Untersuchungen zur Übertragung von Clavibacter michiganensis ssp. sepedonicus auf gesunde Kartoffelknollen

Zusammenfassung In den Jahren 1999 bis 2003 wurde in Freiland-, Klimakammer- und Lagerungsversuchen überprüft, ob ein Risiko für die Übertragung des Erregers der Bakteriellen Ringfäule der Kartoffel (Clavibacter michiganensis ssp. sepedonicus) besteht, wenn (a) gesunde Kartoffelknollen in Kontakt mit Maschinen und Geräten kommen, die mit dem Erreger kontaminiert sind (indirekter Kontakt) und (b) gesunde Kartoffelknollen direkt in Kontakt mit infizierten Knollen kommen (direkter Kontakt). Nach indirektem Kontakt konnte nur beim nachfolgenden Anbau der kontaminierten Knollen in der Klimakammer Befall in Kraut und Knollen festgestellt werden. Im Freiland konnte der Erreger, auch bei wiederholtem Nachbau der geernteten Knollen, nicht nachgewiesen werden. Nach direktem Kontakt und nachfolgendem Anbau der kontaminierten Knollen in der Klimakammer und im Freiland, wurde der Erreger in allen Fällen in den geerntete Knollen nachgewiesen. Befall im Kraut wurde nur in dem Klimakammerversuch und in einem Freilandversuch ermittelt. Wurden durch direkten Kontakt kontaminierte Knollen eingelagert, konnte der Erreger in allen untersuchten Knollen festgestellt werden. Insgesamt besteht ein hohes Risiko, dass gesunde Knollen infiziert werden, wenn oberflächliche Kontaminationen mit dem Erreger erfolgen. Die Wahrscheinlichkeit von Infektionen steigt mit zunehmender Kontaminationsstärke.     

Überleben von Ralstonia solanacearum Biovar 2 (Rasse 3), dem Erreger der Schleimkrankheit der Kartoffel, in Boden und auf verschiedenen Materialien (Mikrokosmos)

Microcosm studies were carried out to test the survival of Ralstonia solanacearum biovar 2 (race 3) in soil at the permanent wilting point (wp) water content and at field capacity (fc) water content and on various material. Soils were placed at permanent ?5°C, 4°C, 15°C and 20°C and weekly fluctuating ?10/0/+10°C and the material at 5, 15 °C, 20°C with relative humidity (rh) uncontrolled or at constant 10% or 90%. In soil, survival was clearly dependent on temperature independent of water content. At 20°C Ralstonia solanacearum could be reisolated up to 364 days, at 15°C up to 290 days, at 4°C up to 209 days and at fluctuating temperatures (?10/0/+10°C) only up to 18 days. The lower the temperature, the more the population declined. At 15°C and 20°C appr. 10⁷ cfu/g soil were detected after 100 days, whereas at ?5°C only 10² cfu/g soil were detected after only 18 days. The pathogen was longer detectable in sandy-clay loam than in lighter sandy soil. It could be longer reisolated at wilting point and the populations did not decline as rapidly as at field capacity. Ralstonia solanacearum could best survive on material surfaces like rubber, plastic and varnished metal with maximum survival of 40 days at 5°C and 10% rh. In general there is a low risk of Ralstonia solanacearum overwintering under European climatic conditions when the fields are cleared of plant debris and the soil is frozen. Contamined material surfaces pose the risk of pathogen transmission to healthy tubers.     

Expression of porcine endogenous retroviruses (PERVs) in melanomas of Munich miniature swine (MMS) Troll

Porcine endogenous retroviruses (PERVs) are integrated in the genome of all pig breeds. Since some of them are able to infect human cells, they might represent a risk for xenotransplantation using pig cells or organs. However, the expression and biological role of PERVs in healthy pigs as well as in porcine tumours is largely unknown. Since we and others have recently shown overexpression of a human endogenous retrovirus, HERV-K, in human melanomas, we studied the expression of PERVs in melanomas of selectively bred Munich miniature swine (MMS) Troll. This breeding herd of MMS Troll is characterised by a high prevalence of melanomas, which histologically resemble various types of cutaneous melanomas in humans. Several genetic factors have been defined when studying inheritance of melanomas and melanocytic nevi in MMS Troll. Here we show that the polytropic PERV-A and PERV-B as well as the ecotropic PERV-C are present in the genome of all melanoma bearing MMS Troll investigated. Most interestingly, in the spleen, but not in other organs, recombinant PERV-A/C proviruses were found. PERV expression was found elevated in melanomas when compared to normal skin and viral proteins were expressed in melanomas and pulmonary metastasis-derived melanoma cell cultures. During passaging of these cells in vitro the expression of PERV mRNA and protein increased and virus particles were released as shown by RT activity in the supernatant and by electron microscopy. Genomic RNA of PERV-A, -B and -C were found in pelleted virus particles. Although PERV expression was elevated in melanomas and pulmonary metastasis-derived cell cultures, the function of the virus in tumour development is still unclear.     

Photodynamic therapy of feline cutaneous squamous cell carcinoma using a newly developed liposomal photosensitizer: preliminary results concerning drug safety and efficacy

BACKGROUND: Squamous cell carcinomas are common skin tumors in cats. We investigated photodynamic therapy (PDT) using a new liposomal photosensitizer as a minimally invasive, effective treatment that can be easily performed while achieving good cosmetic results. AIM: The goal of this study was to assess and describe possible toxicities using a liposomal formulation of the photosensitizer meta-(Tetrahydroxyphenyl)Chlorin (m-THPC) and investigate if favorable pharmacokinetics translate into favorable tumor response and control. ANIMALS: Eighteen client-owned cats with 20 spontaneous cutaneous squamous cell carcinomas were included in the study. METHODS: PDT was performed using a new, liposomal formulation of the photosensitizer. Toxicity, tumor response, and tumor control were evaluated retrospectively. RESULTS: No general adverse effects were observed in cats treated with the new liposomal formulation. Mild local toxicity such as erythema and edema were seen in 15% of the patients. All cats responded to therapy, with a complete response rate of 100%. The overall 1-year control rate was 75%. The tumor recurrence rate was 20% with a median time to recurrence of 172.25 +/-87.1) days. CONCLUSIONS AND CLINICAL IMPORTANCE: A new liposomal photosensitizer was successfully used for squamous cell carcinoma in cats and was well tolerated. There were no systemic adverse effects observed with the liposomal formulation. The favorable pharmacokinetics of the liposomal drug resulted in a favorable tumor response.     

Extracellular heat shock protein 70 levels in tumour-bearing dogs and cats treated with radiation therapy and hyperthermia

Hyperthermia is a form of a cancer treatment which is frequently applied in combination with radiotherapy (RT) to improve therapy responses and radiosensitivity. The mode of action of hyperthermia is multifactorial; the one hand by altering the amount of the blood circulation in the treated tissue, on the other hand by modulating molecular pathways involved in cell survival processes and immunogenic interactions. One of the most dominant proteins induced by hyperthermia is the major stress-inducible heat shock protein 70 (Hsp70). Hsp70 can be found in the blood either as a free-protein (free HSP70) derived from necrotic cells, or lipid-bound (liposomal Hsp70) when it is actively released in extracellular vesicles (EVs) by living cells. The aim of the study was to evaluate the levels of free and liposomal Hsp70 before and after treatment with RT alone or hyperthermia combined with radiotherapy (HTRT) in dogs and cats to evaluate therapy responses. Peripheral blood was collected from feline and canine patients before and at 2, 4, 6 and 24 h after treatment with RT or HTRT. Hsp70 enzyme-linked immunosorbent assays (ELISAs) were performed to determine the free and liposomal Hsp70 concentrations in the serum. The levels were analysed after the first fraction of radiation to study immediate effects and after all applied fractions to study cumulative effects. The levels of free and liposomal Hsp70 levels in the circulation were not affected by the first singular treatment and cumulative effects of RT in cats however, after finalizing all treatment cycles with HTRT free and liposomal Hsp70 levels significantly increased. In dogs, HTRT, but not treatment with RT alone, significantly affected liposomal Hsp70 levels during the first fraction. Free Hsp70 levels were significantly increased after RT, but not HTRT, during the first fraction in dogs. In dogs, on the other hand, RT alone resulted in a significant increase in liposomal Hsp70, but HTRT did not significantly affect the liposomal Hsp70 when cumulative effects were analysed. Free Hsp70 was significantly induced in dogs after both, RT and HTRT when cumulative effects were analysed. RT and HTRT treatments differentially affect the levels of free and liposomal Hsp70 in dogs and cats. Both forms of Hsp70 could potentially be further investigated as potential liquid biopsy markers to study responses to RT and HTRT treatment in companion animals.     

Assessing spillover from marine protected areas and its drivers: A meta‐analytical approach

Overfishing may seriously impact fish populations and ecosystems. Marine protected areas (MPAs) are key tools for biodiversity conservation and fisheries management, yet the fisheries benefits remain debateable. Many MPAs include a fully protected area (FPA), restricting all activities, within a partially protected area (PPA) where potentially sustainable activities are permitted. An effective tool for biodiversity conservation, FPAs, can sustain local fisheries via spillover, that is the outward export of individuals from FPAs. Spillover refers to both: “ecological spillover”: outward net emigration of juveniles, subadults and/or adults from the FPA; and “fishery spillover”: the fraction of ecological spillover that directly benefits fishery yields and revenues through fishable biomass. Yet, how common is spillover remains controversial. We present a meta‐analysis of a unique global database covering 23 FPAs worldwide, using published literature and purposely collected field data, to assess the capacity of FPAs to export biomass and whether this response was mediated by specific FPA features (e.g. size, age) or species characteristics (e.g. mobility, economic value). Results show fish biomass and abundance outside FPAs was higher: (a) in locations close to FPA borders (<200 m) than further away (>200 m); (b) for species with a high commercial value; and (c) in the presence of PPA surrounding the FPA. Spillover was slightly higher in FPAs that were larger and older and for more mobile species. Based on the broadest data set compiled to date on marine species ecological spillover beyond FPAs' borders, our work highlights elements that could guide strategies to enhance local fishery management using MPAs.     

Short-term storage and cryopreservation of milt from Atlantic salmon and sea trout

Experiments on short-term preservation of sperm were performed with Atlantic salmon (Salmo salar). Fertility was maintained for up to 10 days when 2 mm thick samples were stored at 0° C under an oxygen atmosphere in the presence of antibiotics (125 IU penicillin and 125 μg streptomycin per ml sperm). Fertility was completely lost after 24 days. Sperm stored without antibiotics fertilized 100% of eggs after 6 days.

Cryopreservation was carried out with milt from Atlantic salmon and sea trout (Salmo trutta). Semen mixed with extender was frozen on dry ice (pellets) with subsequent storage in liquid nitrogen. Sperm pellets were thawed in a 0.12-M NaHCO₃-solution (10° C) before insemination. The suitability of an extender as previously described by Stoss and Holtz and of a 0.3-M glucose solution with the addition of 10% DMSO, was tested on two different batches of sperm and eggs in Atlantic salmon and sea trout. In addition, the extender earlier reported by Mounib and an aqueous solution of 10% DMSO were only used in Atlantic salmon with one batch of gametes.

Insemination with cryopreserved Atlantic salmon sperm resulted in 36 to 91.3% eyed eggs (control = 100). The differences were caused by the type of extender and the batch of gametes employed. The very simple extender consisting of 0.3 M glucose and 10% DMSO only was the most successful. Results with cryopreserved sea trout sperm ranged between 38.6–54.8% eyed eggs, showing no difference between treatments.     

Is N‐feedback involved in the regulation of nitrogenase activity in Medicago truncatula?

To determine whether senescing leaves provoke an active nitrogen (N) remobilization that results in the reduction of nitrogenase activity, 60% of Medicago truncatula lower leaves were either darkened or individually excised for two weeks. Although a considerable amount of N was remobilized, N₂ fixation activity was found to be increased to maintain the N source/sink balance, indicating an absence of the negative N‐feedback regulation of nitrogenase activity in the senescing M. truncatula.     

Effect of mineral nitrogen fertilizer forms on N2O emissions from arable soils in winter wheat production

Nitrogen fertilizers are supposed to be a major source of nitrous oxide (N₂O) emissions from arable soils. The objective of this study was to compare the effect of N forms on N₂O emissions from arable fields cropped with winter wheat (Triticum aestivum L.). In three field trials in North‐West Germany (two trials in 2011/2012, one trial in 2012/2013), direct N₂O emissions during a one‐year measurement period, starting after application of either urea, ammonium sulfate (AS) or calcium ammonium nitrate (CAN), were compared at an application rate of 220 kg N ha^?1. During the growth season (March to August) of winter wheat, N₂O emission rates were significantly higher in all three field experiments and in all treatments receiving N fertilizer than from the non‐fertilized treatments (control). At two of the three sites, cumulative N₂O emissions from N fertilizer decreased in the order of urea > AS > CAN, with emissions ranging from 522–617 g N ha^?1 (0.24–0.28% of applied fertilizer) for urea, 368–554 g N ha^?1 (0.17–0.25%) for AS, and 242–264 g N ha^?1 (0.11–0.12%) for CAN during March to August. These results suggest that mineral nitrogen forms can differ in N₂O emissions during the growth period of winter wheat. Strong variations in the seasonal dynamics of N₂O emissions between sites were observed which could partly be related to weather events (e.g., precipitation). Between harvest and the following spring (post‐harvest period) no significant differences in N₂O emissions between fertilized and non‐fertilized treatments were detected on two of three fields. Only on one site post‐harvest emissions from the AS treatment were significantly higher than all other fertilizer forms as well as compared to the control treatment. The cumulative one‐year emissions varied depending on fertilizer form across the three field sites from 0.05% to 0.51% with one exception at one field site (AS: 0.94%). The calculated overall fertilizer induced emission averaged for the three fields was 0.38% which was only about 1/3 of the IPCC default value of 1.0%.