Serological and Molecular Characterization of a High Temperature-recovered Virus Belonging to Tospovirus Serogroup IV

A serologically and cytologically distinct gloxinia tospovirus (HT-1) previously isolated from a gloxinia plant infected with Impatiens necrotic spot virus (INSV) when propagated in a high-temperature environment was characterized. Rabbit antisera produced for INSV and Tomato spotted wilt virus (TSWV) nucleocapsids (N) failed to react with HT-1 proteins in western blot analysis. The HT-1 antibodies reacted strongly with homologous antigen but failed to react with INSV and TSWV. However, the HT-1 antiserum reacted in ELISA with Watermelon silver mottle virus (WSMV) from Taiwan and in western blot analysis with the WSMV N protein. A reciprocal test showed that the antiserum prepared against the N protein of WSMV also reacted with the HT-1 N protein in both ELISA and western blot analysis. DNA probes derived from the N gene of HT-1 or WSMV hybridized to RNAs prepared from plants infected with either virus. Stronger signals were obtained with homologous than with heterologous reactions. Neither probe detected INSV or TSWV. The M and S RNAs of HT-1 were sequenced. The M RNA contains two open reading frames (ORF) ; one in the sense orientation encoding a nonstructural (NSm) protein of 308-amino-acids (aa) and the other in the ambisense orientation, a 1122-aa precursor of Gl and G2 glycoproteins. The S RNA also contains two ORFs ; one in the sense orientation encoding a nonstructural (NSs) protein of 439 aa and the other in the ambisense orientation, an N protein of 277 aa. HT-1 is distantly related to INSV and TSWV as shown by low nucleotide (40–52%) and amino acid (28–48%) similarities in the four ORF sequences. The HT-1 virus shares high nucleotide (76–81%) and amino acid (85–92%) similarities with WSMV and peanut bud necrosis virus (PBNV). Based on the serological properties and sequence data, we propose that HT-1 is a distinct species of serogroup IV in the genus Tospovirus. This is the first time that a tospovirus similar to those found in the Far East and in Southeast Asia has been identified in the US. Received 16 October 1999/ Accepted in revised form 20 December 1999     

Evaluation of a novel enzyme-linked immunosorbent assay for detection of antibodies against Salmonella, employing a stable coating of lipopolysaccharide-derived antigens covalently attached to polystyrene microwells.

Polysaccharides derived from Salmonella typhimurium lipopolysaccharide (LPS) representing the O-antigen factors 1, 4, 5, and 12 and the O-antigen factors 6 and 7 from Salmonella choleraesuis LPS were derivatized with the photoreactive compound anthraquinone and subsequently covalently coupled to microtiter polystyrene plates by ultraviolet irradiation. Both polysaccharide antigens could be coupled simultaneously to the same microtiter plate. The coated surface was used in indirect ELISA for the determination of serum antibodies from pigs infected with bacteria of the two Salmonella groups and from uninfected pigs. This ELISA proved itself by having a good long-term durability and a high degree of reproducibility, including low day-to-day variations and low interplate variations. Furthermore, the ELISA showed good specificity and sensitivity when data were compared with the optical density levels of a panel of pig sera as determined by a conventional ELISA on the basis of passive coating of the two Salmonella LPS antigens (the mix-ELISA). The covalent anthraquinone mix-ELISA shows promise as a stable and durable alternative to the existing conventional ELISA for serological surveillance of Salmonella infections in pigs.     

NIR Transmittance Estimation of Free Lipid Content and Its Glycolipid and Digalactosyldiglyceride Contents Using Wheat Flour Lipid Extracts

Free lipids (FL) in 72 hard winter wheat flours were extracted and dissolved in hexane. Absorbance (log 1/T) values were measured using a scanning spectrophotometer with a 2‐mm cuvette, and used to develop calibration models for estimating flour FL content and glycolipid (GL) and digalactosyldiglyceride (DGDG) contents. Fifty‐one calibration samples were selected based on the cutoff point of 0.3 of Mahalanobis distance, and the remaining twenty‐one samples were used for validation. The best model for the estimation of FL content showed coefficients of determination (R²) of 0.95 for the calibration set and R² = 0.89 for the validation set. Glycolipid contents could be estimated by a model which had R² = 0.87 for the calibration set and R² = 0.89 for the validation set. For DGDG, the best model showed R² = 0.94 for the calibration set and R² = 0.88 for the validation set.     

Prediction of Baking Characteristics of Hard Winter Wheat Flours Using Computer‐Analyzed Mixograph Parameters

The objective of this research was to determine whether computer‐analyzed (objective) mixograph parameters could replace conventional mixograph parameters in the evaluation of flour quality. The 642 hard winter wheat flours, collected from federal regional performance nurseries in 1995 and 1996, were analyzed by a conventional and computerized mixograph. Mixograph bandwidths at 6 min (BW6) showed the most significant linear correlation with subjective mixing tolerance scores (r = 0.81, P < 0.1%, n = 642). Prediction models of conventional and experimental baking parameters were developed by continuum regression using computer‐analyzed mixograph parameters of a calibration set (n = 282). The developed models could estimate conventional mixograph mixing time and tolerance scores, baking water absorption and mixing time, and bread loaf volume, showing R² values of 0.86, 0.74, 0.68, 0.80, and 0.51, respectively, for a validation set (n = 380). These results indicated that computer‐analyzed mixograph parameters could be applied to develop prediction models to be used for flour quality evaluation in wheat breeding programs.     

The growth of swine bone marrow cells in the presence of heterologous colony stimulating factor: Characterization of the developing cell population

Conditioned medium prepared from the mouse fibroblast cell line L 929 which is known to produce colony-stimulating factor active on mouse bone marrow cells was also able to stimulate the growth of swine bone marrow cells in a liquid culture system. During the first 4 days of culture mononuclear phagocyte and granulocyte colonies developed. Prolonged cultured cells were classified belonging to the macrophage lineage by morphological and cytochemical criteria. These cells fulfill also functional characteristics for macrophages, like expression of Fc receptors, immune phagocytosis and production of prostaglandins. These bone marrow-derived macrophages could also be activated with LPS and lymphocyte-derived mediators.     

Inactivation of fenitrothion on stored maize at different moisture contents

Biological activity of fenitrothion on stored maize at various moisture contents and at different times after application was measured by biological assay using adults of Tribolium castaneum (Herbst). Inactivation of actual residues over time was then determined after making the necessary allowance for chemical breakdown. At a given moisture content, the inactivation process was substantially completed during the first 6 weeks after application and loss of effectiveness from 6 weeks onwards resulted mainly from chemical breakdown. At a given time after application, residues were less active at higher moisture content (m.c). Differences in activity between moisture contents were apparent within a few hours of application and continued to increase for up to 3 days, with relatively little change thereafter during storage of 24 weeks. Thus after 24 weeks, residues on maize of 18% m.c. had an activity about 20% that of similarly-aged residues at 10% m.c. and 4% that of freshly-applied residues at 10% m.c. These results were in general accord with changes in the proportion of the residue which was collected from the kernels by a surface wash with methanol, this readily-extractable residue presumably representing the insecticide that may be picked up by insects.     

On the origin of bacterial resistance to penicillin: comparison of a beta-lactamase and a penicillin target

Structural data are now available for comparing a penicillin target enzyme, the D-alanyl-D-alanine-peptidase from Streptomyces R61, with a penicillin-hydrolyzing enzyme, the beta-lactamase from Bacillus licheniformis 749/C. Although the two enzymes have distinct catalytic properties and lack relatedness in their overall amino acid sequences except near the active-site serine, the significant similarity found by x-ray crystallography in the spatial arrangement of the elements of secondary structure provides strong support for earlier hypotheses that beta-lactamases arose from penicillin-sensitive D-alanyl-D-alanine-peptidases involved in bacterial wall peptidoglycan metabolism.