Isolation of Pathogenicity Mutants of Fusarium oxysporum f. sp. melonis by Insertional Mutagenesis

Restriction enzyme-mediated integration (REMI) mutagenesis was used to isolate mutants of Fusarium oxysporum f. sp. melonis impaired in pathogenicity. The race 2 strain Mel02010 was transformed with linearized pSH75, conferring resistance to hygromycin B, with or without the enzyme used to linearize the plasmid. Addition of restriction enzymes did not affect the transformation frequency. A total of 2929 REMI transformants were tested for pathogenicity to three melon cultivars, Amus, Ogon 9 and Ohi. The race 2 strains are pathogenic to Amus and Ogon 9, but not to Ohi. Of 43 transformants with reduced pathogenicity on susceptible melon cultivars, 12 mutants were examined in detail for pathogenicity, vegetative growth and integrative mode of pSH75. The levels of pathogenicity varied among these mutants. Two mutants (B48 and B137) almost completely lost pathogenicity to both susceptible cultivars, and the others had reduced pathogenicity. Mutants B48, B241, B886 and X36 were also impaired in vegetative growth. Mutant B809 was a biotin auxotroph. By DNA gel blot analysis, nine mutants were found to contain a single copy of the transformation vector. These mutants may thus be useful in isolating genes involved in pathogenicity. Received 22 December 2000/ Accepted in revised form 16 April 2001     

Characterization of Elicitor-inducible Tobacco Genes Isolated by Differential Hybridization

Inducible responses in plants against pathogen attack play a major role in resistance to disease. The defense responses are mostly associated with the expression of various kinds of inducible genes. We employed differential hybridization to isolate elicitor-inducible genes (EIGs) of tobacco (Nicotiana tabacum cv. Samsun NN) using the tobacco-fungal elicitor system. A cDNA library was constructed from tobacco leaves treated for 12 hr with hyphal wall components (HWC) prepared from Phytophthora infestans, and six EIGs were identified. Expression of all EIGs was induced after inoculation with the soybean pathogen Pseudomonas syringae pv. glycinea (nonpathogenic on tobacco) or treatment with salicylic acid, and a variety of expression patterns of EIG mRNAs was observed. Sequence analysis of EIG cDNAs revealed similarities to genes for SAR8.2 (EIG-B39 and EIG-D14), glycine-rich protein (EIG-G7), extensin (EIG-I30), acyltransferase (EIG-I24) and unknown protein (EIG-J7). Possible roles of EIG products in disease resistance are discussed. Received 30 August 2000/ Accepted in revised form 30 November 2000     

Growth Complementation of hrpXo Mutants of Xanthomonas oryzae pv. oryzae by Virulent Strains in Rice Cultivars Resistant and Susceptible to the Parental Strain

Xanthomonas oryzae pv. oryzae (X. o. pv. oryzae) T7174 is virulent on rice cultivar IR24 and avirulent on IR-BB2. From recent reports, some virulence and avirulence factors of plant pathogenic bacteria are transferred to plant cells through the hrp-dependent type III secretion system. In this study, we investigated the involvement of hrp genes in the compatible and the incompatible interactions between rice and X. o. pv. oryzae after co-inoculation with hrpXo mutants derived from T7174 and virulent strains. Growth of the mutants, named 74ΔHrpXo and 76ΔHrpXo, was repressed in IR24 when the mutants were applied alone. However, growth of the mutants was complemented by co-inoculation with virulent strains. Growth of bioluminescent hrpXo mutant 76ΔHrpXo in IR24 and its growth in IR-BB2 after co-inoculation with T7133, which is virulent on both cultivars, was equally complemented, as detected by bioluminescence from the mutant. On the other hand, only partial complementation of growth of T7174L76, which is a bioluminescent and pathogenic derivative of T7174, by T7133 was observed in IR-BB2. Thus, growth of the hrpXo mutant of X. o. pv. oryzae was complemented by virulent strains in both susceptible and resistant rice leaves with the parental strain. Received 21 July 2000/ Accepted in revised form 26 October 2000     

An Improved Method for Isolating Violet Root Rot Fungus, Helicobasidium mompa, from Basidiocarps

Basidiocarps of the violet root rot fungus, Helicobasidium mompa, are less frequently used for isolation than are mycelial strands and sclerotia even though the basidiocarps are conspicuously produced at the trunk base of diseased plants. Basidiocarps are also more suitable for storage. This paper describes an improved method for obtaining pure cultures from basidiocarps using microcentrifuge tubes to facilitate the awkward steps of rinsing fungal materials under a dissecting microscope. Received 28 June 2000/ Accepted in revised form 4 August 2000     

Comparison of Multistix PRO dipsticks with other biochemical assays for determining urine protein (UP), urine creatinine (UC) and UP:UC ratio in dogs and cats

BACKGROUND: Urine protein: urine creatinine (UP:UC) ratio determined from the quantitative measurement of protein and creatinine in a single urine sample is the best feasible assessment of clinically significant proteinuria in dogs and cats. A dipstick that measures urine protein, urine creatinine, and UP:UC ratio has been used in human medicine and could have application for veterinary practice. OBJECTIVE: The objective of this study was to compare the Multistix PRO dipstick (Bayer Corporation, Elkhart, IN, USA) to other biochemical methods for determination of urine protein and creatinine, and UP:UC ratio in canine and feline urine. METHODS: A complete urinalysis, including sulfosalicylic acid (SSA) precipitation, was performed on urine samples submitted to our laboratory between February and April 2003 from 100 dogs and 49 cats. Urine protein and creatinine concentrations were determined by the Multistix PRO dipstick using a Clinitek 50 analyzer (Bayer) and compared with the results of SSA precipitation and quantitative biochemical analysis. The UP:UC ratios from the dipstick results (calculated by the Clinitek 50 and also manually) were compared with those calculated from quantitative values. Pearson product-moment correlation analysis and diagnostic sensitivity and specificity (using quantitative results as the gold standard) were determined. RESULTS: For both canine and feline urine, protein and creatinine concentrations determined by the Multistix PRO correlated closely with quantitative concentrations for protein (dogs r = .78, P = .0001; cats r = .87, P = .0001) and creatinine (dogs r = .78, P = .0001; cats r = .76, P = .0001). The Multistix PRO was more sensitive and less specific than SSA precipitation for diagnosing clinically significant proteinuria. UP:UC ratios obtained by manual calculation of dipstick results correlated best with quantitative UP:UC ratios in dogs, and had higher specificity but lower sensitivity for the diagnosis of proteinuria. In cats, UP:UC ratios determined by the dipstick method did not correlate (r = -.24, P = .0974) with quantitative values. CONCLUSIONS: The Multistix PRO, with manual calculation of UP:UC, may be a good alternative for the diagnosis of clinically significant proteinuria in dogs, but not cats. Dipstick creatinine concentration should be considered as an estimate.     

玉树县草地退化现状及治理措施

通过调查资料分析,玉树县现有退化草地597533.3hm2,占可利用草场面积的43.56%。草地退化的主要原因是:超载过牧,鼠虫危害以及滥采乱挖等人为因素。并提出了加强草地建设,实施退耕还林(草),退牧还草和三江源生态保护治理等项目工程以及逐步完善和实行以法治草,建立人工草地等措施。     

Comparison of the disease incidences of Finnish Ayrshire and Finnish Black and White dairy cows

Finnish Ayrshire and Finnish Black and White cows were compared regarding the incidences of early and late mastitis, parturient paresis, ketosis, ovarian disorders, metritis and the risk of having a test-day somatic-cell count >200,000 cells ml⁻¹ at any of the first three monthly test days in lactation. In a retrospective cohort study 101,793 cows from 5844 tie stalls and 11,811 cows from 437 loose-housing systems from all over Finland were followed from calving in 2000 until the end of lactation. The observed incidences of those cows were then analysed using generalised linear mixed models. Finnish Black and White cows had higher incidences of all diseases except ovarian disorders. Although the differences were statistically significant in all models except metritis and early mastitis in loose-housing systems, they were, in our view, only important on the national level (for the breeding organisations), and of little importance for the farmers.     

Herd-level risk factors for subclinical Salmonella infection in European finishing-pig herds

Our objective was to find herd factors associated with pigs testing seropositive for Salmonella. Data were collected from 359 finishing-pig herds in Germany, Denmark, Greece, The Netherlands and Sweden, between 1996 and 1998. Pigs fed non-pelleted feed (dry or wet) had 2- and 2.5-times lower odds of seropositivity, compared to pigs fed pelleted feed. The protective effect of non-pelleted feed over pelleted feed may be ascribed to the structure and composition. Also, pigs that were given whey (to drink or as the liquid part of the diet) had 2.6-times lower odds to test seropositive than pigs not getting whey. Pigs produced in batches in herds with hygienic-lock facilities had >3-times lower odds for testing seropositive compared to pigs in herds where only one or neither factor was present. In herds where the caretaker(s) washed hands consistently before tending to the animals, pigs had 1.5-times lower odds of seropositivity than pigs in herds where the caretaker did not. Pigs which were able to have snout contact with pigs in neighbouring pens (because pen separations were either open or too low) had 1.7-times higher odds to test seropositive compared to pigs for which such contact was prevented. Pigs in herds recruiting from more than three supplier herds had three-times higher odds to test seropositive than pigs in herds which breed their own replacement stock or recruit from a maximum of three supplier herds.     

Risk factors for the prevalence of porcine cysticercosis in Mbulu District, Tanzania

To estimate prevalence of and risk factors for the prevalence of porcine cysticercosis in Mbulu District, Tanzania, 770 live pigs were examined by lingual examination in 21 villages. Structured observations and questionnaire interviews were used to assess pig rearing practices and household use of latrines. Associations between factors were analyzed using a Bayesian hierarchical model to obtain prevalence odds ratio (OR) and 95% Bayesian Credible Intervals (95% BCI). Prevalence was 17.4% (village-specific range 3.2–46.7%). Prevalence of porcine cysticercosis was considerably higher in pigs reared in households lacking latrines than in those reared in households that were using latrines (OR=2.04; 95% BCI=1.25, 3.45). About 96% of the pigs were kept under free-range conditions. This study suggests the need for further studies in order to design and implement effective prevention and control measures for porcine cysticercosis in Mbulu District, Tanzania.     

Evaluation of the chemoprophylactic efficacy of 10% long acting injectable moxidectin against gastrointestinal nematode infections in calves in Belgium

The chemoprophylactic efficacy of a single dose of the 10% long acting (LA) injectable formulation of moxidectin on nematode infections in calves, was evaluated. Two similar groups of 11 female, first grazing season Holstein calves were turned out in early May on separate plots of a single, naturally infected pasture. Until 56 days post-treatment (pt), the percentage reduction in faecal egg output was 100%, remaining above 90% during the entire trial, except for day 126 pt. More than 90% of the larvae in the treated group were identified as Cooperia until 140 days after treatment and more than 70% during the rest of the trial, whereas in the control group Cooperia was the most abundant species until day 84 pt and Ostertagia from 126 days pt onwards. The reduction in faecal egg output in the treated group was reflected in the mean pepsinogen levels being below the pathogenic threshold at the end of the grazing season (1.8 units of tysrosine (U tyr)) and the absence of diarrhoea during the second half of the grazing season. In the control group pepsinogen levels remained high (mean: 5.5 U tyr) and prolonged diarrhoea occurred in the second half of the grazing season. Furthermore, the weight gain for the treated group at the end of the grazing season was 41.9 kg higher than for the control group. At necropsy, the reduction in O. ostertagi worm burden in the treated group was 97.5% compared to the control group, while the reduction in C. oncophora worm burden was 57%. An additional benefit of the long acting parasitological control, was reduced pasture contamination.