Changes occurring in freshwater ecosystems seem to be fundamental in the development of all microorganisms, including those pathogenic to fish. This has been especially evident in recent years during which dynamic variations in bacterial fish pathology have been observed. Gram-negative bacteria commonly known to be pathogenic to fish, like Aeromonas spp., Flavobacterium spp., Pseudomonas spp., and Shewanella putrefaciens are replaced by other species, which until now have not been known to be virulent or even conditionally pathogenic to fish. Nowadays, among these other species Acinetobacter spp., Plesiomonas shigelloides , Sphingomonas paucimobilis , and Stenotrophomonas maltophilia are the most frequently isolated from fish exhibiting clinical signs of disease. Two Gram-positive bacteria have become pathogens of particular importance in fish pathology in Poland: Lactococcus garviae and Streptococcus iniae . In addition, infections caused by the Gram-positive bacterium Kocuria rhizophila have appeared in recent years. This bacterium has not been known until now to be pathogenic to fish. Therefore, this infection could be called an emergent disease.

Antimicrobial activities of phytochemicals—trans-cinnamaldehyde (TC), ferulic acid (FA), p-coumaric acid (p-CA), caffeic acid (CA), chlorogenic acid (CHA), Thymus vulgaris essential oil (TO), Eugenia caryophyllus essential oil (ECO), and Melaleuca alternifolia oil (TTO) against Aeromonas species—were assessed. Growth of all Aeromonas salmonicida subsp. salmonicida and almost all Aeromonas sobria strains was inhibited by TC at concentration 0.01 mg/mL, and for most Aeromonas hydrophila strains minimal inhibitory concentrations (MIC) ranged from 0.01 to 0.19 mg/mL. The inhibitory effect of TC against A. salmonicida subsp. salmonicida was comparable to the effect of oxytetracycline, and in the case of A. salmonicida subsp. salmonicida and A. sobria was higher compared to gentamicin. MIC of FA, p-CA, and CA for most strains ranged from 1.56 to 3.12 mg/mL, and MIC values of TO for most strains ranged from 0.39 to 0.78 mg/mL. TO and TC at the concentrations below ½ MIC values used in mixtures exhibited strong synergism. ECO and TC showed synergy in mixture of ⅛ MIC of ECO and ¼ MIC of TC. TC and TO exhibited the strongest inhibitory and bactericidal effect against investigated Aeromonas species, and they are a promising alternative to the use of antibiotics in controlling the growth of these fish pathogens.

Chemical analyses, mass spectrometry, and NMR spectroscopy were applied to study the structure of the lipopolysaccharide (LPS) isolated from Aeromonas veronii strain Bs19, serotype O16. ESI-MS revealed that the most abundant LPS glycoforms have tetra-acylated or hexa-acylated lipid A species, consisting of a bisphosphorylated GlcN disaccharide with an AraN residue as a non-stoichiometric substituent, and a core oligosaccharide composed of Hep₅Hex₃HexN₁Kdo₁P₁. Sugar and methylation analysis together with 1D and 2D ¹H and ¹³C NMR spectroscopy were the main methods used, and revealed that the O-specific polysaccharide (OPS) of A. veronii Bs19 was built up of tetrasaccharide repeating units with the structure: →4)-α-d-Quip3NAc-(1→3)-α-l-Rhap-(1→4)-β-d-Galp-(1→3)-α-d-GalpNAc-(1→. This composition was confirmed by mass spectrometry. The charge-deconvoluted ESI FT-ICR MS recorded for the LPS preparations identified mass peaks of SR- and R-form LPS species, that differed by Δm = 698.27 u, a value corresponding to the calculated molecular mass of one OPS repeating unit (6dHexNAc6dHexHexHexNAc-H₂O). Moreover, unspecific fragmentation spectra confirmed the sequence of the sugar residues in the OPS and allowed to assume that the elucidated structure also represented the biological repeating unit.

Aeromonas species are opportunistic bacteria causing a vast spectrum of human diseases, including skin and soft tissue infections, meningitis, endocarditis, peritonitis, gastroenteritis, and finally hemorrhagic septicemia. The aim of our research was to indicate the molecular alterations in proteins and lipids profiles resulting from Aeromonas sobria and A. salmonicida subsp. salmonicida infection in trout kidney tissue samples. We successfully applied FT-IR (Fourier transform infrared) spectroscopy and MALDI-MSI (matrix-assisted laser desorption/ionization mass spectrometry imaging) to monitor changes in the structure and compositions of lipids, secondary conformation of proteins, and provide useful information concerning disease progression. Our findings indicate that the following spectral bands’ absorbance ratios (spectral biomarkers) can be used to discriminate healthy tissue from pathologically altered tissue, for example, lipids (CH₂/CH₃), amide I/amide II, amide I/CH₂ and amide I/CH₃. Spectral data obtained from 10 single measurements of each specimen indicate numerous abnormalities concerning proteins, lipids, and phospholipids induced by Aeromonas infection, suggesting significant disruption of the cell membranes. Moreover, the increase in the content of lysolipids such as lysophosphosphatidylcholine was observed. The results of this study suggest the application of both methods MALDI-MSI and FT-IR as accurate methods for profiling biomolecules and identifying biochemical changes in kidney tissue during the progression of Aeromonas infection.

Diagnostic tools for the identification and confirmation of animal diseases have been evolving rapidly over the last decade, with diseases of aquatic animals being no exception. Hence, case definitions used in surveillance may now include molecular and genomic components and ultimately be based on the entire genome of a pathogen. While the opportunities brought on by this change in our ability to define and differentiate organisms are manifold, there are also challenges. These include the need to consider typing tool characteristics during sampling design, but also the re-thinking of diagnostic protocols and standards for the meaningful interpretation of the increasingly complex data presented to surveillance managers. These issues are illustrated for aquaculture using the example of multi-country surveillance of antimicrobial resistance of Aeromonas spp. strains isolated from rainbow trouts (Oncorhynchus mykiss) in Europe. In order to fully exploit the opportunities of molecular and genomic information, a multi-disciplinary approach is needed to develop harmonised diagnostic procedures and modified surveillance designs for aquaculture as well as for terrestrial animal production. This will require adjustments in the relevant standards applicable to assess food safety and trade risks.