Safety and Immunogenicity of an In Vivo Muscle Electroporation Delivery System for DNA-hsp65 Tuberculosis Vaccine in Cynomolgus Monkeys

A Bacille Calmette–Guérin (BCG) is still the only licensed vaccine for the prevention of tuberculosis, providing limited protection against Mycobacterium tuberculosis infection in adulthood. New advances in the delivery of DNA vaccines by electroporation have been made in the past decade. We evaluated the safety and immunogenicity of the DNA-hsp65 vaccine administered by intramuscular electroporation (EP) in cynomolgus macaques. Animals received three doses of DNA-hsp65 at 30-day intervals. We demonstrated that intramuscular electroporated DNA-hsp65 vaccine immunization of cynomolgus macaques was safe, and there were no vaccine-related effects on hematological, renal, or hepatic profiles, compared to the pre-vaccination parameters. No tuberculin skin test conversion nor lung X-ray alteration was identified. Further, low and transient peripheral cellular immune response and cytokine expression were observed, primarily after the third dose of the DNA-hsp65 vaccine. Electroporated DNA-hsp65 vaccination is safe but provides limited enhancement of peripheral cellular immune responses. Preclinical vaccine trials with DNA-hsp65 delivered via EP may include a combination of plasmid cytokine adjuvant and/or protein prime–boost regimen, to help the induction of a stronger cellular immune response

Inducible Nitric Oxide Synthase in Heart Tissue and Nitric Oxide in Serum of Trypanosoma cruzi-Infected Rhesus Monkeys: Association with Heart Injury

Chagas disease, a neglected tropical disease caused by the protozoan Trypanosoma cruzi, afflicts from 8 to 15 million people in the Latin America. Chronic chagasic cardiomyopathy (CCC) is the most frequent manifestation of Chagas disease. Currently, patient management only mitigates CCC symptoms. The pathogenic factors leading to CCC remain unknown; therefore their comprehension may contribute to develop more efficient therapies. In patients, high nitric oxide (NO) levels have been associated with CCC severity. In T. cruzi-infected mice, NO, mainly produced via inducible nitric oxide synthase (iNOS/NOS2), is proposed to work in parasite control. However, the participation of iNOS/NOS2 and NO in T. cruzi control and heart injury has been questioned. Here, infected rhesus monkeys and iNOS/NOS2-deficient mice were used to explore the participation of iNOS/NOS2-derived NO in heart injury in T. cruzi infection. Chronically infected monkeys presented electrical abnormalities, myocarditis and fibrosis, resembling the spectrum of human CCC. Moreover, cardiomyocyte lesion correlated with iNOS/NOS2+ cells infiltrating the cardiac tissue. Our findings support that parasite-driven iNOS/NOS2+ cells accumulation in the cardiac tissue and NO overproduction contribute to cardiomyopathy severity, mainly disturbing the pathway involved in electrical synchrony in T. cruzi infection

Thrichomys laurentius (Rodentia; Echimyidae) as a putative reservoir of Leishmania infantum and L. braziliensis: patterns of experimental infection

Submitted by Sandra Infurna ([email protected]) on 2017-08-29T15:55:29Z No. of bitstreams: 1 elisacupolillo_etal_IOC_2010.pdf: 748313 bytes, checksum: 750de936f7c0b976c21696f81a74b5ba (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-08-29T16:06:15Z (GMT) No. of bitstreams: 1 elisacupolillo_etal_IOC_2010.pdf: 748313 bytes, checksum: 750de936f7c0b976c21696f81a74b5ba (MD5)Made available in DSpace on 2017-08-29T16:06:15Z (GMT). No. of bitstreams: 1 elisacupolillo_etal_IOC_2010.pdf: 748313 bytes, checksum: 750de936f7c0b976c21696f81a74b5ba (MD5) Previous issue date: 2010Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia de Tripanosomatídeos. Rio de Janeiro, RJ. Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisas em Leishmaniose. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisas em Leishmaniose. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto de Biotecnologia em Fármacos - Farmanguinhos. Laboratório de Neurovirulência. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia de Tripanosomatídeos. Rio de Janeiro, RJ. Brasil.The importance of the genus Thrichomys in the retention of infection and transmission of Leishmania species is supported by previous studies that describe an ancient interaction between caviomorphs and trypanosomatids and report the natural infection of Thrichomys spp. Moreover, these rodents are widely dispersed in Brazil and recognized as important hosts of other tripanosomatids. Our main purpose was to evaluate the putative role of Thrichomys laurentius in the retention of infection and amplification of the transmission cycle of Leishmania infantum and L. braziliensis. Male and female T. laurentius (n = 24) born in captivity were evaluated for the retention of infection with these Leishmania species and followed up by parasitological, serological, hematological, biochemical, histological, and molecular assays for 3, 6, 9, or 12 months post infection (mpi). T. laurentius showed its competence as maintenance host for the two inoculated Leishmania species. Four aspects should be highlighted: (i) re-isolation of parasites 12 mpi; (ii) the low parasitic burden displayed by T. laurentius tissues; (iii) the early onset and maintenance of humoral response, and (iv) the similar pattern of infection by the two Leishmania species. Both Leishmania species demonstrated the ability to invade and maintain itself in viscera and skin of T. laurentius, and no rodent displayed any lesion, histological changes, or clinical evidence of infection. We also wish to point out the irrelevance of the adjective dermotropic or viscerotropic to qualify L. braziliensis and L. infantum, respectively, when these species are hosted by nonhuman hosts. Our data suggest that T. laurentius may act at least as a maintenance host of both tested Leishmania species since it maintained long-lasting infections. Moreover, it cannot be discarded that Leishmania spp. infection in free-ranging T. laurentius could result in higher parasite burden due the more stressing conditions in the wild. Therefore the tissular parasitism of the skin, infectiveness to the vector, and amplification of the transmission cycle of both Leishmania species could be expected

Kinetics of white blood cell counts in <i>Thrichomys laurentius</i> experimentally infected by <i>Leishmania braziliensis</i> (Lb) or <i>Leishmania infantum</i> (Li).

<p>Each point represents the mean values and standard errors. The * indicates the significant difference between values obtained from non-infected rodents (day 0) and and rodents infected by <i>L. braziliensis</i>.</p

Kinetics of the humoral immune response (IFAT) in <i>Thrichomys laurentius</i> experimentally infected by <i>Leishmania braziliensis</i> (Lb) or <i>Leishmania infantum</i> (Li).

<p>Each point represents the mean value and standard errors of the IgG titers of infected rodents.</p

Photomicrographs of spleen and liver sections from <i>Leishmania braziliensis</i> or <i>L. infantum</i> infected rodents stained with haematoxylin-eosin.

<p>A: <i>L. braziliensis</i>-infected golden hamster (<i>Mesocricetus auratus</i>) 3 months post infection (mpi) – spleen revealed extensive necrosis and inflammation. Note necrotic focus containing amastigotes (arrows). B: <i>L. braziliensis</i>-infected <i>Thrichomys laurentius</i> (7289) 3 mpi – spleen showing architecture of red and white pulp, lack of parasites; C: <i>L. infantum</i>-infected <i>T. laurentius</i> (7548) 12 mpi (liver culture tissue positive) – panoramic view of the liver showing typical architecture with portal (PV) and central vein (CV); D: <i>L. infantum</i>- infected <i>T. laurentius</i> (7538), 12 mpi (liver culture tissue negative) – also illustrated liver showing normal architecture; E: <i>L. infantum</i>-infected <i>T. laurentius</i> (7548), 12 mpi (liver culture tissue positive) – overview of white and red pulp from spleen; F: <i>L. infantum</i>-infected <i>T. laurentius</i> (7538), 12 mpi (liver culture tissue negative) – also illustrated spleen overview of red and white pulp.</p

Detection of <i>Leishmania</i> sp. DNA by polimerase chain reaction (PCR) in different tissues sampled from <i>Thrichomys laurentius</i> experimentally infected by <i>Leishmania braziliensis</i> or <i>Leishmania infantum</i> between months 3 and 12 post infection.

<p>Each point represents the number of positive/total PCR reactions observed for all tissue samples in the different experimental batches.</p><p>mpi. months post infection.</p

Kinetics of red blood cell counts and hemoglobin levels in <i>Thrichomys laurentius</i> experimentally infected by <i>Leishmania braziliensis</i> (Lb) or <i>Leishmania infantum</i> (Li).

<p>Each point represents the mean value and standard errors. The continuous lines indicate the normal range defined by the medium values obtained for each group one-day before the inoculum and two-fold standard errors. The * indicates the significant difference between rodents infected either by <i>L. braziliensis</i> or <i>L. infantum</i>.</p

Kinetics of albumin and total protein levels and albumin/total protein ratio in a <i>Leishmania infantum</i> infected <i>Thrichomys laurentius</i> (7548).

<p>Day 0 indicate the values obtained before the experimental infection.</p