Real-time and multiplex real-time polymerase chain reactions for the detection of Bartonella henselae within cat flea, Ctenocephalides felis, samples.

Bartonella henselae (Rhizobiales: Bartonellacae), the agent of cat-scratch disease, is an emerging bacterial pathogen which can be transmitted via infective faecal material of Ctenocephalides felis Bouché (Siphonaptera: Pulicidae). Worldwide, B. henselae has been identified in 1-53% of felines and 2.9-17.4% of fleas. Although culture is the routine method for detection, the procedure is time-consuming and is rarely used for isolation directly from flea vectors. The current study reports the development of a quantitative real-time polymerase chain reaction (qPCR) to detect and quantify B. henselae organisms from vector samples. The qPCR is specific and detects as few as 2.5 genome copies. To enable direct quantification of Bartonella organisms in different vector samples, we developed a qPCR to detect C. felis DNA that also acts as an extraction control. Combining both PCRs into a multiplex format validates B. henselae results when sampling flea populations, although there is a reduction in sensitivity. This reduction might be counteracted by a different combination of probe fluorophores.

The development of a qPCR assay to detect tick (Ixodida) DNA and its implementation for the study of tick-borne pathogen transmission.

Polymerase chain reaction (PCR) analysis is regularly used to detect pathogens within arthropod vectors, but has also been applied to investigate vector DNA. This study details a novel highly sensitive quantitative PCR (qPCR) which detects and quantifies DNA from Ixodes ricinus, the European vector of Anaplasma phagocytophilum. By pairing this with a qPCR to detect A. phagocytophilum, valid comparisons of pathogen load can be made between different sized tick-tissue samples. These qPCRs were validated in I. ricinus that were fed A. phagocytophilum-infected blood using an artificial membrane feeder. Pathogens were detected in the tick haemolymph within 36h, indicating that successful infection had taken place. This study illustrates the application of vector-targeted qPCRs to confirm and validate pathogen load in samples as part of investigations of vector-pathogen interactions.

Photocatalytic photosystem I/PEDOT composite films prepared by vapor-phase polymerization.

Photosystem I (PSI) achieves photo-induced charge separation with outstanding internal quantum efficiency and has been used to improve the performance of various photoelectrochemical systems. Herein, we describe a fast and versatile technique to assemble composite films containing PSI and a chosen intrinsically conductive polymer (ICP). A mixture of PSI and a Friedel-Crafts catalyst (FeCl3) is drop cast atop a substrate of choice. Contact with ICP monomer vapor at low temperature stimulates polymer growth throughout PSI films in minutes. We assess the effects of PSI loading on the rapid vapor-phase growth of poly(3,4-ethylenedioxythiophene) (PEDOT) within and above PSI multilayer films, and characterize the resulting film's thickness, electrochemical capacitance, and photocatalytic response. Composite films generate cathodic photocurrent when in contact with an aqueous redox electrolyte, confirming retention of the photocatalytic activity of the polymer-entrapped PSI multilayer assembly.

Longitudinal analysis of heritability for dental caries traits.

The role of genetic and environmental factors on dental caries progression in young children was determined. A detailed caries assessment was performed in 2 examinations on 314 pairs of twins initially 1.5 to 8 years old. Surface-based caries prevalence rates (SBCPR) and lesion severity (LSI) were computed. Heritability estimates were calculated by SOLAR software. Analyses were performed on all ages combined and by age group (1.5-< 4; 4-6; > 6). Overall heritability estimates (H) of net increments SBCPRs were H = 30.0 (p < 0.0001), and were greatest for the youngest (H = 30.0) and oldest groups (H = 46.3). Overall LSI heritability estimates [H = 36.1 (p < 0.0001)] were also greatest for the youngest (H = 51.2) and oldest groups (H = 50.6). Similar findings were found for net increments of occlusal surfaces and deep dentinal lesions SBCPRs (H = 46.4-56.2). These findings are consistent with a significant genetic contribution to dental caries progression and severity in both emerging primary and permanent dentitions.

Asymmetric brain edema after cardiac transplantation: cerebroautoregulatory failure and relative hyperperfusion.

Asymmetric brain edema is a rare neurologic complication after cardiovascular surgery. We describe the clinical and imaging features of an asymmetric brain edema syndrome in a 52-year-old man following cardiac transplantation who presented with facial myoclonus and left hemiparesis in the postoperative period. To our knowledge, this is the first case report of asymmetric brain edema syndrome after cardiac transplant and the second following cardiac surgery. Arterial bypass cannula malposition in the ascending aorta or brachiocephalic artery with subsequent cerebral hypoperfusion and subsequent hyperperfusion appears to be the most likely physiologic cause.

AGT and RH blood group polymorphisms affect blood pressure and lipids in Afro-Caribbeans.

Population blood pressure variation is most likely due to multiple genes. This is likely the reason why monogenic testing with the angiotensinogen (AGT) gene polymorphisms on chromosome 1 (1q42-43), especially M235T, has met with negative results, especially in those of African descent. The RH blood group system, also on chromosome 1 (1 p36.2-34), has likewise been associated with blood pressure variation in African-Americans and with the rise in blood pressure with age in whites. Using a random sample of the population, we investigated the combined effects of single and combined variation of the AGTN M235T and RH genotypes on blood pressure, lipids, and lipoprotein concentrations in Afro-Caribbeans aged 18-60 years from the island nation of Dominica. In monogenic analysis, AGT M235T was not associated with blood pressure. However, it was associated with HDL (MM 42+/-23, MT 44+/-12, TT 52+/-14 (P=0.002)). RH genotype was significantly associated with systolic blood pressure (P=0.006) and Apo-A (P=0.003). These effects remained after adjustment for age, gender, weight, and BMI. In the polygenetic analysis, AGT M235T and RH were significantly associated with systolic blood pressure (P=0.037; interaction effects, P=0.068). The association of the AGT M235T with blood pressure across RH blood group haplotypes was then tested. Of the five RH haplotypes available for analysis, the AGT M235T was significantly associated with blood pressure within the "D" haplotype (P=0.01). The RH blood group and gender were significantly associated with systolic blood pressure and Apo-A levels (P=0.005 and 0.012, respectively). All interactions were independent of age and weight. In conclusion, we demonstrate a significant association of AGT M235T with blood pressure and cholesterol metabolism in an Afro-Caribbean population in the "genetic context" of the RH blood group system. Further investigation of these interactions may help understand the effects of genetic factors on cardiovascular risk in African-derived and other populations.

The hospital of the future--planning for change.

This paper describes the hospital planning model developed by the North Eastern Metropolitan Region of the Health Department Victoria to forecast acute public hospital bed-day requirements in the Region. Three age-specific variables: population; separation rate; and length of stay have been used to estimate the level of demand for hospital services. The model also delineates services delivered on a same day or long stay basis. The application of the model to three local government areas demonstrates the importance of population growth and ageing on the type and level of hospital services required and the implications thereof for service delivery and the physical configuration of hospitals.

The pest status and distribution of three cryptic blue oat mite species (Penthaleus spp.) and redlegged earth mite (Halotydeus destructor) in southeastern Australia.

Earth mites are pests of crops and pastures in southeastern Australia. Recent studies show differences between earth mite species in their mode of reproduction, preferred hosts and pesticide tolerance. This paper examines the distribution and pest status of each species. The southeastern Australian distribution for each species is mapped, incorporating new data from eastern New South Wales, South Australia and Tasmania. A new population of an undescribed species previously identified from northwestern Victoria was found in northern New South Wales. CLIMEX was used to identify climatic factors limiting the distribution of P. major and P. falcatus, the most broadly distributed species. This analysis suggests tolerance to heat and desiccation limits the inland distribution of these two species. A three-year survey of agricultural outbreaks indicates that all Penthaleus species are major agricultural pests although their pest status on crop types appears to differ. All species contributed to chemical control failures. However P. falcatus, previously identified in laboratory tests as having increased tolerance to pesticides, was the most common species associated with control failures. A laboratory experiment indicated that mites are sometimes pests on crops on which they cannot persist for a generation. Results are discussed with respect to management of these agricultural pests.

Association of a genetic polymorphism in human apolipoprotein B-100 with intermediate density lipoprotein concentrations.

Immunochemical techniques have been used to identify five antigenic (Ag) sites on apolipoprotein B-100 (apoB), the major protein constituent of very low density (VLDL), intermediate density (IDL), and low density lipoproteins (LDL). Each Ag site results from allelic variation at a specific locus of the apoB gene. In the present study, we assessed whether variations in the five Ag loci were associated with concentrations of plasma lipids or lipoprotein fractions measured by analytical ultracentrifugation in a group of 44 healthy men. Pair-wise analyses of the Ag markers revealed that Ag(a1/d), in association with either Ag(x/y) or Ag(t/z), is significantly related to plasma IDL-mass concentrations. In this cohort we detected no significant associations of the Ag alleles (singly or in combination) with plasma total cholesterol, triglycerides, LDL-cholesterol, HDL-cholesterol, or mass of total VLDL or LDL. These results suggest that genetic variations in the apoB molecule may predispose to variations in concentrations of IDL that could have consequences for atherosclerotic risk.

Anti-apoprotein B monoclonal antibodies detect human low density lipoprotein polymorphism.

Immunochemical polymorphism among human low density lipoproteins (LDL) isolated from different individuals was demonstrated through reduced binding of three monoclonal antibodies to some individual LDL using a solid phase radioimmunoassay. These three antibodies are members of a larger group of monoclonal antibodies previously shown to bind specifically to apoprotein B ( Curtiss , L.K., and Edgington , T. S. (1982) J. Biol. Chem. 257, 15213-15221; Tsao , B.P., Curtiss , L. K., and Edgington , T.S. (1982) J. Biol. Chem. 257, 15222-15228). Those antibodies which distinguished human LDL polymorphism bound to the same or adjacent epitopes on LDL, for they were mutually exclusive in competitive binding experiments. Binding was unaffected by treatment with neuraminidase, with a mixture of glycosidases, or with competing glycoproteins; thus, the carbohydrate moiety of apoprotein B did not appear to influence the epitope recognized by these antibodies. When low density lipoproteins isolated from different individuals were studied, three different phenotypes were recognized; these corresponded to strong, weak, and intermediate binding of the three monoclonal antibodies. This division into three phenotypes is demonstrated to result from a genetic polymorphism; indeed, the data fit a model consisting of two co-dominant apoprotein B alleles, and the three phenotypes then correspond to the two human homozygotes and the heterozygote. The classical Ag antigen phenotype was determined for the LDL isolated from 10 individuals who were also studied with the monoclonal antibodies, and no correspondence was found between the different epitopes recognized by the five presumptive Ag allelic pairs, x/y, a1/d, c/g, t/z, or h/i, and the site recognized by the monoclonal antibodies. All of these data are discussed, and it is concluded that the most likely explanation for the difference in recognition of the two allelic forms of apoprotein B is an alteration in amino acid sequence resulting in a slightly different configuration of a single domain containing the epitopes recognized by the three monoclonal antibodies.

Ag(c): recognition by a monoclonal antibody.

A monoclonal antibody directed against human apolipoprotein B, which was previously shown in family studies to detect allelic variations (J Biol Chem 1984; 259:6423-6430), has now been identified with the Ag(c) factor. This identification allows the location of the Ag system on the structural gene for apolipoprotein B and on the short arm of human chromosome 2. The epitope corresponding to Ag(c) is located within the amino acid sequence common to apolipoproteins B-100 and B-48. Since a single molecule of apolipoprotein B-100 is present on human LDL, individual LDL possesses either the epitope corresponding to Ag(c) or that corresponding to Ag(g). These studies on allelic variation among human apolipoprotein B species parallel similar studies in animals in which a relationship to atherosclerosis was found.

Apolipoprotein B genetic polymorphisms in several human hepatoma derived liver cell lines.

The genetic polymorphism of apoB EcoRI and XbaI restriction sites and the 3' VNTR hypervariable region was examined in nine human hepatoma derived liver cell lines and related to the cells' ability to secrete lipids and apoB. EcoRI and XbaI genotypes appeared to be unrelated to triglyceride, cholesterol and apoB accumulating in the medium. The VNTR consisted of alleles with 47 to 67 repeats; however, these repeats were not associated with elevated concentrations of lipid or apoB. Data suggest that in the hepatoma cell lines, apoB polymorphisms in EcoRI, XbaI and the VNTR hypervariable region are not sufficient in themselves to account for triglyceride, cholesterol and apoB in the medium. It is possible that intracellular apoB synthesis and/or degradation as well as postsecretory apoB binding and uptake are responsible for the variability of apoB and lipid accumulation in the culture medium.