Liquid Swine Manure Can Kill Verticillium dahliae Microsclerotia in Soil by Volatile Fatty Acid, Nitrous Acid, and Ammonia Toxicity

ABSTRACT In previous studies, liquid swine manure (LSM) was sometimes shown to reduce Verticillium wilt of potato caused by Verticillium dahliae. We also observed that microsclerotia of this fungus died within 1 day, or between 3 and 6 weeks, after addition of LSM to some acid soils and within 1 week in some alkaline soils. In this study, we demonstrated that a volatile fatty acid (VFA) mixture with an identical concentration of VFAs as that found in an effective LSM reduced germination in an acid soil (pH 5.1) to the same extent as the LSM after 1 day of exposure. Germination was reduced by 45, 75, and 90% in the 10, 20, and 40% ([wt/wt] soil moisture) treatments, respectively, with the latter being equivalent to an application of 80 hl/ha. Addition to this acid soil of 19 LSMs (30% [wt/wt] soil moisture) collected from different producers resulted in complete kill of microsclerotia with 12 manures. Effective manures had a total concentration of nonionized forms of VFAs in soil solution of 2.7 mM or higher. In some acid soils (pH 5.8), addition of LSM (40% [wt/wt] soil moisture) did not kill microsclerotia until 3 to 6 weeks later. Here, a reduction in viability of microsclerotia was attributed to the accumulation of 0.06 mM nitrous acid in the soil solution at 4 weeks. When an LSM was added (40% [wt/wt] soil moisture) to an alkaline soil (pH 7.9) where VFAs are not toxic, microsclerotia germination was reduced by 80% after 1 week. Here the pH increased to 8.9 and the concentration of ammonia reached 30 mM in the soil solution. An ammonium chloride solution having an equivalent concentration of ammonium as the manure was shown to have the same spectrum of toxicity as the manure in assays ranging from pH 7 to 9, both in solutions and above the solutions. At pH 9, the concentration of ammonia reached 18 mM and 100% mortality of microsclerotia occurred. Thus, in acid soils, LSM can kill microsclerotia of V. dahliae by VFA and/or nitrous acid toxicity and in alkaline soils by ammonia toxicity. In order to take advantage of these mechanisms for disease reduction, the manure chemical composition, rate of addition, and soil characteristics need to be determined for each instance of use.     

Preliminary results on detection of maize treated with electrons for seed dressing and according decontamination and infestation

Saatgut und Getreide kann mit niederenergetischen Elektronen (<300 kev) oder=" hochenergetischen=" elektronen=" (1–10 mev)=" wirksam=" behandelt=" werden,=" um=" mikroorganismen=" und=" insekten=" abzutöten.=" in=" dieser=" vorläufigen=" studie=" wurde=" mais=" mit=" niederenergetischen=" (125 kev)=" und=" hochenergetischen=" elektronen=" (10 mev)=" behandelt.=" um=" diese=" elektronenbehandlung=" nachzuweisen,=" wurden=" verschiedene=" verfahren=" eingesetzt:=" photostimulierte=" lumineszenz=" (psl),=" thermolumineszenz=" (tl)=" und=" dna-kometentest.=" für=" diese=" drei=" nachweismethoden=" existieren=" bereits=" europäische=" normen=" und=" sie=" sind=" als=">Allgemeine Codex Methoden zum Nachweis bestrahlter Lebensmittel etabliert. Die Ergebnisse zeigen, dass PSL und TL geeignete Verfahren sind, um sowohl eine Behandlung von Mais mit niederenergetischen als auch mit hochenergetischen Elektronen zu erkennen. Der DNA-Kometentest erwies sich als weniger geeignet: die Behandlung mit niederenergetischen Elektronen konnte—wie erwartet—nicht nachgewiesen werden. Die Behandlung mit hochenergetischen Elektronen konnte bei einer Maissorte erkannt werden, jedoch nicht bei einer anderen Sorte.     

Participation of Rhipicephalus sanguineus (Acari: Ixodidae) in the epidemiology of canine visceral leishmaniasis

The vectorial competence of the tick Rhipicephalus sanguineus is discussed in relation to the epidemiology of canine visceral leishmaniasis, taking into account its strict association with dogs and the low indices of natural infection presented by its known vector, the phlebotomine sand fly Lutzomyia longipalpis. In order to evaluate natural infection by Leishmania chagasi and the infectivity of these parasites in the tick, 39 specimens (6 females, 11 males and 22 nymphs) of R. sanguineus were removed from 21 dogs showing diverse symptoms of zoonotic visceral leishmaniasis (ZVL). Six ticks (15.4%) gave positive results for the genus Leishmania using the PCR technique. To determine the infectivity of the parasites, 36 hamsters were inoculated orally and peritoneally with macerates of ticks removed from nine dogs symptomatic for visceral leishmaniasis. After 6 months the hamsters were sacrificed and necropsied. Serum was removed for IFAT, as well as spleen and liver fragments to make imprint smears and for PCR. Eight (88.9%) of these dogs presented ticks that were infective for 14 hamsters (41.2%), 12 (85.7%) of them infected peritoneally and two (14.3%) orally. PCR revealed 27 smears (40.9%) to be positive, 20 (62.5%) of them infected peritoneally and seven (20.6%) orally. IFAT showed 14 positive animals (41.2%). Based on these findings, we suggest that the vectorial capacity of R. sanguineus for L. chagasi should be evaluated further, opening new perspectives in the epidemiology of ZVL.     

Osteosarcoma with multiple metastases and subcutaneous involvement in a rabbit (Oryctolagus cuniculus)

BACKGROUND: Osteogenic sarcomas are rare in rabbits, but cases involving the mandible, ribs, frontal bones, and tibia have been reported. OBJECTIVE: The purpose of this study was to describe the clinical and pathologic findings in a rabbit with osteosarcoma and multiple metastases with subcutaneous involvement. METHODS: A 1-year-old female Polish rabbit was evaluated for dysorexia, lameness, gait difficulties with the hind limbs, and the presence of 2 subcutaneous nodules, on the left thigh and in the dorsal region. A hard swelling was also present at the base of the tail. Radiographic findings revealed multiple abnormalities in several bones, which were more severe in the sacrococcygeal region and femur. Several areas of calcification involving the thorax and abdomen as well as skeletal muscles, also were noted. The thigh nodule was surgically removed, and cytologic and histologic evaluations were performed. Following the rabbit's death, postmortem radiographs were taken and necropsy was performed. Samples of tissues and organs, involved bone segments, and other subcutaneous nodules were collected and examined by cytology and histology. RESULTS: Both the subcutaneous nodules and the majority of tissues and organs contained neoplastic cells whose appearance was consistent with osteogenic sarcoma. CONCLUSION: Clinical, radiologic, and pathologic data supported a diagnosis of osteosarcoma with multiple metastases and subcutaneous involvement.     

High-frequency ultrasonography of the skin of clinically normal dogs

OBJECTIVE: To assess the applicability of high-frequency diagnostic ultrasonography for evaluation and accurate measurement of the skin thickness of clinically normal dogs. ANIMALS: 26 healthy dogs (12 sexually intact males, 13 sexually intact females, and 1 spayed female) of various breeds and ages. PROCEDURE: Ultrasonographic examination of the skin and histomorphometric analysis of skin biopsy specimens obtained from the same site were performed. A 13-MHz linear-array transducer was used to obtain a series of ultrasonographic images of the skin in the flank region; images were analyzed and measured by use of imaging software. Cutaneous biopsy specimens were placed in fixative and then stained with H&E and Masson trichrome stains. Histomorphometric analysis was performed by use of an image analyzer. Thickness of the epidermis and dermis of each specimen was evaluated by use of a semiautomatic procedure of quantification. Data obtained from ultrasonographic and histologic measurements were compared by use of the Pearson correlation test. RESULTS: The ultrasonographic pattern of canine skin was consistently characterized by 3 distinct, defined echogenic layers corresponding to the epidermal entry echo, epidermis and dermis, and subcutaneous tissues. A positive correlation was found between ultrasonographic and histologic measurements of skin thickness. CONCLUSIONS AND CLINICAL RELEVANCE: Comparison between ultrasonographic and histologic appearance of the skin revealed that layering of canine skin (ie, epidermis and dermis) and the subcutaneous tissues may be recognized and measured by use of high-frequency ultrasonography. Thus, diagnostic ultrasonography may be a useful tool for the noninvasive evaluation of cutaneous disorders in dogs.     

Residue levels and effectiveness of pyrimethanil vs imazalil when using heated postharvest dip treatments for control of Penicillium decay on citrus fruit

The influence of fungicide concentration and treatment temperature on residue levels of pyrimethanil (PYR) in comparison with the commonly used fungicide imazalil (IMZ) was investigated in orange fruits following postharvest dip treatments. The dissipation rate of PYR residues was recorded as a function of storage conditions. The fungicide efficacy against green and blue molds caused by Penicillium digitatum and Penicillium italicum, respectively, was evaluated on different citrus varieties following the fungicide application at 20 or 50 degrees C. Residue levels of PYR in Salustiana oranges were significantly correlated with the fungicide dosage, but residue concentrations were notably higher (ca. 13-19-fold) after treatment at 50 degrees C as compared to treatments at 20 degrees C. After treatment at temperatures ranging from 20 to 60 degrees C, PYR and IMZ residues in Salustiana oranges were significantly correlated with dip temperatures. Dissipation rates of PYR during storage were negligible in both Salustiana and Tarocco oranges. Results obtained on wounded, noninoculated Miho satsumas revealed that when treatments were performed at 50 degrees C, PYR or IMZ concentrations needed to achieve the complete control of decay were 8- and 16-fold less than by treatment at 20 degrees C. When fruits were inoculated with either P. digitatum or P. italicum, the application of 400 mg L(-1) PYR at 20 degrees C or 100 mg L(-1) PYR at 50 degrees C similarly reduced green and blue mold development. These results were corroborated by storage trials on Marsh grapefruits and Tarocco oranges. The lowest concentration of PYR required to achieve almost total protection of the fruit against decay accounted for 100 mg L(-1) at 50 degrees C and 400 mg L(-1) at 20 degrees C, respectively. Treatments did not affect fruit external appearance, flavor, and taste. It is concluded that postharvest PYR treatment represents an effective option to control green and blue mold in citrus fruit and that integration of fungicide applications and hot water dips may reduce the possibility of selecting fungicide-resistant populations of the pathogen, as a consequence of increased effectiveness of the treatment.     

Genetic analysis and molecular characterisation of laboratory and field mutants of Botryotinia fuckeliana (Botrytis cinerea) resistant to QoI fungicides

BACKGROUND: QoI fungicides, inhibitors of mitochondrial respiration, are considered to be at high risk of resistance development. In several phytopathogenic fungi, resistance is caused by mutations (most frequently G143A) in the mitochondrial cytochrome b (cytb) gene. The genetic and molecular basis of QoI resistance were investigated in laboratory and field mutants of Botryotinia fuckeliana (de Bary) Whetz. exhibiting in vitro reduced sensitivity to trifloxystrobin. RESULTS: B. fuckeliana mutants highly resistant to trifloxystrobin were obtained in the laboratory by spontaneous mutations in wild‐type strains, or from naturally infected plants on a medium amended with 1–3 mg L^?1 trifloxystrobin and 2 mM salicylhydroxamic acid, an inhibitor of alternative oxidase. No point mutations were detected, either in the complete nucleotide sequences of the cytb gene or in those of the aox and Rieske protein genes of laboratory mutants, whereas all field mutants carried the G143A mutation in the mitochondrial cytb gene. QoI resistance was always maternally inherited in ascospore progeny of sexual crosses of field mutants with sensitive reference strains. CONCLUSIONS: The G143A mutation in cytb gene is confirmed to be responsible for field resistance to QoIs in B. fuckeliana. Maternal inheritance of resistance to QoIs in progeny of sexual crosses confirmed that it is caused by extranuclear genetic determinants. In laboratory mutants the heteroplasmic state of mutated mitochondria could likely hamper the G143A detection, otherwise other gene(s) underlying different mechanisms of resistance could be involved. Copyright © 2012 Society of Chemical Industry     

Evaluation of the antifungal susceptibility of Malassezia pachydermatis to clotrimazole, miconazole and thiabendazole using a modified CLSI M27-A3 microdilution method

Background – In this study, we evaluated the antifungal susceptibility of Malassezia pachydermatis to clotrimazole (CTZ), miconazole (MCZ), and thiabendazole (TBD), azole derivatives employed in aural formulations labeled for treatment of canine otitis. Methods – The procedure for in vitro testing was based on the indications of the Clinical and Laboratory Standards Institute (CLSI) M27‐A3 microdilution method. A lipid‐enriched medium was employed to enhance the yeast growth (Christensen’s urea broth, with 0.1% Tween 80 and 0.5% Tween 40 as the lipid sources), while the inoculums size corresponded to approximately 1–5 × 10⁵ yeast cells/mL. Microplates were incubated at 37°C and read 48 h after inoculation. Azole MICs inhibiting fungal growth were the lowest drug concentrations that showed an optical density of ≤50% of the (drug‐free) growth control, as assessed by spectrophotometer (630 nm filter). Results – All isolates were inhibited by the three azoles, with different minimum inhibitory concentration (MIC) values. Most isolates were inhibited by drug concentrations of 2–8 (CTZ), 1–4 (MCZ), or 16–32 (TBD) μg/mL. These results are partially in agreement with the findings of previous studies, in which substantially higher/lower MICs were occasionally reported. This is likely because of the different methodologies employed. Such discrepancies may not apply to clinical situations, where the compounds are applied topically. Conclusion and clinical importance – The concept that clinical failure is linked to increased MICs is debatable, because significantly higher concentrations (in most cases at least 1,000 × the MIC) of the antifungals that were included in our study are routinely used in formulated products.     

Characterization of In Vitro Synthesized Equine Metabolites of the Selective Androgen Receptor Modulators S24 and S4

Several selective androgen receptor modulators (SARMs) have been synthesized and investigated in humans, rats, and dogs in the past, but no data are yet available concerning the metabolism of SARMs in horses. The aryl-propionamide-derived drug candidates S24 and S4 (andarine) have a strong androgen receptor binding affinity and show distinctive specific cell answers. Although no SARM drug candidate (aiming for testosterone replacement therapy) has completed clinical trials yet, S4 has been illicitly available via the Internet. These facts led to the prohibition of SARMs by the German equestrian federation, and the (mis)use of such compounds would further represent a doping rule violation in horse racing. In this study, the drug candidates S24 and S4 were subjected to in vitro metabolism experiments with equine liver microsomal preparations from a female Quarter Horse to obtain information about potential target analytes in equine doping control analysis. The enzymatically synthesized metabolites were characterized by liquid chromatography–tandem mass spectrometry and –high-resolution/high-accuracy mass spectrometry. All observed S24 and S4 equine metabolites are in agreement with earlier in vitro and in vivo studies in humans and dogs. Nevertheless, the relative percentage of generated equine metabolites (as determined from the analytes’ response in full-scan chromatography–tandem mass spectrometry and –high-resolution/high-accuracy mass spectrometry measurements) differs considerably from the reported profiles. Although the S24 metabolite pattern is comparably balanced concerning glucuronidated and sulfonated conjugates, the major S4 metabolite was found to be the unconjugated dephenylated compound, with a proportion of more than 90%.     

Digital Agenesia in Martina Franca Donkey Foal: A Case Report

A 4-day-old male Martina Franca donkey foal was evaluated for a forelimb alteration. Clinical examination and radiographs revealed the agenesia of the distal digit. Biochemical parameters were normal, and ultrasonographic evaluation did not identify any relievable organ alteration. Karyotype study revealed an abnormality on chromosome 1. The foal was discharged with a distal limb bandage in which a palmar splint was applied. A poor prognosis for the functionality of the limb was given. In endangered species, such as the Martina Franca donkey, the excessive inbreeding could result in an increase in genetic disorders. These findings shed new light on the possible pathogenesis of the digital dysgenesia. The study of the karyotype could be a useful approach to detect genetic alterations that could or could not be expressed in the animal, especially in endangered species in which a risk of an excessive inbreeding is considerable. These defects should be considered in the choice and selection of the breeders.