Complex Reorganization and Predominant Non-Homologous Repair Following Chromosomal Breakage in Karyotypically Balanced Germline Rearrangements and Transgenic Integration

We defined the genetic landscape of balanced chromosomal rearrangements at nucleotide resolution by sequencing 141 breakpoints from cytogenetically-interpreted translocations and inversions. We confirm that the recently described phenomenon of “chromothripsis” (massive chromosomal shattering and reorganization) is not unique to cancer cells but also occurs in the germline where it can resolve to a karyotypically balanced state with frequent inversions. We detected a high incidence of complex rearrangements (19.2%) and substantially less reliance on microhomology (31%) than previously observed in benign CNVs. We compared these results to experimentally-generated DNA breakage-repair by sequencing seven transgenic animals, and revealed extensive rearrangement of the transgene and host genome with similar complexity to human germline alterations. Inversion is the most common rearrangement, suggesting that a combined mechanism involving template switching and non-homologous repair mediates the formation of balanced complex rearrangements that are viable, stably replicated and transmitted unaltered to subsequent generations

The genomic landscape of balanced cytogenetic abnormalities associated with human congenital anomalies

Despite the clinical significance of balanced chromosomal abnormalities (BCAs), their characterization has largely been restricted to cytogenetic resolution. We explored the landscape of BCAs at nucleotide resolution in 273 subjects with a spectrum of congenital anomalies. Whole-genome sequencing revised 93% of karyotypes and demonstrated complexity that was cryptic to karyotyping in 21% of BCAs, highlighting the limitations of conventional cytogenetic approaches. At least 33.9% of BCAs resulted in gene disruption that likely contributed to the developmental phenotype, 5.2% were associated with pathogenic genomic imbalances, and 7.3% disrupted topologically associated domains (TADs) encompassing known syndromic loci. Remarkably, BCA breakpoints in eight subjects altered a single TAD encompassing MEF2C, a known driver of 5q14.3 microdeletion syndrome, resulting in decreased MEF2C expression. We propose that sequence-level resolution dramatically improves prediction of clinical outcomes for balanced rearrangements and provides insight into new pathogenic mechanisms, such as altered regulation due to changes in chromosome topology

Cryptic behavior and activity cycles of a small mammal keystone species revealed through accelerometry: a case study of Merriam’s kangaroo rats (Dipodomys merriami)

Abstract Background Kangaroo rats are small mammals that are among the most abundant vertebrates in many terrestrial ecosystems in Western North America and are considered both keystone species and ecosystem engineers, providing numerous linkages between other species as both consumers and resources. However, there are challenges to studying the behavior and activity of these species due to the difficulty of observing large numbers of individuals that are small, secretive, and nocturnal. Our goal was to develop an integrated approach of miniaturized animal-borne accelerometry and radiotelemetry to classify the cryptic behavior and activity cycles of kangaroo rats and test hypotheses of how their behavior is influenced by light cycles, moonlight, and weather. Methods We provide a proof-of-concept approach to effectively quantify behavioral patterns of small bodied (< 50 g), nocturnal, and terrestrial free-ranging mammals using large acceleration datasets by combining low-mass, miniaturized animal-borne accelerometers with radiotelemetry and advanced machine learning techniques. We developed a method of attachment and retrieval for deploying accelerometers, a non-disruptive method of gathering observational validation datasets for acceleration data on free-ranging nocturnal small mammals, and used these techniques on Merriam’s kangaroo rats to analyze how behavioral patterns relate to abiotic factors. Results We found that Merriam’s kangaroo rats are only active during the nighttime phases of the diel cycle and are particularly active during later light phases of the night (i.e., late night, morning twilight, and dawn). We found no reduction in activity or foraging associated with moonlight, indicating that kangaroo rats are actually more lunarphilic than lunarphobic. We also found that kangaroo rats increased foraging effort on more humid nights, most likely as a mechanism to avoid cutaneous water loss. Conclusions Small mammals are often integral to ecosystem functionality, as many of these species are highly abundant ecosystem engineers driving linkages in energy flow and nutrient transfer across trophic levels. Our work represents the first continuous detailed quantitative description of fine-scale behavioral activity budgets in kangaroo rats, and lays out a general framework for how to use miniaturized biologging devices on small and nocturnal mammals to examine behavioral responses to environmental factors

How to study a predator that only eats a few meals a year: high-frequency accelerometry to quantify feeding behaviours of rattlesnakes (Crotalus spp.)

Abstract Background Many snakes are low-energy predators that use crypsis to ambush their prey. Most of these species feed very infrequently, are sensitive to the presence of larger vertebrates, such as humans, and spend large portions of their lifetime hidden. This makes direct observation of feeding behaviour challenging, and previous methodologies developed for documenting predation behaviours of free-ranging snakes have critical limitations. Animal-borne accelerometers have been increasingly used by ecologists to quantify activity and moment-to-moment behaviour of free ranging animals, but their application in snakes has been limited to documenting basic behavioural states (e.g., active vs. non-active). High-frequency accelerometry can provide new insight into the behaviour of this important group of predators, and here we propose a new method to quantify key aspects of the feeding behaviour of three species of viperid snakes (Crotalus spp.) and assess the transferability of classification models across those species. Results We used open-source software to create species-specific models that classified locomotion, stillness, predatory striking, and prey swallowing with high precision, accuracy, and recall. In addition, we identified a low cost, reliable, non-invasive attachment method for accelerometry devices to be placed anteriorly on snakes, as is likely necessary for accurately classifying distinct behaviours in these species. However, species-specific models had low transferability in our cross-species comparison. Conclusions Overall, our study demonstrates the strong potential for using accelerometry to document critical feeding behaviours in snakes that are difficult to observe directly. Furthermore, we provide an ‘end-to-end’ template for identifying important behaviours involved in the foraging ecology of viperids using high-frequency accelerometry. We highlight a method of attachment of accelerometers, a technique to simulate feeding events in captivity, and a model selection procedure using biologically relevant window sizes in an open-access software for analyzing acceleration data (AcceleRater). Although we were unable to obtain a generalized model across species, if more data are incorporated from snakes across different body sizes and different contexts (i.e., moving through natural habitat), general models could potentially be developed that have higher transferability

Estrogen-related receptor gamma implicated in a phenotype including hearing loss and mild developmental delay

Analysis of chromosomal rearrangements has been highly successful in identifying genes involved in many congenital abnormalities including hearing loss. Herein, we report a subject, designated DGAP242, with congenital hearing loss (HL) and a de novo balanced translocation 46,XX,t(1;5)(q32;q15)dn. Using multiple next-generation sequencing techniques, we obtained high resolution of the breakpoints. This revealed disruption of the orphan receptor ESRRG on chromosome 1, which is differentially expressed in inner ear hair cells and has previously been implicated in HL, and disruption of KIAA0825 on chromosome 5. Given the translocation breakpoints and supporting literature, disruption of ESRRG is the most likely cause for DGAP242's phenotype and implicates ESRRG in a monogenic form of congenital HL, although a putative contributory role for KIAA0825 in the subject's disorder cannot be excluded

HIV RNA Screening Reduces Integrase Strand Transfer Inhibitor Resistance Risk in Persons Receiving Long-Acting Cabotegravir for HIV Prevention.

BackgroundThe HPTN 083 trial demonstrated that long-acting cabotegravir (CAB-LA) was superior to tenofovir-disoproxil fumarate/emtricitabine for human immunodeficiency virus (HIV) preexposure prophylaxis (PrEP). Integrase strand transfer inhibitor (INSTI) resistance-associated mutations (RAMs) were detected in some participants with HIV infection. We used a low viral load INSTI genotyping assay to evaluate the timing of emergence of INSTI RAMs and assessed whether HIV screening with a sensitive RNA assay would have detected HIV infection before INSTI resistance emerged.MethodsSingle-genome sequencing to detect INSTI RAMs was performed for samples with viral loads &lt;500 copies/mL from 5 participants with previously identified INSTI RAMs and 2 with no prior genotyping results.ResultsMajor INSTI RAMs were detected in all 7 cases. HIV RNA testing identified infection before major INSTI RAMs emerged in 4 cases and before additional major INSTI RAMs accumulated in 1 case. Most INSTI RAMs were detected early when the viral load was low and CAB concentration was high.ConclusionsWhen using CAB-LA PrEP, earlier detection of HIV infection with a sensitive RNA assay may allow for earlier treatment initiation with the potential to reduce INSTI resistance risk. Further studies are needed to evaluate the value and feasibility of HIV RNA testing with CAB-LA PrEP

Defining the diverse spectrum of inversions, complex structural variation, and chromothripsis in the morbid human genome

Background: Structural variation (SV) influences genome organization and contributes to human disease. However, the complete mutational spectrum of SV has not been routinely captured in disease association studies. Results: We sequenced 689 participants with autism spectrum disorder (ASD) and other developmental abnormalities to construct a genome-wide map of large SV. Using long-insert jumping libraries at 105X mean physical coverage and linked-read whole-genome sequencing from 10X Genomics, we document seven major SV classes at ~5 kb SV resolution. Our results encompass 11,735 distinct large SV sites, 38.1% of which are novel and 16.8% of which are balanced or complex. We characterize 16 recurrent subclasses of complex SV (cxSV), revealing that: (1) cxSV are larger and rarer than canonical SV; (2) each genome harbors 14 large cxSV on average; (3) 84.4% of large cxSVs involve inversion; and (4) most large cxSV (93.8%) have not been delineated in previous studies. Rare SVs are more likely to disrupt coding and regulatory non-coding loci, particularly when truncating constrained and disease-associated genes. We also identify multiple cases of catastrophic chromosomal rearrangements known as chromoanagenesis, including somatic chromoanasynthesis, and extreme balanced germline chromothripsis events involving up to 65 breakpoints and 60.6 Mb across four chromosomes, further defining rare categories of extreme cxSV. Conclusions: These data provide a foundational map of large SV in the morbid human genome and demonstrate a previously underappreciated abundance and diversity of cxSV that should be considered in genomic studies of human disease. Electronic supplementary material The online version of this article (doi:10.1186/s13059-017-1158-6) contains supplementary material, which is available to authorized users

Characterization of Human Immunodeficiency Virus (HIV) Infection in Cisgender Men and Transgender Women Who Have Sex With Men Receiving Injectable Cabotegravir for HIV Prevention: HPTN 083.

BackgroundThe HIV Prevention Trials Network (HPTN) 083 trial demonstrated that long-acting cabotegravir (CAB-LA) was more effective than tenofovir disoproxil fumarate-emtricitabine (TDF/FTC) in preventing human immunodeficiency virus (HIV) in cisgender men and transgender women who have sex with men. We characterized HIV infections that occurred in the blinded phase of HPTN 083.MethodsRetrospective testing included HIV testing, viral load testing, quantification of study drugs, and HIV drug resistance testing.ResultsFifty-eight infections were evaluated, including 51 incident infections (12 in CAB arm and 39 in TDF/FTC arm). In many cases (5 in CAB arm and 37 in TDF/FTC arm), infection was associated with low or unquantifiable study drug concentrations. In 4 cases, infection occurred with on-time CAB-LA injections and expected plasma CAB concentrations. CAB exposure was associated with prolonged viral suppression and delayed antibody expression. In some cases, delayed HIV diagnosis resulted in CAB provision to participants with undetected infection, delayed antiretroviral therapy, and emergence of drug resistance; most of these infections would have been detected earlier with viral load testing.ConclusionsEarly detection of HIV infection and prompt antiretroviral therapy initiation could improve clinical outcomes in persons who become infected despite CAB-LA prophylaxis. Further studies are needed to elucidate the correlates of HIV protection in persons receiving CAB-LA