Evaluation of an Electrolyte Analyser for Measurement of Concentrations of Ionized Calcium and Magnesium in Cats

The goal of this study was to evaluate the Nova CRT 8 electrolyte analyser for determination of concentrations of ionized calcium (Cai) and magnesium (Mgi) in cats, todetermine the effects of sample handling and storage and to establish reference ranges. The precision and analytical accuracy of the Nova CRT 8 analyser were good. The concentrations of Cai and Mgi were significantly lower in aerobically handled serum samples than in those handled anaerobically. The concentrations of Cai and Mgi differed significantly among whole blood, plasma and serum. In anaerobically handled serum, the concentration of Cai was stable for 8 h at 22°C, for 5 days at 4°C and for 1 week at −20°C. The concentration of Mgi was stable for 4 h at 22°C but for less than 24 h at 4°C and for less than 1 week at −20°C. In serum from 36 cats, the reference ranges were 1.20-1.35 mmol/L for Cai and 0.47-0.59 mmol/L for Mgi. The Nova CRT 8 electrolyte analyser is suitable for determination of Cai and Mgi concentrations in cats. Anaerobically handled serum samples are recommended and, stored at room temperature, they yield accurate results when analysed within 4

The fecal microbiome in dogs with acute diarrhea and idiopathic inflammatory bowel disease.

Recent molecular studies have revealed a highly complex bacterial assembly in the canine intestinal tract. There is mounting evidence that microbes play an important role in the pathogenesis of acute and chronic enteropathies of dogs, including idiopathic inflammatory bowel disease (IBD). The aim of this study was to characterize the bacterial microbiota in dogs with various gastrointestinal disorders. Fecal samples from healthy dogs (n = 32), dogs with acute non-hemorrhagic diarrhea (NHD; n = 12), dogs with acute hemorrhagic diarrhea (AHD; n = 13), and dogs with active (n = 9) and therapeutically controlled idiopathic IBD (n = 10) were analyzed by 454-pyrosequencing of the 16S rRNA gene and qPCR assays. Dogs with acute diarrhea, especially those with AHD, had the most profound alterations in their microbiome, as significant separations were observed on PCoA plots of unweighted Unifrac distances. Dogs with AHD had significant decreases in Blautia, Ruminococcaceae including Faecalibacterium, and Turicibacter spp., and significant increases in genus Sutterella and Clostridium perfringens when compared to healthy dogs. No significant separation on PCoA plots was observed for the dogs with IBD. Faecalibacterium spp. and Fusobacteria were, however, decreased in the dogs with clinically active IBD, but increased during time periods of clinically insignificant IBD, as defined by a clinical IBD activity index (CIBDAI). Results of this study revealed a bacterial dysbiosis in fecal samples of dogs with various GI disorders. The observed changes in the microbiome differed between acute and chronic disease states. The bacterial groups that were commonly decreased during diarrhea are considered to be important short-chain fatty acid producers and may be important for canine intestinal health. Future studies should correlate these observed phylogenetic differences with functional changes in the intestinal microbiome of dogs with defined disease phenotypes

Domänenarchitektur inhibitorischer Ionenkanäle der Cys-Loop-Rezeptorfamilie

The spontaneous mouse mutant Oscillator carries a mutation in the Glra1 gene and shows a phenotype similar to human patients suffering from hyperekplexia. Thus, Oscillator is an excellent model to study neuromotor disorders, which are associated with glycine receptor (GlyR) mutations. Based on the Oscillator protein, a truncated GlyRα1 subunit was generated in vitro, which lacks most of the TM3-4 loop, the TM4 and the C-terminus. If the missing C-terminal complementation construct was cotransfected in HEK293-cells together with the truncated α1 variant, GlyR-domains were able to assemble into a functional ion channel. To investigate the underlying mechanism the N-terminus of the complementation construct was successive shortened. A functional complementation of GlyR-domains was observed up to a deletion of 49 amino acids from the N-terminus of the complementation construct in contrast to larger truncations. A wrong orientation into the plasma membrane as well as a defect in oligomerisation was excluded as a reason for the loss of function. Hence, the truncations led to a deletion of a specific interaction motif or reduced the intracellular domain until no steric contact was possible between the domains. A deletion of the negative charged amino acids of the complementation construct resulted most probably in a disappearance of functional ion channels. Therefore, these negative charges probably form an ionic interaction with the basic “RRKRR” motif at the end of the truncated α1-variant. Neither any functional receptor was observed if both termini carried positive charges. Nevertheless, a minimum length of the intracellular domain seemed to be required, because the addition of negative charges to the most truncated tails did not lead to a recovery of GlyR function. The reconstitution was, however, successful using the TM3-4 loop as a third independent construct together with the truncated α1-variant and the C-terminal tail. Thus, the loop represents an important factor in the process of conformational rearrangements following ligand binding, which results in channel opening. Beside the direct interaction of the GlyR domains via ionic interaction, an indirect binding mediated by unknown proteins is also possible. The next step was to bring these complementation experiments in an in vivo situation. Therefore, it was intended to rescue the neurological phenotype of Oscillator mice using a transgene consisting of the missing C-terminal GlyR part. However, the transgene was not expressed on the protein level, which was shown with various methods. The topology of the cys-loop receptor superfamily is very similar. For that reason it was investigated on the basis of the α1β2γ2 GABAA receptor if the mechanism of a complementation is transferable to other family members. In patients of a particular form of epilepsy (GEFS+) a mutation was identified, which lead to a truncated γ2-subunit comparable to the Oscillator-mutation. This truncated γ2-trc variant and an appropriate complementation construct (γ2-iD-TM4) were cloned. The expression of all GABA receptor domains were demonstrated in HEK293-cells, but the γ2-trc and γ2-iD-TM4 were not transported to the plasma membrane. Thus, no functional α1/β2/γ2-trc/γ2_iD-TM4 receptor complexes were formed.Die spontane Mausmutante Oscillator trägt eine Mutation im Glra1-Gen und eignet sich hervorragend zur Untersuchung von motorischen Bewegungsstörungen, die mit Glycinrezeptor- (GlyR) mutationen assoziiert sind, da die Mäuse ähnliche phänotypische Symptome aufweisen wie Hyperekplexie-Patienten. In Anlehnung an die Oscillator-Proteinvariante wurde eine verkürzte GlyRα1 Untereinheit in vitro generiert, der ein Großteil der TM3-4 Schleife, die TM4 sowie der C-Terminus fehlte. Wurde dieser fehlende C-terminale Bereich als ein unabhängiges Komplementationskonstrukt in HEK293-Zellen mit der verkürzten α1 Variante kotransfiziert, waren die GlyR-Domänen in der Lage, sich zu einem funktionellen Ionenkanal zu assemblieren. Um den zugrunde liegenden Mechanismus zu untersuchen, wurde das Komplementationskonstrukt am intrazellulären N-Terminus sukzessive verkürzt. Bei Verwendung der bis zu 49 Aminosäuren verkürzten Konstrukte konnte, im Gegensatz zu stärker verkürzten Konstrukten, eine funktionelle Komplementation beobachtet werden. Als Ursachen für das Abbrechen der Funktionalität wurden sowohl eine falsche Orientierung der Konstrukte in der Plasmamembran als auch eine fehlerhafte Pentamerisierung experimentell ausgeschlossen. So mussten die Trunkierungen entweder ein bestimmtes Interaktionsmotiv deletieren oder die intrazelluläre Domäne so stark verkleinern, dass kein sterischer Kontakt zwischen den Domänen mehr möglich war. Vermutlich bilden die negativ geladenen Aminosäuren des Komplementationskonstruktes eine ionische Wechselwirkung mit dem basischen Motiv „RRKRR“ am Ende der verkürzten α1-Variante aus, da die Funktionalität mit der Deletion dieser negativen Ladungen abbrach. Weiterhin wurden ebenfalls keine funktionellen Kanäle komplementiert, sobald beide Termini positive Ladungen trugen. Dennoch schien zusätzlich eine Mindestlänge der intrazellulären Domäne erforderlich zu sein, da auch das Anbringen von negativen Aminosäuren an die stark verkürzten Konstrukte nicht zu einer Wiederherstellung der GlyR Funktion führte. Dies gelang hingegen durch Zugabe der TM3-4 Schleife, als eine dritte, unabhängige Domäne. Diese fungierte vermutlich als Brücke zwischen den beiden weiteren Domänen, um die durch Ligandenbindung ausgelöste Konformationsänderung bis zur TM3 auf die TM4 und den C-Terminus zu übertragen und so eine Kanalöffnung zu ermöglichen. Neben einer direkten Interaktion der GlyR Domänen mittels ionischer Wechselwirkungen ist auch eine indirekte Interaktion vorstellbar, die über noch unbekannte Proteine vermittelt wurde. Der nächste Schritt bestand in der Überführung der Komplementationsexperimente in eine in vivo-Situation. Hierfür sollte der neurologische Phänotyp der Oscillator-Mäuse durch Einbringen der fehlenden C-terminalen Domäne in Form eines Transgens aufgehoben werden. Jedoch wurde durch eine Vielzahl von Methoden gezeigt, dass das generierte Transgen nicht auf Proteinebene exprimiert war. Da die Topologie in der gesamten Superfamilie der Cys-Loop-Rezeptoren sehr ähnlich ist, sollte anhand der α1β2γ2-Isoform des GABAA-Rezeptors untersucht werden, ob der Mechanismus einer Komplementation auch auf weitere Subfamilien übertragbar ist. In Epilepsie-Patienten (GEFS+) wurde eine Mutation identifiziert, die zu einer verkürzten γ2-UE führt, vergleichbar der Oscillator-Mutation. Diese verkürzte γ2-trc Variante und ein entsprechendes Komplementationskonstrukt (γ2_iD TM4) wurden kloniert. Die Expression aller GABAAR-Domänen wurde in HEK293-Zellen nachgewiesen, jedoch wurde γ2-trc, auch in Kombination mit dem Komplementationskonstrukt, nicht an die Plasmamembran transportiert. Folglich wurden keine funktionellen α1/β2/γ2-trc/γ2_iD TM4 Rezeptorkomplexe ausgebildet

What Influence Does Residual Magnetism Have on the Transformer Core?

Whenever a power or distribution transformer is isolated from the power system, it is very likely that residual magnetism remains in the core. Residual magnetism also occurs when performing winding resistance test which is also a routine test of the transformer manufacturers and onsite test. This paper discusses the influence of residual magnetism on some diagnostic measurement methods and on the inrush current. It also describes how to overcome the difculties of demagnetisation onsite with a mobile test equipment

Utility of diagnostic tests in vomiting dogs presented to an internal medicine emergency service

IntroductionVomiting is a common sign in dogs presenting to emergency services. It can be self-limiting, a sign of a life-threatening extraintestinal, or intestinal disorder. Reasonable diagnostics should be performed to determine the underlying cause. This study aimed to assess the utility of diagnostic tests in vomiting dogs, and its correlation with patient history, and physical examination results. Additionally, parameters to differentiate uncomplicated vomiting from complicated vomiting were investigated.MethodsIn this prospective, observational, clinical study, data from 99 client-owned dogs with vomiting, presenting as first opinion cases, were evaluated. History, physical examination, duration of clinical signs, overall number of episodes of vomiting, appetite, and additional clinical signs were recorded. The standardized diagnostic evaluation of all patients included venous blood gas analysis, complete blood count, serum biochemistry profile, canine pancreatic lipase, abdominal radiographs, ultrasound, and urinalysis. Follow-up was performed 4–5 days later. Based on severity of disease and clinical course, dogs were categorized to “uncomplicated vomiting” (UN), or “complicated vomiting” (COM). The utility of each test for diagnosing the cause of vomiting was evaluated. Spearman correlation coefficient, Chi-squared-, unpaired t-, and Mann–Whitney U-test were used. Statistical significance was defined as p ≤ 0.05.ResultsOut of the 99 dogs, 34 had uncomplicated courses of disease (UN). In 60/99 cases, a diagnosis was obtained, and in 39/99 cases, the cause for vomiting remained unknown. Longer duration of clinical signs, and reduced appetite were associated with higher utility of abdominal ultrasound. A poor mentation was associated with a higher utility of blood examinations and abdominal radiographs. Dogs presenting with an impaired mentation or with additional clinical signs other than diarrhea, were more likely to be in the COM group.DiscussionBased on this investigation, general recommendations concerning the diagnostic approach for patients with vomiting could not be provided. For dogs who have exclusively vomiting as a clinical sign, and present in good mentation, further investigations might not be beneficial, and these dogs may recover with symptomatic treatment alone. Additional diagnostics could be indicated in dogs with additional clinical signs other than diarrhea

C-reactive protein as a tool for monitoring response to treatment in dogs with acute hemorrhagic diarrhea syndrome

ObjectivesC-reactive protein (CRP) is an established marker for systemic inflammation in dogs that is especially elevated in dogs with sepsis. Some dogs with acute hemorrhagic diarrhea syndrome (AHDS) develop bacterial translocation and consequent sepsis during hospitalization. This study aimed to evaluate the course of CRP plasma concentrations during hospitalization and its correlation with clinical and other laboratory variables in dogs with AHDS.MethodsIn this prospective, observational study, CRP was evaluated on days 0, 1, 2, and 3 in 27 client-owned dogs who presented with AHDS. Clinical examination data, blood pressure, acute patient physiologic and laboratory evaluation (APPLE) full and APPLE fast scores, and canine hemorrhagic diarrhea severity (CHDS) index were measured on the same days to evaluate the severity of the disease.ResultsTwenty-five of the 27 dogs were discharged from hospital. Nineteen dogs received antimicrobial treatment due to sepsis or neutropenia. CRP values were mildly elevated on day 0 (median 27.3 mg/L; 1.0–125.8 mg/L) and markedly elevated on day 1 (median 88.9 mg/L; 1.4–192.7 mg/L). CRP concentrations decreased gradually over the following days. Moreover, CRP concentrations correlated moderately with albumin, leucocyte count, neutrophil count, and APPLE full and fast scores, but not with antimicrobial treatment.Conclusion and relevanceCRP concentrations were significantly elevated in patients with AHDS. In this study population, CRP did not help in detecting the requirement of antimicrobial treatment in dogs with AHDS. Nevertheless, as CRP can monitor the response to treatment, regular analysis can guide treatment

Effects of dietary cellulose on clinical and gut microbiota recovery in dogs with uncomplicated acute diarrhea: a randomized prospective clinical trial

OBJECTIVE: To assess the impact of dietary fiber supplementation with cellulose on clinical course, fecal consistency, and intestinal microbiota composition in dogs with uncomplicated acute diarrhea (AD). METHODS: From September 2022 to November 2023, a total of 19 dogs presenting with uncomplicated AD were included in this prospective, randomized, and double-blinded clinical trial. The time to resolution of diarrhea was evaluated via owner surveys and a fecal scoring chart. The client-owned dogs were randomly assigned to a cellulose group (CG) or control group. The intestinal microbiota was analyzed via quantitative PCR. RESULTS: A marginally significant, faster improvement in stool consistency on day 1 was observed in the CG (P = .09). All dogs improved clinically, with a median recovery time of 3.0 days in the CG and 3.2 days in the control group (range, 1 to 6 days in both groups). There was no significant difference regarding the Canine Acute Diarrhea Severity index or composition of the intestinal microbiota during the study. CONCLUSIONS: All dogs with uncomplicated AD exhibited rapid clinical improvement and recovery of the core intestinal microbiota within the first few days. Cellulose improved the fecal consistency in a subset of dogs, and intestinal dysbiosis was mild and self-limiting. CLINICAL RELEVANCE: The administration of dietary cellulose has the potential to accelerate improvements of stool consistency. Mild changes in pathobionts, such as an increased amount of Clostridium perfringens, are self-limiting; thus, antibiotic intervention is not warranted

New hyperekplexia mutations provide insight into glycine receptor assembly, trafficking, and activation mechanisms

Background: Hyperekplexia mutations have provided much information about glycine receptor structure and function. Results: Weidentified and characterized nine new mutations. Dominant mutations resulted in spontaneous activation, whereas recessive mutations precluded surface expression. Conclusion: These data provide insight into glycine receptor activation mechanisms and surface expression determinants. Significance: The results enhance our understanding of hyperekplexia pathology and glycine receptor structure-function. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc. Published in the U.S.A