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Confirmation of the inhibitory effect of cystic fibrosis sera on oyster cilia
508
September, 1971 The Journal o[ P E D I A T R I C S
Letters to the Editor
Fluid intake, renal solute load, and ~vater balance in infancy To the Editor: The recent article by Ziegler and Fomon, "Fluid intake, renal solute load, and water balance in infancy" (J. PEDIAT. 78: 561, 1971), certainly clarified many aspects of infant feeding. The authors state that "formulas should not be calorically concentrated above 100 Kcal per 100 ml. (30 Kcal per ounce). At this level of concentration, it becomes important to avoid diets that present high renal solute loads: Protein should account for no more that 10 per cent of caloric i n t a k e . . . " It should be remembered that when adding carbohydrate to any formula to increase the calories, one is also diluting the protein and the electrolytes. This will not present a problem in nutrition unless the protein is diluted to the extent that it represents less than 5 or 6 per cent of the total caloric intake. If this occurs, marasmic kwashiorkor may develop. Therefore, it is not always the total amount of calories which is important but the percentage distribution of these calories between protein, fat, and carbohydrates.
Gilbert Martin, M.D. Loring Air Force Base Hospital Limestone, Maine This cautionary observation is appreciated. "r
E. N.
Ulcerative ileitis in infancy To the Editor: With respect to the recent Clinical-Pathological Conference, Chronic idiopathic ulcerative ileitis in infancy (J. PeoraT. 78: 538, 1971), I would like to suggest that two important aspects were not discussed. First, the pathologist describes as incidental the autopsy findings of bilateral hydroureter, hydronephrosis, and dilation of the bladder. There is no record of a microscopic urinalysis or of urine cultures so that the diagnosis of urinary
Vol. 79, No. 3, pp. 508-512
tract infection is not eliminated and it could certainly cause vomiting and diarrhea. Second, the patient had received some dietary manipulation before being admitted to Johns Hopkins Hospital and received more dietary manipulation at the latter hospital. It seems to me academic to try, in a severely malnourished child, to go through the gamut of elimination diets. It would seem more to the point to have started this child immediately on intravenous feeding with lipids, amino acids, and carbohydrates. The fact that intravenous alimentation was not discussed was a major omission in my opinion.
Michael S. Brown, M.D. The Montreal Children's Hospital 2300 Tupper St. Montreal 108, Quebec, Canada
Confirmation of the inhibitory effect of cystic fibrosis sera on oyster cilia* To the Editor: The effect of serum on cilia systems has been used by several investigators to identify a specific factor (so called CF factor) in patients with cystic fibrosis. Spock and associates1 described a CF serum factor responsible for a differential effect on ciliary activity in explants of rabbit tracheal mucosa. Later, others 2-5 attempted simpler bioassay methods, using the gills from the freshwater mussel or the oyster to study ciliary inhibition. Findings presented in these studies have been contradictory; therefore, we tested the effect of CF serum on the ciliary activity of oyster gills. The oysters (Crassostrea virginica) were obtained from the Atlantic coastal areas of New Jersey and Chesapeake Bay during the months of May through July, 1969. After this time, the ~Supported by a grant from The National Cystic Fibrosis Research Foundation.
Volume 79 Number 3
accumulation of large amounts of mucus at the borders of the cilia interfered with ciliary motility and terminated this phase of the experiment. The oyster gill preparations were suspended in filtered seawater in the wells of culture slides which were sealed with a microcover glass. In addition to using the cessation of the ciliary beat as an end point, as described by Bowman and associates, 3 ,we also measured the time of appearance of expelled debris for the same purpose. Serum samples were obtained from 17 children with cystic fibrosis and from 19 children who did not have pulmonary disease; sera were tested fresh. With 16 of the 17 CF sera, debris appeared in less than an hour; with 12 of them there was cessation of ciliary beat in less than an hour. Only 3 of the 19 control sera were responsible for the appearance of any debris; this also occurred in less than an hour. Four of the control sera interfered with the ciliary beat in less than an hour. Using a x 2 test procedure, the results with the CF sera were significantly different from those with the control sera in respect to cessation of beat (0.05 > P > 0.025) and time required :For the appearance of debris (P < 0.005). Our findings tend to confirm the observations of Bowman and associates 3 that CF sera do have a ciliostatic effect which is different from that of normal sera. The time required for the appearance of debris is a more definitive end point than ~s the cessation of ciliary beat. There are, however, a number of variables inherent in the method which are responsible for considerable overlap of results. Until standardized, these variables limit the usefulness of the test in diagnosis, in genetic counseling, and in research.
Victor F. Iacocca, Ph.D. LeRoy I. Braddock, Ph.D. Giulio ]. Barbero, M.D. Department o[ Pediatrics Hahnemann Medical College and Hospital Philadelphia, Pa. REFERENCES 1. Spook, A., Heich, H. M. C., Cress, H., and Logan, W. S.: Abnormal serum factor in patients with cystic fibrosis of the pancreas, Pediat. Res. 1: I73, 1969. 2. Patrick, A. D.: In discussion, Cystic Fibrosis, Porter, R., and O'Connor, M., editors, CIBA, Foundation Study Group No. 32, London, 1968, Churchill, p. 14. 3. Bowman, B. H., Lochhart, Z. H., and MeCombs, M. L.: Oyster ciliary inhibition by cystic fibrosis factor, Science 164: 325, 1969.
Letters to the Editor
509
4. Crawford, M. d'A.: Experience of the cilia test for a factor in the serum of patients and carriers of cystic fibrosis, Proceedings of the Fifth International Cystic Fibrosis Conference, David Lawson, editor, London, 1969, p. 42. 5. Christensen, J., and Morrevang, A.: Studies on the effect of CF serum on fresh tissue from Dreissena gills, oyster gills and rabbit tracheal mucosa, Proceedings of the Fifth In ternational Cystic Fibrosis Conference, David Lawson, editor, Loncton, 1969, p. 49.
Unusual skin reation to sterile diluent in tuberculin test To the Editor: Recently, at Sinai Hospital in Baltimore, a patient in the pediatric outpatient department was observed to have a significant skin test response to the sterile diluent which is used in preparing the intermedlate-strength tuberculin test. An intradermal tuberculin test was administered to an 11-year-old Negro boy. Though controis using diluent are not standard practice at Sinai Hospital, several unconfirmed reports of equlvocaI skin test reactions (i.e., indurations of 5 to 9 mm.) had been noted two weeks before the patient was seen, and it was for this reason that 0.1 e.c. of sterile, buffered diluent was applied. Forty-eight hours later both sites showed 5 mm. of induration and erythema. A repeat application of diluent produced the same result after 48 hours, while a Tine test was negative, also at 48 hours. All injections were made using disposable plastic syringes. The patient had no known history of allergic diathesis. The phenomenon is not unknown, since Edwards and associates, 1 in testing PPD-S against pure diluent, found 2 out of 79 persons who responded measurably (2 to 4 mm.) to the diluent. The PPD preparation and diluent were returned to the manufacturer (Merck Sharp & Dohme) for analysis, and no indication of impurity or contamination was found. Sterile buffered diluent contains phosphates of sodium and potassium, as well as phenol (0.5 per cent) as a preservative. The inorganic phosphates are unlikely sensitizing agents, but Goodman and Gilman's2 text states that phenol can be markedly toxic to tissues, even unabraded skin. When directly applied to skin, a pellicle of precipitated protein may form which turns red, then brown
September, 1971 The Journal o[ P E D I A T R I C S
Letters to the Editor
Fluid intake, renal solute load, and ~vater balance in infancy To the Editor: The recent article by Ziegler and Fomon, "Fluid intake, renal solute load, and water balance in infancy" (J. PEDIAT. 78: 561, 1971), certainly clarified many aspects of infant feeding. The authors state that "formulas should not be calorically concentrated above 100 Kcal per 100 ml. (30 Kcal per ounce). At this level of concentration, it becomes important to avoid diets that present high renal solute loads: Protein should account for no more that 10 per cent of caloric i n t a k e . . . " It should be remembered that when adding carbohydrate to any formula to increase the calories, one is also diluting the protein and the electrolytes. This will not present a problem in nutrition unless the protein is diluted to the extent that it represents less than 5 or 6 per cent of the total caloric intake. If this occurs, marasmic kwashiorkor may develop. Therefore, it is not always the total amount of calories which is important but the percentage distribution of these calories between protein, fat, and carbohydrates.
Gilbert Martin, M.D. Loring Air Force Base Hospital Limestone, Maine This cautionary observation is appreciated. "r
E. N.
Ulcerative ileitis in infancy To the Editor: With respect to the recent Clinical-Pathological Conference, Chronic idiopathic ulcerative ileitis in infancy (J. PeoraT. 78: 538, 1971), I would like to suggest that two important aspects were not discussed. First, the pathologist describes as incidental the autopsy findings of bilateral hydroureter, hydronephrosis, and dilation of the bladder. There is no record of a microscopic urinalysis or of urine cultures so that the diagnosis of urinary
Vol. 79, No. 3, pp. 508-512
tract infection is not eliminated and it could certainly cause vomiting and diarrhea. Second, the patient had received some dietary manipulation before being admitted to Johns Hopkins Hospital and received more dietary manipulation at the latter hospital. It seems to me academic to try, in a severely malnourished child, to go through the gamut of elimination diets. It would seem more to the point to have started this child immediately on intravenous feeding with lipids, amino acids, and carbohydrates. The fact that intravenous alimentation was not discussed was a major omission in my opinion.
Michael S. Brown, M.D. The Montreal Children's Hospital 2300 Tupper St. Montreal 108, Quebec, Canada
Confirmation of the inhibitory effect of cystic fibrosis sera on oyster cilia* To the Editor: The effect of serum on cilia systems has been used by several investigators to identify a specific factor (so called CF factor) in patients with cystic fibrosis. Spock and associates1 described a CF serum factor responsible for a differential effect on ciliary activity in explants of rabbit tracheal mucosa. Later, others 2-5 attempted simpler bioassay methods, using the gills from the freshwater mussel or the oyster to study ciliary inhibition. Findings presented in these studies have been contradictory; therefore, we tested the effect of CF serum on the ciliary activity of oyster gills. The oysters (Crassostrea virginica) were obtained from the Atlantic coastal areas of New Jersey and Chesapeake Bay during the months of May through July, 1969. After this time, the ~Supported by a grant from The National Cystic Fibrosis Research Foundation.
Volume 79 Number 3
accumulation of large amounts of mucus at the borders of the cilia interfered with ciliary motility and terminated this phase of the experiment. The oyster gill preparations were suspended in filtered seawater in the wells of culture slides which were sealed with a microcover glass. In addition to using the cessation of the ciliary beat as an end point, as described by Bowman and associates, 3 ,we also measured the time of appearance of expelled debris for the same purpose. Serum samples were obtained from 17 children with cystic fibrosis and from 19 children who did not have pulmonary disease; sera were tested fresh. With 16 of the 17 CF sera, debris appeared in less than an hour; with 12 of them there was cessation of ciliary beat in less than an hour. Only 3 of the 19 control sera were responsible for the appearance of any debris; this also occurred in less than an hour. Four of the control sera interfered with the ciliary beat in less than an hour. Using a x 2 test procedure, the results with the CF sera were significantly different from those with the control sera in respect to cessation of beat (0.05 > P > 0.025) and time required :For the appearance of debris (P < 0.005). Our findings tend to confirm the observations of Bowman and associates 3 that CF sera do have a ciliostatic effect which is different from that of normal sera. The time required for the appearance of debris is a more definitive end point than ~s the cessation of ciliary beat. There are, however, a number of variables inherent in the method which are responsible for considerable overlap of results. Until standardized, these variables limit the usefulness of the test in diagnosis, in genetic counseling, and in research.
Victor F. Iacocca, Ph.D. LeRoy I. Braddock, Ph.D. Giulio ]. Barbero, M.D. Department o[ Pediatrics Hahnemann Medical College and Hospital Philadelphia, Pa. REFERENCES 1. Spook, A., Heich, H. M. C., Cress, H., and Logan, W. S.: Abnormal serum factor in patients with cystic fibrosis of the pancreas, Pediat. Res. 1: I73, 1969. 2. Patrick, A. D.: In discussion, Cystic Fibrosis, Porter, R., and O'Connor, M., editors, CIBA, Foundation Study Group No. 32, London, 1968, Churchill, p. 14. 3. Bowman, B. H., Lochhart, Z. H., and MeCombs, M. L.: Oyster ciliary inhibition by cystic fibrosis factor, Science 164: 325, 1969.
Letters to the Editor
509
4. Crawford, M. d'A.: Experience of the cilia test for a factor in the serum of patients and carriers of cystic fibrosis, Proceedings of the Fifth International Cystic Fibrosis Conference, David Lawson, editor, London, 1969, p. 42. 5. Christensen, J., and Morrevang, A.: Studies on the effect of CF serum on fresh tissue from Dreissena gills, oyster gills and rabbit tracheal mucosa, Proceedings of the Fifth In ternational Cystic Fibrosis Conference, David Lawson, editor, Loncton, 1969, p. 49.
Unusual skin reation to sterile diluent in tuberculin test To the Editor: Recently, at Sinai Hospital in Baltimore, a patient in the pediatric outpatient department was observed to have a significant skin test response to the sterile diluent which is used in preparing the intermedlate-strength tuberculin test. An intradermal tuberculin test was administered to an 11-year-old Negro boy. Though controis using diluent are not standard practice at Sinai Hospital, several unconfirmed reports of equlvocaI skin test reactions (i.e., indurations of 5 to 9 mm.) had been noted two weeks before the patient was seen, and it was for this reason that 0.1 e.c. of sterile, buffered diluent was applied. Forty-eight hours later both sites showed 5 mm. of induration and erythema. A repeat application of diluent produced the same result after 48 hours, while a Tine test was negative, also at 48 hours. All injections were made using disposable plastic syringes. The patient had no known history of allergic diathesis. The phenomenon is not unknown, since Edwards and associates, 1 in testing PPD-S against pure diluent, found 2 out of 79 persons who responded measurably (2 to 4 mm.) to the diluent. The PPD preparation and diluent were returned to the manufacturer (Merck Sharp & Dohme) for analysis, and no indication of impurity or contamination was found. Sterile buffered diluent contains phosphates of sodium and potassium, as well as phenol (0.5 per cent) as a preservative. The inorganic phosphates are unlikely sensitizing agents, but Goodman and Gilman's2 text states that phenol can be markedly toxic to tissues, even unabraded skin. When directly applied to skin, a pellicle of precipitated protein may form which turns red, then brown