Hemoglycin visible fluorescence induced by x rays.

Hemoglycin, a 1494 Da polymer composed of iron and glycine, has been detected in several carbonaceous meteorites. Iron atoms close out the ends of a 5 nm anti-parallel glycine beta sheet and contribute visible and near infrared absorptions that are not present with glycine alone. The 483 nm absorption of hemoglycin was discovered in theory and then observed on beamline I24 at Diamond Light Source. Light absorption in a molecule involves a coupled lower set of states receiving light energy that causes a transition into an upper set of states. In the reverse process, some energy source, such as an x-ray beam, populates the upper set of molecular states, which then radiates light as it returns to the lower "ground" set of states. We report on visible light re-emission during x-ray irradiation of a hemoglycin crystal. The emission is dominated by bands centered at 489 and 551 nm.

Human Angiostrongylus cantonensis: an update.

Angiostrongylus cantonensis was first discovered in 1935 and has become an important emerging pathogen causing human angiostrongyliasis. Major outbreaks of human angiostrongyliasis have been reported in endemic regions. Thousands of cases of human angiostrongyliasis have been documented worldwide. A. cantonensis has spread from its traditional endemic regions of the Pacific islands and Southeast Asia to the American continent including the USA, Caribbean islands and Brazil. Humans acquire A. cantonensis by consumption of raw or undercooked intermediate snail hosts or paratenic hosts. The main clinical manifestations of human angiostrongyliasis are eosinophilic meningitis and ocular angiostrongyliasis. The treatment of this disease includes supportive treatment, corticosteroid therapy, and combined therapy with corticosteroids and anthelminthics. The most effective method for prevention is to persuade people not to eat raw or undercooked intermediate and paratenic hosts.

Serial femtosecond and serial synchrotron crystallography can yield data of equivalent quality: A systematic comparison.

For the two proteins myoglobin and fluoroacetate dehalogenase, we present a systematic comparison of crystallographic diffraction data collected by serial femtosecond (SFX) and serial synchrotron crystallography (SSX). To maximize comparability, we used the same batch of micron-sized crystals, the same sample delivery device, and the same data analysis software. Overall figures of merit indicate that the data of both radiation sources are of equivalent quality. For both proteins, reasonable data statistics can be obtained with approximately 5000 room-temperature diffraction images irrespective of the radiation source. The direct comparability of SSX and SFX data indicates that the quality of diffraction data obtained from these samples is linked to the properties of the crystals rather than to the radiation source. Therefore, for other systems with similar properties, time-resolved experiments can be conducted at the radiation source that best matches the desired time resolution.

Time zero determination for FEL pump-probe studies based on ultrafast melting of bismuth.

A common challenge for pump-probe studies of structural dynamics at X-ray free-electron lasers (XFELs) is the determination of time zero (T0)-the time an optical pulse (e.g., an optical laser) arrives coincidently with the probe pulse (e.g., a XFEL pulse) at the sample position. In some cases, T0 might be extracted from the structural dynamics of the sample's observed response itself, but generally, an independent robust method is required or would be superior to the inferred determination of T0. In this paper, we present how the structural dynamics in ultrafast melting of bismuth can be exploited for a quickly performed, reliable and accurate determination of T0 with a precision below 20 fs and an overall experimental accuracy of 50 fs to 150 fs (estimated). Our approach is potentially useful and applicable for fixed-target XFEL experiments, such as serial femtosecond crystallography, utilizing an optical pump pulse in the ultraviolet to near infrared spectral range and a pixelated 2D photon detector for recording crystallographic diffraction patterns in transmission geometry. In comparison to many other suitable approaches, our method is fairly independent of the pumping wavelength (UV-IR) as well as of the X-ray energy and offers a favorable signal contrast. The technique is exploitable not only for the determination of temporal characteristics of the experiment at the interaction point but also for investigating important conditions affecting experimental control such as spatial overlap and beam spot sizes.

Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography.

We present a crystallography chip enabling in situ room temperature crystallography at microfocus synchrotron beamlines and X-ray free-electron laser (X-FEL) sources. Compared to other in situ approaches, we observe extremely low background and high diffraction data quality. The chip design is robust and allows fast and efficient loading of thousands of small crystals. The ability to load a large number of protein crystals, at room temperature and with high efficiency, into prescribed positions enables high throughput automated serial crystallography with microfocus synchrotron beamlines. In addition, we demonstrate the application of this chip for femtosecond time-resolved serial crystallography at the Linac Coherent Light Source (LCLS, Menlo Park, California, USA). The chip concept enables multiple images to be acquired from each crystal, allowing differential detection of changes in diffraction intensities in order to obtain high signal-to-noise and fully exploit the time resolution capabilities of XFELs.

Mucosal abnormalities in microsporidiosis.

OBJECTIVE: To determine the prevalence of microsporidiosis in HIV-infected patients with and without diarrhoea and to characterize alterations in mucosal architecture and brush border enzyme activities in patients with microsporidiosis. PATIENTS: A total of 259 HIV-infected patients undergoing oesophago-gastroduodenoscopy because of diarrhoea (n = 123) or other symptoms (n = 136) were studied. METHODS: Patients were evaluated for the presence of microsporidia by electron microscopy of duodenal biopsies. Brush border enzyme activities were measured by histochemistry and mucosal architecture was determined by three-dimensional morphometry in biopsies from patients with microsporidiosis and compared with biopsies from a subgroup of HIV-infected patients with or without other enteropathogens. RESULTS: Enterocytozoon bieneusi was detected in 17 patients and Encephalitozoon intestinalis was detected in two patients. Microsporidiosis was significantly more frequent in patients with chronic diarrhoea (19.1%; P < 0.0001) or in patients with acute diarrhoea (7.2%; P = 0.04) than in patients without diarrhoea (1.5%). Microsporidiosis was associated with lactase deficiency (P = 0.03) and a reduced activity of alkaline phosphatase (P = 0.028) and alpha-glucosidase (P = 0.025) at the basal part of the villus compared with brush border enzymes in patients without enteropathogens. Patients with microsporidia had reduced villus height (P = 0.043) and a villus surface reduced by 40% (P = 0.004) compared with patients with enteropathogens other than microsporidia. CONCLUSIONS: Our study confirms the association between microsporidia and diarrhoea. The pathophysiologic mechanism by which microsporidia cause diarrhoea appears in part to be malabsorption, caused by a reduction of absorptive mucosal surface and impairment of enterocyte function.

Aging impairs intestinal immunity.

The elderly are characterized by immunosenescence accompanied by high rates of morbidity and mortality associated with infectious diseases. Despite suggestions that the mucosal immune compartment is relatively unaffected by aging, there are marked deficits in the intestinal mucosal immune responses of old animals and elderly humans. Little is known about the mechanism(s) whereby aging disrupts intestinal immunity. However, several events in the genesis of the intestinal immune response may be perturbed during aging. The first step is the uptake of antigens by specialized epithelial cells (M cells) that overlie the domes of Peyer's patches. We are unaware of any studies on the efficacy of antigen uptake in the intestine as a function of age. The effects of aging on the next step, antigen presentation by dendritic cells and lymphocyte isotype switching, have not been resolved. The third event is the maturation of immunoglobulin A (IgA) immunoblasts and their migration from the Peyer's patches to the intestinal mucosa. Quantitative immunohistochemical analyses suggest that the migration of these putative plasma cells to the intestinal effector site is compromised in old animals. Local antibody production by mature IgA plasma cells in the intestinal mucosa constitutes the fourth step. We recently reported that in vitro IgA antibody secretion by intestinal lamina propria lymphocytes from young and senescent rats is equivalent. The last event is the transport of IgA antibodies across the epithelial cells via receptor-mediated vesicular translocation onto the mucosal surface of the intestine. Receptor-binding assays did not detect age-associated declines in receptor number or binding affinity in either rodent or primate enterocytes as a function of donor age. Efforts to identify the mechanism(s) responsible for the age-related decline in intestinal mucosal immune responsiveness may benefit by focusing on the homing of IgA immunoblasts to the effector site.

Modulation of lymphocyte subsets in Peyer's patches of mice treated with monoclonal antibody against helper T-cells.

Treatment of mice with anti-L3T4, a monoclonal antibody directed against helper T-cells, impairs clearance of intestinal Giardia muris infection. The present study examined the effect of anti-L3T4 treatment on mouse Peyer's patch cytoarchitecture and on the distribution of T-cell subsets within microenvironments of the follicle. Female BALB/c mice, aged 8 weeks, were given 4-7 weekly injections of either anti-L3T4 (1 mg/wk) or PBS (control group), and Peyer's patches were examined by immunohistochemistry or flow cytometry. In anti-L3T4-treated mice, Peyer's patch follicles (B-cell areas) were about two thirds the size of follicles in controls, and virtually all the size difference occurred in germinal centers. Peyer's patches were depleted of L3T4+ cells, yet the proportion of Thy-1.2+ (all T) cells was not decreased correspondingly, and the distribution of Thy-1.2+ cells in the patches was similar to that in control mice. In anti-L3T4-treated mice, Thy-1.2+ cells consisted of (a) Ly-2+ (cytotoxic/suppressor T) cells, and (b) a population of Thy-1.2+ cells that were neither L3T4+ nor Ly-2+. After treatment, Ly-2+ cells accounted for most of the T-cells in interfollicular areas and were also scattered in follicles, in germinal centers, and below the dome epithelium--in areas where L3T4+ cells predominated in control mice. Thy-1.2+ cells that were L3T4- and Ly-2- were mainly localized below the dome epithelium. These shifts indicate complex interrelationships among different lymphocyte subsets in Peyer's patches.

Immunohistochemical localization of peptidergic nerve fibers and neuropeptide receptors in Peyer's patches of the cat ileum.

This study examined the distribution of peptidergic nerve fibers in Peyer's patches to determine whether appropriate receptors were present. Vasoactive intestinal peptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP) and receptors for VIP and SP were localized in lymphoid follicles of the cat ileum using a combined indirect horseradish peroxidase and streptavidin-biotin method. The margins of follicles were innervated by nerve fibers containing VIP, SP and CGRP. Nerve fibers were predominantly around lymphatics and high endothelial venules at the edges of follicles. Specific receptors for VIP and SP were present at the margins of follicles and in the lamina propria around crypts. VIP receptors were numerous on T cells within and around high endothelial venules and lymphatic vessels and at the margins of follicles. SP receptors were identified on a small number of T and B cells, granulocytes and macrophages, restricted to the margins of follicles. The defined distribution in ileal lymphoid tissue of nerve fibers containing VIP and SP and the corresponding localization of their appropriate receptors support immunoregulatory roles for neuropeptides in mucosal immunity.

Premature trypsinogen activation during cerulein pancreatitis in rats occurs inside pancreatic acinar cells.

Although it is widely accepted that trypsinogen activation is an initiating event in the development of acute pancreatitis, its location inside the pancreas is not known. In our studies, acute edematous pancreatitis was induced in rats by one or two intraperitoneal injections of 50 microg cerulein/kg body weight. The pancreas was removed for examination 1 or 2 h after the first and the second cerulein injection, respectively. The cleavage product of trypsinogen activation, trypsinogen activation peptide, was specifically labeled on pancreatic tissue sections by a corresponding antibody, the signal enhanced by a biotin-avidin conjugate, and the site then visualized by coupled peroxidase activity on diaminobenzidine. The sections were examined by light microscopy. Trypsinogen activation peptide, reflecting activation of the pancreatic digestive enzyme trypsinogen, was detected inside pancreatic acinar cells in this animal model of acute pancreatitis. As early as 1 h after the first injection of cerulein, protease activation was seen within the apical pole of acinar cells. Protease activation was increased 2 h after the latter of two injections of cerulein and more evenly distributed within the cells. For the first time morphologic evidence confirms that the activation originates within the acinar cell, rather than from the interstitium or the duct lumen. The location of this activation at the apical site of the acinar cell indicates its origin from subcellular compartments involving the late steps in the secretory pathway.

Small intestine pathogens in AIDS: conventional and opportunistic.

The small intestine, coming in direct contact with ingested potential pathogens, depends on active mucosal immunity to withstand invasion and damage. In patients with AIDS and severe impairment of immunoregulatory lymphocytes, proliferation of protozoal, viral, bacterial, and fungal pathogens produces diarrhea and malabsorption. When noninvasive tests of stool and blood fail to identify responsible organisms, endoscopy can reveal mucosal lesions which are suggestive if not diagnostic. Cryptosporidium, cf2E. intestinalis, cf1CMV, MAC, and other infections can be identified by intestinal biopsy quicker and often at lower overall cost than they can be by culture.

Use of a hypobaric chamber as a model system for investigating ascites in broilers.

In a series of experiments, broilers were reared at simulated altitudes of 3000, 3500, and 5000 meters utilizing a hypobaric chamber. The purpose was to evaluate the chamber as a model for ascites syndrome and to further study the pathophysiology of this disease. Results show that this model can effectively reproduce the disease as it is seen in the field. Birds reared at the simulated altitude of 5000 meters had a statistically significant increase in EKG amplitude, packed cell volume, and heart-to-body-weight and lung-to-body-weight ratios when compared with controls reared at 366 meters. Additionally, there was a trend for birds dying of ascites to have larger-amplitude EKGs than survivors when reared at 5000 meters.

An infectious bronchitis virus isolated from chickens experiencing a urolithiasis outbreak. I. In vitro characterization studies.

An infectious bronchitis virus (IBV) strain isolated from commercial layers experiencing urolithiasis was 50-100 nm in size and possessed widely spaced, club-shaped surface projections. It was sensitive to lipid solvents and exhibited responses characteristic of IBV when exposed to heat, divalent cations, and trypsin. Reciprocal virus-neutralization tests demonstrated it to be closely related to Gray, JMK, Delaware 2868, and Delaware 2897 strains of IBV.

A case of acute pulmonary edema, splenomegaly, and ascites in guinea fowl.

Acute pulmonary edema, splenomegaly, and ascites were observed in a disease outbreak in adult white and pearl guinea fowl. The clinical history and gross and microscopic lesions resembled those described for marble spleen disease of pheasants and avian adenovirus group II splenomegaly of chickens. A small number of intranuclear inclusion bodies were found in liver, spleen, and lung sections of affected guinea fowl. Attempts to isolate virus and serological tests to detect the presence of viral antigens were unsuccessful. Adult female pearl guinea fowl experimentally exposed to pheasant and turkey isolates of type II avian adenoviruses developed gross and microscopic lesions similar to those seen in the field outbreak. The pheasant isolate was the more virulent. Intranuclear inclusion bodies were observed in liver, spleen, and lung sections of pearl guinea fowl inoculated with either of the virus isolates, and direct immunofluorescent examination revealed viral antigen in the spleen and lung.

Physiologic and electrocardiographic changes occurring in broilers reared at simulated high altitude.

An experiment was conducted to determine whether differences in the electrocardiograms (EKGs) of broilers reared at simulated high altitude from the day of hatch can be used to predict which birds are developing ascites. In three replicate experiments, conducted with 100 broilers per replicate, birds were reared at a simulated altitude of 3000 meters or at ambient atmospheric pressure. Lead I, II, and III EKGs were obtained from all birds on days 0, 14, 28, and 42. No consistent significant differences were seen on day 0 in the amplitude of the R or S wave or total amplitude of the QRS complex when broilers that developed ascites while being reared at simulated high altitude were compared with unaffected birds reared at simulated high altitude and with birds reared at ambient atmospheric pressure. On days 14 and 28, the average amplitude of the S wave and the total amplitude of the QRS complex were significantly higher in the ascites group than in the two other groups. Packed cell volumes were significantly higher in birds reared at simulated high altitude at all sampling days (days 14, 28, and 42) than in those reared at ambient atmospheric pressure, and they were significantly higher in the ascites group on day 28 than in the two other groups. Birds in the ascites group weighed significantly less than the two other groups by day 14, and this trend persisted.

Porcine focal symmetrical poliomyelomalacia: test for an interaction between dietary selenium and niacin.

Experiments were conducted to test the hypothesis that dietary supplementation with nicotinamide would retard or eliminate the signs of selenium induced porcine focal symmetrical poliomyelomalacia (PFSP). Mixed-sex feeder pigs, approximately five weeks old, were divided into four groups and daily received, by oral capsule, the following treatments: no supplementation (control); 2.86 mg sodium selenite per kg body wt (selenium only); 44 mg nicotinamide per kg body wt (niacin only); or both the niacin and selenium (niacin + selenium). Over the ten day treatment body weights and behavior scores were recorded, as well as collection of fluid (blood, serum, urine) samples. Upon death, tissue samples (kidney, liver, brain, spinal cord and muscle) were obtained. All of these samples were analyzed for total selenium and bioactive niacin compounds. After gross pathological analysis, 11 samples from specific brain and spinal cord regions were taken for fixation and processing for histological analysis by light microscopy. The selenium only group showed behavior signs related to PFSP after two days of treatment with the average time of death at 6.5 days. Tissue levels of selenium were elevated and histological analyses established the expected lesions of PFSP. No disorders were noted in the control and niacin only groups. The niacin + selenium groups had slightly retarded changes in behavior scores (first differences from controls on day 4) but their mean day of death (7.5 days of treatment) did not differ from that of the selenium only groups. Histological analyses of these tissues revealed similar lesions to the selenium only group, but they may have been of lesser magnitude. The data were consistent with, but only partially supportive of, the above hypothesis.

Changes in pulmonary arterial and femoral arterial blood pressure upon acute exposure to hypobaric hypoxia in broiler chickens.

An experiment was conducted to investigate the pulmonary arterial and femoral arterial pressure responses to acute hypobaric hypoxia. Twenty-four, 7-wk-old Hubbard x Hubbard male chickens were lightly anesthetized and catheters were introduced into the right femoral artery and the pulmonary artery. The birds were then placed in a hypobaric chamber, and blood pressure responses were monitored during acute (15 min) exposures to simulated altitudes of 2,000 and 4,000 m. The pulmonary artery pressure increased .7 and 4% during the first and second exposures to a simulated altitude of 2,000 m, whereas the femoral artery pressure decreased 6 and 8% during exposures to this altitude. The pulmonary artery pressure increased 7% on the first exposure and 23% (P < .05) on the second exposure to a simulated altitude of 4,000 m. The femoral arterial pressure decreased (P < .05) on both exposures to this altitude (29 and 24%, respectively). The initial femoral and pulmonary artery pressures and changes in these pressures upon exposure to hypobaric hypoxia were not consistently correlated with the characteristics of the electrocardiogram, packed cell volume, body weight, or the right:total ventricular weight ratio. These results indicate that acute hypobaric hypoxia elicits a hypotensive response in the systemic arterial circulation and a hypertensive response in the pulmonary arterial circulation of broiler chickens.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic unilateral occlusion of an extrapulmonary primary bronchus induces pulmonary hypertension syndrome (ascites) in male and female broilers.

Previously, it was demonstrated that acute (4 min) and chronic (12 d) occlusion of an extrapulmonary primary bronchus triggers pulmonary hypertension but not pulmonary hypertension syndrome (PHS, ascites) in broilers. The present study was conducted to determine whether a more prolonged period of bronchus occlusion causes PHS similar to that induced by clamping one pulmonary artery. Male and female broiler chicks, 14 to 18 d old, were anesthetized, the thoracic inlet was opened, and a silver clip was positioned to fully obstruct the left extrapulmonary primary bronchus (BRONCHUS CLAMP group) or the left pulmonary artery (PA-CLAMP group). Sham-operated chicks were anesthetized and the thoracic inlet was opened; however, neither the pulmonary artery nor the bronchus was clamped (SHAM group). An electrocardiogram (ECG) was obtained whenever clinical ascites became apparent in individual broilers, or prior to the final necropsy for broilers surviving to the end (Day 36) of the experiment. The right:total ventricular weight ratio (RV:TV) was evaluated as an index of pulmonary arterial pressure. Early post-surgical mortality (up to 21 d of age) was higher in the PA-CLAMP group (27% for males and females combined) than in the BRONCHUS CLAMP (10%) and SHAM (2%) groups. Cumulative ascites mortality (Days 22 to 36) also was higher in the PA-CLAMP group (86% for males, 77% for females) than in the BRONCHUS CLAMP (69% for males, 41% for females) and SHAM (23% for males, 0% for females) groups. Ascitic birds in all treatment groups had higher RV:TV ratios and more negative ECG Lead II S-wave amplitudes than nonascitic birds, reflecting the right ventricular hypertrophy and generalized ventricular dilation typically associated with PHS. These results demonstrate that unilateral bronchus occlusion is an effective experimental model for triggering ascites at a lower incidence than that obtained by occluding one pulmonary artery. Following the onset of pulmonary hypertension, the pathophysiological progression leading to ascites appears to be similar for broilers with either unilateral bronchus or pulmonary artery occlusion.