Impact of diet on faecal output and caecotroph consumption in rabbits.

OBJECTIVES: To assess the impact of four rabbit diets (hay only, extruded diet with hay, muesli with hay and muesli only) on faecal pellet size, faecal output and caecotrophy. MATERIALS AND METHODS: Thirty-two Dutch rabbits were studied over 17 months. Faecal pellet size and weight were measured in weeks 3, 9, 21 and 43 and faecal output in weeks 10, 22 and 45. Number of uneaten caecotrophs was recorded weekly. RESULTS: Faecal pellets were consistently smaller and lighter in rabbits fed muesli only, and the size of pellets produced by those fed muesli with hay decreased over the course of the study. Faecal output was greatest in rabbits with the highest hay intake. Uneaten caecotrophs were found in greatest frequency in rabbits fed muesli. CLINICAL SIGNIFICANCE: Muesli diets have a negative effect on faecal output and caecotroph ingestion and may therefore predispose to digestive disorders. Higher hay intake is associated with greater faecal output and fewer uneaten caecotrophs and may assist in preventing the gastrointestinal stasis.

MaxiK channel interactome reveals its interaction with GABA transporter 3 and heat shock protein 60 in the mammalian brain.

Large conductance voltage and calcium-activated potassium (MaxiK) channels are activated by membrane depolarization and elevated cytosolic Ca(2+). In the brain, they localize to neurons and astrocytes, where they play roles such as resetting the membrane potential during an action potential, neurotransmitter release, and neurovascular coupling. MaxiK channels are known to associate with several modulatory proteins and accessory subunits, and each of these interactions can have distinct physiological consequences. To uncover new players in MaxiK channel brain physiology, we applied a directed proteomic approach and obtained MaxiK channel pore-forming α subunit brain interactome using specific antibodies. Controls included immunoprecipitations with rabbit immunoglobulin G (IgG) and with anti-MaxiK antibodies in wild type and MaxiK channel knockout mice (Kcnma1(-/-)), respectively. We have found known and unreported interactive partners that localize to the plasma membrane, extracellular space, cytosol and intracellular organelles including mitochondria, nucleus, endoplasmic reticulum and Golgi apparatus. Localization of MaxiK channel to mitochondria was further confirmed using purified brain mitochondria colabeled with MitoTracker. Independent proof of MaxiK channel interaction with previously unidentified partners is given for GABA transporter 3 (GAT3) and heat shock protein 60 (HSP60). In human embryonic kidney 293 cells containing SV40 T-antigen (HEK293T) cells, both GAT3 and HSP60 coimmunoprecipitated and colocalized with MaxiK channel; colabeling was observed mainly at the cell periphery with GAT3 and intracellularly with HSP60 with protein proximity indices of ∼ 0.6 and ∼ 0.4, respectively. In rat primary hippocampal neurons, colocalization index was identical for GAT3 (∼ 0.6) and slightly higher for HSP60 (∼ 0.5) association with MaxiK channel. The results of this study provide a complete interactome of MaxiK channel the mouse brain, further establish the localization of MaxiK channel in the mouse brain mitochondria and demonstrate the interaction of MaxiK channel with GAT3 and HSP60 in neurons. The interaction of MaxiK channel with GAT3 opens the possibility of a role of MaxiK channel in GABA homeostasis and signaling.

Impact of diet on incisor growth and attrition and the development of dental disease in pet rabbits.

OBJECTIVES: To assess the impact of four rabbit diets (hay only; extruded diet with hay; muesli with hay; muesli only) on length and curvature of cheek teeth and eruption and attrition rates of incisors. MATERIALS AND METHODS: Thirty-two Dutch rabbits, randomly divided into four diet groups, had length and saggital plane curvature of the first cheek teeth measured radiographically at 1, 9 and 17 months. Eruption/attrition of the left upper incisor was directly measured at weeks 30, 32 and 35. RESULTS: Eruption rates matched attrition rates in all groups, but were higher in the hay only group than in both groups fed muesli. By month 9, a greater degree of tooth curvature was present in rabbits fed muesli only than in those fed hay only and extruded diet with hay. After 17 months, rabbits fed muesli only and muesli with hay had longer lower first cheek teeth and larger interdental spaces between the first two molars than rabbits fed extruded diet and hay and hay only. Three rabbits fed muesli only developed evidence of dental disease. CLINICAL SIGNIFICANCE: Presence of increased tooth length, curvature and interdental spaces indicated early dental pathology in rabbits fed muesli. Muesli diets cannot be recommended for pet rabbits.

Bodyweight and body condition score in rabbits on four different feeding regimes.

OBJECTIVES: The aim of this study was to assess the effects of four diet regimes (extruded diet with ad lib hay, muesli with ad lib hay, ad lib hay only, ad lib muesli only) on bodyweight and body condition score in rabbits. METHODS: Thirty-two Dutch rabbits were studied over 9 months. Bodyweight and body condition score were recorded weekly. RESULTS: All groups gained weight with age, but relative to the ad lib hay only group (mean, 1 · 77 ± 0 · 13 kg), after 9 months rabbits in the ad lib muesli only group were 146% heavier (2 · 59 ± 0 · 32 kg); extruded diet with ad lib hay group 125% heavier (2 · 21 ± 0 · 10 kg); and muesli with ad lib hay group 123% heavier (2 · 18 ± 0 · 13 kg). Median body condition score of the ad lib muesli only group was obese (4 · 5) and was higher than that in both the extruded diet with ad lib hay and muesli with ad lib hay (median = 3 · 5) groups (P < 0 · 001). Both the extruded diet with ad lib hay and muesli with ad lib hay groups had above-ideal body condition score despite having bodyweights within the breed standard range. The ad lib hay only group had a median body condition score of 3 (ideal). There was an overall positive correlation between body condition score and weight (qs = 0 · 814, P < 0 · 001). CLINICAL SIGNIFICANCE: The feeding of muesli without hay should be avoided, to prevent obesity.

Coxiella burnetii (Q-Fever) Seroprevalence in Prey and Predators in the United Kingdom: Evaluation of Infection in Wild Rodents, Foxes and Domestic Cats Using a Modified ELISA.

Coxiella burnetii, the agent of Q-fever, is recognized as a worldwide zoonosis with a wide host range and potentially complex reservoir systems. Infected ruminants are the main source of infection for humans, but cats and other mammals, including wild rodents, also represent potential sources of infection. There has been a recent upsurge of reported cases in humans, domestic ruminants and wildlife in many parts of the world, and studies have indicated that wild brown rats may act as true reservoirs for C. burnetii and be implicated in outbreaks in livestock and humans. However, investigation of reservoir systems is limited by lack of validated serological tests for wildlife or other non-target species. In this study, serum samples from 796 wild rodents (180 bank voles, 309 field voles, 307 wood mice) 102 wild foxes and 26 domestic cats from three study areas in the UK were tested for the presence of antibodies to C. burnetii using a commercial indirect ELISA kit modified for use in multiple wildlife species. Test thresholds were determined for each species in the absence of species-specific reference sera using a bi-modal latent class mixture model to discriminate between positive from negative results. Based on the thresholds determined, seroprevalence in the wild rodents ranged from 15.6% to 19.1% depending on species (overall 17.3%) and was significantly higher in both foxes (41.2%) and cats (61.5%) than in rodents. This is the first report to quantify seroprevalence to C. burnetii in bank voles, field voles, wood mice, foxes and cats in the UK and provides evidence that predator species could act as indicators for the presence of C. burnetii in rodents. The study demonstrates that wildlife species could be significant reservoirs of infection for both livestock and humans, and the high seroprevalence in domestic cats highlights the potential zoonotic risk from this species.

Seroprevalence of Encephalitozoon cuniculi in wild rodents, foxes and domestic cats in three sites in the United Kingdom.

Encephalitozoon cuniculi is an obligate intracellular microsporidian that is the causal agent of encephalitozoonosis, an important and emerging disease in both humans and animals. Little is known about its occurrence in wildlife. In this study, serum samples from 793 wild rodents [178 bank voles (BV), 312 field voles (FV) and 303 wood mice (WM)], 96 foxes and 27 domestic cats from three study areas in the UK were tested for the presence of antibodies to E. cuniculi using a direct agglutination test (DAT). Seroprevalence in the wild rodents ranged from 1.00% to 10.67% depending on species (overall 5.31%) and was significantly higher in foxes [49.50% (50/96)]. None of the 27 cats sampled were found to be seropositive. This is the first report of seroprevalence to E. cuniculi in BV, FV, WM, foxes and cats in the UK and provides some evidence that foxes could act as sentinels for the presence of E. cuniculi in rodents. The study demonstrates that wildlife species could be significant reservoirs of infection for both domestic animals and humans.

Food and water intake and selective feeding in rabbits on four feeding regimes.

Diet plays an important role in maintaining rabbit health. Feeding an incorrect diet, particularly a low fibre diet, has been linked with digestive, dental and urinary tract disease. However, food intake and dietary requirements have been poorly studied in pet rabbits. This study assessed the effects of two commonly fed pet rabbit diets [extruded nuggets with hay (EH) and muesli with hay (MH)] alongside hay only (HO) and muesli only (MO) on food and water intake in 32 Dutch rabbits. Dry matter (DM) intake was greater in the HO group and lower in the MO group than in the EH and MH groups (p < 0.001). The portion of the diet made of hay was greater in the EH group than in the MH group (p < 0.001). Water intake was positively correlated with DM intake and was greatest in the HO group (p < 0.001). Selective feeding occurred in all rabbit groups fed muesli, whether or not hay was also available. Pellets were rejected, and grains and extrudates selected. The presence of selective feeding in all rabbits fed muesli leads to the consumption of an unbalanced diet. In addition, hay intake and water intake were lower when muesli was fed. Conclusions drawn from this study are based on general recommendations for pet rabbits, and clinical disease may have developed by feeding the study diets over a longer time period, as many diet-related conditions typically present in older rabbits. However, the study demonstrates that the feeding of muesli diets cannot be recommended.

γ-Glutamyl-transferase (GGT) activity in the urine of clinically healthy domestic rabbits (Oryctolagus cuniculus).

Free-catch urine samples were collected from forty-one clinically normal domestic rabbits of various ages, breeds and both sexes. The Test γ GT Liquid-0018257640 was used for the in vitro quantitative determination of γ-Glutamyl-transferase (GGT) and reference intervals for γ-glutamyl transferase (γ-glutamyl transpeptidase, γ-GT, GGT) and GGT index (γ-glutamyl transferase to creatinine ratio) were established in fresh urine samples. Possible correlations of GGT and GGT index with sex and age were also explored. The stability of GGT after storage at +4°C for one week and -20°C for one month was investigated. The GGT and the GGT index reference intervals in fresh urine samples of healthy domestic rabbits were found to be 2.7-96.5 IU/l and 0.043-1.034, respectively. The urine GGT activity and the GGT index did not differ significantly between sexes in fresh urine samples. Nevertheless, a statistically significant difference was found in the GGT index with neutered status. Short-term storage at 4°C did not alter the enzyme stability, whereas, freezing did. Further investigations are needed to determine whether these parameters may be useful for early detection of renal tubular damage in rabbits, and in enabling better clinical management of affected animals.

Epidemiological and postmortem findings in 262 red squirrels (Sciurus vulgaris) in Scotland, 2005 to 2009.

Postmortem and virological examinations for squirrelpox virus (SQPV) were carried out on 262 red squirrels (Sciurus vulgaris) found dead or moribund in Scotland between September 2005 and July 2009, to determine the likely causes of death and highlight factors that might be threats to the red squirrel population. Most of the squirrels were submitted from Dumfries and Galloway, and 71 per cent of them were adults. Road traffic accidents, squirrelpox, trauma or starvation were responsible for death in a large proportion (73 per cent) of the squirrels. Thin or emaciated body condition was associated with deaths resulting from pneumonia SQPV infection and starvation, and with the presence of external parasites. There were differences between age groups with regard to the cause of death; a large proportion of juveniles died of starvation, whereas a large proportion of subadults and adults died in road traffic accidents. SQPV infection was associated with the presence of external parasites, but was not associated with the sex of the animals.

Retrospective detection by negative contrast electron microscopy of faecal viral particles in free-living wild red squirrels (Sciurus vulgaris) with suspected enteropathy in Great Britain.

Transmission electron microscopy identified adenovirus particles in 10 of 70 (14.3 per cent) samples of large intestinal content collected at postmortem examination from free-living wild red squirrels (Sciurus vulgaris) across Great Britain between 2000 and 2009. Examination was limited to cases in which an enteropathy was suspected on the basis of predetermined macroscopic criteria such as semi-solid or diarrhoeic faeces, suspected enteritis or the presence of intussusception. In most cases, meaningful histological examination of enteric tissue was not possible due to pronounced autolysis. Two (2.9 per cent) of the samples were negative for adenovirus but were found to contain rotavirus particles, a novel finding in this species.

Mosaicism in autosomal dominant polycystic kidney disease revealed by genetic testing to enable living related renal transplantation.

Patients with end-stage renal disease (ESRD) secondary to autosomal dominant polycystic kidney disease (ADPKD) receive fewer living-related kidney (LRK) transplants than other groups with ESRD. This relates to the difficulties in excluding the disease in potential donors. We report a case which highlights these difficulties and, by discovery of mosaicism for a new mutation, illustrates the role of clinical and molecular genetic resources in assessing young related kidney donors for patients with ADPKD.

Urodynamic properties and neurotransmitter dependence of urinary bladder contractility in the BK channel deletion model of overactive bladder.

Overactive bladder and incontinence are major medical issues, which lack effective therapy. Previously, we showed (Meredith AL, Thornloe KS, Werner ME, Nelson MT, and Aldrich RW. J Biol Chem 279: 36746-36752, 2004) that the gene mSlo1 encodes large-conductance Ca2+-activated K+ (BK) channels of urinary bladder smooth muscle (UBSM) and that ablation of mSlo1 leads to enhanced myogenic and nerve-mediated contractility and increased urination frequency. Here, we examine the in vivo urodynamic consequences and neurotransmitter dependence in the absence of the BK channel. The sensitivity of contractility to nerve stimulation was greatly enhanced in UBSM strips from Slo-/- mice. The stimulation frequency required to obtain a 50% maximal contraction was 8.3 +/- 0.9 and 19.1 +/- 1.8 Hz in Slo-/- and Slo+/+ mice, respectively. This enhancement is at least partially due to alterations in UBSM excitability, as muscarinic-induced Slo-/- contractility is elevated in the absence of neuronal activity. Muscarinic-induced Slo-/- contractility was mimicked by blocking BK channels with iberiotoxin (IBTX) in Slo+/+ strips, whereas IBTX had no effect on Slo-/- strips. IBTX also enhanced purinergic contractions of Slo+/+ UBSM but was without effect on purinergic contractions of Slo-/- strips. In vivo bladder pressure and urine output measurements (cystometry) were performed on conscious, freely moving mice. Slo-/- mice exhibited increased bladder pressures, pronounced pressure oscillations, and urine dripping. Our results indicate that the BK channel in UBSM has a very significant role in urinary function and dysfunction and as such likely represents an important therapeutic target.

Sertoli cell tumour in an Amur tiger.

The histological and immunohistochemical characteristics of a malignant Sertoli cell tumour in a 17-year-old Amur tiger (Panthera tigris altaica) are described. Histological examination of the primary lesion in the right testis and metastatic lesions throughout the internal organs showed a variable cellular pattern with an admixture of tubular structures divided by fine stroma filled with fusiform to stellate cells, and sheets of polygonal cells with abundant vacuolated cytoplasm. Immunohistochemical techniques demonstrated strong positive staining for neuron-specific enolase and variable positive staining for vimentin in neoplastic cells, supporting a diagnosis of a tumour of Sertoli cell origin.

Age-associated increase in rat ventricular ANP gene expression correlates with cardiac hypertrophy.

Atrial natriuretic peptide (ANP), a cardiac-specific hormone, is stored in the atria and released in response to atrial stretch. During cardiac hypertrophy, ANP gene expression is markedly upregulated in the left ventricle (LV). Because the hearts of normotensive senescent rats exhibit left atrial (LA) and left ventricular (LV) hypertrophy and dilatation, we examined ANP mRNA levels by Northern blot analysis and ANP peptide concentrations by radioimmunoassay in atria, LVs, and plasma of rats at 2, 6, 18, and 22-24 mo of age. Compared with LVs of 6-mo-old rats, the LV-to-body weight ratio was elevated 30% by 18 mo of age, whereas levels of ANP mRNA were elevated twofold (not significant) and sevenfold (P < 0.05) in the LV of 18- and 22- to 24-mo-old rats, respectively. The concentration of immunoreactive ANP (ir-ANP) exhibited a four- to fivefold increase in LVs of 18- and 22- to 24-mo-old rats compared with values for 6-mo-old rats (43 +/- 4 pmol/g wet wt; means +/- SE). Among 18-and 22- to 24-mo-old rats a significant correlation was observed between ANP peptide concentration and LV hypertrophy (r 2 = 0.64). Levels of ANP mRNA and ir-ANP in the atria exhibited only modest changes with aging.(ABSTRACT TRUNCATED AT 250 WORDS)

Cystic fibrosis mutation analysis: report from 22 U.K. regional genetics laboratories.

We have collated the results of cystic fibrosis (CF) mutation analysis conducted in 22 laboratories in the United Kingdom. A total of 9,807 CF chromosomes have been analysed, demonstrating 56 different mutations so far observed and accounting for 86% of CF genes in the native Caucasian population of the United Kingdom. delta F508 is the most common at 75.3% of CF mutations (range 56.5-83.7%), followed by G551D (3.08%; range 0.71-7.60%), G542X (1.68%; range 0.85-3.66%), 621 + 1 (G > T) (0.93%; range 0.41-3.16%), 1717-1(G > A) (0.57%; range 0.17-1.14%), 1898 + 1)(G > A) (0.46%), R117H (0.46%), N1303K (0.46%), and R553X (0.46%). The data show a clear geographical variation in the distribution of some of the mutations, most notably a marked regional variation in the distribution of 621 + 1 (G > T) and 1989 + 1(G > A), which are both apparently more frequent in Wales. R560T and R117H appear to be more frequent in Ireland and Scotland, and G551D more frequent in Scotland. In summary, these data illustrate that the mutations present within a particular population need to be defined in order to provide meaningful carrier screening and testing for rare mutations in affected individuals. Furthermore, it is apparent that the ethnic origin of a patient, even within a small country such as the United Kingdom, should be taken into account.

Alterations in cardiac gene expression during the transition from stable hypertrophy to heart failure. Marked upregulation of genes encoding extracellular matrix components.

The failing heart is characterized by impaired cardiac muscle function and increased interstitial fibrosis. Our purpose was to determine whether the functional impairment of the failing heart is associated with changes in levels of mRNA encoding proteins that modulate parameters of contraction and relaxation and whether the increased fibrosis observed in the failing heart is related to elevated expression of genes encoding extracellular matrix components. We studied hearts of 18- to 24-month-old spontaneously hypertensive rats with signs and symptoms of heart failure (SHR-F) or without evidence of failure (SHR-NF) and of age-matched normotensive Wistar-Kyoto (WKY) rats. Compared with WKY rats, SHR-NF exhibited left ventricular (LV) hypertrophy (2.2-fold) and right ventricular (RV) hypertrophy (1.5-fold), whereas SHR-F were characterized by comparable LV hypertrophy (2.1-fold) and augmented RV hypertrophy (2.4-fold; all P < .01). Total RNA was isolated from ventricles and subjected to Northern blot analysis. In SHR-F hearts, the level of alpha-myosin heavy chain mRNA was decreased in both ventricles to 1/3 and 1/5 of the SHR-NF and WKY values, respectively (both P < .01). Levels of beta-myosin heavy chain, alpha-cardiac actin, and myosin light chain-2 mRNAs were not significantly altered in hearts of SHR-NF or SHR-F. Levels of alpha-skeletal actin were twofold greater in SHR-NF hearts compared with WKY hearts and were intermediate in SHR-F hearts. Levels of atrial natriuretic factor (ANF) mRNA were elevated threefold in the LV of SHR-NF (P < .05) but were not significantly increased in the RV of SHR-NF compared with WKY rats. During the transition to failure (SHR-F versus SHR-NF), ANF mRNA levels increased an additional 1.6-fold in the LV and were elevated 4.7-fold in the RV (both P < .05). Levels of sarcoplasmic reticulum Ca(2+)-ATPase (SRCA) mRNA were maintained in the LV of hypertensive and failing hearts at levels not significantly different from WKY values. In contrast, the level of RV SRCA mRNA was 24% less in SHR-NF compared with WKY rats, and during the transition to failure, this difference was not significantly exacerbated (29% less than the WKY value). The levels of fibronectin and pro-alpha 1(I) and pro-alpha 1(III) collagen mRNAs were not significantly elevated in either ventricle of the SHR-NF group but were fourfold to fivefold higher in both ventricles of SHR-F (all P < .05).(ABSTRACT TRUNCATED AT 400 WORDS)

Direct sequencing of the complete CFTR gene: the molecular characterisation of 99.5% of CF chromosomes in Wales.

We have performed an extensive mutation analysis on 184 CF families in Wales. In our previous study, mutations on 329/369 CF chromosomes were identified after screening for delta F508 and sixteen other mutations. To identify the mutations on the remaining 40 uncharacterized CF chromosomes, we have carried out direct DNA sequencing over the complete coding region, intron splice sites, and part of the promoter region of the CFTR gene. During this study we have designed a set of internal sequencing primers which allow clear sequencing through the aforementioned regions. Sequence analysis revealed 15 further mutations (4 of which are novel), and 10 previously described polymorphisms. In total, we have identified 29 mutations, the distribution of which provides further insight into the functional domains of the CFTR protein. We have characterised 99.5% of the CF chromosomes (365/367, one sample degraded). In order to ascertain accurate frequency data for the Welsh population, CF families with at least 3 'Welsh' grandparents were strictly regarded as 'Welsh'. Of these 91 families, delta F508 accounts for 71.6%, 621 + 1G-->T 6.6% and 1898 + 1G-->A 5.5%. The implications for CF population screening in Wales are discussed.