bluuebunny/crossref_metadata_2025 huggingface dataset:
Early Diagnosis and Treatment of Patients with Symptomatic Acute Q Fever Do Not Prohibit IgG Antibody Responses to Coxiella burnetii
Authors: Wielders C. C. H. , Kampschreur L. M. , Schneeberger P. M. , Jager M. M. , Hoepelman A. I. M. , Leenders A. C. A. P. , Hermans M. H. A. , Wever P. C.
Published: August 2012

Little is known about the effect of timing of antibiotic treatment on development of IgG antibodies following acute Q fever. We studied IgG antibody responses in symptomatic patients diagnosed either before or during development of the serologic response toCoxiella burnetii. Between 15 and 31 May 2009, 186 patients presented with acute Q fever, of which 181 were included in this retrospective study: 91 early-diagnosed (ED) acute Q fever patients, defined as negative IgM phase II enzyme-linked immunosorbent assay (ELISA) and positive PCR, and 90 late-diagnosed (LD) acute Q fever patients, defined as positive/dubious IgM phase II ELISA and positive immunofluorescence assay (IFA). Follow-up serology at 3, 6, and 12 months was performed using IFA (IgG phase I and II). High IgG antibody titers were defined as IgG phase II titers of ≥1:1,024 together with IgG phase I titers of ≥1:256. At 12 months, 28.6% of ED patients and 19.5% of LD patients had high IgG antibody titers (P= 0.17). No statistically significant differences were found in frequencies of IgG phase I and IgG phase II antibody titers at all follow-up appointments for adequately and inadequately treated patients overall, as well as for ED and LD patients analyzed separately. Additionally, no significant difference was found in frequencies of high antibody titers and between early (treatment started within 7 days after seeking medical attention) and late timing of treatment. This study indicates that early diagnosis and antibiotic treatment of acute Q fever do not prohibit development of the IgG antibody response.
Pilot Phase IB Clinical Trial of an Alhydrogel-Adsorbed Recombinant Ricin Vaccine
Authors: Vitetta Ellen S. , Smallshaw Joan E. , Schindler John
Published: August 2012

There is no FDA-approved vaccine for the potent plant toxin ricin. We have developed a recombinant ricin vaccine, RiVax. Without adjuvant it is safe and immunogenic in mice, rabbits, and humans. Based on our studies in mice, we now report the results of a small clinical trial with Alhydrogel-adsorbed RiVax.
Evaluation of Gamma Interferon and Antibody Tuberculosis Tests in Alpacas
Authors: Rhodes Shelley , Holder Tom , Clifford Derek , Dexter Ian , Brewer Jacky , Smith Noel , Waring Laura , Crawshaw Tim , Gillgan Steve , Lyashchenko Konstantin , Lawrence John , Clarke John , de la Rua-Domenech Ricardo , Vordermeier Martin
Published: August 2012

We describe the performance of cell-based and antibody blood tests for the antemortem diagnosis of tuberculosis (TB) in South American camelids (SAC). The sensitivity and specificity of the gamma interferon (IFN-γ) release assay, two lateral flow rapid antibody tests (Stat-Pak and Dual Path Platform [DPP]), and two enzyme-linked immunosorbent assay (ELISA)-based antibody tests (Idexx and Enferplex) were determined using diseased alpacas fromMycobacterium bovisculture-confirmed breakdown herds and TB-free alpacas from geographical areas with no history of bovine TB, respectively. Our results show that while the sensitivities of the IFN-γ and antibody tests were similar (range of 57.7% to 66.7%), the specificity of the IFN-γ test (89.1%) was lower than those of any of the antibody tests (range of 96.4% to 97.4%). This lower specificity of the IFN-γ test was at least in part due to undisclosedMycobacterium microtiinfection in the TB-free cohort, which stimulates a positive purified protein derivative (PPD) response. The sensitivity of infection detection could be increased by combining two antibody tests, but even the use of all four antibody tests failed to detect all diseased alpacas. These antibody-negative alpacas were IFN-γ positive. We found that the maximum sensitivity could be achieved only by the combination of the IFN-γ test with two antibody tests in a “test package,” although this resulted in decreased specificity. The data from this evaluation of tests with defined sensitivity and specificity provide potential options for antemortem screening of SAC for TB in herd breakdown situations and could also find application in movement testing and tracing investigations.
Microbiome Diversity in the Bronchial Tracts of Patients with Chronic Obstructive Pulmonary Disease
Authors: Cabrera-Rubio Raúl , Garcia-Núñez Marian , Setó Laia , Antó Josep M. , Moya Andrés , Monsó Eduard , Mira Alex
Published: August 2012

Culture of bacteria from bronchial secretions in respiratory patients has low sensitivity and does not allow for complete assessment of microbial diversity across different bronchial compartments. In addition, a significant number of clinical studies are based on sputum samples, and it is not known to what extent they describe the real diversity of the mucosa. In order to identify previously unrecognized lower airway bacteria and to investigate the complexity and distribution of microbiota in patients with chronic obstructive pulmonary disease (COPD), we performed PCR amplification and pyrosequencing of the 16S rRNA gene in patients not showing signs or symptoms of infection. Four types of respiratory samples (sputum, bronchial aspirate, bronchoalveolar lavage, and bronchial mucosa) were taken from each individual, obtaining on average >1,000 16S rRNA sequences per sample. The total number of genera per patient was >100, showing a high diversity, with
Streptococcus
,
Prevotella
,
Moraxella
,
Haemophilus
,
Acinetobacter
,
Fusobacterium
, and
Neisseria
being the most commonly identified. Sputum samples showed significantly lower diversity than the other three sample types. Lower-bronchial-tree samples, i.e., bronchoalveolar lavage and bronchial mucosa, showed a very similar bacterial compositions in contrast to sputum and bronchial aspirate samples. Thus, sputum and bronchial aspirate samples are upper bronchial tree samples that are not representative of the lower bronchial mucosa flora, and bronchoalveolar lavage samples showed the results closest to those for the bronchial mucosa. Our data confirm that the bronchial tree is not sterile in COPD patients and support the existence a different microbiota in the upper and lower compartments.
Standardized Methods and Quality Control Limits for Agar and Broth Microdilution Susceptibility Testing of Mycoplasma pneumoniae, Mycoplasma hominis, and Ureaplasma urealyticum
Authors: Waites Ken B. , Duffy Lynn B. , Bébéar Cécile M. , Matlow Anne , Talkington Deborah F. , Kenny George E. , Totten Patricia A. , Bade Donald J. , Zheng Xiaotian , Davidson Maureen K. , Shortridge Virginia D. , Watts Jeffrey L. , Brown Steven D.
Published: August 2012

An international multilaboratory collaborative study was conducted to develop standard media and consensus methods for the performance and quality control of antimicrobial susceptibility testing ofMycoplasma pneumoniae,Mycoplasma hominis, andUreaplasma urealyticumusing broth microdilution and agar dilution techniques. A reference strain from the American Type Culture Collection was designated for each species, which was to be used for quality control purposes. Repeat testing of replicate samples of each reference strain by participating laboratories utilizing both methods and different lots of media enabled a 3- to 4-dilution MIC range to be established for drugs in several different classes, including tetracyclines, macrolides, ketolides, lincosamides, and fluoroquinolones. This represents the first multilaboratory collaboration to standardize susceptibility testing methods and to designate quality control parameters to ensure accurate and reliable assay results for mycoplasmas and ureaplasmas that infect humans.
Nostrils of Healthy Volunteers Are Independent with Regard to Staphylococcus aureus Carriage
Authors: Kildow Beau J. , Conradie Johan P. , Robson Rachel L.
Published: August 2012

The right and left nares of healthy adults (
n
= 251) were swabbed separately to determine carriage of
Staphylococcus aureus
in each nostril. Carriers were significantly more likely to carry
S. aureus
in one nostril than in both. Of those carrying
S. aureus
in both nostrils, 20% carried genetically distinct strains in each. Nostrils belonging to a single individual should not be assumed to be homogenous with respect to carriage of
S. aureus
.
A Novel Dirofilaria Species Causing Human and Canine Infections in Hong Kong
Authors: To Kelvin K. W. , Wong Samson S. Y. , Poon Rosana W. S. , Trendell-Smith Nigel J. , Ngan Antonio H. Y. , Lam Jacky W. K. , Tang Tommy H. C. , AhChong Ah-Kian , Kan Joshua Chi-Hang , Chan Kwok-Hung , Yuen Kwok-Yung
Published: August 2012

Dirofilariasis is globally the commonest manifestation of zoonotic filariasis. We report the detection of a novel canine species causing human and canine dirofilariasis in Hong Kong. Three human cases occurring over 10 months were identified, one presenting with cervical lymphadenopathy, one with an abdominal subcutaneous mass, and one with a subconjunctival nodule. Transected worms recovered from the resected abdominal subcutaneous mass were morphologically compatible withDirofilaria. Thecox1gene sequences of the three human isolates were identical; however, they were only 96.2% and 89.3% identical to thecox1gene ofDirofilaria repensandDirofilaria immitis, respectively. Sequencing of the 18S-ITS1-5.8S gene cluster was successful in the intact worm, and the nucleotide sequences were 94.0% and 94.9% identical to those ofD. repensandD. immitis, respectively. Screening of the blood samples from 200 dogs and 100 cats showed the presence of the novelDirofilariaspecies in 3% (6/200) of the dogs' but none of the cats' blood samples. Nucleotide sequences of thecox1gene and 18S-ITS1-5.8S gene clusters of the dogs' samples were identical to those in the human samples. The sera of canines infected by this novelDirofilariaspecies were negative when tested with the SNAP 4DxD. immitisdetection kit, except in the case of dogs with a mixed infection withD. immitisas detected by PCR. The results from this study suggest that this novelDirofilariaspecies is a cause of filarial infection in humans and dogs in Hong Kong. We propose to name thisDirofilariaspecies “Candidatus Dirofilaria hongkongensis.”
Cutaneous Phaeohyphomycosis Caused by Paraconiothyrium cyclothyrioides
Authors: Gordon Rachel A. , Sutton Deanna A. , Thompson Elizabeth H. , Shrikanth Vandana , Verkley Gerard J. M. , Stielow J. Benjamin , Mays Rana , Oleske Deanna , Morrison L. Katie , Lapolla Whitney J. , Galfione Sarah , Tyring Stephen , Samathanam Christina A. , Fu Jianmin , Wickes Brian L. , Mulanovich Victor , Wanger Audrey , Arias Cesar A.
Published: August 2012

Paraconiothyrium cyclothyrioides
is a recently described coelomycetous fungal species. We present a case in a renal transplant patient with chronic skin lesions of the lower extremities caused by
P. cyclothyrioides
. Treatment with posaconazole led to complete resolution of the lesions.
P. cyclothyrioides
should be considered an opportunistic human pathogen in immunocompromised patients.
Evaluation of Two Hepatitis C Virus Genotyping Assays Based on the 5′ Untranslated Region (UTR): the Limitations of 5′ UTR-Based Assays and the Need for a Supplementary Sequencing-Based Approach
Authors: Hong Sung Kuk , Cho Sung Im , Ra Eun Kyung , Kim Eui-Chong , Park Joon Sung , Park Sung Sup , Seong Moon-Woo
Published: August 2012

We evaluated two genotyping methodologies that characterize the 5′ untranslated region (5′ UTR) of the hepatitis C virus (HCV) genome. The limitations of these genotype assays need to be thoroughly evaluated, and sequencing-based approaches may be needed to complement these methods in clinical settings.
Sensing and Control of Bluetongue Virus Infection in Epithelial Cells via RIG-I and MDA5 Helicases
Authors: Chauveau Emilie , Doceul Virginie , Lara Estelle , Adam Micheline , Breard Emmanuel , Sailleau Corinne , Viarouge Cyril , Desprat Alexandra , Meyer Gilles , Schwartz-Cornil Isabelle , Ruscanu Suzana , Charley Bernard , Zientara Stéphan , Vitour Damien
Published: August 2012

Bluetongue virus (BTV), an arthropod-borne member of theReoviridaefamily, is a double-stranded RNA virus that causes an economically important livestock disease that has spread across Europe in recent decades. Production of type I interferon (alpha/beta interferon [IFN-α/β]) has been reportedin vivoandin vitroupon BTV infection. However, the cellular sensors and signaling pathways involved in this process remain unknown. Here we studied the mechanisms responsible for the production of IFN-β in response to BTV serotype 8. Upon BTV infection of A549 cells, expression of IFN-β and other proinflammatory cytokines was strongly induced at both the protein and mRNA levels. This response appeared to be dependent on virus replication, since exposure to UV-inactivated virus failed to induce IFN-β. We also demonstrated that BTV infection activated the transcription factors IFN regulatory factor 3 and nuclear factor κB. We investigated the role of several pattern recognition receptors in this response and showed that expression of IFN-β was greatly reduced after small-interfering-RNA-mediated knockdown of the RNA helicase encoded by retinoic acid-inducible gene I (RIG-I) or melanoma differentiation-associated gene 5 (MDA5). In contrast, silencing of MyD88, Toll-like receptor 3, or the recently described DexD/H-box helicase DDX1 sensor had no or a weak effect on IFN-β induction, suggesting that the RIG-I-like receptor pathway is specifically engaged for BTV sensing. Moreover, we also showed that overexpression of either RIG-I or MDA5 impaired BTV expression in infected A549 cells. Overall, this indicates that RIG-I and MDA5 can both contribute to the recognition and control of BTV infection.
Cytomegalovirus CC Chemokine Promotes Immune Cell Migration
Authors: Vomaske Jennifer , Denton Michael , Kreklywich Craig , Andoh Takeshi , Osborn Jessica M. , Chen Daniel , Messaoudi Ilhem , Orloff Susan L. , Streblow Daniel N.
Published: August 2012

Cytomegaloviruses manipulate the host chemokine/receptor axis by altering cellular chemokine expression and by encoding multiple chemokines and chemokine receptors. Similar to human cytomegalovirus (HCMV), rat cytomegalovirus (RCMV) encodes multiple CC chemokine-analogous proteins, including r129 (HCMV UL128 homologue) and r131 (HCMV UL130 and MCMV m129/130 homologues). Although these proteins play a role in CMV entry, their function as chemotactic cytokines remains unknown. In the current study, we examined the role of the RCMV chemokine r129 in promoting cellular migration and in accelerating transplant vascular sclerosis (TVS) in our rat heart transplant model. We determined that r129 protein is released into culture supernatants of infected cells and is expressed with late viral gene kinetics during RCMV infection and highly expressed in heart and salivary glands during
in vivo
rat infections. Using the recombinant r129 protein, we demonstrated that r129 induces migration of lymphocytes isolated from rat peripheral blood, spleen, and bone marrow and from a rat macrophage cell line. Using antibody-mediated cell sorting of rat splenocytes, we demonstrated that r129 induces migration of naïve/central memory CD4
+
T cells. Through ligand-binding assays, we determined that r129 binds rat CC chemokine receptors CCR3, CCR4, CCR5, and CCR7. In addition, mutational analyses identified functional domains of r129 resulting in recombinant proteins that fail to induce migration (r129-ΔNT and -C31A) or alter the chemotactic ability of the chemokine (r129-F43A). Two of the mutant proteins (r129-C31A and -ΔNT) also act as dominant negatives by inhibiting migration induced by wild-type r129. Furthermore, infection of rat heart transplant recipients with RCMV containing the r129-ΔNT mutation prevented CMV-induced acceleration of TVS. Together our findings indicate that RCMV r129 is highly chemotactic, which has important implications during RCMV infection and reactivation and acceleration of TVS.
HIV-1 Replication and APOBEC3 Antiviral Activity Are Not Regulated by P Bodies
Authors: Phalora Prabhjeet K. , Sherer Nathan M. , Wolinsky Steven M. , Swanson Chad M. , Malim Michael H.
Published: August 2012

The APOBEC3 cytidine deaminases play a critical role in host-mediated defense against exogenous viruses, most notably, human immunodeficiency virus type-1 (HIV-1) and endogenous transposable elements. APOBEC3G and APOBEC3F interact with numerous proteins that regulate cellular RNA metabolism, including components of the RNA-induced silencing complex (RISC), and colocalize with a subset of these proteins to mRNA processing bodies (P bodies), which are sites of mRNA translational repression and decay. We sought to determine the role of P bodies and associated proteins in HIV-1 replication and APOBEC3 antiviral activity. While we established a positive correlation between APOBEC3 protein incorporation into virions and localization to P bodies, depletion of the P-body components DDX6 or Lsm1 did not affect HIV-1 replication, APOBEC3 packaging into virions or APOBEC3 protein mediated inhibition of HIV-1 infectivity. In addition, neither HIV-1 genomic RNA nor Gag colocalized with P-body proteins. However, simultaneous depletion of multiple Argonaute family members, the effector proteins of RISC, could modestly increase viral infectivity. Because some APOBEC3 proteins interact with several Argonaute proteins, we also tested whether they could modulate microRNA (miRNA) activity. We found no evidence for the specific regulation of miRNA function by the APOBEC3 proteins, though more general effects on transfected gene expression were observed. In sum, our results indicate that P bodies and certain associated proteins do not regulate HIV-1 replication or APOBEC3 protein antiviral activity. Localization to P bodies may therefore provide a means of sequestering APOBEC3 enzymatic activity away from cellular DNA or may be linked to as yet unidentified cellular functions.
Monitoring of S Protein Maturation in the Endoplasmic Reticulum by Calnexin Is Important for the Infectivity of Severe Acute Respiratory Syndrome Coronavirus
Authors: Fukushi Masaya , Yoshinaka Yoshiyuki , Matsuoka Yusuke , Hatakeyama Seisuke , Ishizaka Yukihito , Kirikae Teruo , Sasazuki Takehiko , Miyoshi-Akiyama Tohru
Published: August 2012

Severe acute respiratory syndrome coronavirus (SARS-CoV) is the etiological agent of SARS, a fatal pulmonary disorder with no effective treatment. We found that SARS-CoV spike glycoprotein (S protein), a key molecule for viral entry, binds to calnexin, a molecular chaperone in the endoplasmic reticulum (ER), but not to calreticulin, a homolog of calnexin. Calnexin bound to most truncated mutants of S protein, and S protein bound to all mutants of calnexin. Pseudotyped virus carrying S protein (S-pseudovirus) produced by human cells that were treated with small interfering RNA (siRNA) for calnexin expression (calnexin siRNA-treated cells) showed significantly lower infectivity than S-pseudoviruses produced by untreated and control siRNA-treated cells. S-pseudovirus produced by calnexin siRNA-treated cells contained S protein modified with N-glycan side chains differently from other two S proteins and consisted of two kinds of viral particles: those of normal density with little S protein and those of high density with abundant S protein. Treatment with peptide-N-glycosidase F (PNGase F), which removes all types of N-glycan side chains from glycoproteins, eliminated the infectivity of S-pseudovirus. S-pseudovirus and SARS-CoV produced in the presence of α-glucosidase inhibitors, which disrupt the interaction between calnexin and its substrates, showed significantly lower infectivity than each virus produced in the absence of those compounds. In S-pseudovirus, the incorporation of S protein into viral particles was obviously inhibited. In SARS-CoV, viral production was obviously inhibited. These findings demonstrated that calnexin strictly monitors the maturation of S protein by its direct binding, resulting in conferring infectivity on SARS-CoV.
N-Glycans on the Nipah Virus Attachment Glycoprotein Modulate Fusion and Viral Entry as They Protect against Antibody Neutralization
Authors: Biering Scott B. , Huang Andrew , Vu Andy T. , Robinson Lindsey R. , Bradel-Tretheway Birgit , Choi Eric , Lee Benhur , Aguilar Hector C.
Published: August 2012

Nipah virus (NiV) is the deadliest known paramyxovirus. Membrane fusion is essential for NiV entry into host cells and for the virus' pathological induction of cell-cell fusion (syncytia). The mechanism by which the attachment glycoprotein (G), upon binding to the cell receptors ephrinB2 or ephrinB3, triggers the fusion glycoprotein (F) to execute membrane fusion is largely unknown. N-glycans on paramyxovirus glycoproteins are generally required for proper protein conformational integrity, transport, and sometimes biological functions. We made conservative mutations (Asn to Gln) at the seven potential N-glycosylation sites in the NiV G ectodomain (G1 to G7) individually or in combination. Six of the seven N-glycosylation sites were found to be glycosylated. Moreover, pseudotyped virions carrying these N-glycan mutants had increased antibody neutralization sensitivities. Interestingly, our results revealed hyperfusogenic and hypofusogenic phenotypes for mutants that bound ephrinB2 at wild-type levels, and the mutant's cell-cell fusion phenotypes generally correlated to viral entry levels. In addition, when removing multiple N-glycans simultaneously, we observed synergistic or dominant-negative membrane fusion phenotypes. Interestingly, our data indicated that 4- to 6-fold increases in fusogenicity resulted from multiple mechanisms, including but not restricted to the increase of F triggering. Altogether, our results suggest that NiV-G N-glycans play a role in shielding virions against antibody neutralization, while modulating cell-cell fusion and viral entry via multiple mechanisms.
Biochemical, Conformational, and Immunogenic Analysis of Soluble Trimeric Forms of Henipavirus Fusion Glycoproteins
Authors: Chan Yee-Peng , Lu Min , Dutta Somnath , Yan Lianying , Barr Jennifer , Flora Michael , Feng Yan-Ru , Xu Kai , Nikolov Dimitar B. , Wang Lin-Fa , Skiniotis Georgios , Broder Christopher C.
Published: August 2012

The henipaviruses, Hendra virus (HeV) and Nipah virus (NiV), are paramyxoviruses discovered in the mid- to late 1990s that possess a broad host tropism and are known to cause severe and often fatal disease in both humans and animals. HeV and NiV infect cells by a pH-independent membrane fusion mechanism facilitated by their attachment (G) and fusion (F) glycoproteins. Here, several soluble forms of henipavirus F (sF) were engineered and characterized. Recombinant sF was produced by deleting the transmembrane (TM) and cytoplasmic tail (CT) domains and appending a glycosylphosphatidylinositol (GPI) anchor signal sequence followed by GPI-phospholipase D digestion, appending a trimeric coiled-coil (GCNt) domain (sF
GCNt
), or deleting the TM, CT, and fusion peptide domain. These sF glycoproteins were produced as F
0
precursors, and all were apparent stable trimers recognized by NiV-specific antisera. Surprisingly, however, only the GCNt-appended constructs (sF
GCNt
) could elicit cross-reactive henipavirus-neutralizing antibody in mice. In addition, sF
GCNt
constructs could be triggered
in vitro
by protease cleavage and heat to transition from an apparent prefusion to postfusion conformation, transitioning through an intermediate that could be captured by a peptide corresponding to the C-terminal heptad repeat domain of F. The pre- and postfusion structures of sF
GCNt
and non-GCNt-appended sF could be revealed by electron microscopy and were distinguishable by F-specific monoclonal antibodies. These data suggest that only certain sF constructs could serve as potential subunit vaccine immunogens against henipaviruses and also establish important tools for further structural, functional, and diagnostic studies on these important emerging viruses.
Broad-Spectrum Antivirals against 3C or 3C-Like Proteases of Picornaviruses, Noroviruses, and Coronaviruses
Authors: Kim Yunjeong , Lovell Scott , Tiew Kok-Chuan , Mandadapu Sivakoteswara Rao , Alliston Kevin R. , Battaile Kevin P. , Groutas William C. , Chang Kyeong-Ok
Published: August 2012

Phylogenetic analysis has demonstrated that some positive-sense RNA viruses can be classified into the picornavirus-like supercluster, which includes picornaviruses, caliciviruses, and coronaviruses. These viruses possess 3C or 3C-like proteases (3Cpro or 3CLpro, respectively), which contain a typical chymotrypsin-like fold and a catalytic triad (or dyad) with a Cys residue as a nucleophile. The conserved key sites of 3Cpro or 3CLpro may serve as attractive targets for the design of broad-spectrum antivirals for multiple viruses in the supercluster. We previously reported the structure-based design and synthesis of potent protease inhibitors of Norwalk virus (NV), a member of the
Caliciviridae
family. We report herein the broad-spectrum antiviral activities of three compounds possessing a common dipeptidyl residue with different warheads, i.e., an aldehyde (GC373), a bisulfite adduct (GC376), and an α-ketoamide (GC375), against viruses that belong to the supercluster. All compounds were highly effective against the majority of tested viruses, with half-maximal inhibitory concentrations in the high nanomolar or low micromolar range in enzyme- and/or cell-based assays and with high therapeutic indices. We also report the high-resolution X-ray cocrystal structures of NV 3CLpro-, poliovirus 3Cpro-, and transmissible gastroenteritis virus 3CLpro- GC376 inhibitor complexes, which show the compound covalently bound to a nucleophilic Cys residue in the catalytic site of the corresponding protease. We conclude that these compounds have the potential to be developed as antiviral therapeutics aimed at a single virus or multiple viruses in the picornavirus-like supercluster by targeting 3Cpro or 3CLpro.
Internal Catalase Protects Herpes Simplex Virus from Inactivation by Hydrogen Peroxide
Authors: Newcomb William W. , Brown Jay C.
Published: August 2012

Herpes simplex virus 1 (HSV-1) was shown to contain catalase, an enzyme able to detoxify hydrogen peroxide by converting it to water and oxygen. Studies with a catalase inhibitor indicated that virus-associated catalase can have a role in protecting the virus from oxidative inactivation. HSV-1 was found to be more sensitive to killing by hydrogen peroxide in the presence of a catalase inhibitor than in its absence. The results suggest a protective role for catalase during the time HSV-1 spends in the oxidizing environment outside a host cell.
Isolation of Novel Synthetic Prion Strains by Amplification in Transgenic Mice Coexpressing Wild-Type and Anchorless Prion Proteins
Authors: Raymond Gregory J. , Race Brent , Hollister Jason R. , Offerdahl Danielle K. , Moore Roger A. , Kodali Ravindra , Raymond Lynne D. , Hughson Andrew G. , Rosenke Rebecca , Long Dan , Dorward David W. , Baron Gerald S.
Published: August 2012

Mammalian prions are thought to consist of misfolded aggregates (protease-resistant isoform of the prion protein [PrP
res
]) of the cellular prion protein (PrP
C
). Transmissible spongiform encephalopathy (TSE) can be induced in animals inoculated with recombinant PrP (rPrP) amyloid fibrils lacking mammalian posttranslational modifications, but this induction is inefficient in hamsters or transgenic mice overexpressing glycosylphosphatidylinositol (GPI)-anchored PrP
C
. Here we show that TSE can be initiated by inoculation of misfolded rPrP into mice that express wild-type (wt) levels of PrP
C
and that synthetic prion strain propagation and selection can be affected by GPI anchoring of the host's PrP
C
. To create prions
de novo
, we fibrillized mouse rPrP in the absence of molecular cofactors, generating fibrils with a PrP
res
-like protease-resistant banding profile. These fibrils induced the formation of PrP
res
deposits in transgenic mice coexpressing wt and GPI-anchorless PrP
C
(wt/GPI
−
) at a combined level comparable to that of PrP
C
expression in wt mice. Secondary passage into mice expressing wt, GPI
−
, or wt plus GPI
−
PrP
C
induced TSE disease with novel clinical, histopathological, and biochemical phenotypes. Contrary to laboratory-adapted mouse scrapie strains, the synthetic prion agents exhibited a preference for conversion of GPI
−
PrP
C
and, in one case, caused disease only in GPI
−
mice. Our data show that novel TSE agents can be generated
de novo
solely from purified mouse rPrP after amplification in mice coexpressing normal levels of wt and anchorless PrP
C
. These observations provide insight into the minimal elements required to create prions
in vitro
and suggest that the PrP
C
GPI anchor can modulate the propagation of synthetic TSE strains.
Porcine Circovirus Type 2 Induces Autophagy via the AMPK/ERK/TSC2/mTOR Signaling Pathway in PK-15 Cells
Authors: Zhu Binglin , Zhou Yingshan , Xu Fei , Shuai Jiangbing , Li Xiaoliang , Fang Weihuan
Published: August 2012

Porcine circovirus type 2 (PCV2) uses autophagy machinery to enhance its replication in PK-15 cells. However, the underlying mechanisms are unknown. By the use of specific inhibitors, RNA interference, and coimmunoprecipitation, we show that PCV2 induces autophagy in PK-15 cells through a pathway involving the kinases AMP-activated protein kinase (AMPK) and extracellular signal-regulated kinase 1/2 (ERK1/2), the tumor suppressor protein TSC2, and the mammalian target of rapamycin (mTOR). AMPK and ERK1/2 positively regulate autophagy through negative control of the mTOR pathway by phosphorylating TSC2 in PCV2-infected PK-15 cells. Thus, PCV2 might induce autophagy via the AMPK/ERK/TSC2/mTOR signaling pathway in the host cells, representing a pivotal mechanism for PCV2 pathogenesis.
Comparative Study of Influenza Virus Replication in MDCK Cells and in Primary Cells Derived from Adenoids and Airway Epithelium
Authors: Ilyushina Natalia A. , Ikizler Mine R. , Kawaoka Yoshihiro , Rudenko Larisa G. , Treanor John J. , Subbarao Kanta , Wright Peter F.
Published: August 2012

Although clinical trials with human subjects are essential for determination of safety, infectivity, and immunogenicity, it is desirable to know in advance the infectiousness of potential candidate live attenuated influenza vaccine strains for human use. We compared the replication kinetics of wild-type and live attenuated influenza viruses, including H1N1, H3N2, H9N2, and B strains, in Madin-Darby canine kidney (MDCK) cells, primary epithelial cells derived from human adenoids, and human bronchial epithelium (NHBE cells). Our data showed that despite the fact that all tissue culture models lack a functional adaptive immune system, differentiated cultures of human epithelium exhibited the greatest restriction for all H1N1, H3N2, and B vaccine viruses studied among three cell types tested and the best correlation with their levels of attenuation seen in clinical trials with humans. In contrast, the data obtained with MDCK cells were the least predictive of restricted viral replication of live attenuated vaccine viruses in humans. We were able to detect a statistically significant difference between the replication abilities of the U.S. (A/Ann Arbor/6/60) and Russian (A/Leningrad/134/17/57) cold-adapted vaccine donor strains in NHBE cultures. Since live attenuated pandemic influenza vaccines may potentially express a hemagglutinin and neuraminidase from a non-human influenza virus, we assessed which of the three cell cultures could be used to optimally evaluate the infectivity and cellular tropism of viruses derived from different hosts. Among the three cell types tested, NHBE cultures most adequately reflected the infectivity and cellular tropism of influenza virus strains with different receptor specificities. NHBE cultures could be considered for use as a screening step for evaluating the restricted replication of influenza vaccine candidates.
A Genetic Polymorphism of
FREM1
Is Associated with Resistance against HIV Infection in the Pumwani Sex Worker Cohort
Authors: Luo Ma , Sainsbury James , Tuff Jeffrey , Lacap Philip A. , Yuan Xin-Yong , Hirbod Taha , Kimani Joshua , Wachihi Charles , Ramdahin Sue , Bielawny Thomas , Embree Joanne , Broliden Kristina , Ball T. Blake , Plummer Francis A.
Published: August 2012

A subgroup of women enrolled in the Pumwani sex worker cohort remain seronegative and PCR negative for human immunodeficiency virus type 1 despite repeated exposure through high-risk sex work. Studies have shown that polymorphisms of genes involved in antigen presentation and viral restriction factors are associated with resistance to HIV infection. To discover other possible genetic factors underlying this HIV-resistant phenotype, we conducted an exploratory nonbiased, low-resolution, genome-wide single-nucleotide polymorphism (SNP) analysis comparing 60 HIV-resistant women to 48 HIV-infected controls. The SNP minor allele rs1552896, in an intron of
FREM1
, was significantly associated with the resistant phenotype (
P
= 1.68 × 10
−5
; adjusted
P
= 2.37 × 10
−4
; odds ratio [OR], 9.51; 95% confidence interval [CI], 2.82 to 32.05). We expanded the sample size by genotyping rs1552896 in the Pumwani cohort and comparing 114 HIV-resistant women to 609 HIV-infected controls and confirmed the association (
P
= 1.7 × 10
−4
; OR, 2.67; 95% CI, 1.47 to 4.84). To validate the association in a second cohort, we genotyped 783 women enrolled in a mother-child health study and observed the minor allele of rs1552896 enriched in HIV-uninfected women (
n
= 488) compared to HIV-infected enrollees (
n
= 295) (
P
= 0.036; OR, 1.69; 95% CI, 0.98 to 2.93). Quantitative reverse transcription-PCR showed that
FREM1
mRNA was highly expressed in tissues relevant for HIV-1 infection, and immunohistochemical analysis revealed that FREM1 protein is expressed in the ectocervical mucosa of HIV-resistant women. The significant association of rs1552896 with an HIV-resistant phenotype, together with the expression profile of FREM1 in tissues relevant to HIV infection, suggests that
FREM1
is a potentially novel candidate gene for resistance to HIV infection.
Inclusion Bodies Are a Site of Ebolavirus Replication
Authors: Hoenen Thomas , Shabman Reed S. , Groseth Allison , Herwig Astrid , Weber Michaela , Schudt Gordian , Dolnik Olga , Basler Christopher F. , Becker Stephan , Feldmann Heinz
Published: August 2012

Inclusion bodies are a characteristic feature of ebolavirus infections in cells. They contain large numbers of preformed nucleocapsids, but their biological significance has been debated, and they have been suggested to be aggregates of viral proteins without any further biological function. However, recent data for other viruses that produce similar structures have suggested that inclusion bodies might be involved in genome replication and transcription. In order to study filovirus inclusion bodies, we fused mCherry to the ebolavirus polymerase L, which is found in inclusion bodies. The resulting L-mCherry fusion protein was functional in minigenome assays and incorporated into virus-like particles. Importantly, L-mCherry fluorescence in transfected cells was readily detectable and distributed in a punctate pattern characteristic for inclusion bodies. A recombinant ebolavirus encoding L-mCherry instead of L was rescued and showed virtually identical growth kinetics and endpoint titers to those for wild-type virus. Using this virus, we showed that the onset of inclusion body formation corresponds to the onset of viral genome replication, but that viral transcription occurs prior to inclusion body formation. Live-cell imaging further showed that inclusion bodies are highly dynamic structures and that they can undergo dramatic reorganization during cell division. Finally, by labeling nascent RNAs using click technology we showed that inclusion bodies are indeed the site of viral RNA synthesis. Based on these data we conclude that, rather than being inert aggregates of nucleocapsids, ebolavirus inclusion bodies are in fact complex and dynamic structures and an important site at which viral RNA replication takes place.
Complete Genome Sequence of Salmonella Bacteriophage SS3e
Authors: Kim Sung-Hun , Park Jeong-Hyun , Lee Bok-Kwon , Kwon Hyuk-Joon , Shin Ji-Hyun , Kim Jungmin , Kim Shukho
Published: August 2012

A
Salmonella
lytic bacteriophage, SS3e, was isolated, and its genome was sequenced completely. This phage is able to lyse not only various
Salmonella
serovars but also
Escherichia coli
,
Shigella sonnei
,
Enterobacter cloacae
, and
Serratia marcescens
, indicating a broad host specificity. Genomic sequence analysis of SS3e revealed a linear double-stranded DNA sequence of 40,793 bp harboring 58 open reading frames, which is highly similar to
Salmonella
phages SETP13 and MB78.
Complete Genome Sequence of Two Coxsackievirus A1 Strains That Were Cytotoxic to Human Rhabdomyosarcoma Cells
Authors: Sun Qiang , Zhang Yong , Zhu Shuangli , Cui Hui , Tian Huifang , Yan Dongmei , Huang Guohong , Zhu Zhen , Wang Dongyan , Li Xiaolei , Jiang Huafang , An Hongqiu , Xu Wenbo
Published: August 2012

Coxsackievirus A1 (CVA1) belongs to human enterovirus species C within the family
Picornaviridae
, order
Picornavirales
. Two Chinese CVA1 isolates, HT-THLH02F/XJ/CHN/2011 and KS-ZPH01F/XJ/CHN/2011, were isolated from stool specimens of two healthy children in the Xinjiang Uygur autonomous region of China. They were found to elicit cytopathic effects in a human rhabdomyosarcoma cell line, and complete genome sequences of these two CVA1 isolates revealed that natural intertypic recombination events occurred between CVA1 and CVA22.
Biology of a Novel Mycobacteriophage, SWU1, Isolated from Chinese Soil as Revealed by Genomic Characteristics
Authors: Fan Xiangyu , Teng Tieshan , Wang Honghai , Xie Jianping
Published: August 2012

Mycobacteriophage SWU1 is a newly isolated phage from a soil sample collected at Gongping village, Pingchang County, Sichuan Province, China, using
Mycobacterium smegmatis
mc
2
155 as a host. Plaques of SWU1 appear as a unique bull's-eye on an
M. smegmatis
lawn. In this paper, we report the complete genome sequence of SWU1 and some major findings from the analysis result.
Complete Genome of Staphylococcus aureus Phage SA11
Authors: Kim Min Soo , Myung Heejoon
Published: August 2012

A novel lytic bacteriophage, SA11, infecting
Staphylococcus aureus
was isolated, and the whole genome was sequenced. It belongs to the siphoviridae based on electron microscopic observation. It has a linear double-stranded DNA genome of 136,326 bp. Genomic analysis showed that it is distantly related to
Staphylococcus
phages A5W, K, ISP, Sb-1, and G1.
Complete Genome Sequence of Caulobacter crescentus Bacteriophage φCbK
Authors: Panis Gaël , Lambert Christophe , Viollier Patrick H.
Published: August 2012

φCbK is a B3 morphotype bacteriophage of the
Siphoviridae
family that infects
Caulobacter crescentus
, the preeminent model system for bacterial cell cycle studies. The last 4 decades of research with φCbK as a genetic and cytological tool to study the biology of the host warrant an investigation of the phage genome composition. Herein, we report the complete genome sequence of φCbK and highlight unusual features that emerged from its annotation. The complete genome analysis of the φCbK phage provides new insight into its characteristics and potential interactions with its
Caulobacter crescentus
host, setting the stage for future functional studies with φCbK.
Complete Genome Sequence of a Bovine Viral Diarrhea Virus 2 from Commercial Fetal Bovine Serum
Authors: Liu Hua , Li Yan , Gao Mingchun , Wen Kai , Jia Ying , Liu Xiaomei , Zhang Wenlong , Ma Bo , Wang Junwei
Published: August 2012

We isolated a bovine viral diarrhea virus (BVDV) from commercial fetal bovine serum and designated it HLJ-10. The complete genome is 12,284 nucleotides (nt); the open reading frame is 11,694 nt, coding 3,898 amino acids. Phylogenetic analysis indicated that this strain belongs to BVDV group 2.
Complete Genome Sequence of a Novel Porcine Epidemic Diarrhea Virus in South China
Authors: Fan Huiying , Zhang Jie , Ye Yu , Tong Tiezhu , Xie Kangshang , Liao Ming
Published: August 2012

Since early 2010, outbreaks of porcine epidemic diarrhea (PED) have been observed frequently in immunized swine herds in southern China. The suckling piglets are particularly susceptible to porcine epidemic diarrhea virus (PEDV), with a high mortality rate (90%). Recently, a virulent PEDV strain, GD-A, was isolated from an immunized-swine breeding farm in Guangdong, China. This report describes the complete genome sequence of GD-A, and the data will provide important insights into the variation of PEDV field isolates in southern China.
Complete Genome Sequence of an Avian-Origin H3N2 Canine Influenza A Virus Isolated in Farmed Dogs in Southern China
Authors: Su Shuo , Cao Nan , Chen Jidang , Zhao Furong , Li Huatao , Zhao Mingxi , Wang Yanjing , Huang Zhen , Yuan Liguo , Wang Heng , Zhang Guihong , Li Shoujun
Published: August 2012

We report here the complete genomic sequence of an avian-origin H3N2 canine influenza A virus containing multiple mutations in farmed dogs in southern China. Phylogenetic analyses of the sequences of all eight viral RNA segments demonstrated that these are wholly avian influenza viruses of the Asia lineage. To our knowledge, this is the first report of interspecies transmission of an avian H3N2 influenza virus to domestic farm dogs under natural conditions in Southern China. The amino acid information provided herein suggests that continued study is required to determine if this virus could be established in the farm dog population and pose potential threats to public health.
Complete Genome Sequence of Ikoma Lyssavirus
Authors: Marston Denise A. , Ellis Richard J. , Horton Daniel L. , Kuzmin Ivan V. , Wise Emma L. , McElhinney Lorraine M. , Banyard Ashley C. , Ngeleja Chanasa , Keyyu Julius , Cleaveland Sarah , Lembo Tiziana , Rupprecht Charles E. , Fooks Anthony R.
Published: August 2012

Lyssaviruses (family
Rhabdoviridae
) constitute one of the most important groups of viral zoonoses globally. All lyssaviruses cause the disease rabies, an acute progressive encephalitis for which, once symptoms occur, there is no effective cure. Currently available vaccines are highly protective against the predominantly circulating lyssavirus species. Using next-generation sequencing technologies, we have obtained the whole-genome sequence for a novel lyssavirus, Ikoma lyssavirus (IKOV), isolated from an African civet in Tanzania displaying clinical signs of rabies. Genetically, this virus is the most divergent within the genus
Lyssavirus
. Characterization of the genome will help to improve our understanding of lyssavirus diversity and enable investigation into vaccine-induced immunity and protection.
Genome Sequence of a Bombyx mori Nucleopolyhedrovirus Strain with Cubic Occlusion Bodies
Authors: Cheng Ruo-Lin , Xu Yi-Peng , Zhang Chuan-Xi
Published: August 2012

Bombyx mori
nucleopolyhedrovirus (BmNPV) is a typical species of
Baculoviridae
. The complete genome sequence of a BmNPV strain with cubic occlusion bodies is reported here. The genome of this strain consists of 127,465 nucleotides with a G+C content of 40.36% and is 97.3% and 97.5% identical to those of BmNPV strain T3 and
Bombyx mandarina
NPV S1, respectively. Despite the abnormal polyhedra it forms, the
polyhedrin
gene of the BmNPV cubic strain is 100% identical to those of the other two strains. Baculovirus repeated ORFs and homologous repeat regions cause the major differences in genome size of these BmNPV isolates.
Complete Genome Sequence of a Novel Duck Hepatitis A Virus Discovered in Southern China
Authors: Wei Chun-ya , Su Shuo , Huang Zhen , Zhu Wan-jun , Chen Ji-dang , Zhao Fu-rong , Wang Yan-jing , Xie Jie-xiong , Wang Heng , Zhang Guihong
Published: August 2012

We report here the complete genomic sequence of a novel duck hepatitis A virus (DHAV) isolated from mixed infections with DHAV type 1 (DHAV-1) and DHAV-3 in ducklings in Southern China. The whole nucleotide sequence had the highest homology with the sequence of DHAV-3 (GenBank accession number
DQ812093
) (96.2%). To our knowledge, this is the first report of gene rearrangement between DHAV-1 and DHAV-3, and it will help to understand the epidemiology and molecular characteristics of duck hepatitis A virus in Southern China.
Murine Gammaherpesvirus 68 LANA Acts on Terminal Repeat DNA To Mediate Episome Persistence
Authors: Habison Aline C. , Beauchemin Chantal , Simas J. Pedro , Usherwood Edward J. , Kaye Kenneth M.
Published: August 2012

Murine gammaherpesvirus 68 (MHV68) ORF73 (mLANA) has sequence homology to Kaposi's sarcoma-associated herpesvirus (KSHV) latency-associated nuclear antigen (LANA). LANA acts on the KSHV terminal repeat (TR) elements to mediate KSHV episome maintenance. Disruption of mLANA expression severely reduces the ability of MHV68 to establish latent infection in mice, consistent with the possibility that mLANA mediates episome persistence. Here we assess the roles of mLANA and MHV68 TR (mTR) elements in episome persistence. mTR-associated DNA persisted as an episome in latently MHV68-infected tumor cells, demonstrating that the mTR elements can serve as acis-acting element for MHV68 episome maintenance. In some cases, both control vector and mTR-associated DNAs integrated into MHV68 episomal genomes. Therefore, we also assessed the roles of mTRs as well as mLANA in the absence of infection. DNA containing both mLANA and mTRs incispersisted as an episome in murine A20 or MEF cells. In contrast, mTR DNA never persisted as an episome in the absence of mLANA. mLANA levels were increased when mLANA was expressed from its native promoters, and episome maintenance was more efficient with higher mLANA levels. Increased numbers of mTRs conferred more efficient episome maintenance, since DNA containing mLANA and eight mTR elements persisted more efficiently in A20 cells than did DNA with mLANA and two or four mTRs. Similar to KSHV LANA, mLANA broadly associated with mitotic chromosomes but relocalized to concentrated dots in the presence of episomes. Therefore, mLANA acts on mTR elements to mediate MHV68 episome persistence.
Evidence for pH-Dependent Protease Activity in the Adeno-Associated Virus Capsid
Authors: Salganik Maxim , Venkatakrishnan Balasubramanian , Bennett Antonette , Lins Bridget , Yarbrough Joseph , Muzyczka Nicholas , Agbandje-McKenna Mavis , McKenna Robert
Published: August 2012

Incubation of highly purified adeno-associated virus (AAV) capsids
in vitro
at pH 5.5 induced significant autocleavage of capsid proteins at several amino acid positions. No autocleavage was seen at pH 7.5. Examination of other AAV serotypes showed at least two different pH-induced cleavage patterns, suggesting that different serotypes have evolved alternative protease cleavage sites. In contrast, incubation of AAV serotypes with an external protease substrate showed that purified AAV capsid preparations have robust protease activity at neutral pH but not at pH 5.5, opposite to what is seen with capsid protein autocleavage. Several lines of evidence suggested that protease activity is inherent in AAV capsids and is not due to contaminating proteins. Control virus preparations showed no protease activity on external substrates, and filtrates of AAV virus preparations also showed no protease activity contaminating the capsids. Further, N-terminal Edman sequencing identified unique autocleavage sites in AAV1 and AAV9, and mutagenesis of amino acids adjacent to these sites eliminated cleavage. Finally, mutation of an amino acid in AAV2 (E563A) that is in a conserved pH-sensitive structural region eliminated protease activity on an external substrate but did not seem to affect autocleavage. Taken together, our data suggested that AAV capsids have one or more protease active sites that are sensitive to pH induction. Further, it appears that acidic pHs comparable to those seen in late endosomes induce a structural change in the capsid that induces autolytic protease activity. The pH-dependent protease activity may have a role in viral infection.