Molecular Prevalence of Anaplasma marginale and Ehrlichia in Domestic Large Ruminants and Rhipicephalus (Boophilus) microplus Ticks From Southern Luzon, Philippines.

Anaplasmosis and ehrlichiosis are tick-borne rickettsial diseases that cause significant economic losses in the livestock industry worldwide. Although bovine anaplasmosis is known to be endemic in the Philippines, epidemiological data is fragmented. Moreover, little is known about bovine ehrlichiosis in the country. In this study, the prevalence of Anaplasma marginale and Ehrlichia in cattle and water buffalo from provinces in the southern part of Luzon, Philippines, was investigated through PCR. Blood samples from 620 animals comprised of 512 cattle and 108 water buffalo and 195 tick samples were subjected to nested PCR targeting the groESL gene of Anaplasmataceae. Positive samples were further subjected to another nested PCR and conventional PCR to amplify the A. marginale groEL gene and the Ehrlichia dsbA gene, respectively. Selected A. marginale-positive samples were also subjected to nested PCR targeting the msp5 gene. Regardless of the animal host, the overall prevalence in blood samples obtained was 51.9% for Anaplasmataceae, 43% for A. marginale, and 1.1% for Ehrlichia. No water buffalo were positive for Ehrlichia. Meanwhile, 15.9, 6.7, and 2% of the tick samples, all morphologically identified as Rhipicephalus (Boophilus) microplus, were positive for Anaplasmataceae, A. marginale, and Ehrlichia, respectively. Sequence analysis of selected A. marginale msp5 amplicons showed that the isolates from the region share 94-98% identity to reported A. marginale from other countries. The phylogenetic tree showed clustering of isolates in the region and a close relationship with A. marginale isolates from other countries. Sequences of Ehrlichia amplicons from cattle and ticks were 97-100% similar to reported Ehrlichia minasensis isolates. This study showed the high prevalence of A. marginale in Luzon, Philippines, and provided the first molecular evidence of E. minasensis in the country.

Molecular Detection of Rickettsia Spp. and Coxiella Burnetii in Cattle, Water Buffalo, and Rhipicephalus (Boophilus) Microplus Ticks in Luzon Island of the Philippines.

Rickettsia and Coxiella burnetii are zoonotic, tick-borne pathogens that can cause febrile illnesses with or without other symptoms in humans, but may cause subclinical infections in animals. There are only a few reports on the occurrence of these pathogens in cattle and water buffalo in Southeast Asia, including the Philippines. In this study, molecular detection of Rickettsia and C. burnetii in the blood and in the Rhipicephalus (Boophilus) microplus ticks of cattle and water buffalo from five provinces in Luzon Island of the Philippines was done. A total of 620 blood samples of cattle and water buffalo and 206 tick samples were collected and subjected to DNA extraction. After successful amplification of control genes, nested PCR was performed to detect gltA of Rickettsia and com1 of C. burnetii. No samples were positive for Rickettsia, while 10 (cattle = 7, water buffaloes = 3), or 1.6% of blood, and five, or 1.8% of tick samples, were C. burnetii-positive. Sequence analysis of the positive amplicons showed 99-100% similarity to reported C. burnetii isolates. This molecular evidence on the occurrence of C. burnetii in Philippine ruminants and cattle ticks and its zoonotic nature should prompt further investigation and surveillance to facilitate its effective control.

Educating anesthesia residents to obtain and document informed consent for epidural labor analgesia: does simulation play a role?

BACKGROUND: Informed consent is required before placing an epidural. At our hospital, teaching of residents about this is done informally at the bedside. This study aimed to assess the ability of anesthesia residents to acquire and retain knowledge required when seeking informed consent for epidural labor analgesia. It assessed how well this knowledge was translated to clinical ability, by assessing the verbal consent process during an interaction with a standardized patient. METHODS: Twenty anesthesia residents were randomized to a 'didactic group' or a 'simulation group'. Each resident was presented with a written scenario and asked to document the informed consent process, as they normally would do (pre-test). The didactic group then had a presentation about informed consent, while the simulation group members interviewed a simulated patient, the scenarios focusing on different aspects of consent. All residents then read a scenario and documented their informed consent process (post-test). Six weeks later all residents interviewed a standardized patient in labor and documented the consent from this interaction (six-week test). RESULTS: There was no significant difference in the baseline performance of the two groups. Both groups showed significant improvement in their written consent documentation at the immediate time point, the improvement in the didactic group being greater. The didactic group performed better at both the immediate time point and the six-week time point. CONCLUSIONS: In this small study, a didactic teaching method proved better than simulation-based teaching in helping residents to gain knowledge needed to obtain informed consent for epidural labor analgesia.

Maintenance of clinical benefit in Crohn's disease patients after discontinuation of infliximab: long-term follow-up of a single centre cohort.

BACKGROUND: Tumour necrosis factor-blockade with infliximab has advanced the treatment of Crohn's disease. While infliximab is efficacious, it remains to be determined whether patients who enter clinical remission with an anti-tumour necrosis factor therapy can have their treatment stopped and retain the state of remission. AIM: To assess in patients with Crohn's disease who obtained infliximab-induced remission, the proportion who relapsed after infliximab discontinuation. METHODS: This longitudinal cohort study examined patients from a University-based IBD referral centre. Forty eight patients with Crohn's disease in full clinical remission and who then discontinued infliximab were followed up for up to 7 years. Crohn's disease relapse was defined as an intervention with Crohn's disease medication or surgery. RESULTS: Kaplan-Meier analysis of the proportion of patients with sustained clinical benefit demonstrated that 50% relapsed within 477 days after infliximab discontinuance. In contrast, 35% of patients remained well, and without clinical relapse, up to the end of the nearly 7-year follow-up. CONCLUSIONS: In patients with Crohn's disease with an infliximab-induced remission, stopping infliximab results in a predictable relapse in a majority of patients. Nevertheless, a small percentage of patients sustain a long-term remission.

Categories of resistance in St. Augustinegrass lines to southern chinch bug (Hemiptera: Blissidae).

The southern chinch bug, Blissus insularis Barber (Hemiptera: Blissidae), is the most important insect pest of St. Augustinegrass, Stenotaphrum secundatum (Walt.) Kuntze, in Florida and other Gulf Coast states. Resistance to southern chinch bug was identified previously in St. Augustinegrass lines 'FX-10' and NUF-76. Choice and no-choice tests and ovipositional and developmental studies were conducted to determine the categories of resistance in FX-10 and NUF-76 to southern chinch bug. When adult chinch bugs had a choice among attached stolons of three susceptible lines ('Floratam', 'Bitter Blue', and 'Palmetto') and the two resistant lines, chinch bugs were found significantly more often over a 5-d period on the susceptible lines. This result indicates the presence of antixenosis in the resistant lines FX-10 and NUF-76. In a no-choice study, chinch bugs produced less than half as many excretory spots on FX-10 as on the susceptible lines. Significantly fewer excretory spots produced by chinch bugs confined on NUF-76 accumulated by days 3 and 5 after release; however, on the first 2 d, the accumulative number of excretory spots was not significantly less than that found on susceptible lines. The no-choice study confirmed a high level of antixenosis in FX-10, a moderate level of antixenosis in NUF-76, and possible antibiosis in NUF-76. Ovipositional and developmental studies were conducted using only Floratam (a widely planted cultivar that was formerly resistant to B. insularis) and the two resistant lines. Adults released on Floratam produced 11 and 5 times more eggs and 18 and 9 times more offspring than adults on FX-10 and NUF-76, respectively. Plant anatomical and biochemical studies are required to investigate the exact cause of antixenosis in FX-10 and NUF-76 and possible antibiosis in NUF-76.

Bacterial 16S ribosomal DNA in house dust mite cultures.

BACKGROUND: Allergen extracts prepared from Dermatophagoides farinae contain significantly more endotoxin than Dermatophagoides pteronyssinus extracts, and extracts from both mite extracts contain more endotoxin than pollen extracts. Attempts to culture bacteria from mite cultures have failed to establish the sources of the endotoxin. OBJECTIVE: To determine the bacterial sources of endotoxin in mite extracts. METHODS: Live mites of both species were obtained from 2 sources, DNA was extracted from the mites, and DNA encoding bacterial 16S ribosomal RNA was amplified by using specific primers. The amount of bacterial DNA in each mite DNA sample was determined by quantitative PCR using an internal standard, and sequence homologies were determined from amplifications performed by using a high-fidelity DNA polymerase. RESULTS: DNA from D farinae appeared to contain between 11-fold and 24-fold more 16S ribosomal gene copies than the genomic DNA from D pteronyssinus (P < or = .003). Sequence analysis indicated the dominant presence of at least 3 phylogenetic clusters of Bartonella species (henselae, quintana, vinsonii, and grahamii), as well as uncharacterized alpha-proteobacteria, from both D farinae and D pteronyssinus. In a few clones, sequences from Escherichia coli, Pseudomonas species, and Acinetobacter species were also identified. CONCLUSION: House dust mite DNA contains evidence of Bartonella and other Gram-negative species. These Gram-negative species are likely to be the sources of the endotoxin found in mite allergenic extracts.

Substance abuse and major trauma in Johannesburg : trauma

Introduction. The abuse of alcohol and drugs is a major public health problem and intoxication is one of the most important risk factors for violence and other causes of injury. The aim of the current study was to evaluate the prevalence of drug and alcohol use associated with injury in adult trauma patients in an inner city trauma unit in Johannesburg. Methods. A cohort of 105 adult trauma patients admitted to our university trauma unit and its allied medicolegal laboratory were studied in order to provide prospective data and enable us to correlate the prevalence and type of substance abuse with the demographics and injury patterns after major trauma. Results. Forty-five patients required resuscitation at the Johannesburg Hospital Trauma Unit (JHTU) and 60 patients were examined at the Johannesburg Medicolegal Laboratory (JMLL); giving a total of 105 patients. Blood alcohol concentration (BAC) was positive in 59of the patients. The average BAC in the trauma unit patients was 37 mmol/l (0.17 g/dl) ; more than three times the legal limit for driving. Of the patients 43.7were positive for urinary cannabis; women were statistically significantly more likely to have taken cannabis than men (p = 0.039). There was no difference in rates of exposure to cannabis or alcohol in patients who were the victims of interpersonal violence compared with the victims of accidents (p = 0.17). Only 3 patients had taken other drugs of abuse; 2 had taken Mandrax (methaqualone) and 1 amphetamine. Conclusion. Alcohol and cannabis are commonly misused by trauma patients in Johannesburg; the degree of misuse of other drugs appears to be low. Intoxication is a significant risk factor for violence and accidents and the resultant injuries are a massive burden on our society. Doctors have a responsibility to highlight the association between substance misuse and trauma and should also attempt to persuade individual trauma patients to reduce future alcohol consumption

Co-localization of cell surface receptors at high spatial resolution by single-particle fluorescence imaging.

Dual-wavelength single-particle fluorescence imaging has been used to quantify the co-localization of receptors and/or ligands on cells by widefield microscopy. Methods for correction of chromatic aberration and identification of submicroscopic artefacts are presented, with data for the lipopolysaccharide/CD14 and MHC class II/CD74 systems.

Measurements of associations of cell-surface receptors by single-particle fluorescence imaging.

SPFI (single-particle fluorescence imaging) uses the high sensitivity of fluorescence to visualize individual molecules that have been selectively labelled with small fluorescent particles. The images of particles are diffraction-limited spots that are analysed by fitting with a two-dimensional Gaussian function. The spot intensities depend on whether they arise from one or more particles; this provides the basis for determining self-association of cell-surface receptors. We have used this approach to determine dimerization of MHC class II molecules and its disruption by interface peptides. We have also exploited the positional information obtained from SPFI to detect co-localization of cell-surface molecules. This involves labelling two different molecules with different coloured fluorophores and determining their positions separately by dual wavelength imaging. The images are analysed to quantify the overlap of the particle images and hence determine the extent of co-localization of the labelled molecules. The technique provides quantification of the extent of co-localization and can detect whether co-localized molecules occur singly or in clusters. We have obtained preliminary data for co-localization of lipopolysaccharide and CD14 on intact cells. We also show that HLA-DR (human leukocyte antigen-DR) and CD74 are partially co-localized and that interaction between these molecules involves the peptide-binding groove of HLA-DR.

Carpal tunnel syndrome despite negative neurophysiological studies.

The purpose of the present study was to compare the results of conservative and operative treatment for patients with carpal tunnel syndrome having normal neurophysiological studies. We studied 125 patients with normal neurophysiological studies and analysed eight symptoms and signs as "prognostic factors". Ninety-six patients were treated conservatively (splintage, steroid injection, antiinflammatory medications, activity modification) and 29 were treated surgically (open decompression). One year after initiation of treatment we assessed the outcome and statistically analysed (chi-square test) the differences between the two groups. We did not find any statistically significant correlation between "prognostic factors" and outcome. Twenty four percent of the group treated non-operatively had a good or excellent outcome, whereas 90% of the group treated operatively had a good or excellent outcome. This difference was statistically significant (p < 0.0001). Our study supports the view that the diagnosis of carpal tunnel syndrome is clinical and not neurophysiological. We now recommend operative treatment for these patients.

Current opinion regarding indications for emergency department thoracotomy.

BACKGROUND: Emergency department thoracotomy (EDT) is a dramatic but rarely lifesaving intervention. Clinical variability regarding indications for EDT has yet to be quantified. Members of the Eastern and American Associations for the Surgery of Trauma were questioned by mail to evaluate which clinical and demographic factors influence the decision to perform EDT and whether physicians perform EDT in accordance with current practice guidelines. METHODS: A single mailing of an anonymous survey was sent to 1,124 surgeons to collect institutional and physician demographics as well as indications for EDT on the basis of variable mechanisms of trauma, duration of arrest, and signs of life (SOL). Statistical analysis included the Pearson and linear-by-linear association chi(2) tests, independent samples t test, and univariate and multivariate analyses of variance; p values of < 0.05 were considered significant. RESULTS: Completed surveys were received from 358 respondents. After 54 surveys were excluded that were incomplete, late, or from noneligible respondents, 304 surveys were analyzed. There were no significant differences in EDT indications among institutions of differing caseload volume, exposure to penetrating trauma, trauma level designation, American College of Surgeons verification status, or residency program affiliation. In addition, neither the respondent's position nor whether attendings versus residents performed the majority of EDTs influenced clinical decision-making. Performance criteria for EDT were liberal in comparison with established guidelines, especially for blunt trauma. The presence or recent loss of SOL influenced responses, but respondents varied greatly in their definition of SOL. CONCLUSION: A lack of agreement exists regarding the indications for EDT in multiple clinical scenarios as well as in defining SOL. Indications for EDT were liberal, especially for blunt trauma-related indications, and were determined by clinical parameters, not by physician or institutional factors. Our results suggest that clinical practice is at variance with Advanced Trauma Life Support guidelines. We recommend that practice guidelines for EDT be established on the basis of a consensus definition of SOL to allow for a more uniform and selective approach to EDT.

Fluorescence recovery after photobleaching reveals that LPS rapidly transfers from CD14 to hsp70 and hsp90 on the cell membrane.

Although CD14 has been implicated in the immune recognition of bacterial lipopolysaccharide (LPS) from Gram-negative bacteria and also peptidoglycan (PGN) and lipoteichoic acid (LTA) from the outer cell wall of Gram-positive bacteria, accumulating evidence has suggested the possible existence of other functional receptor(s). In this study, we have used fluorescence recovery after photobleaching (FRAP) in order to get the first dynamic picture of the innate recognition of bacteria. We have found that the diffusion coefficient of CD14 remains unaffected after LPS ligation and that the diffusion coefficients of FITC-LPS and FITC-LTA bound to cells differ from that of CD14. Furthermore, FITC-LPS/LTA rapidly become immobile when bound to cells, suggesting that FITC-LPS/LTA must briefly associate with CD14 in the initial attachment process and rapidly move on to an immobile receptor or to a complex of receptors. Further FRAP experiments revealed that heat shock protein 70 (hsp70) and hsp90 are immobile in cell membranes, and antibodies against them were found to block the transfer of LPS to the immobile receptor and to inhibit interleukin 6 production upon LPS stimulation. These experiments indicated that LPS transfers from CD14 to hsp70 and hsp90, which may be part of an LPS/LTA multimeric receptor complex. Thus, hsps are implicated as mediators of the innate activation by bacteria.

Fat embolism syndrome after cementless total hip arthroplasty.

There are few reports in the literature of fat embolism syndrome after cementless total hip arthroplasty (THA). Most reported cases have occurred after fracture or cemented THA. We report a case of a healthy 51-year-old woman who underwent THA for osteoarthritis under spinal anesthesia. A press-fit cup and extensively porous-coated diaphyseal locking stem were used and inserted without cement. In the recovery room, the patient became hypoxemic and hypotensive and developed cortical blindness. The next day, a petechial rash was evident. Gurd's criteria for fat embolism syndrome were fulfilled. Her symptoms resolved over a 2-week period. Patients undergoing cementless THA are at risk for fat embolism syndrome, and this must be considered in the differential diagnosis for postoperative hypoxemia and neurologic deficits.

Investigations of spectrin-lipid interactions using fluoresceinphosphatidylethanolamine as a membrane probe.

The binding of human erythrocyte spectrin to large unilamellar vesicles (LUVET) formed by the extrusion technique has been studied using fluoresceinphosphatidylethanolamine (FPE) as a reporter of electrostatic membrane potential. Spectrin aliquots were added to a suspension of FPE-labelled LUVETs to elucidate both the type of charge involved and the dissociation constants for spectrin binding to various lipids. All binding experiments showed serial increases in FPE fluorescence intensity upon serial additions of spectrin, indicative of increasing positive charge at the membrane surface. This proves for the first time that although exhibiting an overall net negative charge, spectrin binds to lipid surfaces by presenting positive charges to the lipid surface. Binding curves were obtained from the change in fluorescence intensity upon each spectrin addition and analysed to determine dissociation constants. A K(d) of 0.14+/-0.12 microM was found for spectrin binding to FPE-labelled phosphatidylcholine/phosphatidylserine (PC/PS) LUVETs at 22 degrees C in high salt conditions. A similar K(d) of 0.17+/-0.11 microM was obtained for spectrin binding to neutral LUVETs composed of PC. However, binding was found to be much weaker for PC/PS LUVETs under low salt conditions with a K(d) of 1.22+/-0.48 microM.

Digital fluorescence imaging of trafficking of endosomes containing low-density lipoprotein in brain astroglial cells.

We have used digital fluorescence microscopy to examine transport of LDL-containing endosomes in rat brain astroglial cells to show that individual middle endosomes undergo rapid transitions between forward/backward movements and immobile states over short distances. The population of rapidly moving endosomes (>0.04 microm/sec) was 35. 9%, and the remaining endosomes were slowly moving or temporarily immobile (<0.04 microm/sec). The averaged motion was, however, a very slow perinuclear motion with a velocity of 3.25 microm/h. This small velocity is mainly due to frequent changing of directions in movements, requiring 6 h for a significant concentration around the circumference of the cell nuclei. The application of both anti-dynein antibodies and vanadate in permeabilized cells resulted in peripherally concentrated distribution of endosomes, probably due to inhibition of perinuclear motion by dynein-like motor proteins. These results imply that both dynein-like and kinesin-like proteins bind to the same endosome resulting in both perinuclear and peripherally directed movements.

Ambient pollution and heart rate variability.

BACKGROUND: We investigated associations between ambient pollution levels and cardiovascular function in a repeated measures study including 163 observations on twenty-one 53- to 87-year-old active Boston residents observed up to 12 times from June to September 1997. Particles with aerodynamic diameter

Anticipatory strategy training: implications for the postlingually hearing-impaired adult.

The present study was designed to determine if speech recognition performance will improve after subjects prepare for an unfamiliar communication situation as opposed to a familiar one. Forty-five normal-hearing subjects were divided into three groups: one trained using a well-known fairytale, one trained using an obscure fable, and one without training. Post-training, all groups performed similarly when tested on the familiar tale. When test material involved the unfamiliar fable, only the group trained on that material obtained significantly better scores than the other groups. Results support teaching clients that increasing their knowledge of upcoming unfamiliar events can improve subsequent speech recognition.

Intracellular and cell surface heterotypic associations of human leukocyte antigen-DR and human invariant chain.

The intracellular and cell-surface heterotypic associations of HLA-DR in the presence and absence of the invariant chain were investigated. Simultaneous confocal microscopy imaging of the Golgi apparatus and HLA-DR molecules revealed that cells transfected only with HLA-DR and not the invariant chain or HLA-DM, accumulate class II molecules mostly in the Golgi apparatus, proximal to the cell nucleus. In contrast, in cells transfected with both HLA-DR and the invariant chain, or HLA-DR, the invariant chain and HLA-DM, the class II molecules are more evenly distributed in intracellular compartments. Confocal microscopy and flow cytometry revealed that in the absence of the invariant chain, a greater number of HLA-DR molecules are transported to the cell surface. Biochemical experiments and nonequilibrium pH gradient electrophoresis revealed that HLA-DR associates with surface invariant chain in the presence of HLA-DM. In cells that lack HLA-DM, no cell-surface association of HLA-DR and Ii was observed. Taken together, these results reveal two separate and distinct functions for surface and intracellular invariant chain subsets. The intracellular invariant chain "arrests" the class II molecules in the endocytic pathway. In contrast, cell-surface invariant chain associates with class II molecules at the cell surface, possibly playing a role in recycling empty class II molecules or as an accessory molecule.

Sites in the United States contaminated with radioactivity.

Over the century that radioactive materials have been mined, processed, produced, and utilized, many sites across the United States have become contaminated. Such sites include bases and installations of the Department of Defense, weapons production and research facilities of the Department of Energy, properties under the authority of other Federal agencies, privately-owned and governmental facilities that are licensed by the Nuclear Regulatory Commission and its Agreement States, and sites licensed by or the responsibility of states. This review reports on aspects of work by the Environmental Protection Agency, the Department of Defense, the Department of Energy, the Nuclear Regulatory Commission, and others to identify sites contaminated with radioactive materials. It also describes the principal programs that have been instituted to deal with them.