Regulation of biotechnology.
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Biomedical subjects
Publications and source records attributed to A K Vidaver.
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Pseudomonas cepacia strains of plant and clinical origin were compared with the type strains of P. cepacia, P. kingii and P. multivorans. Conventional biochemical tests and antibiotic sensitivity patterns supported the previous proposals of synonymy between P. cepacia, P. kingii and P. multivorans. However, bacteriocin production patterns, onion maceration tests and hydrolysis of low pH pectate agar clearly differentiated strains of clinical and plant origin into two distinct groups; these tests may therefore be helpful in epidemiological studies. In contrast, plant and clinical strains were of equal lethality to mice. Agarose gel electrophoresis indicated the presence of one or more plasmids (molecular weights 9 X 10(6) to 120 X 10(6)) in 15 out of 16 strains of both types examined.
The majority (85% of all strains tested) of 12 phytopathogenic Corynebacterium species produced bacteriocin(s) on nutrient broth--yeast extract (NBY) medium. All C. nebraskense, C. michiganense, C. insidiosum, C. oortii, and C. iranicum strains produced bacteriocin(s). The optimal conditions for production of 23 distinct bacteriocins by eight species of Corynebacterium generally were 20 degrees C and 4 days of incubation on NBY or on modified Burkholder's agar that lacked peptone (MBAL). Production in liquid was marginal and not augmented by adding mitomycin C. Bacteriocins generally had little effect on other strains within a species but were inhibitory to other species. Most bacteriocins appeared to be bactericidal proteins resistant to heat (75 to 80 degrees C, 30 min) but sensitive to proteolytic enzymes. Some strains of C. nebraskense, C. michiganense, C. insidiosum, and C. flaccumfaciens produced two bacteriocins which were clearly differentiated by varying or testing one or more of the following: conditions for production, the indicator, heat stability, and susceptibility to proteolysis. Within certain limitations, a convenient and reproducible typing scheme was devised for strain and species differentiation of most phytopathogenic corynebacteria.
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Root segments and root-soil cores (6.5-cm diameter) from fields and nurseries of winter wheat and sorghum were tested for N2 fixation by using the acetylene reduction assay. Wheat samples (approximately 1,200) from 109 sites generally had low or no activity (0 to 3.1 nmol of C2H4 produced per h per g [dry weight] of root segments), even after 24 h of incubation. However, a commercial field of Scout 66, located in western Nebraska, exhibited appreciable activity (290 nmol of C2H4 produced per h per g [dry weight] of root segments). Of 400 sorghum lines and crosses, grain sorghums (i.e., CK-60A, Wheatland A, B517, and NP-16) generally exhibited higher nitrogenase activity than forage sorghums or winter wheats. CK-60A, a male sterile grain sorghum, was sampled at four locations and had the most consistent activity of 24 to 1,100 nmol of C2H4 produced per h per core. The maximum rate extrapolated to 2.5 g of N per hectare per day. Numerous N2-fixing bacterial isolates were obtained from wheat and sorghum roots that exhibited high nitrogenase activity. Most isolates were members of the Enterobacteriacae, i.e., Klebsiella pneumoniae, Enterobacter cloacae, and Erwinia herbicola.
Bacteriocin-like substances were commonly produced by slow-growing Rhizobium japonicum and cowpea rhizobia on an L-arabinose medium. Antagonism between strains of R. japonicum was not detected in vitro; however, such strains were often sensitive to some bacteriocins produced by cowpea rhizobia. Inhibitory zones (2 to 8 mm from colony margins), produced by 58 of 66 R. japonicum test strains, were reproducibly detected with Corynebacterium nebraskense as an indicator. Quantitative production was not related to symbiotic properties of effective strains, since nine noninfective strains and one ineffective strain produced bacteriocin. Eight R. japonicum strains that did not produce bacteriocin nevertheless formed effective nodules on soybeans. R. japonicum strains that produced bacteriocin in vitro had no antagonistic effect on nonproducer strains during soybean nodulation. Under controlled conditions, a nonproducer (3I1b135) predominated over a bacteriocin producer (3I1b6) when inoculated at 1:1 and 1:9 ratios. Depending on the particular ratio, up to 38% of the total nodules formed were infected with mixed combinations. The bacteriocin(s) had a restricted host range and antibiotic-like properties which included the ability to be dialyzed and resistance to heat (75 to 80 degrees C, 30 min), Pronase, proteinase K, trypsin, ribonuclease, and deoxyribonuclease. R. japonicum strains representing genetic, serological, cultural, and geographic diversity were differentiated into three groups on the basis of bacteriocin production.
Several phytopathogenic fluorescent Pseudomonas species, primarily P. syringae, could be grouped into 16 bacteriocin producer groups, including new and previously described groups. At least 86% of the P. syringae strains could be typed by bacteriocin production. There was poor correlation between bacteriocin type and host plant origin. No correlation was detected between syringomycin (a phytotoxin) production and bacteriocin type.
Pseudomonas phaseolicola cells synchronously infected with phi 6 were fixed and embedded by several procedures. The diameter of phi 6 measured 75 nm and its nucleocapsid 60 nm. Virus nucleocapsids were icosahedral and surrounded by a double membrane. In planar sections the nucleic acid appeared as a hexagonal ring and in cross section as two angular structures.
Uracil was incorporated into all three bacteriophage phi6 dsRNA segments throughout the infection cycle; the rates of incorporation into each of the three segments were approx. constant for the first 15 to 20 min and then increased rapidly until 50 min after infection. The medium and small dsRNA segments were produced in greater amounts than the large dsRNA segment at all times in the infection cycle. Inhibition of host RNA and protein synthesis with rifampin and chloramphenicol revealed that virus dsRNA synthesis immediately after infection was independent of either host function.
Pseudomonas phaseolicola infected with bacteriophage phi 6 synthesized all three viral double-stranded RNA segments, three single-stranded RNAs, and three replicative intermediate-like RNAs in the presence of rifampin. The single-stranded RNA intermediates sedimented and electrophoresed along with melted viral double-stranded RNA, annealed with melted viral double-stranded RNA, and were transient in nature. The relative amounts of the single-stranded RNA intermediates varied during the infection cycle and were altered in the presence of chloramphenicol. The replicative intermediate-like RNAs sedimented faster than double-stranded RNA, failed to enter 2.5% polyacrylamide gels, eluted with double-stranded RNA from a CF-11 cellulose column, were precipitated with single-stranded RNA in 2 M LiC1, and yielded three genome-size pieces of double-stranded RNA upon digestion with RNase. These results are consistent with the hypothesis that complementary strands of the phi 6 double-stranded RNAs are synthesized asynchronously during the infection cycle.