Autofluorescence spectroscopy of optically trapped cells.

Resistance to pneumococcal infection was tested in T-lymphocyte-deficient, nude (nu/nu) mice. Pneumococcal serum opsonizing activity, in vivo phagocytosis ofthe pneumococcus, and the mean lethal dose for the pneumococcus were all found to be the same in nude mice as in control (+/+) mice. T-lymphocytes do not appear to play a significant role in the host's defense against the pneumococcus.

Resistance to infection with the pneumococ- cus (Streptococcus pneumoniae) is a complex process known to involve a number of different aspects of immunity.Surface phagocytosis (12), opsonizing antibody (11), and the third (C3) (10) and fifth (C5) (8) components of complement have all been shown to participate in the host's defense against the pneumococcus.Information is incomplete, however, on the role of thymusderived (T) lymphocytes in resistance to pneu- mococcal infections.Although the evidence ob- tained from patients with primary T-lymphocyte deficiencies suggests that, if T-lymphocytes play a role in the host's defense against the pneumococcus, it is a minor one, there is no direct experimental data on this point.
The nude mouse has a genetically deter- mined absence of the thymus and a severe deficiency of T-lymphocyte functions (4)(5)(6).As a result, it provides the opportunity to study di- rectly, and in vivo, the role of T-lymphocytes in the host's defense against pneumococcal infec- tions.
Nude (nu/nu) mice were kindly supplied by James Hansen of the National Institutes of Health, Bethesda, Md.The nude mice were demonstrated to have 2% or less theta-bearing spleen lymphocytes by R. F. Mortensen (3).Swiss-Webster (+/+) mice were purchased from Microbiological Associates, Bethesda, Md.The colony of nude mice was maintained by mating nude (nu/nu) males with heterozygous (nu/+) females.Only male homozygous mice were used in experiments.Serum was obtained by bleeding the mice from their tails, pooled, and stored at -70 C.
Serum opsonizing activity is an important determinant in the host's defense against the pneumococcus.Accordingly, pneumococcal opso- nizing activity was measured in the pooled se- rum of nude mice.Briefly, 1.25 x 10W exudate leukocytes obtained from Swiss-Webster mice and 6.25 x 108 log-phase type 25 pneumococci were added to the desired dilution of pooled test serum and the mixture was rotated at 12 rpm at 37 C for 30 min (9).Serum opsonizing activity was determined by counting the percentage of polymorphonuclear leukocytes that contained pneumococci on a stained smear.The results are expressed as percentage of phagocytosis.As can be seen in Fig. 1, pneumococcal opsonizing activity was normal in the serum of nude mice when measured over a wide range of serum concentrations.
The phagocytosis of pneumococci was also measured in vivo in nude mice.Mice were in- jected intraperitoneally with 2 ml of a starch- gluten suspension (10).Eighteen hours later, the peritoneal exudates from either nude mice or normal mice were found to contain 2 x 107 leukocytes.Other animals were then injected intraperitoneally with 4 x 108 log-phase type 25 pneumococci.Thirty minutes later each animal was sacrificed, the exudate was recovered and stained, and the percentage of phagocytosis was determined.As can be seen in Table 1, the in vivo phagocytosis of pneumococci was the same in the nude mice as in the control mice.
The mean lethal dose (LD50) for type 3 pneu- mococci, strain IIIS-IR6, of intermediate viru- lence for the mouse (Pn 3-int), was also measured in nude mice.Groups of 10 mice, weighing 16 to 18 g each, were injected intraperitoneally with 10-fold dilutions of log-phase Pn 3-int.The mice were observed for 5 days.The LD50 was calculated by the Reed-Muench method (7) and the probability (P) value was determined as in reference 13.As can be seen in Table 2, the LD50 in the nude mice was the same as that in the control mice.It should also be noted that the cumulative mortality was the same in the two groups on each day of the LD50 study.Both clinical and laboratory observations have suggested that T-lymphocytes might play a role in antipneumococcal immunity.Al- though patients with primary deficiencies of T- lymphocyte function are most notably susceptible to viruses and fungi, they also may have pneumococcal infections (2).In addition, studies have shown that the magnitude of the im- mune response to pneumococcal capsular polysaccharide is influenced by both "suppressor" and "amplifier" T-lymphocytes (1).
In the above studies, pneumococcal serum opsonizing activity and phagocytosis of the pneumococcus in vivo, both manifestations of antipneumococcal host defense, were found to be normal in nude mice.Most importantly, the LD50 for the pneumococcus was the same in nude mice as in control mice.Thus, T cells do not appear to play a significant role in the host's defense against the pneumococcus.

TABLE 1 .
In vivophagocytosis ofPn25 in preformed peritoneal exudates of nude or normal mice

TABLE 2 .
LD50 for Pn 3-int in nude mice