When present in coding regions, tandem repeats (TRs) may have large effects on protein structure and function contributing to health and disease. We use a family-based design to identify de novo TRs and assess their impact at the population level in 148,607 European ancestry participants from the UK Biobank. The 427 loci with de novo TR mutations are enriched for targets of microRNA-184 (21.1-fold, P = 4.30 × 10-5, FDR = 9.50 × 10-3). There are 123 TR-phenotype associations with posterior probabilities > 0.95. These relate to body structure, cognition, and cardiovascular, metabolic, psychiatric, and respiratory outcomes. We report several loci with large likely causal effects on tissue microstructure, including the FAN1-[TG]N and carotid intima-media thickness (mean thickness: beta = 5.22, P = 1.22 × 10-6, FDR = 0.004; maximum thickness: beta = 6.44, P = 1.12 × 10-6, FDR = 0.004). Two exonic repeats FNBP4-[GGT]N and BTN2A1-[CCT]N alter protein structure. In this work, we contribute clear and testable hypotheses of dose-dependent TR implications linking genetic variation and protein structure with health and disease outcomes.

BACKGROUND: Currently, FLT3 internal tandem duplication (ITD) is tested by fragment analysis. With next-generation sequencing (NGS), however, not only FLT3 ITD but also other mutations can be detected, which can provide more genetic information on disease.
METHODS: We retrospectively reviewed the results of two tests-fragment analysis and a custom-designed, hybridization capture-based, targeted NGS panel-performed simultaneously. We used the Pindel algorithm to detect FLT3 ITD mutations.
RESULTS: Among 277 bone marrow aspirate samples tested by NGS and fragment analysis, the results revealed 99.6% concordance in FLT3 ITD detection. Overall, the allele frequency (AF) attained by NGS positively correlated with the standard allelic ratio (AR) attained by fragment analysis, with a Spearman correlation coefficient (r) of 0.757 (95% confidence interval: 0.627-0.846; p < 0.001). It was concluded that an AF of 0.11 attained by NGS is the most appropriate cutoff value (with 85.3% sensitivity and 86.7% specificity) for high mutation burden criterion presented by guidelines.
CONCLUSIONS: Sensitive FLT3 ITD detection with comprehensive information of other mutation offered by NGS could be a useful tool in clinical laboratories. Future studies will be needed to evaluate and standardize NGS AF cutoff to predict actual clinical outcomes.

UNASSIGNED: Certain genetic mutations could have a role in the etiology of acute myeloid leukemia (AML). Hereby, in this study, we primarily aimed to investigate the distribution of genetic mutations in AML patients. We also attempted to analyze the incidence of genetic mutations in AML patients from Turkey.This retrospective study included a total of 126 patients diagnosed with AML, who had molecular mutation test results or records in their patient files. The patients who were not citizens of the Republic of Turkey were not included in the study.It was observed that analyses for at least 1 c-kit exon mutation had been carried out on 76 patients, which detected no c-kit mutation among the types of genetic mutations investigated in all of those 76 patients. We found the frequency of FMS-like tyrosine kinase 3-internal tandem duplication mutation as 25%. The prevalence of translocation(15;17) was approximately 11% and the prevalence of translocation(8;21) was % 6.25. In addition, we also showed that the frequency of inversion16 was nearly 3.7%.Lastly, the possibility of c-kit mutation in AML patients from Turkey might actually be low.

In acute myeloid leukemia (AML) internal tandem duplications of the FLT3 gene (FLT3-ITD) are associated with poor prognosis. Retrospectively, we investigated the prognostic and predictive impact of FLT3-ITD insertion site (IS) in 452 patients randomized within the RATIFY trial, which evaluated midostaurin additionally to intensive chemotherapy. Next-generation sequencing identified 908 ITDs, with 643 IS in the juxtamembrane domain (JMD) and 265 IS in the tyrosine kinase domain-1 (TKD1). According to IS, patients were categorized as JMDsole (n = 251, 55%), JMD and TKD1 (JMD/TKD1; n = 117, 26%), and TKD1sole (n = 84, 19%). While clinical variables did not differ among the 3 groups, NPM1 mutation was correlated with JMDsole (P = 0.028). Overall survival (OS) differed significantly, with estimated 4-year OS probabilities of 0.44, 0.50, and 0.30 for JMDsole, JMD/TKD1, and TKD1sole, respectively (P = 0.032). Multivariate (cause-specific) Cox models for OS and cumulative incidence of relapse using allogeneic hematopoietic cell transplantation (HCT) in first complete remission as a time-dependent variable identified TKD1sole as unfavorable and HCT as favorable factors. In addition, Midostaurin exerted a significant benefit only for JMDsole. Our results confirm the distinct molecular heterogeneity of FLT3-ITD and the negative prognostic impact of TKD1 IS in AML that was not overcome by midostaurin.

Galectin-9 is a β-galactoside-binding lectin which could modulate a variety of biological functions including recognition, aggregation and clearance of pathogen. In this study, one Galectin-9 (named PoGalectin-9) was identified from Japanese flounder Paralichthys olivaceus. PoGalectin-9 belongs to the tandem-repeat type, containing one 127-amino acids CRD domain within N terminal and one 122-amino acids CRD domain within C-terminal. The open reading frame of PoGalectin-9 cDNA was 921 bp encoding 306 amino acids. Sequence similarity comparison confirmed that PoGalectin-9 shared high homology with other Galectin-9. The tissue distribution and expression profiles after bacterial infection were also investigated. PoGalectin-9 was widely distributed in all of the examined tissues of Japanese flounder but was predominantly expressed in the spleen, kidney and intestine. After Edwardsiella tarda challenge, the expression of PoGalectin-9 was up-regulated in spleen and down regulated in kidney. ELISA experiment showed that recombinant PoGalectin-9 (rPoGalectin-9) exhibit binding capacity to lipopolysaccharide (LPS) and peptidoglycan (PGN), which is significantly correlated with the concentration of rPoGalectin-9. Meanwhile, the rPoGalectin-9 protein showed strong agglutinating activities against both Gram-negative bacteria and Gram-positive bacteria. Bacterial binding experiments showed that rPoGalectin-9 could bind all examined bacteria. In conclusion, the present study indicate that PoGalectin-9 might play important roles during the immune responses of Japanese flounder against bacterial pathogens.

Findings of retrospective studies suggest that sorafenib maintenance post-transplantation might reduce relapse in patients with FLT3 internal tandem duplication (FLT3-ITD) acute myeloid leukaemia undergoing allogeneic haematopoietic stem-cell transplantation. We investigated the efficacy and tolerability of sorafenib maintenance post-transplantation in this population.
We did an open-label, randomised phase 3 trial at seven hospitals in China. Eligible patients (aged 18-60 years) had FLT3-ITD acute myeloid leukaemia, were undergoing allogeneic haematopoietic stem-cell transplantation, had an Eastern Cooperative Oncology Group performance status of 0-2, had composite complete remission before and after transplantation, and had haematopoietic recovery within 60 days post-transplantation. Patients were randomly assigned (1:1) to sorafenib maintenance (400 mg orally twice daily) or non-maintenance (control) at 30-60 days post-transplantation. Randomisation was done with permuted blocks (block size four) and implemented through an interactive web-based randomisation system. The primary endpoint was the 1-year cumulative incidence of relapse in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, NCT02474290; the trial is complete.
Between June 20, 2015, and July 21, 2018, 202 patients were enrolled and randomly assigned to sorafenib maintenance (n=100) or control (n=102). Median follow-up post-transplantation was 21·3 months (IQR 15·0-37·0). The 1-year cumulative incidence of relapse was 7·0% (95% CI 3·1-13·1) in the sorafenib group and 24·5% (16·6-33·2) in the control group (hazard ratio 0·25, 95% CI 0·11-0·57; p=0·0010). Within 210 days post-transplantation, the most common grade 3 and 4 adverse events were infections (25 [25%] of 100 patients in the sorafenib group vs 24 [24%] of 102 in the control group), acute graft-versus-host-disease (GVHD; 23 [23%] of 100 vs 21 [21%] of 102), chronic GVHD (18 [18%] of 99 vs 17 [17%] of 99), and haematological toxicity (15 [15%] of 100 vs seven [7%] of 102). There were no treatment-related deaths.
Sorafenib maintenance post-transplantation can reduce relapse and is well tolerated in patients with FLT3-ITD acute myeloid leukaemia undergoing allogeneic haematopoietic stem-cell transplantation. This strategy could be a suitable therapeutic option for patients with FLT3-ITD acute myeloid leukaemia.
None.

Cholera remains a major global public health threat and continuous emergence of new Vibrio cholerae strains is of major concern. We conducted a molecular epidemiological study to detect virulence markers and antimicrobial resistance patterns of V. cholerae isolates obtained from the 2012-2015 cholera outbreaks in Ghana. Archived clinical isolates obtained from the 2012, 2014 and 2015 cholera outbreaks in Ghana were revived by culture and subjected to microscopy, biochemical identification, serotyping, antibiotic susceptibility testing, molecular detection of distinct virulence factors and Multi-Locus Variable-Number of Tandem-Repeat Analysis (MLVA). Of 277 isolates analysed, 168 (60.6%) were confirmed to be V. cholerae and 109 (39.4%) isolates constituted other bacteria (Escherichia coli, Aeromonas sobria, Pseudomonas aeruginosa, Enterobacter cloacae and Enterococci faecalis). Serotyping the V. cholerae isolates identified 151 (89.9%) as Ogawa, 3 (1.8%) as Inaba and 14 (8.3%) as non-O1/O139 serogroup. The O1 serogroup isolates (154/168, 91.7%) carried the cholera toxin ctxB gene as detected by PCR. Additional virulence genes detected include zot, tcpA, ace, rtxC, toxR, rtxA, tcpP, hlyA and tagA. The most common and rare virulence factors detected among the isolates were rtxC (165 isolates) and tcpP (50 isolates) respectively. All isolates from 2014 and 2015 were multidrug resistant against the selected antibiotics. MLVA differentiated the isolates into 2 large unique clones A and B, with each predominating in a particular year. Spatial analysis showed clustering of most isolates at Ablekuma sub-district. Identification of several virulence genes among the two different genotypes of V. cholerae isolates and resistance to first- and second-line antibiotics, calls for scaleup of preventive strategies to reduce transmission, and strengthening of public health laboratories for rapid antimicrobial susceptibility testing to guide accurate treatment. Our findings support the current WHO licensed cholera vaccines which include both O1 Inaba and Ogawa serotypes.

Mycobacterium tuberculosis complex (MTBC) is divided into several major phylogenetic lineages, with differential distribution globally. Using population-based data collected over a three year period, we performed 24-locus Mycobacterial Interspersed Repeat Unit - Variable Number Tandem Repeat (MIRU-VNTR) genotyping on all culture isolates from two districts of the country that differ in tuberculosis (TB) incidence (Gaborone, the capital, and Ghanzi in the Western Kalahari). The study objective was to characterize the molecular epidemiology of TB in these districts. Overall phylogenetic diversity mirrored that reported from neighboring Republic of South Africa, but differences in the two districts were marked. All four major lineages of M. tuberculosis were found in Gaborone, but only three of the four major lineages were found in Ghanzi. Strain diversity was lower in Ghanzi, with a large proportion (38%) of all isolates having an identical MIRU-VNTR result, compared to 6% of all isolates in Gaborone with the same MIRU-VNTR result. This study demonstrates localized differences in strain diversity by two districts in Botswana, and contributes to a growing characterization of MTBC diversity globally.

Friedreich ataxia is a progressive neurodegenerative disorder with reported abnormalities in cerebellar, brainstem, and cerebral white matter. White matter structure can be measured using in vivo neuroimaging indices sensitive to different white matter features. For the first time, we examined the relative sensitivity and relationship between multiple white matter indices in Friedreich ataxia to more richly characterize disease expression and infer possible mechanisms underlying the observed white matter abnormalities. Diffusion-tensor, magnetization transfer, and T1-weighted structural images were acquired from 31 individuals with Friedreich ataxia and 36 controls. Six white matter indices were extracted: fractional anisotropy, diffusivity (mean, axial, radial), magnetization transfer ratio (microstructure), and volume (macrostructure). For each index, whole-brain voxel-wise between-group comparisons and correlations with disease severity, onset age, and gene triplet-repeat length were undertaken. Correlations between pairs of indices were assessed in the Friedreich ataxia cohort. Spatial similarities in the voxel-level pattern of between-group differences across the indices were also assessed. Microstructural abnormalities were maximal in cerebellar and brainstem regions, but evident throughout the brain, while macroscopic abnormalities were restricted to the brainstem. Poorer microstructure and reduced macrostructural volume correlated with greater disease severity and earlier onset, particularly in peri-dentate nuclei and brainstem regions. Microstructural and macrostructural abnormalities were largely independent. Reduced fractional anisotropy was most strongly associated with axial diffusivity in cerebral tracts, and magnetization transfer in cerebellar tracts. Multiple mechanisms likely underpin white matter abnormalities in Friedreich ataxia, with differential impacts in cerebellar and cerebral pathways.

FLT3-internal tandem duplication occurs in 20-30% of acute myeloid leukemia and confers an adverse prognosis with its allelic ratio being a key risk stratifier. The US Food and Drug Administration recently approved FLT3 inhibitors midostaurin and gilteritinib in FLT3 mutation-positive acute myeloid leukemia. Historically, FLT3 was tested by fragment analysis, which has become the standard method endorsed by international guidelines. However, next generation sequencing is increasingly used at acute myeloid leukemia diagnosis given its ability to simultaneously evaluate multiple clinically informative markers. As FLT3-internal tandem duplication detection was known to be challenging by next generation sequencing and the results carry profound prognostic and therapeutic implications, it is important to thoroughly examine its performance in FLT3-internal tandem duplication detection and allelic ratio classification. In a comparative study with fragment analysis, we retrospectively reviewed our experience using a custom-designed, hybridization capture-based, targeted next generation sequencing panel. Among 7902 cases, FLT3-internal tandem duplication was detected in 335 with variable sizes (3-231 bp) and insertion sites. Fragment analysis was also performed in 402 cases, demonstrating 100% concordance in FLT3-internal tandem duplication detection. In 136 dual-tested, positive cases, 128/136 (94%) exhibited concordant high/low allelic ratio classifications. The remaining 6% showed borderline low allelic ratio by next generation sequencing. The two methods were concordant in FLT3-tyrosine kinase domain mutation detection at the hotspot D835/I836 targeted by fragment analysis. Furthermore, seven mutations which may benefit from FLT3 inhibitor therapy were detected by next generation sequencing, in regions not covered by fragment analysis. Our study demonstrates that using a hybridization capture-based chemistry and optimized bioinformatics pipeline, next generation sequencing can reliably detect FLT3-internal tandem duplication and classify its allelic ratio for acute myeloid leukemia risk stratification. Next generation sequencing also exhibits superior comprehensiveness in FLT3 mutation detection and may further improve personalized, targeted therapy in acute myeloid leukemia.