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Biomedical subjects

K Nielsen

Publications and source records attributed to K Nielsen.

At least 19 recordsLinked to original sources
PubMed2006

Phi c31 integrase induces chromosomal aberrations in primary human fibroblasts.

Phi c31 integrase is investigated as a novel tool for nonviral gene therapy as the enzyme can direct site-specific integration into a host chromosome. In order to investigate effects of phi c31 integrase expression in normal human cells, we have generated stably transfected primary human fibroblasts expressing the enzyme. All control cells were cytogenetically normal, but in cells expressing phi c31 integrase, numerous chromosomal abnormalities including various translocations were found, suggesting that the enzyme itself acts as a mutagen.

Attachment Sites, Microbiological↗
PubMed2006

Development of an indirect enzyme-linked immunosorbent assay for detecting equine serum antibodies to the lipopolysaccharide of Salmonella abortusequi.

An indirect enzyme-linked immunosorbent assay (IELISA) was developed for the detection of equine serum antibodies to lipopolysaccharide of Salmonella enterica subsp. enterica serovar Abortusequi (LPS), a causative organism of Equine Paratyphoid. The data presented demonstrates that horses immunized with S. abortusequi LPS developed antibodies detectable by the IELISA. By comparison, the tube agglutination test (TAT) did not detect antibody to S. abortusequi LPS as consistently as the IELISA. The data suggests that the IELISA may be a more suitable test for the detection of serum antibodies to S. abortusequi than the TAT.

Agglutination Tests↗
PubMed2006

Rapid, field-adapted indirect enzyme-linked immunosorbent assay for detection of antibodies in bovine whole blood and serum to Brucella abortus.

A simple, rapid, field-adapted indirect enzyme-linked immunoassay (FldELISA) for the detection of antibodies to Brucella abortus in whole blood and serum has been developed. This assay detects antibodies to B. abortus in approximately 15 min or less. Over a 3-month period, this assay has consistently identified immunized and nonimmunized animals, while the percent coefficient of variation for each immunized animal has been less than 20%. As with any indirect enzyme-linked immunoassay, quality control can be established and maintained. Using defined positive and negative sera, the sensitivity and specificity of the FldELISA was 100% and 94.2%, respectively. As a model, this test can be readily extended to other disease applications that use lipopolysaccharide or other stable antigens for the detection of antibodies, such as those to Salmonella spp., Escherichia coli, or Yersinia spp.

Animals↗
PubMed2005

Analysis of artificially degraded DNA using STRs and SNPs--results of a collaborative European (EDNAP) exercise.

Recently, there has been much debate about what kinds of genetic markers should be implemented as new core loci that constitute national DNA databases. The choices lie between conventional STRs, ranging in size from 100 to 450 bp; mini-STRs, with amplicon sizes less than 200 bp; and single nucleotide polymorphisms (SNPs). There is general agreement by the European DNA Profiling Group (EDNAP) and the European Network of Forensic Science Institutes (ENFSI) that the reason to implement new markers is to increase the chance of amplifying highly degraded DNA rather than to increase the discriminating power of the current techniques. A collaborative study between nine European and US laboratories was organised under the auspices of EDNAP. Each laboratory was supplied with a SNP multiplex kit (Foren-SNPs) provided by the Forensic Science Service, two mini-STR kits provided by the National Institute of Standards and Technology (NIST) and a set of degraded DNA stains (blood and saliva). Laboratories tested all three multiplex kits, along with their own existing DNA profiling technique, on the same sets of degraded samples. Results were collated and analysed and, in general, mini-STR systems were shown to be the most effective. Accordingly, the EDNAP and ENFSI working groups have recommended that existing STR loci are reengineered to provide smaller amplicons, and the adoption of three new European core loci has been agreed.

Analysis of Variance↗
PubMed2005

Comparison of fluorescence polarization assay with card and complement fixation tests for the diagnosis of goat brucellosis in a high-prevalence area.

An evaluation of fluorescence polarization assay (FPA) to detect antibodies against Brucella melitensis according to the Mexican Official Norm (NOM) was performed. In this study, a total of 2582 goat serum samples from a high-prevalence area in northeast Mexico where vaccination is applied, were used. Of these, 1094 were classified as NOM negatives (card test (CT) negatives or CT positives/complement fixation test (CFT) negatives) and 1488 as NOM positives (CT and CFT positives). The receiver operator characteristics (ROC) curve analysis was used to obtain the FPA sensitivity (83.5%), specificity (82.2%) and accuracy (88.2%) compared with NOM criteria, using a cut-off value of 89mP for positive samples. In addition, FPA produced 84.1% of negative results versus 65.7% of CT using 1094 CFT negative samples, which indicated that FPA performance was better than CT to detect negative samples or differentiate samples from vaccinated animals. Finally, FPA showed 95.8% sensitivity when using 702 negative non-vaccinated samples. Taken together, these results suggested that FPA might replace CT as a screening test for its better performance compared with CFT, its adjustable cut-off useful in different epidemiological situations, and for its reliability, ease of performance, comparable cost with CT regimen, and potential application in field and high-throughput laboratories. The use of FPA as screening test will help to reduce the percentage of goats wrongly slaughtered because of brucellosis misdiagnosis. More studies on FPA are required for its approval as diagnostic tool for goat brucellosis.

Agglutination Tests↗
PubMed2005

Serological discrimination by indirect enzyme immunoassay between the antibody response to Brucella sp. and Yersinia enterocolitica O:9 in cattle and pigs.

A rapid, inexpensive and rugged serological test that distinguishes cattle and swine infected with Brucella sp. or Yersinia enterocolitica O:9 is described. The test protocol, which is an indirect enzyme immunoassay uses a high concentration of divalent cation chelating agents to minimize binding of Y. enterocolitica O:9 antibody to rough lipopolysaccharide antigen derived from B. abortus RB51. No false positive reactions were observed when testing 100 Canadian cattle and swine without any evidence of brucellosis. The assay detected 91.6% of cattle (n=155) and 93.5% (n=31) of swine infected with Brucella sp. Sera from 58 cattle and 38 swine exposed to Y. enterocolitica O:9 were negative while only 20 sera from 121 'false positive' reactors of unspecified origin gave low level positive reactions, eliminating 84% of the false positive reactions.

Animals↗
PubMed2005

Combined ilioinguinal blockade and local infiltration anaesthesia for groin hernia repair--a double-blind randomized study.

BACKGROUND: Local infiltration anaesthesia for inguinal hernia repair is cost-effective, but fear of intra-operative pain may hinder its widespread use. It is unknown whether a combined ilioinguinal blockade and local infiltration anaesthesia improves intra-operative analgesia. METHODS: We performed a double-blind randomized study in 160 patients undergoing inguinal hernia mesh repair under local infiltration anaesthesia with or without additional ilioinguinal blockade. Intra-operative pain and pain at 24 and 48 h postoperatively and analgesic requirements (acetaminophen, ibuprofen, and tramadol) were assessed. RESULTS: Median intra-operative pain scores were reduced (P=0.02) from 13 to 9 with additional ilioinguinal blockade, with no differences in requirement for sedation. There were significantly (P<0.05) more patients with intra-operative visual analogue pain scale >/=30 in the placebo group vs the ilioinguinal blockade group. Postoperative pain scores and analgesic requirements were similar. CONCLUSION: Combined ilioinguinal blockade and local infiltration anaesthesia is recommended for groin hernia repair to reduce intra-operative pain.

Adult↗
PubMed2005

Persistence of animal and human glycopeptide-resistant enterococci on two Norwegian poultry farms formerly exposed to avoparcin is associated with a widespread plasmid-mediated vanA element within a polyclonal enterococcus faecium population.

The evolutionary processes responsible for the long-term persistence of glycopeptide-resistant Enterococcus faecium (GREF) in nonselective environments were addressed by genetic analyses of E. faecium populations in animals and humans on two Norwegian poultry farms that were previously exposed to avoparcin. A total of 222 fecal GREF (n = 136) and glycopeptide-susceptible (n = 86) E. faecium (GSEF) isolates were obtained from farmers and poultry on three separate occasions in 1998 and 1999. Pulsed-field gel electrophoresis (PFGE) and plasmid DNA analyses discerned 22 GREF and 32 GSEF PFGE types within shifting polyclonal animal and human E. faecium populations and indicated the presence of transferable plasmid-mediated vanA resistance, respectively. Examples of dominant, persistent GREF PFGE types supported the notion that environmentally well-adapted GREF types may counteract the reversal of resistance. PFGE analyses, sequencing of the purK housekeeping gene, and partial typing of vanA-containing Tn1546 suggested a common animal and human reservoir of glycopeptide resistance. Inverse PCR amplification and sequence analyses targeting the right end of the Tn1546-plasmid junction fragment strongly indicated the presence of a common single Tn1546-plasmid-mediated element in 20 of 22 GREF PFGE types. This observation was further strengthened by vanY-vanZ hybridization analyses of plasmid DNAs as well as the finding of a physical linkage between Tn1546 and a putative postsegregation killing system for seven GREF PFGE types. In conclusion, our observations suggest that the molecular unit of persistence of glycopeptide resistance is a common mobile plasmid-mediated vanA-containing element within a polyclonal GREF population that changes over time. In addition, we propose that "plasmid addiction systems" may contribute to the persistence of GREF in nonselective environments.

Animal Husbandry↗
PubMed2005

A comparison of four serologic assays in screening for Brucella exposure in Hawaiian monk seals.

A survey for Brucella spp. antibodies was undertaken on 164 serum samples from 144 Hawaiian monk seals (Monachus schauinslandi) from the northwestern Hawaiian Islands collected between 1995 and 2002. The buffered antigen plate agglutination test (BPAT), the indirect enzyme immunoassay (I-ELISA), the competitive enzyme immunoassay (C-ELISA), and the fluorescence polarization assay (FPA) were compared with regard to their ability in detecting antibodies to Brucella spp. in the serum samples. Overall, antibodies were detected in 28 (17.1%) animals, using the BPAT test, 25 (15.2%) by the C-ELISA, and 19 (11.6%) in the I-ELISA and the FPA test, using thresholds established for cattle. No evidence of gross pathology consistent with clinical brucellosis was noted in any of the seropositive animals tested. Although further work would be necessary to validate these tests for use with monk seals it appears that both the C-ELISA and the FPA tests would be appropriate as diagnostic screening tests for detection of antibodies to Brucella spp. in this species.

Agglutination Tests↗
PubMed2005

Towards single screening tests for brucellosis.

This paper describes an indirect enzyme-linked immunosorbent assay (I-ELISA) and a fluorescence polarisation assay (FPA), each capable of detecting antibody in several species of hosts to smooth and rough members of the genus Brucella. The I-ELISA uses a mixture of smooth lipopolysaccharide (SLPS) and rough lipopolysaccharide (RLPS) as the antigen, and a recombinant protein A/G conjugated with horseradish peroxidase as the detection reagent. When using individually determined cutoff values, the SLPS/RLPS combined-antigen I-ELISA detected antibody in slightly more animals exposed to SLPS or to RLPS than did I-ELISA procedures using each individual antigen separately. Similarly, the assay using combined antigens detected antibody in slightly fewer animals not exposed to Brucella sp. When a universal cutoff of 10% positivity was used (relative to strongly positive control sera of each species), the overall performance index (percentage sensitivity plus percentage specificity) value decreased by 1.0 (from 199.4 to 198.4). In the FPA, it was not possible to use a universal cutoff without significant loss of performance. The overall sensitivity value for the FPA using the combined FPA antigen was 1.0% lower than using the O-polysaccharide (OPS) from SLPS and 9.1% higher than using the core antigen (CORE) from RLPS. When the combined antigen was used, the FPA specificity was slightly higher (1.2%) than from only the OPS, and considerably higher (12.6%) than the CORE. Overall, both the I-ELISA and the FPA with combined antigens were suitable as screening tests for all species of Brucella in the animal species tested.

Animal Diseases↗
PubMed2004

Gene expression profiles during beta cell maturation and after IL-1beta exposure reveal important roles of Pdx-1 and Nkx6.1 for IL-1beta sensitivity.

AIM/HYPOTHESIS: Maturation of the beta cells in the islets of Langerhans is dependent upon sequential activation of different transcription factors such as Pdx-1 and Nkx6.1. This maturation is associated with an acquired sensitivity to cytokines and may eventually lead to type 1 diabetes. The aims of this study were to characterise changes in mRNA expression during beta cell maturation as well as after interleukin-1beta (IL-1beta) exposure. METHODS: Transcriptome analyses were performed on two phenotypes characterised as a glucagon-producing pre-beta-cell phenotype (NHI-glu), which matures to an IL-1beta-sensitive insulin-producing beta cell phenotype (NHI-ins). Beta cell lines over-expressing Pdx-1 or Nkx6.1, respectively, were used for functional characterisation of acquired IL-1beta sensitivity. RESULTS: During beta cell maturation 98 fully annotated mRNAs changed expression levels. Of these, 50 were also changed after 24 h of IL-1beta exposure. In addition, 522 and 197 fully annotated mRNAs, not affected by maturation, also changed expression levels following IL-1beta exposure of the beta cell and the pre-beta-cell phenotype, respectively. Beta cell maturation was associated with an increased expression of Nkx6.1, whereas both Pdx-1 and Nkx6.1 expression were decreased following IL-1beta exposure. Over-expression of Nkx6.1 or Pdx-1 in cell lines resulted in a significantly increased sensitivity to IL-1beta. CONCLUSIONS/INTERPRETATION: These results suggest that the final beta cell maturation accompanied by increased IL-1beta sensitivity is, in part, dependent upon the expression of genes regulated by Pdx-1 and Nkx6.1. Future classification of the genes regulated by these transcription factors and changed during beta cell maturation should elucidate their role in the acquired sensitivity to IL-1beta and may be helpful in identifying new targets for intervention/prevention strategies.

Animals↗
PubMed2004

Enzyme immunoassay for the diagnosis of brucellosis: chimeric Protein A-Protein G as a common enzyme labeled detection reagent for sera for different animal species.

A recombinant protein combining the immunoglobulin binding sites of Proteins A and G, conjugated with horseradish peroxidase was used as a universal detection reagent for the assessment of antibodies against Brucella spp. The reagent was applied in an indirect enzyme immunoassay for detection of antibodies to smooth lipopolysaccharide antigen in sera from Brucella spp. exposed and non-exposed cattle, sheep, goats and pigs and to antibodies to rough lipopolysaccharide in sheep, dogs and cattle. The results were similar to those obtained when murine monoclonal antibody-enzyme conjugates were used. An added advantage was that a universal cut-off for all tests using the proteins A and G detection reagent could be established, simplifying diagnostic interpretation of the data.

Animals↗
PubMed2004

Crystal structure and spectroscopic properties of CsVO2SO4.

Dark crystals of the V(V) compound CsVO(2)SO(4), suitable for X-ray investigations have been obtained from the catalytically important Cs(2)S(2)O(7)-V(2)O(5) system. By cooling of the mixture with the composition X(V)2(O)5 = 0.5, some crystals were obtained in the otherwise glassy sample. The compound crystallizes in the orthorhombic space group Pbca with a = 6.6688(13) A, b = 10.048(2) A, and c = 17.680(4) A at 20 degrees C and Z = 8. It contains a coordination sphere with a short V-O bond of 1.595(2) A and trans to this the closest VO distance at 3.4 A and four equatorial V-O bonds in the range 1.725(1)-1.984(2) A. The deformation of the VO(6) octahedron is thus much more pronounced compared to other known oxo sulfato V(V) compounds, and the coordination polyhedron of V(V) should be regarded as a tetragonal pyramid with the vanadium atom in the center. Each VO(2)(+) group is coordinated to the neighboring groups by oxygen and sulfate double bridges in a zigzag structure where two sulfate oxygens virtually remain uncoordinated-one is found at the very long nonbonding V-O distance from the neighboring chain. This is the first time that we find pentacoordination of vanadium in the 12 different V(III), V(IV), and V(V) compounds examined so far. The FTIR and Raman spectra of the compound are in agreement with the simple formula unit of the investigated compound.

Journal Article↗
PubMed2004

Antibody response and antigen-specific gamma-interferon profiles of vaccinated and unvaccinated pregnant sheep experimentally infected with Brucella melitensis.

It is well known that the immune response in sheep against Brucella melitensis is subject to individual variation, depending on diverse factors. It bears asking whether these factors (e.g. clinical disease, active infection, state of previous immunity), when affecting a group, can cause variation in the performance of different diagnostic tests. To clarify some of the circumstances in which this immune response can vary, we examine the immune-response profile of sheep protected against the clinical disease by prior vaccination with strain Rev. 1 in comparison with the profile of unprotected females showing the classical brucellosis symptoms. An experimental infection was provoked at midpregnancy under controlled conditions of both non-vaccinated (n=7) and previously Rev.1-vaccinated ewes (n=5). Their immune response was monitored from 7 to 9 weeks before abortion or normal birth to 30 weeks afterwards. Antibody response was assessed by classical tests (Rose Bengal test, complement fixation test (CFT)) in comparison with other diagnostic tests (indirect ELISA (iELISA), competitive ELISA (cELISA), fluorescence polarization assay (FPA), immunocapture test (ICT)). In addition, the cell-mediated immune response was indirectly evaluated by the in vitro antigen-specific release of gamma-interferon. The antibody levels and antigen-specific gamma-IFN profile of the non-vaccinated ewes having the disease and excreting the pathogen was notably high and differed significantly (P<0.05 or P<0.01) from those of vaccinated ewes that neither contracted brucellosis nor excreted the pathogen. In general, all the tests detect the infection in the non-vaccinated ewes with substantial effectiveness. It can be concluded that the high levels of circulating antibodies and of antigen-specific gamma-IFN are related to active Brucella infection. Similarly, the state of protection against the disease, but not necessarily against infection, due to a previous immunization with the Rev. 1 vaccination, appears to be responsible for a low level of detectable immune response. Nevertheless, the design of the study limits conclusions to pregnant ewes and cannot be extrapolated to non-pregnant ewes or rams. Likewise, the study provides no information on animals which are carriers of B. melitensis.

Animals↗
PubMed2004

Serological relationship between cattle exposed to Brucella abortus, Yersinia enterocolitica O:9 and Escherichia coli O157:H7.

Sera from cattle naturally infected with Brucella abortus (n = 160), vaccinated with B. abortus S19 (n = 88) or immunized with Yersinia enterocolitica O:9 (n = 25) or Escherichia coli O157:H7 (n = 80) were collected. The sera were compared for antibody content to the same bacteria by indirect enzyme immunoassay (IELISA), fluorescence polarization assay (FPA) and competitive enzyme immunoassay (CELISA). Cattle sera (n = 523) collected randomly from across Canada were tested in the same tests. Sera from the B. abortus infected group reacted positively in the brucellosis IELISA (IELISA(Br)), CELISA and FPA (FPA(Br)) and the Y. enterocolitica IELISA (IELISA(Ye)) while the Y. enterocolitica FPA (FPA(Ye)) detected antibody in 93.8% and the E. coli IELISA (IELISA(Ec)) 86.9% and the E. coli FPA (FPA(Ec)) 48.1%. About 70% of the sera from B. abortus S19 vaccinated animals reacted in the three IELISAs, 45% in the CELISA, and 37.7% in the FPA(Ec), 21.6% in the FPA(Br) and 5.7% in the FPA(Ye). Sera from E. coli O:157 exposed cattle reacted mainly in the IELISA(Ec) and FPA(Ec) although surprisingly 87.5% reacted in the IELISA(Ye) and only 3.8% in the IELISA(Br). No reactions were observed with these sera in the FPA(Br) and FPA(Ye) but one serum gave a low positive reaction in the CELISA. All sera from Y. enterocolitica O:9 exposed cattle reacted in the IELISA(Br) and IELISA(Ye) and 80% in the IELISA(Ec). In the CELISA, 44% gave a positive reaction and 64% were positive in the FPA(Br), 28% in the FPA(Ye) and 12% in the FPA(Ec). Of the 523 Canadian sera, about 50% reacted in the E. coli tests with only minor reactions in the Y. enterocolitica O:9 and B. abortus assays. From the data, the cross reaction between E. coli O157:H7, Y. enterocilitica O:9 and B. abortus is dependent on the test used. Thus, extensive cross reaction was observed with the IELISA with much less reactivity in the FPA and the CELISA.

Animals↗
PubMed2004

Rough lipopolysaccharide of Brucella abortus RB51 as a common antigen for serological detection of B. ovis, B. canis, and B. abortus RB51 exposure using indirect enzyme immunoassay and fluorescence polarization assay.

Rough lipopolysaccharide (RLPS) antigens were prepared from cultures of Brucella abortus RB51, B. ovis, and B. canis. The preparations were standardized by weight and tested with sera from cattle immunized with B. abortus RB51, sheep infected with B. ovis, and dogs infected with B. canis. Populations of unexposed animals of each species were also tested. The tests used were the indirect enzyme immunoassay (IELISA) using RLPS and the fluorescence polarization assay (FPA) using RLPS core fractions, labeled with fluorescein isothiocyanate. The IELISA using B. abortus RB51 RLPS antigen resulted in sensitivity and specificity values of 94.8% and 97.3%, respectively, when testing bovine sera, 98.5% and 97.8% when testing ovine sera, and 95.8% and 100% when testing dog sera. The IELISA using B. ovis RLPS antigen gave sensitivity and specificity values of 80.5% and 91.7%, respectively with bovine sera, 98.9% and 93.8% with sheep sera, and 70.8% and 79.8% with dog sera. The IELISA using B. canis RLPS antigen resulted in sensitivity and specificity values of 97.0% and 97.4%, respectively, with bovine sera, 96.2% and 96.3% with sheep sera, and 95.8% and 98.8% with dog sera. Labeling RLPS core from B. ovis and B. canis with fluorescein was not successful. B. abortus RB51 core labeled with fluorescein resulted in sensitivity and specificity values of 93.5% and 99.8%, respectively, with bovine sera and 78.1% and 99.0% with sheep sera. It was not possible to test the dog sera in the FPA.

Animals↗
PubMed2004

Melanoma and nonmelanoma skin cancer in patients with multiple tumours--evidence for new syndromes in a population-based study.

BACKGROUND: The hypotheses that Swedish patients with four or more primary tumours [including at least one cutaneous malignant melanoma (CMM)] harbour an increased number of CDKN2A (formerly p16) germline mutations, and that this group of patients show a predisposition to other tumours, e.g. nonmelanoma skin cancer (NMSC), were studied descriptively. So far the mutation 113insArg explains all CDKN2A-associated CMM in ethnic Swedes. OBJECTIVES: All patients with four or more primary tumours, of which at least one was a CMM, from the Southern Swedish Regional Tumour Registry, between 1958 and 1999, were included in this study. METHODS: Forty-four patients were found and subdivided into three groups according to having multiple CMM (group A) or single CMM +/- NMSC (groups B and C). Screening for the presence of the Swedish founder mutation 113insArg in blood or in tissue blocks was performed. RESULTS: Patients in group A were younger at the time of the first CMM diagnosis than patients in group B and group C. The 113insArg mutation was found in four of 44 patients (9%), three with multiple CMM. In group C (n = 14) no founder mutation was evident, while in group B (n = 15) one mutation carrier was found. Nonmutation carriers with multiple CMM (group A) also had a predilection for meningiomas and neurinomas (four patients) or multiple NMSC (three patients). In group B CMM were especially associated with adenocarcinomas but in group C CMM were associated with multiple NMSC. CONCLUSION: The association between meningiomas and neurinomas (no acoustic neurinoma was seen) might indicate a new syndrome. Patients in groups B and C may harbour unknown genetic defects, which could interact with different environmental risk factors.

Adenocarcinoma↗