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Mycoplasma Ovipneumoniae - A Primary Cause Of Severe Pneumonia Epizootics In The Norwegian Muskox (Ovibos Moschatus) Population, Torstein Tengs Dr.

Dr. Torstein Tengs

The Norwegian muskox (Ovibos moschatus) population lives on the high mountain plateau of Dovre and originates from animals introduced from Greenland. In the late summers of 2006 and 2012, severe outbreaks of pneumonia with mortality rates of 25-30% occurred. During the 2012 epidemic high quality samples from culled sick animals were obtained for microbiological and pathological examinations. High throughput sequencing (pyrosequencing) of pneumonic lung tissue revealed high concentrations of Mycoplasma ovipneumoniae in all six animals examined by this method and Pasteurella multocida subsp. multocida in four animals, whereas no virus sequences could be identified. Mycoplasma ovipneumoniae and P. multocida multocida …

Ultra-Deep Pyrosequencing Of Partial Surface Protein Genes From Infectious Salmon Anaemia Virus (Isav) Suggest Novel Mechanisms Involved In Transition To Virulence, Torstein Tengs Dr.

Dr. Torstein Tengs

Uncultivable HPR0 strains of infectious salmon anaemia viruses (ISAVs) infecting gills are non-virulent putative precursors of virulent ISAVs (vISAVs) causing systemic disease in farmed Atlantic salmon (Salmo salar). The transition to virulence involves two molecular events, a deletion in the highly polymorphic region (HPR) of the hemagglutinin-esterase (HE) gene and a Q266→L266 substitution or insertion next to the putative cleavage site (R267) in the fusion protein (F). We have performed ultra-deep pyrosequencing (UDPS) of these gene regions from healthy fish positive for HPR0 virus carrying full-length HPR sampled in a screening program, and a vISAV strain from an ISA outbreak …

Sequence Analysis Of The Genome Of Piscine Orthoreovirus (Prv) Associated With Heart And Skeletal Muscle Inflammation (Hsmi) In Atlantic Salmon (Salmo Salar), Torstein Tengs Dr.

Dr. Torstein Tengs

Piscine orthoreovirus (PRV) is associated with heart- and skeletal muscle inflammation (HSMI) of farmed Atlantic salmon (Salmo salar). We have performed detailed sequence analysis of the PRV genome with focus on putative encoded proteins, compared with prototype strains from mammalian (MRV T3D)- and avian orthoreoviruses (ARV-138), and aquareovirus (GCRV-873). Amino acid identities were low for most gene segments but detailed sequence analysis showed that many protein motifs or key amino acid residues known to be central to protein function are conserved for most PRV proteins. For M-class proteins this included a proline residue in μ2 which, for MRV, has been …

A Mobile Genetic Element With Unknown Function Found In Distantly Related Viruses, Torstein Tengs Dr.

Dr. Torstein Tengs

BACKGROUND

The genetic element s2m seems to represent one of very few examples of mobile genetic elements in viruses. The function remains obscure and a scattered taxonomical distribution has been reported by numerous groups.

METHODS

We have searched GenBank in order to identify all viral accessions that have s2m(-like) sequence motifs. Rigorous phylogenetic analyses and constrained tree topology testing were also performed in order to investigate the apparently mobile nature of s2m.

RESULTS

The stem-loop s2m structure can be found in four families of + ssRNA viruses; Astroviridae, Caliciviridae, Picornaviridae and Coronaviridae. In all of these virus families, with the …

Piscine Myocarditis Virus (Pmcv) In Wild Atlantic Salmon Salmo Salar, Torstein Tengs Dr.

Dr. Torstein Tengs

Cardiomyopathy syndrome (CMS) is a severe cardiac disease of sea-farmed Atlantic salmon Salmo salar L., but CMS-like lesions have also been found in wild Atlantic salmon. In 2010 a double-stranded RNA virus of the Totiviridae family, provisionally named piscine myocarditis virus (PMCV), was described as the causative agent of CMS. In the present paper we report the first detection of PMCV in wild Atlantic salmon. The study is based on screening of 797 wild Atlantic salmon by real-time RT-PCR. The samples were collected from 35 different rivers along the coast of Norway, and all individuals included in the study were …

Prevalence Of Tick Borne Encephalitis Virus In Tick Nymphs In Relation To Climatic Factors On The Southern Coast Of Norway, Torstein Tengs Dr.

Dr. Torstein Tengs

BACKGROUND

Tick-borne encephalitis (TBE) is among the most important vector borne diseases of humans in Europe and is currently identified as a major health problem in many countries. TBE endemic zones have expanded over the past two decades, as well as the number of reported cases within endemic areas. Multiple factors are ascribed for the increased incidence of TBE, including climatic change. The number of TBE cases has also increased in Norway over the past decade, and the human cases cluster along the southern coast of Norway. In Norway the distribution and prevalence of TBE virus (TBEV) in tick populations …

A Strain Of Piscine Myocarditis Virus (Pmcv) Infecting Argentina Silus (Ascanius), Torstein Tengs Dr.

Dr. Torstein Tengs

No abstract.

Quantification Of Piscine Reovirus (Prv) At Different Stages Of Atlantic Salmon Salmo Salar Production, Torstein Tengs Dr.

Dr. Torstein Tengs

The newly described piscine reovirus (PRV) appears to be associated with the development of heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon Salmo salar L. PRV seems to be ubiquitous among fish in Norwegian salmon farms, but high viral loads and tissue distribution support a causal relationship between virus and disease. In order to improve understanding of the distribution of PRV in the salmon production line, we quantified PRV by using real-time PCR on heart samples collected at different points in the life cycle from pre-smolts to fish ready for slaughter. PRV positive pre-smolts were found in about …

Prevalence Of Piscine Myocarditis Virus (Pmcv) In Marine Fish Species, Torstein Tengs Dr.

Dr. Torstein Tengs

No abstract.

A Novel Totivirus And Piscine Reovirus (Prv) In Atlantic Salmon (Salmo Salar) With Cardiomyopathy Syndrome (Cms), Torstein Tengs

Dr. Torstein Tengs

BACKGROUNDCardiomyopathy syndrome (CMS) is a severe disease affecting large farmed Atlantic salmon. Mortality often appears without prior clinical signs, typically shortly prior to slaughter. We recently reported the finding and the complete genomic sequence of a novel piscine reovirus (PRV), which is associated with another cardiac disease in Atlantic salmon; heart and skeletal muscle inflammation (HSMI). In the present work we have studied whether PRV or other infectious agents may be involved in the etiology of CMS.RESULTSUsing high throughput sequencing on heart samples from natural outbreaks of CMS and from fish experimentally challenged with material from fish diagnosed with CMS …

Heart And Skeletal Muscle Inflammation Of Farmed Salmon Is Associated With Infection With A Novel Reovirus, Torstein Tengs

Dr. Torstein Tengs

Atlantic salmon (Salmo salar L.) mariculture has been associated with epidemics of infectious diseases that threaten not only local production, but also wild fish coming into close proximity to marine pens and fish escaping from them. Heart and skeletal muscle inflammation (HSMI) is a frequently fatal disease of farmed Atlantic salmon. First recognized in one farm in Norway in 1999, HSMI was subsequently implicated in outbreaks in other farms in Norway and the United Kingdom. Although pathology and disease transmission studies indicated an infectious basis, efforts to identify an agent were unsuccessful. Here we provide evidence that HSMI is associated …

Non-Prejudiced Detection And Characterization Of Genetic Modifications, Torstein Tengs

Dr. Torstein Tengs

The application of gene technology is becoming widespread much thanks to the rapid increase in technology, resource, and knowledge availability. Consequently, the diversity and number of genetically modified organisms (GMOs) that may find their way into the food chain or the environment, intended or unintended, is rapidly growing. From a safety point of view the ability to detect and characterize in detail any GMO, independent of publicly available information, is fundamental. Pre-release risk assessments of GMOs are required in most jurisdictions and are usually based on application of technologies with limited ability to detect unexpected rearrangements and insertions. We present …

Comparison Of Nine Different Real-Time Pcr Chemistries For Qualitative And Quantitative Applications In Gmo Detection, Torstein Tengs

Dr. Torstein Tengs

Several techniques have been developed for detection and quantification of genetically modified organisms, but quantitative real-time PCR is by far the most popular approach. Among the most commonly used realtime PCR chemistries are TaqMan probes and SYBR green, but many other detection chemistries have also been developed. Because their performance has never been compared systematically, here we present an extensive evaluation of some promising chemistries: sequenceunspecific DNA labeling dyes (SYBR green), primer-based technologies (AmpliFluor, Plexor, Lux primers), and techniques involving double-labeled probes, comprising hybridization (molecular beacon) and hydrolysis (TaqMan, CPT, LNA, and MGB) probes, based on recently published experimental data. …

Uniqueprimer - A Web Utility For Design Of Specific Pcr Primers And Probes, Torstein Tengs

Dr. Torstein Tengs

We have developed a web-based tool for design of specific PCR primers and probes. The program allows you to enter primer sequence information as well as an optional probe, and sequence similarity searches (MegaBLAST) will be performed to see if the sequences match the same sequence entry in the specified database. If primers (and probe) match, this will be reported. The program can handle overlapping amplicons, amplification from a single primer, ambiguous bases and other problematic cases.

A Quantitative Taqman Mgb Real-Time Polymerase Chain Reaction Based Assay For Detection Of The Causative Agent Of Crayfish Plague Aphanomyces Astaci, Torstein Tengs

Dr. Torstein Tengs

Here we present the development and first validation of a TaqMan minor groove binder (MGB) real-time polymerase chain reaction (RT-PCR) method for quantitative and highly specific detection of Aphanomyces astaci, the causative agent of crayfish plague. The assay specificity was experimentally assessed by testing against DNA representative of closely related oomycetes, and theoretically assessed by additional sequence similarity analyses comparing the primers and probe sequences to available sequences in EMBL/GenBank. The target of the assay is a 59 bp unique sequence motif of A. astaci found in the internal transcribed spacer 1 of the nuclear ribosomal gene cluster. A standard …

A Statistical Approach For Evaluation Of Pcr Results To Improve The Practical Limit Of Quantification (Loq) Of Gmo Analyses (Simquant), Torstein Tengs

Dr. Torstein Tengs

The predominant approach for quantification of genetically modified organisms (GMO) is the application of quantitative real-time PCR. However, for a large number of processed food and feed products, this approach is unsuitable, because they contain low amounts (mass) of amplifiable DNA. Here we present a novel approach, ‘‘Single molecule quantification’’ (SIMQUANT) for GMO quantification of samples with extremely low amounts of DNA. The approach is based on statistics and application of multiple qualitative parallel PCRs. Here the qualitative PCRs were done using real-time PCR setup, but this is not a requirement. The difference is that the quantitative real-time PCR requires …

Francisella Philomiragia Subsp. Noatunensis Subsp. Nov., Isolated From Farmed Atlantic Cod (Gadus Morhua L.), Torstein Tengs

Dr. Torstein Tengs

Seven bacterial isolates from farmed Atlantic cod displaying chronic granulomatous disease were characterized by phenotypic and molecular taxonomic methods. The isolates were Gram-negative, facultatively intracellular, non-motile, strictly aerobic coccobacilli which produced H2S from cysteine-supplemented media and are therefore phenotypically consistent with members of the genus Francisella. Comparison of 16S rRNA gene sequences and six partial housekeeping gene sequences (groEL, shdA, rpoB, rpoA, pgm and atpA) confirmed the organism as a member of the genus Francisella, with Francisella philomiragia as its closest relative (99.3% 16S rRNA gene sequence similarity, 92.2–99.0% housekeeping gene sequence similarity). Despite the close relationship with F. philomiragia, …

Phenotypically Different Microalgal Morphospecies With Identical Ribosomal Rna: A Case Of Rapid Adaptive Evolution?, Torstein Tengs

Dr. Torstein Tengs

The agents driving the divergence and speciation of freeliving microbial populations are still largely unknown. We investigated the dinoflagellate morphospecies Scrippsiella hangoei and Peridinium aciculiferum, which abound in the Baltic Sea and in northern temperate lakes, respectively. Electron microscopy analyses showed significant interspecific differences in the external cellular morphology, but a similar plate pattern in the characteristic dinoflagellate armor. Experimentally, S. hangoei grew in a wide range of salinities (0–30), whereas P. aciculiferum only grew in low salinities (0–3). Despite these phenotypic differences and the habitat segregation, molecular analyses showed identical ribosomal DNA sequences (ITS1, ITS2, 5.8S, SSU, and partial …

Raphidophyceae [Chadefaud Ex Silva] Systematics And Rapid Identification: Sequence Analyses And Real–Time Pcr Assays, Torstein Tengs

Dr. Torstein Tengs

Species within the class Raphidophyceae were associated with fish kill events in Japanese, European, Canadian, and U.S. coastal waters. Fish mortality was attributable to gill damage with exposure to reactive oxygen species (peroxide, superoxide, and hydroxide radicals), neurotoxins, physical clogging, and hemolytic substances. Morphological identification of these organisms in environmental water samples is difficult, particularly when fixatives are used. Because of this difficulty and the continued global emergence of these species in coastal estuarine waters, we initiated the development and validation of a suite of real-time polymerase chain reaction (PCR) assays. Sequencing was used to generate complete data sets for …

Equal Performance Of Taqman, Mgb, Molecular Beacon, And Sybr Green-Based Detection Assays In Detection And Quantification Of Roundup Ready Soybean, Torstein Tengs

Dr. Torstein Tengs

We have tested and compared the performance of 12 different assays representing four different real-time polymerase chain reaction (PCR) chemistries in the context of genetically modified organism detection. Several different molecular beacon, SYBR Green, TaqMan, and MGB assays were designed for the event specific detection and quantification of the 3' integration junction of GTS 40-3-2 (Roundup Ready) soybean. Sensitivity as well as robustness in the presence of background DNA were tested. None of the PCR-based approaches appeared to be significantly better than any of the other, but the molecular beacon assays had the lowest efficiency and also seemed more sensitive …

Heterotachy Processes In Rhodophyte-Derived Secondhand Plastid Genes: Implications For Addressing The Origin And Evolution Of Dinoflagellate Plastids, Torstein Tengs

Dr. Torstein Tengs

Serial transfer of plastids from one eukaryotic host to another is the key process involved in evolution of secondhand plastids. Such transfers drastically change the environment of the plastids and hence the selection regimes, presumably leading to changes over time in the characteristics of plastid gene evolution and to misleading phylogenetic inferences. About half of the dinoflagellate protists species are photosynthetic and unique in harboring a diversity of plastids acquired from a wide range of eukaryotic algae. They are therefore ideal for studying evolutionary processes of plastids gained through secondary and tertiary endosymbioses. In the light of these processes, we …

Sensitive Mutation Detection In Heterogeneous Cancer Specimens By Massively Parallel Picoliter Reactor Sequencing, Torstein Tengs

Dr. Torstein Tengs

The sensitivity of conventional DNA sequencing in tumor biopsies is limited by stromal contamination and by genetic heterogeneity within the cancer. Here, we show that microreactor-based pyrosequencing can detect rare cancer-associated sequence variations by independent and parallel sampling of multiple representatives of a given DNA fragment. This technology can thereby facilitate accurate molecular diagnosis of heterogeneous cancer specimens and enable patient selection for targeted cancer therapies.

A Group Of Dinoflagellates Similar To Pfiesteria As Defined By Morphology And Genetic Analysis, Torstein Tengs

Dr. Torstein Tengs

Eight novel pfiesteria-similar dinoflagellates are described that form a genetically well-defined group with distinct morphological features, and are capable of feeding myzocytotically on cryptophytes and other algae. Taxonomically, there was concordance between the genetic analysis and plate tabulation features among these taxa. It is suggested that these dinoflagellates are important grazers of other phytoplankton in the environment, and their total ecological significance has been underestimated.

Detection Of Oncogenic Mutations In The Egfr Gene In Lung Adenocarcinoma With Differential Sensitivity To Egfr Tyrosine Kinase Inhibitors, Torstein Tengs

Dr. Torstein Tengs

The complete sequencing of the human genome and the development of molecularly targeted cancer therapy have promoted efforts to identify systematically the genetic alterations in human cancer. By high-throughput sequencing of tyrosine kinase genes in human non-small-cell lung cancer, we identified somatic mutations in the kinase domain of the epidermal growth factor receptor tyrosine kinase gene (EGFR) that are correlated with clinical response to EGFR tyrosine kinase inhibitors (TKIs). We have shown that these mutant forms of EGFR induce oncogenic transformation in different cellular systems. Cells whose growth depends on EGFR with mutations in exons 19 and 21 are sensitive …

Highly Divergent Ssu Rrna Genes Found In The Marine Ciliates Myrionecta Rubra And Mesodinium Pulex, Torstein Tengs

Dr. Torstein Tengs

Myrionecta rubra and Mesodinium pulex are among the most commonly encountered planktonic ciliates in coastal marine and estuarine regions throughout the world. Despite their widespread distribution, both ciliates have received little attention by taxonomists. In order to better understand the phylogenetic position of these ciliates, we determined the SSU rRNA gene sequence from cultures of M. rubra and M. pulex. Partial sequence data were also generated from isolated cells of M. rubra from Chesapeake Bay. The M. rubra and M. pulex sequences were very divergent from all other ciliates, but shared a branch with 100% bootstrap support. Both species had …

Genomic Representations Using Concatenates Of Type Iib Restriction Endonuclease Digestion Fragments, Torstein Tengs

Dr. Torstein Tengs

We have developed a method for genomic representation using Type IIB restriction endonucleases. Representation by concatenation of restriction digests, or RECORD, is an approach to sample the fragments generated by cleavage with these enzymes. Here, we show that the RECORD libraries may be used for digital karyotyping and for pathogen identification by computational subtraction.

Development Of Real–Time Pcr Assays For The Detection Of Chattonella Species In Culture And Environmental Samples, Torstein Tengs

Dr. Torstein Tengs

Raphidophytes have been associated with fish kill events in Japanese, European and U.S. coastal waters, and many produce toxins that can pose a threat to human health. The recognition of raphidophytes as HAB species in mid-Atlantic estuarine waters led us to initiate molecular phylogenetic analyses of these taxa and to develop real-time PCR assays for rapid detection of important species. The molecular studies and PCR detection methods will enhance ongoing taxonomic, toxicologic and ecological of these organisms and will be a useful tool in HAB monitoring programs.

Kryptoperidinium Foliaceum Blooms In South Carolina: A Multi–Analytical Approach To Identification, Torstein Tengs

Dr. Torstein Tengs

Observations following the discovery of Kryptoperidinium foliaceum blooms in South Carolina (SC), USA, suggest that a multi-analytical approach, using a standard, minimal set of criteria, should be adopted for determining dinoflagellate species identity and taxonomic placement. A combination of morphological, molecular, and biochemical analyses were used to determine the identity of this “red tide” dinoflagellate, first documented in SC waters in the spring of 1998. Results from thecal plate tabulations (based on scanning electron and epifluorescence microscopy), gene sequence data, species-specific PCR probe assays, and microalgal pigment profiles were analyzed and compared to reference cultures of K. foliaceum. Comparative data …

No Difference Found In Ribosomal Dna Sequences From Physiologically Diverse Clones Of Karenia Brevis (Dinophyceae) From The Gulf Of Mexico, Torstein Tengs

Dr. Torstein Tengs

Maximum growth rate and toxin content were significantly different among five strains of Karenia brevis isolated from Texas and Florida when grown under identical culture conditions. Sequence analysis of the 18S rRNA gene and internal transcribed spacer (ITS) regions revealed, however, that all five strains were identical. Consequently, a clear genetic basis for physiological variability among various geographical isolates of K. brevis from the Gulf of Mexico could not be assessed using these genetic markers. Both the ITS and 18S rRNA regions may be useful in species-specific probe selection. At the intra-specific level, however, an alternative marker will be needed …

Discovery Of The Toxic Dinoflagellate Pfiesteria In Northern European Waters, Torstein Tengs

Dr. Torstein Tengs

Several dinoflagellate strains of the genus Pfiesteria were isolated by culturing techniques from sediment samples taken in the Oslofjord region of Norway. Pfiesteria piscicida, well known as a fish killer from the Atlantic coast of America, was identified by genetic methods and light microscopy. The related species Pfiesteria shumwayae was attracted from the sediment by the presence of fish, and has proved toxic. This present survey demonstrates the wide distribution of these potentially harmful species, but so far they have not been connected with fish kills in Europe.