Serological identification of oral Bacteroides spp. by enzyme-linked immunosorbent assay

A rapid method for identifying black-pigmented oral Bacteroides spp. is described. Species-specific rabbit antisera to Bacteroides gingivalis, B. intermedius, and B. melaninogenicus were used in an enzyme-linked immunosorbent assay to identify clinical isolates of black-pigmented Bacteroides spp. from humans. The results showed excellent agreement with biochemical identification of B. gingivalis and B. intermedius. Only 36% of the B. melaninogenicus isolates were identified with the enzyme-linked immunosorbent assay, suggesting that this group of black-pigmented Bacteroides spp. is made up of more than one serotype. The serological enzyme-linked immunosorbent assay should enable rapid identification of black-pigmented Bacteroides spp. isolated from sites of oral diseases and may also be used to identify the presence of these organisms in complex bacterial mixtures from oral sites.     

Detection of binding antibodies to native and recombinant human immunodeficiency virus type 1 envelope glycoproteins following recombinant gp160 immunization measured by flow cytometry and enzyme immunoassays. The AIDS Vaccine Clinical Trials Network.

The ability of antibody induced by vaccination with recombinant gp160 (rgp160) to bind to native and recombinant human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins was measured. Thirty-three HIV-1-seronegative healthy adult volunteers were injected four times with 40 or 80 micrograms of an HIV-1LAV envelope glycoprotein candidate vaccine per dose. The vaccine consisted of rgp160 produced in insect tissue culture cells infected with a recombinant baculovirus which contains the gp160 gene from the HIV-1LAV strain. By using a flow cytometric indirect immunofluorescence assay (FIFA) to detect vaccine-induced antibody to native envelope glycoprotein expressed by target cells infected with HIV-1IIIB, sera from 9 of the 33 vaccinees were positive. These included sera from eight vaccinees which stained HIV-1IIIB-infected cells and sera from two vaccinees which stained target cells infected with HIV-1MN, a heterologous virus strain. None of the sera stained cells infected with the HIV-1RF strain. Envelope glycoprotein-binding antibody was more frequently detectable in an enzyme-linked immunosorbent assay (ELISA) by using rgp160 compared with that which was detectable in the FIFA with uninfected target cells which were pulsed with rgp160 antigen. Positive correlations were observed between the rgp160 FIFA and a whole-virus-lysate enzyme immunoassay, between the rgp160 FIFA and the rgp160 ELISA, and between the rgp160 ELISA and the whole-virus-lysate enzyme immunoassay. The ability of sera from some volunteers who received rgp160 vaccine to bind to HIV-1-infected cells suggests that further studies with this vaccine should be done.     

Dissociation between lymph node and peripheral reactivity in immunological unresponsiveness to DNCB

Contact sensitivity and production of anti-DNP-antibodies induced by epicutaneously applied dinitrochlorobenzene (DNCB) can be permanently suppressed by a previous intravenous injection of a high dose of dinitrobenzene-sulphonate (DNBSO₃). The animals fail to develop a paracortical area rich in pyroninophilic cells, in lymph nodes regional to the local application of hapten, characteristic of sensitized controls.

Following the injection of lymphoid cells from normal syngeneic donors into these unresponsive animals, large pyroninophilic cells develop in the paracortical area 4 days after sensitization and in some cases antibody production is restored. However, contact sensitivity remains suppressed.

If a high dose of DNBSO₃ (600 mg/kg) is injected intravenously on the same day as DNCB is applied locally, or when a low dose of DNBSO₃ (60 mg/kg) is administered 12 weeks before DNCB, contact sensitivity and production of anti-DNP-antibodies are also suppressed but in such animals the development of a paracortical area containing large pyroninophilic cells is not inhibited.

In both cases an immune reaction can be suppressed in the presence of a paracortical area containing large pyroninophilic cells. Therefore it appears that this state of unresponsiveness is not exclusively conditioned by the absence of large pyroninophilic cells in paracortical areas and may be associated with other factors.

     

Hemodynamic and hormonal responses to lower body negative pressure in men with varying profiles of strength and aerobic power

Hemodynamic, cardiac, and hormonal responses to lower-body negative pressure (LBNP) were examined in 24 healthy men to test the hypothesis that responsiveness of reflex control of blood pressure during orthostatic challenge is associated with interactions between strength and aerobic power. Subjects underwent treadmill tests to determine peak oxygen uptake ( O_2max) and isokinetic dynamometer tests to determine knee extensor strength. Based on predetermined criteria, subjects were classified into one of four fitness profiles of six subjects each, matched for age, height, and body mass: (a) low strength/average aerobic fitness, (b) low strength/high aerobic fitness, (c) high strength/average aerobic fitness, and (d) high strength/high aerobic fitness. Following 90 min of 0.11 rad (6°) head-down tilt (HDT), each subject underwent graded LBNP to –6.7 kPa or presyncope, with maximal duration 15 min, while hemodynamic, cardiac, and hormonal responses were measured. All groups exhibited typical hemodynamic, hormonal, and fluid shift responses during LBNP, with no intergroup differences between high and low strength characteristics. Subjects with high aerobic power exhibited greater (P < 0.05) stroke volume and lower (P < 0.05) heart rate, vascular peripheral resistance, and mean arterial pressure during rest, HDT, and LBNP. Seven subjects, distributed among the four fitness profiles, became presyncopal. These subjects showed greatest reduction in mean arterial pressure during LBNP, had greater elevations in vasopressin, and lesser increases in heart rate and peripheral resistance. Neither O_2max nor leg strength were associated with fall in arterial pressure or with syncopal episodes. We conclude that interactions between aerobic and strength fitness characteristics do not influence responses to LBNP challenge.     

Localization of two epitopes recognized by monoclonal antibody PCG-4 on Clostridium difficile toxin A.

The toxin A gene of Clostridium difficile contains a 2.5-kb region encoding a series of contiguous repeating units located at the COOH terminus of the molecule. We previously showed that the monoclonal antibody (MAb) PCG-4, which neutralizes the enterotoxic activity of toxin A, binds to epitopes located within these repeating units. In the present study, we subcloned a series of fragments from this portion of the gene. The recombinant peptides expressed from the gene fragments were examined for reactivity with MAb PCG-4 to identify the epitopes involved in binding. Our results showed that MAb PCG-4 recognizes epitopes in amino acid residues 2097 through 2141 and amino acid residues 2355 through 2398.     

Immunohistochemical evidence for a spinothalamic pathway co-containing cholecystokinin- and galanin-like immunoreactivities in the rat

Using indirect immunofluorescence technique combined with retrograde tracing as well as surgical lesions, a system of spinothalamic neurons containing both galanin- and cholecystokinin-like immunoreactivity has been defined. The cell bodies are located in the lumbar segments L1-L5 with a preferential localization dorsal to the central canal at rostral levels and lateral to the canal at caudal levels. The cells project via the ventral part of the lateral funiculus to the most ventral and posterior parts of thalamus. Here a distinct, varicose terminal network was seen extending caudally from an area lateral to the medial lemniscus, running medially over the medial lemniscus, traversing the parafascicular nucleus and running dorsal to the fasciculus retroflexus into the periventricular gray matter. Transection of various parts of the spinal cord as well as retrograde tracing experiments indicate that the spinothalamic galanin cholecystokinin system represents a crossed pathway. The present results demonstrate that a spinothalamic system can be characterized by its content of galanin- and cholecystokinin-like peptides, two putative messenger molecules. It is only a minor component of the total spinothalamic projection.     

Crystalline and disordered state of poly(dihexylsilylene) copolymers

A systematic comparison of random copolymers, derived from poly(dihexylsilylene) (PDHS) by incorporation of monomeric units with shorter unbranched alkyl side chains, has been carried out based on calorimetry, variable temperature UV spectroscopy, and ²⁹Si MAS (magic angle spinning) solid state NMR investigations. Also, hexylmethylsilylene units and branched monomers have been copolymerized. Up to 10% comonomer with shorter linear side chains (i. e., pentyl to propyl) could be incorporated into PDHS without impeding the all-trans order of the crystalline phase. In this case, the UV absorption maximum of the crystalline low-temperature phase was affected only slightly according to the length and fraction of the comonomer side chains. A less ordered crystal structure (λ_max = 345–355 nm) was observed when the content of comonomers with shorter side chains was about 20%. Yet, all these materials form conformationally disordered mesophases. A clear disordering transition and corresponding thermochromism was not observed any more when 50% of propyl side chains were incorporated. The order of the crystalline and the mesophase is also strongly perturbed if only a small fraction (4%) of the side chains are branched at C².     

Characterization of PaxA and its operon: a cohemolytic RTX toxin determinant from pathogenic Pasteurella aerogenes

Pasteurella aerogenes is known as a commensal bacterium or as an opportunistic pathogen, as well as a primary pathogen found to be involved in abortion cases of humans, swine, and other mammals. Using broad-range DNA probes for bacterial RTX toxin genes, we cloned and subsequently sequenced a new operon named paxCABD encoding the RTX toxin PaxA in P. aerogenes. The pax operon is organized analogous to the classical RTX operons containing the activator gene paxC upstream of the structural toxin gene paxA, which is followed by the secretion protein genes paxB and paxD. The highest sequence similarity of paxA with known RTX toxin genes is found with apxIIIA (82%). PaxA is structurally similar to ApxIIIA and also shows functional analogy to ApxIIIA, since it shows cohemolytic activity with the sphingomyelinase of Staphylococcus aureus, known as the CAMP effect, but is devoid of direct hemolytic activity. In addition, it shows to some extent immunological cross-reactions with ApxIIIA. P. aerogenes isolated from various specimens showed that the pax operon was present in about one-third of the strains. All of the pax-positive strains were specifically related to swine abortion cases or septicemia of newborn piglets. These strains were also shown to produce the PaxA toxin as determined by the CAMP phenomenon, whereas none of the pax-negative strains did. This indicated that the PaxA toxin is involved in the pathogenic potential of P. aerogenes. The examined P. aerogenes isolates were phylogenetically analyzed by 16S rRNA gene (rrs) sequencing in order to confirm their species. Only a small heterogeneity (<0.5%) was observed between the rrs genes of the strains originating from geographically distant farms and isolated at different times.     

Detection of DNA damage after hyperbaric oxygen (HBO) therapy

Hyperbaric oxygen HBO therapy is successfully used for the treatmentof a variety of conditions. However, exposure to high concentrationsof oxygen is known to induce damage to cells, possibly due toan increased oxygen radical production. As reactive oxygen speciesalso cause DNA damage, we investigated the DNA-damaging effectof HBO with the alkaline version of the single cell gel testcomet assay. Oxidative DNA base modifications were determinedby converting oxidized DNA bases to strand breaks using bacterialformamidopyrimidine-DNA glycosylase FPG, a DNA repair enzyme,which specifically nicks DNA at sites of 8-oxo-guanines andformamidopyrimidines. HBO treatment under therapeutic conditionsclearly and reproducibly induced DNA damage in leukocytes ofall test subjects investigated. Increased DNA damage was foundimmediately at the end of the treatment, while 24 h later, noeffect was found. Using FPG protein we detected significantoxidative base damage after HBO treatment DNA damage was detectedonly after the first treatment and not after further treatmentsunder the same conditions, indicating an increase in antioxidantdefences. DNA damage did not occur when the HBO treatment wasstarted with a reduced treatment time which was then increasedstepwise. ³To whom correspondence should be addressed