Age-induced changes in cellular membranes of soybean seed axes. -Physiol. Plant. 73: 85-91. The physical and chemical properties of microsomal membranes and cellular antioxidant systems were investigated in imbibed soybean (Glycine m u L. Merr. cv.Maple Arrow) seeds following aging for 5 years at room temperature. The loss of germination capacity in aged seeds was associated with increased solute leakage during imbibition and with a loss of membrane phospholipid. Higher levels of free fatty acids were observed in the microsomal membranes from aged seeds. However, there was no change in fatty acid saturation. Wide angle X-ray diffraction studies indicated the presence of gel phase in addition to liquid-crystalline phase lipid domains in the membranes of aged seeds. Those from fresh seeds were exclusively liquid-crystalline. Fluorescence depolarization, using diphenylhexatriene. suggested that the microviscosity of the membrane bilayer was increased by aging. Aged seeds had a lower antioxidant potential in the lipid fraction, lower tocopherol content, and reduced ascorbate:dehydroxyascorbate ratio indicating that the aging process was associated with exposure to an oxidative stress.

Increasing evidence exists for the role that cattle play in the epidemiology of campylobacteriosis. In this study, the prevalence and distribution of Campylobacter jejuni were longitudinally examined at the subspecies level in the beef cattle production continuum. Animals were subdivided into two groups: those that were not administered antibiotics and those that were administered the antimicrobial growth promoter chlortetracycline and sulfamethazine (AS700). Samples were longitudinally collected throughout the confined feeding operation (CFO) period and during the slaughter process, and C. jejuni was isolated and genotyped to assess subtype richness and to elucidate transmission dynamics from farm to fork. The bacterium was frequently isolated from cattle, and the bacterial densities shed in feces increased over the CFO period. Campylobacter jejuni was also isolated from digesta, hides, the abattoir environment, and carcasses. The administration of AS700 did not conspicuously reduce the C. jejuni densities in feces or within the intestine but significantly reduced the bacterial densities and the diversity of subtypes on abattoir samples. All cattle carried multiple subtypes, including clinically relevant subtypes known to represent a risk to human health. Instances of intra-animal longitudinal transmission were observed. Although clinically relevant subtypes were transmitted to carcasses via direct contact and aerosols, the bacterium could not be isolated nor could its DNA be detected in ground beef regardless of treatment. Although the evidence indicated that beef cattle represent a significant reservoir for C. jejuni, including high-risk subtypes strongly associated with the bovine host, they do not appear to represent a significant risk for direct foodborne transmission. This implicates alternate routes of human transmission. IMPORTANCE Limited information is available on the transmission of Campylobacter jejuni subtypes in the beef production continuum and the foodborne risk posed to humans. Cattle were colonized by diverse subtypes of C. jejuni, and the densities of the bacterium shed in feces increased during the confined feeding period. Campylobacter jejuni was readily associated with the digesta, feces, and hides of cattle entering the abattoir, as well as the local environment. Moreover, C. jejuni cells were deposited on carcasses via direct contact and aerosols, but the bacterium was not detected in the ground beef generated from contaminated carcasses. We conclude that C. jejuni bacterial cells associated with beef cattle do not represent a significant risk through food consumption and suggest that clinically relevant subtypes are transmitted through alternate routes of exposure.

The impacts of the antimicrobial growth promoter (AGP), chlortetracycline with sulfamethazine (AS700), on the development of antimicrobial resistance and longitudinal transmission of Campylobacter jejuni within the beef production continuum were empirically determined. Carriage of tetracycline resistance determinants in the enteric bacterial community increased at a greater rate for AS700-treatment cattle. The majority of the bacteria from animals administered AS700 carried tetW. Densities of C. jejuni shed in feces increased over the confined feeding period, and the administration of AS700 did not conspicuously reduce C. jejuni densities in feces or within the intestine. The majority of C. jejuni isolates recovered were resistant to tetracycline, but the resistance rates to other antibiotics was low (≤20.1%). The richness of C. jejuni subtypes recovered from AS700-treated animals that were either resistant or susceptible to tetracycline was reduced, indicating selection pressure due to AGP administration. Moreover, a degree of subtype-specific resistance to tetracycline was observed. tetO was the primary tetracycline resistance determinant conferring resistance in C. jejuni isolates recovered from cattle and people. Clinically-relevant C. jejuni subtypes (subtypes that represent a risk to human health) that were resistant to tetracycline were isolated from cattle feces, digesta, hides, the abattoir environment, and carcasses, but not from ground beef. Thus, study findings indicate that clinically-relevant C. jejuni subtypes associated with beef cattle, including those resistant to antibiotics, do not represent a significant foodborne risk.

Numerous chromomsome-length polymorphisms (CLPs) associated with chromosome IV were detected in an inbred line of race 8 of Ustilago hordei (teliospore line 1279). Polymorphisms for chromosome IV were observed in the 1600-1900-kb range in approximately 8% of the haploid sporidia originating from this teliospore collection. A monosporidial strain, 1279Ca2, exhibited a new 1620-kb chromosome band and a concurrent loss of the 1950-kb chromosome IV. A ribosomal DNA probe from Armillaria mellea specifically hybridized to both the variant 1620-kb chromosome and to the 1950-kb rDNA chromosome IV in parental strains. Following digestion of chromosome IV with HinfI, the telomere fragments of the 1620-kb chromosome were similar to those of the 1950-kb chromosome IV, indicating that the 1620-kb chromosome arose following a deletion of approximately 330 kb from chromosome IV. The Hinf1 digest of chromosome IV, when probed with the rDNA probe, revealed that much of the rDNA of chromosome IV was lost in the 1279Ca2 line. rDNA sequences were coincidentally recorded in chromosomes I and II in the sister sporidial line, 1279Ca4. When the 1279Ca2 line was mated to other members within the same tetrad and inoculated onto susceptible barley, karyotypes of tetrads and random sporidia originating from the F1 progeny possessed variant chromosomes ranging in size from 1536 to 2110 kb. Among sporidia in 5 of the 12 ordered tetrads sampled, a 1:1 ratio of the 1950-kb parental chromosome IV to the variant chromosome was observed. Within these tetrads, the two polymorphic chromosomes were identical in size and larger than that of the original variant 1620-kb chromosome suggesting that a chromosome expansion, averaging 150 kb, had occurred. In 5 of the 12 tetrads, a 1:1:1:1 ratio representing the two original parents and two recombinant chromosomes was observed, suggesting that normal or unequal recombination had occurred during meiosis. In 2 of the 12 F1 progeny tetrads, the 1950-kb chromosome IVs were apparently eliminated. In karyotypes of these sporidial lines, we observed rDNA sequences in chromosomes I and III that were translocated from chromosome IV. Among 78 random sporidia in the F1 generation, duplication of the variant chromosome IV was observed in three strains. These results suggest that polymorphisms in the rDNA chromosome IV, which consist of chromosome expansion, translocation, and chromosome elimination or duplication, are common in the 1279 strain of U. hordei and its progeny, and that this variability appears to be associated with the tandem-repeat nature of the rDNA sequences.

The objectives of this study were the identification of genes that show relatively strong levels of expression in the rumen protozoan, Isotricha intestinalis, and the demonstration that promoters from such genes can be used in the construction of recombinant expression vectors. In order to identify highly expressed genes, a cDNA library was constructed for I. intestinalis, and RNA expression analysis conducted on 62 clones using a filter array hybridization assay. Expression levels for individual clones ranged from easily detectable to below the detection threshold of the technique. Eleven cDNAs showed relatively intense hybridization signals, and the gene for one of these clones, I87, was characterized in detail. The ability of the I87 promoter to drive the expression of recombinant genes was tested by linking it to the luciferase reporter gene in a yeast shuttle vector and transforming Saccharomyces cerevisiae cells for expression analysis. The results showed that a rumen protozoal gene promoter is capable of directing the expression of a reporter gene in S. cerevisiae.