Pseudomonas syringae pv. coryli, the Causal Agent of Bacterial Twig Dieback of Corylus avellana

ABSTRACT Thirty-eight bacterial strains isolated from hazelnut (Corylus avellana) cv. Tonda Gentile delle Langhe showing a twig dieback in Piedmont and Sardinia, Italy, were studied by a polyphasic approach. All strains were assessed by fatty acids analysis and repetitive sequence-based polymerase chain reaction (PCR) fingerprinting using BOX and ERIC primer sets. Representative strains also were assessed by sequencing the 16S rDNA and hrpL genes, determining the presence of the syrB gene, testing their biochemical and nutritional characteristics, and determining their pathogenicity to hazelnut and other plants species or plant organs. Moreover, they were compared with reference strains of other phytopathogenic pseudomonads. The strains from hazelnut belong to Pseudomonas syringae (sensu latu), LOPAT group Ia. Both fatty acids and repetitive-sequence-based PCR clearly discriminate such strains from other Pseudomonas spp., including P. avellanae and other P. syringae pathovars as well as P. syringae pv. syringae strains from hazelnut. Also, the sequencing of 16S rDNA and hrpL genes differentiated them from P. avellanae and from P. syringae pv. syringae. They did not possess the syrB gene. Some nutritional tests also differentiated them from related P. syringae pathovars. Upon artificial inoculation, these strains incited severe twig diebacks only on hazelnut. Our results justify the creation of a new pathovar because the strains from hazelnut constitute a homogeneous group and a discrete phenon. The name of P. syringae pv. coryli is proposed and criteria for routine identification are presented.     

Transmission of Phytophthora ramorum in Mixed-Evergreen Forest in California

ABSTRACT During 2001 to 2003, the transmission biology of Phytophthora ramorum, the causal agent of sudden oak death, was studied in mixedevergreen forest, a common forest type in northern, coastal California. Investigation of the sources of spore production focused on coast live oak (Quercus agrifolia) and bay laurel (Umbellularia californica), dominant hosts that comprised 39.7 and 46.2% of the individuals at the study site, respectively. All tests for inoculum production from the surface of infected coast live oak bark or exudates from cankers were negative. In contrast, sporangia and chlamydospores were produced on the surface of infected bay laurel leaves. Mean number of zoospores produced from infected bay laurel leaves under natural field conditions during rainstorms was 1,173.0 +/- SE 301.48, and ranged as high as 5,200 spores/leaf. P. ramorum was recovered from rainwater, soil, litter, and streamwater during the mid- to late rainy season in all 3 years of the study. P. ramorum was not recovered from sporadic summer rains or soil and litter during the hot, dry summer months. Concentrations of inoculum in rainwater varied significantly from year to year and increased as the rainy season progressed for the two complete seasons that were studied. Potential dispersal distances were investigated for rainwater, soil, and streamwater. In rainwater, inoculum moved 5 and 10 m from the inoculum source. For soil, transmission of inoculum was demonstrated from infested soil to bay laurel green leaf litter, and from bay laurel green leaf litter to aerial leaves of bay laurel seedlings. One-third to one-half of the hikers tested at the study site during the rainy season also were carrying infested soil on their shoes. In streamwater, P. ramorum was recovered from an unforested site in pasture 1 km downstream of forest with inoculum sources. In total, these studies provide details on the production and spread of P. ramorum inoculum in mixed-evergreen forest to aid forecasting and managing disease transmission of this environmentally destructive pathogen.     

Spatial Patterns of Viable Spore Deposition of Gibberella zeae in Wheat Fields

ABSTRACT An increased understanding of the epidemiology of Gibberella zeae will contribute to a rational and informed approach to the management of Fusarium head blight (FHB). An integral phase of the FHB cycle is the deposition of airborne spores, yet there is no information available on the spatial pattern of spore deposition of G. zeae above wheat canopies. We examined spatial patterns of viable spore deposition of G. zeae over rotational (lacking cereal debris) wheat fields in New York in 2002 and 2004. Viable, airborne spores (ascospores and macroconidia) of G. zeae were collected above wheat spikes on petri plates containing a selective medium and the resulting colonies were counted. Spores of G. zeae were collected over a total of 68 field environments (three wheat fields during 54 day and night sample periods over 2 years) from spike emergence to kernel milk stages of local wheat. Spatial patterns of spore deposition were visualized by contour plots of spore counts over entire fields. The spatial pattern of spore deposition was unique for each field environment during each day and night sample period. Spore deposition patterns during individual sample periods were classified by spatial analysis by distance indices (SADIE) statistics and Mantel tests. Both analyses indicated that the majority (93%) of the spore deposition events were random, with the remainder being aggregated. All of the aggregated patterns were observed during the night. Observed patterns of spore deposition were independent of the mean number of viable spores deposited during individual sample periods. The spatial pattern for cumulative spore deposition during anthesis in both years became aggregated over time. Contour maps of daily and cumulative spore deposition could be compared with contour maps of FHB incidence to gain insights into inoculum thresholds and the timing of effective inoculum for infection.     

Induced resistance for plant disease control: maximizing the efficacy of resistance elicitors

ABSTRACT Plants can be induced to develop enhanced resistance to pathogen infection by treatment with a variety of abiotic and biotic inducers. Biotic inducers include infection by necrotizing pathogens and plant-growth-promoting rhizobacteria, and treatment with nonpathogens or cell wall fragments. Abiotic inducers include chemicals which act at various points in the signaling pathways involved in disease resistance, as well as water stress, heat shock, and pH stress. Resistance induced by these agents (resistance elicitors) is broad spectrum and long lasting, but rarely provides complete control of infection, with many resistance elicitors providing between 20 and 85% disease control. There also are many reports of resistance elicitors providing no significant disease control. In the field, expression of induced resistance is likely to be influenced by the environment, genotype, and crop nutrition. Unfortunately, little information is available on the influence of these factors on expression of induced resistance. In order to maximize the efficacy of resistance elicitors, a greater understanding of these interactions is required. It also will be important to determine how induced resistance can best fit into disease control strategies because they are not, and should not be, deployed simply as "safe fungicides". This, in turn, will require information on the interaction of resistance elicitors with crop management practices such as appropriate-dose fungicide use.     

Use of gyroscopic sensors for objective evaluation of trimming and shoeing to alter time between heel and toe lift-off at end of the stance phase in horses walking and trotting on a treadmill

OBJECTIVE: To determine whether a shoe with an axialcontoured lateral branch would induce greater lateral roll of the forelimb hoof during the time between heel and toe lift-off at end of the stance phase (breakover). Animals-10 adult horses. PROCEDURE: A gyroscopic transducer was placed on the hoof of the right forelimb and connected to a transmitter. Data on hoof angular velocity were collected as each horse walked and trotted on a treadmill before (treatment 1, no trim-no shoe) and after 2 treatments by a farrier (treatment 2, trim-standard shoe; and treatment 3, trim-contoured shoe). Data were converted to hoof angles by mathematical integration. Breakover duration was divided into 4 segments, and hoof angles in 3 planes (pitch, roll, and yaw) were calculated at the end of each segment. Multivariable ANOVA was performed to detect differences among treatments and gaits. RESULTS: Trimming and shoeing with a shoe with contoured lateral branches induced greater mean lateral roll to the hoof of 3.2 degrees and 2.5 degrees during the first half of breakover when trotting, compared with values for no trim-no shoe and trim-standard shoe, respectively. This effect dissipated during the second half of breakover. When horses walked, lateral roll during breakover was not significantly enhanced by use of this shoe. CONCLUSIONS AND CLINICAL RELEVANCE: A shoe with an axial-contoured lateral branch induced greater lateral roll during breakover in trotting horses, but change in orientation of the hoof was small and limited to the first half of breakover.     

In vitro cytopathic effects of a cysteine protease of Tritrichomonas foetus on cultured bovine uterine epithelial cells

OBJECTIVE: To evaluate the cytopathic effects of Tritrichomonas foetus and a purified cysteine protease (ie, CP30) of T foetus on cultured bovine uterine epithelial cells (BUECs) in vitro. SAMPLE POPULATION: 10 reproductive tracts were obtained from late-term bovine fetuses at a commercial abattoir. PROCEDURE: An in vitro culture system of BUECs was developed to study the cytopathic effects of T foetus and purified CP30 of T foetus on host cells. Cytotoxicity of T foetus or CP30 on exposed BUECs was determined. Fluorescence microscopy and flow cytometry analyses were used to detect apoptosis. A fluorometric assay was used to detect BUEC caspase 3 activation. The CP inhibitor E-64 and a caspase inhibitor were used to inhibit apoptosis. RESULTS: Cytopathic effects were observed in BUECs treated with parasites or CP30 and were concentration and time dependent. The BUECs underwent apoptosis in the presence of parasites or CP30. The specific CP inhibitor E-64 abolished the induction of apoptosis in BUECs by CP30. The caspase inhibitor reduced the amount of apoptosis in BUECs. CONCLUSIONS AND CLINICAL RELEVANCE: T foetus and its CP30 induce apoptosis in cultured BUECs in vitro. Induction of apoptosis by CP30 is correlated with protease activity. Endometrial cell death as a result of a T foetus infection is likely to be more important in mediating infertility than a direct effect on the conceptus. Provoking an apoptotic reaction in the host may mitigate an inflammatory reaction or immune response and therefore favor survival of the parasite in a chronic infection.     

Relative sensitivity of polymerase chain reaction assays used for detection of feline herpesvirus type 1 DNA in clinical samples and commercial vaccines

OBJECTIVE: To determine relative detection rates and detection limits for 6 published polymerase chain reaction (PCR) assays used for detection of feline herpesvirus type 1 (FHV-1) DNA. SAMPLE POPULATION: 5 vaccines licensed for use in preventing FHV-1-associated disease; 15 conjunctival biopsy specimens collected from cats with keratitis, conjunctivitis, or both; and a plaque-purified field isolate of FHV-1 cultured in vitro. PROCEDURE: Vaccines and clinical samples were assessed for FHV-1 DNA by use of all 6 assays. Detection rates were calculated by assuming that any sample in which FHV-1 DNA was detected was a true-positive result. Detection limits were estimated by use of serial dilutions of DNA extracted from cultured FHV-1 and 1 clinical sample. RESULTS: Testing by use of all 6 assays resulted in detection of FHV-1 DNA in all 5 vaccines. Testing by use of all 6 assays yielded concordant results for 9 of 15 conjunctival biopsy specimens (8 with negative results and 1 with a positive result). Calculated detection rates for clinical samples ranged from 29% to 86%. Assay sensitivity was ranked similarly by use of detection rate or detection limit. CONCLUSIONS AND CLINICAL RELEVANCE: Testing by use of all assays was equally likely to detect vaccine virus. Therefore, a positive PCR result in a cat may reflect vaccine virus rather than wild-type virus. Test sensitivity as assessed by detection limits and detection rates varied greatly. Because FHV-1 can be shed in clinically normal animals, high detection rate will not necessarily correlate with high diagnostic sensitivity.     

Evaluation of enzyme-linked immunosorbent assays performed on milk and serum samples for detection of paratuberculosis in lactating dairy cows

OBJECTIVE: To determine whether results obtained for milk and serum samples with ELISAs intended for diagnosis of paratuberculosis in dairy cows were comparable to results obtained by means of mycobacterial culture of fecal samples. DESIGN: Cross-sectional study. ANIMALS: 689 lactating dairy cows in 9 Ontario herds. PROCEDURE: Milk, serum, and fecal samples were obtained from all cows. Fecal samples were submitted for mycobacterial culture. Serum samples were tested with a commercially available ELISA for antibodies against Mycobacterium paratuberculosis, and preserved milk samples were tested with an indirect ELISA for antibodies against M paratuberculosis. RESULTS: Results were positive for 130 of the 689 (18.9%) serum samples, 77 of the 689 (11.1%) milk samples, and 72 of the 689 (10.4%) fecal samples. The level of agreement between results for milk and serum samples was only moderate. Proportions of positive results for serum and fecal samples were significantly different, but proportions of positive results for milk and fecal samples were not significantly different. In addition, results for milk samples had a higher level of agreement with results of mycobacterial culture than did results for serum samples. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that the indirect ELISA used on milk samples may be a convenient method of detecting paratuberculosis in dairy herds.     

Evaluation of the association between initial proteinuria and morbidity rate or death in dogs with naturally occurring chronic renal failure

OBJECTIVE: To determine whether urine protein-to-creatinine ratio (UP:C) > or = 1.0 at initial diagnosis of chronic renal failure (CRF) is associated with greater risk of development of uremic crises, death, and progression of renal failure in dogs. DESIGN: Prospective cohort study. ANIMALS: 45 dogs with CRF PROCEDURE: Dogs were prospectively assigned to 2 groups on the basis of initial UP:C < 1.0 or 2 > or = 1.0. The association between magnitude of proteinuria and development of uremic crises and death was determined before and after dogs with initial UP:C > or =1.0 were assigned to 3 subgroups and compared with dogs with initial UP:C < 1.0. Changes in reciprocal serum creatinine concentration were used to estimate decrease in renal function. RESULTS: Initially, dogs had similar clinical characteristics with the exception of systolic blood pressure and UP:C. Relative risks of development of uremic crises and death were approximately 3 times higher in dogs with UP:C > or =1.0, compared with dogs with UP:C < 1.0. Relative risk of adverse outcome was approximately 1.5 times higher for every 1-unit increment in UP:C. The decrease in renal function was of greater magnitude in dogs with UP:C > or =1.0, compared with dogs with UP:C < 1.0. CONCLUSIONS AND CLINICAL RELEVANCE: Initial UP:C > or =1.0 in dogs with CRF was associated with greater risk of development of uremic crises and death, compared with dogs with UP:C < 1.0. Initial determinations of UP:C in dogs with naturally occurring CRF may be of value in refining prognoses.