Oligonucleotide indexing of DNA barcodes: identification of tuna and other scombrid species in food products

<p>Abstract</p> <p>Background</p> <p>DNA barcodes are a global standard for species identification and have countless applications in the medical, forensic and alimentary fields, but few barcoding methods work efficiently in samples in which DNA is degraded, e.g. foods and archival specimens. This limits the choice of target regions harbouring a sufficient number of diagnostic polymorphisms. The method described here uses existing PCR and sequencing methodologies to detect mitochondrial DNA polymorphisms in complex matrices such as foods. The reported application allowed the discrimination among 17 fish species of the Scombridae family with high commercial interest such as mackerels, bonitos and tunas which are often present in processed seafood. The approach can be easily upgraded with the release of new genetic diversity information to increase the range of detected species.</p> <p>Results</p> <p>Cocktail of primers are designed for PCR using publicly available sequences of the target sequence. They are composed of a fixed 5' region and of variable 3' cocktail portions that allow amplification of any member of a group of species of interest. The population of short amplicons is directly sequenced and indexed using primers containing a longer 5' region and the non polymorphic portion of the cocktail portion. A 226 bp region of <it>CytB </it>was selected as target after collection and screening of 148 online sequences; 85 SNPs were found, of which 75 were present in at least two sequences. Primers were also designed for two shorter sub-fragments that could be amplified from highly degraded samples. The test was used on 103 samples of seafood (canned tuna and scomber, tuna salad, tuna sauce) and could successfully detect the presence of different or additional species that were not identified on the labelling of canned tuna, tuna salad and sauce samples.</p> <p>Conclusions</p> <p>The described method is largely independent of the degree of degradation of DNA source and can thus be applied to processed seafood. Moreover, the method is highly flexible: publicly available sequence information on mitochondrial genomes are rapidly increasing for most species, facilitating the choice of target sequences and the improvement of resolution of the test. This is particularly important for discrimination of marine and aquaculture species for which genome information is still limited.</p

Synthesis, Binding and Antiviral Properties of Potent Core-Extended Naphthalene Diimides Targeting the HIV-1 Long Terminal Repeat Promoter G-Quadruplexes

We have previously reported that stabilization of the G-quadruplex structures in the HIV-1 long terminal repeat (LTR) promoter suppresses viral transcription. Here we sought to develop new G-quadruplex ligands to be exploited as antiviral compounds by enhancing binding toward the viral G-quadruplex structures. We synthesized naphthalene diimide derivatives with a lateral expansion of the aromatic core. The new compounds were able to bind/stabilize the G-quadruplex to a high extent, and some of them displayed clear-cut selectivity toward the viral G-quadruplexes with respect to the human telomeric G-quadruplexes. This feature translated into low nanomolar anti-HIV-1 activity toward two viral strains and encouraging selectivity indexes. The selectivity depended on specific recognition of LTR loop residues; the mechanism of action was ascribed to inhibition of LTR promoter activity in cells. This is the first example of G-quadruplex ligands that show increased selectivity toward the viral G-quadruplexes and display remarkable antiviral activity

RNA-Sequence analysis of primary alveolar macrophages after in vitro infection with porcine reproductive and respiratory syndrome virus strains of differing virulence

Porcine reproductive and respiratory syndrome virus (PRRSV) mainly infects porcine alveolar macrophages (PAMs), resulting in porcine reproductive and respiratory syndrome (PRRS) in pigs. Most of the transcriptomic studies on PAMs infected with PRRSV conducted thus far have made use of microarray technology. Here, we investigated the transcriptome of PAMs in vitro at 12 h post-infection with two European PRRSV strains characterized by low (Lelystad, LV) and high (Lena) virulence through RNA-Seq. The expression levels of genes, isoforms, alternative transcription start sites (TSS) and differential promoter usage revealed a complex pattern of transcriptional and post-transcriptional gene regulation upon infection with the two strains. Gene ontology analysis confirmed that infection of PAMs with both the Lena and LV strains affected signaling pathways directly linked to the innate immune response, including interferon regulatory factors (IRF), RIG1-like receptors, TLRs and PKR pathways. The results confirmed that interferon signaling is crucial for transcriptional regulation during PAM infection. IFN-beta 1 and IFN-alpha omega, but not IFN-alpha, were up-regulated following infection with either the LV or Lena strain. The down-regulation of canonical pathways, such as the interplay between the innate and adaptive immune responses, cell death and TLR3/TLR7 signaling, was observed for both strains, but Lena triggered a stronger down-regulation than LV. This analysis contributes to a better understanding of the interactions between PRRSV and PAMs and outlines the differences in the responses of PAMs to strains with different levels of virulence, which may lead to the development of new PRRSV control strategies

Sensitive Detection and Quantification of Anisakid Parasite Residues in Food Products

Anisakids are nematodes whose larval stages are often present in fish, molluscs, and crustaceans. Members of the family Anisakidae belonging to the genera Anisakis and Pseudoterranova are implicated in human infections caused by the consumption of raw or undercooked fish. Adequate cooking will kill anisakid larvae, however, killed or inactivated larvae can still cause sensitization and immunoglobulin E-dependent hypersensitivity in human. This work describes the development of DNA-based tests to detect and quantify the presence of Anisakis spp. and Pseudoterranova spp. larvae in fish and fish-derived products, including fish fillets, surimi, fish sticks, canned fish, and baby food. Primers and TaqMan MGB probes recognizing only Anisakis spp. and Pseudoterranova spp. were designed on the first internal transcribed spacer 1 regions of rDNA for a real-time polymerase chain reaction assay. A commercial probe for 18S rDNA was used to detect and quantify the total eukaryotic DNA of the samples. The specificity and sensitivity of the assays were tested using reference samples prepared from mixtures made of Anisakis larvae in different quantity of codfish, and subsequent dilutions. Studies were performed to assess the ability of the test to detect and quantify anisakids in various products. Results showed that this test is able to detect anisakid DNA contained in a proportion of 1:10(5) in 1 ng of total DNA. The high prevalence of anisakids reported in main fishery species was confirmed by frequently detecting anisakids DNA in fish muscle and fish-derived products. A partial correlation was found between the number of larvae present in the viscera and the level of contamination of fish fillets. In conclusion, this molecular test is useful to detect the presence of Anisakis spp. and Pseudoterranova spp. in fish and fish-derived products and to quantify the level of contamination along the food chain, with potential applications for fish farms, fish markets, and food producers

Pig immune response to general stimulus and to porcine reproductive and respiratory syndrome virus infection: a meta-analysis approach

BACKGROUND: The availability of gene expression data that corresponds to pig immune response challenges provides compelling material for the understanding of the host immune system. Meta-analysis offers the opportunity to confirm and expand our knowledge by combining and studying at one time a vast set of independent studies creating large datasets with increased statistical power. In this study, we performed two meta-analyses of porcine transcriptomic data: i) scrutinized the global immune response to different challenges, and ii) determined the specific response to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) infection. To gain an in-depth knowledge of the pig response to PRRSV infection, we used an original approach comparing and eliminating the common genes from both meta-analyses in order to identify genes and pathways specifically involved in the PRRSV immune response. The software Pointillist was used to cope with the highly disparate data, circumventing the biases generated by the specific responses linked to single studies. Next, we used the Ingenuity Pathways Analysis (IPA) software to survey the canonical pathways, biological functions and transcription factors found to be significantly involved in the pig immune response. We used 779 chips corresponding to 29 datasets for the pig global immune response and 279 chips obtained from 6 datasets for the pig response to PRRSV infection, respectively. RESULTS: The pig global immune response analysis showed interconnected canonical pathways involved in the regulation of translation and mitochondrial energy metabolism. Biological functions revealed in this meta-analysis were centred around translation regulation, which included protein synthesis, RNA-post transcriptional gene expression and cellular growth and proliferation. Furthermore, the oxidative phosphorylation and mitochondria dysfunctions, associated with stress signalling, were highly regulated. Transcription factors such as MYCN, MYC and NFE2L2 were found in this analysis to be potentially involved in the regulation of the immune response. The host specific response to PRRSV infection engendered the activation of well-defined canonical pathways in response to pathogen challenge such as TREM1, toll-like receptor and hyper-cytokinemia/ hyper-chemokinemia signalling. Furthermore, this analysis brought forth the central role of the crosstalk between innate and adaptive immune response and the regulation of anti-inflammatory response. The most significant transcription factor potentially involved in this analysis was HMGB1, which is required for the innate recognition of viral nucleic acids. Other transcription factors like interferon regulatory factors IRF1, IRF3, IRF5 and IRF8 were also involved in the pig specific response to PRRSV infection. CONCLUSIONS: This work reveals key genes, canonical pathways and biological functions involved in the pig global immune response to diverse challenges, including PRRSV infection. The powerful statistical approach led us to consolidate previous findings as well as to gain new insights into the pig immune response either to common stimuli or specifically to PRRSV infection

Using SNP array data to test for host genetic and breed effects on Porcine Reproductive and Respiratory Syndrome Viremia

<p>Abstract</p> <p>Background</p> <p>The effect of breed on Porcine Reproductive and Respiratory Syndrome Viremia (PRRSV) was tested using data collected in 17 Italian commercial pig farms and 1096 genotypes obtained by the PorcineSNP60 BeadChip. A binomial logistic model was used to investigate the relationship between breed-clusters and PRRSV susceptibility. Breed-clusters were defined using the matrix of genomic kinship between all pairs of piglets.</p> <p>Results</p> <p>Only the contemporary group effect, defined as all piglets reared in the same herd, in the same year and whose samples were collected in the same season, was significant. Sex, age and breed-cluster showed no statistically significant effect on PRRS viremia, although the Landrace and Cross breed-clusters showed the lowest Odds-Ratio</p> <p>Conclusions</p> <p>The model failed to detect a significant breed-cluster effect, highlighting the impact of environment and management on PRRS viremia incidence. Incomplete exposure over the observed period may have masked possible breed differences.</p

Agreement technologies and their use in cloud computing environments

The final publication is available at Springer via http://dx.doi.org/10.1007/s13748-012-0031-9[EN] Nowadays, cloud computing is revolutionizing the services provided through the Internet to adapt itself in order to keep the quality of its services. Recent research foresees the advent of a new discipline of agent-based cloud computing systems that can make decisions about adaption in an uncertain environment. This paper discusses the role of argumentation in the next generation of agreement technologies and its use in cloud computing environments.This work is supported by the Spanish government (MICINN), project reference: TIN2012-36586-C03-01.Heras Barberá, SM.; De La Piedra, F.; Julian Inglada, VJ.; Rodríguez, S.; Botti Navarro, VJ.; Bajo, J.; Corchado, JM. (2012). Agreement technologies and their use in cloud computing environments. Progress in Artificial Intelligence. 1(4):277-290. https://doi.org/10.1007/s13748-012-0031-9S27729014European Comission: The Future of Cloud Computing. Technical report (2010)Barham, P., Dragovic, B., Fraser, K., Hand, S., Harris, T., Ho, A., Neugebauer, R., Pratt, I., Warfield, A.: Xen and the art of virtualization. In: SOSP03 Proceedings of the Nineteenth ACM Symposium on Operating Systems Principles, pp. 164–177. ACM, New York (2003)Wang, L., et. al.: Scientific cloud computing: early definition and experience. In: 10th IEEE International Conference on High Performance Computing and Communications (HPCC-08), pp. 825–830. IEEE Press (2008)Talia, D.: Clouds meet agents: toward intelligent cloud services. Internet Comput. IEEE 16(2), 78–81 (2012). doi: 10.1109/MIC.2012.28Heras, S.: Case-Based Argumentation Framework for Agent Societies. PhD thesis, Universitat Politècnica de València. http://hdl.handle.net/10251/12497 (2011)Ashton, K.: That ‘internet of things’ thing. RFID J. (2009). http://www.rfidjournal.com/article/view/4986Klusch, M.: Information agent technology for the Internet: a Survey. Data Knowl. Eng. 36, 337–372 (2001)Schaffer, H.E.: X as a Service. Cloud Computing, and the Need for Good Judgment IT Professional 11(5), 4–5 (2009). doi: 10.1109/MITP.2009.112Richardson, L., Ruby, S.: RESTful Web Services, Web services for the real world O’Reilly, Media, May, p. 454 (2007)GlusterFS Developers. The Gluster web site. http://www.gluster.org (2012)Chodorow, K., Dirolf, M.: The Definitive Guide. O’Reilly Media, MongoDB (2010)Fuentes-Fernandez, R., Hassan, S., Pavon, J., Galan, J.M., Lopez-Paredes, A.: Metamodels for role-driven agent-based modelling. Comput. Math. Organ. Theory 18(1), 91–112 (2012)Jordán, J., et al.: A customer support application using argumentation in multi-agent systems. In: 14th International Conference on, Information Fusion, pp. 772–778 (2011)Heras, S., Jordán, J., Botti, V., Julián, V.: Argue to agree: a case-based argumentation approach. Int. J. Approx. Reasoning (2012, in press)Walton, D., Reed, C., Macagno, F.: Argumentation Schemes. Cambridge University Press, Cambridge (2008)Bench-Capon, T., Sartor, G.: A model of legal reasoning with cases incorporating theories and values. Artif. Intell. 150(1–2), 97–143 (2003)Dignum, F., Weigand, H.: Communication and deontic logic. In: Information Systems Correctness and Reusability, pp. 242–260. World Scientific, Singapore (1995)Wooldridge, M., Jennings, N.R.: Intelligent agents: theory and practice. Knowl. Eng. Rev. 10(2), 115–152 (1995)Lopez-Rodriguez, I., Hernandez-Tejera, M.: Software agents as cloud computing services. In: 9th International Conference on Practical Applications of Agents and Multiagent Systems. Advances in Intelligent and Soft Computing, vol. 88, pp. 271–276. Springer, Berlin (2011)Sim, K.M.: Towards complex negotiation for cloud economy. In: 5th International Conference on Advances in Grid and Pervasive Computing. LNCS, vol. 6104, pp. 395–406. Springer, Berlin (2010)Aversa, R., et al.: Cloud agency: a mobile agent based cloud system. In: International Conference on Complex, Intelligent and Software Intensive Systems, pp. 132–137. IEEE Computer Society Press, Washington, DC (2010)Cao, B., et al.: A service-oriented qos-assured and multi-agent cloud computing architecture. In: 1st International Conference on Cloud Computing. LNCS, vol. 5931, pp. 644–649. Springer, Berlin (2009)Rahwan, I., Simari, G. (eds.): Argumentation in Artificial Intelligence. Springer, Berlin (2009

Genetic diversity and breed-informative SNPs identification in domestic pig populations using coding SNPs

Background: The use of breed-informative genetic markers, specifically coding Single Nucleotide Polymorphisms (SNPs), is crucial for breed traceability, authentication of meat and dairy products, and the preservation and improvement of pig breeds. By identifying breed informative markers, we aimed to gain insights into the genetic mechanisms that influence production traits, enabling informed decisions in animal management and promoting sustainable pig production to meet the growing demand for animal products.Methods: Our dataset consists of 300 coding SNPs genotyped from three Italian commercial pig populations: Landrace, Yorkshire, and Duroc. Firstly, we analyzed the genetic diversity among the populations. Then, we applied a discriminant analysis of principal components to identify the most informative SNPs for discriminating between these populations. Lastly, we conducted a functional enrichment analysis to identify the most enriched pathways related to the genetic variation observed in the pig populations.Results: The alpha diversity indexes revealed a high genetic diversity within the three breeds. The higher proportion of observed heterozygosity than expected revealed an excess of heterozygotes in the populations that was supported by negative values of the fixation index (FIS) and deviations from the Hardy-Weinberg equilibrium. The Euclidean distance, the pairwise FST, and the pairwise Nei’s GST genetic distances revealed that Yorkshire and Landrace breeds are genetically the closest, with distance values of 2.242, 0.029, and 0.033, respectively. Conversely, Landrace and Duroc breeds showed the highest genetic divergence, with distance values of 2.815, 0.048, and 0.052, respectively. We identified 28 significant SNPs that are related to phenotypic traits and these SNPs were able to differentiate between the pig breeds with high accuracy. The Functional Enrichment Analysis of the informative SNPs highlighted biological functions related to DNA packaging, chromatin integrity, and the preparation of DNA into higher-order structures.Conclusion: Our study sheds light on the genetic underpinnings of phenotypic variation among three Italian pig breeds, offering potential insights into the mechanisms driving breed differentiation. By prioritizing breed-specific coding SNPs, our approach enables a more focused analysis of specific genomic regions relevant to the research question compared to analyzing the entire genome

Agent-based virtual organization architecture

The purpose of this paper is to present the applicability of THOMAS, an architecture specially designed to model agent-based virtual organizations, in the development of a multiagent system for managing and planning routes for clients in a mall. In order to build virtual organizations, THOMAS offers mechanisms to take into account their structure, behaviour, dynamic, norms and environment. Moreover, one of the primary characteristics of the THOMAS architecture is the use of agents with reasoning and planning capabilities. These agents can perform a dynamic reorganization when they detect changes in the environment. The proposed architecture is composed of a set of related modules that are appropriate for developing systems in highly volatile environments similar to the one presented in this study. This paper presents THOMAS as well as the results obtained after having applied the system to a case study