A new site-specific endonuclease from Neisseria cinerea.
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Biomedical subjects
Publications and source records attributed to S M Martin.
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Fifteen patients with recent influenza B infection were admitted to three Dallas hospitals in the first 11 weeks of 1977. The patients' ages ranged from five to 73 years, with a median of 18 years. Most had no significant underlying disease. The spectrum of clinical illness included postinfluenzal bacterial pneumonia (three cases), other severe chest disease (two cases), hyperpyrexia and possible rhabdomyolysis in the elderly (two cases), onset of toxemia of pregnancy, thyroid dysfunction, Stevens-Johnson syndrome, neurologic disorders (two cases), and Reye's syndrome (three cases). Two patients died. Two elderly men with high fever and weakness entered the hospital within three days of illness and two of three patients with Reye's syndrome were admitted four days after the onset of influenza, but the 11 other patients gave a history of seven to 31 days of symptoms before being hospitalized. Morbidity and mortality with influenza B are neither trivial nor restricted to debilitated hosts.
Defined media are described that support 14-20 h generation times for Acanthamoeba castellanii and A. rhysodes in monolayer cultures. The media differ in minor ways from previously described media, but the growth rates are greatly improved over previously reported values. Maximum growth rates were observed for A. castellanii in a complex medium containing 21 amino acids, but near-maximum rates could be achieved in relatively simple media containing 9 amino acids. Growth occurred with 6 amino acids, as reported by others, but generation times exceeded 30 h. Amitosis was a common problem during early subcultures in defined media, defined media by glucose and acetate starvation. The rate of encystment varied with cell density at the time of starvation and was optimal at initial densities of 400-800 amebae/mm2.
Two regimens for transformation have been devised for the phage group 2 strains UT0002-19 and UT0017 of Staphylococcus aureus. Strain UT0002-19 produces exfoliative toxin, which is responsible for the clinical manifestations of the staphylococcal scalded skin syndrome, whereas strain UT0017 does not produce exfoliative toxin. A large pool of antibiotic-resistant and auxotrophic mutants from strains UT0002-19 and UT0017 were isolated by using (i) antibiotic gradient plates, (ii) trimethoprim selection, and (iii) nitrosoguanidine mutagenesis, which sometimes was coupled by enrichment with penicillin or methicillin. Transformation frequencies of genetic markers in mutant strains ranged from 10(-6) to 10(-8). Three genetic linkage groups were identified on the strain UT0017 chromosome. The first linkage group was thy-4-lys-5-trp-21-thr-4, the second was pyr-26-nov-9-his-3, and the third consisted of ilv-9 and pen-1, a genetic determinant for beta-lactamase synthesis. Two linkage groups were detected on the strain UT0002-19 CHROMOSOMe. The first was defined as thy-1-lys-5-trp-3-thr-4-ala-8, whereas the second consisted of nov-9 and his-3 markers. A chromosomal locus that controlled synthesis of exfoliative toxin could not be mapped.
A large pool of antibiotic resistant and auxotrophic mutants was isolated from the Staphylococcus aureus phage group 2 strains UT0002-19 and UT0017 by (1) antibiotic gradient plates, (2) trimethoprim selection, and (3) nitrosoguanidine mutagenesis, which sometimes was coupled by enrichment with either penicillin or methicillin. Strain UT0002-19 has a chromosomal determinant for exfoliative toxin (ET), which causes "scalded skin syndrome" in man. A few mutants were isolated from the phage group 1 strain UT0080, which also produces ET. Two transformation regimens, called the broth and plate methods, were devised for the phage group 2 strains. They employed 80 alpha as helper phage, and recipient cells were incubated with transforming DNA in the presence of Ca2+. Strain UT0080 was transformed using phage 55 as helper. Maximum competence of the phage group 2 strains occurred during early logarithmic growth in trypticase soy broth, but cells grown overnight on heart infusion agar were also competent. Transformation frequencies of all markers ranged from 10(-6) to 10(-8). For phage 80 alpha, a multiplicity of infection of 4 was optimal in transforming a mutant of strain UT0002-19. Transformation of gly, lin, met, ole, rif, and ser markers in S. aureus is reported for the first time. Ery and ole markers in all three strains exhibited cross-resistance. Mapping studies, similar to those performed by DNA-mediated transformation in the phage group 3 strain 8325, can now be commenced for phage group 2 strains of S. aureus in order to elucidate the molecular genetics of this medically important bacterium.
An examination of conditions for the growth of Desulfovibrio desulfuricans, with the aim of optimizing hydrogenase production, is reported. An ammonium sulfate-lactate-yeast extract medium gave 5 to 10 times as much hydrogenase activity as a peptone-yeast extract medium. It made little if any difference whether the gas used for sparging was nitrogen, hydrogen, or a mixture thereof but increasing the rate of sparging and agitation did result in a slight decrease in activity. Control of pH during culture development was of little benefit to hydrogenase production. At least two hydrogenases were present in D. desulfuricans: one periplasmic, the other membrane bound.
The periplasmic hydrogenase of Desulfovibrio desulfuricans was isolated and purified. Cells were washed with Tris-EDTA and the enzyme precipitated from the wash with ammonium sulfate. Absorption chromatography on DEAE and hydroxyapatite yielded the enzyme at better than 95% purity as judged by gel electrophoresis. The hydrogenase catalyzed the production of more than 9000 mumol H2/min mg protein(-1) from reduced methyl viologen at 37 degrees C. It is very stable and resists inactivation by heat (50% activity remained after 5 min in air at 65 degrees C) and by enzyme inhibitors (except N-ethylmaleimide and potassium ferricyanide). After storage in air at 4 degrees C for 1 month no activity was lost. The enzyme activity is sensitive to ionic environmental changes. With methyl viologen the optimum pH was 5.5 but with p-xylene polymeric viologen the optimum was about pH 7 but less sharp. The molecular weight was 47 X 10(3)(+/- 2 X 10(3), 52 X 10(3)(+/- X 10(3), and 56 X 19(3)(+/- 2 X 10(3) by SDS-gel electrophoresis, gel chromatography, and sedimentation equilibrium, respectively, and the isoelectric point was at pH 6.0. They enzyme might be useful in the production of hydrogen from water and solar energy.
A mesophilic anaerobe, Acetivibrio cellulolyticus, isolated from a cellulose-enrichment culture, degraded cellulose by the secretion of cellulolytic enzymes into the culture medium. Both exo- and endoglucanase activities were detected and shown to be regulated by induction and catabolite repression. Endoglucanase synthesis was induced by cellulose, cellobiose and salicin. Synthesis induced during growth on cellobiose was inhibited, although not completely repressed by glucose. Exoglucanase activity was enhanced when cells were grown on insoluble or soluble poly beta-glucoside polymers. Activity was progressively inhibited by supplementing microcrystalline substrates with increasing amounts of cellobiose.
A review of the medical records of 123 persons with Legionnaires' disease hospitalized in the 1976 Philadelphia epidemic showed that the manifestations of infection ranged from mild grippe to a severe pneumonia that also involved other organ systems. Early in the illness, constitutional symptoms predominated. Fever, malaise, myalgia, rigors, confusion, headache, and diarrhea were usually followed by nonproductive cough and dyspnea. Physical examination showed few abnormalities other than rales. Moderate leukocytosis with left shift, elevated erythrocyte sedimentation rate, elevation of serum levels of liver enzymes, and hematuria and proteinuria were characteristic. Chest radiograph showed patchy, often nodular, areas of consolidation. Progression of pneumonia led to respiratory failure and the need for mechanical ventilatory assistance for 19 patients; renal failure, primarily after shock, occurred in 18 persons. Twenty-six patients died. Treatment with erythromycin or tetracycline resulted in the lowest case-fatality ratios, but the associations were not statistically significant.
Since January, 1970, the Center for Disease Control (CDC) has corridnated surveillance of nosocomial infections in a group of voluntarily cooperating hispitals in the United States. In 1970, this surveillance system failed to realize one of its major goals: detection of a nationwide epidemic of septicemia caused by contaminated intravenous products. However, retrospective review of infections reported to CDC revealed that the data received were sufficient for the outbreak to have been recognized. Beginning in July, 1970, one month after the contaminated products were first distributed and five months before the outbreak was actually detected. CDC data showed a persistent increase in the incidence of Enterobacter and Erwinia (presently designated Enterobacter agglomerans) bacteremia. Furthermore, monthly rates of cases of bacteremia caused by these organisms were higher in hospitals using the contaminated intravenous products than for hospitals not using them. Failure to detect this outbreak at the time of its occurrence was due to delays in data processing and insufficiently sophisticated data analysis. Based on this experience, CDC has modified the surveillance system to aid recognition of future outbreaks.
An explosive, common-source outbreak of pneumonia caused by a previously unrecognized bacterium affected primarily persons attending an American Legion convention in Philadelphia in July, 1976. Twenty-nine of 182 cases were fatal. Spread of the bacterium appeared to be air borne. The source of the bacterium was not found, but epidemiologic analysis suggested that exposure may have occurred in the lobby of the headquarters hotel or in the area immediately surrounding the hotel. Person-to-person spread seemed not to have occurred. Many hotel employees appeared to be immune, suggesting that the agent may have been present in the vicinity, perhaps intermittently, for two or more years.
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In September 1974, the largest outbreak of foodborne salmonellosis ever reported to the Center for Disease Control--affecting an estimated 3,400 persons--occurred on the Navajo Nation Indian Reservation. The responsible agent was Salmonella newport and the vehicle of transmission was potato salad served to an estimated 11,000 persons at a free barbecue. The cooked ingredients of the potato salad had been stored for up to 16 hours at improper holding temperatures. The magnitude of the outbreak allowed us to study secondary transmission by calculating the rates of diarrheal illness during the 2 weeks following the outbreak in persons who did not attend the barbecue and by examining the results of stool cultures obtained after the outbreak. We found no secondary transmission. We conclude that a health official should monitor food preparation and service at large social gatherings and that person-to-person transmission of salmonellosis probably does not normally occur even in settings considered highly conductive to cross-infection.
The activity and sedimentation of acid phosphatase (APase), acid deoxyribonuclease (DNase), and acid ribonuclease (RNase) were investigated throughout growth and encystment in Acanthamoeba castellanii. The activities/mg protein of all 3 hydrolases are high in young cultures and decrease to constant levels in postlog cells. The RNase activity/ameba decreases 50% during growth, whereas the activity/cell of both APase and DNase remains constant. The percent sedimentation at 20,000 g of all 3 enzymes gradually increases from about 40% in midlog to a plateau of 80% in postlog cells. During encystment, the sedimentation behavior of RNase differs from that of APase and DNase. Encystment is characterized by a differential decrease in the activity/cell of the 3 hydrolases, with RNase decreasing most rapidly and APase least rapidly. APase is unique in that a transient increase of its specific activity is noted during encystment, even though its activity/cell is decreasing.
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