College of American Pathologists
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  Clinical Abstracts





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December 2001

Use of amplification and serologic tests to diagnose minimal tuberculosis
The most widely used rapid test for the diagnosis of active tuberculosis is direct microscopic examination of a smear of sputum for acid-fast bacilli (AFB smear). The preparation and reading of smears, however, is time-consuming and detects only 40 to 80 percent of pulmonary tuberculosis cases, and only the more advanced cases. Patients at any earlier stage of the disease, while still negative by smear, are less contagious and have lower morbidity and mortality; thus diagnosis of these patients at this earlier stage would be advantageous. Culture mycobacterium is a highly sensitive test, but it takes at least two weeks if solid media are used. Chest x-ray is commonly used to diagnose active TB, but it has poor sensitivity, specificity, and reproducibility. Tuberculin skin testing has very poor specificity, and false-negatives occur in 20 to 30 percent of patients diagnosed with active TB. The newer nucleic acid amplification techniques have specificities in the range of 95 percent as well as 95 percent sensitivity in smear-positive specimens. The tests, however, are expensive and require sophisticated equipment, and the sensitivity is only 50 to 71 percent in patients with smear-negative TB. This, of course, is the setting in which a rapid diagnostic test other than AFB smear is most needed. Serologic tests using an enzyme-linked immunosorbent assay technique to detect antibodies to Mycobacterium tuberculosis are relatively simple and inexpensive but have demonstrated poor sensitivity. The authors conducted a prospective study to evaluate the sensitivity, specificity, and clinical utility of nucleic acid amplification tests and a new commercially available multiple-antigen serologic test for the diagnosis of minimal pulmonary TB. The study involved 500 consecutive patients at a clinic in Montreal who were referred for sputum induction for diagnosis of possible active TB. Patients underwent sputum induction, chest x-ray, and tuberculin testing, and had blood drawn for serologic tests. Fluorescent microscopy and PCR (by the Roche Amplicor MTB) were performed, and the specimens were cultured for mycobacteria using the Bactec liquid media and solid media. Sixty cases of active TB emerged from the group of screened patients. The sensitivity and specificity, respectively, of the following diagnostic tests were: mycobacterial culture, 73 percent and 100 percent; PCR, 42 percent and 100 percent; chest x-ray, 67 to 77 percent and 66 to 76 percent; tuberculin testing, 94 percent and 20 percent; and serology, 33 percent and 87 percent. The authors concluded that no test has sensitivity and specificity high enough for the accurate diagnosis of these minimal pulmonary TB cases. They suggest a combination of tests, together with consideration of key clinical characteristics, to improve diagnostic accuracy.

Al Zahrani K, Al Jahdali H, Poirier L, et al. Accuracy and utility of commercially available amplification and serologic tests for the diagnosis of minimal pulmonary tuberculosis. Am J Resp Crit Care Med. 2000;162:1323-1329.

Reprints: Dr. Dick Menzies, Montreal Chest Institute, 3650 St. Urbain St., Montreal, Quebec, H2X 2P4 Canada;

Interaction of plasma cholesterol and red blood cells
The literature has indicated that symptoms of angina pectoris diminish within weeks after blood cholesterol levels are lowered through drug therapy. In vitro study of hypercholesterolemic rabbit blood has shown that red blood cell membrane cholesterol content is in dynamic equilibrium with plasma cholesterol content. High blood cholesterol concentrations reduce red cell oxygen transport capacity, and, in the chronic, steady state, these high plasma cholesterol levels shift the hemoglobin dissociation curve of the red cells to the left. A similar human red blood cell transmembrane barrier to oxygen diffusion in the presence of hypercholesterolemia has been demonstrated in a pilot study involving four humans. The authors attempted to determine whether and to what extent the plasma cholesterol concentration is an associated factor in red cell oxygen release and cellular oxygen availability. They performed red cell membrane oxygen diffusion analysis on samples from 100 patients and analyzed these findings with respect to the plasma, red cell, and red cell membrane cholesterol concentrations. Forty-nine of the 100 patients had plasma cholesterol in the range of 175 to 229 mg/dL. The other 59 patients had plasma cholesterol in the range of 230 to 299 mg/dL. The high cholesterol group had plasma cholesterol concentrations that were on average 24.35 percent higher than those of the low cholesterol group. Red cell cholesterol concentration was 14 percent higher in the high cholesterol group. For red cell membranes, concentrations were 31.7 percent higher. A highly significant difference was noted between the oxygen diffusion curves in the blood of the high and low cholesterol groups (P<0.05). Blood oxygen diffusion defined by these diffusion curves was inversely proportional to the plasma, red cell, and red cell membrane cholesterol concentrations. The relative tissue oxygen availability after a circulation period of more than three minutes in the diffusion system showed a decrease of 17.5 percent between the plasma cholesterol groups. There was a difference of 35 percent in the relative tissue oxygen availability between the two extremes of plasma cholesterol concentration. The authors concluded that the plasma cholesterol concentration may be an influencing factor in red cell cholesterol membrane content, which in turn may regulate red cell membrane oxygen transport, oxygen release, and cellular availability of oxygen. Therefore, one reason to modify lipid levels may be to alleviate angina pectoris symptoms.

Buchwald H, Menchaca HJ, Michalek VN, et al. Plasma cholesterol: an influencing factor in red blood cell oxygen release and cellular oxygen availability. J Am Coll of Surg. 2000;191;490-497.

Reprints: Dr. Henry Buchwald, University of Minnesota, 420 Delaware St. SE, 290 MMC, Minneapolis, MN 55455

Use of adenine-saline-preserved red cell transfusions in premature infants
Concerns about transfusing red cells in premature infants in the neonatal intensive care unit include using red cells collected in additive solutions, number of donor exposures, and hyperkalemia. This study examined the safety and efficacy of transfusing split packs of red cells collected in adenine-saline additive solution (AS-3) and stored for up to 35 days. Type O, Rh(D)-negative, CMV-negative RBC units were collected in AS-3. The units were packed and separated into four split packs (85 mL each) using a sterile docking device and designated for one patient only to minimize donor exposures to the recipient. The RBC units were not washed or irradiated and were not subject to extra manipulation prior to transfusion. Blood was transfused to neonates in volumes of 7 mL/kg and administered by syringe pump over one to three hours. Fifty-six babies received a total of 263 small-volume transfusions from AS-3-preserved split RBC packs (mean, 4.7 transfusions per infant; range, one to 25). There was a mean of 1.7 donor exposures per infant (range, one to eight). The mean overall increase in hematocrit level was 0.04 (four percent) following a 7 mL/kg packed RBC transfusion. The response to transfusion was at least as good with the older units as with the fresher ones. No significant differences were seen in pre- and post-transfusion potassium and bilirubin values. The mean change in potassium was -0.16 mmol/L. The mean change in total bilirubin was +1.86 µmol/L. No significant increases in potassium or bilirubin were noted when calculated according to the age of the RBC unit. The authors concluded that designated AS-3-preserved RBC packs limit donor exposures, can be used safely for small-volume neonatal transfusions until their date of expiration, and give a constant transfusion effect up to 35 days of storage.

Mangel J, Goldman M, Garcia C, et al. Reduction of donor exposures in premature infants by the use of designated adenine-saline preserved split red blood cell packs. J Perinatol. 2001;21:363-367.

Correspondence: Dr. Gwendoline Spurll, Dept. of Hematology, C-686, Royal Victoria Hospital, 687 Avenue des Pins, Montreal, Quebec, H3A 1A1, Canada

Is serum hyaluronic acid a marker for fibromyalgia?
The etiology of fibromyalgia is unclear. It may not be a discrete disorder as no consistent abnormalities of skeletal muscle have been described in the condition. Fibromyalgia may occur in connection with rheumatic conditions or as a primary complaint. Alterations in turnover of connective tissue theoretically could relate to the characteristic symptoms of pain, tenderness, and muscle weakness. Findings of marginal decreases in collagen propeptides and abnormal collagen deposition around nerve endings have been described. One study showed a 10-fold increase in serum hyaluronic acid levels, but this finding could not be replicated in a small followup study. The authors of this study measured hyaluronic acid in serum from 53 Danish patients with clinically established fibromyalgia and 55 control subjects using a radiometric assay. They found no association between hyaluronic acid levels and fibromyalgia; all results were within reference range.

Bliddal H, Møller HJ, Schaadt M, et al. Patients with fibromyalgia have normal serum levels of hyaluronic acid. J Rheumatol. 2000; 27:2658-2659.

Reprints: Dr. H. Bliddal, Dept. of Rheumatology, Frederiksberg Hospital, DK-2000 Copenhagen F, Denmark;

CD8+ T-cell participation in the primary immune response to Epstein-Barr virus
Recent research has shown that a massive proliferation of antigen-specific T-cells is an early part of the CD8+ T-cell response to a foreign antigen. Many of these cells die as antigen levels fall, and small numbers remain to mediate the response to subsequent exposures to the antigen. Details are lacking as to mechanisms of downregulation and selection of the surviving memory cells. The authors used blood specimens from 14 HLA-A2+ and HLA-B8+ individuals in the acute phase of infectious mononucleosis to study the primary immune response to the Epstein-Barr virus infection. In this setting, very large numbers of cells from expanded T-cell clones dominate the response. These cells are highly cytotoxic, expressing high levels of perforin. They are also heterogeneous with respect to the secretion of proinflammatory cytokines—some subpopulations being hyporesponsive. The majority of cells are programmed to die via cytokine-rescuable pathway, and the clones dominating the primary response are heavily culled during downregulation of the primary response and establishment of T-cell memory.

Callan MFC, Fazou C, Yang H, et al. CD8+ T-cell selection, function, and death in the primary immune response in vivo. J Clin Invest. 2000;106:1251-1261.

Reprints: Andrew J. McMichael, Medical Research Council Human Immunology Unit, Institute of Molecular Medicine, Headington, Oxford OX3 9DS, United Kingdom;