Quantitative galactomannan detection is superior to PCR in diagnosing and monitoring invasive pulmonary aspergillosis in an experimental rat model.

Two diagnostic tests, an Aspergillus-specific PCR and an enzyme-linked immunosorbent assay (ELISA) for the quantitative determination of galactomannan, were compared for diagnosing and monitoring invasive pulmonary aspergillosis. Persistently neutropenic rats with left-sided invasive pulmonary aspergillosis were sacrificed at regular intervals after inoculation. Blood samples and bronchoalveolar lavage (BAL) fluid were cultured and tested by PCR as well as by ELISA. Disseminated fungal infection in extrapulmonary organs was determined. The sensitivity of the ELISA was higher than that of the PCR on all days of measurements, in both blood and BAL fluid. Positive PCR or ELISA results in blood were not significantly associated with disseminated fungal infection. Serial testing in a separate group of rats showed consistently increasing concentrations of circulating galactomannan during the course of disease, while a positive PCR could be followed by negative results. The concentration of galactomannan was highly predictive for the time of survival (P < 0.0001). It was concluded that, in this model, quantitative galactomannan detection is superior to PCR in diagnosing and monitoring invasive pulmonary aspergillosis.

Ancient "dental implants": a recently proposed example from France evaluated with other spurious examples.

The recent announcement of the discovery of a first or second century AD Roman example of a dental implant appears to fall into a category of spurious claims regarding ancient dentistry. Several earlier claims of the discovery of impressive examples of ancient dentistry are reviewed, followed by a discussion of the recent discovery from France. This survey of all of these claims regarding finds of ancient dental implants, plus data from the modern and rapidly evolving field of implant dentistry, suggests that these "ancient implants" are the product of the imaginations of 20th-century archaeologists.

Scintigraphic imaging of bacterial and fungal infection in granulocytopenic rats.

UNLABELLED: Scintigraphic imaging in granulocytopenic patients can be very useful to detect and localize infections, which often do not show localizing signs and symptoms. We studied the potential of 99mTc-labeled polyethylene glycol (PEG)-coated liposomes and 99mTc-labeled IgG to image bacterial and fungal infection in a granulocytopenic rat model. 67Ga-citrate was used as a reference agent. METHODS: 99mTc-PEG-liposomes, 99mTc-hydrazinonicotinate (HYNIC)-IgG or 67Ga-citrate was administered to granulocytopenic rats with a Staphylococcus aureus abscess or with unilateral invasive pulmonary aspergillosis. Imaging and biodistribution studies were performed. RESULTS: All agents visualized the S. aureus infection from 1 h after injection onward. However, only with 99mTc-PEG-liposomes and with 99mTc-HYNIC-IgG did activity in the infectious foci increase with time up to 24 h. 99mTc-PEG-liposomes and 99mTc-HYNIC-IgG showed significantly higher accumulation in the infectious focus compared with 67Ga-citrate (1.33+/-0.31 and 1.40+/-0.16 percentage injected dose per gram [%ID/g], respectively, versus 0.31+/-0.04 %ID/g 24 h after injection; P<0.05). At 24 h after injection, abscess-to-muscle ratios were highest for 99mTc-liposomes (72.1+/-19.1), followed by 99mTc-HYNIC-IgG (18.3+/-3.3) and 67Ga-citrate (4.4+/-0.7). In pulmonary aspergillosis, both 99mTc-PEG-liposomes and 99mTC-HYNIC-IgG showed significantly higher uptake in the infected lung than did 67Ga-citrate (3.6+/-0.4 and 8.3+/-0.8 %ID/g, respectively, versus 1.3 %ID/g at 24 h after injection; P<0.05). CONCLUSION: 99mTc-PEG-liposomes and 99mTc-HYNIC-IgG performed better than did 67Ga-citrate in the localization of peripheral bacterial infection and fungal infection in the lung in granulocytopenic rats. The high focal uptake and high target-to-nontarget ratios of 99mTc-PEG-liposomes and 99mTc-HYNIC-IgG indicate that both radiopharmaceuticals may become valuable agents to image infection in granulocytopenic patients.

Proficiency testing of selected antigen and antibody tests for use in dogs and cats.

OBJECTIVE: To determine the correlation of seroimmunologic test results between reference and nonreference laboratories. DESIGN: Retrospective data analysis. PROCEDURE: Serum samples obtained from naturally infected dogs and cats were distributed to reference and nonreference laboratories for seroimmunologic testing. Correlation of test results was evaluated by use of nonparametric analysis. RESULTS: Correlation coefficients were high between laboratory groups for samples tested for feline immunodeficiency virus antibodies, FeLV antigen, and toxoplasmosis antibodies in cats. Results for feline immunodeficiency virus antibody tests from reference laboratories were more likely to be positive than results from nonreference laboratories. Test results for feline infectious peritonitis antibodies, antinuclear antibodies, and Borrelia antibodies in cats were not significant. Coefficient correlations were significant for results of heart-worm antigen, Brucella antibodies, Toxoplasma antibodies, antinuclear antibodies, and rheumatoid factor in dogs. Results for Borrelia antibodies were not correlated between laboratory groups. CLINICAL IMPLICATIONS: Results were highly correlated between reference and nonreference laboratories for 8 of 14 seroimmunologic tests. Seroimmunologic tests for use in cats were less correlated as a group than those for use in dogs. Poor correlation of results between laboratories was attributed to variations in control agents, antigens, reagents, technical expertise, and cutoff values and end-point titers used for diagnosis.

An automated enzymatic method for measurement of D-arabinitol, a metabolite of pathogenic Candida species.

An automated enzymatic method was developed for the measurement of D-arabinitol in human serum. The assay is based on a novel, highly specific D-arabinitol dehydrogenase from Candida tropicalis. This enzyme catalyzes the oxidation of D-arabinitol to D-ribulose and the concomitant reduction of NAD+ to NADH. The NADH produced is used in a second reaction to reduce p-iodonitrotetrazolium violet (INT) to INT-formazan, which is measured spectrophotometrically. The entire reaction sequence can be performed automatically on a COBAS MIRA-S clinical chemistry analyzer (Roche Diagnostic Systems, Inc., Montclair, N.J.). Replicate analyses of human sera supplemented with D-arabinitol over a concentration range of 0 to 40 microM demonstrated that the pentitol could be measured with an accuracy of +/- 7% and a precision (standard deviation) of +/- 0.4 microM. Serum D-arabinitol measurements correlated with those determined by gas chromatography (r = 0.94). The enzymatic method is unaffected by L-arabinitol, D-mannitol, or other polyols commonly found in human serum. Any of 17 therapeutic drugs potentially present in serum did not significantly influence assay performance. Data illustrating the application of the assay in patients for possible diagnosis of invasive candidiasis and the monitoring of therapeutic intervention are presented. The automated assay described here was developed to facilitate the investigation of D-arabinitol as a serum marker for invasive Candida infections.

Transfer of tumor immunity by both CD4+ and CD8+ tumor infiltrating T lymphocytes activated in vivo by IL-2 therapy of tumor bearing mice.

DBA/2 mice were inoculated i.p. with syngeneic SL2 lymphoma or P815 mastocytoma on day 0 and treated i.p. with 20,000 units IL-2/day on day 10-14. This treatment is curative for 70-90% of the tumor bearing mice. Peritoneal cells and/or spleen cells were isolated from responding mice at the last day of IL-2 therapy. The in vivo antitumor activity of these cells was tested in Winn Assays (i.p.) and by adoptive transfer (i.v.) into mice injected s.c. with tumor previously. Peritoneal exudate cells isolated on day 14 (PEC14) from mice cured of SL2 tumor were highly effective in Winn Assays. Up to 5 x 10(7) SL2 cells could be eliminated in naive mice when injected i.p. together with 2 x 10(7) PEC14. Adoptive transfer (i.v.) of PEC14, without the addition of IL-2, into mice s.c. injected with SL2 tumor cells 1 or 3 days earlier, also prevented outgrowth of the tumor cells. T cells isolated from the spleens were less effective. Only at E:T ratios of 100:1 and 10:1 was tumor outgrowth inhibited. The adoptive transfer of PEC14 resulted in a long lasting immunity of the recipient mice. Furthermore, it was shown that depletion of both the CD8+ and CD4+ T cells from the suspensions used in the transfer studies, resulted in a significant decrease of tumor inhibition. However, the effect was not abrogated completely. PEC14 isolated from IL-2-treated DBA/2 mice cured of P815 tumor, protected naive mice against P815 tumor at E:T ratio 20:1. Adoptive transfer (i.v.) of these PEC14 into mice bearing s.c. P815 did not have an antitumor effect. In conclusion, low dose i.p. IL-2 therapy predominantly induces locally both CD4+ and CD8+ T cells with a strong antitumor activity in vivo. The potency of the IL-2-induced immunity seems related to the type of tumor used.

Effector cells of low-dose IL-2 immunotherapy in tumor bearing mice: tumor cell killing by CD8+ cytotoxic T lymphocytes and macrophages.

The presence and cytotoxicity of tumor infiltrating cells is described in mice during effective immunotherapy with interleukin 2 (IL-2). DBA/2 mice were inoculated i.p. with 2 x 10(4) tumor cells on day 0 and treated with daily i.p. injections with 20,000 units IL-2 on days 10-14. Mice bearing a large syngeneic i.p. tumor burden (SL2 lymphoma, P815 mastocytoma, L5178Y lymphoma, or L1210 lymphoma) could be cured by i.p. immunotherapy with these low doses of IL-2. In the peritoneal cavity of these mice an infiltrate of mononuclear cells was present. Similar numbers of lymphocytes (10(6)-10(7)) and macrophages (+/- 10(7)) were present in control tumor bearing mice and IL-2 treated tumor bearing mice. The ratio of CD4+/CD8+ T lymphocytes in the peritoneal cavity of mice rejecting the SL2 tumor was smaller than 0.5, whereas this ratio is about 2 in naive mice. In the spleens of IL-2 treated tumor bearing mice only a minor decrease of CD4+/CD8+ ratio was observed from 2.1-2.4 to 0.9-1.9. T cells isolated from the peritoneal cavity of mice inoculated with SL2 tumor cells and treated with IL-2, were highly cytotoxic to SL2 cells: at E:T ratio 2:1 the cytotoxicity index was 37 +/- 3. This cytotoxicity was specific and mediated by CD8+ T lymphocytes. Macrophages that were present in the peritoneal cavity of mice treated with IL-2 were also highly cytotoxic. The C.I. of these cells was 63-76% at E:T ratio 1:1. Cytotoxic macrophages were also present in untreated tumor bearing mice. The i.p. injections of IL-2 (20,000 units/day) caused a four-fold increase in the local NK-activity in the peritoneal cavity in naive mice. These IL-2 injections did not generate LAK-activity in vivo. Specificity of the in vivo tumor rejection was tested by injection SL 2 i.p. on day 0 and P815 i.p. on day 10, or vice versa, followed by IL-2 treatment. Only the tumor cells that were injected on day 0 were rejected. These in vivo experiments point to specific tumor rejection. In conclusion, both cytotoxic macrophages and CTL's are present in a sufficient number and with sufficient cytotoxicity to explain the killing of tumor cells in the peritoneal cavity. The CTL-activity seems of decisive importance for tumor rejection as this is induced by IL-2.

Mandibular symphysis (medial suture) closure in modern Homo sapiens: preliminary evidence from archaeological populations.

Four archaeologically derived populations of human infants provide evidence for age at closure of the mandibular suture. These data suggest fusion by 7-8 months of age, with a range from 6 to 9 months. This provides a useful tool for age identification of the remains of young children recovered from archaeological and other contexts.

Fiber optic probe cytometer.

An optical fiber probe is used to both excite and collect fluorescence from a suspension of cells. The configuration of the probe is such that one or a few cells are sensed at a time, with a convenient cell concentration. With fluorescently labeled antibodies to cellular antigens, the fiber optic cytometer is able to identify the presence of a specific set of cells with high sensitivity.

Electrodiagnostic tests in the detection of malignant hyperthermia.

Malignant hyperthermia (MH) can be a catastrophic and sometimes fatal reaction in certain susceptible individuals who are exposed to skeletal muscle relaxants such as succinylcholine and/or certain inhalation anesthetics such as halothane. In an attempt to develop a safe, simple in vivo test to identify the at-risk patients, repetitive nerve stimulation (RNS) and electromyographic (EMG) studies were performed on 9 patients with MH, 15 patients suspected of having MH, and 9 control subjects. Serial recordings were made following injections of caffeine sodium benzoate, succinylcholine, and halothane at the test site. Measurements were done on the amplitude of the evoked compound motor action potentials (CMAP), on RNS, and on the number of negative peaks of motor unit potentials (MUP) on maximum EMG recruitment. The results showed the effects of the drugs on amplitudes of the CMAPs to be significant (P less than 0.001) in all three groups, but no significant differences were found among the groups (P = 0.93). The negative peaks of MUP on EMG recording showed a significant difference upon introducing succinylcholine (P less than 0.001), and significant interaction effects of the treatment by the patient group. Preliminary comparisons of in vivo tests with in vitro contracture tests in nine patients showed good correlation.

Brain stem infusion of the gamma-aminobutyric acid antagonist bicuculline increases plasma insulin levels in the rat.

Previous neuroanatomical and physiological studies have indicated that nucleus ambiguus (Amb) is one source of vagal motoneurons in the brain stem that innervates the pancreas and which, when stimulated, increases insulin release. To investigate one of the neurotransmitter inputs to Amb neurons and its relation to insulin secretion, bicuculline, a specific gamma-aminobutyric acid (GABA) antagonist, was infused bilaterally into the Amb region as well as into a neighboring area, the rostral level of the lateral nucleus tractus solitarius (nts) of anesthetized male rats. Experiments were carried out in the presence or absence of the alpha-adrenergic blocker phentolamine. In the absence of phentolamine, no increases in plasma insulin levels were seen after bicuculline or vehicle infusion into the Amb region or after bicuculline infusion into the nts, while plasma glucose levels were significantly increased. In the presence of phentolamine, bicuculline infusion into the Amb region led to a prompt and significant increase in plasma insulin levels that could not be accounted for by changes in glycemia. The infusion of vehicle into Amb or of bicuculline into nts produced small or insignificant increases in plasma insulin levels. These results suggest that Amb neurons capable of modulating plasma insulin levels are under tonic GABA inhibition, an effect that appears to be specific for Amb neurons, since bicuculline infusion into another brain stem nucleus (nts) had no effect on insulin release. The fact that phentolamine pretreatment was necessary to reveal the bicuculline-induced effects corroborates previous studies showing that in addition to a central nervous system inhibition of vagal motoneurons by GABA, there is a tonic sympathetic inhibitory input to the endocrine pancreas capable of masking any disinhibition of vagal motoneurons. The physiological role of GABA inhibition of Amb neurons that innervate the pancreatic beta-cells remains to be determined.

Monocyte-mediated regulation of cellular immunity in humans: loss of suppressor activity with ageing.

The effect of ageing on cellular immunity in humans was investigated. Human T cell cytotoxicity (CMC) measured by an in vitro xenogeneic assay was found previously to be depressed in individuals greater than 60 years old (group 2) compared to individuals less than 50 years old (group 1). Removal of plastic-adherent cells prior to sensitization in the xenogeneic CMC assay of human peripheral blood mononuclear cells (HPBM) resulted in a significantly higher rise (P less than 0.001, Wilcoxon rank test) in CMC activity in group 1 compared to group 2. Replacement of plastic-adherent cells (greater than 35% monocytes) to HPBM depleted of monocytes returned the CMC activity to the level observed with unseparated HPBM. Separation of HPBM on Percoll density gradients into monocyte-enriched (less than 75%) and monocyte-depleted (less than 3%) fractions indicated that monocytes were responsible for suppressing CMC in group 1. These results are consistent with the hypothesis that monocyte suppressor function for CMC response declines in ageing humans. When indomethacin (1 microgram/ml) was added to HPBM the Con A- and PHA-induced DNA synthetic response rose in groups 1 and 2. Indomethacin induced a significantly larger (P less than 0.01) rise in suboptimal PHA mitogenesis in group 1 compared to group 2. Individuals whose CMC response rose following adherent cell depletion were examined for the effect of indomethacin on the CMC response of their HPBM. In nine of 13 cases, addition of indomethacin also resulted in increased CMC activity.

Age-related changes in antibody-dependent cell-mediated cytotoxicity in mouse spleen.

The age-related variation in antibody-dependent cell-mediated cytotoxicity (ADCC) in spleen was examined in four different mouse strains (DBA/2, Balb/c, NZB and NZB/NZW). The ADCC effector activity against antibody-coated chicken red blood cells in untreated spleen from all four strains was roughly comparable. An initial rise in activity with increasing age until two to three months was followed by a sharp decline in effector activity. Treatment of the spleen cells with poly-l-lysine-coated carbonyl iron to remove phagocytes resulted in a decrease in ADCC activity in NZB, NZB/NZW and DBA/2 strains at all ages examined. The same general pattern of age-dependent decline was still seen in these depleted spleens. However, the same treatment essentially abolished ADCC in Balb/c spleen at all ages studied.

Immune complexes in hepatitis B antigen-associated periarteritis nodosa. Detection by antibody-dependent cell-mediated cytotoxicity and the Raji cell assay.

A subpopulation of peripheral blood lymphocytes with the ability to lyse target cells coated with specific antibody (antibody-dependent cell-mediated cytotoxicity, ADCC) was serially studied in a patient with hepatitis B antigen-associated periarteritis nodosa. The effector lymphocytes possess FC and complement receptors but do not require complement for functional activity. We found that the patient's ADCC was decreased during periods of disease activity and was almost normal during remission. The patient's serum could block ADCC in normal lymphocytes, and the blocking ability correlated with the concentration of immune complexes as determined by the Raji cell assay (a radioimmunoassay using complement receptors on human cultured lymphoblastoid cells). The concentration of immune complexes and the ADCC blocking ability of ther serum both correlated with disease activity. Serum from five other patients with active vasculitis was found to contain significant amounts of immune complexes and was able to block normal ADCC. It appears that the ADCC assay can be used to detect the presence of circulating immune complexes and to monitor disease activity in periarteritis nodosa.