The deoxyribonucleic acid (DNA) of Streptococcus lactis C2, S. cremoris B1, and S. diacetilactis 18-16 was labeled by growing cells in Trypticase soy broth containing 3H-labeled thymine. The cells were gently lysed with lysozyme, ethylenediaminetetraacetic acid, and sodium lauryl sulfate. The chromosomal DNA was separated from plasmid DNA by precipitation with 1.0 M sodium chloride. The existence of covalently closed circular DNA in the three organisms was shown by cesium chloride-ethidium bromide equilibrium density gradient centrifugation of the cleared lysate material. In an attempt to correlate the loss of lactose metabolism with the loss of plasmid DNA, lactose-negative mutants of these organisms were examined for the presence of extrachromosomal particles. Covalently closed circular DNA was detected in the lactose-negative mutants of S. lactis C2 and S. diacetilactis 18-16. In S. cremoris B1, however, no covalently closed circular DNA was observed by using cesium chloride-ethidium bromide gradients. Electron micrographs of the satellite band material from S. lactis C2 and its lactose-negative mutant confirmed the presence of plasmid DNA. Three distinct plasmids having approximate molecular weights of 1.3 x 106, 2.1 x 106, and 5.1 x 10' were observed in both organisms.

In a series of experiments we investigated effects of several factors on intake of urea by lactating dairy cows. Cows given an unfamiliar ingredient or altered moisture in the ration reduced their intake, and this effect was attributed to a newness factor. Addition of urea to a ration may have a similar effect. An experimental design providing both no-choice and two-choice data was devised to minimize the effect of newness. When urea was isolated physically by pellets coated with ground corn, cows selected against urea-containing pellets on a two-choice basis and reduced intake on a no-choice basis. When the choice was between two urea-containing rations, cows preferred the pellets in which urea flavor and odor should have been most evident. Urea odor in the feedbox or urea in the drinking water did not reduce feed intake. Eating rate of a 2.5% urea-containing ration over two daily 30-min feeding periods was about one-half that of a non-urea ration. This effect was evident during the first 5-min eating interval. Administration of urea solution into the rumen prior to feeding a nonurea ration markedly reduced subsequent intake. Reticulum pH and ammonia indicated possible sublethal ammonia toxicity. Intake and eating rate were depressed and reticulum pH was elevated when cows were fed a ration with 2.5% urea versus 1% urea or nonurea rations. Elevated reticulum ammonia occurred on both 1 and 2.5% urea-containing rations. Cows not previously exposed to urea-containing rations reduced intake and eating rate when fed rations with 2.0 and 2.5% urea. Intake reduction was not comparable to that by cows preconditioned to urea rations. Upon third exposure to 2.5% urea in the ration, cows reduced and ceased intake but readily consumed a nonurea ration. Cows require preconditioning to develop a negative aversion to rations containing high urea, perhaps through a mechanism of sublethal ammonia toxicity.

Partial lactose-fermenting revertants from lactose-negative (lac-) mutants of Streptococcus lactis C2 appeared on a lawn of laccells after 3 to 5 days of incubation at 25 C. The revertants grew slowly on lactose with a growth response similar to that for cryptic cells. In contrast to lac+ S. lactis C2, the revertants were defective in the accumulation of [4C ]thiomethyl-fl-D-galactoside, indicating that they were devoid of a transport system. Hydrolysis of o-nitrophenyl-fl-D-8:30 on July 7, 2020 by guest http://jb.asm.org/ Downloaded from o-nitrophenyl-p-D-galactoside (ONPG) hydrolysis in

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Ultraviolet (UV)-induced phage lysates, from lactose-positive (lac+) Streptococcus lactis C2, transduced lactose fermenting ability to lac-recipient cells of this organism. Although the phage titer could not be determined due to the absence of an appropriate indicator strain, the number of transductants was proportional to the amount of phage lysate added. Treatment of the lysate with deoxyribonuclease had no effect on this conversion, indicating the observed genetic change was not mediated by free deoxyribonucleic acid. When the lac+ transductants were isolated and exposed to UV irradiation, lysates with higher transducing ability were obtained. The transducing ability of this lysate was about 100-fold higher than that observed in the original lysates. The lac+ transductants were unstable since lacsegregants occurred at high frequency. The phage lysate from S. lactis C2 also transduced maltose and mannose metabolism to the respective negative recipient cells. The results demonstrate the transduction of carbohydrate markers by a streptococcal phage and establish a genetic transfer system in group N streptococci.on August 1, 2020 by guest