Parasitolo~ Today, vol. 9. no. I0. 1993 349

Immunology and Molecular Biology of Tropical Infectious Diseases

J.R, David and D.A, Ham

Since it is not possible to summarize reports from all 48 speakers at the Second Annual Meeting of the Inter- national Centers for Tropical Disease Research, this review is selective, em- phasizing talks on protective antigens, cytokines and the immune response and presentation of antigens for de- veloping vaccines. Highlights in several other areas will be mentioned briefly.

Interleukin I 0

The importance of interleukin 10 (IL-10) in suppressing the immune re sponse in Leishmania was brought up by several speakers. Following stimulation with Leishmania antigen, lymphocytes from patients with acute visceral leish- maniasis produced IL-10, but failed to do so after therapy (Steve Reed, Seattle Biomedical Research Institute, Seattle, WA, USA; Edgar Carvalho, Federal University of Bahia Medical School, Salvador, Bahia, Brazil; and Richard Pearson, University of Virginia Medical School, Charlottesville, VA, USA). The antigen-induced proliferation, diminished in lymphocytes from acute patients, was markedly enhanced by anti IL-10 (SR and EC). Some cloned L. braziliensis in- duced both THI and TH2 lymphokine mRNAs, whereas the Leishmania hom- ologue of elongation initiation factor stimulated only THI mRNA (SR). Patients exhibiting delayed hypersensi- tivity failed to produce IL-10 (RP).

In relation to IL-10, Donald Ham (Harvard School of Public Health, Boston, MA, USA) discussed the role of certain oligosaccharides on the pro- tective immune response. Antibodies to lacto-N-fucopentaose (LNFIII, found on schistosome eggs) were present in highly protective sera. LNFIII contains the Lewis × trisaccharide, the ligand for P-selectins. LNFIII was shown to strongly stimulate B cells to proliferate and produce IL-10 (which inhibits THI cells), thus switching the immune response from THI to TH2. As LNFIII or structurally similar sugars are also on the surface of Trypanosomo cruzi, HIV gpl20 and certain tumour cells, this 0 1993 Elsevier Science Pubhshers L td, I LJK)

mechanism may be used by micro- organisms and tumor cells to subvert the protective immune response, and it should be amenable to attack.

Interleukin 12

Interleukin 12 (IL-12), a cytokine which stimulates natural killer (NK) cells and T cells to produce IFN-~/ appears to be critical in THI differen- tiation and immunity to Leishmonia. Phillip Scott (University of Pennsylvania, Philadelphia, PA, USA) reported that spleens from L. major-infected C3H/HeN resistant mice made more IL-12 than cells from BALB/c suscep- tible mice. Further, the levels of IL-12 correlated directly with those of inter- feron gamma (IFN-~) in the cultures, and that anti-lL-12 monoclonal anti- bodies decreased the level of IFN-~. Finding that injecting IL-12 with soluble Leishrnania antigen (SLA) blocked IL-4 production and increased IFN-'y in BALB/c mice, he vaccinated BALB/c mice with SLA and IL-12 and showed that, following challenge, they were markedly protected, similar to C3HNeH mice at eight weeks. Fred Heinzel (Case Western Reserve Uni- versity, Cleveland, OH, USA) reported that BALB/c mice injected IP with IL-12 for the first seven days of infection showed 80% protection. This protec- tion was partially blocked by anti- IFN-~y. Infected macrophages show no mRNA for IL-12, but do if stimulated with LPS. Sergio Romagnani (Instituto di Clinica Medica III, Firenze, Italy) reported that IL-12 and IFN-~ favor Tnl differentiation of human T cells. Indeed, IL-12 can change the profile of established TH2 clones to produce IFN- ~/. Thus, IL-12 appears to play an important role in THI differentiation and activity and may be useful in vac- cine development.

Reverse Genetics

Adrian Hill (University of Oxford, UK) described reverse genetics as a

novel route to identify protective anti- gens in infectious organisms. Studies of a large case-control study in the Gambia showed a significant associ- ation between HLA class I molecule HLA-B53 and resistance to severe malaria, suggesting that HLA class I restricted cytotoxic T lymphocytes (CTLs) were playing a role. To test this prediction, analysis of self-peptides eluted from HLA-B53 indicated a bind- ing motif of a proline at position 2 and hydrophobic amino acid at position 9. Synthesized peptides corresponding to this motif from four pre-e~throcytic antigens were tested for binding to HLA-B53. Although all bound with high affinity, only one from liver-stage- specific-antigen- I (LSA-I) was the target of CTL in Gambians exposed to malaria. This epitope appears to be conserved in wild-type isolates from the Oambia. This approach can be applied to infectious diseases for which protective HLA associations have been identified and can probably be extended to HLA Class II.

Malaria Vaccine

Other studies on vaccine develop- ment focused on specific modes of immunization and specific antigens. Ruth Hussenzweig (New York Univer- sity Medical Center, New York, NY, USA) found that immunizing mice with a recombinant influenza virus contain- ing the cytotoxic T-cell epitope of the circumsporozoite (CS) protein con- ferred good protection against sporo- zoite challenge, 60% being completely protected. The protective response was CD8+ T-cell dependent. Further, Multiple Antigen Peptides (MAPs) based on the B- and T-cell epitopes of the CS protein of Plasmodium falci parum were given with alum to mice. Another group got the MAPs with complete Freund's adjuvant. Both groups showed similarly high levels of anti-sporozoite antibody. Accordingly, these MAPs are being constructed for human trials. Stephen Hoffman (Naval Medical Research Institute, Rockville,

350 Parasitology Today. vol. 9, no. I0. 1993

MD, USA) has been immunizing mice with naked DNA, using a vector with a CMV promoter that has been successful for other systems.

Schistosome Vaccine

Mette Strand (Johns Hopkins Uni- versity School of Medicine, Baltimore, MD, USA) vaccinated mice against Schistosoma mansoni using a recombi- nant 62 kDa protein, a portion of a 200 kDa protein (related to myosin) identified by sera from mice that had been made highly resistant by exposure to irradiated cercariae. Mice vaccinated three times with antigen in proteo- somes exhibited 42-83% protection. The level of protection in rats was 94-97%. Protection in baboons ranged from 0-54% with mean of 28%. Of special interest, there was a direct cor- relation, p = 0.967, between the anti- body titer to the 62 kDa protein and percent reduction in worm burden. The antibody was IgG~. No antifecundity effect was seen. Bernadette L. Ramirez (Research Institute for Tropical Medi- cine, Manila, The Philippines) present- ing results on vaccination studies in mice with S. japonicum, reported that paramyosin given several times without adjuvant gave 73% protection, whereas glutathione S-transferase (GST) was not protective.

Improved Diagnostics

Exciting new developments in im- proved diagnostics were presented. Janice Kolberg (Chiton Corporation, Emeryville, CA, USA) reported a method to detect DNA or RNA by signal amplification using branched DNA and a chemiluminescent detection system. Clinical samples are placed di- rectly into microtiter plates; the 'hands- on' time is four hours, and the results are quantitative. Qualitative results can be obtained by exposure to photo- graphic film. The assay has been used

to detect a number of viruses including HIV and is being developed for Mycobacterium tuberculosis. John Chan (Albert Einstein College of Medicine, Bronx, MY, USA) reported on a luciferase phage assay for the diagnosis of resistant tuberculosis. Mycobacterium tuberculosis is infected by a specific recombinant mycobacteriophage con- taining the luciferase gene (FFlux) and can be detected in one to two hours. If M. tuberculosis have been previously treated with drugs for 48 hours, the mycobacteriophage will only enter living bacteria. Thus, they have great potential for rapidly (in 48 h) identifying drug resistance in mycobacteria.

Polymerase Chain Reaction and Diagnostics

There were several reports on improved diagnosis by PCR. Thomas Wellems (Laboratory for Malaria Research, NIAID, NIH, Bethesda, MD, USA) having identified a single mutation in the P. falciparum dihydrofolate re- ductase gene (DHFR) associated with pyrimethamine resistance and an ad- ditional mutation for cycloguanil (two in- hibitors of DHFR), constructed primers to detect drug resistance to these two drugs by PCR. When applied to P. falciparum isolates from Rondonia, a site of epidemic malaria in Brazil, most of the isolates showed the pyrimeth- amine resistant phenotype but none showed the cycloguanil phenotype. PCR diagnosis was useful in surveying re- infection of black flies for the Oncho- cerciasis Control Program in Africa (Thomas Unnash, University of Alabama at Birmingham, AL, USA), the diagnosis of post-treatment re-lapse in visceral leishmaniasis (Jorge Arevelo, Instituto de Medicina Tropical Alexander von Humboldt, Lima, Peru), and in the diag- nosis of cutaneous leishmaniasis (Kristen Weigle, University of North Carolina, Chapel Hill, NC, USA) and tuberculosis (Robert Barker, Harvard School of Public Health, Boston, MA, USA).

Immunodiagnosis

Reports on improvements using immunodiagnosis for clinical samples included the simple detection of Crypto sporidium and Giardia by monoclonal antibodies using immunofluorescence (Charles Sterling, University of Arizona, Tucson, AZ, USA) differentiating patho- genic from nonpathogenic Entamoeba histotytica by monoclonal antibody- based enzyme-linked immunosorbent assay (ELISA) (William Petri, University of Virginia, Charlottesville, VA, USA), the use of the FAST~ELISA for field work (Victor Tsang, Center of Disease Control and Prevention, Atlanta, AL, USA), and the detection of fecal lacto- ferrin by latex agglutination for the diagnosis of inflammatory diarrhea (Richard Guerrant, University of Virginia School of Medicine, Charlottesville, VA, USA).

Transfection of Malaria

A report not directly related to the main theme of the program needs to be mentioned. This is the work by Dyann Wirth, Kamini Mendis and associ- ates (Harvard School of Public Health, Boston, MA, USA) on the first success- ful transfection of the malaria parasite. Firefly luciferase was transiently expressed under the control of pgp28 in the sexual stage of Plasmodium galinacium.

Acknowledgements The Second Annual Meeting of the Inter- national Centers for Tropical Disease Research of the National Institute of Allergy and Infectious Disease was held 28-30 April 1993, at the National Institutes of Health in Bethesda, MD, USA, with an emphasis on vaccine development and new technologies for diagnosis.

John R. David and Donald A. Ham are at the Department of Tropical Public Health, Har- vard School of Public Health, 665 Huntington Avenue, Boston, MA 02115, USA.

O r g a n i z i n g a M e e t i n g ?

Are you organizing a meeting that is of interest to parasitologists? If so, please supply Parasitology Today with the derails (dates, tide, location) and a contact name, and we will include your meeting in our Diary page, without charge.

Please make sure we receive the derails at least 8-10 weeks prior to the meeting.

A t t e n d i n g a M e e t i n g ?

Are you attending a meeting that is of interest to parasitologists? If so, perhaps you would like to consider writing a short reort of the meeting, capturing its main messages and the future directions indicated for the topic.

Please contact the Editor of Parasitology Today at Elsevier Trends Journals, 68 Hills Road, Cambridge, UK CB2 I LA.