Klaus Nielsen

Bio: Klaus Nielsen is an academic researcher from Canadian Food Inspection Agency. The author has contributed to research in topics: Immunoassay & Complement fixation test. The author has an hindex of 36, co-authored 106 publications receiving 3837 citations. Previous affiliations of Klaus Nielsen include Nielsen Holdings N.V. & Agriculture and Agri-Food Canada.

Diagnosis of brucellosis by serology.

Abstract: Serological diagnosis of brucellosis began more than 100 years ago with a simple agglutination test. It was realized that this type of test was susceptible to false positive reactions resulting from, for instance, exposure to cross reacting microorganisms. It was also realized that this test format was inexpensive, simple and could be rapid, although results were subjectively scored. Therefore, a number of modifications were developed along with other types of tests. This served two purposes: one was to establish a rapid screening test with high sensitivity and perhaps less specificity and a confirmatory test, usually more complicated but also more specific, to be used on sera that reacted positively in screening tests. This led to another problem: if a panel of tests were performed and they did not all agree, which interpretation was correct? This problem was further compounded by the extensive use of a vaccine which gave rise to an antibody response similar to that resulting from field infection. This led to the development of an assay that could distinguish vaccinal antibody, starting with precipitin tests. These tests did not perform well, giving rise to the development of primary binding assays. These assays, including the competitive enzyme immunoassay and the fluorescence polarization assay are at the apex of current development, providing high sensitivity and specificity as well as speed and mobility in the case of the fluorescence polarization assay.

Diagnosis of Brucellosis in Livestock and Wildlife

Abstract: Aim To describe and discuss the merits of various direct and indirect methods applied in vitro (mainly on blood or milk) or in vivo (allergic test) for the diagnosis of brucellosis in animals. Methods The recent literature on brucellosis diagnostic tests was reviewed. These diagnostic tests are applied with different goals, such as national screening, confirmatory diagnosis, certification, and international trade. The valida tion of such diagnostic tests is still an issue, particularly in wildlife. The choice of the testing strategy depends on the prevailing brucellosis epidemiological situation and the

Review of detection of Brucella spp. by polymerase chain reaction.

Abstract: Here we present a review of most of the currently used polymerase chain reaction (PCR)-based methods for identification of Brucella bacteria in biological samples. We focused in particular on methods using single-pair primers, multiplex primers, real-time PCRs, PCRs for marine Brucella, and PCRs for molecular biotyping. These methods are becoming very important tools for the identification of Brucella, at the species level and recently also at the biovar level. These techniques require minimum biological containment and can provide results in a very short time. In addition, genetic fingerprinting of isolates aid in epidemiological studies of the disease and its control. PCR-based methods are more useful and practical than conventional methods used to identify Brucella spp., and new methods for Brucella spp identification and typing are still being developed. However, the sensitivity, specificity, and issues of quality control and quality assurance using these methods must be fully validated on clinical samples before PCR can be used in routine laboratory testing for brucellosis.

Diagnosis of Brucellosis

Abstract: Brucellosis is an important zoonosis and a significant cause of reproductive losses in animals. Abortion, placentitis, epididymitis, and orchitis are the most common clinical manifestations in animals. In humans, brucellosis is a debilitating and chronic disease, which may affect a variety of organs. Clinical diagnosis of brucellosis is not easily achieved. Laboratory testing is therefore very important for a correct identification of the disease in humans and for the detection and confirmation in animals. Definitive diagnosis is normally done by isolation and identification of the causative agent. While definitive, isolation is time-consuming, must be performed by highly skilled personnel, and it is hazardous. For these reasons, serological tests are normally preferred. Brucellosis serology have advanced considerably in the last decades with very sensitive and specific new tests available. Modern genetic characterization of Brucellae using molecular DNA technology have been developed. Several PCR-based assays have been proposed, from the rapid recognition of genus to differential identification of species and strains. This review describes bacteriological, serological, and molecular methods used for the diagnosis of human and animal brucellosis.

A homogeneous fluorescence polarization assay for detection of antibody to Brucella abortus

Abstract: A homogeneous fluorescence polarization (FP) assay (FPA) was developed for detection of antibody in bovine sera to Brucella abortus The assay used O-polysaccharide prepared from B abortus lipopolysaccharide in the molecular weight range of 20-30 kDa which was conjugated with fluorescein isothiocyanate and used as a tracer Fluorescence polarization was measured with a FPM-1 fluorescence polarization analyzer Sample (20 microliters) was added to 20 ml of diluent buffer at ambient temperature A serum blank reading was taken and tracer (10 microliters) to yield approx 15 nM fluorescein equivalents was added The FP of the tracer was determined after a period of greater than 2 min A positive reaction was indicated by a significant elevation of the FP reading over the negative control In a blind study, 9480 bovine sera were tested in addition to sets of four controls which were included with each lot of 100 samples tested The controls were a strong positive, a weak positive, a negative and a serum derived from a B abortus strain 19 vaccinated cow Test sera included 8669 sera from Canadian cattle which were negative by routine serological tests, 561 sera from cows from which B abortus had been isolated either from tissues or milk and 250 sera from cattle previously vaccinated with B abortus strain 19 at various times One lot of O-polysaccharide tracer was used for all tests The initial cut-off for negative samples in the fluorescence polarization assay was set at 1072 mP This resulted in a sensitivity estimate of 981 +/- 11% and the specificity was 998 +/- 009% After decoding the samples and retesting false positive and negative reactions, the sensitivity estimate was 985 +/- 10% and the specificity was 100% It became evident that the initial cut-off value was set too high and, using ROC analysis, a cut-off of 90 mP increased the sensitivity to 9902% while the specificity decreased to 9996% Of the 250 sera from vaccinated cattle, 248 were negative giving a point specificity value of 992%

The ferritins: molecular properties, iron storage function and cellular regulation☆

Abstract: The iron storage protein, ferritin, plays a key role in iron metabolism. Its ability to sequester the element gives ferritin the dual functions of iron detoxification and iron reserve. The importance of these functions is emphasised by ferritin's ubiquitous distribution among living species. Ferritin's three-dimensional structure is highly conserved. All ferritins have 24 protein subunits arranged in 432 symmetry to give a hollow shell with an 80 A diameter cavity capable of storing up to 4500 Fe(III) atoms as an inorganic complex. Subunits are folded as 4-helix bundles each having a fifth short helix at roughly 60° to the bundle axis. Structural features of ferritins from humans, horse, bullfrog and bacteria are described: all have essentially the same architecture in spite of large variations in primary structure (amino acid sequence identities can be as low as 14%) and the presence in some bacterial ferritins of haem groups. Ferritin molecules isolated from vertebrates are composed of two types of subunit (H and L), whereas those from plants and bacteria contain only H-type chains, where ‘H-type’ is associated with the presence of centres catalysing the oxidation of two Fe(II) atoms. The similarity between the dinuclear iron centres of ferritin H-chains and those of ribonucleotide reductase and other proteins suggests a possible wider evolutionary linkage. A great deal of research effort is now concentrated on two aspects of fenitin: its functional mechanisms and its regulation. These form the major part of the review. Steps in iron storage within ferritin molecules consist of Fe(II) oxidation, FE(III) migration and the nucleation and growth of the iron core mineral. H-chains are important for Fe(II) oxidation and L-chains assist in core formation. Iron mobilisation, relevant to ferritin's role as iron reserve, is also discussed. Translational regulation of mammalian ferritin synthesis in response to iron and the apparent links between iron and citrate metabolism through a single molecule with dual function are described. The molecule, when binding a [4Fe-4S] cluster, is a functioning (cytoplasmic) aconitase. When cellular iron is low, loss of the [4Fe-4S] cluster allows the molecule to bind to the 5′-untranslated region (5′-UTR) of the ferritin m-RNA and thus to repress translation. In this form it is known as the iron regulatory protein (IRP) and the stem-loop RNA structure to which it binds is the iron regulatory element (IRE). IREs are found in the 3′-UTR of the transferrin receptor and in the 5′-UTR of erythroid aminolaevulinic acid synthase, enabling tight co-ordination between cellular iron uptake and the synthesis of ferritin and haem. Degradation of ferritin could potentially lead to an increase in toxicity due to uncontrolled release of iron. Degradation within membrane-encapsulated ‘secondary lysosomes’ may avoid this problem and this seems to be the origin of another form of storage iron known as haemosiderin. However, in certain pathological states, massive deposits of ‘haemosiderin’ are found which do not arise directly from ferritin breakdown. Understanding the numerous inter-relationships between the various intracellular iron complexes presents a major challenge.

Iron metabolism in pathogenic bacteria.

Abstract: ▪ Abstract The ability of pathogens to obtain iron from transferrins, ferritin, hemoglobin, and other iron-containing proteins of their host is central to whether they live or die. To combat invading bacteria, animals go into an iron-withholding mode and also use a protein (Nramp1) to generate reactive oxygen species in an attempt to kill the pathogens. Some invading bacteria respond by producing specific iron chelators—siderophores—that remove the iron from the host sources. Other bacteria rely on direct contact with host iron proteins, either abstracting the iron at their surface or, as with heme, taking it up into the cytoplasm. The expression of a large number of genes (>40 in some cases) is directly controlled by the prevailing intracellular concentration of Fe(II) via its complexing to a regulatory protein (the Fur protein or equivalent). In this way, the biochemistry of the bacterial cell can accommodate the challenges from the host. Agents that interfere with bacterial iron metabolism may prove ex...

Lateral flow (immuno)assay: its strengths, weaknesses, opportunities and threats. A literature survey.

Abstract: Lateral flow (immuno)assays are currently used for qualitative, semiquantitative and to some extent quantitative monitoring in resource-poor or non-laboratory environments. Applications include tests on pathogens, drugs, hormones and metabolites in biomedical, phytosanitary, veterinary, feed/food and environmental settings. We describe principles of current formats, applications, limitations and perspectives for quantitative monitoring. We illustrate the potentials and limitations of analysis with lateral flow (immuno)assays using a literature survey and a SWOT analysis (acronym for “strengths, weaknesses, opportunities, threats”). Articles referred to in this survey were searched for on MEDLINE, Scopus and in references of reviewed papers. Search terms included “immunochromatography”, “sol particle immunoassay”, “lateral flow immunoassay” and “dipstick assay”.

Diseases of Poultry

Abstract: VERY few veterinary surgeons have thought fit to write a book on diseases of poultry. Mr. Ernest Gray has done justice to the subject and is to be congratulated on his effort. A book of this size, written by one with specialized knowledge, will add to the value of any library or private bookshelf. Diseases of Poultry Their Aetiology, Diagnosis, Treatment and Control; with a Section on the Normal Anatomy and Physiology of the Fowl. By Ernest Gray. (Lockwood's Agricultural and Horticultural Handbooks.) Pp. x + 198 + 16 plates. (London: Crosby Lockwood and Son, Ltd., 1940.) 9s. 6d. net.

OIE Manual of Diagnostic Tests and Vaccines for Terrestrial Animals

Abstract: The most recent version of the OIE Manual of Diagnostic Tests and Vaccines for Terrestrial Animals (the Terrestrial Manual) is online: http://www.oie.int/en/international...