Effect of acute Plasmodium falciparum malaria on reactivation and shedding of the eight human herpes viruses

Human herpes viruses (HHVs) are widely distributed pathogens. In immuno-competent individuals their clinical outcomes are generally benign but in immuno-compromised hosts, primary infection or extensive viral reactivation can lead to critical diseases. Plasmodium falciparum malaria profoundly affects the host immune system. In this retrospective study, we evaluated the direct effect of acute P. falciparum infection on reactivation and shedding of all known human herpes viruses (HSV-1, HSV-2, VZV, EBV, CMV, HHV-6, HHV-7, HHV-8). We monitored their presence by real time PCR in plasma and saliva of Ugandan children with malaria at the day of admission to the hospital (day-0) and 14 days later (after treatment), or in children with mild infections unrelated to malaria. For each child screened in this study, at least one type of HHV was detected in the saliva. HHV-7 and HHV-6 were detected in more than 70% of the samples and CMV in approximately half. HSV-1, HSV-2, VZV and HHV-8 were detected at lower frequency. During salivary shedding the highest mean viral load was observed for HSV-1 followed by EBV, HHV-7, HHV-6, CMV and HHV-8. After anti-malarial treatment the salivary HSV-1 levels were profoundly diminished or totally cleared. Similarly, four children with malaria had high levels of circulating EBV at day-0, levels that were cleared after anti-malarial treatment confirming the association between P. falciparum infection and EBV reactivation. This study shows that acute P. falciparum infection can contribute to EBV reactivation in the blood and HSV-1 reactivation in the oral cavity. Taken together our results call for further studies investigating the potential clinical implications of HHVs reactivation in children suffering from malaria.     

Prevalence of human herpesvirus (HHV)-8 infection among carriers of cardiovascular disease

Infectious agents, such as herpesviruses, have been hypothesized to be involved in development of atheromatous plaque. The study aim was to evaluate the possibility that HHV-8 infection could be an additional risk factor for the establishment of cardiovascular disease. HHV-8 seroprevalence was determined by immunofluorescence in a population of cardiovascular disease patients (n=50) as compared to an age- and sex-matched group of control subjects (n=47); HHV-8 genome was detected in DNA extracted from circulating PBMC and from atheromatous lesions by PCR with primers specific for the minor virus capsid gene (ORF 26). The seroprevalence of HHV-8 was significantly increased in the patients as compared to the control population, while the presence of HHV-8 genome was observed in PBMC from 2 patients and 1 control. Virus-specific DNA was found in 2 out of 4 atheromatous plaques. The higher seroprevalence in patients suffering from vascular diseases as compared to age-and sex-matched controls suggests that HHV-8 infection could be an additional risk factor for the establishment of cardiovascular disease, although the data on the persistence of viral DNA in PBMC or in the arterial lesions are too exiguous to definitively support this hypothesis. More extensive studies are needed to define the exact role of HHV-8 infection in the establishment and progression of atheromatous lesions.     

In vitro acyclovir and cidofovir susceptibilities of human herpesvirus type 1 clinical isolates

Infections with human herpesviruses types 1 and 2 (HHV-1 and HHV-2) are common worldwide and cause a wide range of signs and symptoms. Antiviral drugs, in particular aciclovir are used in therapy of herpetic infections. The aim of the study was determination of susceptibilities of HHV-1 isolates (n+46) for antiviral drugs (acyclovir and cidofovir) in vitro. Swabs taken from different lesions were used for infection of Vero cells and cythopathic effect was observed. Viruses from cell cultures with positive CPE were later identified with in-house PCR and efficacy of acyclovir and cidofovir in HHV-1 infected Vero cell monolayer cultures was tested by the yield reduction assay. Obtained data indicate, that aciclovir ID50 average value for HHV-1 clinical isolates was 0.74 microg/ml--the value about 10% greater then described in literature. Similarly in vitro analysis of sensitivity of viruses for cidofovir, shows that concentrating is over ten-fold higher in comparison for aciclovir.     

Experimental cutaneous primary infection with Cryptococcus neoformans (Sanfelice) Vuillemin

Summary 1. Subcutaneous inoculation into the forelegs of hamsters withC. neoformans following trauma as well as subcutaneous inoculation into the abdominal wall led to infection with dissemination.2. Active skin lesions persisted during the 39 days of the experiment without regression of skin lesions.3. Involvement of lymph nodes was observed in (a few) animals infected with strains 220 and 216 only.4. In addition to the virulence of the strain, the dose of the organisms also affected the degree of infection in the tissues.5 Intratesticular inoculation led to infection with faster and more severe dissemination.6. Schaumann bodies were observed in skin lesions.7. Cortisone accelerated the course of infection.8. Primary skin infection with subsequent I.P. challenge produced a partial protection insofar as the hematogenous spread was concerned.     

Human Herpesvirus-6 U14 Induces Cell-Cycle Arrest in G2/M Phase by Associating with a Cellular Protein,EDD

The human herpesvirus-6 (HHV-6) infection induces cell-cycle arrest. In this study, we found that the HHV-6-encoded U14 protein induced cell-cycle arrest at G2/M phase via an association with the cellular protein EDD, a mediator of DNA-damage signal transduction. In the early phase of HHV-6 infection, U14 colocalized with EDD dots in the nucleus, and similar colocalization was also observed in cells transfected with a U14 expression vector. When the carboxyl-terminal region of U14 was deleted, no association of U14 and EDD was observed, and the percentage of cells in G2/M decreased relative to that in cells expressing wild-type U14, indicating that the C-terminal region of U14 and the U14–EDD association are critical for the cell-cycle arrest induced by U14. These results indicate that U14 is a G2/M checkpoint regulator encoded by HHV-6.     

Rabbitpox virus and vaccinia virus infection of rabbits as a model for human smallpox

The threat of smallpox release and use as a bioweapon has encouraged the search for new vaccines and antiviral drugs, as well as development of new small-animal models in which their efficacy can be determined. Here, we reinvestigate a rabbit model in which the intradermal infection of rabbits with very low doses of either rabbitpox virus (RPV) or vaccinia virus Western Reserve (VV-WR) recapitulates many of the clinical features of human smallpox. Following intradermal inoculation with RPV, rabbits develop systemic disease characterized by extensive viremia, numerous secondary lesions on the skin and mucocutaneous tissues, severe respiratory disease, death by 9 days postinfection, and, importantly, natural aerosol transmission between animals. Contrary to previous reports, intradermal infection with VV-WR also resulted in a very similar lethal systemic disease in rabbits, again with natural aerosol transmission between animals. When sentinel and index animals were cohoused, transmission rates approached 100% with either virus, with sentinel animals exhibiting a similar, severe disease. Lower rates of transmission were observed when index and sentinel animals were housed in separate cages. Sentinel animals infected with RPV with one exception succumbed to the disease. However, the majority of VV-WR-infected sentinel animals, while becoming seriously ill, survived. Finally, we tested the efficacy of the drug 1-O-hexadecyloxypropyl-cidofovir in the RPV/rabbit model and found that an oral dose of 5 mg/kg twice a day for 5 days beginning 1 day before infection was able to completely protect rabbits from lethal disease.     

Common Avian Infection Plagued the Tyrant Dinosaurs

Background

Tyrannosaurus rex and other tyrannosaurid fossils often display multiple, smooth-edged full-thickness erosive lesions on the mandible, either unilaterally or bilaterally. The cause of these lesions in the Tyrannosaurus rex specimen FMNH PR2081 (known informally by the name ‘Sue’) has previously been attributed to actinomycosis, a bacterial bone infection, or bite wounds from other tyrannosaurids.

Methodology/Principal Findings

We conducted an extensive survey of tyrannosaurid specimens and identified ten individuals with full-thickness erosive lesions. These lesions were described, measured and photographed for comparison with one another. We also conducted an extensive survey of related archosaurs for similar lesions. We show here that these lesions are consistent with those caused by an avian parasitic infection called trichomonosis, which causes similar abnormalities on the mandible of modern birds, in particular raptors.

Conclusions/Significance

This finding represents the first evidence for the ancient evolutionary origin of an avian transmissible disease in non-avian theropod dinosaurs. It also provides a valuable insight into the palaeobiology of these now extinct animals. Based on the frequency with which these lesions occur, we hypothesize that tyrannosaurids were commonly infected by a Trichomonas gallinae-like protozoan. For tyrannosaurid populations, the only non-avian dinosaur group that show trichomonosis-type lesions, it is likely that the disease became endemic and spread as a result of antagonistic intraspecific behavior, consumption of prey infected by a Trichomonas gallinae-like protozoan and possibly even cannibalism. The severity of trichomonosis-related lesions in specimens such as Tyrannosaurus rex FMNH PR2081 and Tyrannosaurus rex MOR 980, strongly suggests that these animals died as a direct result of this disease, mostly likely through starvation.     

Activation of the Epstein-Barr virus replicative cycle by human herpesvirus 6.

One common attribute of herpesviruses is the ability to establish latent, life-long infections. The role of virus-virus interaction in viral reactivation between or among herpesviruses has not been studied. Preliminary experiments in our laboratory had indicated that infection of Epstein-Barr virus (EBV) genome-positive human lymphoid cell lines with human herpesvirus 6 (HHV-6) results in EBV reactivation in these cells. To further our knowledge of this complex phenomenon, we investigated the effect of HHV-6 infection on expression of the viral lytic cycle proteins of EBV. Our results indicate that HHV-6 upregulates, by up to 10-fold, expression of the immediate-early Zebra antigen and the diffuse and restricted (85 kDa) early antigens (EA-D and EA-R, respectively) in both EBV producer and nonproducer cell lines (i.e., P3HR1, Akata, and Raji). Maximal EA-D induction was observed at 72 h post-HHV-6 infection. Furthermore, expression of late EBV gene products, namely, the viral capsid antigen (125 kDa) and viral membrane glycoprotein gp350, was also increased in EBV producer cells (P3HR1 and Akata) following infection by HHV-6. By using dual-color membrane immunofluorescence, it was found that most of the cells expressing viral membrane glycoprotein gp350 were also positive for HHV-6 antigens, suggesting a direct effect of HHV-6 replication on induction of the EBV replicative cycle. No expression of late EBV antigens was observed in Raji cells following infection by HHV-6, implying a lack of functional complementation between the deleted form of EBV found in Raji cells and the superinfecting HHV-6. The susceptibility of the cell lines to infection by HHV-6 correlated with increased expression of various EBV proteins in that B95-8 cells, which are not susceptible to HHV-6 infection, did not show an increase in expression of EBV antigens following treatment with HHV-6. Moreover, UV light-irradiated or heat-inactivated HHV-6 had no upregulating effect on the Zebra antigen or EA-D in Raji cells, indicating that infectious virus is required for the observed effects of HHV-6 on these EBV products. These results show that HHV-6, another lymphotropic human herpesvirus, can activate EBV replication and may thus contribute to the pathogenesis of EBV-associated diseases.     

T-cell immune response to human herpesvirus-6 in healthy adults.

The proliferative response of peripheral blood mononuclear cells (PBMC) from healthy adults to human herpesvirus-6 (HHV-6) was examined. It was found that PBMC from 13 of 14 HHV-6-seropositive adults apparently proliferated in response to stimulation with HHV-6 antigen in contrast to the lack of response of cord blood mononuclear cells. In order to confirm the presence of HHV-6-specific memory T cells in the peripheral blood of healthy adults, we established HHV-6-specific T-cell clones from an HHV-6-seropositive individual. CD4+ T-cell clones generated from HHV-6-stimulated PBMC were found to proliferate upon stimulation with HHV-6 in the presence of autologous antigen-presenting cells, but not in response to herpes simplex virus type 1 antigen or mock-infected control antigen. These results indicate that a T-cell immune response against HHV-6 infection is generally present in healthy adult populations.     

Viral interactions in human lymphoid tissue: Human herpesvirus 7 suppresses the replication of CCR5-tropic human immunodeficiency virus type 1 via CD4 modulation

Human immunodeficiency virus (HIV) infection is often accompanied by infection with other pathogens that affect the clinical course of HIV disease. Here, we identified another virus, human herpesvirus 7 (HHV-7) that interferes with HIV type 1 (HIV-1) replication in human lymphoid tissue, where critical events of HIV disease occur. Like the closely related HHV-6, HHV-7 suppresses the replication of CCR5-tropic (R5) HIV-1 in coinfected blocks of human lymphoid tissue. Unlike HHV-6, which affects HIV-1 by upregulating RANTES, HHV-7 did not upregulate any CCR5-binding chemokine. Rather, the inhibition of R5 HIV-1 by HHV-7 was associated with a marked downregulation of CD4, the cellular receptor shared by HHV-7 and HIV-1. HHV-7-induced CD4 downregulation was sufficient for HIV-1 inhibition, since comparable downregulation of CD4 with cyclotriazadisulfonamide, a synthetic macrocycle that specifically modulates expression of CD4, resulted in the suppression of HIV infection similar to that seen in HHV-7-infected tissues. In contrast to R5 HIV-1, CXCR4-tropic (X4) HIV-1 was only minimally suppressed by HHV-7 coinfection. This selectivity in suppression of R5 and X4 HIV-1 is explained by a suppression of HHV-7 replication in X4- but not in R5-coinfected tissues. These results suggest that HIV-1 and HHV-7 may interfere in lymphoid tissue in vivo, thus potentially affecting the progression of HIV-1 disease. Knowledge of the mechanisms of interaction of HIV-1 with HHV-7, as well as with other pathogens that modulate HIV-1 replication, may provide new insights into HIV pathogenesis and lead to the development of new anti-HIV therapeutic strategies.     

Monocytes in Kaposi's sarcoma lesions are productively infected by human herpesvirus 8.

PCR analysis and serological studies demonstrated a close association between Kaposi's sarcoma (KS)-associated herpesvirus, or human herpesvirus 8 (HHV-8), and the development of Kaposi's sarcoma (KS). The majority of the KS cells were shown to be latently infected by the virus. In this study we investigated which type of cell is productively infected in KS lesions. In situ hybridization was performed with strand-specific RNA probes complementary to the sequences coding for the minor capsid protein (VP23) of HHV-8. The VP23 gene is specifically expressed during the lytic or replicative period of the virus life cycle, and therefore it is a useful marker to detect productively infected cells. By in situ hybridization of KS lesions, a strong hybridization signal was detected only in a small subset of the KS cells of the lesions. Simultaneous application of immunohistochemical staining and in situ hybridization identified the virus-replicating cells to be of monocytic origin. Productively infected monocytes may be an important reservoir for transmission of the virus and for the increase and maintenance of the high load of HHV-8 generally observed in nodular KS lesions during late stages of infection.     

Human herpesvirus 6 open reading frame U14 protein and cellular p53 interact with each other and are contained in the virion

A mass spectroscopic analysis of proteins from human herpesvirus 6 (HHV-6)-infected cells showed that the HHV-6 U14 protein coimmunoprecipitated with the tumor suppressor p53. The binding of U14 to p53 was verified by coimmunoprecipitation experiments in both Molt-3 cells infected with HHV-6 and 293 cells cotransfected with U14 and p53 expression vectors. Indirect immunofluorescence assays (IFAs) showed that by 18 h postinfection (hpi) U14 localized to the dot-like structures observed in both the nucleus and cytoplasm where p53 was partly accumulated. Despite Northern blotting evidence that U14 follows late kinetics, the U14 protein was detected immediately after infection (at 3 hpi) by IFA. In addition, by Western blotting, U14 was detected at 0 hpi or in the presence of cycloheximide which completely abolished the expression of IE1 protein. In addition to U14, p53 was detected at 0 hpi although it was not detected in mock-infected cells. Furthermore, both U14 and p53 were clearly detected in the viral particles by Western blotting and immunoelectron microscopy, supporting the idea that U14 and p53 are incorporated into virions. Our study provides the first evidence of the incorporation of cellular p53 into viral particles and suggests that p53 may play an important role in viral infection.     

In vitro susceptibility of T lymphocytes from chimpanzees (Pan troglodytes) to human herpesvirus 6 (HHV-6): a potential animal model to study the interaction between HHV-6 and human immunodeficiency virus type 1 in vivo.

The in vitro susceptibility of several nonhuman primate species to human herpesvirus 6 (HHV-6) was investigated. Only peripheral blood mononuclear cells from chimpanzees (Pan troglodytes) were found permissive to productive infection by HHV-6, indicating that the host range of HHV-6, albeit limited, may not be restricted to Homo sapiens. However, natural HHV-6 infection in chimpanzees, as well as in the other species tested, could not be documented by serological analysis. As previously observed with human cells, HHV-6 infection of chimpanzee peripheral blood mononuclear cells was highly cytopathic and the infected cells exhibited phenotypic features of activated T lymphocytes. Although in humans the majority of HHV-6-infected lymphocytes displayed the CD4 antigen, in chimpanzees a mixed CD4+ and CD8+ phenotype was observed. HHV-6 was also shown to productively coinfect individual chimpanzee T cells with human immunodeficiency virus type 1, resulting in an accelerated induction of cytopathicity. In light of these findings, we propose the utilization of chimpanzees as a potential animal model system to investigate the in vivo interaction between HHV-6 and human immunodeficiency virus type 1 and its relevance to the development of acquired immune deficiency syndrome.     

E6 and E7 from beta HPV38 cooperate with ultraviolet light in the development of actinic keratosis-like lesions and squamous cell carcinoma in mice

Cutaneous beta human papillomavirus (HPV) types appear to be involved in the development of non-melanoma skin cancer (NMSC); however, it is not entirely clear whether they play a direct role. We have previously shown that E6 and E7 oncoproteins from the beta HPV type 38 display transforming activities in several experimental models. To evaluate the possible contribution of HPV38 in a proliferative tissue compartment during carcinogenesis, we generated a new transgenic mouse model (Tg) where HPV38 E6 and E7 are expressed in the undifferentiated basal layer of epithelia under the control of the Keratin 14 (K14) promoter. Viral oncogene expression led to increased cellular proliferation in the epidermis of the Tg animals in comparison to the wild-type littermates. Although no spontaneous formation of tumours was observed during the lifespan of the K14 HPV38 E6/E7-Tg mice, they were highly susceptible to 7,12-dimethylbenz(a)anthracene (DMBA)/12-0-tetradecanoylphorbol-13-acetate (TPA) two-stage chemical carcinogenesis. In addition, when animals were exposed to ultraviolet light (UV) irradiation, we observed that accumulation of p21(WAF1) and cell-cycle arrest were significantly alleviated in the skin of Tg mice as compared to wild-type controls. Most importantly, chronic UV irradiation of Tg mice induced the development of actinic keratosis-like lesions, which are considered in humans as precursors of squamous cell carcinomas (SCC), and subsequently of SCC in a significant proportion of the animals. In contrast, wild-type animals subjected to identical treatments did not develop any type of skin lesions. Thus, the oncoproteins E6 and E7 from beta HPV38 significantly contribute to SCC development in the skin rendering keratinocytes more susceptible to UV-induced carcinogenesis.     

Oral mucosal lesions and temporomandibular joint impairment of elderly people in the South East Local Government Area of Ibadan

Objective: To determine the prevalence of oral mucosal lesions, temporomandibular joint (TMJ) impairment and oral health‐related habits in the elderly people in South East Local Government Area (SELGA) in Ibadan. Background data: SELGA is one of the largest local government areas in Oyo State, Nigeria and has a population of 225 800. Design: A cross‐sectional survey. Methodology: A randomly selected sample of 690 elderly people from 23 wards in SELGA were examined by two trained and calibrated examiners using mirror, probe and natural light. Results: One hundred and fifty‐five (22.46%) of the elderly examined had one or more oral pathology lesions, representing infection‐related swelling, non‐infection‐related swelling, pre‐malignant lesions, denture stomatitis, non‐denture based ulcers, angular cheilitis, geographic tongue, scrotal tongue, lichen planus, hyper‐pigmentation and TMJ impairment. Fifty‐five (35.5%) of these pathologies were infection‐related swellings. Six (3.9%) of the pathologies were denture‐related stomatitis, 14(9.03%) were pre‐malignant lesions, 14(9.03%) were non‐infection‐related swelling, while 27 (17.4%) were TMJ impairment. The rest of the lesions constituted the remaining 25.1%. Only 15.9% of the participants had tobacco‐related habits, 10.7% drank alcohol, whilst 41.6% chewed kola nuts. Conclusion: Oral pathoses prevalence (22.46%) was similar to findings in developed countries (29.9%) but with a reversal of causal factors, infection‐related swellings were more common, whereas pre‐malignant lesions were less common. The infections were mostly complications from periodontal disease, which culminated in abscess formation. Tobacco‐related habits were not highly prevalent among the elderly people in SELGA. However, oral pathoses resulting from nutritional deficiencies were the third most frequent lesions observed.     

Genetic linkage analysis of oral lichen planus in a Chinese family

Oral lichen planus (OLP) is a common oral inflammatory disease affecting about 1-2% of the general adult population. As with European families who are diagnosed with OLP, the Chinese family who we studied was diagnosed with a severe form of oral reticular and erosive lesions; moreover, two of the five affected individuals developed oral cancer at an early age. A whole-genome genotyping scan with linkage analysis was performed using the 10K SNP array to investigate the genetic susceptibility of the Chinese family to OLP, which revealed one maximal nonparametric LOD score of 2.32 (P = 0.0156) for SNP marker rs2372736, defined at the chromosome 3p14-3q13 region encompassing 19 SNPs. Blood samples were obtained from 10 members of the family, which included the grandmother, father and mother, and the children altogether. The grandfather is dead, but the family members remembered he also suffered from the same disease. Chromosome 3p14-3q13 was identified as the candidate gene region for OLP; this information provides a foundation for further identification of the gene responsible for OLP.     

Accumulation of macrophages expressing MRP8 and MRP14 in skin lesions during Leishmania major infection in BALB/c and RAG-2 knockout mice

Migration inhibitory factor-related protein 8 (MRP8) and MRP14 are expressed by myeloid cells and especially known as marker proteins of an immature and inflammatory subtype of macrophages. In this study, we immunohistochemically examined an accumulation of MRP8+ and MRP14+ macrophages in skin lesions during Leishmania major infection in susceptible BALB/c and RAG-2-/- mice. L. major infection caused the development of a nodular type of skin lesion at the infection site in mice and a massive accumulation of macrophages was observed in the lesions at four weeks after the infection. Immunohistochemical analyses showed MRP8+ and MRP14+ macrophages are predominant cell types in the skin lesions in both mouse strains. In contrast, F4/80+ cells, which correspond to mature macrophages, were rarely found in the skin lesions. These data suggest that the accumulation of inflammatory subtype of macrophages in BALB/c mice during L. major infection can be induced without acquired immune responses.     

Prevalence of human herpesvirus-6B in Korean hematopoietic stem cell transplantation recipients

Human herpesvirus-6 (HHV-6) is a major pathogen associated with diseases of recipients of hematopoietic stem cell transplants (HSCT). We have isolated HHV-6 in Korean HSCT recipients and carried out a prospective investigation of its prevalence. We obtained peripheral blood from HSCT recipients who had signs of HHV-6 infection. Cord blood mononuclear cells (CBMC) and Sup-T1 cells were used to culture the HHV-6. Indirect immunofluorescence assays (IFA), and the polymerase chain reaction (PCR) were employed to detect HHV-6. The prevalence of HHV-6 infection in HSCT recipients was calculated on the basis of the PCR results. HHV-6 was isolated from four clinical samples. After culturing the HHV-6 in CBMC, the standard strain and the four clinical isolates were propagated in Sup-T1 cells. The infected cells became grossly enlarged and multinucleate after 7-21 days. The virus was identified primarily on the basis of the morphological changes of the cultured cells, and confirmed by specific IFA with monoclonal antibody to HHV-6. HHV-6 was detected in each sample by PCR with primers specific for the major immediate early gene. Sequencing of the standard strain and PCR products confirmed identification of the HHV-6B variant. By PCR we detected 415 instances of HHV-6 in 3966 samples (14.6% of peripheral blood mononuclear cells and 6.3% of sera), and HHV-6 DNAemia was most frequent from the second to the fourth week after HSCT.