Microbiological analysis of skin lesions of cod (Gadus morhua) from the southern part of the Baltic Sea.

Introduction: Since the middle of the 1980s, severe skin disorders have been observed in Baltic cod (Gadus morhua) each year. Available data on the spectrum of bacteria isolated from the clinical cases being limited, and evaluation of the microbial background of fish skin lesions being useful, a bacteriological examination has been undertaken. Material and Methods: A total of 1,381 cod were caught during two voyages of the Baltica research vessel in the Polish exclusive economic zone of the southern Baltic Sea. After an examination which found lesions in 164 of the fish, a microbiological analysis was performed to isolate bacteria from them. The collected strains were phenotyped and genotyped, and their antimicrobial resistance was analysed by minimum inhibitory concentration (MIC) techniques. Results: Bacteriological examinations provided 850 isolates. The dominant microorganisms were mesophilic Aeromonas spp., Pseudomonas spp. and Shewanella baltica. Opportunistic bacteria potentially hazardous to human health were also isolated, e.g. Alcaligenes faecalis, Staphylococcus epidermidis, Stenotrophomonas maltophilia and Vibrio sp. The MIC analysis determined the highest number of bacteria to resist sulphamethoxazole and amoxicillin and clavulanic acid. Conclusion: Most of the collected bacteria were opportunistic pathogens for fish, widespread in the aquatic environment, and potentially threatening to humans.

Etiology of Ulcerative Dermal Necrosis (UDN) in Brown Trout (Salmo trutta Morpha trutta)-Preliminary Results.

Every year, ulcerative dermal necrosis (UDN) affects salmonids that spend most of their lives in the sea during their migration to the rivers of northern Poland to spawn. The clinical form of the disease manifests itself in ulcerative skin lesions, which lead to significant weakening of the fish and, in most cases, result in their death. This study was carried out on samples taken from sea trout in the Slupia River in northern Poland. In order to identify the pathogen, experiments on the transmission of the disease were carried out, and additional histopathological, microbiological and electron microscopic examinations were performed. As a result of these studies, it was possible to experimentally transfer the disease from sick to healthy fish. The results indicate a complex etiology of the disease (lack of a clearly defined pathogen), in which the change in the environment from salty to freshwater triggers the related changes in skin physiology, which are the main causes of increased susceptibility to the development of the disease.

Phylogenetic analysis of Shewanella spp. isolated from fish.

Different species of Shewanella spp. widely inhabit freshwater and marine environments. Some of them are opportunistic fish pathogens. The application of high-throughput sequencing enabled the characterization and taxonomic reclassification of many Shewanella spp. species. Still, some strains collected from fish need to be better recognized. The aim of the present study was to classify and determine the phylogenetic relationships of Shewanella spp. collected from fish. The complete genomes of 94 strains of Shewanella spp. from different fish species were sequenced using Illumina platform (MiSeq). The 16S rRNA gene, genomic features and whole-genome relationships of those bacteria were comprehensively analysed in comparison to reference strains. Whole-genome analysis showed that the tested Shewanella spp. strains were clustered into six groups similar to reference strains of S. xiamenensis, S. oneidensis, S. glacialipiscicola, S. hafniensis, S. baltica and S. oncorhynchi. Our study indicates that the whole-genome sequence analysis enabled taxonomic classification and assessment of the diversity of the Shewanella spp. strains, as opposed to recently the gold standard method of 16S rRNA amplicon sequencing. The high genetic diversity and low similarity to the reference genome of S. oneidensis indicate that the group of strains may be a subspecies or even new species. Furthermore, we showed that the most frequent Shewanella spp. species occurring in freshwater fish in our study is the recently described species S. oncorhynchi.

Could mycotoxigenic Fusarium sp. play a role in ulcerative dermal necrosis (UDN) of brown trout (Salmo trutta morpha trutta)?

Fusarium infections have been reported in aquatic animals, but are still poorly investigated in wild salmonids. The aim of the study was to determine the impact of the fungi and their toxins on the health status of brown trout (Salmo trutta morpha trutta) migrating from the Baltic Sea to the freshwater. Individuals from the wild brown trout population exhibiting ulcerative skin lesions were collected from the Slupia River in Poland and subjected to microbiological, histopathological, and hematological examinations, as well as toxicological analysis for a presence of mycotoxins. The results of microflora isolation from the brown trout skin samples revealed the presence of conditionally pathogenic bacteria and fungi classified by molecular techniques as Fusarium spp. Toxicological analysis allowed for detection of zearalenone (ZEN) in the liver, kidney, and gastrointestinal tract of the fish. In several cases, there was α-zearalenone (α-ZEL) identified at trace levels in the liver, as well as sterigmatocystin and enniatin B at low levels in the kidney and the liver. Histopathological examination revealed the presence of fungal hyphae disrupting the epidermis and penetrating into the necrotic dermis and hypodermis. The decreased values of the blood parameters, i.e., hemoglobin concentration (HGB), packed cell volume (PCV), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and white blood cell count (WBC), were indicative of osmoregulation failure being a consequence of the skin damage. The results of the study provide new information regarding Fusarium sp. infection in brown trout and serve as the basis for further research on the potential impact of the fungi and their mycotoxins on the Baltic salmonid population, including their role in ulcerative dermal necrosis.

Phenotypic Diversity and Potential Virulence Factors of the Shewanella Putrefaciens Group Isolated from Freshwater Fish.

INTRODUCTION: The Shewanella putrefaciens group are ubiquitous microorganisms recently isolated from different freshwater fish species and causing serious health disorders. The purpose of the study was to characterise isolates of the S. putrefaciens group with special emphasis on elucidating serological diversity and determining putative virulence factors. MATERIAL AND METHODS: Isolates collected from freshwater fish (n = 44) and reference strains were used. The identification of bacteria was carried out using biochemical kits and 16S rRNA sequencing. Polyclonal antibodies were prepared against the S. putrefaciens group. The bacterium's susceptibility to antimicrobial agents, its enzymatic properties, and its adhesion ability to fish cell lines were also tested. Finally, selected isolates were used in challenge experiments in common carp and rainbow trout. RESULTS: Excluding six isolates undeterminable for species, the bacteria were classified to three species: S. putrefaciens, S. xiamenensis, and S. oneidensis, and showed some phenotypic diversity. Fourteen serological variants of the S. putrefaciens group were determined with the newly developed serotyping scheme. CONCLUSION: Serodiversity may play an important role in the virulence of particular isolates. Further, S. putrefaciens group members adhere to epithelial cells and produce enzymes which may contribute to their virulence. Challenge tests confirmed the pathogenicity of the S. putrefaciens group for fish.

Genotypic diversity among Shewanella spp. collected from freshwater fish.

Different Shewanella species are isolated both from healthy and from diseased fish. To date, contemporary methods do not provide sufficient insight to determine species and detail differentiation between tested strains. Bacteria isolated from cultured (n = 33), wild (n = 12) and ornamental (n = 6) fish, as well as several reference strains, were tested by 16S rRNA gene sequencing, ERIC-PCR and pulsed-field gel electrophoresis (PFGE) assays. Our study indicates that isolates collected from freshwater fish were genetically diverse. Based on 16S rRNA gene sequences, bacteria were clustered into groups S. putrefaciens, S. xiamenensis and S. oneidensis. Some isolates were classified only to genus Shewanella; thus, 16S rRNA gene analyses were not enough to determine the species. ERIC-PCR revealed 49 different genotype profiles indicating that the method might be useful for differentiation of Shewanella isolates irrespectively to species identification, contrary to PFGE which is not suitable for Shewanella typing.