David J. Weiner

Fate of Litomosoides carinii adults transplanted into the pleural or peritoneal cavity of infected and naive multimammate rats (Mastomys natalensis).

In each of 4 experiments, 58 multimammate rats (Mastomys natalensis) were subdivided into 5 groups. Thirty-four rats were infected with Litomosoides carinii and infections were allowed to become patent. Ten days after patency adult worms were surgically transferred from donor rats to each of 1 group of infected rats and 1 group of naive rats. Groups of infected and naive rats served as controls. Transfers were made either intrapleurally or intraperitoneally. Samples of blood and tissues were taken from each of 2 animals necropsied from each group at intervals to 31 days. At necropsy, the transferred worms and the original population (if any) were examined and samples were fixed. Infected rats accepted new worms with a minimum of reaction while naive rats rejected worms beginning at day 10 (intrapleural) of 17 (intraperitoneal). Rejections were complete leaving a white fibrous mass by day 24 or 31, respectively. Hemagglutination antibody titers fell into 2 groups: infected and naive. IHA titers of naive recipient rats increased into the infected range by day 3 (intrapleural) or day 10 (intraperitoneal). Microfilaria counts presented a more varied pattern but a similar lag in the intraperitoneal recipient group was observed. It was concluded that a preparation period is necessary for successful residence of adult worms, and that this preparation is not restricted to the pleural cavity.

Fine structure and cytochemical evidence for the presence of polysaccharide surface coat of Dirofilaria immitis microfilariae.

Abstract Cherian P. V. , Stromberg B. E. , Weiner D. J. and Soulsby E. J. L. 1980. Fine structure and cytochemical evidence for the presence of polysaccharide surface coat of Dirofilaria immitis microfilariae . International Journal for Parasitology 10 : 227–233. Cytochemical staining techniques were employed at the fine structural level using ruthenium red, ruthenium violet and Alcian blue-lanthanum nitrate to demonstrate the polysaccharide rich surface coat of Dirofilaria immitis microfilariae. The coat matrix present at the external surface of the cuticle of microfilariae stained densely with each of the polycationic dyes. The reaction products were restricted to the outer surface of the cuticle suggesting that the polycationic dyes did not penetrate the cuticle. The junctions of the cuticular annulations lacked surface coat matrix and reaction products which might be indicative of the absence of carbohydrate residues or the masking of reactive sugar molecules in these areas. The speciflcity of the reaction was indicated by the absence of reaction products in untreated organisms. These carbohydrate moieties probably represent glycoproteins as structural constituents of the parasite surface. Ultrastructural analysis of the surface of microfilariae is of signiflcance in elucidating both the molecular dynamics of the parasite surface and its immunological function in the host.

Litomosoides carinii: effect of splenectomy on the ability of naive Mastomys natalensis to accept transplanted adults.

Abstract Fifty-eight male rats Mastomys natalensis , were used in each of two experiments to study the effect of splenectomy on the rejection of transplnted nematodes, Litomosoides carinii . The rats were divided into five groups: splenectomized recipients (SR), normal recipients (NR), splenectomized (S), normal (N) and donor (D). Each group had 12 animals except for the D group which had 10. After patency of the D group (tank infected with Litomosoides carinii ), groups SR and S were splenectomized. Fourteen days later, (three to five L. Carinii derived from the D group) were surgically transplanted into the peritoneal cavity of the SR and NR groups. In the first experiment, weekly sacrifices were made starting at Day 3. In the second experiment, all groups were sacrificed at Day 32. Worms transplanted into the SR group were accepted while those transplanted into the NR group were rejected. Sequentially examined antibody titers after Day 3 fell into two groups, those that were recipients of transplants (SR and NR) and those that were not (NR and SR). After becoming positive on Day 3, the microfilaremia of the SR group rose by Day 31 while that of the NR group fell to near 0. It was concluded that the spleen is necessary for the rejection of transplanted L. carinii by naive M. natalensis .

Immunotherapy with VGX-3100 (HPV16 and HPV18 plasmids) + INO-9012 (DNA encoding IL-12) in human papillomavirus (HPV) associated head and neck squamous cell carcinoma (HNSCCa): interim safety and immunogenicity results

Meeting abstracts Oropharyngeal HNSCCa is frequently associated with HPV infection. DNA-based Immunotherapy with plasmids encoding HPV16 and HPV18 E6/E7 antigens has been shown to generate robust immune responses in women with HPV-driven high-grade cervical dysplasia. We hypothesize that HPV-

Litomosoides carinii in jirds ( Meriones unguiculatus ): ability to retard development of challenge larvae can be transferred with cells and serum

To test the ability of cells and/or serum from jirds (Meriones unguiculatus) infected with Litomosoides carinii to transfer the ability to retard development of challenge larvae, a series of transfer experiments were done. Groups of jirds received larval challenge preceded by one of eight preparations: spleen cells and/or serum from 10-day-patent infected jirds; normal spleen cells and/or normal serum; primary larvae; challenge larvae only. No significant differences in size or numbers of larvae recovered were found among groups receiving either cells or serum only. However, significant differences in larval size were found between groups receiving both cells and serum from infected donors and those receiving normal cells and serum. These comparisons indicate that the ability of infected jirds to retard development of challenge infection larvae can be transferred with cells and serum together but not separately.

The effect of diethylcarbamazine on microfilariae ofLitomosoides carinii in vitro and in vivo

Culture-derivedLitomosoides carinii microfilariae (MFF) were used in in vitro and in vivo systems to investigate the effect of diethylcar-bamazine (DEC) on these MFF. In vivo: Male rats,Mastomys natalensis, all of the same age, were injected intrathoracically (12) or intraperitoneally (36) with 103 or 104 MFF. After 30 min one half of each group of rats was given DEC per os. At 30, 60, and 120 min after DEC administration, two rats from the treated and two from the untreated group were bled and killed. The pleural or peritoneal cavities were rinsed with warm saline (0.15m NaCl) to recover MFF. In both the intrathoracic and intraperitoneal experiments, equal numbers of MFF were recovered from treated and control rats at 30 and 120 min. However, at 60 min 85.5% fewer were recovered from the treated than from the nontreated animals. MFF were not found in the blood. In vitro: MFF were added to tissue culture dish wells (Linbro Div., Flow Labs, Hamden, Conn) prepared as follows: DEC-Serum (serum from normal rats given DEC at 500 mg/kg), DEC+Serum (serum with added DEC), serum only, RPMI 1640 only, and RPMI 1640+DEC. Furthermore, the five treatments were prepared either with or without unstimulated peritoneal exudate (PE) cells. At 30 min in the DEC-Serum wells 45% of the MFF had adherent PE cells; in the remaining wells these cells adhered to 11% or fewer MFF. We interpret the aforementioned phenomena as representing the first step in the trapping and elimination of MFF after DEC treatment ofL. carinii-infectedM. natalensis.

The 2-Mercaptoethanol Labile Immunoglobulin Response of Beagles Experimentally Infected with Dirofilaria immitis

Seventeen purebred beagles, maintained in isolation from mosquitoes, were divided into 3 groups: Group I (noninfected controls), Group II (infected once), and Group III (infected twice). Groups II and III were inoculated subcutaneously with 30 Dirofilaria immitis infective larvae. Upon patency of the first infection (31 weeks), Group III was inoculated a 2nd time with 30 infective larvae. Throughout the 78-week experimental period, weekly blood serum samples were tested by indirect hemagglutination (IHA) for humoral antibodies using D. immitis extract as antigen. Selected serum samples were tested by single radial diffusion to estimate the relative amounts of immune globulin M (IgM) and by 2-mercaptoethanol (2-ME) sensitivity and IHA for specific antibody of the 2-ME labile type (presumably IgM). The beginning of a strong antibody response was noted in Groups II and III by the 10th week after inoculation, which was decreasing by the 34th week. After receiving the 2nd inoculation of larvae the mean titer of Group III continued at the postpatent level while the titers of Group II (which received only a single dose of larvae) became variable and were seen to decline slightly. In both infected groups, however, the mean titers were similar by the 74th week after infection. The immunodiffusion results indicated a fairly constant level of IgM, but the 2-ME sensitivity studies indicated an IgM response in Groups II and III (peak at 25 weeks) which decreased in Group II and persisted in Group III. In both groups a 2-ME nonlabile antibody response persisted as late as 60 weeks after infection. It was concluded that the antibody titers seen in this study after primary and secondary infections with D. immitis were composed mainly of IgM. This was probably due to the difference in efficiency of the 2 immunoglobulins in the IHA test. No evidence of an increase in antibody titer was seen after the 2nd infection, but this may not apply to reinfections given at a later time than those in the present work. The characteristics and dynamics of the immunologic response of the dog to Dirofilaria immitis infection have been reported recently (Pacheco, 1966; Tulloch et al., 1970), but these studies considered only primary infections and data have not been published on the activity of the various immunoglobulins during the course of infection. Weiner and Bradley (1972) reported that the humoral antibody response of the dog, measured by indirect hemagglutination (IHA), is variable. Specific antibody was detected 10 weeks after infection with 30 infective larvae of D. immitis, but antibody titers declined with the appearance of microfilariae in the peripheral circulation. Following reinfection at week 31, antibody titers failed to increase, but those of the singly infected group decreased. Received for publication 2 March 1973. * This investigation was supported in part by NIH Training Grant No. 5 TO1-A100383-02 from the NIAID. Florida Agricultural Experiment Stations Journal Series No. 4524. t Present address: Laboratories of Parasitology, The School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pa. 19104. The purpose of the present study was to examine the pattern of IgM production during the course of both primary and secondary infections of D. immitis in order to attempt to clarify the apparent lack of a classical secondary response following reinfection of the dog. MATERIALS AND METHODS The detailed experimental protocol is reported elsewhere (Weiner and Bradley, 1972). Seventeen purebred beagles (Hazelton-Saunders Labs, Midlothian, Va.) were treated as follows: Group I (6 normal controls), Group II (5 dogs, infected once), and Group III (6 dogs, infected twice). Dogs were reared helminth-free and maintained in mosquito-free isolation with food and water ad lib. A 12-ml blood sample was drawn from each dog weekly, 3 ml of which were collected in EDTAtreated tubes for blood cellular work and 9 ml allowed to clot for serum collection. Serum was stored at -12 C.

Litomosoides carinii : retardation of worm growth and of migration of challenge infections in jirds ( Meriones unguiculatus )

Inbred jirds (Meriones unguiculatus) were divided into three groups; each animal in two of the groups was infected with 30 infective larvae (L3) of Litomosoides carinii. When these infections were patent, the jirds of one of the two infected groups plus those of the third group were injected with 30 L3 L. carinii each. All animals were killed either on day 14 or 24 after the second infection for the recovery, enumeration and measurement of all worms and developing larvae. Challenge larvae were stunted (smaller) and fewer than control larvae. Additionally, fewer challenge larva were recovered on day 14 than on day 24, indicating that migration to the pleural cavity was retarded.