Disseminated cryptococcosis with gastrointestinal involvement and false-negative cryptococcal antigen testing due to postzone phenomenon: a case report and review of the literature.

BACKGROUND: Cryptococcosis is an increasingly common infection given the growing immunocompromised population worldwide. Cryptococcal antigen (CrAg) testing demonstrates excellent sensitivity and specificity and is the mainstay of diagnosis. However, there may be rare instances in which false-negative CrAg results can delay diagnosis and early treatment, which are critical to ensure positive outcomes. CASE PRESENTATION: A 31-year-old man living with HIV/AIDS who was not taking antiretroviral therapy was hospitalized with fever, diarrhea, and headaches. CD4 count on presentation was 71 cells/uL, and HIV viral load was 3,194,949 copies/mL. Serum CrAg testing was initially negative, however CSF CrAg performed several days later was positive at 1:40 and blood and CSF cultures grew Cryptococcus neoformans. Colonoscopy revealed mucosal papules throughout the sigmoid colon, and tissue biopsy showed yeast within the lamina propria consistent with GI cryptococcosis. Given the high burden of disease, the original serum CrAg specimen was serially diluted and subsequently found to be positive at 1:2,560, confirming the postzone phenomenon. CONCLUSION: Cryptococcosis has a wide array of presentations including intraluminal GI disease, as seen in this patient. While serum CrAg testing displays excellent test characteristics, it is important for clinicians to be aware of the rare instances in which false-negative results may occur in the presence of excess antigen, as in this case.

Multiorgan WU Polyomavirus Infection in Bone Marrow Transplant Recipient.

WU polyomavirus (WUPyV) was detected in a bone marrow transplant recipient with severe acute respiratory distress syndrome who died in 2001. Crystalline lattices of polyomavirus-like particles were observed in the patient's lung by electron microscopy. WUPyV was detected in the lung and other tissues by real-time quantitative PCR and identified in the lung and trachea by immunohistochemistry. A subset of WUPyV-positive cells in the lung had morphologic features of macrophages. Although the role of WUPyV as a human pathogen remains unclear, these results clearly demonstrate evidence for infection of respiratory tract tissues in this patient.

WU polyomavirus in respiratory epithelial cells from lung transplant patient with Job syndrome.

We detected WU polyomavirus (WUPyV) in a bronchoalveolar lavage sample from lungs transplanted into a recipient with Job syndrome by using immunoassays specific for the WUPyV viral protein 1. Co-staining for an epithelial cell marker identified most WUPyV viral protein 1-positive cells as respiratory epithelial cells.

Immunohistochemical detection of KI polyomavirus in lung and spleen.

Little is known about the tissue tropism of KI polyomavirus (KIPyV), and there are no studies to date describing any specific cell types it infects. The limited knowledge of KIPyV tropism has hindered study of this virus and understanding of its potential pathogenesis in humans. We describe tissues from two immunocompromised patients that stained positive for KIPyV antigen using a newly developed immunohistochemical assay targeting the KIPyV VP1 (KVP1) capsid protein. In the first patient, a pediatric bone marrow transplant recipient, KVP1 was detected in lung tissue. Double immunohistochemical staining demonstrated that approximately 50% of the KVP1-positive cells were CD68-positive cells of the macrophage/monocyte lineage. In the second case, an HIV-positive patient, KVP1 was detected in spleen and lung tissues. These results provide the first identification of a specific cell type in which KVP1 can be detected and expand our understanding of basic properties and in vivo tropism of KIPyV.

Cache valley virus in a patient diagnosed with aseptic meningitis.

Cache Valley virus was initially isolated from mosquitoes and had been linked to central nervous system-associated diseases. A case of Cache Valley virus infection is described. The virus was cultured from a patient's cerebrospinal fluid and identified with real-time reverse transcription-PCR and sequencing, which also yielded the complete viral coding sequences.

Serologic evidence of frequent human infection with WU and KI polyomaviruses.

WU polyomavirus (WUPyV) and KI polyomavirus (KIPyV) are novel human polyomaviruses. They were originally identified in human respiratory secretions, but the extent of human infection caused by these viruses has not been described to date. To determine the seroepidemiology of WUPyV and KIpyIV, we used an ELISA to screen serum samples from 419 patients at the St. Louis Children's Hospital and Barnes-Jewish Hospital during 2007-2008. The age-stratified deidentified samples were examined for antibodies to the major capsid proteins of WUPyV and KIPyV. Seropositivity for each virus was similar; antibody levels were high in the youngest age group (<6 months), decreased to a nadir in the next age group (6 to <12 months), and then steadily increased with subsequent age groups, eventually reaching a plateau of approximate, equals 80% for WUPyV and approximate, equals 70% for KIPyV. These results demonstrate that both KIPyV and WUPyV cause widespread infection in the human population.

Nyamanini and midway viruses define a novel taxon of RNA viruses in the order Mononegavirales.

Here, we report the sequencing and classification of Nyamanini virus (NYMV) and Midway virus (MIDWV), two antigenically related viruses that were first isolated in 1957 and 1966, respectively. Although these viruses have been cultured multiple times from cattle egrets, seabirds, and their ticks, efforts to classify them taxonomically using conventional serological and electron microscopic approaches have failed completely. We used a random shotgun sequencing strategy to define the genomes of NYMV and MIDWV. Contigs of 11,631 and 11,752 nucleotides, representing the complete genome of NYMV and the near-complete genome of MIDWV, respectively, were assembled. Each virus genome was predicted to carry six open reading frames (ORFs). BLAST analysis indicated that only two of the ORF proteins of each virus, the putative nucleocapsid and polymerase, had detectable sequence similarity to known viral proteins. Phylogenetic analysis of these ORF proteins demonstrated that the closest relatives of NYNV and MIDWV are negative-stranded-RNA viruses in the order Mononegavirales. On the basis of their very limited sequence similarity to known viruses, we propose that NYMV and MIDWV define a novel genus, Nyavirus, in this order.

Nuclear localization sequences in cytomegalovirus capsid assembly proteins (UL80 proteins) are required for virus production: inactivating NLS1, NLS2, or both affects replication to strikingly different extents.

Scaffolding proteins of spherical prokaryotic and eukaryotic viruses have critical roles in capsid assembly. The primary scaffolding components of cytomegalovirus, called the assembly protein precursor (pAP, pUL80.5) and the maturational protease precursor (pPR, pUL80a), contain two nuclear localization sequences (NLS1 and NLS2), at least one of which is required in coexpression experiments to translocate the major capsid protein (MCP, pUL85) into the nucleus. In the work reported here, we have mutated NLS1 and NLS2, individually or together, in human cytomegalovirus (HCMV, strain AD169) bacmid-derived viruses to test their effects on virus replication. Consistent with results from earlier transfection/coexpression experiments, both single-mutant bacmids gave rise to infectious virus but the double mutant did not. In comparisons with the wild-type virus, both mutants showed slower cell-to-cell spread; decreased yields of infectious virus (3-fold lower for NLS1(-) and 140-fold lower for NLS2(-)); reduced efficiency of pAP, pPR, and MCP nuclear translocation (sixfold lower for NLS1(-) and eightfold lower for NLS2(-)); increased amounts of a 120-kDa MCP fragment; and reduced numbers of intranuclear capsids. All effects were more severe for the NLS2(-) mutant than the NLS1(-) mutant, and a distinguishing feature of cells infected with the NLS2(-) mutant was the accumulation of large, UL80 protein-containing structures within the nucleus. We conclude that these NLS assist in the nuclear translocation of MCP during HCMV replication and that NLS2, which is unique to the betaherpesvirus UL80 homologs, may have additional involvements during replication.

The amino-conserved domain of human cytomegalovirus UL80a proteins is required for key interactions during early stages of capsid formation and virus production.

Assembly of many spherical virus capsids is guided by an internal scaffolding protein or group of proteins that are often cleaved and eliminated in connection with maturation and incorporation of the genome. In cytomegalovirus there are at least two proteins that contribute to this scaffolding function; one is the maturational protease precursor (pUL80a), and the other is the assembly protein precursor (pUL80.5) encoded by a shorter genetic element within UL80a. Yeast GAL4 two-hybrid assays established that both proteins contain a carboxyl-conserved domain that is required for their interaction with the major capsid protein (pUL86) and an amino-conserved domain (ACD) that is required for their self-interaction and for their interaction with each other. In the work reported here, we demonstrate that when the ACD is deleted (deltaACD) or disrupted by a point mutation (L47A), the bacterially expressed mutant protein sediments as a monomer during rate-velocity centrifugation, whereas the wild-type protein sediments mainly as oligomers. We also show that the L47A mutation reduces the production of infectious virus by at least 90%, results in the formation of irregular nuclear capsids, gives rise to tube-like structures in the nucleus that resemble the capsid core in cross-section and contain UL80 proteins, slows nuclear translocation of the major capsid protein, and may slow cleavage by the maturational protease. We provide physical corroboration that mutating the ACD disrupts self-interaction of the UL80 proteins and biological support for the proposal that the ACD has a critical role in capsid assembly and production of infectious virus.

Vaccine development against HIV-1: current perspectives and future directions.

The development of an efficacious vaccine against the human immunodeficiency virus (HIV) is of great urgency, because it is accepted that vaccination is the only means capable of controlling the AIDS pandemic. The foundation of HIV vaccine development is the analysis of immune responses during natural infection and the utilization of this knowledge for the development of protective immunization strategies. Initial vaccine development and experimentation are usually in animal models, including murine, feline, and nonhuman primates. Experimental vaccine candidates are closely studied for both efficacy and safety before proceeding to human clinical trials. There are a number of different therapeutic and prophylactic vaccine strategies currently being studied in human clinical trials. Vaccine strategies that are being tested, or have previously been tested, in humans include subunit, DNA plasmid, and viral vector, and combinations of these various strategies. Some of the results of these trials are promising, and additional research has focused on the development of appropriate chemical and genetic adjuvants as well as methods of vaccine delivery to improve the host immune response. This review summarizes the vaccine strategies that have been tested in both animal models and human clinical trials.