Metabolism and evolution of Haemophilus influenzae deduced from a whole-genome comparison with Escherichia coli.

BACKGROUND: The 1.83 Megabase (Mb) sequence of the Haemophilus influenzae chromosome, the first completed genome sequence of a cellular life form, has been recently reported. Approximately 75 % of the 4.7 Mb genome sequence of Escherichia coli is also available. The life styles of the two bacteria are very different - H. influenzae is an obligate parasite that lives in human upper respiratory mucosa and can be cultivated only on rich media, whereas E. coli is a saprophyte that can grow on minimal media. A detailed comparison of the protein products encoded by these two genomes is expected to provide valuable insights into bacterial cell physiology and genome evolution. RESULTS: We describe the results of computer analysis of the amino-acid sequences of 1703 putative proteins encoded by the complete genome of H. influenzae. We detected sequence similarity to proteins in current databases for 92 % of the H. influenzae protein sequences, and at least a general functional prediction was possible for 83 %. A comparison of the H. influenzae protein sequences with those of 3010 proteins encoded by the sequenced 75 % of the E. coli genome revealed 1128 pairs of apparent orthologs, with an average of 59 % identity. In contrast to the high similarity between orthologs, the genome organization and the functional repertoire of genes in the two bacteria were remarkably different. The smaller genome size of H. influenzae is explained, to a large extent, by a reduction in the number of paralogous genes. There was no long range colinearity between the E. coli and H. influenzae gene orders, but over 70 % of the orthologous genes were found in short conserved strings, only about half of which were operons in E. coli. Superposition of the H. influenzae enzyme repertoire upon the known E. coli metabolic pathways allowed us to reconstruct similar and alternative pathways in H. influenzae and provides an explanation for the known nutritional requirements. CONCLUSIONS: By comparing proteins encoded by the two bacterial genomes, we have shown that extensive gene shuffling and variation in the extent of gene paralogy are major trends in bacterial evolution; this comparison has also allowed us to deduce crucial aspects of the largely uncharacterized metabolism of H. influenzae.

Bacterial start site prediction.

With the growing number of completely sequenced bacterial genes, accurate gene prediction in bacterial genomes remains an important problem. Although the existing tools predict genes in bacterial genomes with high overall accuracy, their ability to pinpoint the translation start site remains unsatisfactory. In this paper, we present a novel approach to bacterial start site prediction that takes into account multiple features of a potential start site, viz., ribosome binding site (RBS) binding energy, distance of the RBS from the start codon, distance from the beginning of the maximal ORF to the start codon, the start codon itself and the coding/non-coding potential around the start site. Mixed integer programing was used to optimize the discriminatory system. The accuracy of this approach is up to 90%, compared to 70%, using the most common tools in fully automated mode (that is, without expert human post-processing of results). The approach is evaluated using Bacillus subtilis, Escherichia coli and Pyrococcus furiosus. These three genomes cover a broad spectrum of bacterial genomes, since B.subtilis is a Gram-positive bacterium, E.coli is a Gram-negative bacterium and P. furiosus is an archaebacterium. A significant problem is generating a set of 'true' start sites for algorithm training, in the absence of experimental work. We found that sequence conservation between P. furiosus and the related Pyrococcus horikoshii clearly delimited the gene start in many cases, providing a sufficient training set.

Computer survey for likely genes in the one megabase contiguous genomic sequence data of Synechocystis sp. strain PCC6803.

Using the computer program GeneMark, the open reading frames (ORFs) previously assigned within the one megabase sequence data of the genome of the cyanobacterium, Synechocystis sp. strain PCC6803 (Kaneko et al., DNA Res. 2: 153-166, 1995), were re-examined. Matrices required by GeneMark for its statistical calculation were generated and modified by running a script termed GeneMark-Genesis that performed recursive application of GeneMark against the Synechocystis data and evaluated the probability scores for optimization. Based on the matrices thus generated, 752 of the 818 previously assigned ORFs (92%) were supported by GeneMark as likely coding sequences, of which 26 were predicted to start at more internal positions than previously assigned. In addition, 50 ORFs were newly identified as likely coding sequences, most of them being shorter than 300 bp. Thus, the procedure was proven to be very powerful to locate likely coding regions within the genomic sequence data of Synechocystis without having prior information concerning their similarity to the genes of other organisms. However, GeneMark did not predict 66 previously assigned ORFs as likely genes: 14 of them showed significant degrees of similarity to known genes and 10 others were found within IS-like elements. It seems that these genes, many of which appear to be exogenous origin, escaped detection by GeneMark as in the case of "class 3 (horizontally transferred) genes" of E. coli, which in turn suggests that genes of different phylogenetic origins might also be detected as such by modifying the matrices.

A telemedicine consultative service for the evaluation of patients with urolithiasis.

OBJECTIVES: A 6-month pilot teleconsultative project linking Georgetown University Medical Center (GUMC) in Washington, DC, and City Hospital in Martinsburg, West Virginia, 90 miles away, was designed to assess the effectiveness of telemedicine on the clinical decision-making process for patients with urolithiasis. METHODS: The telemedicine system designed and tested for this project was based on a PC-based platform. Videoconferencing and review of the patient's imaging studies were performed over an Integrated Service Digital Network (ISDN) with 3 Basic Rate (BRI) ISDN lines providing a 336-kilobytes/s bandwidth through an Inverse Multiplexor (IMUX). Treatment options were recorded for the clinical trial group and a simulated study group by the consulting urologist after the initial telephone consultation, after the telemedicine consultation, and after examination of those patients transferred to GUMC. RESULTS: A total of 32 telemedicine consultations were performed: 14 in the clinical trial group and 18 in the simulated study group. The recommendation of the consulting urologist at the tertiary center was altered in 12 patients (37.5%) after the telemedicine consultation compared with the recommended treatment after the initial telephone consultation. CONCLUSIONS: In the evaluation of patients with urolithiasis, this telemedicine application enhanced the clinical decision-making process by allowing for improved quality of care through immediate access and effective transfer of information between the referring urologist, the patient, and the stone center specialist.

Management of locally advanced prostate cancer.

The staging and treatment of prostate cancer are complex, particularly in patients with clinical disease that has advanced locally beyond the confines of the gland. Management choices are made more difficult by a paucity of quality randomized and controlled studies. Staging has traditionally relied on digital rectal examination, which is now being augmented by improved noninvasive radiologic studies. Radiation is the most common form of treatment today, and newer techniques are being examined and compared to external-beam therapy. Surgical intervention as monotherapy has failed to show a survival advantage. Current approaches treatment appear to be evolving toward combination therapies, potentially incorporating hormonal manipulation. Patients with locally advanced disease should be encouraged to enter prospective clinical trials.

Media for an integrated nursing curriculum.

In conclussion, the media decision-making process for an integrated nursing curriculum is based upon the following steps: - An analysis of the philosophy and unifying strands in the conceptual framework of the curriculum model, - An evaluation of currently owned media in terms of the theoretical framework and educational objectives of the model, - An assessment of commercial media with realistic planning for purchase and/or production, and - The development of a systematic ongoing evaluation process by students, faculty, and nursing service personnel. The focus of this discussion has been clarification of the assessment and planning steps necessary in the media decision-making process for an integrated nursing curriculum. While these steps are applicable to any educational model, they have particular relevance to a curriculum model in which concept learning and interrelationships are stressed.

Pneumococcal conjugate vaccine for young children.

UNLABELLED: Pneumococcal disease is a common cause of morbidity and mortality in the pediatric population. Pneumococcal infections, which account for most serious bacterial disease in infancy and early childhood, are a major cause of acute otitis media, sinusitis, pneumonia, bacterial meningitis, and bacteremia. Streptococcus pneumoniae is the causative agent in a large percentage of these infections, although other microorganisms also play a role. The recent emergence of drug-resistant strains has provided a strong incentive for preventing pneumococcal infections by vaccination. However, the capsular polysaccharide pneumococcal vaccines used to immunize adults are neither immunogenic nor protective in young children due to poor antibody responses. Therefore, research has focused on development of additional immunogenic pneumococcal vaccines to provide long-term immunity in children < 2 years of age. The most promising approach has been the development of a protein-polysaccharide conjugate vaccine for the seven serotypes (4, 6B, 9V, 14, 18C, 19F, and 23F) that most commonly cause infections in childhood. An effective conjugate vaccine that protects against these serotypes has the potential to prevent 85 percent of bacteremia episodes, 83 percent of meningitis episodes, and 65 percent of otitis media cases in the U.S. among children younger than 6 years. The Food and Drug Administration (FDA) recently approved the first protein-polysaccharide conjugate vaccine to prevent invasive pneumococcal diseases in infants and toddlers < 2 years of age. This conjugated vaccine against pneumococcus uses the same technology as the successful vaccine against Haemophilus influenzae type b. It consists of an immunogenic but inert protein coupled covalently to the polysaccharide coat of the selected strains of pneumococci. The conjugated antigen induces a more powerful, T-cell-based immune response in infants, which is developed by the time they are 2 months of age. Some important questions regarding this vaccine for children < 2 years of age: Is the vaccine safe? Is it immunogenic? Is it efficacious in preventing invasive pneumococcal disease and controlling otitis media? FINDINGS: Results of three randomized double-blind trials designed to evaluate the safety and immunogenicity of this vaccine in healthy children < 2 years of age were reported within the last three years. The studies found that the vaccine is safe and highly immunogenic for all seven serotypes. The most recent study, involving over 37,000 young children, also evaluated the vaccine's efficacy, and reported that the vaccine is highly effective in preventing invasive disease and has had an impact on otitis media. CONCLUSIONS: The heptavalent pneumococcal conjugate vaccine is safe and highly effective in preventing pneumococcal meningitis and bacteremic pneumonia in young children < 2 years of age; it is less effective in preventing otitis media. Based on the results of three well-designed studies demonstrating the vaccine's safety, immunogenicity, and efficacy, the vaccine is safe and effective for active immunization of children < 2 years of age against invasive disease caused by seven Streptococcus pneumoniae serotypes included in the vaccine. At this time, there is no clear medical consensus regarding its safety and efficacy for control of otitis media in children < 2 years of age. This application has not been evaluated by the FDA. The pneumococcal conjugate vaccine should be considered experimental, and has not been shown to be safe or efficacious for Streptococcus pneumoniae disease other than that caused by the serotypes included in the vaccine and for invasive infection, such as bacteremia or meningitis, caused by other microorganisms.

An evidence-based evaluation of percutaneous vertebroplasty.

BACKGROUND INFORMATION: Percutaneous vertebroplasty is a therapeutic, interventional radiologic procedure that involves injection of bone cement into a cervical, thoracic, or lumbar vertebral body lesion for the relief of pain and the strengthening of bone. This procedure only recently has been introduced, and is being used for patients with lytic lesions due to bone metastases, aggressive hemangiomas, or multiple myeloma, and for patients who have medically intractable debilitating pain resulting from osteoporotic vertebral collapse. FINDINGS: Results from two uncontrolled prospective studies and several case series reports, including one with 187 patients, indicate that percutaneous vertebroplasty can produce significant pain relief and increase mobility in 70 percent to 80 percent of patients with osteolytic lesions in the vertebrae from hemangiomas, metastases, or myeloma, or with osteoporotic compression fractures. In these reports, pain relief was apparent within one to two days after injection, and persisted for at least several months up to several years. While experimental studies and preliminary clinical results suggest that percutaneous vertebroplasty can also strengthen the vertebral bodies and increase mobility, it remains to be proven whether this procedure can prevent additional fractures in the injected vertebrae. In addition, the duration of effect is not known; there were no long-term follow-up data on most of these patients, and these data may be difficult to obtain and interpret in patients with an underlying malignant process, because disease progression may confound evaluation of the treatment effect. Complications were relatively rare, although some studies reported a high incidence of clinically insignificant leakage of bone cement into the paravertebral tissues. In a few cases, the leakage of polymer caused compression of spinal nerve roots or neuralgia. Several instances of pulmonary embolism were also reported. Although patient selection criteria have not been definitely established, percutaneous vertebroplasty is considered appropriate treatment for patients with vertebral lesions resulting from osteolytic metastasis and myeloma, hemangioma, and painful osteoporotic compression fractures if the following criteria have been met: o Severe debilitating pain or loss of mobility that cannot be relieved by correct medical therapy. o Other causes of pain, such as herniated intervertebral disk have been ruled out by computed tomography or magnetic resonance imaging. o The affected vertebra has not been extensively destroyed and is at least one third of its original height. o Radiation therapy or concurrent surgical interventions, such as laminectomy, may also be required in patients with compression of the spinal cord due to ingrowth of a tumor. CONCLUSIONS: Percutaneous vertebroplasty has only recently been introduced as a treatment for osteolytic lesions and osteoporotic compression fractures of the vertebrae, but early results are promising. Up to 80 percent of patients with pain unresponsive to correct medical treatment experience a significant degree of pain relief, and few serious complications have been reported. However, relatively few patients have undergone this procedure, and there are no data from controlled clinical trials or from studies with long-term follow-up. At the present time this procedure is still in the investigational stages, but may be appropriate for patients with no other reasonable options for medical treatment.

Alcohol abuse in adolescents.

The problem of alcohol abuse among teenagers is a major health concern in this nation. The primary care provider's role in prevention, early detection and referral is diversified. This role is tailored in response to the clinician's assessment of his or her target population, the practice setting and the community resources and referral possibilities. Prevention and early detection are primary goals, with the aim of achieving health coping strategies, improving school performance, preventing accidents, increasing meaningful socialization, and preventing serious pathological changes and death as realistic outcomes for adolescents.

From the Archives of the AFIP. Microcystic adenoma of the pancreas.

Microcystic adenoma is a benign pancreatic neoplasm typically seen in elderly women. Presenting symptoms and signs such as pain, weight loss, jaundice, and a palpable mass are usually observed, but the lesion may be incidental and does not necessarily require excision. The tumor is generally large and well demarcated. It contains innumerable small cysts, giving it a honeycombed appearance. On computed tomographic scans, the hypervascular tumor has attenuation values close to those of water before contrast material is administered, but it usually enhances afterward. On sonograms, it is often predominantly echogenic, although some larger cysts may be seen. A calcified central scar may be seen with either modality.

How to interpret an anonymous bacterial genome: machine learning approach to gene identification.

In this report we address the problem of accurate statistical modeling of DNA sequences, either coding or noncoding, for a bacterial species whose genome (or a large portion) was sequenced but not yet characterized experimentally. Availability of these models is critical for successful solution of the genome annotation task by statistical methods of gene finding. We present the method, GeneMark-Genesis, which learns the parameters of Markov models of protein-coding and noncoding regions from anonymous bacterial genomic sequence. These models are subsequently used in the GeneMark and GeneMark.hmm gene-finding programs. Although there is basically one model of a noncoding region for a given genome, several models of protein-coding region are automatically obtained by GeneMark-Genesis. The diversity of protein-coding models reflects the diversity of oligonucleotide compositions, particularly the diversity of codon usage strategies observed in genes from one and the same genome. In the simplest and the most important case, there are just two gene models-typical and atypical ones. We show that the atypical model allows one to predict genes that escape identification by the typical model. Many genes predicted by the atypical model appear to be horizontally transferred genes. The early versions of GeneMark-Genesis were used for annotating the genomes of Methanoccocus jannaschii and Helicobacter pylori. We report the results of accuracy testing of the full-scale version of GeneMark-Genesis on 10 completely sequenced bacterial genomes. Interestingly, the GeneMark.hmm program that employed the typical and atypical models defined by GeneMark-Genesis was able to predict 683 new atypical genes with 176 of them confirmed by similarity search.

Deriving ribosomal binding site (RBS) statistical models from unannotated DNA sequences and the use of the RBS model for N-terminal prediction.

Accurate prediction of the position of translation initiation (N-terminal prediction) is a difficult problem. N-terminal prediction from DNA sequence alone is ambiguous is several candidate start sites are close to each other. Protein similarity search is usually unable to indicate the true start of a gene as it would require a strong protein sequence similarity at the N-terminal portion of a protein where conservative regions are rarely situated. With the aid of the GeneMark program for gene identification, we extract DNA sequence fragments presumably containing ribosome binding sites (RBS) from unannotated complete genomic sequences. These DNA segments are aligned to generate the RBS model using the Gibbs' sampling method. N-terminal prediction is then performed by using the RBS model in conjunction with the GeneMark start codon prediction to aid in determining the true N-terminal site.

From the Archives of the AFIP. Xanthogranulomatous pyelonephritis.

Xanthogranulomatous pyelonephritis is a form of chronic infection of the kidney and surrounding tissues characterized by destruction and replacement of renal parenchyma by lipid-laden macrophages. Gross pathologic features include massive renal enlargement, lithiasis, peripelvic fibrosis, hydronephrosis, and lobulated yellow masses replacing renal parenchyma. Typically, the disease is diffuse and has characteristic imaging features. Less commonly, the process is focal and is difficult to differentiate from malignant disease on radiologic studies. Ultrasound demonstrates renal enlargement with multiple anechoic or hypoechoic masses replacing the normal corticomedullary differentiation and a contracted pelvis. Peripelvic fibrosis may obscure acoustic shadowing from a central staghorn calculus. On computed tomographic scans, a staghorn calculus may be seen in a contracted renal pelvis of an enlarged kidney, with characteristic low-attenuation, peripherally enhancing rounded masses. Extrarenal extension of the inflammatory process is frequently seen.

Applications of GeneMark in multispecies environments.

This paper is supposed to bridge the gap between practical experience in using GeneMark for a rapidly widening repertoire of genomes, and the available publications that determine and compare the gene prediction accuracy of the GeneMark method for different genomes. Here we focus on the genome-specific variability of prediction error rates and their sources. DNA sequence inhomogeneity is present both in training and control sets of coding and non-coding regions. Coding region inhomogeneity, caused by differences in sequence composition between "native" and horizontally transferred genes or between genes expressed at different levels, contributes to the false negative error rate. Inhomogeneity of non-coding region may frequently be caused by the presence of unnoticed genes and contributes to the false positive error rate. We have documented such unnoticed genes in GenBank sequences for several species Some of protein products of these genes have been characterized by similarity search methods. For others, which we call "pioneer genes", no significant similarity has been found at a protein sequence level although the confidence of GeneMark prediction is high. For instance, to date a majority of those pioneer gene predictions made for E. coli now show strong similarity to more recently characterized proteins that have been added to protein sequence database. Another practical question is related to genomic sequence inhomogeneity at interspecies level: if GeneMark has not been trained for a particular species, is it possible to apply models derived for phylogenetically close genomes? The answer is, yes. The results of cross-species gene prediction experiments show that cross-species prediction can often be reasonably accurate.

GeneLynx: a gene-centric portal to the human genome.

GeneLynx is a meta-database providing an extensive collection of hyperlinks to human gene-specific information in diverse databases available on the Internet. The GeneLynx project is based on the simple notion that given any gene-specific identifier (accession number, gene name, text, or sequence), scientists should be able to access a single location that provides a set of links to all the publicly available information pertinent to the specified human gene. GeneLynx was implemented as an extensible relational database with an intuitive and user-friendly Web interface. The data are automatically extracted from more than 40 external resources, using appropriate approaches to maximize coverage of the available data. Construction and curation of the system is mediated by a custom set of software tools. An indexing utility is provided to facilitate the establishment of hyperlinks in external databases. A unique feature of the GeneLynx system is a communal curation system for user-aided annotation. GeneLynx can be accessed freely at http://www.genelynx.org.

Primary extragonadal germ cell tumors of the retroperitoneum: differentiation of primary and secondary tumors.

Primary extragonadal germ cell tumors (EGCTs) of the retroperitoneum are rarely encountered. In most cases, they arise from remnants of the genital ridge, whereas the majority of retroperitoneal germ cell tumors are metastases from primary testicular tumors. Differentiating between these primary and secondary tumors can be difficult because gonadal germ cell tumors can regress or develop slowly. Careful examination of the testes with ultrasound (US) is crucial for this determination, but asynchronous or metachronous lesions in both the testes and retroperitoneum can occur. Symptoms are often nonspecific, and diagnosis is frequently delayed. Serum markers such as beta-human chorionic gonadotropin and alpha-fetoprotein are often helpful in the diagnosis and follow-up of these tumors. Imaging techniques such as US and computed tomography play a major role in the localization and differentiation of primary and secondary retroperitoneal EGCTs. Treatment of EGCTs usually requires a combination of surgery, irradiation, and chemotherapy.

From the archives of the AFIP. Malignant germ cell tumors of the ovary: radiologic-pathologic correlation.

Malignant germ cell tumors are the most common malignant ovarian neoplasms in girls and young women. The most well known of these neoplasms are dysgerminoma, immature teratoma, endodermal sinus tumor, and a mixture of two or more of these elements. All malignant germ cell tumors, especially immature teratoma, can be associated with mature (benign) teratoma. These tumors are generally large and have a complex but predominantly solid appearance on cross-sectional images. Elevated serum alpha-fetoprotein and human chorionic gonadotropin levels can help establish the diagnosis. Cystic areas of calcifications suggestive of mature teratoma do not exclude a co-existing malignant neoplasm.

Extraadrenal retroperitoneal paraganglioma: clinical, pathologic, and CT findings.

Paragangliomas of the retroperitoneum arise from specialized neural crest cells distributed along the aorta in association with the sympathetic chain. In order to ascertain characteristic CT features of extraadrenal retroperitoneal paragangliomas to differentiate them from other retroperitoneal tumors, 31 discrete tumors and two cases of paragangliomatosis in 28 patients were reviewed retrospectively, and the CT features were correlated with clinical and pathologic findings. There were 16 men and 12 women. Average age was 37 years (range, 11-70 years). Twenty-four patients (86%) had hypertension. Of these, catecholamine levels were elevated in all 18 patients who had biochemical studies. Four patients (14%) had malignant paragangliomas. The discrete tumors were classified by location as suprarenal (26%), renal hilar (32%), or infrarenal (42%). Suprarenal paragangliomas could not be distinguished from the ipsilateral adrenal gland on CT. The average size of functional tumors was smaller (7.0 cm) than that of nonfunctional tumors (12.0 cm), but the sizes of the two groups overlapped. Smaller tumors were more likely to be homogeneous and have well-defined margins than were larger tumors. Our findings indicate that extraadrenal retroperitoneal paragangliomas are functionally active more often than previously reported and that they are readily detected by CT as soft-tissue masses closely associated with the entire length of the abdominal aorta. However, no CT feature was found that was unique for paraganglioma.

From the archives of the AFIP. Leiomyosarcoma of the retroperitoneum and inferior vena cava: radiologic-pathologic correlation.

Leiomyosarcoma is the second most common primary retroperitoneal tumor in adults. Retroperitoneal leiomyosarcoma exhibits three major growth patterns: (a) completely extravascular (extraluminal) (62% of cases), (b) completely intravascular (intraluminal) (5% of cases), and (c) extra- and intraluminal (33% of cases). The usual clinical manifestation is a large abdominal mass. Intraluminal leiomyosarcoma may be accompanied by symptoms referable to venous thrombosis. The variable gross features and potential for intravascular extension result in various radiologic appearances, the most common being a large, partially necrotic soft-tissue mass in the retroperitoneum with or without extension into the inferior vena cava. Computed tomography and magnetic resonance imaging typically show a nonfatty, necrotic retroperitoneal mass and a vascular component when it is present. Ultrasound and angiography may also be useful, especially if vascular involvement is suspected from other imaging studies. Treatment of choice is surgical excision, which is frequently impossible due to the invasiveness of the tumor. Prognosis is related to extent of invasion and the adequacy of resection. Long-term prognosis is poor, and most patients die of local recurrence or distant metastasis.

MR and CT evaluation of intracranial sarcoidosis.

Fourteen patients with CNS manifestations of neurosarcoidosis were evaluated by MR imaging and CT. Evaluations were done on a 0.5-T superconductive magnet with T1- and T2-weighted sequences. CT with contrast was obtained in all patients. The granulomatous lesions were classified by location into basilar, convexity, intrahemispheric, and periventricular white-matter involvement. Hydrocephalus with or without an associated lesion was also noted. MR determined the presence of disease in all patients (100%), but was less accurate than CT in depicting disease in two patients (14%). CT determined the presence of disease in 12 patients (85%) and was less accurate than MR in delineating hypothalamic involvement in two patients and periventricular white-matter disease in three patients. There was great variability in the appearance of intracranial sarcoidosis on MR. Three patients had lesions that were isointense or hypointense (relative to cerebral cortex) on both T1- and T2-weighted images while nine patients had lesions that were hyperintense on T2-weighted images. Convexity involvement and hydrocephalus were well documented by both CT and MR. These results indicate that both MR and CT are helpful in fully evaluating a patient with suspected intracranial sarcoidosis.