Antioxidant prevention of Heinz body formation and oxidative injury in cats

OBJECTIVE: To determine the effectiveness of 3 antioxidants in preventing Heinz body anemia in cats. DESIGN: Prospective study. ANIMALS: 44 specific-pathogen-free healthy cats. PROCEDURE: Cats were housed individually, divided randomly into 4 groups, and given the following orally every 12 hours: empty gelcaps (control cats), N-acetylcysteine (NAC, 100 mg/kg of body weight), vitamin E (d,l-alpha-tocopherol; 400 IU), or ascorbate (250 mg). After 2 weeks, Heinz bodies were induced by dietary onion powder (OP; 1% or 3% of dry matter) or propylene glycol (PG, 8% wt/vol in drinking water) for an additional 3 weeks. Intake of treated water or food was recorded daily. Body weight, PCV, Heinz body and reticulocyte percentages, reduced glutathione concentration, and total antioxidant status were measured twice weekly in all cats. RESULTS: Heinz body percentage and degree of anemia did not differ significantly among cats receiving antioxidants and control cats except in cats that ingested water containing PG, in which antioxidant supplementation was associated with a decrease in water intake. Of cats that were fed a diet that contained OP, cats that received NAC had significantly higher reduced glutathione concentrations, compared with other cats in the experiment. Total antioxidant status did not consistently correlate with antioxidant supplementation or type of oxidant administered (ie, OP or PG). CONCLUSIONS AND CLINICAL RELEVANCE: Although the effect of antioxidant supplementation on Heinz body anemia in cats was minimal, antioxidants may have subclinical biochemical effects such as GSH sparing that may be important against milder forms of oxidative stress.     

Liver sonography and ultrasound-guided puncture of the gallbladder in pigs: new possibilities for the diagnosis of mycotoxicosis

60 pigs representing all age groups (suckling pigs, weaner pigs, hogs, gilts and sows; thereof 37 females, 2 males, 21 castrated males) were examined by ultrasound of the liver and by ultrasound guided gallbladder puncture. The visibility of the liver and gallbladder was strongly influenced by the size of the animals. The thickness of the abdominal and thoracic walls in older animals proved to be a highly ultrasound-absorptive medium, which limited the ability to assess the underlying tissue structures. As a result, gallbladder puncture of these animals was possible only with a certain degree of technical difficulty. The gallbladder puncture procedure itself posed little risk to the animal. The primary risk resulted from the general anesthesia required. Autopsy showed no pathological findings due to gallbladder puncture with the exception of minimal, rapidly healing, local infectious processes. The bile proved to be a suitable medium of the detection of mycotoxins and their metabolites. Selective accumulation of these toxins in bile provide a more reliable diagnostic tool than the standard mycotoxicological tests of feed.     

Studies on the antibody response of Lama glama--evaluation of the binding capacity of different IgG subtypes in ELISAs for clenbuterol and BSA

Camelidae are known to produce three subtypes of immunoglobulin G (IgG), two of which are devoid of light chains. Two llamas (Lama glama) were immunised against clenbuterol-bovine serum albumin (BSA). Enzyme-linked immunosorbent assays (ELISAs) for clenbuterol and BSA on the basis of protein A-coated microtitration plates were established to investigate the titre development. Three subclasses of IgG (IgG(1): 29+66KDD, IgG(2): 52KDD, IgG(3): 56KDD) depending on their different binding properties to protein A and protein G could be separated chromatographically. Only IgG(1), which consists of conventional four-chain antibodies, bound to clenbuterol, whereas all forms of heavy-chain antibodies merely bound BSA.     

Immune cell populations in the duodenal mucosa of cats with inflammatory bowel disease

The aim of this study was to characterize the phenotype of leukocytes infiltrating the duodenal mucosa of cats with inflammatory bowel disease (IBD) by using immunohistochemistry and computer-aided morphometry to assess whether immunologic markers would aid in characterization of IBD. Frozen and formalin-fixed duodenal biopsies were collected from cats referred for investigation of chronic vomiting, diarrhea, or both (n = 34). Reference ranges were previously established by using duodenal samples from healthy cats (n = 16). No significant difference was found in the number of immunoglobulin G+ (IgG+) or IgA+ in either the villous lamina propria or the crypt lamina propria between cats with IBD and control cats. T cells (CD3+) increased in number from crypt to the tip of the villi in biopsies from both diseased (mean +/- SD for each group was 18.8 +/- 6.6 and 17.7 +/- 4.2 cells/ 10,000 m2 in cryptal areas to 25.2 +/- 9.5 and 29.1 +/- 13.3 cells/10,000m2 in villous areas) and healthy animals (17.9 +/- 3.9 cells/10,000 microm2 in cryptal areas to 24.1 +/- 9.3 cells/10,000 microm2 in villous areas) and no significant difference was found between diseased and control cats. By contrast, major histocompatibility complex (MHC) class II expression by leukocytes with dendritic cell or macrophage morphology in the lamina propria was significantly greater in cats with IBD (13.3 +/- 4.2 cells/10,000 microm2 in cryptal area; P = .016) than in healthy cats (11.9 +/- 3.0 cells/10,000 microm2) and MHC class II expression by enterocytes also was more pronounced in these cats showing an overall intensity of expression of 7.1 +/- 4.0 cells/10,000 microm2 in cats with IBD as opposed to 0.0 +/- 0.0 cells/10,000 microm2 to 0.3 +/- 0.7 cells/10,000 microm2 in healthy cats. These findings suggest that a subtle immunologic dysregulation occurs in spontaneously arising feline IBD.     

A One-Step, Immunochromatographic Lateral Flow Device Specific to Rhizoctonia solani and Certain Related Species, and Its Use to Detect and Quantify R. solani in Soil

ABSTRACT A murine hybridoma cell line GD2 secreting an immunoglobulin (Ig)M monoclonal antibody (MAb) was produced against surface antigens from an anastomosis group (AG) 4 isolate of Rhizoctonia solani (teleomorph: Thanatephorus cucumeris). Ascites were produced in mice using GD2 hybridoma cells and used to develop a rapid immunochromatographic lateral flow device (LFD) for the detection of antigens from R. solani and certain related Rhizoctonia spp. The LFD was tested for specificity against surface antigens from related and unrelated soil fungi. Antigens from representative isolates of R. solani AGs 1, 2-1, 2-3, 2-t, 3, 4, 5, 6, 7, 8, 9, 10, 11, and BI gave a positive response in LFD tests, as did antigens from Thanatephorus orchidicola, T. praticola, R. fragariae (teleomorph: Ceratorhiza fragariae), Ceratorhiza goodyerae-repentis, Ceratobasidium cornigerum, and binucleate AGE. Antigens from R. solani AGs 2-2, 2-2IIIB, and 2-2IV and from the related fungi R. carotae, R. cerealis (teleomorph: Ceratobasium cereale), R. crocorum (teleomorph: Helicobasidium brebissonii), R. oryzae (teleomorph Waitea circinata), and R. zeae gave negative responses, as did antigens from a range of unrelated fungi and oomycetes including Fusarium, Gliocladium, Trichoderma, Pythium, and Phytophthora spp. The usefulness of the LFD to detect R. solani was demonstrated in soils naturally infested with R. solani AG3. There was close agreement between results of LFD tests and conventional plate enrichment tests employing selective medium. The specificity of the technique was confirmed by polymerase chain reaction PCR using R. solani AG3-specific primers and by analyses based on sequences of the internal transcribed spacer (ITS)1-5.8S-ITS2 rRNA-encoding regions of unrelated fungi recovered from soil samples. The LFD was used to quantify R. solani AG4 in artificially infested soil samples (chopped potato soil inoculum). Estimates of CFU per gram of soil were derived using a most-probable number technique, which was based on the presence or absence of a detectable signal in the LFD. Estimates of CFU obtained in LFD tests and those obtained in a plate-trapped antigen enzyme-linked immunosorbent assay incorporating MAb GD2 were identical (449 CFU g(-1) of soil).     

Ellipsomyxa gobii (Myxozoa: Ceratomyxidae) in the common goby Pomatoschistus microps (Teleostei: Gobiidae) uses Nereis spp. (Annelida: Polychaeta) as invertebrate hosts

Nereis diversicolor O.F. Müller and N. succinea Frey et Leuckart (Polychaeta, Nereidae) living in brackish shallow areas in Denmark are naturally infected with tetractinomyxon actinospores. Infected Nereis spp. were experimentally fed to various potential fish hosts, and the actinosporean stages developed into myxosporean stages of Ellipsomyxa gobii K?ie, 2003 (Ceratomyxidae) in the gallbladder of the common goby Pomatoschistus microps (Kr?yer) (Gobiidae). The European eel Anguilla anguilla (L.), three-spined stickleback Gasterosteus aculeatus L., small sand eel Ammodytes tobianus L., flounder Platichthys flesus (L.), European plaice Pleuronectes platessa L. and common sole Solea solea (L.) did not become experimentally infected. In Danish shallow brackish areas P. microps is naturally infected with E. gobii, in some areas with a prevalence >90%. We compared small subunit ribosomal DNA sequences of the actinosporean with E. gobii from P. microps. Sequences were identical, which further verifies that both forms belong to the same organism. This is the first myxozoan two-host life cycle in the marine environment.     

Internal Transcribed Spacer Sequence-Based Phylogeny and Polymerase Chain Reaction-Restriction Fragment Length Polymorphism Differentiation of Tilletia walkeri and T. indica

ABSTRACT A polymerase chain reaction-restriction fragment length polymorphism assay to distinguish Tilleita walkeri, a rye grass bunt fungus that occurs in the southeastern United States and Oregon, from T. indica, the Karnal bunt fungus, is described. The internal transcribed spacer (ITS) region of the ribosomal DNA repeat unit was amplified and sequenced for isolates of T. indica, T. walkeri, T. horrida, and a number of other taxa in the genus Tilletia. A unique restriction digest site in the ITS1 region of T. walkeri was identified that distinguishes it from the other taxa in the genus. Phylogenetic analysis of the taxa based on ITS sequence data revealed a close relationship between T. indica and T. walkeri, but more distant relationships between these two species and other morphologically similar taxa.     

A combination of different polymerase chain reaction (PCR) assays for the presumptive identification of Yersinia pestis

A combination of four polymerase chain reaction (PCR) assays targeting the Yersinia pestis-specific plasmoidal genes of the fraction 1 capsular antigen and plasminogen activator/coagulase, the gene of the V antigen of the Yersinia virulence plasmid, and the chromosomal 16S rRNA gene was evaluated for the identification of Y. pestis isolates. All four assays were subjected to the same sample preparation technique, reagents and cycling conditions. Eighteen Y. pestis, 66 Y. pseudotuberculosis, 40 Y. enterocolitica strains, the type strains of the other Yersinia species, and 20 other pathogenic bacterial strains were investigated. By using the proposed combination of PCR assays all Y. pestis strains were identified correctly. The applicability of this combination of PCR assays was demonstrated by the detection of Y. pestis DNA in spiked tissues from Rattus norwegicus and fleas (Xenopsylla cheopis and Ctenocephalides spp.). As little as 60 genome equivalents were detected. This system is applicable for monitoring Y. pestis and its vectors in enzootic natural foci and in the diagnosis of plague in humans and animals.     

Occurrence of zearalenone, alpha- and beta-zearalenol in bile of breeding sows in relation to reproductive performance

Although zearalenone-induced reproductive disorders and the clinical appearance of hyperestrogenism were reproduced and documented quite often the role of zearalenone-contaminated fodder as a cause for fertility problems in sow breeding is still discussed controversial. Therefore the correlation of zearalenone and zearalenone-derivatives in bile (n = 794) and feed (n = 158) with fertility problems of unknown origin was investigated in this study. For the analysis of zearalenone and its derivatives in bile a HPLC/EIA combination was used. On the one hand, this procedure guaranteed the quantitatively reproducible detection, on the other hand, the investigation expenditure could be kept small with regard to a later effort in the routine diagnostics. The detection limits for zearalenone, alpha- and beta-zearalenol in bile were at 1.0 ng/ml, 1.0 ng/ml, and 3.0 ng/ml, respectively. Results were confirmed by GC-MS. Zearalenone and zearalenone-derivatives were detected in almost every bile analysed. The contamination rate was 96.2%. In opposition to recent investigations beta-zearalenol was perceived as a relevant metabolite in swine. The contamination rate of feeding stuffs was 25.9%. Incubation of samples with beta-glucosidase did not elevate the detected amounts of zearalenone. As the measurable concentrations in bile and fodder were only slightly correlated the analysis of bile represents a reasonable alternative for fodder investigation. However, a correlation between the occurrence of zearalenone, alpha- and beta-zearalenol in bile of sows and non-infectious reproductive disorders could not be established at the loading level found. These results are in line with those statements obtained in feeding experiments regarding the risk evaluation of zearalenone in sow reproduction.