Malnutrition is a risk factor for the development of visceral leishmaniasis. However, the immunological basis for this susceptibility is unknown. We have developed a mouse model to study the effect of malnutrition on innate immunity and early visceralization following Leishmania donovani infection. Three deficient diets were studied, including 6, 3, or 1% protein; these diets were also deficient in iron, zinc, and calories. The control diet contained 17% protein, was zinc and iron sufficient, and was provided ab libitum. Three days after infection with L. donovani promastigotes, the total extradermal (lymph nodes, liver, and spleen) and skin parasite burdens were equivalent in the malnourished (3% protein) and control mice, but in the malnourished group, a greater percentage (39.8 and 4.0%, respectively; P ‫؍‬ 0.009) of the extradermal parasite burden was contained in the spleen and liver. The comparable levels of parasites in the footpads in the two diet groups and the higher lymph node parasite burdens in the well-nourished mice indicated that the higher visceral parasite burdens in the malnourished mice were not due to a deficit in local parasite killing but to a failure of lymph node barrier function. Lymph node cells from the malnourished, infected mice produced increased levels of prostaglandin E 2 (PGE 2 ) and decreased levels of interleukin-10. Inducible nitric oxide synthase activity was significantly lower in the spleen and liver of the malnourished mice. Thus, malnutrition causes a failure of lymph node barrier function after L. donovani infection, which may be related to excessive production of PGE 2 and decreased levels of IL-10 and nitric oxide.Globally, protein-energy malnutrition is the most frequent cause of immunodeficiency (58). Epidemiologic and experimental studies have documented an increased risk for visceral leishmaniasis, caused by intracellular protozoan parasites of the Leishmania donovani complex, in the malnourished host (1, 2, 26). However, the immunologic basis for this association has not been established and standardized experimental models have not addressed this important issue.In this study, our goal was to investigate the mechanisms of the malnutrition-related susceptibility to visceral leishmaniasis. There were three components of the study. First, we needed to create a murine model of malnutrition that was relevant to human malnutrition in developing countries. Although the mouse has been extensively used in animal models of malnutrition, there is no standard murine model of protein-energy malnutrition (69). Human malnutrition is complex, typically involving deficiency of protein and energy with superimposed deficits of other nutrients. Zinc deficiency usually accompanies protein-energy malnutrition (19). Iron deficiency is highly prevalent in developing countries and may accompany zinc deficiency due to a common risk factor, cereal-based diets with little meat (61). Thus, in this model, in addition to protein and energy, zinc and iron were selected as deficient nutrients.M...

The acquisition of immunity following subclinical or resolved infection with the intracellular parasite Leishmania donovani suggests that vaccination could prevent visceral leishmaniasis (VL). The LACK (Leishmania homolog of receptors for activated C kinase) antigen is of interest as a vaccine candidate for the leishmaniases because of its immunopathogenic role in murine L. major infection. Immunization of mice with a truncated (24-kDa) version of the 36-kDa LACK antigen, delivered in either protein or DNA form, was found previously to protect against cutaneous L. major infection by redirecting the early T-cell response away from a pathogenic interleukin-4 (IL-4) response and toward a protective Th1 response. The amino acid sequence of the Leishmania p36(LACK) antigen is highly conserved, but the efficacy of this vaccine antigen in preventing disease caused by strains other than L. major has not been determined. We investigated the efficacy of a p36(LACK) DNA vaccine against VL because of the serious nature of this form of leishmaniasis and because it was unclear whether the LACK vaccine would be effective in a model where there was not a dominant pathogenic IL-4 response. We demonstrate here that although the LACK DNA vaccine induced a robust parasite-specific Th1 immune response (IFN-␥ but not IL-4 production) and primed for an in vivo T-cell response to inoculated parasites, it did not induce protection against cutaneous or systemic L. donovani challenge. Coadministration of IL-12 DNA with the vaccine did not enhance the strong vaccine-induced Th1 response or augment a protective effect.

The present study demonstrates that HPV may infect the glandular mucosa of the colon and suggests a possible association between HPV and colorectal cancer.

Although the implication of genetic factors in cervical cancer development remains to be elucidated, accumulative epidemiological evidence suggests that polymorphisms of cytokine genes may be involved in the etiology of cervical carcinoma. Tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10) are two multifunctional cytokines implicated in inflammation, immunity, and cellular organization, and were proposed to play important roles in cancer biology. In order to determine whether IL-10 -1082 (G/A) and TNF-α -238 (G/A) and -308 (G/A) polymorphisms are associated with susceptibility to cervical cancer, a case-control study of 122 cancer patients and 176 healthy controls was conducted. Cervical samples were genotyped for both TNF-α polymorphisms by PCR-RFLP assay. SNP-1082 from IL-10 gene was genotyped using pyrosequencing technology. The association between cervical cancer risk and the studied SNPs was evaluated by logistic regression. Under univariate analysis, none of these polymorphisms appeared associated with susceptibility of cervical cancer development or HPV infection. However, individuals carrying heterozygous genotype for TNF-α -238 polymorphism seem to be at lower risk for cervical cancer development, with borderline significance (OR = 0.42, P = 0.069), as well as those carrying heterozygous genotypes for IL-10 and TNF-α -238 (OR = 0.40, P = 0.08). In conclusion, these results suggest a potential effect of TNF-α -238 G/A in the reduction of cervical cancer risk in Argentine women, but not TNF-α -308 or IL-10. Larger studies are needed to fully understand the genetic predisposition for the development of cervical cancer.

Progressive disease in the hamster model of visceral leishmaniasis, caused by Leishmania donovani, in contrast to infection in mice, mimics the progressive disease observed in untreated humans. During progressive infection in hamsters, there was a vigorous type 1 cellular immune response, which is typically associated with control of infection, suggesting that there was ineffective IFN-γ-mediated macrophage activation. Indeed, at the site of infection, hamsters did not express NO synthase 2 (NOS2), which is the primary mechanism for control of infection in mice. Furthermore, in striking contrast to mouse macrophages, IFN-γ-activated hamster macrophages did not did not express NOS2 nor generate NO, and were unable to restrict the replication of intracellular L. donovani. The absent hamster NOS2 expression was not the result of NOS2 gene deletion and the NOS2 cDNA had an intact open reading frame. Furthermore, the impaired transcription of NOS2 mRNA was selective and not due to global impairment of IFN-γ signaling (members of the IFN-γ-signaling pathway were expressed and functional and IFN-γ up-regulated several primary and secondary response genes). Strikingly, the proximal hamster NOS2 promoter, like the human ortholog, had >20-fold less basal and IFN-γ/LPS-inducible activity than the corresponding mouse promoter. Thus, reduced basal and IFN-γ-induced activity of the hamster NOS2 transcriptional unit, which is unique to this small animal and similar to the human counterpart, accompanies the inability of the animal to control an intracellular pathogen.

The aim of the present study is to determine the presence and molecular integrity of high-risk HPV types in colorectal adenocarcinomas and to assess whether viral DNA is related to common proto-oncogene alterations, such as k-ras mutations and c-myc gene amplification, in colorectal cancer. Seventy-five colorectal adenocarcinomas were screened for HPV infection using nested-PCR (MY09/11-GP5+/6+). HPV typing was performed by type-specific PCR for HPV 16 and HPV 18 DNA. Unidentified samples were subsequently sequenced to determine the viral genotype. The physical status of HPV was determined by a nested PCR approach for type-specific E2 sequences. C-myc amplification was assessed by co-amplification with β-globin as control locus, and mutation in k-ras codons 12 and 13 by ARMS-PCR. Overall, HPV was detected in thirty-three colorectal specimens (44%). HPV 16 was the prevalent type (16/75), followed by HPV 18 (15/75), HPV 31 (1/75) and HPV 66 (1/75). E2 disruption was detected in 56.3% of HPV 16 and in 40% of HPV 18 positive tumors. C-myc amplification was detected in 29.4% of cases, while k-ras mutations in 30.7%. There was no significant trend for HPV infection in tumors harboring either k-ras or c-myc alterations. This study demonstrates HPV DNA and viral integration in colorectal tumors, suggesting a potential role of this virus in colorectal carcinogenesis. There was no concurrence, however, of k-ras and c-myc activation with viral infection.

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BackgroundHuman papillomavirus (HPV) is the carcinogen of almost all invasive cervical cancer and a major cause of oral and other anogenital malignancies. HPV genotyping by dideoxy (Sanger) sequencing is currently the reference method of choice for clinical diagnostics. However, for samples with multiple HPV infections, genotype identification is singular and occasionally imprecise or indeterminable due to overlapping chromatograms. Our aim was to explore and compare HPV metagenomes in abnormal cervical cytology by deep sequencing for correlation with disease states.ResultsLow- and high-grade intraepithelial lesion (LSIL and HSIL) cytology samples were DNA extracted for PCR-amplification of the HPV E6/E7 genes. HPV+ samples were sequenced by dideoxy and deep methods. Deep sequencing revealed ~60% of all samples (n = 72) were multi-HPV infected. Among LSIL samples (n = 43), 27 different genotypes were found. The 3 dominant (most abundant) genotypes were: HPV-39, 11/43 (26%); -16, 9/43 (21%); and -35, 4/43 (9%). Among HSIL (n = 29), 17 HPV genotypes were identified; the 3 dominant genotypes were: HPV-16, 21/29 (72%); -35, 4/29 (14%); and -39, 3/29 (10%). Phylogenetically, type-specific E6/E7 genetic distances correlated with carcinogenic potential. Species diversity analysis between LSIL and HSIL revealed loss of HPV diversity and domination by HPV-16 in HSIL samples.ConclusionsDeep sequencing resolves HPV genotype composition within multi-infected cervical cytology. Biodiversity analysis reveals loss of diversity and gain of dominance by carcinogenic genotypes in high-grade cytology. Metagenomic profiles may therefore serve as a biomarker of disease severity and a population surveillance tool for emerging genotypes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-017-3612-y) contains supplementary material, which is available to authorized users.