Zoonotic origin and transmission of Middle East respiratory syndrome coronavirus in the UAE.

Since the emergence of Middle East respiratory syndrome coronavirus (MERS-CoV) in 2012, there have been a number of clusters of human-to-human transmission. These cases of human-to-human transmission involve close contact and have occurred primarily in healthcare settings, and they are suspected to result from repeated zoonotic introductions. In this study, we sequenced whole MERS-CoV genomes directly from respiratory samples collected from 23 confirmed MERS cases in the United Arab Emirates (UAE). These samples included cases from three nosocomial and three household clusters. The sequences were analysed for changes and relatedness with regard to the collected epidemiological data and other available MERS-CoV genomic data. Sequence analysis supports the epidemiological data within the clusters, and further, suggests that these clusters emerged independently. To understand how and when these clusters emerged, respiratory samples were taken from dromedary camels, a known host of MERS-CoV, in the same geographic regions as the human clusters. Middle East respiratory syndrome coronavirus genomes from six virus-positive animals were sequenced, and these genomes were nearly identical to those found in human patients from corresponding regions. These data demonstrate a genetic link for each of these clusters to a camel and support the hypothesis that human MERS-CoV diversity results from multiple zoonotic introductions.

Molecular cloning, sequencing, and expression of Eimeria tenella HSP70 partial gene.

Members of the Eimeria genus are protozoan parasites of the subphylum Apicomplexa (Eimeriidae family), and belong to the coccidia group. Eimeria tenella is one of the most pathogenic species owing to its ability to penetrate the mucosa, and cause inflammation and damage. It is an obligate intracellular parasite that causes disease by destroying the host cells during multiplication. Heat shock protein 70 (HSP70) is a molecular chaperone that prevents cellular stress. The objective of this study was to clone, sequence, and express E. tenella HSP70 protein. After selecting the region of highest hydrophilicity in the hsp70 gene, we cloned complementary DNA (cDNA) into a pTrcHis2-TOPO vector and transformed it into TOP10 Escherichia coli cells; after induction, the bacteria expressed a 23-kDa protein with insoluble expression levels of approximately 5 mg/L. In summary, the partial hsp70 gene was successfully expressed in E. coli, producing a 23-kDa protein under insoluble conditions, and the antigen characteristics predicted by hydrophilicity analysis suggest the development of a vaccine for use in avian coccidiosis.

[Specific aspects of treatment for women with bipolar affliction]. / Spezifische Behandlungsaspekte bipolar erkrankter Frauen.

This manuscript summarizes specific issues in the disease course and pharmacological treatment of women with bipolar disorders. Gender differences relevant to the female biology manifest in symptoms, outcome, and course. The preponderance of depressive symptoms is typical, and the risk of rapid cycling is estimated to be eight times higher for women than for men. Comorbid anxiety and eating disorders occur more frequently in female patients. In planning treatment it is important to take fertility, contraception, and pregnancy into consideration and adjust the pharmacotherapy to harmonize with the patient's current phase of life. Little is known about potential sexual dysfunctions of bipolar women. Further research should include clinical and observational studies focusing on gender-specific differences in symptomatology, treatment, and long-term outcome of bipolar disorders.

Disseminated multiorgan MDR-TB resistant to virtually all first-line drugs.

The emergence of multidrug-resistant tuberculosis (MDR-TB) is a major global concern since, despite a complex treatment regime, it still remains a lethal threat. A 21-yr-old male HIV-negative migrant from Burma presented with a disseminated tuberculosis affecting the lung, spleen, liver, mediastinal and abdominal lymph nodes. This particular strain of Mycobacterium tuberculosis proved to be resistant to all but one (pyrazinamide) of the first-line drugs, i.e. rifampicin, isoniazid and ethambutol, plus streptomycin, rifabutin and ofloxacin. On the mere account of its susceptibility concerning kanamycin it could not be labelled as extensively drug-resistant tuberculosis. After 1month of a standard first-line four-drug regimen and a subsequent 4months of second-line treatment with amikacin, moxifloxacin, terizidone, protionamide, linezolid and pyrazinamide, sputum cultures eventually yielded constantly negative results. Likewise, the organ manifestations decreased significantly, so as to be virtually undetectable in computed tomography scans after 1yr of continuous treatment. A moderate pancytopenia reversed completely after dose adjustment of linezolid. Disseminated tuberculosis manifestations without typical pulmonary cavernous lesions are likely to represent primary infection rather than reactivation. Even a multiorgan disseminated MDR-TB with an extensive resistance pattern (including fluoroquinolones) can be successfully treated with an individual second-line treatment and result in considerably few adverse events.

Xenogenic oogenesis of chicken (Gallus domesticus) female primordial germ cells in germline chimeric quail (Coturnix japonica) ovary.

In present study, chicken primordial germ cells (PGCs) were transferred into quail embryos to investigate the development of these germ cells in quail ovary. Briefly, 2 microl of chicken embryonic blood (stage 14) or about 100 purified circulating PGCs were transferred into quail embryo. Contribution of chicken PGCs were detected in gonads of chimeric quail embryos (stage 28) by immunocytochemical staining of cell surface antigen SSEA-1, and by in situ hybridization (ISH) with female chicken specific DNA probe. As a result, 52.0+/-43.2 (n=18) and 42.7+/-27.3 (n=17) chicken PGCs were found in the gonads of chimeric quail embryo that was injected with chicken embryonic blood (stage 14) and about 100 purified circulating PGCs, respectively. Furthermore, the ovaries of 81.8% (9/11) 12 days post incubation (dpi) chimeric quail embryos were observed with a mean of 457.6+/-237.1 female chicken PGCs-derived oogonia scattered in ovarian cortex area. In 9 out of 12 newly hatched and one week old chimeric quail chicks, on average of 2883.0+/-1924.1 primary oocytes and 3 follicles derived from chicken PGCs were found, respectively. The present results suggest that chicken female PGCs are able to migrate, colonize, proliferate and differentiate into oogonia, primary oocytes in chimeric quail ovary.

A 40-basepair VNTR polymorphism in the dopamine transporter (DAT1) gene and the rapid response to antidepressant treatment.

Finding predictors of the response to antidepressant therapy is a major goal of molecular psychiatry. The genes encoding the serotonin (SERT) and dopamine (DAT1) transporters are among the possible candidate genes modulating an individual's antidepressant response. In a naturalistic prospective cohort study with a total of 190 fully assessed patients, improvement of depression symptoms during the 3 weeks following initiation of antidepressant therapy was recorded using the 21-item Hamilton Depression Rating Scale (HDRS). The SLC6A3 3' UTR 40-bp variable number of tandem repeats (VNTR) and the SLC6A4 5' 44-bp insertion/deletion polymorphism were analyzed by polymerase chain reaction. There was a significantly smaller number of rapid responders among homozygous carriers of the DAT1 9-repeat allele (9/9) than among heterozygous (9/10) and homozygous (10/10) carriers of the 10-repeat allele (19 versus 37 versus 52%, respectively, P=0.0037). Median decline in HDRS score was 35, 40, and 52% in patients with the 9/9, 9/10, and 10/10 genotypes, respectively (P=0.013). The effect was found in all classes of medications (selective serotonin reuptake inhibitors (SSRIs), tricyclics, mirtazapine, venlafaxine) and statistically significant also within the subgroup of patients having received SSRIs. The serotonin promoter insertion/deletion genotype had no effect in the entire study group, but there was an insignificant trend of better response in the l/l and l/s carriers who received SSRIs or mirtazapine. In conclusion, the dopamine transporter VNTR polymorphism influenced rapid response to antidepressant therapy. Compared with homozygous carriers of the 10-repeat allele, carriers of the 9/10 genotype had an odds ratio (OR) calculated by logistic regression analysis of 1.6 (95% CI 0.8-3.2) and carriers of the 9/9 genotype had an OR of 6.0 (1.5-24.4) for no or poor response. Further studies are required to confirm this clinical association and to elucidate the underlying mechanisms.

[The value of pharmacogenetic tests in antidepressive medication therapy]. / Pharmakogenomik in der klinischen Praxis. Der Stellenwert pharmakogenetischer Tests in der antidepressiven Arzneitherapie.

The pharmacokinetics and effect of antidepressants are influenced by genetic factors. Modern methods of genotyping allow fast and inexpensive identification of genetic variants and thus can be used in clinical diagnostics to improve the tolerance to drug therapy. Numerous studies have investigated the significance of genetic variants in drug-metabolizing enzymes, drug and natural substrate transporters, neurotransmitter receptors, and molecules involved in signal transduction. While the interindividual differences in oral clearance, half-life, and bioavailability caused by genetic variants in the cytochrome P450 liver enzymes can be overcome by individual adjustment of dosage according to certain genotypes, the effects of genetic variants in antidepressive target structures are more difficult to translate into clinical recommendations. This article gives an overview of the currently available literature and points to situations in which the determination of pharmacogenetic variants might change drug therapy or therapeutic strategies for the individual patient. Dose adjustments for common antidepressant drugs based upon differences in pharmacokinetic parameters caused by genetic variability will be given.

[Bipolar disorders--how to recognize and treat them]. / Bei jeder Depression immer auch an bipolare Erkrankungen denken. Die verkannte Manie.

Bipolar disorders are often diagnosed too late with an average of ten years elapsing between the first disease episode and the correct diagnosis and treatment. The most common misdiagnoses are unipolar depression, schizophrenia and ADHD (Attention Deficit Hyperactivity Disorder). The suicide rate associated with bipolar disease is very high. Treatment consists in the administration of mood stabilizers, in the first instance lithium, but also atypical neuroleptics or lamotrigine. In the depressive phase, additional antidepressants or lamotrigine, in the manic phase valproate or an antipsychotic agent may be needed. Medication must be continued unchanged for several months beyond acute treatment. The subsequent relapse prophylaxis depends on effectiveness, tolerability, comorbidity, suicidal risk and compliance. Pharmacotherapy is supplemented by psychotherapy and psycho-education.

Lithium augmentation in treatment-resistant depression: clinical evidence, serotonergic and endocrine mechanisms.

For now more than 50 years, lithium has been the gold standard for the pharmacologic treatment of bipolar disorder. However, its utility is not restricted to acute mania and prophylactic treatment of bipolar disorder. A relatively new indication for its use is the addition to an antidepressant in the acute treatment phase of unipolar major depression. To date, this treatment approach called lithium augmentation is the best-documented approach in the treatment of refractory depression. In international treatment guidelines and algorithms, lithium augmentation is considered a first-line treatment strategy for patients with a major depressive episode who do not adequately respond to standard antidepressant treatment. In a recent double-blind, placebo-controlled trial, lithium augmentation has demonstrated to also be effective in the continuation treatment phase to prevent early relapses. From animal studies there is robust evidence that lithium augmentation increases serotonin (5-HT) neurotransmission, possibly by a synergistic action of lithium and the antidepressant on brain 5-HT pathways. In contrast to the established decline of HPA system activity during treatment with tricyclic antidepressants, neuroendocrine studies on the effects of lithium augmentation on the HPA system showed an unexpected and marked increase in the ACTH and cortisol response in the combined DEX/CRH test. Here we review new data on the efficacy and mechanism of action of lithium augmentation.

Evaluation of a newly developed real-time PCR for the detection of Taylorella equigenitalis and discrimination from T. asinigenitalis.

A 'culture-LightCycler PCR' assay has been developed for the detection of Taylorella equigenitalis, the causative agent of contagious equine metritis (CEM) in horses. The primers and hybridisation probes were derived from the 16S rDNA sequence. Their specificity was determined in two closely related organisms and six commensal bacteria of the genital tract. The assay was specific for T. equigenitalis and discriminates T. asinigenitalis isolates. It also avoids misidentifications of morphologically and phenotypically similar organisms. The sensitivity was evaluated in comparison to a standard bacteriological culture method. It detected T. equigenitalis in 10 of 52 samples that had not been identified bacteriologically. The results indicated that the assay had a greater sensitivity. This is the first real-time PCR for the detection of T. equigenitalis and avoids PCR carry-over contamination. The 'culture-LightCycler PCR' assay is specific, sensitive and reproducible, and can be used effectively for the detection of T. equigenitalis infections.

[Primary subcutaneous hematogenous osteomyelitis in childhood]. / Die primär subakute hämatogene Osteomyelitis im Kindesalter.

The most frequent form of bone infection is haematogenous osteomyelitis (HOM), typically affecting infants and children. Dependent on the virulence of the pathogen and the patients immune response, one can distinguish between the acute (AHOM) and primary subacute haematogenous osteomyelitis (PSHO). In contrast to AHOM, diagnosis of PSHO is severely impeded in that clinical and blood-chemistry findings usually do not enable differentiation from primary malignant bone tumors. With a comparable age predilection and clinical symptoms, as well as very similar conventional radiographic, MRI- and bone-scan-findings, the most important differential diagnosis is Ewing's-sarcoma. The here demonstrated case of a 12 year-old girl shows that PSHA may imitate a sarcoma very closely, even concerning such usually fairly reliable radiographic aspects like osteolysis and lamellar periostal bone reaction. Despite the use of MRI, the diagnosis initially remained uncertain and a malignant bony lesion could only be ruled out after open biopsy and histopathological evaluation.

Analysis of ploidy (in megabase chromosomes) in Trypanosoma brucei after genetic exchange.

The megabase chromosomes of Trypanosoma brucei are normally diploid, but the extent to which this ploidy is maintained when parasites undergo genetic exchange is not known. To investigate this issue, a panel of 30 recombinant clones resulting from the co-transmission through tsetse flies of three different parental T. brucei lines in all pair-wise combinations (STIB 247, STIB 386 and TREU 927/4) were examined. These clones are products of 28 different mating events; four of them result from self-fertilisation and the others are F1 hybrids. DNA contents of the three parental lines were determined by flow cytometry and shown to differ only slightly with DNA content increasing in the order 927/4 < 247 < 386. Flow cytometry of the recombinant clones indicated DNA contents were similar to the parents in 28 clones and raised approximately 1.5 times the parental values in only two. The two F1 hybrid progeny with raised DNA contents were shown by marker analysis to be trisomic for seven independent loci indicating that they were probably triploid whereas progeny with DNA contents similar to parental values inherited a single allele from each parent for four independent loci indicating that they were diploid.

Evidence that FGF receptor signaling is necessary for endoderm-regulated development of precardiac mesoderm.

Endoderm cells in the heart forming region (HFR endoderm) of stage 6 chicken embryos are required to support the proliferation and terminal differentiation of precardiac mesoderm cells in vitro. The endoderm's effect can be substituted by growth factors, including members of the fibroblast growth factor (FGF) family. However, direct implication of FGFs in this process requires evidence that inhibition of FGF signaling interferes with proliferation and/or terminal differentiation. This report examines the consequences of treating endoderm/precardiac mesoderm co-explants with agents that inactivate FGF receptors. Using sodium chlorate, which prevents FGF ligand-receptor interaction, it was observed that the percentage of S-phase precardiac mesoderm cells was markedly reduced, suggesting that cell proliferation was inhibited. To more specifically affect FGF signaling, the explants were treated with an antibody that recognizes an extracellular domain of FGF receptor-1 (FGFR-1). This treatment similarly inhibited cell proliferation. Although both agents modestly delayed cardiac myocyte differentiation as indicated by the contractile function, expression of alpha-sarcomeric actin was not affected. These findings provide additional evidence that an intact FGF signaling pathway is required during heart development.

Regulation of new blood vessel growth into ischemic skeletal muscle.

PURPOSE: In a rabbit model, transposition of a muscle pedicle flap to an ischemic hind limb has been shown to result in the development of new blood vessels that connect the arterial circulation of the flap to the circulation of the limb. The hypothesis that exogenous recombinant basic fibroblast growth factor (bFGF) would enhance the development of this new blood supply was examined and the regulation of bFGF in this process was investigated. METHODS: The right common iliac artery was ligated in 12 male New Zealand white rabbits. An abdominal wall muscle flap based on the left inferior epigastric artery was transposed to the right thigh. bFGF in phosphate-buffered saline (PBS) at 3 ng/h (n = 6), or PBS alone (n = 6), was infused for 7 days via mini-osmotic pumps with an infusion catheter positioned at the flap-muscle interface. The flap-muscle interface was immunostained with anti-alpha-actin antibody to determine blood vessel density (number of vessels/mm) and with anti-bFGF antibody to evaluate bFGF distribution. RNA was isolated from these sections, and polymerase chain reaction (PCR) was used to examine endogenous bFGF messenger RNA (mRNA) expression. RESULTS: Blood vessel density was significantly increased in animals receiving exogenous bFGF (22. 0 +/- 10.6 vessels/mm vs. 10.7 +/- 8.8 vessels/mm, P =.009). In the controls, neovessels were arranged in clusters with endogenous bFGF concentrated around these clusters. In bFGF-treated animals, vessels were diffusely scattered throughout the flap-limb interface, corresponding to the distribution pattern of infused bFGF. There was no difference in bFGF mRNA expression between the control and the bFGF-treated groups. CONCLUSION: Exogenous bFGF infusion significantly augmented new blood vessel development at the flap-limb interface. Endogenous bFGF was up-regulated around the newly developed microvessels in control animals, and vessel growth correlated with the diffuse distribution of exogenous bFGF, implicating bFGF as an important factor in angiogenesis. Exogenous bFGF did not affect bFGF mRNA expression, suggesting that the regulation of bFGF is not under autocrine control.

Mutational analysis of acute-phase response factor/Stat3 activation and dimerization.

Signal transducer and transcription (STAT) factors are activated by tyrosine phosphorylation in response to a variety of cytokines, growth factors, and hormones. Tyrosine phosphorylation triggers dimerization and nuclear translocation of these transcription factors. In this study, the functional role of carboxy-terminal portions of the STAT family member acute-phase response factor/Stat3 in activation, dimerization, and transactivating potential was analyzed. We demonstrate that truncation of 55 carboxy-terminal amino acids causes constitutive activation of Stat3 in COS-7 cells, as is known for the Stat3 isoform Stat3beta. By the use of deletion and point mutants, it is shown that both carboxy- and amino-terminal portions of Stat3 are involved in this phenomenon. Dimerization of Stat3 was blocked by point mutations affecting residues both in the vicinity of the tyrosine phosphorylation site (Y705) and more distant from this site, suggesting that multiple interactions are involved in dimer formation. Furthermore, by reporter gene assays we demonstrate that carboxy-terminally truncated Stat3 proteins are incapable of transactivating an interleukin-6-responsive promoter in COS-7 cells. In HepG2 hepatoma cells, however, these truncated Stat3 forms transmit signals from the interleukin-6 signal transducer gp130 equally well as does full-length Stat3. We conclude that, dependent on the cell type, different mechanisms allow Stat3 to regulate target gene transcription either with or without involvement of its putative carboxy-terminal transactivation domain.

Evidence that fibroblast growth factors 1 and 4 participate in regulation of cardiogenesis.

Previous studies in this laboratory have indicated that the early embryonic chick heart depends on fibroblast growth factor-2 (FGF-2; bFGF), sequentially utilized in paracrine and autocrine fashion, for its growth and development (Sugi and Lough, [1995] Dev. Biol. 168-567-574). This view emanated from immunohistochemical detection of FGF-like antigens in endoderm cells at stage 6, and later in the early myocardium at stage 9+ (Parlow et al. [1991] Dev. Biol. 146:139-147). To identify other members of the FGF family that are expressed by these cells, we have used peptide-generated antisera that specifically recognize FGFs 1 and 4. Like FGF-2, FGFs 1 and 4 were exclusively detected in the endoderm at stage 5+ and later in the myocardium, appearing as punctate cytoplasmic deposits. However, whereas FGF-2 is first detected at stage 9+, FGFs 1 and 4 did not appear until stages 11 and 15, respectively. Expression of all FGFs peaked at stages 18-24, decreasing thereafter in parallel with reduced myocardial cell proliferation. To determine these isoproteins' ability to facilitate the completion of terminal cardiac myocyte differentiation, stage 5+ precardiac mesoderm was cultured in defined medium with purified FGFs. Like FGF-2, as little as 5-10 ng/ml FGF-1 or FGF-4 supported the proliferation and differentiation of precardiac myoblasts, resulting in the formation of a vesicle containing an adherent multilayer of synchronously contractile cells. Evidence that this represented FGF receptor-mediated signaling rather than a nonspecific effect of exogenous FGF was indicated by the ability of sodium chlorate to inhibit FGF-mediated cardiogenesis. These findings are consistent with the hypothesis that, like FGF-2, FGFs 1 and 4 participate in the regulation of early heart development via paracrine and autocrine mechanisms.