bluuebunny/crossref_metadata_2025 huggingface dataset:
Hormonally Induced Spawning, Embryonic Development, and Larval Rearing of the Southern Temperate Banded Morwong, Cheilodactylus spectabilis
Authors: Ritar Arthur J. , Pribadi Tyas A.
Published: November 2006

Banded morwong (Cheilodactylus spectabilis) are of interest for marine finfish aquaculture in temperate southern Australia. To improve their ovulatory response, adult females were implanted during the autumn spawning season with slow‐release pellets containing 0–400 μg luteinizing‐hormone‐releasing hormone analogue (LHRHa)/kg body weight within 24 h of capture from the wild. Compared to the sham control group, animals treated with LHRHa produced significantly more eggs on each day after implantation for the following 7 d (91 ± 39 and 290 ± 38 mL) and a higher proportion ovulated (8/12 and 27/27). Of fish treated with LHRHa, 93% ovulated 2 d after implantation and 79% ovulated three times at 2‐d intervals, whereas control animals showed no cyclicity of ovulation and few ovulated more than once. Egg production was highest at the first ovulation after LHRHa treatment and declined at subsequent ovulations. In a second experiment investigating the range 100–400 μg LHRHa, there was no effect of dose rate on ovulation parameters, which additionally examined implantation either immediately after capture or after a 5‐d delay. Compared to immediate implantation, a delay resulted in a lower proportion of animals that could be stripped after implantation (100 and 50%, respectively) and the volume of eggs was lower (135 ± 15  and 107 ± 10 mL). The egg quality was poor following delayed implantation, resulting in no fertilization after artificial insemination compared with immediate implantation in which fertilization and hatch rates were higher for eggs collected on Day 2 after implantation (79 ± 8% and 58 ± 9%) than on Day 4 (23 ± 7% and 15 ± 6%). Thus, it is important to implant animals as soon as possible after capture to ensure optimum egg quality. Good‐quality eggs were buoyant and spherical and had a diameter of 1050 ± 25 μm with a single pigmented oil droplet of 190 ± 9 μm. When a separate large batch of eggs collected 2 d after implantation with 100 μg LHRHa was inseminated and cultured at 18 C, larvae hatched after 63 ± 2 h at a standard length of 2.6 ± 0.4 mm. Newly hatched larvae were buoyant and transparent with only a few melanophores, eyes were nonpigmented and jaws were nonfunctional. By the fourth day, jaws were functional and eyes were fully pigmented. Utilization of the endogenous yolk and oil was completed by Day 6, and swimming commenced with exogenous feeding. Larvae, initially fed lipid‐enriched rotifers followed by Artemia, reached 8.9 ± 0.7 mm length on Day 55, after which they metamorphosed to the postlarval paperfish stage of development, 22 ± 0.9 mm on Day 100, and 43 ± 1.0 mm at 6 mo of age. The results show that treatment of wild‐caught females with slow‐release pellets containing LHRHa is effective for the production of eggs for hatchery rearing.
Efficacy of Three Methionine Sources in Diets for Pacific White Shrimp, Litopenaeus vannamei
Authors: Forster Ian P. , Dominy Warren G.
Published: November 2006

Abstract.— To examine the need for supplemental dietary methionine and to determine the ability of different methionine sources to meet this need for Pacific white shrimp, Litopenaeus vannamei, a growth trial was conducted. A control diet (30% crude protein) was formulated to be deficient in methionine by inclusion of high levels of dehulled, solvent‐extracted soybean meal and with no fish meal (methionine level was 0.45% of diet as fed, 1.5% of total amino acids). Three test diets were manufactured based on the control diet but supplemented with l‐methionine, dl‐methionine, or an analog of methionine (2‐hydroxy‐4‐methylthio butanoic acid [HMTBA], calcium salt) at the level of 0.5% as fed (equivalent to 1.5% of total amino acids). Cystine was constant among these diets at 0.5% of diet as fed. A commercial feed (30% crude protein) was included as a reference. The culture vessels were flat‐bottomed, cylindrical, black fiberglass tanks (1.5‐m diameter) initially filled with 1000 L seawater and supplied with aeration and seawater from a well at a rate of 2 L/min. At the end of the trial, all surviving shrimp were counted and weighed. The trial was run outdoors, with a mean water temperature of 25.9 C. The mean survival, final weight, growth rate, and feed efficiency of shrimp in the control group were 95.9%, 10.8 g, 0.93 g/wk, and 0.44 g/g, respectively. The mean values of all the test treatments were 96.9%, 11.6 g, 1.00 g/wk, and 0.47 g/g, respectively. The final weight and growth rate of shrimp fed the control diet were significantly lower than those of shrimp fed the methionine‐supplemented diets, indicating that this species has a requirement for dietary methionine. There were no significant differences (P > 0.05) in any of the parameters among the test treatments, indicating that l‐methionine, dl‐methionine, and HMTBA can be used to meet the methionine requirement of this species under these experimental conditions. It is concluded that the three sources of methionine examined in this trial, l‐methionine, dl‐methionine, and HMTBA, are similar in their ability to meet the methionine requirement of this species.
Taurine Supplementation of All‐plant Protein Diets for Rainbow Trout (Oncorhynchus mykiss)
Authors: Gaylord T. Gibson , Teague April M. , Barrows Frederic T.
Published: November 2006

Abstract.— Taurine has been demonstrated to be conditionally indispensable for several carnivorous fish species. Current trends in trout production include decreasing levels of fish‐meal content in feeds, along with faster growing strains of fish. Taurine may be a limiting nutrient in support of elevated planes of growth for rainbow trout. A 9‐wk feeding trial was conducted using a factorial treatment design with protein source (fish meal or plant) and taurine supplementation (four levels) as the main effects. The fish‐meal diet series included 23% herring meal and contained 1.76% total sulfur amino acids (TSAA). The plant diet series did not contain any animal proteins and substituted protein from soy protein concentrate in place of the herring‐meal protein and contained 1.5% TSAA. Taurine was supplemented at 0, 5, 10, and 15 g/kg dry diet to each of the diets in the plant series and the fish‐meal series of diets. All diets were formulated to contain 43.8% crude protein and 20% lipid with an estimated physiological fuel value of 4.2 kcal/g. Fifteen fish were stocked in each of 24 tanks with a mean initial weight of approximately 26.8 g per fish. The unsupplemented fish‐meal diet contained 2 g/kg taurine, and the unsupplemented plant diet had taurine levels below the detection limit of 0.1 g/kg diet. Taurine supplementation improved growth, feed conversion ratios, protein retention efficiencies, and energy retention efficiencies of fish fed the plant protein diets. No effects of taurine supplementation were observed for these response factors in fish fed the fish‐meal series diets. This study demonstrates that taurine supplementation may be necessary for rainbow trout fed plant‐protein‐based feeds.
Characterization of Vibrio spp. Associated with Diseased Shrimp from Culture Ponds of Andhra Pradesh (India)
Authors: Jayasree L. , Janakiram P. , Madhavi R.
Published: November 2006

Abstract.— Surveys undertaken on diseases caused by Vibrio spp. in Penaeus monodon from culture ponds of coastal Andhra Pradesh recorded the occurrence of five types of diseases: tail necrosis, shell disease, red disease, loose shell syndrome (LSS), and white gut disease (WGD). Among these, LSS, WGD, and red disease caused mass mortalities in shrimp culture ponds. Six species of Vibrio—V. harveyi, V. parahaemolyticus, V. alginolyticus, V. anguillarum, V. vulnificus, and V. splendidus—are associated with the diseased shrimp. The number of Vibrio spp. associated with each disease ranged from two to five. Additionally, shrimp with red disease had concurrent infections with white spot syndrome virus. Vibrio harveyi in the case of LSS and WGD, V. parahaemolyticus for red disease, and V. alginolyticus for shell disease are the major etiologcal agents. Differences occur in the degree of virulence of different species of Vibrio and also different isolates of the same species. Vibrio harveyi isolated from LSS shrimp is the most virulent. In general, all the Vibrio isolates from LSS shrimp tend to be more virulent as compared to their counterparts from other diseased shrimp. It is apparent that the degree of virulence of various Vibrio isolates depends on its source and the pond environmental conditions. Most of the Vibrio isolates showed susceptibility to oxytetracycline, norfloxacin, and ciprofloxacin. The luminous V. harveyi exhibited resistance to many antibiotics and susceptibility to only three drugs. Considering the emergence of antimicrobial resistant strains of Vibrio, the need for using probiotics in place of antibiotics for disease control is stressed.
Sufism – What Is It Exactly?
Authors: Heck Paul L.
Published: November 2006

Sufism is commonly called mysticism, the mysticism of Islam, but is it mysticism and what is its relation to Islam? Despite diverse expressions and modernist and reformist attempts to disassociate it from Islam, Sufism is the spirituality of Islam. Sometimes saint based, sometimes text based, it aims to bring the soul into relation with the sanctity of the other world, thus orienting it to divine truth. Sufism thus sees itself as the completion of Islam, its living embodiment, in contrast to legal formalism and theological scholasticism, but not in opposition to Muslim laws and doctrines. Its goal is sanctity, embodiment of the godly holiness described by the Qur’an. It is thus a path to saintliness not as perfection of human virtue but as extension of the prophecy of Islam, standing in an integral relation to the ethical and theological outlook of Islam, with which, this article argues, Sufism is finely interwoven.
C57BL/6 and Congenic Interleukin-10-Deficient Mice Can Serve as Models ofCampylobacter jejuniColonization and Enteritis
Authors: Mansfield L. S. , Bell J. A. , Wilson D. L. , Murphy A. J. , Elsheikha H. M. , Rathinam V. A. K. , Fierro B. R. , Linz J. E. , Young V. B.
Published: November 2006

Campylobacter jejuniis a globally distributed cause of human food-borne enteritis and has been linked to chronic joint and neurological diseases. We hypothesized thatC. jejuni11168 colonizes the gastrointestinal tract of both C57BL/6 mice and congenic C57BL/6 interleukin-10-deficient (IL-10−/−) mice and that C57BL/6 IL-10−/−mice experienceC. jejuni11168-mediated clinical signs and pathology. Individually housed mice were challenged orally withC. jejuni11168, and the course of infection was monitored by clinical examination, bacterial culture,C. jejuni-specific PCR, gross pathology, histopathology, immunohistochemistry, and anti-C. jejuni-specific serology. Ceca ofC. jejuni11168-infected mice were colonized at high rates: ceca of 50/50 wild-type mice and 168/170 IL-10−/−mice were colonized. In a range from 2 to 35 days after infection withC. jejuni11168, C57BL/6 IL-10−/−mice developed severe typhlocolitis best evaluated at the ileocecocolic junction. Rates of colonization and enteritis did not differ between male and female mice. A dose-response experiment showed that as little as 106CFU produced significant disease and pathological lesions similar to responses seen in humans. Immunohistochemical staining demonstratedC. jejuniantigens within gastrointestinal tissues of infected mice. Significant anti-C. jejuniplasma immunoglobulin levels developed by day 28 after infection in both wild-type and IL-10-deficient animals; antibodies were predominantly T-helper-cell 1 (Th1)-associated subtypes. These results indicate that the colonization of the mouse gastrointestinal tract byC. jejuni11168 is necessary but not sufficient for the development of enteritis and that C57BL/6 IL-10−/−mice can serve as models for the study ofC. jejunienteritis in humans.
Cross-Protection against a Human Enteric Coronavirus and a Virulent Bovine Enteric Coronavirus in Gnotobiotic Calves
Authors: Han Myung Guk , Cheon Doo-Sung , Zhang Xuming , Saif Linda J.
Published: November 2006

A group 2 human coronavirus designated HECV-4408 was isolated from a child with acute diarrhea and is antigenically and genetically more closely related to bovine coronavirus (BCoV) than to human coronavirus OC43 (X. M. Zhang, W. Herbst, K. G. Kousoulas, and J. Storz, J. Med. Virol.44:152-161, 1994). To determine whether HECV-4408 infects gnotobiotic calves and induces cross-protective immunity against the virulent enteric BCoV DB2 strain, gnotobiotic calves (n= 4) were orally inoculated with HECV-4408 and then challenged with BCoV DB2 at postinoculation day (PID) 21. All calves inoculated with HECV-4408 developed diarrhea at PID 3 to 4 lasting 5 to 9 days. Fecal and nasal virus shedding were first detected by reverse transcription-PCR at PID 3 to 4 and at PID 2 to 4, respectively. After challenge with bovine coronavirus, no diarrhea or virus shedding was detected in calves inoculated with HECV-4408, but a mock-inoculated calf developed diarrhea and fecal and nasal shedding. Fecal immunoglobulin A (IgA) and serum IgG antibodies were first detected at PID 7 and PID 14, respectively. At postchallenge day 7, serum IgG and fecal IgA antibody titers remained the same or increased only twofold compared to prechallenge titers. An additional two gnotobiotic calves were inoculated with HECV-4408 and euthanized at PID 5. Moderate villous atrophy was observed in the small intestines, and viral antigen was detected in villous enterocytes of the small and large intestines by immunohistochemistry. These results support and extend the previous report that HECV-4408 is likely a variant of bovine coronavirus. They confirm its infectivity for calves and complete cross-protection against a bovine coronavirus (DB2 strain) showing 98.2% amino acid identity to HECV-4408 in the S protein.
Nuclear Export of Simian Immunodeficiency Virus Vpx Protein
Authors: Singhal Prabhat K. , Rajendra Kumar P. , Subba Rao Malireddi R. K. , Mahalingam Sundarasamy
Published: November 2006

Lentiviruses, human immunodeficiency viruses (HIVs), and simian immunodeficiency
viruses (SIVs) are distinguished from oncoretroviruses by their ability
to infect nondividing cells such as macrophages. Retroviruses must gain
access to the host cell nucleus for replication and propagation. HIV
and SIV preintegration complexes (PIC) enter nuclei after traversing
the central aqueous channel of the limiting nuclear pore complex
without membrane breakdown. Among the nucleophilic proteins, namely,
matrix, integrase, Vpx, and Vpr, present in HIV type 2/SIV PIC, Vpx is
implicated in nuclear targeting and is also available for incorporation
into budding virions at the plasma membrane. The mechanisms of these
two opposite functions are not known. We demonstrate that Vpx is a
nucleocytoplasmic shuttling protein and contains two novel noncanonical
nuclear import signals and a leptomycin B-sensitive nuclear export
signal. In addition, Vpx interacts with the cellular tyrosine kinase
Fyn through its C-terminal proline-rich motif. Furthermore, our data
indicate that Fyn kinase phosphorylates Vpx and regulates its export
from nucleus. Replacement of conserved tryptophan residues within
domain 41 to 63 and tyrosine residues at positions 66, 69, and 71 in
Vpx impairs its nuclear export, virion incorporation, and SIV
replication in macrophages. Nuclear export is essential to ensure the
availability of Vpx in the cytoplasm for incorporation into virions,
leading to efficient viral replication within nondividing
cells.
Induction of Robust Immune Responses against Human Immunodeficiency Virus Is Supported by the Inherent Tropism of Adeno-Associated Virus Type 5 forDendritic Cells
Authors: Xin Ke-Qin , Mizukami Hiroaki , Urabe Masashi , Toda Yoshihiko , Shinoda Kaori , Yoshida Atsushi , Oomura Kenji , Kojima Yoshitsugu , Ichino Motohide , Klinman Dennis , Ozawa Keiya , Okuda Kenji
Published: November 2006

The ability of adeno-associated virus serotype 1 to 8 (AAV1 to AAV8) vectors expressing the human immunodeficiency virus type 1 (HIV-1) Env gp160 (AAV-HIV) to induce an immune response was evaluated in BALB/c mice. The AAV5 vector showed a higher tropism for both mouse and human dendritic cells (DCs) than did the AAV2 vector, whereas other AAV serotype vectors transduced DCs only poorly. AAV1, AAV5, AAV7, and AAV8 were more highly expressed in muscle cells than AAV2. An immunogenicity study of AAV serotypes indicates that AAV1, AAV5, AAV7, and AAV8 vectors expressing the Env gp160 gene induced higher HIV-specific humoral and cell-mediated immune responses than the AAV2 vector did, with the AAV5 vector producing the best responses. Furthermore, mice injected with DCs that had been transduced ex vivo with an AAV5 vector expressing the gp160 gene elicited higher HIV-specific cell-mediated immune responses than did DCs transduced with AAV1 and AAV2 vectors. We also found that AAV vectors produced by HEK293 cells and insect cells elicit similar levels of antigen-specific immune responses. These results demonstrate that the immunogenicity of AAV vectors depends on their tropism for both antigen-presenting cells (such as DCs) and non-antigen-presenting cells (such as muscular cells) and that AAV5 is a better vector than other AAV serotypes. These results may aid in the development of AAV-based vaccine and gene therapy.
Kaposi's Sarcoma-Associated Herpesvirus
ori-Lyt
-Dependent DNA Replication: DualRole of Replication and Transcription Activator
Authors: Wang Yan , Tang Qiyi , Maul Gerd G. , Yuan Yan
Published: November 2006

Lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) is
essential for viral propagation and pathogenicity. In Kaposi's sarcoma
lesions, constant lytic replication plays a role in sustaining the
population of latently infected cells that otherwise are quickly lost
by segregation of latent viral episomes as spindle cells divide. Lytic
DNA replication initiates from an origin (
ori-Lyt
) and
requires
trans
-acting elements. Two functional
ori-Lyt
s have been identified in the KSHV genome.
Some
cis
-acting and
trans
-acting elements for
ori-Lyt
-dependent DNA replication have been found. Among
these, K8 binding sites, a cluster of C/EBP binding motifs, and a
replication and transcription activator (RTA) responsive element (RRE)
are crucial
cis
-acting elements. Binding of K8 and RTA
proteins to these motifs in
ori-Lyt
DNA was demonstrated to be
absolutely essential for DNA replication. In the present study,
functional roles of RTA in
ori-Lyt
-dependent DNA replication
have been investigated. Two distinct functions of RTA were revealed.
First, RTA activates an
ori-Lyt
promoter and initiates
transcription across GC-rich tandem repeats. This RTA-mediated
transcription is indispensable for DNA replication. Second, RTA is a
component of the replication compartment, where RTA interacts with
prereplication complexes composed of at least six core machinery
proteins and K8. The prereplication complexes are recruited to
ori-Lyt
DNA through RTA, which interacts with the RRE, as well
as K8, which binds to a cluster of C/EBP binding motifs with the aid of
C/EBP α. The revelation of these two functions of RTA, together
with its role in initiation of a transcriptional cascade that leads to
transcription of all viral lytic genes, shows that RTA is a critical
initiator and regulator of KSHV lytic DNA replication and viral
propagation.
Identification of an Essential Domain in the Herpesvirus VP1/2 Tegument Protein: the Carboxy Terminus Directs Incorporation into Capsid Assemblons
Authors: Lee Joy I-Hsuan , Luxton G. W. Gant , Smith Gregory Allan
Published: November 2006

The herpesvirus tegument is a layer of viral and cellular proteins located between the capsid and envelope of the virion. The VP1/2 tegument protein is critical for the propagation of all herpesviruses examined. Using an infectious clone of the alphaherpesvirus pseudorabies virus, we have made a collection of truncation and in-frame deletion mutations within the VP1/2 gene (UL36) and examined the resulting viruses for spread between cells. We found that the majority of the VP1/2 protein either was essential for virus propagation or did not tolerate large deletions. A recently described amino-terminal deubiquitinase-encoding domain was dispensable for alphaherpesvirus propagation, but the rate of propagation in an epithelial cell line and the frequency of transport in axons of primary sensory neurons were both reduced. We mapped one essential domain to a conserved sequence at the VP1/2 carboxy terminus and demonstrated that this domain sufficient to redirect the green fluorescent protein to capsid assemblons in nuclei of infected cells.
Rotavirus Infection Enhances Lipopolysaccharide-Induced Intussusception in a Mouse Model
Authors: Warfield Kelly L. , Blutt Sarah E. , Crawford Sue E. , Kang Gagandeep , Conner Margaret E.
Published: November 2006

Unexpected reports of intussusception after vaccination with the live tetravalent rotavirus vaccine RotaShield resulted in voluntary withdrawal of the vaccine. Intussusception, a condition in which the intestine acutely invaginates upon itself, is the most common cause of intestinal obstruction in children. We report here the development of a mouse model to study rotavirus-induced intussusception. In this model, both homologous murine and heterologous simian rotavirus strains significantly enhanced the rate of lipopolysaccharide (LPS)-induced intussusception, and this enhancement was replication dependent, requiring rotavirus doses of greater than one 50% infectious dose. Rotavirus-induced intussusceptions did not have observable lymphoid lead points, despite the induction of intestinal lymphoid hyperplasia after rotavirus infection. Intussusceptions are also postulated to result from altered intestinal motility, but rotavirus infection had no effect on gastrointestinal transit. LPS-induced intussusception is associated with the induction of inflammatory mediators, and intussusception rates can be modified by inflammatory antagonists. We show that rotavirus infection significantly enhanced serum tumor necrosis factor alpha and gamma interferon cytokine levels after LPS treatment compared to uninfected mice. Together, these data suggest that rotavirus infection sensitized mice to the inflammatory effects of subsequent LPS treatment to enhance intussusception rates.
Divergences in Protein Activity and Cellular Localization within theCampoletis sonorensisIchnovirus Vankyrin Family
Authors: Kroemer Jeremy A. , Webb Bruce A.
Published: November 2006

Ichnoviruses (IVs) occur in obligate symbiotic associations with endoparasitic ichneumonid wasps. IVs are injected with eggs during parasitization, where viral infection and gene expression alter host physiology to ensure endoparasitoid survival. The sevenCampoletis sonorensisIV (CsIV)vankyringenes encode proteins that possess ankyrin repeat domains resembling the inhibitory domains of NF-κB transcription factor inhibitors (IκBs). The CsIVvankyrinsare divided into two subclasses: those expressed primarily in the host fat body (three genes) and those expressed in host hemocytes (four genes). CsIV vankyrin proteins showed limited antigenic similarity when analyzed by Western blotting. Cellular localization and expression patterns of recombinant vankyrin proteins in High Five and Sf9 insect cells differed within and between the subclasses and in cells exposed to lipopolysaccharide, laminarin, or viral immune challenge. In unstimulated Sf9 cells, five vankyrins were detected in cell nuclei. The remaining two proteins localized predominantly to cytoplasmic granules. Immune stimulation of cells resulted in a nuclear-to-cytoplasmic shift of three vankyrins but did not affect localization of other variants. When expressed from recombinantAutographa californicamultiple nucleopolyhedroviruses (AcMNPVs), all vankyrins showed a nuclear localization during early stages of infection with patterns resembling those of immune-challenged cells as the infection progressed. Two fat body vankyrins also produced unique biological effects when expressed from recombinant AcMNPV. Insect cells infected with these viruses exhibited enhanced longevity compared to those infected with viruses expressing other vankyrins. Together, these data suggest that vankyrin proteins in CsIV have divergent physiological functions.
Characterization of a Novel 5′ Subgenomic RNA3a Derived from RNA3 of Brome Mosaic Bromovirus
Authors: Wierzchoslawski Rafal , Urbanowicz Anna , Dzianott Aleksandra , Figlerowicz Marek , Bujarski Jozef J.
Published: November 2006

The synthesis of 3′ subgenomic RNA4 (sgRNA4) by initiation from an internal sg promoter in the RNA3 segment was first described for Brome mosaic bromovirus (BMV), a model tripartite positive-sense RNA virus (W. A. Miller, T. W. Dreher, and T. C. Hall, Nature 313:68-70, 1985). In this work, we describe a novel 5′ sgRNA of BMV (sgRNA3a) that we propose arises by premature internal termination and that encapsidates in BMV virions. Cloning and sequencing revealed that, unlike any other BMV RNA segment, sgRNA3a carries a 3′ oligo(A) tail, in which respect it resembles cellular mRNAs. Indeed, both the accumulation of sgRNA3a in polysomes and the synthesis of movement protein 3a in in vitro systems suggest active functions of sgRNA3a during protein synthesis. Moreover, when copied in the BMV replicase in vitro reaction, the minus-strand RNA3 template generated the sgRNA3a product, likely by premature termination at the minus-strand oligo(U) tract. Deletion of the oligo(A) tract in BMV RNA3 inhibited synthesis of sgRNA3a during infection. We propose a model in which the synthesis of RNA3 is terminated prematurely near the sg promoter. The discovery of 5′ sgRNA3a sheds new light on strategies viruses can use to separate replication from the translation functions of their genomic RNAs.
Functional Characterization of Kaposi's Sarcoma-Associated Herpesvirus ORF45 byBacterial Artificial Chromosome-Based Mutagenesis
Authors: Zhu Fan Xiu , Li Xiaojuan , Zhou Fuchun , Gao Shou-Jiang , Yuan Yan
Published: November 2006

Open
reading frame 45 (ORF45) of Kaposi's sarcoma-associated herpesvirus
(KSHV) encodes an immediate-early protein. This protein is also present
in virions as a tegument protein. ORF45 protein interacts with
interferon regulatory factor 7 (IRF-7) and inhibits virus-induced type
I interferon production by blocking activation of IRF-7. To define
further the function of ORF45 and the mechanism underlying its action,
we constructed an ORF45-null recombinant virus genome (BAC-stop45) by
using a bacterial artificial chromosome (BAC) system. Stable 293T cells
carrying the BAC36 (wild type) and BAC-stop45 genomes were generated.
When monolayers of 293T BAC36 and 293T BAC-stop45 cells were induced
with 12-
O
-tetradecanoylphorbol-13-acetate and sodium butyrate,
no significant difference was found between them in overall viral gene
expression and lytic DNA replication, but induced 293T BAC-stop45 cells
released 10-fold fewer virions to the medium than did 293T BAC36 cells.
When ORF45-null virus was used to infect cells, lower infectivity was
observed than for wild-type BAC36. These results suggest that KSHV
ORF45 plays roles in both early and late stages of viral infection,
probably in viral ingress and
egress.
The African Swine Fever Virus Nonstructural Protein pB602L Is Required for Formation of the Icosahedral Capsid of the Virus Particle
Authors: Epifano Carolina , Krijnse-Locker Jacomine , Salas María L. , Rodríguez Javier M. , Salas José
Published: November 2006

African swine fever virus (ASFV) protein pB602L has been described as a molecular chaperone for the correct folding of the major capsid protein p72. We have studied the function of protein pB602L during the viral assembly process by using a recombinant ASFV, vB602Li, which inducibly expresses the gene coding for this protein. We show that protein pB602L is a late nonstructural protein, which, in contrast with protein p72, is excluded from the viral factory. Repression of protein pB602L synthesis inhibits the proteolytic processing of the two viral polyproteins pp220 and pp62 and leads to a decrease in the levels of protein p72 and a delocalization of the capsid protein pE120R. As shown by electron microscopy analysis of cells infected with the recombinant virus vB602Li, the viral assembly process is severely altered in the absence of protein pB602L, with the generation of aberrant “zipper-like” structures instead of icosahedral virus particles. These “zipper-like” structures are similar to those found in cells infected under restrictive conditions with the recombinant virus vA72 inducibly expressing protein p72. Immunoelectron microscopy studies show that the abnormal forms generated in the absence of protein pB602L contain the inner envelope protein p17 and the two polyproteins but lack the capsid proteins p72 and pE120R. These findings indicate that protein pB602L is essential for the assembly of the icosahedral capsid of the virus particle.
Adenovirus Fiber Shaft Contains a Trimerization Element That Supports Peptide Fusion for Targeted Gene Delivery
Authors: Li Jiali , Lad Sonya , Yang Guang , Luo Yunping , Iacobelli-Martinez Milena , Primus F. James , Reisfeld Ralph A. , Li Erguang
Published: November 2006

Adenoviral (Ad) vectors have been widely used in human gene therapy clinical trials. However, their application has frequently been restricted by the unfavorable expression of cell surface receptors critical for Ad infection. Infections by Ad2 and Ad5 are largely regulated by the elongated fiber protein that mediates its attachment to a cell surface receptor, coxsackie and adenovirus receptor (CAR). The fiber protein is a homotrimer consisting of an N-terminal tail, a long shaft, and a C-terminal knob region that is responsible for high-affinity receptor binding and Ad tropism. Consequently, the modification of the knob region, including peptide insertion and C-terminal fusion of ligands for cell surface receptors, has become a major research focus for targeting gene delivery. Such manipulation tends to disrupt fiber assembly since the knob region contains a stabilization element for fiber trimerization. We report here the identification of a novel trimerization element in the Ad fiber shaft. We demonstrate that fiber fragments containing the N-terminal tail and shaft repeats formed stable trimers that assembled onto Ad virions independently of the knob region. This fiber shaft trimerization element (FSTE) exhibited a capacity to support peptide fusion. We showed that Ad, modified with a chimeric protein by direct fusion of the FSTE with a growth factor ligand or a single-chain antibody, delivered a reporter gene selectively. Together, these results indicate that the shaft region of Ad fiber protein contains a trimerization element that allows ligand fusion, which potentially broadens the basis for Ad vector development.
Epstein-Barr Virus Selectively Deregulates DNA Damage Responses in Normal B Cells but Has No Detectable Effect on Regulation of the Tumor Suppressor p53
Authors: O'Nions Jenny , Turner Abigail , Craig Richard , Allday Martin J.
Published: November 2006

To determine whether latent Epstein-Barr virus (EBV) modifies DNA damage responses in B lymphocytes, cells were treated with agents either producing DNA cross-links and adducts or generating double-strand breaks. The cyclin-dependent kinase inhibitor p21
WAF1
accumulated in mitogen-stimulated primary B cells following exposure to all genotoxins tested. In contrast, when proliferation was EBV driven, p21
WAF1
failed to accumulate after treatment with the DNA adduct-producing agents. The tumor suppressor p53 was stabilized and phosphorylated after all treatments, irrespective of whether latent EBV was present. This suggests that regulatory pathways upstream of p53 are unaffected by latent EBV but downstream effectors are altered if DNA adducts or distortions are involved.
A New Group B Adenovirus Receptor Is Expressed at High Levels on Human Stem and Tumor Cells
Authors: Tuve Sebastian , Wang Hongjie , Ware Carol , Liu Ying , Gaggar Anuj , Bernt Kathrin , Shayakhmetov Dmitry , Li ZongYi , Strauss Robert , Stone Daniel , Lieber André
Published: November 2006

CD46
is used by human group B adenoviruses (Ads) as a high-affinity
attachment receptor. Here we show evidence that several group B Ads
utilize an additional receptor for infection of human cells, which is
different from CD46. We tentatively named this receptor receptor X.
Competition studies with unlabeled and labeled Ads, recombinant Ad
fiber knobs, and soluble CD46 and CD46 antibodies revealed three
different subgroups of group B Ads, in terms of their receptor usage.
Group I (Ad16, -21, -35, and -50) nearly exclusively uses CD46. Group
II (Ad3, -7p, and -14) utilizes receptor X and not CD46. Group III
(Ad11p) uses both CD46 and the alternative receptor X. Interaction of
group II and III Ads with receptor X occurs via the fiber knob.
Receptor X is an abundantly expressed glycoprotein that interacts with
group II and III Ads at relatively low affinity in a
Ca
2+
-dependent manner. This receptor is expressed at
high levels on human mesenchymal and undifferentiated embryonic stem
cells, as well as on human cancer cell lines. These findings have
practical implications for stem cell and gene
therapy.
Rotavirus Nonstructural Glycoprotein NSP4 Is Secreted from the Apical Surfaces of Polarized Epithelial Cells
Authors: Bugarc̀ić Andrea , Taylor John A.
Published: November 2006

NSP4, a nonstructural glycoprotein encoded by rotavirus, is involved in the morphogenesis of virus particles in the endoplasmic reticulum of infected cells. NSP4 is also implicated in the pathophysiology of rotavirus-induced diarrhea by acting as an enterotoxin. To mediate enterotoxic effects in vivo, NSP4 must be secreted or released from rotavirus-infected cells in a soluble form; however, previous studies have indicated that NSP4 is a transmembrane glycoprotein localized within endomembrane compartments in infected cells. In this study, we examined the fate of NSP4 synthesized in Caco-2 cells infected with bovine rotavirus. Our studies reveal that NSP4 is actively secreted into the culture medium, preferentially from the infected-cell apical surface. The secretion of NSP4 is dramatically inhibited by brefeldin A and monensin, suggesting that a Golgi-dependent pathway is involved in release of the protein. In agreement with the proposed involvement of the Golgi apparatus during secretion, secreted NSP4 appears to undergo additional posttranslational modification compared to its cell-associated counterpart and is partially resistant to deglycosylation by endoglycosidase H. Our experiments identify a novel, soluble form of NSP4 secreted from virus-infected cells with the potential to carry out the enterotoxigenic role previously attributed to recombinant forms of the protein.
Gene Structure and Expression of Kaposi's Sarcoma-Associated Herpesvirus ORF56, ORF57, ORF58, and ORF59
Authors: Majerciak Vladimir , Yamanegi Koji , Zheng Zhi-Ming
Published: November 2006

Though similar to those of herpesvirus saimiri and Epstein-Barr virus (EBV), the Kaposi's sarcoma-associated herpesvirus (KSHV) genome features more splice genes and encodes many genes with bicistronic or polycistronic transcripts. In the present study, the gene structure and expression of KSHV ORF56 (primase), ORF57 (MTA), ORF58 (EBV BMRF2 homologue), and ORF59 (DNA polymerase processivity factor) were analyzed in butyrate-activated KSHV
+
JSC-1 cells. ORF56 was expressed at low abundance as a bicistronic ORF56/57 transcript that utilized the same intron, with two alternative branch points, as ORF57 for its RNA splicing. ORF56 was transcribed from two transcription start sites, nucleotides (nt) 78994 (minor) and 79075 (major), but selected the same poly(A) signal as ORF57 for RNA polyadenylation. The majority of ORF56 and ORF57 transcripts were cleaved at nt 83628, although other nearby cleavage sites were selectable. On the opposite strand of the viral genome, colinear ORF58 and ORF59 were transcribed from different transcription start sites, nt 95821 (major) or 95824 (minor) for ORF58 and nt 96790 (minor) or 96794 (major) for ORF59, but shared overlapping poly(A) signals at nt 94492 and 94488. Two cleavage sites, at nt 94477 and nt 94469, could be equally selected for ORF59 polyadenylation, but only the cleavage site at nt 94469 could be selected for ORF58 polyadenylation without disrupting the ORF58 stop codon immediately upstream. ORF58 was expressed in low abundance as a monocistronic transcript, with a long 5′ untranslated region (UTR) but a short 3′ UTR, whereas ORF59 was expressed in high abundance as a bicistronic transcript, with a short 5′ UTR and a long 3′ UTR similar to those of polycistronic ORF60 and ORF62. Both ORF56 and ORF59 are targets of ORF57 and were up-regulated significantly in the presence of ORF57, a posttranscriptional regulator.
Walleye Dermal Sarcoma Virus Retroviral Cyclin Directly Contacts TAF9
Authors: Rovnak Joel , Quackenbush Sandra L.
Published: November 2006

Walleye dermal sarcoma virus (WDSV) is a complex retrovirus associated with dermal sarcomas in walleye fish. A WDSV accessory gene encodes a cyclin homolog or retroviral cyclin (rv-cyclin). WDSV rv-cyclin was found to be associated with transcription complexes and to affect transcription in a cell-type and promoter-dependent manner. It inhibited the WDSV promoter in walleye fibroblasts and activated transcription from GAL4 promoters when fused to the GAL4 DNA binding domain, and an activation domain (AD) has been localized to 30 amino acids in the carboxyl region. rv-cyclin can block the pulldown of transcription coactivators by the AD of VP16, and the isolated rv-cyclin AD interferes specifically with the interaction between the carboxyl halves of the VP16 AD, VP16C, and TATA-binding protein-associated factor 9 (TAF9). The carboxyl region and isolated AD can bind TAF9 directly in assays of protein-protein interaction in vitro. Furthermore, rv-cyclin and the isolated rv-cyclin AD interfere specifically with the function of VP16C in transcription assays. A previously identified motif within the VP16C sequence mediates TAF9 binding, and this motif is present in the activation domains of a variety of TAF9-binding transcriptional activators. A similar motif is present in the rv-cyclin AD, and point mutations within this motif affect rv-cyclin function and protein-protein interactions. The results support a model of transcription regulation by direct interaction with TAF9.
Conformational Changes in the VP1-Unique Region of Native Human Parvovirus B19 Lead to Exposure of Internal Sequences That Play a Role in Virus Neutralization and Infectivity
Authors: Ros Carlos , Gerber Marco , Kempf Christoph
Published: November 2006

The unique region of the capsid protein VP1 (VP1u) of human parvovirus B19 (B19) elicits a dominant immune response and has a phospholipase A
2
(PLA
2
) activity, which is necessary for the infection. In contrast to the rest of the parvoviruses, the VP1u of B19 is thought to occupy an external position in the virion, making this region a promising candidate for vaccine development. By using a monoclonal antibody against the most-N-terminal portion of VP1u, we revealed that this region rich in neutralizing epitopes is not accessible in native capsids. However, exposure of capsids to increasing temperatures or low pH led to its progressive accessibility without particle disassembly. Although unable to bind free virus or to block virus attachment to the cell, the anti-VP1u antibody was neutralizing, suggesting that the exposure of the epitope and the subsequent virus neutralization occur only after receptor attachment. The measurement of the VP1u-associated PLA
2
activity of B19 capsids revealed that this region is also internal but becomes exposed in heat- and in low-pH-treated particles. In sharp contrast to native virions, the VP1u of baculovirus-derived B19 capsids was readily accessible in the absence of any treatment. These results indicate that stretches of VP1u of native B19 capsids harboring neutralizing epitopes and essential functional motifs are not external to the capsid. However, a conformational change renders these regions accessible and triggers the PLA
2
potential of the virus. The results also emphasize major differences in the VP1u conformation between natural and recombinant particles.
Contribution of T-Cell Receptor Repertoire Breadth to the Dominance of Epitope-Specific CD8+T-Lymphocyte Responses
Authors: Manuel Edwin R. , Charini William A. , Sen Pritha , Peyerl Fred W. , Kuroda Marcelo J. , Schmitz Jörn E. , Autissier Patrick , Sheeter Dennis A. , Torbett Bruce E. , Letvin Norman L.
Published: November 2006

Dominant epitope-specific CD8+T-lymphocyte responses play a central role in controlling viral spread. We explored the basis for the development of this focused immune response in simian immunodeficiency virus (SIV)- and simian-human immunodeficiency virus (SHIV)-infected rhesus monkeys through the use of two dominant (p11C and p199RY) and two subdominant (p68A and p56A) epitopes. Using real-time PCR to quantitate T-cell receptor (TCR) variable region beta (Vβ) family usage, we show that CD8+T-lymphocyte populations specific for dominant epitopes are characterized by a diverse Vβ repertoire, whereas those specific for subdominant epitopes employ a dramatically more focused Vβ repertoire. We also demonstrate that dominant epitope-specific CD8+T lymphocytes employ TCRs with multiple CDR3 lengths, whereas subdominant epitope-specific cells employ TCRs with a more restricted CDR3 length. Thus, the relative dominance of an epitope-specific CD8+T-lymphocyte response reflects the clonal diversity of that response. These findings suggest that the limited clonal repertoire of subdominant epitope-specific CD8+T-lymphocyte populations may limit the ability of these epitope-specific T-lymphocyte populations to expand and therefore limit the ability of these cell populations to contribute to the control of viral replication.
Retrotransposon Suicide: Formation of Ty1 Circles and Autointegration via a Central DNA Flap
Authors: Garfinkel David J. , Stefanisko Karen M. , Nyswaner Katherine M. , Moore Sharon P. , Oh Jangsuk , Hughes Stephen H.
Published: November 2006

Despite their evolutionary distance, the
Saccharomyces cerevisiae
retrotransposon Ty1 and retroviruses use similar strategies for replication, integration, and interactions with their hosts. Here we examine the formation of circular Ty1 DNA, which is comparable to the dead-end circular products that arise during retroviral infection. Appreciable levels of circular Ty1 DNA are present with one-long terminal repeat (LTR) circles and deleted circles comprising major classes, while two-LTR circles are enriched when integration is defective. One-LTR circles persist when homologous recombination pathways are blocked by mutation, suggesting that they result from reverse transcription. Ty1 autointegration events readily occur, and many are coincident with and dependent upon DNA flap structures that result from DNA synthesis initiated at the central polypurine tract. These results suggest that Ty1-specific mechanisms minimize copy number and raise the possibility that special DNA structures are a targeting determinant.
Increased Frequency of Circulating CCR5+CD4+T Cells in Human Immunodeficiency Virus Type 2 Infection
Authors: Soares Rui , Foxall Russell , Albuquerque Adriana , Cortesão Catarina , Garcia Miguel , Victorino Rui M. M. , Sousa Ana E.
Published: November 2006

CCR5 expression determines susceptibility to infection, cell tropism, and the rate of human immunodeficiency virus type 1 (HIV-1) disease progression. CCR5 is also considered the major HIV-2 coreceptor in vivo, in spite of broad coreceptor use in vitro. Here we report a significantly increased proportion of memory-effector CD4 T cells expressing CCR5 in HIV-2-infected patients correlating with CD4 depletion. Moreover, HIV-2 proviral DNA was essentially restricted to memory-effector CD4, suggesting that this is the main target for HIV-2. Similar levels of proviral DNA were found in the two infection categories. Thus, the reduced viremia and slow rate of CD4 decline that characterize HIV-2 infection seem to be unrelated to coreceptor availability.
The Betaretrovirus Mason-Pfizer Monkey Virus Selectively Excludes Simian APOBEC3G from Virion Particles
Authors: Doehle Brian P. , Bogerd Hal P. , Wiegand Heather L. , Jouvenet Nolwenn , Bieniasz Paul D. , Hunter Eric , Cullen Bryan R.
Published: November 2006

The APOBEC3 protein family can constitute a potent barrier to the successful infection of mammalian species by retroviruses. Therefore, any retrovirus that has evolved the ability to replicate in a given animal must have developed mechanisms that allow it to avoid or inhibit the APOBEC3 proteins expressed in that animal. Here, we demonstrate that Mason-Pfizer monkey virus (MPMV) is resistant to inhibition by the APOBEC3G protein expressed in its normal host, the rhesus macaque, but highly susceptible to inhibition by murine APOBEC3 (mA3). MPMV virion particles fail to package rhesus APOBEC3G (rA3G), and MPMV Gag binds rA3G poorly in coexpressing cells. In contrast, MPMV virions package mA3 efficiently and MPMV Gag-mA3 complexes are readily detected. Moreover, mA3, but not rA3G, partially colocalizes with MPMV Gag in the cytoplasm of coexpressing cells. Previously, we have demonstrated that murine leukemia virus also escapes inhibition by APOBEC3 proteins by avoiding virion incorporation of its cognate APOBEC3 protein, mA3, yet is inhibited by primate APOBEC3G proteins, which it packages effectively (B. P. Doehle, A. Schäfer, H. L. Wiegand, H. P. Bogerd, and B. R. Cullen, J. Virol. 79:8201-8207, 2005). The finding that two essentially unrelated beta- and gammaretroviruses use similar mechanisms to escape inhibition by the APOBEC3 proteins found in their normal host species suggests that the selective exclusion of APOBEC3 proteins from virion particles may be a general mechanism used by simple mammalian retroviruses.
Proviral Progeny of Heterodimeric Virions Reveal a High Crossover Rate for Human Immunodeficiency Virus Type 2
Authors: Mukherjee Sayandip , Lee Hui-Ling Rose , Ron Yacov , Dougherty Joseph P.
Published: November 2006

Human immunodeficiency virus type 1 (HIV-1), the causative agent of AIDS in humans, exhibits a very high rate of recombination. Bearing in mind the significant epidemiological and clinical contrast between HIV-2 and HIV-1 as well as the critical role that recombination plays in viral evolution, we examined the nature of HIV-2 recombination. Towards this end, a strategy was devised to measure the rate of crossover of HIV-2 by evaluating recombinant progeny produced exclusively by heterodimeric virions. The results showed that HIV-2 exhibits a crossover rate similar to that of HIV-1 and murine leukemia virus, indicating that the extremely high rate of crossover is a common retroviral feature.