Characterization of two unusual clinically significant Francisella strains.

We have isolated two phenotypically distinct nonfastidious Francisella strains (Fx1 and Fx2) from the blood of compromised patients with pneumonia and compared them with eight other Francisella strains, including Francisella tularensis biovar tularensis, F. tularensis biovar novicida, and F. philomiragia. Our isolates grew well on sheep blood agar, chocolate agar, modified Thayer-Martin agar, and Trypticase soy agar. Fx1 and Fx2 were determined to be within the Francisella genus by cellular fatty acid analysis and by the utilization of glucose, production of H2S and catalase, and lack of motility, oxidase, nitrate reductase, and gelatinase. They were additionally shown to belong to the species F. tularensis by sequencing of two variable regions comprising approximately 500 nucleotides of the 16S rRNA gene. Also, RNA probe hybridization confirmed their belonging to the species F. tularensis. However, the new strains, which are not identical, are distinguished from other F. tularensis strains by growth characteristics, repetitive extragenic palindromic PCR fragment pattern, and some biochemical tests. Key biochemical differences included the findings that Fx1 was positive for beta-galactosidase and arabinose hydrolysis and that both strains were citrulline ureidase positive and glycerol negative. Commercial F. tularensis antiserum agglutinated stock F. tularensis strains but not Fx1, Fx2, F. tularensis biovar novicida, or F. philomiragia; serum from either patient failed to agglutinate or only weakly agglutinated commercial antigen but showed agglutination when tested against each patient's respective isolate. Fx1 and Fx2 produced beta-lactamase. Because of their good growth, negative serology, and biochemical profile, the organisms could be misidentified in the clinical laboratory if standard strategies or commercial identification systems are used.     

Recruitment of fibre types and quadriceps muscle portions during repeated, intense knee-extensor exercise in humans

To investigate recruitment of slow-twitch (ST) and fast-twitch (FT) muscle fibres, as well as the involvement of the various quadriceps femoris muscle portions during repeated, intense, one-legged knee-extensor exercise, 12 healthy male subjects performed two 3-min exercise bouts at ~110% maximum thigh O₂ consumption (EX1 and EX2) separated by 6 min rest. Single-fibre metabolites were determined in successive muscle biopsies obtained from the vastus lateralis muscle (n=6) and intra-muscular temperatures were continuously measured at six quadriceps muscle sites (n=6). Creatine phosphate (CP) had decreased (P<0.05) by 27, 73 and 88% in ST fibres and 25, 71 and 89% in FT fibres after 15 and 180 s of EX1 and after 180 s of EX2, respectively. CP was below resting mean–1 SD in 15, 46, 84 and 100% of the ST fibres and 9, 48, 85 and 100% of the FT fibres at rest, after 15 and 180 s of EX1 and after 180 s of EX2, respectively. A significant muscle temperature increase (T_m) occurred within 2–4 s at all quadriceps muscle sites. T_m varied less than 10% between sites during EX1, but was 23% higher (P<0.05) in the vastus lateralis than in the rectus femoris muscle during EX2. T_m in the vastus lateralis was 101 and 109% of the mean quadriceps value during EX1 and EX2, respectively. We conclude that both fibre types and all quadriceps muscle portions are recruited at the onset of intense knee-extensor exercise, that essentially all quadriceps muscle fibres are activated during repeated intense exercise and that metabolic measurements in the vastus lateralis muscle provide a good indication of the whole-quadriceps muscle metabolism during repeated, intense, one-legged knee-extensor exercise.     

Prevalence, co-morbidity and correlates of mental disorders in the general population: results from the German Health Interview and Examination Survey (GHS)

BACKGROUND: The German National Health Interview and Examination Survey (GHS) is the first government mandated nationwide study to investigate jointly the prevalence of somatic and mental disorders within one study in the general adult population in Germany. This paper reports results from its Mental Health Supplement (GHS-MHS) on 4-week 12-month, and selected lifetime prevalence of a broad range of DSM-IV mental disorders, their co-morbidity and correlates in the community. METHODS: The sample of the GHS-MHS (n=4181; multistage stratified random sample drawn from population registries; conditional response rate: 87.6%) can be regarded as representative for the German population aged 18-65. Diagnoses are based on fully structured computer assisted clinical interviews (M-CIDI), conducted by clinically trained interviewers. RESULTS: 12-month prevalence for any DSM-IV study disorder is 31% (lifetime: 43%; 4-week: 20%) with anxiety disorders, mood disorders and somatoform syndromes being the most frequent diagnoses. Retrospective age of onset information reveals that most disorders begin early in life. Co-morbidity rates among mental disorders range from 44% to 94%. Correlates of increased rates of mental disorders and co-morbidity were: female gender (except for substance disorders), not being married, low social class, and poor somatic health status. Health care utilization for mental disorders depended on co-morbidity (30% in 'pure', 76% in highly co-morbid cases) and varied from 33% for substance use disorders to 75% for panic disorder. CONCLUSIONS: Results confirm and extend results from other national studies using the same assessment instruments with regard to prevalence, co-morbidity and sociodemographic correlates, covering a broader range of DSM-IV disorders [i.e. somatoform disorders, all anxiety disorders (except PTSD), mental disorders due to substance or general medical factor, eating disorders]. Intervention rates were higher than in previous studies, yet still low overall.     

Dissecting karyotypic patterns in renal cell carcinoma: an analysis of the accumulated cytogenetic data

Molecular cytogenetic analysis frequently shows human papillomavirus (HPV) integration near translocation breakpoints in cervical cancer cells. We have recently described a cluster of HPV18 integrations in the distal end of the common fragile site FRA8C at 8q24 in primary cervical carcinoma samples. Chromosome band 8q24 contains the MYC gene (alias c-MYC), FRA8C, and FRA8D. The MYC gene is frequently deregulated--usually by translocation or amplification--in various tumor types. In the present study, we performed a molecular cytogenetic analysis of HPV18 integration patterns and the 8q24 translocation in a primary cervical carcinoma and in HeLa cells using combined binary ratio-fluorescence in situ hybridization. Our aim was to determine how the chromosomal breaks involved in these events relate physically to the MYC gene; whether they map to the FRA8C site, the FRA8D site, or both; and how they correlate with the occurrence of DNA flexibility domains. The 8q24 translocation breakpoints mapped between stretches of integrated HPV18 sequences in the distal end of FRA8C. This region contained DNA helix flexibility clusters, several of which mapped in the vicinity of HPV integration sites and translocation breakpoints in cervical carcinomas. DNA helix flexibility clusters were also found near known MYC translocation breakpoints in Burkitt lymphomas (BL), but most BL breakpoints mapped clearly outside FRA8C. Our data revealed that FRA8C is involved in HPV integration and chromosomal translocations in cervical carcinoma; however, this fragile site is not involved in classical MYC translocations in most BLs. In the context of the familial nature of cervical cancer, FRA8C may be considered a candidate susceptibility region for cervical carcinoma.     

Glycoprotein B of bovine herpesvirus type 4: Its phylogenetic relationship to gB equivalents of the herpesviruses

In order to estimate the phylogenetic relationship of BHV-4 among the herpesviruses, we have cloned and sequenced its glycoprotein B (gB). The 2.6 kb open reading frame codes for a 874 amino acid long protein. The comparison of its deduced amino acid sequence with those of its counterparts in 19 distinct herpesviruses groups BHV-4 into the -herpesvirinae. The calculation of an evolutionary tree emphasized that BHV-4 is more closely related to herpesvirus saimiri (HVS) than to Epstein-Barr virus (EBV). However, in contrast to EBV and HVS, the gB of BHV-4 contains a putative protease cleavage site and 20 potential N-glycosylation sites. The alignment of the amino acid sequences revealed that 10 cysteine and 7 proline residues, as well as the motifs SPF and GQLG, were completely conserved among the 20 investigated gBs.     

Eccentric and concentric torque-velocity characteristics of the quadriceps femoris in man

Summary The primary purpose of this investigation was to study the eccentric and concentric torque-velocity characteristics of the quadriceps femoris in man using a recently developed combined isometric, concentric and eccentric controlled velocity dynamometer (the SPARK System). A secondary purpose was to compare the method error associated with maximal voluntary concentric and eccentric torque output over a range of testing velocities. 21 males (21–32 years) performed on two separate days maximal voluntary isometric, concentric and eccentric contractions of the quadriceps femoris at 4 isokinetic lever arm velocities of 0° · s^–1 (isometric), 30° · s^–1 120° · s^–1 and 270° · s^–1. Eccentric peak torque and angle-specific torques (measured every 10° from 30° to 70°) did not significantly change from 0° · s^–1 to 270° · s^–1 (p>0.05) (with the exception of angle-specific 40° torque, which significantly increased;p<0.05). The mean method error was significantly higher for the eccentric tests (10.6%±1.6%) than for the concentric tests (8.1%±1.7%) (p<0.05). The mean method error decreased slightly with increasing concentric velocity (p>0.05), and increased slightly with increasing eccentric velocity (p>0.05). A tension restricting neural mechanism, if active during maximal eccentric contractions, could possibly account for the large difference seen between the present eccentric torque-velocity results and the classic results obtained from isolated animal muscle.     

Further analysis of ATP-mediated activation of K+ channels in renal epitheloid Madin Darby canine kidney (MDCK) cells

ATP activates K⁺ channels by increasing intracellular calcium activity in Madin Darby canine kidney (MDCK) cells. The present study has been performed to test for the involvement of G-proteins and of protein kinase C in the intracellular transmission of these effects. To this end, the effect of ATP on intracellular calcium and K⁺ channel activity has been studied in cells pretreated with the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) and/or pertussis toxin. The ATP-induced increase of intracellular calcium is not significantly affected by pretreatment with pertussis toxin, is significantly blunted by pretreatment with TPA and is abolished by pretreatment with both pertussis toxin and the phorbol ester. The ATP activation of K⁺ channels is similarly blunted by pretreatment with TPA, but is not abolished by pretreatment with both the phorbol ester and pertussis toxin. Furthermore, the ATP-induced hyperpolarization is not abolished in cells pretreated with both pertussis toxin and TPA. In those cells, ATP may activate K⁺ channels by calcium-independent mechanisms or lead to localized increases of intracellular calcium sufficient to activate the K⁺ channels but escaping detection with fura-2 fluorescence.     

Rheogenic sodium-bicarbonate cotransport in the peritubular cell membrane of rat renal proximal tubule

The mechanism of bicarbonate transport across the peritubular cell membrane was investigated in rat kidney proximal tubules in situ by measuring cell pH and cell Na⁺ activity in response to sudden reduction of peritubular Na⁺ and/or HCO ₃ ^– . The following observations were made: 1. sudden peritubular reduction of either ion concentration produced the same transient depolarizing potential response; 2. bicarbonate efflux in response to peritubular reduction of bicarbonate was accompanied by sodium efflux; 3. sodium efflux in response to peritubular sodium removal was accompanied by cell acidification indicating bicarbonate efflux; 4. all aforementioned phenomena were inhibited by SITS (10^–3 mol/l) except for a small SITS-independent sodium efflux and depolarization which occurred in response to peritubular sodium removal and was not accompanied by cell pH changes; 5. bicarbonate efflux and accompanying potential changes in response to reduction of peritubular bicarbonate virtually vanished in sodium-free solutions. From these observations we conclude that bicarbonate efflux proceeds as rheogenic sodium-bicarbonate cotransport with a stoichiometry of bicarbonate to sodium greater than 1. The question which of the charged species of the bicarbonate buffer system moves cannot yet be decided. Attempts to determine the stoichiometry from the SITS-inhibitable initial cell depolarization and from the SITS-inhibitable initial fluxes suggest a stoichiometry of 3 HCO ₃ ^– : 1 Na⁺. In addition to sodium-dependent bicarbonate flux, evidence was obtained for a sodium-independent transport system of acids or bases which is able to regulate cell pH even in sodium-free solutions.     

Human papillomavirus type 16 DNA sequence

The complete nucleotide sequence of HPV16 DNA (7904 bp) cloned from an invasive cervical carcinoma was determined. Homology comparisons allowed us to align the major open reading frames with the other published papilloma virus DNA sequences. The general organization of the open reading frames is similar to that of the other four papillomavirus (BPV1, HPV1a, HPV6b, CRPV) already sequenced. The sequence reveals an interruption of the reading frame coding for a suspected E1 protein.     

The 49 K subunit of NADH: ubiquinone reductase (complex I) from Neurospora crassa mitochondria: primary structure of the gene and the protein

Summary The primary structure of the 49 K subunit of the respiratory chain NADH:ubiquinone reductase (complex I) from Neurospora crassa was determined by sequencing cDNA, genomic DNA and the N-terminus of the mature protein. The sequence lengths correlate to a molecular mass of 54002 daltons for the preprotein and 49239 daltons for the mature protein. The presequence consists of 42 amino acids of typical composition for sequences which target nuclear-encoded proteins into mitochondria. The mature protein consists of 436 amino acids and shows 64% similarity to a 49 K subunit of bovine heart NADH:ubiquinone reductase and 33% to a predicted translation product of an open reading frame in the chloroplast DNAs of Marchantia polymorpha and Nicotiana tabacum. Evidence for an iron-sulfur cluster in the subunit is discussed.