bacteria, two major groups of bacteria that differ in structure and macromolecular composition. RESULTS: Transcriptional profiling of D. discoideum revealed sets of genes whose expression is enriched in amoebae interacting with different species of bacteria, including sets that appear specific to amoebae interacting with Gram(+) or with Gram(-) bacteria. In a genetic screen utilizing the growth of mutant amoebae on a variety of bacteria as a phenotypic readout, we identified amoebal genes that are only required for growth on Gram(+) bacteria, including one that encodes the cell-surface protein gp130, as well as several genes that are only required for growth on Gram(-) bacteria, including one that encodes a putative lysozyme, AlyL. These genes are required for parts of the transcriptional response of wild-type amoebae, and this allowed their classification into potential response pathways. CONCLUSIONS: We have defined genes that are critical for amoebal survival during feeding on Gram(+), or Gram(-), bacteria that we propose form part of a regulatory network that allows D. discoideum to elicit specific cellular responses to different species of bacteria in order to optimize survival.

Diagnostic protein discovery using liquid chromatography/mass spectrometry for proteolytic peptide targeting.

A peptide targeting method has been developed for diagnostic protein discovery, which combines proteolytic digestion of fractionated plasma proteins and liquid chromatography coupled to electrospray time-of-flight mass spectrometry (LC/ESI-TOFMS) profiling. Proteolysis prior to profiling overcomes molecular weight limitations and compensates for the poor sensitivity of matrix-assisted laser desorption/ionization (MALDI) protein profiling. LC/MS increases the peak capacity compared to crude fractionation techniques or single sample MALDI analysis. Differentially expressed peptides are targeted in the mass chromatograms using bioinformatic techniques and subsequently sequenced with MALDI tandem MS. In a model study comparing pancreatic cancer patients to controls, 74% of the peptide targets were successfully sequenced. This profiling method was superior to previous experiments using single sample MALDI analysis for protein profiling or proteolytic peptide profiling, because more potential protein markers were identified.