Characterization of the Urinary Metagenome and Virome in Healthy Children.

Recent advances in next-generation sequencing and metagenomic studies have provided insights into the microbial profile of different body sites. However, research on the microbial composition of urine is limited, particularly in children. The goal of this study was to optimize and develop reproducible metagenome and virome protocols using a small volume of urine samples collected from healthy children. We collected midstream urine specimens from 40 healthy children. Using the metagenomics shotgun approach, we tested various protocols. Different microbial roots such as Archaea, Bacteria, Eukaryota, and Viruses were successfully identified using our optimized urine protocol. Our data reflected much variation in the microbial fingerprints of children. Girls had significantly higher levels of Firmicutes, whereas boys had significantly higher levels of Actinobacteria. The genus Anaerococcus dominated the urinary bacteriome of healthy girls, with a significant increase in Anaerococcus prevotii, Anaerococcus vaginalis, and Veillonella parvula (p-value < 0.001) when compared with that of boys. An increased relative abundance of Xylanimonas and Arthrobacter, with a significantly high abundance of Arthrobacter sp. FB24 (p-value 0.0028) and Arthrobacter aurescences (p-value 0.015), was observed in boys. The urinary mycobiome showed a significant rise in the genus Malassezia and Malassezia globose fungus (p-value 0.009) in girls, whereas genus Saccharomyces (p-value 0.009) was significantly high in boys. The beta diversity of the urinary mycobiome was found to differ between different age groups. Boys had significantly more Mastadenovirus and Human mastadenovirus-A in their urinary virome than girls. With increasing age, we noticed an increase in the relative abundance of the order Caudovirales. Our optimized protocols allowed us to identify the unique microbes for each sex by using an adequate volume of urine (3−10 mL) to screen for the bacteriome, mycobiome, and virome profiles in the urine of healthy children. To the best of our knowledge, this is the first study to characterize the metagenomics profiles of urine in a healthy pediatric population.

iPSCs derived from insulin resistant offspring of type 2 diabetic patients show increased oxidative stress and lactate secretion.

BACKGROUND: The genetic factors associated with insulin resistance (IR) are not well understood. Clinical studies on first-degree relatives of type 2 diabetic (T2D) patients, which have the highest genetic predisposition to T2D, have given insights into the role of IR in T2D pathogenesis. Induced pluripotent stem cells (iPSCs) are excellent tools for disease modeling as they can retain the genetic imprint of the disease. Therefore, in this study, we aimed to investigate the genetic perturbations associated with insulin resistance (IR) in the offspring of T2D parents using patient-specific iPSCs. METHODS: We generated iPSCs from IR individuals (IR-iPSCs) that were offspring of T2D parents as well as from insulin-sensitive (IS-iPSCs) individuals. We then performed transcriptomics to identify key dysregulated gene networks in the IR-iPSCs in comparison to IS-iPSCs and functionally validated them. RESULTS: Transcriptomics on IR-iPSCs revealed dysregulated gene networks and biological processes indicating that they carry the genetic defects associated with IR that may lead to T2D. The IR-iPSCs had increased lactate secretion and a higher phosphorylation of AKT upon stimulation with insulin. IR-iPSCs have increased cellular oxidative stress indicated by a high production of reactive oxygen species and higher susceptibility to H2O2 -induced apoptosis. CONCLUSIONS: IR-iPSCs generated from offspring of diabetic patients confirm that oxidative stress and increased lactate secretion, associated with IR, are inherited in this population, and may place them at a high risk of T2D. Overall, our IR-iPSC model can be employed for T2D modeling and drug screening studies that target genetic perturbations associated with IR in individuals with a high risk for T2D.

The Human Microbiome in Chronic Kidney Disease: A Double-Edged Sword.

Chronic kidney disease (CKD) is an increasing global health burden. Current treatments for CKD include therapeutics to target factors that contribute to CKD progression, including renin-angiotensin-aldosterone system inhibitors, and drugs to control blood pressure and proteinuria control. Recently, associations between chronic disease processes and the human microbiota and its metabolites have been demonstrated. Dysbiosis-a change in the microbial diversity-has been observed in patients with CKD. The relationship between CKD and dysbiosis is bidirectional; gut-derived metabolites and toxins affect the progression of CKD, and the uremic milieu affects the microbiota. The accumulation of microbial metabolites and toxins is linked to the loss of kidney functions and increased mortality risk, yet renoprotective metabolites such as short-chain fatty acids and bile acids help restore kidney functions and increase the survival rate in CKD patients. Specific dietary interventions to alter the gut microbiome could improve clinical outcomes in patients with CKD. Low-protein and high-fiber diets increase the abundance of bacteria that produce short-chain fatty acids and anti-inflammatory bacteria. Fluctuations in the urinary microbiome are linked to increased susceptibility to infection and antibiotic resistance. In this review, we describe the potential role of the gut, urinary and blood microbiome in CKD pathophysiology and assess the feasibility of modulating the gut microbiota as a therapeutic tool for treating CKD.

Molecular characterization of extended spectrum ß -lactamases enterobacteriaceae causing lower urinary tract infection among pediatric population.

Background: The ß-lactam antibiotics have traditionally been the main treatment of Enterobacteriaceae infections, nonetheless, the emergence of species producing ß- Lactamases has rendered this class of antibiotics largely ineffective. There are no published data on etiology of urinary tract infections (UTI) and antimicrobial resistance profile of uropathogens among children in Qatar. The aim of this study is to determine the phenotypic and genotypic profiles of antimicrobial resistant Enterobacteriaceae among children with UTI in Qatar. Methods: Bacteria were isolated from 727 urine positive cultures, collected from children with UTI between February and June 2017 at the Pediatric Emergency Center, Doha, Qatar. Isolated bacteria were tested for antibiotic susceptibility against sixteen clinically relevant antibiotics using phoenix and Double Disc Synergy Test (DDST) for confirmation of extended-spectrum beta-lactamase (ESBL) production. Existence of genes encoding ESBL production were identified using polymerase chain reaction (PCR). Statistical analysis was done using non-parametric Kappa statistics, Pearson chi-square test and Jacquard's coefficient. Results: 201 (31.7%) of samples were confirmed as Extended Spectrum ß -Lactamases (ESBL) Producing Enterobacteriaceae. The most dominant pathogen was E. coli 166 (83%) followed by K. pneumoniae 22 (11%). Resistance was mostly encoded by bla CTX-M (59%) genes, primarily bla CTX-MG1 (89.2%) followed by bla CTX-MG9 (7.7%). 37% of isolated bacteria were harboring multiple bla genes (2 genes or more). E. coli isolates were categorized into 11 clusters, while K. pneoumoniae were grouped into five clonal clusters according to the presence and absence of seven genes namely bla TEM, bla SHV, bla CTX-MG1, bla CTX-MG2, bla CTX-MG8 bla CTX-MG9,bla CTX-MG25. Conclusions: Our data indicates an escalated problem of ESBL in pediatrics with UTI, which mandates implementation of regulatory programs to reduce the spread of ESBL producing Enterobacteriaceae in the community. The use of cephalosporins, aminoglycosides (gentamicin) and trimethoprim/sulfamethoxazole is compromised in Qatar among pediatric population with UTI, leaving carbapenems and amikacin as the therapeutic option for severe infections caused by ESBL producers.

Antibiotic Resistance Profile of Commensal Escherichia coli Isolated from Broiler Chickens in Qatar.

Antibiotic resistance (AR) is a growing public health concern worldwide, and it is a top health challenge in the 21st century. AR among Enterobacteriaceae is rapidly increasing, especially in third-generation cephalosporins and carbapenems. Further, strains carrying mobilized colistin resistance ( mcr) genes 1 and 2 have been isolated from humans, food-producing animals, and the environment. The uncontrolled use of antibiotics in food-producing animals is a major factor in the generation and spread of AR. No studies have been done to evaluate AR in the veterinary sector of Qatar. This study aimed at establishing primary baseline data for the prevalence of AR among food-producing animals in Qatar. Fecal samples (172) were obtained from two broiler farms and one live bird market in Qatar, and 90 commensal Escherichia coli bacteria were isolated and subjected to susceptibility testing against 16 clinically relevant antibiotics by using the E-test method. The results found that 81 (90%) of 90 isolates were resistant to at least one antibiotic, 14 (15.5%) of 90 isolates were colistin resistant, 2 (2.2%) of 90 isolates were extended-spectrum ß-lactamase producers, and 2 (2.2%) of 90 isolates were multidrug resistant to four antibiotic classes. Extended-spectrum ß-lactamase-producing E. coli and colistin-resistant isolates were confirmed by using double-disc susceptibility testing and PCR, respectively. Such a high prevalence of antibiotic-resistant E. coli could be the result of a long application of antibiotic treatment, and it is an indicator of the antibiotic load in food-producing animals in Qatar. Pathogens carrying AR can be easily transmitted to humans through consumption of undercooked food or noncompliance with hygiene practices, mandating prompt development and implementation of a stewardship program to control and monitor the use of antibiotics in the community and agriculture.