Project description:IntroductionThe bacterium Elizabethkingia miricola is a multispecies pathogen associated with meningitis-like disease that has been isolated from several amphibian species, including the bullfrog, but this is the first isolation in Guangxi. In the present study, the dominant bacteria were isolated from the brains of five bullfrogs with meningitis-like disease on a South China farm in Guangxi.MethodsThe NFEM01 isolate was identified by Gram staining; morphological observations; 16S rRNA, rpoB, and mutT-based phylogenetic tree analysis; and physiochemical characterization and was subjected to drug sensitivity and artificial infection testing.Results and discussionAs a result of identification, the NFEM01 strain was found to be E. miricola. An artificial infection experiment revealed that NFEM01 infected bullfrogs and could cause symptoms of typical meningitis-like disease. As a result of the bacterial drug sensitivity test, NFEM01 is highly sensitive to mequindox, rifampicin, enrofloxacin, nitrofural, and oxytetracycline and there was strong resistance to gentamicin, florfenicol, neomycin, penicillin, amoxicillin, doxycycline, and sulfamonomethoxine. This study provides a reference to further study the pathogenesis mechanism of E. miricola-induced bullfrog meningitislike disease and its prevention and treatment.
Project description:BackgroundViruses included in the family Iridoviridae are large, icosahedral, dsDNA viruses that are subdivided into 5 genera. Frog virus 3 (FV3) is the type species of the genus Ranavirus and the best studied iridovirus at the molecular level. Typically, antibodies directed against a virus act to neutralize the virus and limit infection. Antibody dependent enhancement occurs when viral antibodies enhance infectivity of the virus rather than neutralize it.ResultsHere we show that anti-FV3 serum present at the time of FV3 infection enhances infectivity of the virus in two non-immune teleost cell lines. We found that antibody dependent enhancement of FV3 was dependent on the Fc portion of anti-FV3 antibodies but not related to complement. Furthermore, the presence of anti-FV3 serum during an FV3 infection in a non-immune mammalian cell line resulted in neutralization of the virus. Our results suggest that a cell surface receptor specific to teleost cell lines is responsible for the enhancement.ConclusionsThis report represents the first evidence of antibody dependent enhancement in iridoviruses. The data suggests that anti-FV3 serum can either neutralize or enhance viral infection and that enhancement is related to a novel antibody dependent enhancement pathway found in teleosts that is Fc dependent.
Project description:Klebsiella pneumoniae is a major cause of nosocomial infection and is considered a clinically important bacterium with antibiotic-resistant strains. There are few reports of K. pneumoniae infections in cultured aquatic animals, and no natural infection has been reported in amphibians. From September to October 2021, a high-mortality disease outbreak occurred in a pond-raised American bullfrog farm in Guangzhou, China. The infected bullfrogs were characterized by multiple organ congestive enlargement and inflammation. A pathogenic bacterium was isolated from the viscera of infected bullfrogs and confirmed to be K. pneumoniae by morphological, biochemical, and phylogenetic analyses. Infection experiments confirmed the virulence of the pathogenic strain against bullfrogs and tadpoles. A histopathological examination showed that the strain was harmful to multiple organs. Antibiotic resistance experiments indicated the isolate was a carbapenemase-producing multidrug-resistant K. pneumoniae (MDR-KP) strain. This study is the first report of K. pneumoniae infected American bullfrogs (Rana catesbeiana) and amphibians. These results will shed light on the pathogenicity of K. pneumoniae and help prevent and control K. pneumoniae infections in bullfrogs. IMPORTANCE Klebsiella pneumoniae is recognized as the most common multidrug-resistant bacterial pathogen in humans, and little is known about its pathogenicity in aquatic animals. Recently, K. pneumoniae was found to cause substantial mortality and morbidity in American farm frogs. This was the first report of K. pneumoniae infecting amphibians. In this study, we analyzed the biochemical, growth, and phylogenetic characteristics of the K. pneumoniae strain and described the symptoms and pathological features of infected bullfrogs and tadpoles; this will provide useful data for the prevention and control of infectious diseases, which has been suggested to decrease economic losses in bullfrog farming and reduce the potential threat to public health posed by K. pneumoniae.
Project description:Ranaviruses (family Iridoviridae) cause important diseases in cold-blooded vertebrates. In addition, some occurrences indicate that, in this genus, the same virus can infect animals from different taxonomic groups. A strain isolated from a Ranavirus outbreak (2012) in the state of Sao Paulo, Brazil, had its genome sequenced and presented 99.26% and 36.85% identity with samples of Frog virus 3 (FV3) and Singapore grouper iridovirus (SGIV) ranaviruses, respectively. Eight potential recombination events among the analyzed sample and reference FV3 samples were identified, including a recombination with Bohle iridovirus (BIV) sample from Oceania. The analyzed sample presented several rearrangements compared to FV3 reference samples from North America and European continent. We report for the first time the complete genome of Ranavirus FV3 isolated from South America, these results contribute to a greater knowledge related to evolutionary events of potentially lethal infectious agent for cold-blooded animals.
Project description:These data and analyses support the research article "Culture, cryobanking and passaging of karyotypically validated native Australian amphibian cells" Mollard (2018) [1]. The data and analyses presented here include: (1) three additional karyomaps of cells from the cryobanked and passaged frog and tadpole species Litoria infrafrenata; and (2) combined short-to-long arm ratios of the four karyomaps measured from each respective animal here and in Ref [1].
Project description:Invasive species pose a major threat to global biodiversity. The effects of invasive species can be strongly influenced and potentially mediated by their reproductive characteristics, such as fecundity, egg production, and duration and number of reproductive events. Selection for smaller body size at first reproduction can also play a role in their establishment, facilitating colonization and spread. The American bullfrog, native to the eastern U.S. (Lithobates catesbeianus), is a species that has invaded more than 40 countries across 4 continents. This species has become especially prevalent in the western United States since its introduction in the early 1900s. This study characterized reproductive characteristics of bullfrogs with emphasis on the minimum size at which males and females reach sexual maturity in the Willamette Valley, Oregon, USA invasion range. We collected and dissected 121 individuals in 2013 and 2017, quantifying characteristics of sexual maturity including snout-vent length, total length, sex, tympanum diameter, presence of distended oviducts or eggs for females, and testes length and sperm activity in males. Our results showed that the minimum reproductive size of both males and females was smaller relative to bullfrogs in their native range as well as in populations across their invasive range. Reduction in size at reproductive maturity is likely impacting the invasive success of American bullfrogs and this study gives us insight on management actions to control the invasion. Applying this insight, managers can adjust their definition of reproductively active adults, increasing the target population of culling and other control methods.
Project description:The fungal pathogen Batrachochytrium dendrobatidis (Bd) is the causative agent of chytridiomycosis and has been a key driver in the catastrophic decline of amphibians globally. While many strategies have been proposed to mitigate Bd outbreaks, few have been successful. In recent years, the use of probiotic formulations that protect an amphibian host by killing or inhibiting Bd have shown promise as an effective chytridiomycosis control strategy. The North American bullfrog (Lithobates catesbeianus) is a common carrier of Bd and harbours a diverse skin microbiota that includes lactic acid bacteria (LAB), a microbial group containing species classified as safe and conferring host benefits. We investigated beneficial/probiotic properties: anti-Bd activity, and adhesion and colonisation characteristics (hydrophobicity, biofilm formation and exopolysaccharide-EPS production) in two confirmed LAB (cLAB-Enterococcus gallinarum CRL 1826, Lactococcus garvieae CRL 1828) and 60 presumptive LAB (pLAB) [together named as LABs] isolated from bullfrog skin.We challenged LABs against eight genetically diverse Bd isolates and found that 32% of the LABs inhibited at least one Bd isolate with varying rates of inhibition. Thus, we established a score of sensitivity from highest (BdGPL AVS7) to lowest (BdGPL C2A) for the studied Bd isolates. We further reveal key factors underlying host adhesion and colonisation of LABs. Specifically, 90.3% of LABs exhibited hydrophilic properties that may promote adhesion to the cutaneous mucus, with the remaining isolates (9.7%) being hydrophobic in nature with a surface polarity compatible with colonisation of acidic, basic or both substrate types. We also found that 59.7% of LABs showed EPS synthesis and 66.1% produced biofilm at different levels: 21% weak, 29% moderate, and 16.1% strong. Together all these properties enhance colonisation of the host surface (mucus or epithelial cells) and may confer protective benefits against Bd through competitive exclusion. Correspondence analysis indicated that biofilm synthesis was LABs specific with high aggregating bacteria correlating with strong biofilm producers, and EPS producers being correlated to negative biofilm producing LABs. We performed Random Amplified Polymorphic DNA (RAPD)-PCR analysis and demonstrated a higher degree of genetic diversity among rod-shaped pLAB than cocci. Based on the LAB genetic analysis and specific probiotic selection criteria that involve beneficial properties, we sequenced 16 pLAB which were identified as Pediococcus pentosaceus, Enterococcus thailandicus, Lactobacillus pentosus/L. plantarum, L. brevis, and L. curvatus. Compatibility assays performed with cLAB and the 16 species described above indicate that all tested LAB can be included in a mixed probiotic formula. Based on our analyses, we suggest that E. gallinarum CRL 1826, L. garvieae CRL 1828, and P. pentosaceus 15 and 18B represent optimal probiotic candidates for Bd control and mitigation.