Gene expression analysis of children with acute hematogenous osteomyelitis caused by Methicillin-resistant Staphylococcus aureus: correlation with clinical severity of illness.

Children with acute hematogenous osteomyelitis (AHO) demonstrate a broad spectrum of clinical manifestations, ranging from mild to severe. Several advances have been achieved in the study of host immune response to acute invasive Staphylococcus aureus infections through gene expression analysis. However, previous research has neither attempted to evaluate the response of children with AHO specific to Methicillin-resistant Staphylococcus aureus (MRSA) nor to correlate gene expression with clinical phenotype. Study objective was to correlate gene expression of children with AHO due to MRSA with clinical severity of illness. Whole blood samples were obtained in Tempus tubes from 12 children with osteomyelitis once cultures obtained directly from the site of infection confirmed to be positive for MRSA. Using an Illumina platform and a systems-wide modular analysis, microarray findings from ten of these children were compared to that of nine healthy (age, ethnicity and gender) matched controls and correlated with clinical severity of illness. Children with AHO from MRSA demonstrated over-expression of innate immunity with respect to neutrophil activity, coagulation, inflammatory response, and erythrocyte development. Concurrently, these children demonstrated under-expression of adaptive immunity with respect to lymphocyte activation and activity of T-cell, cytotoxic or NK cell, and B-cell lines. Three over-expressed genes, P2RX1, SORT1, and RETN, and two under-expressed genes, LOC641788 and STAT 4, were significantly correlated with severity of illness. STAT 4 showed the strongest correlation (R2 = -0.83). STAT4 downregulation could potentially explain under-expression of genes related to adaptive immunity in this cohort of patients with AHO. This study identified specific genes which correspond to disease severity during the early hospitalization of children with AHO from MRSA. Pattern recognition of this combination of genes could help to identify in the future severe clinical phenotypes before the disease is fully manifest and direct appropriate attention and resources to those children

Genomic Heterogeneity of Methicillin Resistant Staphylococcus aureus Associated with Variation in Severity of Illness among Children with Acute Hematogenous Osteomyelitis.

IntroductionThe association between severity of illness of children with osteomyelitis caused by Methicillin-resistant Staphylococcus aureus (MRSA) and genomic variation of the causative organism has not been previously investigated. The purpose of this study is to assess genomic heterogeneity among MRSA isolates from children with osteomyelitis who have diverse severity of illness.Materials and methodsChildren with osteomyelitis were prospectively studied between 2010 and 2011. Severity of illness of the affected children was determined from clinical and laboratory parameters. MRSA isolates were analyzed with next generation sequencing (NGS) and optical mapping. Sequence data was used for multi-locus sequence typing (MLST), phylogenetic analysis by maximum likelihood (PAML), and identification of virulence genes and single nucleotide polymorphisms (SNP) relative to reference strains.ResultsThe twelve children studied demonstrated severity of illness scores ranging from 0 (mild) to 9 (severe). All isolates were USA300, ST 8, SCC mec IVa MRSA by MLST. The isolates differed from reference strains by 2 insertions (40 Kb each) and 2 deletions (10 and 25 Kb) but had no rearrangements or copy number variations. There was a higher occurrence of virulence genes among study isolates when compared to the reference strains (p = 0.0124). There were an average of 11 nonsynonymous SNPs per strain. PAML demonstrated heterogeneity of study isolates from each other and from the reference strains.DiscussionGenomic heterogeneity exists among MRSA isolates causing osteomyelitis among children in a single community. These variations may play a role in the pathogenesis of variation in clinical severity among these children

Effects of glucosamine and chondroitin sulfate supplementation in addition to resistance exercise training and manual therapy in patients with knee osteoarthritis: a randomized controlled trial

Objective: To determine the added benefits of short-term glucosamine and chondroitin sulfate supplementation in combination with manual therapy and resistance exercise training in the management of knee osteoarthritis. Method: The parallel-design, double-blind randomised controlled trial was conducted from January to September 2020 at the Foundation University Institute of Rehabilitation Sciences and Fauji Foundation Hospital, Rawalpindi, Pakistan, and comprised knee osteoarthritis patients of either gender having radiological evidence of grade III or less on Kellgren classification. The subjects were randomly allocated experimental group A and control group B. Both the groups received manual therapy and resistance exercise training, while group A additionally received glucosamine and chondroitin sulfate supplementation for 4 weeks. Study outcomes included pain, function, quality of life, range of motion, strength, fall risk, skeletal muscle mass, visceral fat area, body fat, intracellular water ratio, and segmental lean and fat mass. Data was analysed using SPSS 21. Results: Of the 24 subjects, there were 12(50%) in each of the two groups. Both groups had 9(75%) men and 3(25%) women. In terms knee osteoarthritis grade, there was no significant difference between the groups (p=1.00). No significant differences were observed in any of the outcome measures neither at the halfway mark, nor post-intervention between the groups (p>0.05) except for percentage change in segmental lean mass of the right leg at the 2nd week and of the left leg at the 4th week (p<0.05). ---Continu

Body composition profiling and obesity analysis of healthy adults: a cross-sectional study

Obesity as depicted by changes in the body composition is considered a global epidemic of the 21st century, predisposing to cardiometabolic diseases. This analytical cross-sectional study evaluated body composition parameters in both genders and conducted an obesity analysis of healthy adult Pakistani population. Using non-probability purposive sampling technique, data was collected from 205 healthy adults (aged 18-45 years), who had not been participating in any structured exercise or dietary regime over the last six months. Body composition was assessed using bioelectrical impedance analyser. For data analysis, descriptive statistics, Mann-Whitney U-test and one-sample T-test were applied. The mean body mass index was 24.3±4.93 kg/m2. Body composition components which were significantly higher in males included soft lean mass, fat-free mass index, skeletal muscle mass index and bone mineral content, whereas females had significantly higher percentage of body fat (PBF), visceral fat area and fat mass index (p<0.05). ---Continu

Module Analysis of Gene Expression Profile in Whole Blood From Patients with MRSA AHO.

<p>Gene expression in one of the subjects within the healthy control group revealed an unusual array of cellular expression. This subject was excluded and microarray statistical analysis was repeated without his information. Gene expression levels were compared between healthy controls and patients with MRSA AHO on a module by module analysis. Each column represents a subject and each row a cell type. Colored spots represent the percentage of significant over-expressed (red) or under expressed (blue) transcripts (Mann-Whitney p<0.05). The intensity of the color refers to the number of transcripts within each of the pre-selected modules not the actual fold change that is up or down regulated compared to healthy controls. Modular analysis identified that patients had upregulation of innate immunity and mild downregulation of adaptive immunity.</p

Top 24 Over-expressed and 34 Under-expressed Genes in MRSA AHO.

<p>Genes associated to SIS in bold.</p

Correlation of Gene Over and Under-Expression and Severity of Illness Score (SIS).

<p>Statistical correlation (Pearson, Spearman, Pearson Log Gene and Pearson Log-Log) of severity of illness score and quantitative gene expression was significant for five genes. <b>A)</b> RETN (resistin gene), SORT 1 (sortilin 1 gene) and P2RX1 (purinergic receptor P2X, ligand-gated ion channel 1) showed over-expression. <b>B)</b> LOC641788 (similar to CDC26 subunit of anaphase promoting complex) and STAT 4 (signal transducer and activator of transcription 4) showed under-expression. Of those genes, RETN and STAT 4 had the strongest correlation coefficients (RETN: R2 = 0.8 and STAT 4: R2 = –0.83).</p

Gene Expression Biosignature in Whole Blood From Healthy Controls and Patients With MRSA AHO.

<p>Complete statistical analysis (Welch t-Test with Benjamini - Hochberg false discovery rate, p<0.05) between the two groups (healthy control group with 9 subjects and MRSA AHO group with 10 subjects) identified 58 genes (24 over-expressed and 34 under-expressed) which significantly differentiated children with MRSA AHO from the healthy controls.</p