Introduction
Turtles as invasive species exist in the environment and could ravage their biodiversity by competing with native European pond turtles for breeding sites and food. They could also account for a source of a new commensal and pathogenic bacteria and consequently new diseases. The current knowledge on this subject is scanty1-6 so we have provided a project investigating the influence of alien turtles on the environment and autochthonous animal species and humans.
Materials and Methods
Turtles were trapped in water reservoirs in eastern Poland. Samples were taken during quarantine and after euthanasia. In order to detect multiple bacterial flora, samples were directly plated on 2 agar plates: CHROMagar™ Acinetobacter and CHROMagar™ Orientation. Every distinct colony in context of shape, color, smoothness of edge, polish, and change of color of surrounding agar, etc. was transferred to blood agar for purity check and hemolysis. Every isolate was brought under identification of MALDI-TOF mass spectrometry.
Results
There were 147 samples collected from 74 alien turtles (8 species) captured in 2015–2016: intestines (n=63) and faeces (n=32), ovaries/oviduct (n=44) and eggs (n=1), and transport/quarantine box linings (n=7). The total number 828 isolates were obtained from intestines (n=365), faeces (n=167), ovaries/oviduct (n=272), eggs (n=5), and transport/quarantine box linings (n=18). Acinetobacter CHROMagar™ and CHROMagar™ Orientation yield, 257 and 574 isolates, respectively. Total number of isolates were classified into 10 orders: Enterobacteriales (n=291), Pseudomonadales (n=128), Aeromonadales (n=118), Burkholderiales (n=56), Lactobacillales (n=33), Xanthomonadales (n=13), Bacillales (n=7), Flavobacteriales (n=6), Campylobacteriales (n=1), Caulobacteriales (n=1). The most numerous bacterium was Citrobacter freundii (n=122).
Among these abundant bacteria there was a group of zoonotic potential like Acinetobacter calcoaceticus, A. junii, Aeromonas caviae, Arcobacter butzleri, Citrobacter braakii, Delftia acidovorans, Elisabethkingia meningoseptica, Enterobacter cloacae, Klebsiella oxytoca, Morganella morganii, Myroides odoratimimus, Plesiomonas shigelloides, Providencia alcalifaciens, Pseudomonas aeruginosa, P. monteillii, Stenotrophomonas maltophilia, Staphylococcus haemolyticus. They are usually isolated from soil and water, but their pathogenicity encompasses community and hospital-acquired infections.
The other group contained bacteria that could be pathogenic for freshwater fish, both cultured and ornamental, causing serious disorders of their health. Among them, the most important and frequently isolated from samples collected from diseased fish were: Acinetobacter spp., Aeromonas caviae, A. sobria and A. hydrophila, Citrobacter freundii, Lactococcus garvieae, Pseudomonas spp., Serratia marcescens. Clinical symptoms of fish infections with these bacteria vary depending on the microorganism and fish species. Mesophilic Aeromonas (A. caviae, A. sobria, A. hydrophila) cause two kind of disease affecting, both carps and salmonids: MAI (motile aeromonads infection) where skin ulceration as well as gill and fin lesions are observed, and MAS (motile aeromonads septicemia) which is systemic disease with high mortalities reaching about 80% of the stock. Infections with Serratia spp. in rainbow trout and Pseudomonas spp. in different fish species are mainly connected with skin lesions and fish mortalities. Meanwhile, infections with gram-positive Lactococcus in salmonids are manifested by exophthalmia with petechiaes in the eyes, darkening of the skin, and anal edema; fish mortalities above 70% are also observed.
The third batch is comprised of bacteria, which are considered as safe environmental microorganisms or commensal flora: Lactococcus lactis, Lysinibacillus sphaericus, Hafnia alvei, Enterococcus gallinarum, Achromobacter ruhlandii, Acinetobacter tandoii, A. towneri, Aeromonas media, Bordetella trematum, Pseudomonas fulva, P. otitidis, P. putida.
Some of those bacteria could provide resistance genes, i.e., Klebsiella, Acinetobacter, and Pseudomonas. Therefore, a subset of isolates will be further analyzed for detailed epidemiological and phylogenic characterization as well as antimicrobial resistance (i.e., whole genome sequencing and MIC testing).
Conclusions
Invasive and alien turtles should be considered as a source of potentially pathogenic bacteria, to humans as well as to animals like fish and turtles, including autochthonous pond turtle (Emys orbicularis). The impact of alien turtles both on the environment and epidemiology of infections needs further in-depth studies.
Acknowledgments
This work was supported by the National Science Centre project “Invasive turtle species as a source and vector of animal and human pathogens” (Grant No. 2013/11/B/NZ7/01690).
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