Identification and characterization of two novel genomic RNA segments RNA5 and RNA6 in rose rosette virus infecting roses.

Rose rosette virus (RRV), a negative-strand RNA virus belonging to the genus Emaravirus, has recently been characterized to be the causal agent of rose rosette disease. Roses showing typical symptoms of RRV collected from a rose nursery in Florida were subjected to reverse transcription-PCR (RT-PCR) assay using primers corresponding to the conserved inverted 13 nucleotide long stretches found at the termini of the RRV genomic RNA segments. RT-PCR analysis yielded two novel genomic RNA segments, RNA5 and RNA6, in addition to the previously identified four RNA segments. The RNA5 is 1650 bp long and encodes for a polypeptide of 465 amino acids (54.3 K), while RNA6 is 1400 bp long and encodes for a polypeptide of 233 amino acids (27.05 K). RACE analysis showed that, both the RNA segments posses at their 5' and 3' termini, stretches of conserved inverted complementary13 nucleotides long sequence with two nucleotide mismatches as previously identified in other genomic RNA segments. Northern blot analysis as well as RT-PCR using specific primers showed the presence of the novel genomic RNA segments in infected plants, but absent in the non-infected plants. The GenBank Acc. Nos. for the sequences reported in this paper are KT007556 and KT007557.

First Report of Cucumber mosaic virus Associated with Capsicum chinense var. Scotch Bonnet in Florida.

Scotch bonnet (Capsicum chinense) is a tropical hot pepper variety that is grown in South America, the Caribbean Islands, and in Florida, and is an important cash crop. In Florida, scotch bonnet is grown on ~100 acres annually. Virus-like leaf symptoms including mosaic and yellow mottling were observed on scotch bonnet plants in a field at Quincy, FL, with a disease incidence of ~5%. Two symptomatic and one non-symptomatic plant sample were collected from this field for identification of the causal agent associated with the symptoms. Viral inclusion assays (2) of the epidermal tissues of the symptomatic scotch bonnet samples using Azure A stain indicated the presence of spherical aggregates of crystalline inclusion bodies. Testing of the symptomatic samples using lateral flow immunoassays (Immunostrips, Agdia, Elkhart, IN) specific to Cucumber mosaic virus (CMV), Potato virus Y (PVY), Pepper mild mottle virus (PMMoV), Tobacco mosaic virus (TMV), Zucchini yellow mosaic virus (ZYMV), and Papaya ringspot virus (PRSV), showed a positive reaction only to CMV. The sap from an infected leaf sample ground in 0.01 M Sorensons phosphate buffer (pH 7.0) was used to mechanically inoculate one healthy scotch bonnet plant (tested negative for CMV with Immunostrip) at the 2- to 3-leaf stage. The inoculated plant developed mild mosaic and mottling symptoms 12 to 14 days post inoculation. The presence of CMV in the mechanically inoculated plant was further verified using CMV Immunostrips. Total RNA was extracted (RNeasy Plant Mini Kit, Qiagen, Valencia, CA) from the previously collected two symptomatic and one non-symptomatic scotch bonnet samples. The samples were subjected to reverse-transcription (RT)-PCR assays using SuperScript III One-Step RT-PCR System (Invitrogen, Life Technologies, Grand Island, NY), and using multiplex RT-PCR primer sets (1). The primers were designed to differentiate the CMV subgroup I and II, targeting the partial coat protein gene and the 3'UTR. The RT-PCR assays using the multiplex primers produced an amplicon of 590 bp, with the CMV subgroup I primers. The RT-PCR product was only amplified from the symptomatic leaf samples. The obtained amplicons were gel eluted, and directly sequenced bi-directionally (GenBank Accession Nos. KF805389 and KF805390). BLAST analysis of these sequences showed 97 to 98% nucleotide identities with the CMV isolates in the NCBI database. The isolates collected in Florida exhibited highest identity (98%) with the CMV isolate from tomato (DQ302718). These results revealed the association of CMV subgroup I with symptomatic scotch bonnet leaf samples. Although CMV has been reported from scotch bonnet, this is the first report of its occurrence in Florida. References: (1) S. Chen et al. Acta Biochim Biophys Sin. 43:465, 2011. (2) R. G. Christie and J. R. Edwardson. Plant Dis. 70:273, 1986.

Synergetic bitherapy in mice with xenografts of human prostate cancer using a methional mimic (METLICO) and an aldehyde dehydrogenase 3 inhibitor (MATE): systemic intraperitoneal (IP) and targeted intra-tumoral (IT) administration.

An intraperitoneal (IP) monotherapy in nu/nu mice with subcutaneous xenografts of a human prostate epithelial cancer cell line:DU145 was undertaken with an aldehyde dehydrogenase 3 inhibitor MATE, that is a potent apoptogen on (DU145) in culture but not on their human prostate epithelial normal counterparts [13] . Tumour growth was slowed down but treatment had to be done 5days/week. To try to potentiate the action of MATE in vivo, a bitherapy was undertaken based on the synergetic apoptotic effect that had been observed previously in culture on DU145 treated with a methional mimic METLICO and DIMATE, an inhibitor of ALDH1 and ALDH3 [19]. The bitherapy with METLICO/MATE administered IP was as effective as the monotherapy with MATE alone by IP, but at a 2-fold lower dose of MATE and at a dose of METLICO that had no growth-inhibitory effect as a monotherapy . Hence there was definite synergism with bitherapy. To try to increase the efficacy of bitherapy, it was administered by the intra-tumoral (IT) route using the recently developed 20-bars-pressurized microinjection system from CERMA [16, 17]. IT administration of the bitherapy was indeed more effective than that by IP as regards tumour volumes are concerned. Histopathological analysis of IT-treated tumours confirmed that there were many necrotized zones but intact cells were still present. Approaches for treating a wider zone of tumour tissue by IT-bitherapy are discussed.

A set of glycoproteins recognized by A2B5 antibody with major bands at 55 and 76 kDa is overexpressed in human head and neck squamous cell carcinomas.

A2B5 antibody was found to strongly label frozen sections of human head and neck squamous cell carcinomas. The low amount of glycolipids (c-series gangliosides and sulfatides) purified from the same tumors and reactive with A2B5 by immunostaining on thin-layer plates could not account for the high level of tissue labeling. Proteins were extracted from both normal tissues and squamous cell carcinomas and analyzed by Western blot with A2B5 antibody on PVDF membranes. The antibody was found to stain a set of glycoproteins with two major bands at 55 and 76 kDa present in normal tissues and overexpressed in carcinomas. Staining was abolished by prior treatment of the PVDF membranes either with Arthrobacter ureafaciens neuraminidase or with a solution of 10 mM periodate that is known to destroy carbohydrates. Our results show that the carbohydrate epitope recognized by A2B5 antibody can be displayed by both glycolipids and glycoproteins.

Outcome of patients with idiopathic pulmonary fibrosis admitted to the ICU for respiratory failure.

STUDY OBJECTIVES: To analyze the outcome of acute respiratory failure (ARF) in patients with idiopathic pulmonary fibrosis (IPF), and to evaluate the benefits of invasive and noninvasive mechanical ventilation (MV). DESIGN: Retrospective study. SETTING: University hospital. PATIENTS: Fifteen consecutive patients with IPF referred to the ICU for ARF between January 1989 and June 1998. MEASUREMENTS AND RESULTS: Fifteen patients (mean +/- SD age, 64 +/- 10 years) were included. Eight patients had clinical, functional, and radiologic features of IPF, and the remaining seven patients also had biopsy specimen-proven IPF. The mean duration between diagnosis of IPF and admission to the ICU was 26.5 +/- 28 months. At the time of ICU admission, mean arterial blood gas levels were as follows: PaO(2)/fraction of inspired oxygen, 113 +/- 95; pH, 7.32 +/- 0.10; and PaCO(2), 55 +/- 21 mm Hg. All patients received MV; 12 patients required tracheal intubation, either at the time of ICU admission (n = 10) or after failure of noninvasive ventilation (NIV; n = 2); and 3 patients only received NIV. Three of the five patients receiving NIV died of respiratory failure. Eleven patients died in the ICU, either from hypoxemia (n = 8) or from septic shock (n = 3). Four patients were discharged alive from the ICU, and two of them died shortly thereafter. CONCLUSION: The outcome of patients with IPF referred to the ICU for ARF was very poor and not improved by MV. Without a clearly identified reversible cause of ARF, these patients should not benefit from admission to the ICU.

Hyperintense benign liver lesions on spin-echo T1-weighted MR images: pathologic correlations.

BACKGROUND: To determine the incidence of hyperintensity on T1-weighted spin echo (SE) images in benign liver lesions, value of fat-suppressed magnetic resonance (MR) imaging for the detection of fat within these lesions, and the causes of hyperintensity by correlation to pathologic examinations. METHODS: Five hundred forty-nine patients with 805 benign liver lesions including 585 hemangiomas, 188 focal nodular hyperplasias (FNHs), 14 hepatic adenomas (HAs), 14 focal fatty infiltrations (FFIs), two biliary cystadenomas, and two hemorrhagic cysts were examined by T2-weighted and T1-weighted SE MR imaging. For hyperintense lesions on T1-weighted SE images, fat-suppressed images were obtained by selective presaturation of fat. RESULTS: Thirty-two lesions (four FNHs, 10 HAs, 14 FFIs, two biliary cystadenomas, and two hemorrhagic cysts) appeared hyperintense on T1-weighted SE images; 21 of these became hypointense on the fat-suppressed T1 weighted SE images (one FNH, six HAs, and 14 FFIs) and contained fat at pathological examination. The other 11 lesions remained hyperintense on fat-suppressed T1-weighted SE images and had no fat deposition. Causes of hyperintensity in these cases were sinusoidal dilatation, copper deposition, hemorrhage, and high protein content. CONCLUSION: Among benign liver lesions, hyperintensity on T1-weighted SE images is rare (3.9%). Causes of this hyperintensity are fat deposition, copper accumulation, sinusoidal dilatation, bemorrhage, and high protein content. Fat-suppressed imaging can distinguish fat deposition from other causes of hyperintensity.

[Role of magnetic resonance imaging in liver pathology]. / Place de l'imagerie par résonance magnétique en pathologie hépatique.

The increasingly common use of ultrasound examination of the liver has led to increased detection of hepatic tumours, showing a hitherto unsuspected prevalence. Improved techniques in MRI, such as dynamic sequences and new contrast media, are ameliorating the detection and characterization of the lesions, in particular with regard to computed tomography. In addition, since MRI is more sensitive to different components than in computed tomography, it permits better study of diffuse diseases such as haemochromatosis or fatty infiltration, particularly in pseudotumoural forms. The continuing progress in system improvement makes MRI an excellent method for diagnosis of bile duct disorders, especially in case of obstruction.

Thin-section CT detection of emphysema associated with bronchiectasis and correlation with pulmonary function tests.

PURPOSE: To evaluate, on thin-section CT scans, the prevalence of emphysema in patients with bronchiectasis and to correlate the results of thin-section CT scans with the results of pulmonary function tests, in order to question whether there was a particular functional test profile in this group of patients. PATIENTS AND METHODS: This is a retrospective study including 90 patients having both thin-section CT scans and pulmonary function tests for bronchiectasis. A CT scoring system was established for assessing the airway disease by the severity and extent of bronchiectasis and by the extent of emphysema. CT scans were reviewed independently by two reviewers and final interpretation was obtained by consensus. Results of thin-section CT scans were correlated with results of pulmonary function tests, including FEV1 and FEV1/FVC to assess air-way obstruction, total lung capacity and residual volume to assess air trapping, and diffusing capacity for carbon monoxide/alveolar volume (DCO/VA). RESULTS: CT evidence of emphysema, which was noted in 45% of the patients (n = 41), was mainly localized in the same bronchopulmonary segments as bronchiectasis. The presence of emphysema was in relation to the extent and to the severity of bronchiectasis. Only eight patients with CT evidence of emphysema had functional evidence of emphysema (20%). When comparing the group of patients with CT evidence of emphysema with the group of patients with no CT evidence of emphysema, the group of patients with CT evidence of emphysema had significantly higher airflow obstruction and air trapping, had significantly lesser value of diffusing capacity, but with no decreased gas transfer (DCO/VA > 80%). CONCLUSION: Our series suggests that there is a high prevalence of emphysema in patients with bronchiectasis. Emphysema that was not suggested using pulmonary function tests in most of the cases could explain in part the higher airway obstruction observed in the group of patients with CT evidence of emphysema. This study could support the suggestive notion that emphysema, which was mainly localized in bronchiectatic lobes, could be due to the inflammatory airway process.