PathogenMIPer: a tool for the design of molecular inversion probes to detect multiple pathogens

BACKGROUND: Here we describe PathogenMIPer, a software program for designing molecular inversion probe (MIP) oligonucleotides for use in pathogen identification and detection. The software designs unique and specific oligonucleotide probes targeting microbial or other genomes. The tool tailors all probe sequence components (including target-specific sequences, barcode sequences, universal primers and restriction sites) and combines these components into ready-to-order probes for use in a MIP assay. The system can harness the genetic variability available in an entire genome in designing specific probes for the detection of multiple co-infections in a single tube using a MIP assay. RESULTS: PathogenMIPer can accept sequence data in FASTA file format, and other parameter inputs from the user through a graphical user interface. It can design MIPs not only for pathogens, but for any genome for use in parallel genomic analyses. The software was validated experimentally by applying it to the detection of human papilloma virus (HPV) as a model system, which is associated with various human malignancies including cervical and skin cancers. Initial tests of laboratory samples using the MIPs developed by the PathogenMIPer to recognize 24 different types of HPVs gave very promising results, detecting even a small viral load of single as well as multiple infections (Akhras et al, personal communication). CONCLUSION: PathogenMIPer is a software for designing molecular inversion probes for detection of multiple target DNAs in a sample using MIP assays. It enables broader use of MIP technology in the detection through genotyping of pathogens that are complex, difficult-to-amplify, or present in multiple subtypes in a sample

Soleus Fiber Force and Maximal Shortening Velocity After Non-Weight Bearing with Intermittent Activity

This study examined the effectiveness of intermittent weight bearing (IWB) as a countermeasure to non-weight-bearing (NWB)-induced alterations in soleus type 1 fiber force (in mN), tension (P(sub o); force per fiber cross-sectional area in kN/sq m), and maximal unloaded shortening velocity (V(sub o), in fiber lengths/s). Adult rats were assigned to one of the following groups: normal weight bearing (WB), 14 days of hindlimb NWB (NWB group), and 14 days of hindlimb NWB with IWB treatments (IWB group). The IWB treatment consisted of four 10-min periods of standing WB each day. Single, chemically permeabilized soleus fiber segments were mounted between a force transducer and position motor and were studied at maximal Ca(2+) activation, after which type 1 fiber myosin heavy-chain composition was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. NWB resulted in a loss in relative soleus mass (-45%), with type 1 fibers displaying reductions in diameter (-28%) and peak isometric force (-55%) and an increase in V(sub o) (+33%). In addition, NWB induced a 16% reduction in type 1 fiber P., a 41% reduction in type 1 fiber peak elastic modulus [E(sub o), defined as ((delta)force/(delta)length x (fiber length/fiber cross-sectional area] and a significant increase in the P(sub o)/E(sub o) ratio. In contrast to NWB, IWB reduced the loss of relative soleus mass (by 22%) and attenuated alterations in type 1 fiber diameter (by 36%), peak force (by 29%), and V(sub o)(by 48%) but had no significant effect on P(sub o), E(sub o) or P(sub o)/E(sub o). These results indicate that a modest restoration of WB activity during 14 days of NWB is sufficient to attenuate type 1 fiber atrophy and to partially restore type 1 peak isometric force and V(sub o) to WB levels. However, the NWB-induced reductions in P(sub o) and E(sub o) which we hypothesize to be due to a decline in the number and stiffness of cross bridges, respectively, are considerably less responsive to this countermeasure treatment

The Influence of Food Intake Specificity in Children with Autism on Gut Microbiota

Autism spectrum disorder (ASD) is a complex of neurodevelopmental conditions with increasing incidence. The microbiota of children with ASD is distinct from neurotypical children, their food habits are also different, and it is known that nutrient intake influences microbiota in a specific way. Thus, this study investigates the food habits of children with ASD and their association with the gut microbiota. Children with ASD had their dietary energy intakes similar to controls, but they more often demonstrated food selectivity, which seemed to result in deficiency of micronutrients such as vitamins K, B6, C, iron, cooper, docosahexaenoic and docosapentanoic acid. Using high-throughput sequencing, a DNA library of intestinal microbiota was performed. Core microbiota was similar in children with and without ASD, but Dichelobacter, Nitriliruptor and Constrictibacter were found to be putative markers of ASD. The changes in gut microbiota that we observed in connection to food selectivity, intake of fats and omega-3 in particular, fermented milk products and animal/plant protein consumption had similar character, independent of diagnosis. However, high fibre intake was connected with a decreased α-diversity only in children with ASD. High carbohydrate and fibre intake influenced β-diversity, changing the abundance of Bacteroides and other genera, many of them members of the Clostidiaceae. Modulating food habits of ASD children can influence their gut microbiota composition

ATAD3A-related pontocerebellar hypoplasia: new patients and insights into phenotypic variability

Abstract Background Pathogenic variants in the ATAD3A gene lead to a heterogenous clinical picture and severity ranging from recessive neonatal-lethal pontocerebellar hypoplasia through milder dominant Harel-Yoon syndrome up to, again, neonatal-lethal but dominant cardiomyopathy. The genetic diagnostics of ATAD3A-related disorders is also challenging due to three paralogous genes in the ATAD3 locus, making it a difficult target for both sequencing and CNV analyses. Results Here we report four individuals from two families with compound heterozygous p.Leu77Val and exon 3–4 deletion in the ATAD3A gene. One of these patients was characterized as having combined OXPHOS deficiency based on decreased complex IV activities, decreased complex IV, I, and V holoenzyme content, as well as decreased levels of COX2 and ATP5A subunits and decreased rate of mitochondrial proteosynthesis. All four reported patients shared a strikingly similar clinical picture to a previously reported patient with the p.Leu77Val variant in combination with a null allele. They presented with a less severe course of the disease and a longer lifespan than in the case of biallelic loss-of-function variants. This consistency of the phenotype in otherwise clinically heterogenous disorder led us to the hypothesis that the severity of the phenotype could depend on the severity of variant impact. To follow this rationale, we reviewed the published cases and sorted the recessive variants according to their impact predicted by their type and the severity of the disease in the patients. Conclusion The clinical picture and severity of ATAD3A-related disorders are homogenous in patients sharing the same combinations of variants. This knowledge enables deduction of variant impact severity based on known cases and allows more accurate prognosis estimation, as well as a better understanding of the ATAD3A function