Trends in companion animal access to veterinary care in Canada, 2007 to 2020.

Objective: Assess trends in access to veterinary care for companion animals in Canada. Procedure: Analysis and integration of available data, 2007 to 2020. Results: Cumulative growth in the Canadian veterinary workforce was 38%, and 49% for companion animal veterinarians. Clients per companion animal veterinarian decreased 30% from 2008 to 2020. Absolute client numbers increased 1.3%, compared to pet population growth of 17%. Medicalized pets (those that had received veterinary care in the past year) increased 25%, from 9.02 million in 2007 to 11.24 million in 2020. Non-medicalized pets increased 1.8%, from 4.48 million to 4.56 million. In 2007, 33% of pets were non-medicalized, compared to 29% (15% of dogs and 42% of cats) in 2020. There was a cumulative increase of 31% for total non-medicalized dogs, and a change of -5.6% for cats. Gross and net revenues per client increased by 99 and 112%, respectively, compared to cumulative inflation of 21%. Conclusion and clinical relevance: The analysis identified a large cohort of pets that had not received veterinary care each year. The trends were fewer clients per veterinarian, each paying higher veterinary costs, and suggested a relative, rather than absolute, veterinary capacity shortage overall. Accessible care-provision models must be encouraged, regulated for, and allowed to flourish alongside traditional models.

Tendances en matière d'accès des animaux de compagnie aux soins vétérinaires au Canada, 2007 à 2020. Objectif: Évaluer les tendances en matière d'accès aux soins vétérinaires pour les animaux de compagnie au Canada. Procédure: Analyse et intégration des données disponibles, 2007 à 2020. Résultats: La croissance cumulative de la main-d'œuvre vétérinaire canadienne était de 38 %, et de 49 % pour les médecins vétérinaires d'animaux de compagnie. Le nombre de clients par vétérinaire pour animaux de compagnie a diminué de 30 % entre 2008 et 2020. Le nombre absolu de clients a augmenté de 1,3 %, comparé à une croissance de la population d'animaux de compagnie de 17 %. Les animaux de compagnie médicalisés (ceux qui ont reçu des soins vétérinaires au cours de l'année écoulée) ont augmenté de 25 %, passant de 9,02 millions en 2007 à 11,24 millions en 2020. Les animaux de compagnie non médicalisés ont augmenté de 1,8 %, passant de 4,48 millions à 4,56 millions. En 2007, 33 % des animaux de compagnie étaient non médicalisés, contre 29 % (15 % des chiens et 42 % des chats) en 2020. Il y a eu une augmentation cumulée de 31 % pour le total des chiens non médicalisés, et une variation de ­5,6 % pour les chats. Les revenus bruts et nets par client ont augmenté respectivement de 99 et 112 %, par rapport à une inflation cumulée de 21 %. Conclusion et pertinence clinique: L'analyse a identifié chaque année une large cohorte d'animaux de compagnie qui n'avaient pas reçu de soins vétérinaires. Les tendances étaient moins de clients par vétérinaire, chacun payant des frais vétérinaires plus élevés, et suggéraient une pénurie globale de capacité vétérinaire relative plutôt qu'absolue. Les modèles de prestation de soins accessibles doivent être encouragés, réglementés et autorisés à prospérer aux côtés des modèles traditionnels.(Traduit par Dr Serge Messier).

Efficacy of the Fast Track Friendship Group Program for Peer-rejected Children: A Randomized-Controlled Trial.

OBJECTIVE: To evaluate the benefits of the Fast Track Friendship Group program implemented as a stand-alone school-based intervention on the social cognitions, social behavior, peer and teacher relationships of peer-rejected students. METHOD: Over four successive years, 224 peer-rejected elementary students (57% White, 17% Black, 20% Latinx, 5% multiracial; 68% male; grades 1-4; Mage = 8.1 years old) were identified using peer sociometric nominations and randomly assigned to the intervention (n = 110) or a treatment-as-usual control group (n = 114). Four school districts serving economically-disadvantaged urban and rural communities participated. Intervention involved weekly small group social skills training with classmate partners, with sessions tailored to address individual child needs. Consultation meetings held at the start and mid-point of intervention were designed to help teachers and parents support the generalization of targeted skills. RESULTS: Multi-level linear models, with children nested within schools (controlling for demographics and baseline scores) documented improvements in social-cognitive skills (direct assessments of emotion recognition and competent social problem-solving), social behavior (teacher ratings of social skills and externalizing behavior), and interpersonal relationships (peer sociometric nominations of peer acceptance and friendships, teacher-rated student-teacher closeness). Significant effects were generally small (ds = .19-.36) but consistent across child sex, grade level, and behavioral characteristics. CONCLUSIONS: The intervention proved feasible for high-fidelity implementation in school settings and produced significant improvements in the social adjustment of peer-rejected children, validating the approach as a school-based Tier 2 intervention.

Risk Factors for SARS-CoV-2 Infection and Illness in Cats and Dogs1.

We tested swab specimens from pets in households in Ontario, Canada, with human COVID-19 cases by quantitative PCR for SARS-CoV-2 and surveyed pet owners for risk factors associated with infection and seropositivity. We tested serum samples for spike protein IgG and IgM in household pets and also in animals from shelters and low-cost neuter clinics. Among household pets, 2% (1/49) of swab specimens from dogs and 7.7% (5/65) from cats were PCR positive, but 41% of dog serum samples and 52% of cat serum samples were positive for SARS-CoV-2 IgG or IgM. The likelihood of SARS-CoV-2 seropositivity in pet samples was higher for cats but not dogs that slept on owners' beds and for dogs and cats that contracted a new illness. Seropositivity in neuter-clinic samples was 16% (35/221); in shelter samples, 9.3% (7/75). Our findings indicate a high likelihood for pets in households of humans with COVID-19 to seroconvert and become ill.

Cu2+-based distance measurements by pulsed EPR provide distance constraints for DNA backbone conformations in solution.

Electron paramagnetic resonance (EPR) has become an important tool to probe conformational changes in nucleic acids. An array of EPR labels for nucleic acids are available, but they often come at the cost of long tethers, are dependent on the presence of a particular nucleotide or can be placed only at the termini. Site directed incorporation of Cu2+-chelated to a ligand, 2,2'dipicolylamine (DPA) is potentially an attractive strategy for site-specific, nucleotide independent Cu2+-labelling in DNA. To fully understand the potential of this label, we undertook a systematic and detailed analysis of the Cu2+-DPA motif using EPR and molecular dynamics (MD) simulations. We used continuous wave EPR experiments to characterize Cu2+ binding to DPA as well as optimize Cu2+ loading conditions. We performed double electron-electron resonance (DEER) experiments at two frequencies to elucidate orientational selectivity effects. Furthermore, comparison of DEER and MD simulated distance distributions reveal a remarkable agreement in the most probable distances. The results illustrate the efficacy of the Cu2+-DPA in reporting on DNA backbone conformations for sufficiently long base pair separations. This labelling strategy can serve as an important tool for probing conformational changes in DNA upon interaction with other macromolecules.

Promoting Social and Emotional Learning in Middle School: Intervention Effects of Facing History and Ourselves.

There are fewer evidence-based social and emotional learning programs for middle school students compared to younger grades. This randomized controlled trial tests the effectiveness of Facing History and Ourselves (hereafter, Facing History) with a sample of 694 (Facing History n = 437; Comparison n = 257) students from a low-resourced school district. Youth self-identified as female (59%), Black/African American (61%), Hispanic/Latinx (18%), White (2%), and multi-racial or some other race/ethnicity (19%). Intervention students perceived their classrooms as more caring and democratic than students in the comparison group. They reported higher levels of empathy, prosocial behavior, and stronger participatory citizenship beliefs. This study expands the evidence-based of effective SEL programs available to schools.

Factor VIII prophylaxis effects outweigh other hemostasis contributors in predicting severe haemophilia A joint outcomes.

INTRODUCTION: The Joint Outcome Study (JOS) demonstrated that previously untreated children with severe haemophilia A treated with prophylactic factor VIII (FVIII) concentrate had superior joint outcomes at age 6 years compared to those children treated episodically for bleeding. However, variation in joint outcome within each treatment arm was not well explained. AIM: In this study, we sought to better understand variation in joint outcomes at age 6 years in participants of the JOS. METHODS: We evaluated the influence of FVIII half-life, treatment adherence, constitutional coagulant and anticoagulant proteins, and global assays on joint outcomes (number of joint bleeds, total number of bleeds, total MRI score and joint physical exam score). Logistic regression was used to evaluate the association of variables with joint failure status on MRI, defined as presence of subchondral cyst, surface erosion or joint-space narrowing. Each parameter was also correlated with each joint outcome using Spearman correlations. RESULTS: Prophylaxis treatment arm and FVIII trough were each found to reduce risk of joint failure on univariate logistic regression analysis. When controlling for treatment arm, FVIII trough was no longer significant, likely because of the high level of covariation between these variables. We found no consistent correlation between any laboratory assay performed and any joint outcome parameter measured. CONCLUSION: In the JOS, the effect of prescribed prophylactic FVIII infusions on joint outcome overshadowed the contribution of treatment adherence, FVIII half-life, global assays of coagulation and constitutional coagulation proteins. (ClinicalTrials.gov number, NCT00207597).

Metal Ion Binding at the Catalytic Site Induces Widely Distributed Changes in a Sequence Specific Protein-DNA Complex.

Metal ion cofactors can alter the energetics and specificity of sequence specific protein-DNA interactions, but it is unknown if the underlying effects on structure and dynamics are local or dispersed throughout the protein-DNA complex. This work uses EcoRV endonuclease as a model, and catalytically inactive lanthanide ions, which replace the Mg2+ cofactor. Nuclear magnetic resonance (NMR) titrations indicate that four Lu3+ or two La3+ cations bind, and two new crystal structures confirm that Lu3+ binding is confined to the active sites. NMR spectra show that the metal-free EcoRV complex with cognate (GATATC) DNA is structurally distinct from the nonspecific complex, and that metal ion binding sites are not assembled in the nonspecific complex. NMR chemical shift perturbations were determined for 1H-15N amide resonances, for 1H-13C Ile-δ-CH3 resonances, and for stereospecifically assigned Leu-δ-CH3 and Val-γ-CH3 resonances. Many chemical shifts throughout the cognate complex are unperturbed, so metal binding does not induce major conformational changes. However, some large perturbations of amide and side chain methyl resonances occur as far as 34 Å from the metal ions. Concerted changes in specific residues imply that local effects of metal binding are propagated via a ß-sheet and an α-helix. Both amide and methyl resonance perturbations indicate changes in the interface between subunits of the EcoRV homodimer. Bound metal ions also affect amide hydrogen exchange rates for distant residues, including a distant subdomain that contacts DNA phosphates and promotes DNA bending, showing that metal ions in the active sites, which relieve electrostatic repulsion between protein and DNA, cause changes in slow dynamics throughout the complex.

ESR spectroscopy identifies inhibitory Cu2+ sites in a DNA-modifying enzyme to reveal determinants of catalytic specificity.

The relationship between DNA sequence recognition and catalytic specificity in a DNA-modifying enzyme was explored using paramagnetic Cu(2+) ions as probes for ESR spectroscopic and biochemical studies. Electron spin echo envelope modulation spectroscopy establishes that Cu(2+) coordinates to histidine residues in the EcoRI endonuclease homodimer bound to its specific DNA recognition site. The coordinated His residues were identified by a unique use of Cu(2+)-ion based long-range distance constraints. Double electron-electron resonance data yield Cu(2+)-Cu(2+) and Cu(2+)-nitroxide distances that are uniquely consistent with one Cu(2+) bound to His114 in each subunit. Isothermal titration calorimetry confirms that two Cu(2+) ions bind per complex. Unexpectedly, Mg(2+)-catalyzed DNA cleavage by EcoRI is profoundly inhibited by Cu(2+) binding at these hitherto unknown sites, 13 Å away from the Mg(2+) positions in the catalytic centers. Molecular dynamics simulations suggest a model for inhibition of catalysis, whereby the Cu(2+) ions alter critical protein-DNA interactions and water molecule positions in the catalytic sites. In the absence of Cu(2+), the Mg(2+)-dependence of EcoRI catalysis shows positive cooperativity, which would enhance EcoRI inactivation of foreign DNA by irreparable double-strand cuts, in preference to readily repaired single-strand nicks. Nonlinear Poisson-Boltzmann calculations suggest that this cooperativity arises because the binding of Mg(2+) in one catalytic site makes the surface electrostatic potential in the distal catalytic site more negative, thus enhancing binding of the second Mg(2+). Taken together, our results shed light on the structural and electrostatic factors that affect site-specific catalysis by this class of endonucleases.

FVIII/VWF ratio is not a reliable predictor of VWD in children.

Adults with von Willebrand Disease (VWD) are known to have a ratio of factor VIII activity (FVIII:C) to von Willebrand factor antigen (VWF:Ag) greater than 1. We, however, noted healthy children with ratios that are unexpectedly high. Though the FVIII:C/VWF:Ag ratio differs significantly between healthy children and VWD patients in some age groups, the substantial overlap of observed ranges suggests that a ratio threshold-based screening approach alone cannot reliably discriminate between these groups. The diagnostic performance of this ratio is poor for VWD in children, which may decrease its value as a screening tool in the pediatric population.

Divide and conquer is always best: sensitivity of methyl correlation experiments.

The HMCM [CG]CBCA experiment (Tugarinov and Kay in J Am Chem Soc 125:13868-13878, 2003) correlates methyl carbon and proton shifts to Cγ, Cß, and Cα resonances for the purpose of resonance assignments. The relative sensitivity of the HMCM[CG]CBCA sequence experiment is compared to a divide-and-conquer approach to assess whether it is best to collect all of the methyl correlations at once, or to perform separate experiments for each correlation. A straightforward analysis shows that the divide-and-conquer approach is intrinsically more sensitive, and should always be used to obtain methyl-Cγ, Cß, and Cα correlations. The improvement in signal-to-noise associated with separate experiments is illustrated by the detection of methyl-aliphatic correlations in a 65 kDa protein-DNA complex.

Elevated lipoprotein (a), small apolipoprotein (a), and the risk of arterial ischemic stroke in North American children.

Lipoprotein (a) is a risk factor for adult cardiovascular events, in which the apolipoprotein (a) component is thought to promote atherogenesis and impair fibrinolysis. We investigated whether elevated plasma lipoprotein (a) concentration and small predominant apolipoprotein (a) isoform size (number of kringle-4 domains) are risk factors for childhood arterial ischemic stroke and correlate with plasma fibrinolytic function. Patients who had had an arterial ischemic stroke in childhood (29 days - <21 years at onset; n=43) and healthy controls (n=127) were recruited for plasma sampling and laboratory determinations. Cases were followed for recurrence in a prospective cohort study. The median lipoprotein (a) concentration did not differ between groups [cases: median 18.0 nmol/L (7.5 mg/dL) and observed range 0.9-259 nmol/L (0.38-108.0 mg/dL), controls: 20.4 nmol/L (8.5 mg/dL) and 0.2-282 nmol/L (0.08-117.5 mg/dL); P=0.62]. While odds of incident stroke were not significantly increased, risks of recurrent arterial ischemic stroke were each more than ten-times increased for lipoprotein(a) >90(th) percentile of race-specific reference values and apolipoprotein (a) <10(th) percentiles [odds ratio=14.0 (95% confidence interval: 1.0-184), P=0.05 and odds ratio=12.8 (1.61-101), P=0.02]. Statistically significant but weak correlations were observed between euglobulin lysis time and both lipoprotein (a) level (r=0.18, P=0.03) and apolipoprotein (a) size (r= -0.26, P=0.002). In conclusion, elevated lipoprotein (a) and small apolipoprotein (a) potently increase the risk of recurrent arterial ischemic stroke in children, with a mechanism only partially attributable to impaired fibrinolysis. Collaborative studies are warranted to investigate these findings further and, more broadly, to establish key risk factors for incident and recurrent arterial ischemic stroke in children.

Further variation of the adulticide protocol for the treatment of canine heartworm infection: can it be even shorter and cost less?

BACKGROUND: This retrospective study evaluated modified three-dose melarsomine treatment protocols in a shelter setting and compared them to the American Heartworm Society (AHS)-recommended protocol. METHODS: As compared with the AHS protocol, the shelter protocols utilized doxycycline 10 mg/kg once daily (SID) or twice daily (BID), and varied the time from initiation of doxycycline (day 1) to the first melarsomine injection (M1). Dogs were retrospectively grouped based on the shelter's current protocol (M1 on day 14; Group A) and the AHS protocol (M1 on day 60; Group C), allowing a week on either side of the target M1 day. Treatments that fell outside these ranges formed two additional treatment groups (Groups B and D). Respiratory complications were defined as respiratory signs requiring additional treatment, and were statistically compared for Groups A and C. New respiratory signs and gastrointestinal (GI) signs were compared between dogs receiving SID or BID doxycycline. RESULTS: One hundred fifty-seven dogs with asymptomatic or mild heartworm disease at presentation were included. All dogs survived to discharge. There was no statistically significant difference between Groups A (n = 79) and C (n = 27) for new respiratory signs post-melarsomine (P = 0.73). The time to M1 for 14 dogs that developed new respiratory signs was a median of 19 days, compared with 22 days for 143 dogs without new respiratory signs (P = 0.2). Respiratory complications post-melarsomine were uncommon. New respiratory signs post-melarsomine occurred in 10/109 (9.2%) dogs receiving SID doxycycline and 4/48 (8.3%) dogs receiving BID doxycycline (P > 0.999). GI signs prior to M1 were recorded for 40/109 (36.7%) dogs receiving SID doxycycline and 25/48 (52.1%) receiving BID doxycycline (P = 0.08). Forty-four follow-up antigen test results were available; all tests performed > 3 months after the third melarsomine injection were negative. CONCLUSIONS: This study provided support for initiating melarsomine after 14 days of doxycycline and for a lower doxycycline dose. Shorter and less expensive treatment protocols can increase lifesaving capacity and improve quality of life for shelter dogs by reducing the duration of exercise restriction and length of stay.

A Pandora's box in feline medicine: presenting signs and surgical outcomes in 58 previously hoarded cats with chronic otitis media-interna.

OBJECTIVES: The aim of the present study was to report clinical findings, surgical complications and outcomes for previously hoarded cats treated surgically for otitis media-interna (OMI) and to investigate the risk factors for complications and poor outcomes. METHODS: A retrospective study was conducted of 58 cats from an institutional hoarding environment that underwent ventral bulla osteotomy (VBO). RESULTS: Inappetence was uncommon at presentation (9/58, 16%) compared with pruritus/alopecia (50%), nasopharyngeal signs (45%), otitis externa (OE) (79%) and otitis interna (OI) (ataxia ± head tilt/head excursions) in 40%. Purulent aural discharge occurred in 36% and polyps in 26%. The tympanic bulla wall was moderately or severely thickened radiographically in 38/108 (35%) ears. Cultures were positive for Streptococcus equi subspecies zooepidemicus in 26/48 (54%) cats. Of the 58 cats, 40 (69%) had complications after the first VBO and 19/30 (63%) after the second. Of 101 complications, 56 (55%), from 27/88 (31%) surgeries, were considered serious, including life-threatening perioperative complications in seven, OI in eight, prolonged anorexia in six and worsening of pruritus/alopecia in nine cases. Three cats developed xerostomia (dry mouth) after the second VBO. Pruritus/alopecia, nasopharyngeal signs, OE and purulent aural discharge resolved in a statistically significant proportion of cats but persisted in some. Full resolution of OI was uncommon. OI preoperatively, and surgery performed by a generalist (vs specialist) surgeon, were risk factors for OE at recheck (OI: odds ratio [OR] 4.35; 95% confidence interval [CI] 1.21-15.70; P = 0.02; surgery: OR 3.64; 95% CI 1.03-12.87; P = 0.045). No other prognostic indicators were identified. No variables tested were significantly associated with risk of serious complications or euthanasia. CONCLUSIONS AND RELEVANCE: Surgical management of chronic OMI was successful in most cases but was not benign and not always beneficial. The analysis was unable to identify clinically helpful outcome predictors. Optimal management of chronic feline OMI remains a challenge, particularly for animal shelters. Less invasive approaches and chronic medical management require further investigation.

Differential temperature-dependent multimeric assemblies of replication and repair polymerases on DNA increase processivity.

Differentiation of binding accurate DNA replication polymerases over error prone DNA lesion bypass polymerases is essential for the proper maintenance of the genome. The hyperthermophilic archaeal organism Sulfolobus solfataricus (Sso) contains both a B-family replication (Dpo1) and a Y-family repair (Dpo4) polymerase and serves as a model system for understanding molecular mechanisms and assemblies for DNA replication and repair protein complexes. Protein cross-linking, isothermal titration calorimetry, and analytical ultracentrifugation have confirmed a previously unrecognized dimeric Dpo4 complex bound to DNA. Binding discrimination between these polymerases on model DNA templates is complicated by the fact that multiple oligomeric species are influenced by concentration and temperature. Temperature-dependent fluorescence anisotropy equilibrium binding experiments were used to separate discrete binding events for the formation of trimeric Dpo1 and dimeric Dpo4 complexes on DNA. The associated equilibria are found to be temperature-dependent, generally leading to improved binding at higher temperatures for both polymerases. At high temperatures, DNA binding of Dpo1 monomer is favored over binding of Dpo4 monomer, but binding of Dpo1 trimer is even more strongly favored over binding of Dpo4 dimer, thus providing thermodynamic selection. Greater processivities of nucleotide incorporation for trimeric Dpo1 and dimeric Dpo4 are also observed at higher temperatures, providing biochemical validation for the influence of tightly bound oligomeric polymerases. These results separate, quantify, and confirm individual and sequential processes leading to the formation of oligomeric Dpo1 and Dpo4 assemblies on DNA and provide for a concentration- and temperature-dependent discrimination of binding undamaged DNA templates at physiological temperatures.

Fecal viral DNA shedding following clinical panleukopenia virus infection in shelter kittens: a prospective, observational study.

OBJECTIVES: The aims of this study were to determine the magnitude and duration of fecal viral DNA shedding after diagnosis of feline panleukopenia (FP) in a group of shelter cats using quantitative real-time PCR (qPCR); to assess the utility of a negative point-of-care test or the resolution of diarrhea and systemic signs as proxy measures for qPCR positivity; and to investigate patterns of additional enteric pathogens in relation to feline panleukopenia viral shedding duration. METHODS: Feline panleukopenia virus (FPV) infection in clinically affected shelter cats was confirmed by a commercial qPCR test. Observations were made on days 0, 3, 7, 14 and 21 post-diagnosis. Fecal flotation, FPV qPCR and the canine parvovirus IDEXX SNAP Parvo ELISA (SNAP) test were performed on fecal samples. RESULTS: Forty cats and kittens with confirmed panleukopenia were initially enrolled. Sixteen kittens were sampled until day 14, and 12 were followed to day 21. Median DNA viral copy numbers fell below the diagnostic cut-off by day 7, with 13/16, 6/16, 1/16 and 0/12 testing PCR-positive on days 3, 7, 14 and 21, respectively. The SNAP test was positive in 12/16 kittens on day 0 and only 3/16 on day 3. SNAP test results, diarrhea and systemic signs were inconsistent in relation to qPCR positivity post-diagnosis. Additional enteric pathogens were common. The presence of additional pathogen types was suggestive of a longer PCR shedding duration, but this was not tested statistically owing to the small sample size. CONCLUSIONS AND RELEVANCE: These findings suggest that cats should be isolated for at least 14 days after a diagnosis of FP, but that release from isolation after this point is reasonable, in association with a multifaceted infection control strategy. The study findings did not support using SNAP test results, diarrhea or systemic signs as proxy measures for virus shedding.

Behavior and adoptability of hoarded cats admitted to an animal shelter.

OBJECTIVES: The aim of this study was to analyze the behavioral characteristics and success of adoption for previously hoarded cats. METHODS: Shelter records and post-adoption surveys were analyzed for hoarded cats ⩾6 months old at intake. A non-standard scoring system was used. Intake scores were allocated contemporaneously and socialization scores were applied retrospectively for three time points (TPs): 5-10 days post-intake (shelter TP), ⩽1 week post-adoption (home TP1) and >1 week post-adoption (home TP2). Adoption returns were compared between hoarded and non-hoarded cats. RESULTS: The study included 195 hoarded cats, of which 174 were adopted. Of 164 cats with intake scores, 86 (52%) were scored as 'friendly' at intake. Forty-five cats had socialization scores for all of the TPs, and of these, the percentages of 'supersocial' or 'social' decreased from 87% at the shelter TP to 47% at home TP1, then increased to 84% at home TP2. Most cats that scored as 'tense' at intake had supersocial or social scores at home TP2. Nine of the 88 cats with survey results had out-of-box (OOB) elimination in either the shelter or home but only 1/88 in both. Adopters expressed positive feelings for 42/43 cats for which feelings-based language was used in their survey responses. Notable behaviors, such as neediness, were recorded for 48/88 cats. Relationships with other household pets were typically positive. Eighteen of 174 hoarded (10%) and 188/2662 non-hoarded (7.1%) cats were returned post-adoption. Of these, six hoarded and 87 non-hoarded returns included behavioral reasons. There were no significant differences between hoarded and non-hoarded cats for total or behavioral returns. CONCLUSIONS AND RELEVANCE: Hoarded cats had high adoption rates, high adopter satisfaction and the potential for good emotional well-being in adoptive homes. Behavior at intake and OOB elimination in the shelter may not reflect post-adoption behavior. Behavior-based outcome decisions for these vulnerable animals should be deferred to allow time for habituation.

Specific labeling of threonine methyl groups for NMR studies of protein-nucleic acid complexes.

Specific (13)C labeling of Thr methyl groups has been accomplished via the growth of a standard laboratory strain of Escherichia coli on [2-(13)C]glycerol in the presence of deuterated isoketovalerate, Ile, and Ala. Diversion of the label from the Thr biosynthetic pathway is suppressed by including Lys, Met, and Ile in the growth medium. This method complements the repertoire of methyl labeling schemes for NMR structural and dynamic studies of proteins and is particularly useful for the study of nucleic acid binding proteins because of the high propensity of Thr residues at protein-DNA and -RNA interfaces.

A trimeric DNA polymerase complex increases the native replication processivity.

DNA polymerases are essential enzymes in all domains of life for both DNA replication and repair. The primary DNA replication polymerase from Sulfolobus solfataricus (SsoDpo1) has been shown previously to provide the necessary polymerization speed and exonuclease activity to replicate the genome accurately. We find that this polymerase is able to physically associate with itself to form a trimer and that this complex is stabilized in the presence of DNA. Analytical gel filtration and electrophoretic mobility shift assays establish that initially a single DNA polymerase binds to DNA followed by the cooperative binding of two additional molecules of the polymerase at higher concentrations of the enzyme. Protein chemical crosslinking experiments show that these are specific polymerase-polymerase interactions and not just separate binding events along DNA. Isothermal titration calorimetry and fluorescence anisotropy experiments corroborate these findings and show a stoichiometry where three polymerases are bound to a single DNA substrate. The trimeric polymerase complex significantly increases both the DNA synthesis rate and the processivity of SsoDpo1. Taken together, these results suggest the presence of a trimeric DNA polymerase complex that is able to synthesize long DNA strands more efficiently than the monomeric form.