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Biomedical subjects

D Kemp

Publications and source records attributed to D Kemp.

At least 19 recordsLinked to original sources

World Association for the Advancement of Veterinary Parasitology (W.A.A.V.P.) guidelines for evaluating the efficacy of acaricides against ticks (Ixodidae) on ruminants.

These guidelines have been prepared to assist in the planning, conduct and interpretation of studies for the assessment of the efficacy of acaricides (excluding vaccines and other bio-control agents) against single and multi-host ticks (Ixodidae) on ruminants. Information is provided on the selection of animals, dose determination, dose confirmation and field studies, record keeping and result interpretation. The use of pen facilities is advocated for dose determination and confirmation studies for defining therapeutic and persistent efficacy. A minimum of two studies per tick species for which claims are sought is recommended for each dose determination and dose confirmation investigation. If dose confirmation studies demonstrate greater than 95% efficacy the sponsor may proceed to field studies, where a minimum of two studies per geographical location is preferred to confirm the therapeutic and persistent efficacy under field conditions. If dose confirmation studies demonstrate less than 95% efficacy then longer-term field studies can be conducted over two tick seasons with a minimum of two studies per geographical location. These studies can incorporate other control methods such as tick vaccines, to demonstrate stable long-term tick management. Specific advice is also given on conducting studies with paralysis ticks. These guidelines are also intended to assist investigators on how to conduct specific experiments, to provide specific information for registration authorities involved in the decision-making process, to assist in the approval and registration of new acaricides, and to facilitate the worldwide adoption of standard procedures.

Animals↗

Reduced oviposition of Boophilus microplus feeding on sheep vaccinated with vitellin.

The most abundant protein present in Boophilus microplus eggs, vitellin, was isolated and purified as a non-covalent complex of six glyco-polypeptides of Mr 44-107kDa. The protein complex bound haem. Immuno-blots demonstrated that antibodies raised to vitellin recognised a 200kDa polypeptide in the haemolymph of adult female ticks. This is consistent with the general proposal that in arthropods vitellin is derived by proteolytic processing from a large precursor protein, vitellogenin. In parallel with this study, an 80kDa glycoprotein (GP80) was independently purified from larvae of B. microplus using efficacy in vaccination trials as an assay. Antibodies to GP80 also recognised a 200kDa protein in the haemolymph of ticks and a major 87kDa polypeptide present in the vitellin complex. Conversely, antibodies to purified vitellin recognised GP80. The amino-terminal amino acid sequences of the 87kDa vitellin polypeptide and GP80 were identical for at least the first 11 residues and internal peptide sequences from both polypeptides were co-located in a single but incomplete deduced amino sequence of B. microplus vitellogenin. Thus, GP80 is a processed product from vitellogenin and highly related to but not completely identical with the 87kDa vitellin polypeptide. Vaccination trials in the model host sheep were performed with purified vitellin and GP80. Sheep vaccinated with either purified vitellin or GP80 returned significantly reduced numbers of engorged female ticks with decreased weights and reduced oviposition. In contrast, sheep vaccinated with recombinant hexahis-GP80, which was incorrectly folded and not glycosylated showed no significant effects on ticks. It was concluded that vitellin and GP80 could induce immune responses that partially protect sheep from the tick, B. microplus. However, critical protective epitopes are associated with the folding of the protein and/or the oligosaccharides attached to it.

Animals↗

The mitochondrial 12S gene is a suitable marker of populations of Sarcoptes scabiei from wombats, dogs and humans in Australia.

We sequenced part of the mitochondrial 12S ribosomal RNA gene of 23 specimens of Sarcoptes scabiei from eight wombats, one dog and three humans. Twelve of the 326 nucleotide positions varied among these mites and there were nine haplotypes (sequences) that differed by 1-8 nucleotides. Phylogenetic analyses indicated that these mites were from two lineages: (1) mites from wombats from Victoria, Australia, and mites from the humans and dog from the Northern Territory, Australia (haplotypes 1-4, 9); and (2) mites from the humans and dog from the Northern Territory (haplotypes 5-8). Mites from the three different hosts (wombats, a dog and humans) had not diverged phylogenetically; rather, these mites had similar 12S sequences. Thus, we conclude that these mites from wombats, humans and a dog are closely related, and that they diverged from a common ancestor relatively recently. This conclusion is consistent with the argument that people and/or their dogs introduced to Australia the S. scabiei mites that infect wombats in Australia . So, S. scabiei, which has been blamed for the extinction of populations of wombats in Australia, may be a parasitic mite that was introduced to Australia with people and/or their dogs. These data show that the mitochondrial 12S rRNA gene may be a suitable population marker of S. scabiei from wombats, dogs and humans in Australia.

Animals↗

NK cells are essential for effective BCG immunotherapy.

Adjuvant intravesical bacillus Calmette-Guérin (BCG) therapy is a well-established and successful adjuvant immunotherapy in the treatment of superficial bladder cancer. Although the function of natural killer (NK) cells in other immunotherapeutic regimens (e.g., lymphokine-activated killer [LAK] cell or interleukin-2 [IL-2] therapy) has been established, the contribution of NK cells to effective BCG immunotherapy is not clear. We used a human in vitro system to analyze the role of NK cells in BCG-induced cellular cytotoxicity. After stimulation of mononuclear cells with BCG for 7 days, these BCG-activated killer (BAK) cells displayed substantial cytotoxicity against bladder tumor cells. Magnetic depletion experiments and fluorescence activated cell sorting revealed that NK cells were the major effector cell population. To address NK cell function in vivo, we studied a syngeneic orthotopic murine bladder cancer model and compared BCG immunotherapy in C57BL/6 wild-type mice, NK-deficient beige mice and mice treated with anti-NK1.1 monoclonal antibody. Four weekly instillations of viable BCG significantly prolonged survival in wild-type mice compared with control mice treated with solvent alone. In contrast, BCG therapy was completely ineffective in NK-deficient beige mice and in mice treated with anti-NK1.1 monoclonal antibody. These findings suggest a key role for NK cells during BCG immunotherapy.

Animals↗

Peritrophins of adult dipteran ectoparasites and their evaluation as vaccine antigens.

Several peritrophins of larvae of Lucilia cuprina (sheep blowfly) have demonstrated potential as vaccine antigens, and some have been characterised and cloned. These proteins are tightly associated with the peritrophic matrix, a chitinous tube or sac lining the lumen of the gut of most insects. The peritrophins require strong denaturants for their removal from peritrophic matrix. We now report the preliminary characterisation of peritrophins of the adult stage of L. cuprina and Haematobia irritans exigua (buffalo fly). Similar SDS-PAGE profiles were obtained for proteins extracted in SDS or urea from isolated adult peritrophic matrices of both species. Radioiodination of urea-extracted peritrophins improved sensitivity, indicating numerous proteins of 15-75 kDa. Direct radioiodination of L. cuprina peritrophic matrix preferentially labelled high molecular weight complexes and proteins of 80-90 kDa. Two-dimensional gel analyses of a urea extract of adult L. cuprina peritrophic matrix revealed that most proteins were moderately acidic. Antibodies produced against SDS-extracted peritrophins, or against sonicated peritrophic matrices of these two flies were crossreactive. The sera also appeared to recognise SDS-extracted components of Triton X-100 treated and washed adult peritrophic matrix of the mosquito, Aedes vigilax (Skause). This profile altered as the peritrophic matrix matured. In concordance with extracts from the adult L. cuprina and H.i. exigua peritrophic matrices, proteins in the 50-75 kDa region were immunodominant. The vaccine potential of the peritrophins of these Diptera were examined following vaccination of cattle and rabbits with adult H.i. exigua or L. cuprina peritrophins. When the adult life stages of H.i. exigua or two mosquitoes, A. vigilax and A. aegypti (Linnaeus), were fed on the sera or blood of vaccinated hosts, there were no detrimental effects to any life cycle stages of these Diptera.

Aedes↗

Immunological aspects of nutritional diabetes prevention in NOD mice: a pilot study for the cow's milk-based IDDM prevention trial.

Human epidemiological studies delineated early exposure to intact dietary protein (e.g., most infant formulas) as an environmental risk factor for the development of IDDM. The Trial to Reduce IDDM in the Genetically at Risk (TRIGR), an international IDDM prevention trial, has been designed to determine if avoidance of intact dairy protein in high-risk infants < or =6 months of age can reduce the subsequent diabetes incidence. We here studied the casein hydrolysate-based trial diet (Nutramigen) in NOD mice. When given either continuously or for 10 weeks after weaning, the test diet was highly effective in preventing autoimmune diabetes (32-week incidence: 4.6 vs. 58.8%) and in preserving pancreatic insulin levels, with little effect on islet inflammation. Spleen cells from protected NOD mice failed to adoptively transfer diabetes into irradiated syngeneic recipients. When co-transferred with splenocytes from diabetic donors, cells from diet-protected mice inhibited adoptive diabetes transfer (incidence 50 vs. 94%, P < 0.001). T-cell reactivity to the islet cell autoantigens ICA69 (islet cell antigen 69) and GAD65 developed only in diabetic recipients of spleen cell grafts, indicating that diabetes protection extends to more than one autoantigen. In protected mice, ICA69 T-cell reactivity was not detectable spontaneously nor after priming with this autoantigen; however, priming with the cross-reactive non-self-antigen bovine serum albumin recruited T-cells responsive to ICA69. Thus, diabetes prevention with the clinical trial diet is effective in NOD mice, where it affects some T-cell repertoires and allows development of regulatory cells that interfere with destructive autoimmunity.

Adoptive Transfer↗

Cloning and characterisation of angiotensin-converting enzyme from the dipteran species, Haematobia irritans exigua, and its expression in the maturing male reproductive system.

The angiotensin-converting enzymes (ACE) are involved in the regulation of the specific maturation or degradation of a number of mammalian bioactive peptides. A carboxydipeptidase similar to mammalian ACE has now been identified in the adult stage of the haematophagous fly, Haematobia irritans exigua (buffalo fly), a close relative of the horn fly of North America. The enzyme was purified by lectin-affinity chromatography and ion-exchange chromatography and migrated as a doublet of 70 kDa upon reducing SDS/PAGE. Unlike mammalian ACE, the fly carboxydipeptidase (HieACE) is not membrane bound. The amino acid sequence of an internal peptide from HieACE and a conserved amino acid region present in all mammalian ACE were used to design degenerate oligonucleotide primers suitable for PCR. A DNA fragment amplified from adult buffalo fly cDNA was used to identify a cDNA clone that encoded the enzyme. The cDNA sequence encodes a carboxydipeptidase with 41-42% amino acid identity to the mammalian testicular ACE. The active-site regions of mammalian ACE are conserved in the deduced amino acid sequence of HieACE. Enzymatically, HieACE is very similar to its mammalian counterparts, with comparable Km and V(max) values for the synthetic substrate, benzoylglycylglycylglycine, and similar patterns of inhibition by EDTA, ACE inhibitor peptide and captopril. HieACE also specifically activates angiotensin I to angiotensin II and degrades other mammalian ACE substrates such as bradykinin, substance P and cholecystokinin-8. In the adult fly, HieACE is expressed in the compound ganglion and in the posterior region of the midgut. Similar to the mammalian system, expression of this enzyme is induced in the maturing male reproductive system, which suggests conservation of ACE function in these species.

Amino Acid Sequence↗

RAPD analysis of isolates of Burkholderia pseudomallei from patients with recurrent melioidosis.

Twenty-seven isolates of Burholderia pseudomallei (formerly Pseudomonas pseudomallei) from ten patients with recurrent melioidosis were analysed by RAPD. In two cases RAPD patterns in recurrent isolates differed from the original isolates; one was considered a likely reinfection while the other may represent relapse from one of two strains initially infecting the patient. In two cases where a change in antibiotic resistance had occurred between original and relapse isolates, slight changes in RAPD patterns were found with one of the four primers used. In the other six cases the relapse was clearly due to the original strain re-emerging unchanged, with identical RAPD patterns with all four primers.

Adult↗

Subdivision of Burkholderia pseudomallei ribotypes into multiple types by random amplified polymorphic DNA analysis provides new insights into epidemiology.

Ribotyping has previously been used for epidemiological studies of Burkholderia pseudomallei (previously Pseudomonas pseudomallei). We show here that random amplified polymorphic DNA (RAPD) analysis allows subdivision of strains of the same ribotype. With five different primers, no two epidemiologically unrelated isolates of any single ribotype in this study of 102 isolates from humans, goats, cats, and soil had identical RAPD patterns. Conversely, RAPD analysis showed clonality for isolates from each of two animal outbreaks of melioidosis and from a nontropical focus of animal and human melioidosis spanning 25 years. Some soil isolates were identical to epidemiologically related animal and human isolates as determined by RAPD typing. There was no evidence that the clinical outcome of melioidosis was related to RAPD patterns.

Animals↗

Streptokinase alleles and disease association in group A streptococci.

Allele-specific oligonucleotides were used for PCR-based typing of the streptokinase locus of group A streptococcal strains, including well characterized type strains, isolates from patients with acute poststreptococcal glomerulonephritis and strains from Aboriginal communities in the Northern Territory of Australia. The streptokinase SKN allele, previously thought to be associated with glomerulonephritis, was no more frequent in nephritogenic than in non-nephritogenic streptococcal strains in this collection.

Alleles↗

Sequence- and strand-specific cleavage in oligodeoxyribonucleotides and DNA containing 3'-thiothymidine.

Oligonucleotides containing a 3'-thiothymidine residue (T3's) at the cleavage site for the EcoRV restriction endonuclease (between the central T and A residues of the sequence GATATC) have been prepared on an automated DNA synthesizer using 5'-O-monomethoxytritylthymidine 3'-S-(2-cyanoethyl N,N-diisopropylphosphorothioamidite). The self-complementary sequence GACGAT3'sATCGTC was completely resistant to cleavage by EcoRV, while the heteroduplex composed of 5'-TCTGAT3'sATCCTC and 5'-GAGGATATCAGA (duplex 4) was cleaved only in the unmodified strand (5'-GAGGATATCAGA). In contrast, strands containing a 3'-S-phosphorothiolate linkage could be chemically cleaved specifically at this site with Ag+. A T3's residue has also been incorporated in the (-) strand of double-stranded closed circular (RF IV) M13mp18 DNA at the cleavage site of a unique EcoRV recognition sequence by using 5'-pCGAGCTCGAT3'sATCGTAAT as a primer for polymerization on the template (+) strand of M13mp18 DNA. On treatment of this substrate with EcoRV, only one strand was cleaved to produce the RF II or nicked DNA. Taken in conjunction with the cleavage studies on the oligonucleotides, this result demonstrates that the 3'-S-phosphorothiolate linkage is resistant to scission by EcoRV. Additionally, the phosphorothiolate-containing strand of the M13mp18 DNA could be cleaved specifically at the point of modification using iodine in aqueous pyridine. The combination of enzymatic and chemical techniques provides, for the first time, a demonstrated method for the sequence-specific cleavage of either the (+) or (-) strand.

Base Sequence↗

Postoperative urinary retention: Part I--Overview and implications for the postanesthesia care unit nurse.

Postoperative urinary retention and resulting bladder distention are common complications following surgical procedures. The PACU nurse needs a thorough understanding of bladder distention, causes, and how to assess for signs and symptoms in the PACU patient. Unrelieved bladder distention can lead to discomfort and problems with micturation or chronic urinary retention. The anatomy of the bladder, causes and factors related to bladder distention, and methods with which the PACU nurse can assess and evaluate for bladder distention are presented.

Humans↗

Postoperative urinary retention: Part II--A retrospective study.

Postoperative urinary retention, a potential postoperative complication, can lead to discomfort and contribute to atony of bladder wall, need for catheterization and, sometimes, prolonged hospital stays. A retrospective study was conducted to analyze variables related to postoperative urinary retention. Data was collected by a chart review of 100 postoperative cases in a large Veterans Administration Medical Center. All patients were male, had a mean age of 61.1 years, and had received spinal or general anesthesia. Patients with known urologic problems, paraplegic patients, quadruplegic patients, patients undergoing procedures lasting less than 3 hours, and patients with indwelling Foley catheters were excluded. The medical review form was used to collect data from the medical records. Sixty-one percent of the patients required straight catheterization for their first postoperative urinary output. Sixty-five percent of the spinal anesthetic patients and 57% of the general anesthetic patients required straight catheterization. Orthopedic patients were the highest percentage of patients requiring catheterization. The patients who received large volumes of intravenous (IV) fluid in the operating room were more likely to be catheterized in the PACU. Determining whether physical assessment or IV volume indicated need for catheterization of these patients was difficult because documentation on the PACU record was usually incomplete. Complete documentation of assessment and intervention on the PACU record occurred 10% of the time. A need for full assessment on all postsurgical patients in the PACU and improved documentation is indicated.

Adult↗

Epithelial action potentials in embryos of the Australian lungfish.

The epithelial cells of the skin of embryonic Australian lungfish (Neoceratodus forsteri) between stages 30 and 39, are mechanosensitive and excitable, showing overshooting action potentials 400-800 ms long with a rapid rise followed by a slower repolarization (usually with a shoulder on the repolarizing phase), which propagate at around 10 mm s-1. This skin impulse system is very similar to that found in embryonic and larval Amphibia; the significance of this similarity is discussed.

Action Potentials↗

An advanced cochlear echo technique suitable for infant screening.

An advanced cochlear echo test technique has been specially developed to tackle some specific problems associated with measurements from infants. The test procedure and data processing combination has greatly improved noise immunity and has given very reliable results on poorly co-operating children. Technical details are given and the results of a study of 105 ears from 55 children are presented. The potential of the advanced technique as a screening test is discussed.

Adolescent↗