Genomics to accelerate genetic improvement in tilapia.

Selective breeding of tilapia populations started in the early 1990s and over the past three decades tilapia has become one of the most important farmed freshwater species, being produced in more than 125 countries around the globe. Although genome assemblies have been available since 2011, most of the tilapia industry still depends on classical selection techniques using mass spawning or pedigree information to select for growth traits with reported genetic gains of up to 20% per generation. The involvement of international breeding companies and research institutions has resulted in the rapid development and application of genomic resources in the last few years. GWAS and genomic selection are expected to contribute to uncovering the genetic variants involved in economically relevant traits and increasing the genetic gain in selective breeding programs, respectively. Developments over the next few years will probably focus on achieving a deep understanding of genetic architecture of complex traits, as well as accelerating genetic progress in the selection for growth-, quality- and robustness-related traits. Novel phenotyping technologies (i.e. phenomics), lower-cost whole-genome sequencing approaches, functional genomics and gene editing tools will be crucial in future developments for the improvement of tilapia aquaculture.

Bayesian genome-wide association analysis for body weight in farmed Atlantic salmon (Salmo salar L.).

We performed a genome-wide association study to detect markers associated with growth traits in Atlantic salmon. The analyzed traits included body weight at tagging (BWT) and body weight at 25 months (BW25M). Genotypes of 4662 animals were imputed from the 50K SNP chip to the 200K SNP chip using fimpute software. The markers were simultaneously modeled using Bayes C to identify genomic regions associated with the traits. We identified windows explaining a maximum of 3.71% and 3.61% of the genetic variance for BWT and BW25M respectively. We found potential candidate genes located within the top ten 1-Mb windows for BWT and BW25M. For instance, the vitronectin (VTN) gene, which has been previously reported to be associated with cell growth, was found within one of the top ten 1-Mb windows for BWT. In addition, the WNT1-inducible-signaling pathway protein 3, melanocortin 2 receptor accessory protein 2, myosin light chain kinase, transforming growth factor beta receptor type 3 and myosin light chain 1 genes, which have been reported to be associated with skeletal growth in humans, growth stimulation during the larval stage in zebrafish, body weight in pigs, feed conversion in chickens and growth rate of sheep skeletal muscle respectively, were found within some of the top ten 1-Mb windows for BW25M. These results indicate that growth traits are most likely controlled by many variants with relatively small effects in Atlantic salmon. The genomic regions associated with the traits studied here may provide further insight into the functional regions underlying growth traits in this species.

Biodegradation of roxarsone by a bacterial community of underground water and its toxic impact.

Roxarsone is included in chicken food as anticoccidial and mainly excreted unchanged in faeces. Microorganisms biotransform roxarsone into toxic compounds that leach and contaminate underground waters used for human consumption. This study evaluated roxarsone biotransformation by underground water microorganisms and the toxicity of the resulting compounds. Underground water from an agricultural field was used to prepare microcosms, containing 0.05 mM roxarsone, and cultured under aerobic or anaerobic conditions. Bacterial communities of microcosms were characterized by PCR-DGGE. Roxarsone degradation was measured by HPLC/HG/AAS. Toxicity was evaluated using HUVEC cells and the Toxi-ChromoTest kit. Roxarsone degradation analysis, after 15 days, showed that microcosms of underground water with nutrients degraded 90 and 83.3% of roxarsone under anaerobic and aerobic conditions, respectively. Microcosms without nutrients degraded 50 and 33.1% under anaerobic and aerobic conditions, respectively. Microcosms including nutrients showed more roxarsone conversion into toxic inorganic arsenic species. DGGE analyses showed the presence of Proteobacteria, Firmicutes, Actinobacteria, Planctomycetes and Spirochaetes. Toxicity assays showed that roxarsone biotransformation by underground water microorganisms in all microcosms generated degradation products toxic for eukaryotic and prokaryotic cells. Furthermore, toxicity increased when roxarsone leached though a soil column and was further transformed by the bacterial community present in underground water. Therefore, using underground water from areas where roxarsone containing manure is used as fertilizer might be a health risk.

Evaluation of an immunological score to assess the risk of severe infection in heart recipients.

BACKGROUND: We previously reported how specific humoral and cellular immunological markers that are readily available in clinical practice can be used to identify heart transplant recipients (HTR) at risk of developing severe infections. In this study, we perform an extended analysis to identify immunological profiles that could prove to be superior to individual markers in assessing the risk of infection early after heart transplantation. METHODS: In a prospective follow-up study, we evaluated 100 HTR at 1 week after transplantation. Laboratory tests included determination of immunoglobulin (Ig) levels (IgG, IgA, IgM), complement factors (C3 and C4), and lymphocyte subsets (CD3+, CD4+, CD8+ T cells, B cells, and natural killer [NK] cells). The prevalence of infection during the first 3 months was registered at scheduled visits after transplantation. Severe infections were defined as all infections requiring hospitalization and intravenous antimicrobial therapy. RESULTS: During follow-up, 33 patients (33%) developed severe infections. The individual risk factors of severe infection, according to the Cox regression analysis, were as follows: IgG <600 mg/dL (hazard ratio [HR], 2.41; 95% confidence interval [CI], 1.21-4.78; P = 0.012), C3 <80 mg/dL (HR, 4.65; 95% CI, 2.31-9.38; P < 0.0001), C4 <18 mg/dL (HR 2.30, 95% CI, 1.15-4.59; P = 0.018), NK count <30 cells/µL (HR 4.07, 95% CI, 1.76-9.38; P = 0.001), and CD4 count <350 cells/µL (HR, 3.04; 95% CI, 1.47-6.28; P = 0.0027). An immunological score was created. HRs were used to determine the number of points assigned to each of the 5 previously mentioned individual risk factors. The score was obtained from the sum of these factors. In the multivariate Cox regression analysis, the immunological score was useful for identifying patients at risk of infection and was the only variable that maintained a significant association with the development of infection, after adjustment for the 5 individual factors. CONCLUSION: Patients with an immunological score ≥13 were at the highest risk of severe infections (HR, 9.29; 95% CI, 4.57-18.90; P < 0.0001). This score remained significantly associated with the risk of severe infection after adjustment for clinical risk factors of infection. An immunological score was useful for identifying HTR at risk of developing severe infections. If this score is validated in multicenter studies, it could be easily introduced into clinical practice.

Red blood cell antigen portrait of self-identified black donors in Quebec.

OBJECTIVES: The goal of this study was to establish a red blood cell antigen portrait of self-identified Black donors for the province of Quebec, Canada. BACKGROUND: The demand for extensively phenotyped red blood cells is on the rise. A good example is the sickle cell patient cohort. To better answer their transfusion needs, Héma-Québec put forward great efforts to increase the recruitment of donors among cultural communities. MATERIALS AND METHODS: In October 2009, an optional question was added on the record of donation to indicate the donor's ethnicity. Self-identified Black donors were extensively phenotyped by the Immunohematology Laboratory, whereas the Research and Development team genotyped red blood cell antigens to complete the picture. RESULTS: Approximately 1500 self-identified Black donors have donated blood at least once since the beginning of the programme. Genotyping results predicted rare phenotypes: 18 S-s- (3 U-, 15 U+(w) ), 15 Js(a+b-), 5 Hy-, 3 Jo(a-), 34 hr(B) +(w) /- and 15 hr(B)-. CONCLUSION: These Black donors, with or without a rare phenotype, are precious to the patient cohort depending on blood transfusions and to our organisation as the blood provider for the whole province of Quebec.

Characterization of Mycobacterium salmoniphilum as causal agent of mycobacteriosis in Atlantic salmon, Salmo salar L., from a freshwater recirculation system.

Thirty Atlantic salmon, Salmo salar L., with low corporal condition relative to other fish present in the culture system, were sampled from a freshwater recirculation pisciculture located in Chile. The most characteristic signs and lesions were cachexia and presence of multiple greyish-white granulomas within internal organs. The external and internal lesions, along with the microscopic, histologic and biochemical findings, were consistent with mycobacteriosis. The identification of Mycobacterium salmoniphilum as the causal agent of the lesions was possible through the use of molecular analyses. This study represents the first report of Mycobacterium salmoniphilum in a freshwater recirculation system and the first case of fish mycobacteriosis described in Chile.

Performance, serum amino acid concentrations and expression of selected genes in pair-fed growing pigs exposed to high ambient temperatures.

Heat stress (HS) depresses pig performance mainly because of appetite reduction, although other factors involved in the cellular availability of nutrients may also contribute to that depression. An experiment was conducted with twelve pair-fed pigs (30.3 ± 2.7 kg BW) to examine the effect of severe HS (up to 45 °C) on the expression of genes coding for two cationic amino acid (AA) transporters (b(0,+) AT and CAT-1), leptin, heat-shock protein (Hsp-90) and myosin in several tissues; serum concentrations (SC) of AA; and performance. There were two treatments: Comfort, pigs housed at an average temperature of 22 (±2) °C; and HS, pigs housed in a similar room with no climate control, where temperature was raised up to 45 °C. All pigs received the same wheat-soybean meal diet and had similar daily feed intake. Comfort pigs had a higher daily gain and better gain/feed ratio than HS pigs (p < 0.05). The expression of b(0,+) AT in jejunum and liver, that of myosin in the Semitendinosus muscle, and leptin in adipose tissue was lower, but CAT-1 in jejunum and liver, and Hsp-90 in liver was higher in HS pigs. The SC of Lys and Met in HS pigs were around 55% and 20%, respectively, of that in Comfort pigs (p < 0.05). In conclusion, HS affects the expression of cationic AA transporters, myosin, Hsp-90, leptin; the SC of Lys and Met; and the performance of pair-fed pigs. These results suggest that HS-related changes in gene expression affect the performance of pigs beyond the effect caused by the reduction in voluntary feed intake.

Assessing footprints of selection in commercial Atlantic salmon populations using microsatellite data.

Relatively large rates of response to traits of economic importance have been observed in different selection experiments in salmon. Several QTL have been mapped in the salmon genome, explaining unprecedented levels of phenotypic variation. Owing to the relatively large selection intensity, individual loci may be indirectly selected, leaving molecular footprints of selection, together with increased inbreeding, as its likely relatives will share the selected loci. We used population differentiation and levels of linkage disequilibrium in chromosomes known to be harbouring QTL for body weight, infectious pancreatic necrosis resistance and infectious salmon anaemia resistance to assess the recent selection history at the genomic level in Atlantic salmon. The results clearly suggest that the marker SSA0343BSFU on chromosome 3 (body weight QTL) showed strong evidence of directional selection. It is intriguing that this marker is physically mapped to a region near the coding sequence of DVL2 , making it an ideal candidate gene to explain the rapid evolutionary response of this chromosome to selection for growth in Salmo salar. Weak evidence of diversifying selection was observed in the QTL associated with infectious pancreatic necrosis and infectious salmon anaemia resistance. Overall, this study showed that artificial selection has produced important changes in the Atlantic salmon genome, validating QTL in commercial salmon populations used for production purposes according to the recent selection history.

Restoration of humoral immunity after intravenous immunoglobulin replacement therapy in heart recipients with post-transplant antibody deficiency and severe infections.

IgG hypogammaglobulinemia is a risk factor for infection in heart recipients. We assessed reconstitution of humoral immunity after non-specific intravenous immunoglobulin (IVIg) replacement therapy administered to treat secondary IgG hypogammaglobulinemia in heart recipients with severe infections. The study population comprised 55 heart recipients who were administered IVIg (IVIg group) and 55 heart recipients with no severe infectious complications (control group). An event was defined as a severe infection requiring intravenous drug therapy during the first year after transplantation. The IVIg protocol comprised non-specific 5% pasteurized IVIg at a dose of 300-400 mg/kg/months. IgG titers were lower in the IVIg group than in controls at seven d (577 vs. 778 mg/dL, p < 0.001) and at one month (553 vs. 684, p = 0.003). After IVIg therapy, IgG concentrations were similar in both groups at three months (681 vs. 737, p = 0.25) and at six months (736 vs. 769, p = 0.46). At three months, the IVIg group had higher levels of antitetanus toxoid and anti-HBs (ELISA, 2.07 ± 2.11 vs. 0.60 ± 1.24 mg/dL [p = 0.003] and 42 ± 40 vs. 11 ± 31 IU/mL [p = 0.005], respectively) than controls. The mean number of infectious complications was significantly lower after IVIG therapy in the IVIG group. IVIg was associated with restoration of humoral immunity in heart recipients with post-transplant IgG hypogammaglobulinemia and severe infections.

Decreased levels of serum complement C3 and natural killer cells add to the predictive value of total immunoglobulin G for severe infection in heart transplant recipients.

BACKGROUND: Infection remains a source of mortality in heart recipients. We previously reported that post-transplant immunoglobulin G (IgG) quantification can help identify the risk for infection. We assessed whether other standardized parameters of humoral and cellular immunity could prove useful when identifying patients at risk of infection. METHODS: We prospectively studied 133 heart recipients over a 12-month period. Forty-eight patients had at least one episode of severe infection. An event was defined as an infection requiring intravenous antimicrobial therapy. RESULTS: Cox regression analysis revealed an association between the risk of developing infection and the following: lower IgG2 subclass levels (day 7: relative hazard [RH] 1.71; day 30: RH 1.76), lower IgA levels (day 7: RH 1.61; day 30: RH 1.91), lower complement C3 values (day 7: RH 1.25), lower CD3 absolute counts (day 30: RH 1.10), lower absolute natural killer [NK] cell count (day 7: RH 1.24), and lower IgG concentrations (day 7: RH 1.31; day 30: RH 1.36). Cox regression bivariate analysis revealed that lower day 7 C3 levels, IgG2 concentration, and absolute NK cell count remained significant after adjustment for total IgG levels. CONCLUSIONS: Data suggest that early immune monitoring including C3, IgG2, and NK cell testing in addition to IgG concentrations is useful when attempting to identify the risk of infection in heart transplant recipients.

Arsenic mobilization by epilithic bacterial communities associated with volcanic rocks from Camarones River, Atacama Desert, northern Chile.

The arsenic biogeochemical cycle is greatly dependent on microbial transformations that affect both the distribution and mobility of arsenic species in the environment. In this study, a microbial biofilm from volcanic rocks was characterized on the basis of its bacterial composition and ability to mobilize arsenic under circumneutral pH. Biofilm microstructure was analyzed by scanning electron microscopy (SEM)-energy-dispersive spectroscopy (EDS). Strains were isolated from biofilms and identified by 16S rDNA sequences analysis. Arsenic oxidation and reduction capacity was assayed with high-performance liquid chromatography coupled to gaseous formation performing the detection by atomic absortion in a quartz bucket (HPLC/HG/QAAS), and polymerase chain reaction was used to detect aox and ars genes. Bacterial communities associated with volcanic rocks were studied by denaturing gradient gel electrophoresis (DGGE). The SEM-EDS studies showed the presence of biofilm after 45 days of incubation. The relative closest GenBank matches of the DNA sequences, of isolated arsenic-resistant strains, showed the existence of four different genus: Burkholderia, Pseudomonas, Erwinia, and Pantoea. Four arsenite-resistant strains were isolates, and only three strains were able to oxidize >97% of the As(III) present (500 uM). All arsenate-resistant isolates were able to reduce between 69 and 86% of total As(V) (1000 uM). Analysis of 16S rDNA sequences obtained by DGGE showed the presence of four bacterial groups (∝-proteobacteria, γ-proteobacteria, Firmicutes, and Actinobacteria). Experiments demonstrate that epilithic bacterial communities play a key role in the mobilization of arsenic and metalloids speciation.

Detection of selection signatures in farmed coho salmon (Oncorhynchus kisutch) using dense genome-wide information.

Animal domestication and artificial selection give rise to gradual changes at the genomic level in populations. Subsequent footprints of selection, known as selection signatures or selective sweeps, have been traced in the genomes of many animal livestock species by exploiting variation in linkage disequilibrium patterns and/or reduction of genetic diversity. Domestication of most aquatic species is recent in comparison with land animals, and salmonids are one of the most important fish species in aquaculture. Coho salmon (Oncorhynchus kisutch), cultivated primarily in Chile, has been subjected to breeding programs to improve growth, disease resistance traits, and flesh color. This study aimed to identify selection signatures that may be involved in adaptation to culture conditions and traits of productive interest. To do so, individuals of two domestic populations cultured in Chile were genotyped with 200 thousand SNPs, and analyses were conducted using iHS, XP-EHH and CLR. Several signatures of selection on different chromosomal regions were detected across both populations. Some of the identified regions under selection contained genes such anapc2, alad, chp2 and myn, which have been previously associated with body weight in Atlantic salmon, or sec24d and robo1, which have been associated with resistance to Piscirickettsia salmonis in coho salmon. Findings in our study can contribute to an integrated genome-wide map of selection signatures, to help identify the genetic mechanisms of phenotypic diversity in coho salmon.

Set-up and routine use of a database of 10,555 genotyped blood donors to facilitate the screening of compatible blood components for alloimmunized patients.

BACKGROUND AND OBJECTIVES: Large-scale genotyping of blood donors for red blood cell and platelet antigens has been predicted to replace phenotyping assays in the screening of compatible blood components for alloimmunized patients. Although several genotyping platforms have been described, novel procedures and processes are needed to perform genotyping efficiently and to maximize its benefits for blood banks. MATERIALS AND METHODS: Here we describe the processes and procedures developed to introduce large-scale genotyping in our routine operations. RESULTS: Preliminary cost-benefit analysis indicated that genotyping must target frequent blood donors (> 3 donations/year) to be efficiently used. A custom-designed computer application was developed to manage the whole project. It selects frequent donors among recent donations, prints coded labels to identify blood samples sent to the external genotyping laboratory, and stores genotyping results. It can search for donors compatible for any combination of the 22 genotyped antigens as well as consult the current inventory for the presence of the corresponding blood components. The phenotype of recovered components is confirmed by standard serology techniques prior to shipment to hospitals. CONCLUSION: Since October 2007, 10 555 blood donors have been genotyped. The database is used on a regular basis to find compatible blood components with a genotype-phenotype concordance of 99.6%.

Isolation of arsenite-oxidizing bacteria from a natural biofilm associated to volcanic rocks of Atacama Desert, Chile.

Arsenic is naturally present in rocks, soil, water, and air. It is released to the environment by natural processes such as volcanic eruptions, and rock erosion. In this study, two arsenite-oxidizing strains were isolated from volcanic rocks obtained from the Camarones Valley, Atacama Desert, Chile. Strains were isolated from biofilms and identified by 16s ARNr sequences analysis. aox genes were detected by RT-PCR. The arsenic oxidation ability was assayed with silver nitrate and HPLC-HG-AAS. Four arsenite-resistant strains were isolated (8 mM). RT-PCR analysis showed the presence of aox genes in UC-2 and UC-6 strains. In addition, UC-2 and UC-6 strains were able to oxidize 90 and 95% arsenite present in the medium to arsenate, at a rate of 9.3 and 9.8 microg ml(-1) h(-1 )respectively. Bicarbonate (HCO(3) (-)) was used as unique carbon source. Finally, the significative oxidation capacity shown by both strains opens the way to further studies aimed at implementing biological systems to treat arsenic rich wastewater.

Arsenic resistant bacteria isolated from arsenic contaminated river in the Atacama Desert (Chile).

In this study, arsenic resistant bacteria were isolated from sediments of an arsenic contaminated river. Arsenic tolerance of bacteria isolated was carried out by serial dilution on agar plate. Redox abilities were investigated using KMnO4. arsC and aox genes were detected by PCR and RT-PCR, respectively. Bacterial populations were identified by RapID system. Forty nine bacterial strains were isolated, of these, 55 % corresponded to the reducing bacteria, 4% to oxidizing bacteria, 8% presented both activities and in 33% of the bacteria none activity was detected. arsC gene was detected in 11 strains and aox genes were not detected. The activity of arsenic transforming microorganisms in river sediment has significant implications for the behavior of the metalloid.

Isolation of arsenite-oxidizing bacteria from arsenic-enriched sediments from Camarones river, Northern Chile.

In Northern Chile, high arsenic concentrations are found in natural water, both natural and anthropogenic sources, a significant health risk. Nine bacterial strains were isolated from Camarones river sediments, located in Northern Chile, a river showing arsenic concentrations up to 1,100 microg/L. These strains were identified as Pseudomonas and they can oxidize arsenite (As(III)) to the less mobile arsenate (As(V)). The arsenite oxidase genes were identified in eight out of nine isolates. The arsenite oxidizing ability shown by the nine strains isolated from arsenic enriched sediments open the way to their potential application in biological treatment of effluents contaminated with arsenic.

Genetic (co)variation in skin pigmentation patterns and growth in rainbow trout.

From a physiological-behavioral perspective, it has been shown that fish with a higher density of black eumelanin spots are more dominant, less sensitive to stress, have higher feed intake, better feed efficiency and therefore are larger in size. Thus, we hypothesized that genetic (co)variation between skin pigmentation patterns and growth exists and it is advantageous in rainbow trout. The objective of this study was to determine the genetic relationships between skin pigmentation patterns and BW in a breeding population of rainbow trout. We performed a genetic analysis of pigmentation traits including dorsal color (DC), lateral band (LB) intensity, amount of spotting above (SA) and below (SB) the lateral line, and BW at harvest (HW). Variance components were estimated using a multi-trait linear animal model fitted by restricted maximum likelihood. Estimated heritabilities were 0.08±0.02, 0.17±0.03, 0.44±0.04, 0.17±0.04 and 0.23±0.04 for DC, LB, SA, SB and HW, respectively. Genetic correlations between HW and skin color traits were 0.42±0.13, 0.32±0.14 and 0.25±0.11 for LB, SA and SB, respectively. These results indicate positive, but low to moderate genetic relationships between the amount of spotting and BW in rainbow trout. Thus, higher levels of spotting are genetically associated with better growth performance in this population.

Malignancy after heart transplantation: incidence, prognosis and risk factors.

The Spanish Post-Heart-Transplant Tumour Registry comprises data on neoplasia following heart transplantation (HT) for all Spanish HT patients (1984-2003). This retrospective analysis of 3393 patients investigated the incidence and prognosis of neoplasia, and the influence of antiviral prophylaxis. About 50% of post-HT neoplasias were cutaneous, and 10% lymphomas. The cumulative incidence of skin cancers and other nonlymphoma cancers increased with age at HT and with time post-HT (from respectively 5.2 and 8.9 per 1000 person-years in the first year to 14.8 and 12.6 after 10 years), and was greater among men than women. None of these trends held for lymphomas. Induction therapy other than with IL2R-blockers generally increased the risk of neoplasia except when acyclovir was administered prophylactically during the first 3 months post-HT; prophylactic acyclovir halved the risk of lymphoma, regardless of other therapies. Institution of MMF during the first 3 months post-HT reduced the incidence of skin cancer independently of the effects of sex, age group, pre-HT smoking, use of tacrolimus in the first 3 months, induction treatment and antiviral treatment. Five-year survival rates after first tumor diagnosis were 74% for skin cancer, 20% for lymphoma and 32% for other tumors.

Structural heart disease as the cause of syncope.

We described the clinical evolution of patients with structural heart disease presenting at the emergency room with syncope. Patients were stratified according to their syncope etiology and available scores for syncope prognostication. Cox proportional hazard models were used to investigate the relationship between etiology of the syncope and event-free survival. Of the 82,678 emergency visits during the study period, 160 (0.16%) patients were there due to syncope, having a previous diagnosis of structural heart disease. During the median follow-up of 33.8±13.8 months, mean age at the qualifying syncope event was 68.3 years and 40.6% of patients were male. Syncope was vasovagal in 32%, cardiogenic in 57%, orthostatic hypotension in 6%, and of unknown causes in 5% of patients. The primary composite endpoint death, readmission, and emergency visit in 30 days was 39.4% in vasovagal syncope and 60.6% cardiogenic syncope (P<0.001). Primary endpoint-free survival was lower for patients with cardiogenic syncope (HR=2.97, 95%CI=1.94-4.55; P<0.001). The scores were analyzed for diagnostic performance with area under the curve (AUC) and did not help differentiate patients with an increased risk of adverse events. The differential diagnosis of syncope causes in patients with structural heart disease is important, because vasovagal and postural hypotension have better survival and less probability of emergency room or hospital readmission. The available scores are not reliable tools for prognosis in this specific patient population.