Biofilm bacteria: formation and comparative susceptibility to antibiotics

The Calgary Biofilm Device (CBD) was used to form bacterial biofilms of selected veterinary gram-negative and gram-positive pathogenic bacteria from cattle, sheep, pigs, chicken, and turkeys. The minimum inhibitory concentration (MIC) and minimum biofilm eradication concentration (MBEC) of ampicillin, ceftiofur, cloxacillin, oxytetracycline, penicillin G, streptomycin, tetracycline, enrofloxacin, erythromycin, gentamicin, tilmicosin, and trimethoprim-sulfadoxine for gram-positive and -negative bacteria were determined. Bacterial biofilms were readily formed on the CBD under selected conditions. The biofilms consisted of microcolonies encased in extracellular polysaccharide material. Biofilms composed of Arcanobacterium (Actinomyces) pyogenes, Staphylococcus aureus, Staphylococcus hyicus, Streptococcus agalactiae, Corynebacterium renale, or Corynebacterium pseudotuberculosis were not killed by the antibiotics tested but as planktonic bacteria they were sensitive at low concentrations. Biofilm and planktonic Streptococcus dysgalactiae and Streptococcus suis were sensitive to penicillin, ceftiofur, cloxacillin, ampicillin, and oxytetracycline. Planktonic Escherichia coli were sensitive to enrofloxacin, gentamicin, oxytetracycline and trimethoprim/ sulfadoxine. Enrofloxacin and gentamicin were the most effective antibiotics against E. coli growing as a biofilm. Salmonella spp. and Pseudomonas aeruginosa isolates growing as planktonic populations were sensitive to enrofloxacin, gentamicin, ampicillin, oxytetracycline, and trimethoprim/sulfadoxine, but as a biofilm, these bacteria were only sensitive to enrofloxacin. Planktonic and biofilm Pasteurella multocida and Mannheimia haemolytica had similar antibiotic sensitivity profiles and were sensitive to most of the antibiotics tested. The CBD provides a valuable new technology that can be used to select antibiotics that are able to kill bacteria growing as biofilms.     

Sensitivity,specificity, and predictive values of reagent test strip estimations of blood urea nitrogen

Azostix-reagent-tests(R) strips (Ames, Miles, Inc., Diagnostic Division, Elkhart, IN) were used to measure blood urea nitrogen values in blood samples from 125 dogs and cats at the North Carolina State University, College of Veterinary Medicine. Results of the tests were compared with standard serum urea nitrogen results. Sensitivity, specificity, and negative predictive values were high (86.4, 90.3, and 96.5%, respectively). Positive predictive value was low, 65.5% of the dogs and cats with elevated blood urea nitrogen values were correctly classified as abnormal The test performs well when the prevalence of abnormal values is near 50%.     

Evaluation of a radioimmunoassay for type III procollagen peptide in the dog

Type III procollagen peptide (P-3-P) is a serum marker for hepatic fibrosis in humans. The utility of a commercially available radioimmunoassay for P-3-P was evaluated in the dog. The specificity of the assay was assessed by polyacrylamide gel electrophoresis (PAGE) of canine serum and purified bovine P-3-P, followed by Western immunoblotting with rabbit aniti-P-3-P serum. The sensitivity was assessed by performing the radioimmunoassay on dilutions of sera from 22 dogs. Polyacrylamide gel electrophoresis of purified bovine P-3-P and sera from two dogs suspected of having elevated P-3-P concentrations revealed no homologous bands of staining. Western immunoblotting showed marked cross-reactivity of the high antisera concentrations with several components of the serum proteins, but none corresponding to the purified P-3-P. All tested sera from dogs had minimal competitive binding with radiolabeled P-3-P in the radioimmunoassay. Dilution curves of dog sera did not parallel either the standard curve or the dilution curve of a known test human serum. There were no statistically different P-3-P concentrations in any of the groups of dogs studied. It was concluded that currently available radioimmunoassay kits for the measurement of P-3-P in the human are not applicable in the dog. Seemingly, the structure or metabolism of canine P-3-P may vary significantly from that of the bovine or human, limiting the sensitivity and specificity of this assay in the dog.     

Assessment of the hip reduction angle for predicting osteoarthritis of the hip in the Labrador Retriever

Hip palpation has been used to provide semiquantitative information regarding passive joint laxity and susceptibility to hip dysplasia. The purpose of this study was to: (1) evaluate the intra- and inter-examiner repeatability of the hip reduction angle measured at 4 months of age by three examiners using manual goniometry and an electromagnetic tracking system; (2) compare the hip reduction angle measured with manual goniometry to the hip reduction angle measured with the electromagnetic tracking system; and (3) evaluate the hip reduction angle, distraction index and Ortolani manoeuvre at 4 months of age as predictors of the development of hip osteoarthritis at 12 months of age in 11 Labrador Retriever dogs. Intra- and inter-examiner repeatability was demonstrated for both the manual and electromagnetic goniometric measurement of the hip reduction angle (coefficient of variation < 4.3% and < 6.1%; and P = 0.163 and P = 0.836 respectively). The hip reduction angle measured by manual goniometry was moderately correlated to the hip reduction angle measured by the electromagnetic tracking system (r = 0.603, P < 0.0000). The hip reduction angle measured by manual and electromagnetic goniometry was a poor predictor of osteoarthritis at 12 months of age (r = 0.231, P < 0.062, and r = 0.321, P < 0.01). The distraction index was moderately correlated with the development of osteoarthritis by 12 months of age (r = 0.493, P < 0.0000). The Ortolani sign was sensitive (100%) but not specific (41%) for the development of osteoarthritis at 12 months of age. The hip reduction angle did not further quantify the Ortolani manoeuvre as a predictor of osteoarthritis in Labrador Retrievers.     

Rates and quantities of carbon flux to ectomycorrhizal mycelium following 14C pulse labeling of Pinus sylvestris seedlings: effects of litter patches and interaction with a wood-decomposer fungus

We used a novel digital autoradiographic technique that enabled, for the first time, simultaneous visualization and quantification of spatial and temporal changes in carbon allocation patterns in ectomycorrhizal mycelia. Mycorrhizal plants of Pinus sylvestris L. were grown in microcosms containing non-sterile peat. The time course and spatial distribution of carbon allocation by P. sylvestris to mycelia of its mycorrhizal partners, Paxillus involutus (Batsch) Fr. and Suillus bovinus (L.): Kuntze, were quantified following 14C pulse labeling of the plants. Litter patches were used to investigate the effects of nutrient resource quality on carbon allocation. The wood-decomposer fungus Phanerochaete velutina (D.C.: Pers.) Parmasto was introduced to evaluate competitive and territorial interactions between its mycelial cords and the mycelial system of S. bovinus. Growth of ectomycorrhizal mycelium was stimulated in the litter patches. Nearly 60% of the C transferred from host plant to external mycorrhizal mycelium (> 2 mm from root surfaces) was allocated to mycelium in the patches, which comprised only 12% of the soil area available for mycelial colonization. Mycelia in the litter patch most recently colonized by mycorrhizal mycelium received the largest investment of carbon, amounting to 27 to 50% of the total 14C in external mycorrhizal mycelium. The amount of C transfer to external mycelium of S. bovinus following pulse labeling was reduced from a maximum of 167 nmol in systems with no saprotroph to a maximum of 61 nmol in systems interacting with P. velutina. The 14C content of S. bovinus mycelium reached a maximum 24-36 h after labeling in control microcosms, but allocation did not reach a peak until 56 h after labeling, when S. bovinus interacted with mycelium of P. velutina. The mycelium of S. bovinus contained 9% of the total 14C in the plants (including mycorrhizae) at the end of the experiment, but this was reduced to 4% in the presence of P. velutina. The results demonstrate the dynamic manner in which mycorrhizal mycelia deploy C when foraging for nutrients. The inhibitory effect of the wood-decomposer fungus P. velutina on C allocation to external mycorrhizal mycelium has important implications for nutrient cycling in forest ecosystems.     

Adaptation of Marek's disease virus to the Vero continuous cell line

Marek's disease virus (MDV) is a highly infectious, cell-associated oncogenic herpesvirus. Production of MD vaccines has been limited to primary chicken and duck embryo fibroblast (CEF and DEF) cultures. These have a limited life span and cannot be readily stored in liquid nitrogen. Moreover, the need to prepare CEF and DEF cells on a regular basis from 10 to 11 day-old embryos derived from a flock that must be tested continuously for the presence of avian pathogens adds to the cost of vaccine production. A continuous cell line that would support MDV replication could have significant advantages for the rapid large-scale preparation of MD vaccines. In this report, we describe the adaptation to growth of CEF-grown preparations of serotype 1 and serotype 3 (herpesvirus of turkeys; HVT) strains of MDV in cells of the Vero continuous cell line. Although both viruses produced typical CPE, higher levels of infectious progeny and more extensive virus-specific immunofluorescence were obtained for HVT than for the serotype 1 virus. PCR and pulsed field electrophoresis (PFE) analysis of the DNA from Vero cells infected with either virus confirmed the presence of virus-specific DNA.     

BA和激素对试管番茄愈伤组织形态发生的影响

用番茄‘Bonny Best’品种的子叶和下胚轴作为外植体,在MS附加不同水平的BA和激素的培养基中,愈伤组织形态发生的情况是不同的。本试验结果表明:在相同的植物生长调节剂水平下,番茄子叶外植体较下胚轴外植体更容易形成愈伤组织和生根;番茄子叶和下胚轴愈伤组织的形成要求不同的激素水平,较高的水平适合于子叶愈伤组织的生长,而下胚轴愈伤组织的生长要求较低的激素水平;较高浓度的BA诱导子叶和下胚轴外植体形成白色紧密的愈伤组织,而且茎的分化出现较早,较低水平的BA产生疏松的愈伤组织,分化效迟;在MS BA22.2-44.4μM IAA1-5μM的培养基上,番茄子叶和下胚轴切段可直接分化成茎;在无激素MS培养基上,下胚轴切段可直接成苗;疏松的愈伤组织是进行悬浮培养的良好材料,可利用它们进行原生质体培养和抗性克隆的筛选。     

Renal medullary crest necrosis associated with phenylbutazone therapy in horses

Thirty-five cases of renal medullary crest necrosis morphologically similar to the renal papillary necrosis of analgesic nephropathy as described in man and rats are reported in horses receiving maintenance dosages of phenylbutazone. The primary lesion is a well-demarcated focal medullary necrosis resulting in sequestration of fragments of the renal crest. Renal cortical lesions are considered secondary to the medullary necrosis and consist of segmental pallor as a result of tubular dilatation, filtrate retention, and interstitial edema. Ischemia in concert with phenylbutazone is suggested as the etiology. Renal medullary crest necrosis is presented as more appropriate morphological terminology for this lesion in the equine species than renal papillary necrosis as is used in man and rats.     

Antigenic relationship of turkey coronavirus isolates from different geographic locations in the United States

The purpose of the present study was to examine the antigenicity of turkey coronavirus (TCV) isolates from various geographic areas with antibodies to different viruses. Seventeen isolates of TCV were recovered from intestinal samples submitted to Animal Disease Diagnostic Laboratory, Purdue University, from turkey farms located in different geographic areas. The prototype TCV Minnesota isolate (TCV-ATCC) was obtained from the American Type Culture Collection. Intestinal sections were prepared from turkey embryos infected with different TCV isolates and reacted with polyclonal or monoclonal antibodies to TCV, infectious bronchitis virus (IBV), bovine coronavirus (BCV), transmissible gastroenteritis virus (TGEV), reovirus, rotavirus, adenovirus, or enterovirus in immunofluorescent antibody staining. All 18 TCV isolates have the same antigenic reactivity pattern with the same panel of antibodies. Positive reactivity was seen with polyclonal antibodies to the TCV Indiana isolate, the TCV Virginia isolate, TCV-ATCC, and the IBV Massachusetts strain as well as monoclonal antibodies to the TCV North Carolina isolate or the membrane protein of IBV. Antibodies to BCV or TGEV were not reactive with any of the TCV isolates. Reactivity of antibodies to unrelated virus, rotavirus, reovirus, adenovirus, or enterovirus with different TCV isolates was all negative, except positive response was seen between enterovirus antibody and a TCV western North Carolina isolate, suggesting coinfection of turkeys with TCV and enterovirus in that particular case. The results indicated that the TCV isolates from these geographic locations in the U.S. shared close antigenicity and were antigenically related to IBV.