Ugent faculteit diergeneeskunde, Vakgroep Pathologie, Bacteriologie en Pluimvee
Abstract bachelor project FBT 2020-21: Diagnosis of pathogens in hedgehogs by means of microbiological and molecular research
Several hedgehogs in animal shelters in Geraardsbergen, Ostend and Merelbeke have died of an unknown cause. Previous studies have proven that Corynebacterium ulcerans could be detected in 38 hedgehogs and that this pathogen is the primary cause of death. Corynebacterium ulcerans causes leases in wounds.
This study will investigate if there were any other pathogens present in the hedgehogs, such as Salmonella, Borrelia, Leptospira and Herpes. Also, the correlation between these pathogens and C. ulcerans will be considered. In total one hundred ninety hedgehogs have been transferred from the animal shelters to the lab. In the lab several tissue samples were taken. To examine the presence of Salmonella, the small intestines will be enriched in a non-selective medium and transferred to a selective medium. After this a selective isolation will be performed so colonies can grow. These colonies will eventually be identified by MALDI-TOF spectrometry. The presence of Herpes will be detected in the liver and brains by Nested-PCR. Leptospira will be detected in the kidneys by using PCR. The presence of Borrelia will be detected in the ears, heart and urinary bladder by performing qPCR.
The study has proven Salmonella to be the most common pathogen to be present in the hedgehogs, after C. ulcerans. The other pathogens are less common. Although Salmonella is the most common pathogen, it is not correlated to C. ulcerans because only one hedgehog has both infections. To conclude, all of the infections are coincidental and none are explicitly correlated to C. ulcerans.
Abstract bachelor project FBT 2019-20: The effects of corticosterone on the infectivity of Batrachochytrium salamandrivorans on A6-cells
Since the late 1970’s, herpetologists started to observe unusual disappearances of amphibian populations in protected areas. Because the data was not of high quality and long-term census, it was hard to determine if the data was representing normal fluctuation or true declines. After a lot of research, scientists found a new pathogen that caused these declines. It was called Batrachochytrium dendrobatidis. Later, a new decline was observed in several populations of fire salamanders. Attempts to identify known pathogens resulted in all samples negative. Therefore further research was conducted. The research revealed a new highly pathogenic chytrid fungi that was called Batrachochytrium salamandrivorans. To date, not much is known about this second fungi. That is why the aim of this study is to see what effects corticosterone has on the infectivity of the chytrid fungi. Corticosterone increases due to physiological stress and is in charge for altering the host’s physiology and its susceptibility to a pathogen. In order to test the effects A6-cells were planted in a 96-wellplate and grown in medium supplemented with corticosterone. The corticosterone is originally dissolved in ethanol, therefore the solvent is also tested in this set-up due to its toxicity. After a few days, a cytotoxicity test can be executed. The neutral red test will be used. Due to the lack of time, because of the COVID-19 outbreak, other set-ups to test the effects on the chytrid fungus and its infectivity were not executed. So only a few results were obtained from the cytotoxicity tests. At first sight the results shows a higher percent of viability at higher corticosterone levels than the control group. It also shows a normal to slightly lower percentages at normal and low corticosterone levels. The control of the solvent shows the opposite. There are higher values at normal and low corticosterone levels and show a normal survival rate at high corticosterone levels. Because this test was only performed once, it can not be assumed that these are true values. Further statistics show no differences between the ethanol and corticosterone group, this means that corticosterone is not the factor that shows the results of this test. The experiment must be executed several more times to avoid coincidence, and there must be searched for an alternative solvent. This solvent must have the ability to solve corticosterone and be non-toxic to the cells. It is also possible to dilute the solvent more. Further research can give more information about the interactions of B. salamandrivorans with its host, the effects of corticosterone and other mediators such as host factors and environmental factors.
Abstract bachelor project FBT 2018-19: BATRACHOCHYTRIUM DENDROBATIDIS ON THE 'WILD ALPINE NEWTS'
Batrachochytrium dendrobatidis is a major pathogen of frogs worldwide, associated with declines in amphibian populations. The skin disease caused by this fungus is named chytridiomycosis and affects the vital function of amphibian skin. Not all amphibians respond equally to infection and host responses might range from resistant, over tolerant to susceptible. The clinical outcome of infection is highly dependent on the amphibian host, the fungal virulence and environmental determinants. B. dendrobatidis infects the skin of a large range of anurans, urodeles and caecilians. The aim of this study is to identify the impact of B. dendrobatidis infections in wild alpine newts . Data for this study were obtained from alpine newts that were trapped in fikes. This screening was done at different time points and consisted of a sampling, DNA extraction, preparation for the final qPCR and the qPCR. The results show that there’s only 1 % of the samples positive for B. dendrobatidis. This reveals that the prevalence of the fungus is very low in East Flanders. For the future, there must be keeping an eye on the B. dendrobatidis for further dispersion.
Abstract bachelor project FBT 2016-17: Optimization of PCR with specific primers for SNP detection in fire salamanders
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Samenvatting eindwerk 2014-2015: Screening on lethal chytridiomycosis through quantitative PCR on wild living amphibians
Chytridiomycosis is an infection caused by two different fungi, Batrachochytrium dendrobatidis (Bd) or Batrachochytrium salamandrivorans (Bs). Both fungal species infect amphibians. Bs is more isolated and characterized from salamanders and Bd from frogs / toads.
About ten years ago Bd was isolated for the first time from frog skin and described. This fungus causes epidermal hyperplasia and hyperkeratosis of the amphibian skin. In 2012, another related species was discovered, the salamander-infecting fungus Bs. This fungus causes especially deep skin ulcers.
Both fungi interfere with the physiological functioning of the amphibian skin, whereby they eventually cause in most cases, death of the amphibian. The fast and easy spread of these fungi lately induces a large number of infections. There is also observing a huge decline and even extinction of some amphibian populations. Therefore chytridiomycose is one of the greatest threats to biodiversity.
Nowadays, it is important to screen as many as possible amphibians both living and dead, of the presence of these fungi. Thanks to this kind of screening more information can be discovered about the lethal chytridiomycosis. Such screenings are done through quantitative PCR. At this point, it can’t be avoided that these fungi cause extinction of some wild amphibian populations.
This screening consisted of a sampling, DNA extraction, preparation for the final qPCR and the qPCR. The different screenings were performed according to different sampling methods and DNA extractions.
First, 28 wild yellow-bellied toads were found during different periods screened on the Bd fungus. This has shown that there was only one toad positive. However, it was in fact a small dose, and it was not certain whether the toad was recently infected or whether it was already in a recovery phase. The toad wasn’t found later, there is no certainty that this animal has survived the infection. The next screenings were performed on amphibians that were found in the wild death. These screenings are especially important to quickly detect an infection outbreak. Both dead newt and several ordinary toads were screened. None of these animals had chytridiomycose as cause of death.
These screenings could clarify why and how a chytridiomycosis outbreak takes place specifically on these locations. Also changes in infection status from some animals can be checked. Because of this, a solution to this problem could eventually be found on a long-term perspective.
Prof. Ann Martel