Columnaris disease is responsible for substantial losses throughout the production of many freshwater fish species. One of the ways in which the bacterium Flavobacterium columnare is so effective in ...initiating disease is through the formation of biofilms on fish skin and gills. To further explore the interaction between host factors and bacterial cells, we assayed the ability of vertebrate mucus to enhance F. columnare biofilm development. Different concentrations of catfish, tilapia and pig mucus (5–60 µg/ml) increased biofilm growth at varying degrees among F. columnare isolates. Our data suggest that vertebrate mucus acts as a signalling molecule for the development of F. columnare biofilms; however, there are clear disparities in how individual isolates respond to different mucus fractions to stimulate biofilms. The expression of iron acquisition genes among two genomovar II isolates showed that ferroxidase, TonB receptor and the siderophore synthetase gene were all significantly upregulated among F. columnare biofilms. Interestingly, the siderophore acetyltransferase gene was only shown to be significantly upregulated in one of the genomovar II isolates. This work provides insight into our understanding of the interaction between F. columnare and vertebrate mucus, which likely contributes to the growth of planktonic cells and the transition into biofilms.
Ichthyophthirius multifiliis (Ich), a ciliated protozoan parasite of fish, expresses surface antigens (i-antigens), which react with host antibodies that render them immobile. The nucleotide sequence ...of an i-antigen gene of I. multifiliis strain ARS-6 was deduced. The predicted protein of 47 493 Da is comprised of 460 amino acids (aa's) arranged into five imperfect repeats with periodic cysteine residues with the structure: CX₍₁₉₎₂₀CX₂CX₁₆₋₂₇CX₂CX₂₀₍₂₁₎CX₃. The N-terminal aa's typify a signal peptide motif while a stretch of C-terminal aa's resemble a glycosyl-phosphatidyl-inositol (GPI)-anchor addition site. The degree of deduced i-antigen aa sequence identity of strain ARS-6 (GenBank accession # ACH87654 and # ACH95659) with other I. multifiliis i-antigen sequences present in GenBank ranges from 99% to 36% identity with 52 kDa i-antigens of I. multifiliis strain G5 (accession #s AAK94941 and AAK01661 respectively). Immunoblot analysis of i-antigens following exposure of I. multifiliis theronts to catfish anti-I. multifiliis immune serum did not show any appreciable alteration in i-antigen expression. The mechanism that regulates i-antigen expression in I. multifiliis remains a puzzling question.
Utilizing RNA-seq, this study compared the transcriptomic responses of three improved strains (VSel, PSel, and CSel) of rainbow trout fry during acute stages of challenge with infectious ...hematopoietic necrosis virus (IHNV). The VSel strain has been selected for resistance against the specific strain of IHNV used in our challenge, PSel has undergone selection for utilization of plant-protein based feeds and previously has shown elevated non-specific disease resistance despite no disease related selection pressures, and the final strain, CSel, is a commercial strain that has been domesticated for several years but has not been selected for specific viral disease resistance. Following a 21-day IHNV challenge, Kaplan-Meier survival estimator curves and cumulative percent mortality (CPM) showed significant differences in IHNV resistance across strains: VSel - 19.3 ± 5.0%, PSel - 67. ± 3.03%, CSel - 94.6 ± 4.1% CPM. To evaluate acute responses to IHNV infection, whole blood, as well as samples from the kidney, liver, and intestine, were collected at 0, 4, 12, 24, and 48 h post infection (hpi). Serum lysozyme activity, a marker of non-specific innate immunity, showed strain and temporal effects during the acute infection phase with PSel showing the highest activity at 0 and 48 hpi. Differential gene expression responses were detected, with varying degrees, in all tissues, both between strains, as well as across acute timepoints within strains. The VSel strain showed upregulation for a particular subset of viral recognition genes during early infection timepoints and rather limited upregulation of immune genes later, while maintaining and reactivating metabolic pathways. The CSel strain showed a downregulation of metabolic related genes and a limited upregulation of immune genes, while the PSel strain showed similar downregulation of metabolic genes during acute infection, yet when compared to the CSel strain, showed a more robust innate immune response. Evaluation of upregulated immune response genes, as well as interferon-related genes showed the PSel strain to have the greatest number of uniquely upregulated immune genes in both the kidney and intestine, with CSel and PSel showing a similar number of such genes upregulated in liver. A moderate number of immune response genes were shared between PSel and CSel in all tissues, though both PSel and VSel showed a high number of uniquely overexpressed immune response genes in the kidney, and PSel showed the highest number of uniquely upregulated interferon related genes in the intestine. Overall, the VSel response was unique from the CSel with very little overlap in activated immune responses. Findings from this study highlight the disparity in IHNV resistance among genetic strains of rainbow trout, while identifying molecular mechanisms underlying differences in disease phenotypes. Furthermore, our results on trout strains with distinct selection backgrounds yields comparative insights into the adaptive gains brought about by selection programs for pathogen-specific disease resistance, as well as the non-specific immune enhancement associated with selection for utilization of plant-based diets.
•Trout with disparate selection histories showed differences in resistance to IHNV challenge.•Trout selected for IHNV resistance showed the greatest survival, with rapid and resolute transcriptional responses.•Trout selected for growth on plant-based diets showed intermediate disease resistance and the most robust immune response.•Selection for growth performance on alternative plant-base diets can lead to enhanced non-specific immune responses.
Polyadenylation of eukaryotic transcripts is usually restricted to mRNA, providing transcripts with stability from degradation by nucleases. Conversely, an RNA degradation pathway can be signaled ...through poly (A) tailing in prokaryotic, archeal, and organellar biology. Recently polyadenylated transcripts have also been discovered in rRNA in some eukaryotes including humans and yeast. Here we report the discovery of polyadenylated rRNAs in the ciliate teleost parasite
Ichthyophthirius multifiliis, an important fish pathogen. Through large-scale analysis of ESTs, a large contig composed of the 28S rRNA with poly (A) tails was identified. Analysis using multiple sequence alignments revealed four potential polyadenylation sites including three internal regions and the 3′ end of the rRNA. Further analysis using a polyadenylation test, re-sequencing, and gene-specific PCR using primers flanking the presumed poly (A) sites confirmed the presence of polyadenylated rRNA in this parasite. The functions of polyadenylation of rRNA in this organism are largely unknown at present, but the presence of internal polyadenylation sites, along with the presence of truncated segments of the rRNA, may suggest a role of the polyadenylation in the degradation pathway, a function typical of prokaryotes, archaea, and organelles. These results are in congruence with reports of a similar phenomenon in humans and yeast.
ABSTRACT
The Gram-negative bacterium, Aeromonas hydrophila, has been responsible for extensive losses in the catfish industry for over a decade. Due to this impact, there are ongoing efforts to ...understand the basic mechanisms that contribute to virulent A. hydrophila (vAh) outbreaks. Recent challenge models demonstrated that vAh cultured in the presence of the iron chelating agent deferoxamine mesylate (DFO) were more virulent to channel catfish (Ictalurus punctatus). Interestingly, differential gene expression of select iron acquisition genes was unremarkable between DFO and non-DFO cultures, posing the question: why the increased virulence? The current work sought to evaluate growth characteristics and protein expression of vAh after the addition of DFO. A comparative proteome analysis revealed differentially expressed proteins among tryptic soy broth (TSB) and TSB + DFO treatments. Upregulated proteins identified among the TSB + DFO treatment were enriched for gene ontology groups including iron ion transport, siderophore transport and siderophore uptake transport, all iron acquisition pathways. Protein-protein interactions were also evaluated among the differentially expressed proteins and predicted that many of the upregulated iron acquisition proteins likely form functional physiological networks. The proteome analysis of the vAh reveals valuable information about the basic biological processes likely leading to increased virulence during iron restriction in this organism.
Proteome analysis of virulent Aeromonas hydrophila reveals basic biological processes that lead to increased virulence during iron restricted growth.
Vibrio parahaemolyticus is a Gram-negative bacterium commonly found in marine and estuarine environments and is endemic among the global shrimp aquaculture industry. V. parahaemolyticus proteins PirA ...and PirB have been determined to be major virulence factors that contribute significantly to the development of acute hepatopancreatic necrosis disease. Our previous work had demonstrated the lethality of recombinant PirA and PirB proteins to Pacific white shrimp (Liptopenaeus vannamei). To understand the host response to these proteins, recombinant PirA and PirB proteins were administered using a reverse gavage method and individual shrimp were then sampled over time. Shrimp hepatopancreas libraries were generated and RNA sequencing was performed on the control and recombinant PirA/B-treated samples. Differentially expressed genes were identified among the assayed time points. Differentially expressed genes that were co-expressed at the later time points (2-, 4- and 6-h) were also identified and gene associations were established to predict functional physiological networks. Our analysis reveals that the recombinant PirA and PirB proteins have likely initiated an early host response involving several cell survival signaling and innate immune processes.
•RNA sequencing performed to evaluate the shrimp host response to Vibrio toxin.•Differential gene expression identified in the shrimp hepatopancreas post-exposure to Vibrio toxin.•The shrimp host response activated regulatory pathways, including cell survival signaling and innate immune processes.
Objective
The sunshine bass (female White Bass Morone chrysops × male Striped Bass M. saxatilis) is an important food fish raised in U.S. aquaculture. Spawning is performed by manually stripping and ...fertilizing White Bass eggs with Striped Bass sperm. Embryos hatch in approximately 48 h. Mature sunshine bass females can become fertile and produce viable eggs, which is problematic in commercial food‐fish production; therefore, triploid fish are desired.
Methods
The present nonreplicated study incorporated three warm and three cold temperature shocks, along with three postfertilization initiation times and three exposure times for each temperature, during initial stages of incubation to induce triploidy. After exposure to temperature shocks, fertilized eggs were incubated in experimental hatching systems. The best performing cold and warm temperatures and times (and an ambient diploid control) were then used to incubate fertilized eggs in a commercial‐scale trial using standard industry practices with a single batch of eggs. Larvae were stocked at about 185,000 larvae/ha (75,000 larvae/acre) in newly filled and fertilized ponds. Fry were sampled weekly and harvested at 30 days.
Result
There were 15 survivors from the pond containing fish derived from cold‐shocked fertilized eggs; these individuals were tested via Coulter Counter, and no triploids were detected. Testing of 50 survivors from the pond containing fish derived from warm‐shocked fertilized eggs indicated that 14% were triploid.
Conclusion
Therefore, initial trials indicate that warm temperature shock is preferable to cold shock for inducing a greater percentage of triploid sunshine bass.
Impact statement
Successful use of temperature shock during the initial stages of fertilized egg incubation to produce triploid sunshine bass leads to a new method to improve sustainability and profitability for farmers.
Toll-like receptors (TLRs) are important components of innate immunity. They were found to recognise specific structures on pathogens termed pathogen-associated molecular patterns (PAMPs) and utilise ...conserved signaling pathways to activate pro-inflammatory cytokines and type-1 interferons. In spite of much understanding gained from the mammalian systems, many fish TLRs are unknown. Recent studies in Japanese flounder as well as in zebrafish suggested that the ligand binding and activation of inflammatory responses in fish may be different from and more complex than those found in mammals. In channel catfish, the major aquaculture species in the United States, only partial sequences of TLR3 and TLR5 were reported. As a part of efforts to characterise the innate immune components in channel catfish, here we cloned and sequenced both the cDNA and the gene for TLR2, a receptor believed mostly responsible for recognition of lipopeptides on the surface of most Gram-positive bacteria. However, expression analysis after infection with a Gram-negative bacterium,
Edwardsiella ictaluri indicated that TLR2 was modestly down-regulated in the head kidney tissue of blue catfish, and with a similar pattern in the head kidney of channel catfish though the down-regulation in channel catfish was not statistically significant. In the spleen, an insignificant down-regulation was initially observed early after infection, with an increase of TLR expression later after infection. These results suggest the involvement of TLR2 in the responses after the bacterial infection. As LPS is believed to be the major PAMP for Gram-negative bacteria, additional research is warranted to determine the functions and mechanisms of TLR2 in infections of Gram-negative bacteria.
Replacement of fishmeal (FM) with alternative plant proteins, especially soybean meal (SBM), can cause a diarrhea-like symptom in rainbow trout (RBT), characterized by very fine fecal particles. ...These fines do not settle out in raceway effluent for collection and can contribute to pollution of receiving waters. In this study, two experiments were conducted. Experiment 1 examined effects of nine protein sources (sardine meal, menhaden meal, soy protein concentrates (SPC) (three types), SBM (regular and high protein), corn protein concentrate (CPC), and poultry by-product meal (PBM)) on fecal particle size distribution. Results showed that all five soy-based diets produced feces in RBT having 75.7–89.3% fines and only about 1.0% large particles, while the remaining four diets yielded feces having a balanced particle size distribution. Oligosaccharides present naturally in soy products, thought to contribute fecal fines, were not correlated to fecal particle size classes. Instead, high crude fiber content in soy-based diets was found to be responsible for unbalanced fecal particle distribution in RBT. Experiment 2 examined if improvements in formulation could reduce the negative effect of soy-based ingredients. Eight practical diets (FM, SPC, SPC + 0.3% guar gum, PBM + CPC, PBM + CPC + 20 or 30% SPC, and PBM + CPC + 20 or 30% SPC + 0.3% guar gum) were formulated to contain 40% protein and 20% lipid. Results showed that diets containing mixtures of PBM, CPC, and 20% or 30% SPC plus guar gum produced trout feces with the highest percentage of large particles and lowest of fines, while the diet containing SPC alone (56%) plus guar gum resulted in trout feces having the highest content of mid-size particles. It was concluded that crude fiber in soy protein products (SBM and SPC) caused undesirable fecal particle profiles in RBT, and the addition of guar gum could significantly alleviate this negative effect.
Juvenile rainbow trout were fed a plant‐based diet supplemented with inorganic Mn added at 0, 0.5, 1, 2, 4, 8, 16, or 32 mg/kg diet for 12 wk. Whole‐body Mn concentrations increased with increasing ...dietary levels. Rainbow trout fed 2–8 mg Mn/kg diet exhibited weight gain that was significantly higher than fish fed 0, 0.5, 1, 16, and 32 mg Mn/kg diet. Feed conversion ratio (FCR) showed a similar trend, where FCR values decreased with increasing dietary Mn until concentrations reached 8 mg Mn/kg, after which they increased. A rational nonlinear model (R2 = 0.84) fit to weight‐gain data showed that the optimal predicted supplementation level for Mn was 4.8 mg/kg diet. The optimal predicted supplementation level for Mn was similar for FCR (5 mg Mn/kg diet). The predicted 5 mg/kg diet of supplemental Mn required for the plant‐based diet formulation examined in this study is lower than the National Research Council–recommended supplementation level of 12 mg/kg. However, due to potential differences in bioavailability or retention of Mn among plant‐protein sources, the level of dietary supplementation for optimal growth could be slightly higher or lower depending on the dietary formulation.
PDF
