Sensitive diagnosis of bovine tuberculosis in a farmed cervid herd with use of an MPB70 protein fluorescence polarization assay

After histopathological examination of a lesion found in a herd member returned a diagnosis of mycobacteriosis, a farmed herd (n = 47) of elk (Cervus elaphus nelsoni) and red deer (C. elaphus elaphus) was investigated for bovine tuberculosis with a battery of antemortem and postmortem diagnostic tests. Every animal was tested with the mid-cervical tuberculin skin test; all 47 had negative results. All of the 16 adult animals and 15 of the 31 calves (approximately 2-years-old) were blood-tested with a lymphocyte stimulation test (LST) and a fluorescence polarization assay (FPA), which detects antibody to the MPB70 protein antigen. At necropsy of the 31 blood-tested animals, tissues were harvested for histopathological examination and culture of mycobacteria. Mycobacterium bovis was isolated from 16 of the 31 animals, and a scotochromogen was also isolated from 1 of the 16 whose tissues yielded M. bovis. Each of these 16 animals, 15 of which were calves, also received a histopathological diagnosis of mycobacteriosis. Other species of mycobacteria, including those belonging to the M. avium and M. terrae complexes, were isolated from an additional 7 animals. The FPA was scored “positive” or “suspect” for 16 animals, 13 (81%) of which were culture-positive for M. bovis. The other 3 animals that were culture-positive for M. bovis had negative FPA results. Of the 3 FPA-positive or FPA-suspect animals that were culture-negative, 2 were suspected to have mycobacteriosis on the basis of the histopathological examination. The 7 animals from which Mycobacterium species other than M. bovis were cultured were all FPA-negative. The only animal with positive LST results was also FPA-positive and culture-positive for M. bovis. The M. bovis isolates had an identical spoligotype pattern, with an octal code of 664073777777600. This is the first report of the isolation and identification of this strain type in Canada.     

Agronomic performance and combining ability estimates of yellow maize inbred lines under adequate and deficit moisture conditions

Journal of Crop Science and Biotechnology - Breeding for drought tolerance and increased grain yield is vital in mitigating the threat posed by recurrent drought stress on maize production, as well...     

Quantitative and qualitative influence of inoculation methods on in planta growth of rice blast fungus

ABSTRACT Molecular analyses of early disease events require infected plant tissue in which the pathogen is present in high quantities and interacts with the plant in a way found in the field. In this study, a quantitative polymerase chain reaction (Q-PCR) assay was developed to determine an "infection ratio" of fungal to plant cells in infected tissues. This assay was used to evaluate four inoculation methods (spray, mist, dip, and sheath) as well as use of whole plants or excised parts. Fluorescence stereomicroscopy was used to follow individual lesions developing from appressoria to macroscopic symptoms. Disease progression and outcomes were documented from 24 to 96 h postinoculation (hpi), as well as effectiveness of Pi-ta-mediated resistance. Even at 96 hpi, fungus proliferated well ahead of visible plant damage, especially in veins. Developing lesions sometimes were surrounded by greener areas in detached leaves. Spray inoculation was not sufficient for detecting fungal gene expression in planta before 96 h. Alternatively, a leaf sheath assay produced infected tissues containing 10 to 30% fungal DNA by 34 h. Used together, Q-PCR quantification and fluorescence stereomicroscopy will facilitate studies of early plant invasion because infection density and fungal growth stages are directly observed, not assumed from incubation time.     

Peripheral nerve stimulation under ultrasonographic control to determine the needle‐to‐nerve relationship

ObjectiveTo determine the needle‐to‐nerve distances during electrical nerve location in dogs at different currents and pulse duration using a peripheral nerve stimulator (PNS) under ultrasound control (US), and the minimal electrical thresholds (MET) necessary to obtain a motor response (MR) after achieving needle‐to‐nerve contact.Study designProspective in vivo experimental trial in a clinical settingAnimalsThirty dogs, scheduled for locoregional anaesthesia of the sciatic nerve.MethodsNeedle‐to‐nerve distance was measured ultrasonographically after obtaining the MR of sciatic nerve with 2, 1 and 0.5 mA and pulse duration 0.1 ms (NS0.1). Thereafter the needle was placed in contact with the nerve and MET was determined. The procedure was repeated with 0.3 ms (NS0.3). Finally the needle was reintroduced to contact the sciatic nerve guided only by US, thus MET‐US was determined. Data were analysed using Kruskal–Wallis or Mann–Whitney tests.ResultsNeedle‐to‐nerve distances were greater when MR was obtained with 2 mA than with 1 and 0.5 mA at 0.1 and 0.3 ms. No significant differences were observed between the needle‐to‐nerve distances using 0.1 or 0.3 ms.The MET [median (range)] was 0.4 (0.18–1.3) mA in NS0.1, 0.32 (0.12–0.8) mA in NS0.3; while MET‐US was 0.7 (0.32–1.5) mA. When the needle contacted the nerve, the MR achieved with currents below 0.3 mA was obtained in 17.2, 40 and 0% of cases using NS0.1, NS0.3 and US respectively.Conclusions and clinical relevanceThe electrical current necessary to obtain a MR decreased as the needle moved towards the nerve. However when the needle tip contacted the nerve, an MR with low current intensity could not be obtained. Thus the absence of motor response at currents below 0.3 mA cannot rule out needle‐epineurium contact. When ultrasound is combined with PNS, it is more important to assess the correct needle position than searching for an MR at low currents.     

Viability and infectiousness of eggs of Echinococcus granulosus aged under natural conditions of inferior arid climate

We studied the viability and infectiousness of aged Echinococcus granulosus eggs by in vivo evaluation in ovines. Our results demonstrated that after 41 months of ageing of the eggs under environmental conditions of an inferior arid climate (Patagonia, Argentina), they were still able to produce infection in 4/4 ovines challenged with 1200 eggs per ovine. In the ovines experimentally infected with these aged eggs, the occurrence of hepatic and pulmonary cysts was determined by necropsy, histologic and genetic studies. The eggs were found in a semi-senescent stage, thus keeping their capacity to generate an infection in the intermediary ovine host.     

Spoligotyping of Mycobacterium bovis isolates found in Manitoba

Spoligotyping was applied to 44 isolates of Mycobacterium bovis obtained from the Canadian province of Manitoba. Isolates were obtained from submissions of elk (n = 16), deer (n = 1), and cattle (n = 27) tissues spanning the period of 1990 to early 2003. Two spoligotype profiles were obtained differing only in the reaction with oligonucleotide number 12. Forty of the 44 isolates (90.9%) hybridized with oligonucleotide 12 (MB-1 type), while the remaining 4 of 44 (9.1%) did not show a signal at position 12 (MB-2 type). Octal codes for these 2 types are 656573377603600 and 656473377603600, respectively. These spoligotypes have not been reported as occurring elsewhere worldwide.     

An environmentally sustainable approach for the management of <Emphasis Type="Italic">Phaeoacremonium minimum</Emphasis>, the main agent of wood diseases in <Emphasis Type="Italic">Actinidia deliciosa</Emphasis>

Among the fungi associated with the kiwi wood diseases, the vascular pathogen Phaeoacremonium minimum can infect plants already at nursery stage, without any external symptoms. At the moment, there are not effective control strategies. The effect of soil treatments applicable in organic agriculture was evaluated in two-years experiments on potted kiwi plants artificially inoculated with P. minimum. The soil treatments were based on commercial formulations of iron chelate, silicon, neem paste, Trichoderma afroharzianum strain T22, and cover cropping with selected perennial Poaceae. Cover cropping and iron chelate treatments enhanced the iron availability for the plants and significantly reduced wood necrosis caused by the pathogen. Both treatments also produced an increase of hairy root proliferation, so the plants were able to better cope with stress conditions. Laboratory assays showed the role of iron on the pathogen growth and its pathogenesis enzyme activities.     

Pulse wave Doppler ultrasound of testicular arteries and their relationship with semen characteristics in healthy bulls

Background: Semen evaluation is used to estimate the testicular function. In bulls, the spermatozoa present in the ejaculate are the result of a process that begun more than 2 mo earlier, bequeathing a delayed depiction of the actual function of the testis. Since testis vascularization might be critical for the gonad function, selected pulse wave Doppler ultrasound parameters were assessed in this study, for instance the peak systolic velocity, the end diastolic velocity and the resistive index of the testicular artery along the spermatic cord, the marginal portion of the testicular artery and the intratesticular branches of the testicular artery both in healthy adult and young bulls.Correlations between these parameters and characteristics of semen that was collected numerous times, before and after the Doppler ultrasound examination.Results: The peak systolic velocity and the end diastolic velocity measured in the testicular artery along the spermatic cord(supratesticular artery – SA) were variable among the bulls and within individual bulls, likely due to the convoluted course of the vessel. The resistive index was found highly repeatable in the same bull. A reduction in the resistive index was found between the supratesticular artery and the marginal portion of the testicular artery(P 0.01), and between the marginal portion of the testicular artery and the intratesticular branches of the testicular artery(P 0.05). No differences were recorded for the pulse wave Doppler ultrasound parameters in young bulls compared with adults. A significant correlation was found between the resistive index of the marginal portion of the testicular artery and total sperm in the ejaculate(r = 0.516, P 0.05), the immature sperm(r = 0.462, P 0.05), the teratoid sperm(r = 0.375, P 0.05), and the "Dag defect" sperm(r = 0.389, P 0.05). Similarly, the resistive index of the intratesticular branches of the testicular artery were found correlated with the total sperm number in the ejaculate(r = 0.568, P 0.05), the immature sperm(r = 0.523, P 0.05), the teratoid sperm(r = 0.418, P 0.05), and the "Dag defect" sperm(r = 0.341, P 0.05).Conclusions: The data presented in this study suggest that the resistive index, measured at the marginal portion of the testicular artery, could be an easy-to-perform parameter to evaluate the spermatogenesis quality in young bulls and normal adults.