Validation of a Short Tandem Repeat Multiplex Typing System for Genetic Individualization of Domestic Cat Samples

Aim To conduct developmental validation studies on a polymerase chain reaction (PCR) based short tandem repeat (STR) multiplex typing system, developed for the purpose of genetic individualization and parentage testing in domestic cat samples. Methods To evaluate reproducibility of the typing system, the multiplex was amplified using DNA extracted from hair, blood, and buccal samples obtained from the same individual (n = 13). Additional studies were performed to evaluate the system’s species’ specificity, using 26 North American mammalian species and two prokaryotes Sacchromyces and Escherichia coli, sensitivity, and ability to identify DNA mixtures. Patterns of Mendelian inheritance and mutation rates for the 11 loci were directly examined in a large multi-generation domestic cat pedigree (n = 263). Results Our studies confirm that the multiplex system was species-specific for feline DNA and amplified robustly with as little as 125 picograms of genomic template DNA, demonstrating good product balance. The multiplex generated all components of a two DNA mixture when the minor component was one tenth of the major component at a threshold of 50 relative fluorescence units. The multiplex was reproducible in multiple tissue types of the same individual. Mutation rates for the 11 STR were within the range of sex averaged rates observed for Combined DNA Index System (CODIS) loci. Conclusion The cat STR multiplex typing system is a robust and reliable tool for the use of forensic DNA analysis of domestic cat samples

Genomic Microsatellites as Evolutionary Chronometers: A Test in Wild Cats

Nuclear microsatellite loci (2- to 5-bp tandem repeats) would seem to be ideal markers for population genetic monitoring because of their abundant polymorphism, wide dispersal in vertebrate genomes, near selective neutrality, and ease of assessment; however, questions about their mode of generation, mutation rates and ascertainment bias have limited interpretation considerably. We have assessed the patterns of genomic diversity for ninety feline microsatellite loci among previously characterized populations of cheetahs, lions and pumas in recapitulating demographic history. The results imply that the microsatellite diversity measures (heterozygosity, allele reconstitution and microsatellite allele variance) offer proportionate indicators, albeit with large variance, of historic population bottlenecks and founder effects. The observed rate of reconstruction of new alleles plus the growth in the breadth of microsatellite allele size (variance) was used here to develop genomic estimates of time intervals following historic founder events in cheetahs (12,000 yr ago), in North American pumas (10,000–17,000 yr ago), and in Asiatic lions of the Gir Forest (1000–4000 yr ago)

Recurrent Evolution of Melanism in South American Felids

Morphological variation in natural populations is a genomic test bed for studying the interface between molecular evolution and population genetics, but some of the most interesting questions involve non-model organisms that lack well annotated reference genomes. Many felid species exhibit polymorphism for melanism but the relative roles played by genetic drift, natural selection, and interspecies hybridization remain uncertain. We identify mutations of Agouti signaling protein (ASIP) or the Melanocortin 1 receptor (MC1R) as independent causes of melanism in three closely related South American species: the pampas cat (Leopardus colocolo), the kodkod (Leopardus guigna), and Geoffroy’s cat (Leopardus geoffroyi). To assess population level variation in the regions surrounding the causative mutations we apply genomic resources from the domestic cat to carry out clone-based capture and targeted resequencing of 299 kb and 251 kb segments that contain ASIP and MC1R, respectively, from 54 individuals (13–21 per species), achieving enrichment of ~500–2500-fold and ~150x coverage. Our analysis points to unique evolutionary histories for each of the three species, with a strong selective sweep in the pampas cat, a distinctive but short melanism-specific haplotype in the Geoffroy’s cat, and reduced nucleotide diversity for both ancestral and melanism-bearing chromosomes in the kodkod. These results reveal an important role for natural selection in a trait of longstanding interest to ecologists, geneticists, and the lay community, and provide a platform for comparative studies of morphological variation in other natural populations

Identification and assessment of intimate partner violence in nurse home visitation

Aims and objectives: To develop strategies for the identification and assessment of intimate partner violence in a nurse home visitation programme. Background: Nurse home visitation programmes have been identified as an intervention for preventing child abuse and neglect. Recently, there is an increased focus on the role these programmes have in addressing intimate partner violence. Given the unique context of the home environment, strategies for assessments are required that maintain the therapeutic alliance and minimise client attrition. Design: A qualitative case study. Methods: A total of four Nurse–Family Partnership agencies were engaged in this study. Purposeful samples of nurses (n = 32), pregnant or parenting mothers who had self-disclosed experiences of abuse (n = 26) and supervisors (n = 5) participated in this study. A total of 10 focus groups were completed with nurses: 42 interviews with clients and 10 interviews with supervisors. The principles of conventional content analysis guided data analysis. Data were categorised using the practice–problem–needs analysis model for integrating qualitative findings in the development of nursing interventions. Results: Multiple opportunities to ask about intimate partner violence are valued. The use of structured screening tools at enrolment does not promote disclosure or in-depth exploration of women\u27s experiences of abuse. Women are more likely to discuss experiences of violence when nurses initiate nonstructured discussions focused on parenting, safety or healthy relationships. Nurses require knowledge and skills to initiate indicator-based assessments when exposure to abuse is suspected as well as strategies for responding to client-initiated disclosures. Conclusion: A tailored approach to intimate partner violence assessment in home visiting is required. Relevance to clinical practice: Multiple opportunities for exploring women\u27s experiences of violence are required. A clinical pathway outlining a three-pronged approach to identification and assessment was developed

Endogenous Retrovirus Insertion in the KIT Oncogene Determines White and White spotting in Domestic Cats

The Dominant White locus (W) in the domestic cat demonstrates pleiotropic effects exhibiting complete penetrance for absence of coat pigmentation and incomplete penetrance for deafness and iris hypopigmentation. We performed linkage analysis using a pedigree segregating White to identify KIT (Chr. B1) as the feline W locus. Segregation and sequence analysis of the KIT gene in two pedigrees (P1 and P2) revealed the remarkable retrotransposition and evolution of a feline endogenous retrovirus (FERV1) as responsible for two distinct phenotypes of the W locus, Dominant White, and white spotting. A full-length (7125 bp) FERV1 element is associated with white spotting, whereas a FERV1 long terminal repeat (LTR) is associated with all Dominant White individuals. For purposes of statistical analysis, the alternatives of wild-type sequence, FERV1 element, and LTR-only define a triallelic marker. Taking into account pedigree relationships, deafness is genetically linked and associated with this marker; estimated P values for association are in the range of 0.007 to 0.10. The retrotransposition interrupts a DNAase I hypersensitive site in KIT intron 1 that is highly conserved across mammals and was previously demonstrated to regulate temporal and tissue-specific expression of KIT in murine hematopoietic and melanocytic cells. A large-population genetic survey of cats (n = 270), representing 30 cat breeds, supports our findings and demonstrates statistical significance of the FERV1 LTR and full-length element with Dominant White/blue iris (P \u3c 0.0001) and white spotting (P \u3c 0.0001), respectively

A Mutation in LTBP2 Causes Congenital Glaucoma in Domestic Cats (Felis catus)

The glaucomas are a group of diseases characterized by optic nerve damage that together represent a leading cause of blindness in the human population and in domestic animals. Here we report a mutation in LTBP2 that causes primary congenital glaucoma (PCG) in domestic cats. We identified a spontaneous form of PCG in cats and established a breeding colony segregating for PCG consistent with fully penetrant, autosomal recessive inheritance of the trait. Elevated intraocular pressure, globe enlargement and elongated ciliary processes were consistently observed in all affected cats by 8 weeks of age. Varying degrees of optic nerve damage resulted by 6 months of age. Although subtle lens zonular instability was a common feature in this cohort, pronounced ectopia lentis was identified in less than 10% of cats examined. Thus, glaucoma in this pedigree is attributed to histologically confirmed arrest in the early post-natal development of the aqueous humor outflow pathways in the anterior segment of the eyes of affected animals. Using a candidate gene approach, significant linkage was established on cat chromosome B3 (LOD 18.38, θ = 0.00) using tightly linked short tandem repeat (STR) loci to the candidate gene, LTBP2. A 4 base-pair insertion was identified in exon 8 of LTBP2 in affected individuals that generates a frame shift that completely alters the downstream open reading frame and eliminates functional domains. Thus, we describe the first spontaneous and highly penetrant non-rodent model of PCG identifying a valuable animal model for primary glaucoma that closely resembles the human disease, providing valuable insights into mechanisms underlying the disease and a valuable animal model for testing therapies

An ∼140-kb Deletion Associated with Feline Spinal Muscular Atrophy Implies an Essential LIX1 Function for Motor Neuron Survival

The leading genetic cause of infant mortality is spinal muscular atrophy (SMA), a clinically and genetically heterogeneous group of disorders. Previously we described a domestic cat model of autosomal recessive, juvenile-onset SMA similar to human SMA type III. Here we report results of a whole-genome scan for linkage in the feline SMA pedigree using recently developed species-specific and comparative mapping resources. We identified a novel SMA gene candidate, LIX1, in an ~140-kb deletion on feline chromosome A1q in a region of conserved synteny to human chromosome 5q15. Though LIX1 function is unknown, the predicted secondary structure is compatible with a role in RNA metabolism. LIX1 expression is largely restricted to the central nervous system, primarily in spinal motor neurons, thus offering explanation of the tissue restriction of pathology in feline SMA. An exon sequence screen of 25 human SMA cases, not otherwise explicable by mutations at the SMN1 locus, failed to identify comparable LIX1 mutations. Nonetheless, a LIX1-associated etiology in feline SMA implicates a previously undetected mechanism of motor neuron maintenance and mandates consideration of LIX1 as a candidate gene in human SMA when SMN1 mutations are not found

Effect of Addition of an Intimate Partner Violence Intervention to a Nurse Home Visitation Program on Maternal Quality of Life: A Randomized Clinical Trial

Importance: Intimate partner violence (IPV) is a public health problem with significant adverse consequences for women and children. Past evaluations of a nurse home visitation program for pregnant women and first-time mothers experiencing social and economic disadvantage have not consistently shown reductions in IPV. Objective: To determine the effect on maternal quality of life of a nurse home visitation program augmented by an IPV intervention, compared with the nurse home visitation program alone. Design, Setting, and Participants: Cluster-based, single-blind, randomized clinical trial at 15 sites in 8 US states (May 2011-May 2015) enrolling 492 socially disadvantaged pregnant women (≥16 years) participating in a 2.5-year nurse home visitation program. Interventions: In augmented program sites (n = 229 participants across 7 sites), nurses received intensive IPV education and delivered an IPV intervention that included a clinical pathway to guide assessment and tailor care focused on safety planning, violence awareness, self-efficacy, and referral to social supports. The standard program (n = 263 participants across 8 sites) included limited questions about violence exposure and information for abused women but no standardized IPV training for nurses. Main Outcomes and Measures: The primary outcome was quality of life (WHOQOL-BREF; range, 0-400; higher score indicates better quality of life) obtained through interviews at baseline and every 6 months until 24 months after delivery. From 17 prespecified secondary outcomes, 7 secondary end points are reported, including scores on the Composite Abuse Scale, SPAN (Startle, Physiological Arousal, Anger, and Numbness), Prime-MD Patient Health Questionnaire, TWEAK (Tolerance/Worry About Drinking/Eye-Opener/Amnesia/C[K]ut Down on Drinking), Drug Abuse Severity Test, and the 12-Item Short-Form Health Survey (physical and mental health), version 2. Results: Among 492 participants enrolled (mean age, 20.4 years), 421 (86%) completed the trial. Quality of life improved from baseline to 24 months in both groups (change in WHOQOL-BREF scores from 299.5 [SD, 54.4] to 308.2 [SD, 52.6] in the augmented program group vs from 293.6 [SD, 56.4] to 316.4 [SD, 57.5] in the standard program group). Based on multilevel growth curve analysis, there was no statistically significant difference between groups (modeled score difference, -4.9 [95% CI, -16.5 to 6.7]). There were no statistically significant differences between study groups in any of the secondary participant end points. There were no adverse events recorded in either group. Conclusions and Relevance: Among pregnant women experiencing social and economic disadvantage and preparing to parent for the first time, augmentation of a nurse home visitation program with a comprehensive IPV intervention, compared with the home visitation program alone, did not significantly improve quality of life at 24 months after delivery. These findings do not support the use of this intervention. Trial Registration: ClinicalTrials.gov Identifier: NCT01372098