bluuebunny/crossref_metadata_2025 huggingface dataset:
Diagnostic Usefulness of Ribosomal Protein L7/L12 for Pneumococcal Pneumonia in a Mouse Model
Authors: Sawa Tomoka , Kimura Soichiro , Honda Natsue Hosono , Fujita Kazue , Yoshizawa Sadako , Harada Yoshihiro , Sugiyama Yuya , Matsuyama Kenji , Sohka Takayuki , Saji Tsutomu , Yamaguchi Keizo , Tateda Kazuhiro
Published: October 2012

The capsular antigen detection (CAD) kit is widely used in clinics to detect
Streptococcus pneumoniae
infection from urine, because it is rapid, convenient, and effective. However, there are several disadvantages, including false-positive results in children colonized with
S. pneumoniae
and prolonged positive readings even after the bacteria have been cleared. RP-L7/L12 is a component of the 50S ribosome that is abundant in all bacteria and is specific for each bacterial species. We investigated whether RP-L7/L12 could be used to accurately diagnose pneumococcal pneumonia infection in mouse models of pneumonia and colonization generated by infecting CBA/JN or CBA/N mice, respectively, with
S. pneumoniae
strain 741. RP-L7/L12 detection by enzyme-linked immunosorbent assay accurately assessed active lung infection, as RP-L7/L12 levels decreased simultaneously with the bacterial lung burden after imipenem administration in the pneumonia mouse model. Based on the data, antibodies detecting RP-L7/L12 were applied to rapid immunochromatographic strips (ICS) for urine sample testing. When we compared the ICS test with the CAD kit in the pneumonia model, the results correlated well. Interestingly, however, when the lung bacterial burden became undetectable after antibiotic treatment, the ICS test was correspondingly negative, even though the same samples tested by the CAD kit remained positive. Similarly, while the ICS test exhibited negative results in the nasal colonization model, the CAD kit demonstrated positive results. Bacterial RP-L7/L12 may be a promising target for the development of new methods to diagnose infectious disease. Further studies are warranted to determine whether such a test could be useful in children.
Detection, Identification, and Distribution of Fungi in Bronchoalveolar Lavage Specimens by Use of Multilocus PCR Coupled with Electrospray Ionization/Mass Spectrometry
Authors: Shin Jeong Hwan , Ranken Raymond , Sefers Susan E. , Lovari Robert , Quinn Criziel D. , Meng Shufang , Carolan Heather E. , Toleno Donna , Li Haijing , Lee Jeong Nyeo , Stratton Charles W. , Massire Christian , Tang Yi-Wei
Published: October 2012

As pulmonary fungal infections continue to increase due to an increasing number of immunocompromised patients, rapid detection and accurate identification of these fungal pathogens are critical. A broad fungal assay was developed by incorporating broad-range multilocus PCR amplification and electrospray ionization/mass spectrometry (PCR/ESI-MS) to detect and identify fungal organisms directly from clinical specimens. The aims of this study were to evaluate the performance of PCR/ESI-MS for detection, identification, and determination of the distribution of fungal organisms in bronchoalveolar lavage (BAL) fluid specimens. The BAL fluid specimens submitted for fungal culture at Vanderbilt University Medical Center between May 2005 and October 2011 were included. Cultures and identification were done using standard procedures. In addition, DNA was extracted from BAL fluid specimens, and fungal DNA amplification/identification were performed by PCR/ESI-MS. The results were compared with those of the standard cultures. A total of 691 nonduplicated BAL fluid specimens with sufficient leftover volume for molecular testing were evaluated using PCR/ESI-MS. Among them, 134 specimens (19.4%) were positive for fungi by both culture and PCR/ESI-MS testing. Of the dual-positive specimens, 125 (93.3%) were positive for
Candida
and
Aspergillus
species, with concordances between culture and PCR/ESI-MS results being 84 (67.2%) at the species level and 109 (87.2%) at the genus level. In addition, 243 (35.2%) and 30 (4.3%) specimens were positive only by PCR/ESI-MS or by culture, respectively (odds ratio [OR] = 11.95, 95% confidence interval [CI] = 7.90 to 18.17,
P
= 0.0000). Codetection of fungal organisms was noted in 23 (3.3%) specimens by PCR/ESI-MS, which was significantly higher than the 4 (0.6%) in which they were noted by culture (OR = 5.91, 95% CI = 1.93 to 20.27,
P
= 0.0002). Among 53 specimens in which cultures failed because of bacterial overgrowth, at least one fungus was identified in 26 specimens (47.3%) by PCR/ESI-MS. PCR/ESI-MS provides an advanced tool for rapid and sensitive detection, identification, and determination of the distribution of fungal organisms directly from BAL fluid specimens. Moreover, it detected fungal organisms in specimens in which cultures failed because of bacterial overgrowth. The clinical relevance of the significantly higher detection rate of fungal organisms by PCR/ESI-MS merits further investigation.
A Novel, Sensitive Assay for High-Throughput Molecular Detection of Plasmodia for Active Screening of Malaria for Elimination
Authors: Cheng Zhibin , Sun Xiaodong , Yang Ye , Wang Heng , Zheng Zhi
Published: October 2012

Although malaria remains one of the leading infectious diseases in the world, the decline in malaria transmission in some area makes it possible to consider elimination of the disease. As countries approach elimination, malaria diagnosis needs to change from diagnosing ill patients to actively detecting infections in all carriers, including asymptomatic and low-parasite-load patients. However, few of the current diagnostic methods have both the throughput and the sensitivity required. We adopted a sandwich RNA hybridization assay to detect genus
Plasmodium
18S rRNA directly from whole-blood samples from
Plasmodium falciparum
and
Plasmodium vivax
patients without RNA isolation. We tested the assay with 202 febrile patients from areas where malaria is endemic, using 20 μl of each blood sample in a 96-well plate format with a 2-day enzyme-linked immunosorbent assay (ELISA)-like work flow. The results were compared with diagnoses obtained using microscopy, a rapid diagnostic test (RDT), and genus-specific real-time PCR. Our assay identified all 66 positive samples diagnosed by microscopy, including 49 poorly stored samples that underwent multiple freeze-thaw cycles due to resource limitation. The assay uncovered three false-negative samples by microscopy and four false-negative samples by RDT and agreed completely with real-time PCR diagnosis. There was no negative sample by our assay that would show a positive result when tested with other methods. The detection limit of our assay for
P. falciparum
was 0.04 parasite/μl. The assay's simple work flow, high throughput, and sensitivity make it suitable for active malaria screening.
Development and Application of Real-Time PCR for Detection of Subgroup J Avian Leukosis Virus
Authors: Qin Liting , Gao Yulong , Ni Wei , Sun Meiyu , Wang Yongqiang , Yin Chunhong , Qi Xiaole , Gao Honglei , Wang Xiaomei
Published: October 2012

Subgroup J avian leukosis virus (ALV-J) is an avian retrovirus that causes severe economic losses in the poultry industry. The early identification and removal of virus-shedding birds are important to reduce the spread of congenital and contact infections. In this study, a TaqMan-based real-time PCR method for the rapid detection and quantification of ALV-J with proviral DNA was developed. This method exhibited a high specificity for ALV-J. Moreover, the detection limit was as low as 10 viral DNA copies. The coefficients of variation (CVs) of both interassay and intra-assay reproducibility were less than 1%. The growth curves of ALV-J in DF-1 cells were measured by real-time PCR, yielding a trend line similar to those determined by 50% tissue culture infective dose (TCID
50
) and p27 antigen detection. Tissue samples suspected of ALV infection were evaluated using real-time PCR, virus isolation, and routine PCR, and the positivity rates were 60.1%, 41.6% and 44.5%, respectively. Our data indicated that the real-time PCR method provides a sensitive, specific, and reproducible diagnostic tool for the identification and quantification of ALV-J for clinical diagnosis and in laboratory research.
Comparison of the Staphylococcus QuickFISH BC Test with the Tube Coagulase Test Performed on Positive Blood Cultures for Evaluation and Application in a Clinical Routine Setting
Authors: Carretto E. , Bardaro M. , Russello G. , Mirra M. , Zuelli C. , Barbarini D.
Published: October 2012

Many studies demonstrate that delayed proper therapy in bloodstream infections caused by
Staphylococcus aureus
increases the mortality rate, emphasizing the need to shorten the turnaround time for positive blood cultures. Different techniques are currently available, from phenotypic methods to more complex tests such as matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF), real-time PCR (RT-PCR), and fluorescence in situ hybridization using peptide nucleic acid probes (PNA FISH). This study evaluated the performance of the
Staphylococcus
QuickFISH BC test (QFT), a novel FISH methodology, compared with the direct tube coagulase test (DTCT) on blood cultures exhibiting Gram-positive cocci in clusters. A total of 173 blood cultures collected from 128 different patients were analyzed using the DTCT, evaluated after both 4 and 24 h, and the QFT. A total of 179 isolates were identified using the Vitek2 system. Thirty-five out of 35
Staphylococcus aureus
were correctly identified by the QFT (sensitivity = 100%), with a specificity of 100% (no green fluorescence was detected for strains different from
S. aureus
). The DTCT was positive after 4 h for 28 out of the 35 samples (sensitivity = 80%) and after 24 h for 31 out of the 35 samples (sensitivity = 88.57%). Among the remaining 144 isolates, one was then identified as
Corynebacterium striatum
and two as
Micrococcus luteus
. QFT identified 139 out of the 141 coagulase-negative staphylococci (CoNS) (sensitivity = 98.58%), showing again a specificity of 100% (no fluorescent red signals were detected for strains different from CoNS). We also discuss also the implementation process of this methodology in our setting, with particular emphasis on the workflow and the cost-effectiveness.
Evaluation of the Abbott HBV RUO Sequencing Assay Combined with Laboratory-Modified Interpretive Software
Authors: Germer Jeffrey J. , Abraham Priya , Mandrekar Jayawant N. , Yao Joseph D. C.
Published: October 2012

The Abbott HBV RUO Sequencing assay (Abbott Molecular Inc., Des Plaines, IL), which combines automated sample processing, real-time PCR, and bidirectional DNA sequencing, was evaluated for detection of nucleos(t)ide analogue (NA) resistance-associated mutations located in the hepatitis B virus (HBV) polymerase (Pol) gene. Interpretive software from the assay manufacturer was modified to allow interrogation of the overlapping HBV surface (S) gene sequence for HBV genotype determination and detection of immune escape mutations. Analytical sensitivity (detection and sequencing) of the assay was determined to be 103.9 IU/ml (95% confidence interval [CI], 80.0 to 173.3) for HBV genotype A. Testing of commercially available HBV genotype panels consisting of 23 individual members yielded complete agreement between expected results and results obtained from the laboratory-developed HBV genotype library. Excellent specificity was observed among clinical specimens with serologic or molecular markers for various unrelated blood-borne viruses (n= 6) and sera obtained from healthy, HBV-negative blood donors (n= 20). Retrospectively selected clinical specimens tested by a commercial reference laboratory HBV sequencing assay (n= 54) or the TrugeneHBVGenotyping kit (n= 7) and the Abbott HBV RUO Sequencing assay showed minor differences in detection and reporting of NA resistance-associated mutations in 7 of 61 (11.5%) specimens but complete agreement of genotype results. The Abbott HBV RUO Sequencing assay provided a convenient and efficient assay workflow suitable for routine clinical laboratory use, with the flexibility to be modified for customized detection of NA resistance-associated mutations, HBV genotype determination, and detection of immune escape mutations from a single contiguous HBV sequence.
Ring Infiltrate in Staphylococcal Keratitis
Authors: Wallang Batriti S. , Das Sujata , Sharma Savitri , Sahu Srikant K. , Mittal Ruchi
Published: October 2012

Smear and culture tests of corneal scrapings from a patient with a ring infiltrate confirmed significant growth of a
Staphylococcus
species resistant to fluoroquinolones. Because of nonresponse to medical management, the patient underwent therapeutic penetrating keratoplasty. Staphylococcal infection of the cornea may appear as a ring-like infiltrate that is recalcitrant to medical management.
Human Cryptosporidiosis Caused by
Cryptosporidium tyzzeri
and
C. parvum
Isolates Presumably Transmitted from Wild Mice
Authors: Rašková Veronika , Květoňová Dana , Sak Bohumil , McEvoy John , Edwinson Adam , Stenger Brianna , Kváč Martin
Published: October 2012

We report a case of severe human cryptosporidiosis caused by
Cryptosporidium tyzzeri
and
C. parvum
with an unusually high frequency of liquid stools. Wild mice were the most likely source of infection, demonstrating the potential for wild-mouse-borne
Cryptosporidium
to infect humans and highlighting the health risks associated with synantropic rodents.
APOBEC3G Restricts HIV-1 to a Greater Extent than APOBEC3F and APOBEC3DE in Human Primary CD4
+
T Cells and Macrophages
Authors: Chaipan Chawaree , Smith Jessica L. , Hu Wei-Shau , Pathak Vinay K.
Published: October 2012

APOBEC3 proteins inhibit HIV-1 replication in experimental systems and induce hypermutation in infected patients; however, the relative contributions of several APOBEC3 proteins to restriction of HIV-1 replication in the absence of the viral Vif protein in human primary CD4
+
T cells and macrophages are unknown. We observed significant inhibition of HIV-1Δ
vif
produced in 293T cells in the presence of APOBEC3DE (A3DE), APOBEC3F (A3F), APOBEC3G (A3G), and APOBEC3H haplotype II (A3H HapII) but not APOBEC3B (A3B), APOBEC3C (A3C), or APOBEC3H haplotype I (A3H HapI). Our previous studies showed that Vif amino acids Y
40
RHHY
44
are important for inducing proteasomal degradation of A3G, whereas amino acids
14
DRMR
17
are important for degradation of A3F and A3DE. Here, we introduced substitution mutations of
40
YRHHY
44
and
14
DRMR
17
in replication-competent HIV-1 to generate
vif
mutants NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 to compare the antiviral activity of A3G to the combined antiviral activity of A3F and A3DE in activated CD4
+
T cells and macrophages. During the first 15 days (round 1), in which multiple cycles of viral replication occurred, both the NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 mutants replicated in activated CD4
+
T cells and macrophages, and only the NL4-3 YRHHY>A5 mutant showed a 2- to 4-day delay in replication compared to the wild type. During the subsequent 27 days (round 2) of cultures initiated with peak virus obtained from round 1, the NL4-3 YRHHY>A5 mutant exhibited a longer, 8- to 10-day delay and the NL4-3 DRMR>A4 mutant exhibited a 2- to 6-day delay in replication compared to the wild type. The NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 mutant proviruses displayed G-to-A hypermutations primarily in GG and GA dinucleotides as expected of A3G- and A3F- or A3DE-mediated deamination, respectively. We conclude that A3G exerts a greater restriction effect on HIV-1 than A3F and A3DE.
Influenza A Viruses Grow in Human Pancreatic Cells and Cause Pancreatitis and Diabetes in an Animal Model
Authors: Capua Ilaria , Mercalli Alessia , Pizzuto Matteo S. , Romero-Tejeda Aurora , Kasloff Samantha , De Battisti Cristian , Bonfante Francesco , Patrono Livia V. , Vicenzi Elisa , Zappulli Valentina , Lampasona Vito , Stefani Annalisa , Doglioni Claudio , Terregino Calogero , Cattoli Giovanni , Piemonti Lorenzo
Published: October 2012

Influenza A viruses commonly cause pancreatitis in naturally and experimentally infected animals. In this study, we report the results of
in vivo
investigations carried out to establish whether influenza virus infection could cause metabolic disorders linked to pancreatic infection. In addition,
in vitro
tests in human pancreatic islets and in human pancreatic cell lines were performed to evaluate viral growth and cell damage. Infection of an avian model with two low-pathogenicity avian influenza isolates caused pancreatic damage resulting in hyperlipasemia in over 50% of subjects, which evolved into hyperglycemia and subsequently diabetes. Histopathology of the pancreas showed signs of an acute infection resulting in severe fibrosis and disruption of the structure of the organ. Influenza virus nucleoprotein was detected by immunohistochemistry (IHC) in the acinar tissue. Human seasonal H1N1 and H3N2 viruses and avian H7N1 and H7N3 influenza virus isolates were able to infect a selection of human pancreatic cell lines. Human viruses were also shown to be able to infect human pancreatic islets.
In situ
hybridization assays indicated that viral nucleoprotein could be detected in beta cells. The cytokine activation profile indicated a significant increase of MIG/CXCL9, IP-10/CXCL10, RANTES/CCL5, MIP1b/CCL4, Groa/CXCL1, interleukin 8 (IL-8)/CXCL8, tumor necrosis factor alpha (TNF-α), and IL-6. Our findings indicate that influenza virus infection may play a role as a causative agent of pancreatitis and diabetes in humans and other mammals.
Naturally Acquired Picornavirus Infections in Primates at the Dhaka Zoo
Authors: Oberste M. Steven , Feeroz Mohammed M. , Maher Kaija , Nix W. Allan , Engel Gregory A. , Begum Sajeda , Hasan Kamrul M. , Oh Gunwha , Pallansch Mark A. , Jones-Engel Lisa
Published: October 2012

The conditions in densely populated Bangladesh favor picornavirus transmission, resulting in a high rate of infection in the human population. Data suggest that nonhuman primates (NHP) may play a role in the maintenance and transmission of diverse picornaviruses in Bangladesh. At the Dhaka Zoo, multiple NHP species are caged in close proximity. Their proximity to other species and to humans, both zoo workers and visitors, provides the potential for cross-species transmission. To investigate possible interspecies and intraspecies transmission of picornaviruses among NHP, we collected fecal specimens from nine NHP taxa at the Dhaka Zoo at three time points, August 2007, January 2008, and June 2008. Specimens were screened using real-time PCR for the generaEnterovirus,Parechovirus,and Sapelovirus, and positive samples were typed by VP1 sequencing. Fifty-two picornaviruses comprising 10 distinct serotypes were detected in 83 fecal samples. Four of these serotypes, simian virus 19 (SV19), baboon enterovirus (BaEV), enterovirus 112 (EV112), and EV115, have been solely associated with infection in NHP. EV112, EV115, and SV19 accounted for 88% of all picornaviruses detected. Over 80% of samples from cages housing rhesus macaques, olive baboons, or hamadryas baboons were positive for a picornavirus, while no picornaviruses were detected in samples from capped langurs or vervet monkeys. In contrast to our findings among synanthropic NHP in Bangladesh where 100% of the picornaviruses detected were of human serotypes, in the zoo population, only 15% of picornaviruses detected in NHP were of human origin. Specific serotypes tended to persist over time, suggesting either persistent infection of individuals or cycles of reinfection.
Differential Effects of Human Immunodeficiency Virus Type 1 Capsid and Cellular Factors Nucleoporin 153 and LEDGF/p75 on the Efficiency and Specificity of Viral DNA Integration
Authors: Koh Yasuhiro , Wu Xiaolin , Ferris Andrea L. , Matreyek Kenneth A. , Smith Steven J. , Lee KyeongEun , KewalRamani Vineet N. , Hughes Stephen H. , Engelman Alan
Published: October 2012

Retroviruses integrate into cellular DNA nonrandomly. Lentiviruses such as human immunodeficiency virus type 1 (HIV-1) favor the bodies of active genes and gene-enriched transcriptionally active regions of chromosomes. The interaction between lentiviral integrase and the cellular protein lens epithelium-derived growth factor (LEDGF)/p75 underlies the targeting of gene bodies, whereas recent research has highlighted roles for the HIV-1 capsid (CA) protein and cellular factors implicated in viral nuclear import, including transportin 3 (TNPO3) and nucleoporin 358 (NUP358), in the targeting of gene-dense regions of chromosomes. Here, we show that CA mutations, which include the substitution of Asp for Asn74 (N74D), significantly reduce the dependency of HIV-1 on LEDGF/p75 during infection and that this difference correlates with the efficiency of viral DNA integration. The distribution of integration sites mapped by Illumina sequencing confirms that the N74D mutation reduces integration into gene-rich regions of chromosomes and gene bodies and reveals previously unrecognized roles for NUP153 (another HIV-1 cofactor implicated in viral nuclear import) and LEDGF/p75 in the targeting of the viral preintegration complex to gene-dense regions of chromatin. A role for the CA protein in determining the dependency of HIV-1 on LEDGF/p75 during infection highlights a connection between the viral capsid and chromosomal DNA integration.
Quantification of the Dynamics of Enterovirus 71 Infection by Experimental-Mathematical Investigation
Authors: Fukuhara Mitsuko , Iwami Shingo , Sato Kei , Nishimura Yorihiro , Shimizu Hiroyuki , Aihara Kazuyuki , Koyanagi Yoshio
Published: October 2012

Enterovirus 71 (EV71) is the causative agent of hand-foot-and-mouth disease and can trigger neurological disorders. EV71 outbreaks are a major public health concern in Asia-Pacific countries. By performing experimental-mathematical investigation, we demonstrate here that viral productivity and transmissibility but not viral cytotoxicity are drastically different among EV71 strains and can be associated with their epidemiological backgrounds. This is the first report demonstrating the dynamics of nonenveloped virus replication in cell culture using mathematical modeling.
A Novel Function of RNAs Arising From the Long Terminal Repeat of Human Endogenous Retrovirus 9 in Cell Cycle Arrest
Authors: Xu Lai , Elkahloun Abdel G. , Candotti Fabio , Grajkowski Andrzej , Beaucage Serge L. , Petricoin Emanuel F. , Calvert Valerie , Juhl Hartmut , Mills Frederick , Mason Karen , Shastri Neal , Chik Josh , Xu Cynthia , Rosenberg Amy S.
Published: October 2012

The human genome contains approximately 50 copies of the replication-defective human endogenous retrovirus 9 (ERV-9) and thousands of copies of its solitary long term repeat (sLTR) element. While some sLTRs are located upstream of critical genes and have enhancer activity, other sLTRs are located within introns and may be transcribed as RNAs. We found that intronic RNAs arising from U3 sLTRs of ERV-9 were expressed as both sense (S) and antisense (AS) transcripts in all human cells tested but that expression levels differed in malignant versus nonmalignant cells. In nonmalignant cells, AS was expressed at higher levels than S and at higher levels than in malignant cells; in malignant cells, AS was expressed at amounts equivalent to those of S RNA. Critically, U3 AS RNA was found to physically bind to key transcription factors for cellular proliferation, including NF-Y, p53, and sp1, indicating that such RNA transcripts may function as decoy targets or traps for NF-Y and thus inhibit the growth of human cancer cells. Indeed, short U3 oligodeoxynucleotides (ODNs) based on these RNA sequences ably inhibited proliferation of cancer cell lines driven by cyclins B1/B2, the gene targets of NF-Y.
Cellular LITAF Interacts with Frog Virus 3 75L Protein and Alters Its Subcellular Localization
Authors: Eaton Heather E. , Ferreira Lacerda Andressa , Desrochers Guillaume , Metcalf Julie , Angers Annie , Brunetti Craig R.
Published: October 2012

Iridoviruses are a family of large double-stranded DNA (dsDNA) viruses that are composed of 5 genera, including the
Lymphocystivirus
,
Ranavirus
,
Megalocytivirus
,
Iridovirus
, and
Chloriridovirus
genera. The frog virus 3 (FV3) 75L gene is a nonessential gene that is highly conserved throughout the members of the
Ranavirus
genus but is not found in other iridoviruses. FV3 75L shows high sequence similarity to a conserved domain found in the C terminus of LITAF, a small cellular protein with unknown function. Here we show that FV3 75L localizes to early endosomes, while LITAF localizes to late endosomes/lysosomes. Interestingly, when FV3 75L and LITAF are cotransfected into cells, LITAF can alter the subcellular localization of FV3 75L to late endosomes/lysosomes, where FV3 75L then colocalizes with LITAF. In addition, we demonstrated that virally produced 75L colocalizes with LITAF. We confirmed a physical interaction between LITAF and FV3 75L but found that this interaction was not mediated by two PPXY motifs in the N terminus of LITAF. Mutation of two PPXY motifs in LITAF did not affect the colocalization of LITAF and FV3 75L but did change the location of the two proteins from late endosomes/lysosomes to early endosomes.
Cooperativity in Virus Neutralization by Human Monoclonal Antibodies to Two Adjacent Regions Located at the Amino Terminus of Hepatitis C Virus E2 Glycoprotein
Authors: Keck Zhenyong , Wang Wenyan , Wang Yong , Lau Patrick , Carlsen Thomas H. R. , Prentoe Jannick , Xia Jinming , Patel Arvind H. , Bukh Jens , Foung Steven K. H.
Published: October 2012

A challenge for hepatitis C virus (HCV) vaccine development is defining conserved epitopes that induce protective antibodies against this highly diverse virus. An envelope glycoprotein (E2) segment located at amino acids (aa) 412 to 423 contains highly conserved neutralizing epitopes. While polyclonal antibodies to aa 412 to 423 from HCV-infected individuals confirmed broad neutralization, conflicting findings have been reported on polyclonal antibodies to an adjacent region, aa 434 to 446, that may or may not interfere with neutralization by antibodies to aa 412 to 423. To define the interplay between these antibodies, we isolated human monoclonal antibodies (HMAbs) to aa 412 to 423, designated HC33-related HMAbs (HC33 HMAbs), and characterized their interactions with other HMAbs to aa 434 to 446. A subset of the HC33 HMAbs neutralized genotype 1 to 6 infectious cell culture-derived HCV virions (HCVcc) with various activities. Although nonneutralizing HC33 HMAbs were isolated, they had lower binding affinities than neutralizing HC33 HMAbs. These antibodies could be converted to neutralizing antibodies by affinity maturation. Unidirectional competition for binding to E2 was observed between HC33 HMAbs and HMAbs to aa 434 to 446. When HMAbs to aa 434 to 446, which mediated neutralization, were combined with neutralizing HC33 HMAbs, biphasic patterns in neutralization were observed. A modest degree of antagonism was observed at lower concentrations, and a modest degree of synergism was observed at higher concentrations. However, the overall effect was additive neutralization. A similar pattern was observed when these antibodies were combined to block E2 binding to the HCV coreceptor, CD81. These findings demonstrate that both of these E2 regions participate in epitopes mediating virus neutralization and that the antibodies to aa 412 to 423 and aa 434 to 446 do not hinder their respective virus-neutralizing activities.
The C-Terminal Region of Rift Valley Fever Virus NSm Protein Targets the Protein to the Mitochondrial Outer Membrane and Exerts Antiapoptotic Function
Authors: Terasaki Kaori , Won Sungyong , Makino Shinji
Published: October 2012

The NSm nonstructural protein of Rift Valley fever virus (family
Bunyaviridae
, genus
Phlebovirus
) has an antiapoptotic function and affects viral pathogenesis. We found that NSm integrates into the mitochondrial outer membrane and that the protein's N terminus is exposed to the cytoplasm. The C-terminal region of NSm, which contains a basic amino acid cluster and a putative transmembrane domain, targeted the protein to the mitochondrial outer membrane and exerted antiapoptotic function.
Enterovirus 71 Uses Cell Surface Heparan Sulfate Glycosaminoglycan as an Attachment Receptor
Authors: Tan Chee Wah , Poh Chit Laa , Sam I-Ching , Chan Yoke Fun
Published: October 2012

Enterovirus 71 (EV-71) infections are usually associated with mild hand, foot, and mouth disease in young children but have been reported to cause severe neurological complications with high mortality rates. To date, four EV-71 receptors have been identified, but inhibition of these receptors by antagonists did not completely abolish EV-71 infection, implying that there is an as yet undiscovered receptor(s). Since EV-71 has a wide range of tissue tropisms, we hypothesize that EV-71 infections may be facilitated by using receptors that are widely expressed in all cell types, such as heparan sulfate. In this study, heparin, polysulfated dextran sulfate, and suramin were found to significantly prevent EV-71 infection. Heparin inhibited infection by all the EV-71 strains tested, including those with a single-passage history. Neutralization of the cell surface anionic charge by polycationic poly-
d
-lysine and blockage of heparan sulfate by an anti-heparan sulfate peptide also inhibited EV-71 infection. Interference with heparan sulfate biosynthesis either by sodium chlorate treatment or through transient knockdown of
N
-deacetylase/
N
-sulfotransferase-1 and exostosin-1 expression reduced EV-71 infection in RD cells. Enzymatic removal of cell surface heparan sulfate by heparinase I/II/III inhibited EV-71 infection. Furthermore, the level of EV-71 attachment to CHO cell lines that are variably deficient in cell surface glycosaminoglycans was significantly lower than that to wild-type CHO cells. Direct binding of EV-71 particles to heparin-Sepharose columns under physiological salt conditions was demonstrated. We conclude that EV-71 infection requires initial binding to heparan sulfate as an attachment receptor.
Efficient Herpes Simplex Virus 1 Replication Requires Cellular ATR Pathway Proteins
Authors: Mohni Kareem N. , Dee Alexander R. , Smith Samantha , Schumacher April J. , Weller Sandra K.
Published: October 2012

Herpes simplex virus 1 (HSV-1) is a double-stranded DNA virus that replicates in the nucleus of the host cell and is known to interact with several components of the cellular DNA-damage-signaling machinery. We have previously reported that the DNA damage response kinase, ATR, is specifically inactivated in HSV-1-infected cells. On the other hand, we have also shown that ATR and its scaffolding protein, ATRIP, are recruited to viral replication compartments, where they play beneficial roles during HSV-1 replication. In order to better understand this apparent discrepancy, we tested the hypothesis that some of the components of the ATR pathway may exert an antiviral effect on infection. In fact, we learned that all 10 of the canonical ATR pathway proteins are stable in HSV-infected cells and are recruited to viral replication compartments; furthermore, short hairpin RNA (shRNA) knockdown shows that several, including ATRIP, RPA70, TopBP1, Claspin, and CINP, are required for efficient HSV-1 replication. We also determined that activation of the ATR kinase prior to infection did not affect virus yield but did result in reduced levels of recombination between coinfecting viruses. Together, these data suggest that ATR pathway proteins are not antiviral
per se
but that activation of ATR signaling may have negative consequences during viral replication, such as inhibiting recombination.
The Host Proteins Transportin SR2/TNPO3 and Cyclophilin A Exert Opposing Effects on HIV-1 Uncoating
Authors: Shah Vaibhav B. , Shi Jiong , Hout David R. , Oztop Ilker , Krishnan Lavanya , Ahn Jinwoo , Shotwell Matthew S. , Engelman Alan , Aiken Christopher
Published: October 2012

Following entry of the HIV-1 core into target cells, productive infection depends on the proper disassembly of the viral capsid (uncoating). Although much is known regarding HIV-1 entry, the actions of host cell proteins that HIV-1 utilizes during early postentry steps are poorly understood. One such factor, transportin SR2 (TRN-SR2)/transportin 3 (TNPO3), promotes infection by HIV-1 and some other lentiviruses, and recent studies have genetically linked TNPO3 dependence of infection to the viral capsid protein (CA). Here we report that purified recombinant TNPO3 stimulates the uncoating of HIV-1 cores
in vitro
. The stimulatory effect was reduced by RanGTP, a known ligand for transportin family members. Depletion of TNPO3 in target cells rendered HIV-1 less susceptible to inhibition by PF74, a small-molecule HIV-1 inhibitor that induces premature uncoating. In contrast to the case for TNPO3, addition of the CA-binding host protein cyclophilin A (CypA) inhibited HIV-1 uncoating and reduced the stimulatory effect of TNPO3 on uncoating
in vitro
. In cells in which TNPO3 was depleted, HIV-1 infection was enhanced 4-fold by addition of cyclosporine, indicating that the requirement for TNPO3 in HIV-1 infection is modulated by CypA-CA interactions. Although TNPO3 was localized primarily to the cytoplasm, depletion of TNPO3 from target cells inhibited HIV-1 infection without reducing the accumulation of nuclear proviral DNA, suggesting that TNPO3 facilitates a stage of the virus life cycle subsequent to nuclear entry. Our results suggest that TNPO3 and cyclophilin A facilitate HIV-1 infection by coordinating proper uncoating of the core in target cells.
MICRO-FINITE ELEMENT MODELING OF WRINKLE FORMATION FOR CELL LOCOMOTION APPLICATIONS
Authors: MOHAMMADI HADI , HERZOG WALTER , MEQUANINT KIBRET
Published: October 2012

We developed a generic numerical procedure using the finite element method (FEM) for modeling wrinkles on an elastic thin membrane. The geometric characterization of wrinkles formed in an elastic membrane, which is caused by a crawling cell, may help in understanding the forces acting between a cell and its compliant substrate. In this study, the equivalent nodal forces, obtained from the virtual changes in the element internal energy, were differentiated to calculate the stiffness matrix. Micro-wrinkles can be computed as a function of membrane thickness which affects the stiffness matrix. This model may be used for applications in cell mechanobiology.
PREDICTION OF MUSCLE FORCES USING STATIC OPTIMIZATION FOR DIFFERENT CONTRACTILE CONDITIONS
Authors: ARSLAN YUNUS ZIYA , JINHA AZIM , KAYA MOTOSHI , HERZOG WALTER
Published: October 2012

In this study, we introduced a novel cost function for the prediction of individual muscle forces for a one degree-of-freedom musculoskeletal system. Unlike previous models, the new approach incorporates the instantaneous contractile conditions represented by the force-length and force-velocity relationships and accounts for physiological properties such as fiber type distribution and physiological cross-sectional area (PCSA) in the cost function. Using this cost function, it is possible to predict experimentally observed features of force-sharing among synergistic muscles that cannot be predicted using the classical approaches. Specifically, the new approach allows for predictions of force-sharing loops of agonistic muscles in one degree-of-freedom systems and for simultaneous increases in force in one muscle and decreases in a corresponding agonist. We concluded that the incorporation of the contractile conditions in the weighting of cost functions provides a natural way to incorporate observed force-sharing features in synergistic muscles that have eluded satisfactory description.
MOTION IN GAUGE THEORIES OF GRAVITY
Authors: TRESGUERRES ROMUALDO
Published: October 2012

A description of motion is proposed, adapted to the composite bundle interpretation of Poincaré Gauge Theory. Reference frames, relative positions and time evolution are characterized in gauge-theoretical terms. The approach is illustrated by an appropriate formulation of the familiar example of orbital motion induced by Schwarzschild spacetime.
DARK MATTER PRODUCTION BY TOPOLOGY CHANGE
Authors: TANAKA IZUMI , NAGAMI SEIJI
Published: October 2012

The purpose of this study is to investigate the new dark matter candidates and their production mechanisms. In this study, we concentrated on the dark matter production by analyzing the topology change. Our results revealed the variety of the dark matter productions.
A NEW GEOMETRICAL FRAMEWORK FOR THE DE BROGLIE–BOHM QUANTUM THEORY
Authors: HURLEY D. J. , VANDYCK M. A.
Published: October 2012

A geometrical framework for the de Broglie–Bohm quantum theory is presented, in which the trajectories of an N-particle system are interpretable as the integral curves of a particular vector field defined on a 3N-dimensional manifold [Formula: see text] constructed from physical space M. It is mathematically valid even when M is curved. If M is flat, the usual theory is recovered and automatically expressed in whatever curvilinear coordinates one may wish to choose. The general construction is illustrated by the case of a free particle moving on the surface of a sphere. (A modified Bohr quantization condition for angular momentum is obtained, with a first correction proportional to the curvature.) The Zeeman effect and some bound states on the sphere are also considered.
Corpus luteal contribution to maternal pregnancy physiology and outcomes in assisted reproductive technologies
Authors: Conrad Kirk P. , Baker Valerie L.
Published: October 2012

Investigations in the rat model of pregnancy indicate an important role for the corpus luteal (CL) hormone relaxin in the maternal circulatory and osmoregulatory changes in pregnancy, which are epitomized by profound vasodilation and modest hypoosmolality, respectively. In a pilot study of infertile women who became pregnant through donor eggs, in vitro fertilization, and embryo transfer, the gestational rise in glomerular filtration and fall in plasma osmolality were markedly subdued. Because these women were infertile, they lacked a CL and circulating relaxin (and possibly other vasoactive CL hormones). Based on these findings in pregnant rats and women, we hypothesize that infertile women conceiving through donor eggs will have overall subdued circulatory changes (e.g., attenuated reduction in systemic vascular resistance and subdued increase in cardiac output) particularly during early pregnancy when CL hormones predominate before the full development and maturation of the placenta. In contrast, infertile women conceiving by autologous eggs retrieved after ovarian stimulation and fresh embryo transfer may have a relatively hyperdynamic circulation due to the presence of many CL (up to 20 or more) and higher circulating levels of vasodilatory ovarian hormones such as relaxin. Emerging evidence suggests that women undergoing Assisted Reproductive Technologies (ART) have increased risk for adverse pregnancy outcomes such as preeclampsia and small for gestational-age babies. This increased risk may be partly caused by the maternal milieu, which is not physiological in ART pregnancies due to the abnormal status of the CL.
Aryl hydrocarbon receptor agonist, leflunomide, protects the ischemic-reperfused kidney: role of Tregs and stem cells
Authors: Baban Babak , Liu Jun Yao , Mozaffari Mahmood S.
Published: October 2012

The aryl hydrocarbon receptor (AHR) has emerged as a major modulator of inflammatory processes. We tested the hypothesis that AHR activation protects the ischemic-reperfused kidney in association with the suppression of the inflammatory response. Accordingly, male mice were treated with the nondioxin AHR agonist, leflunomide (40 mg/kg ip); vehicle-treated animals served as controls. Thereafter, the right kidney was subjected to an ischemia (45 min)-reperfusion (4 h) insult, while the left kidney served as a sham control. Renal cells prepared from ischemic-reperfused kidneys of leflunomide-treated mice displayed preservation of mitochondrial membrane potential (Ψm) and decreased apoptosis and necrosis compared with vehicle-treated ischemic-reperfused kidneys. Leflunomide treatment increased regulatory T cells (Tregs; forkhead box P3+) and IL-10-positive cells but reduced IL-17- and IL-23-expressing cells in both the peripheral blood and kidney cells, indicative of down-regulation of inflammatory responses. Leflunomide treatment also increased mobilization of stems cells subsets (i.e., mesenchymal and hematopoietic stem cells and endothelial progenitor cells) in the peripheral blood and promoted their recruitment into the ischemic-reperfused kidney. Collectively, the results indicate that AHR stimulation may represent a novel renoprotective mechanism likely involving mobilization and recruitment of Tregs and stem cells into the damaged kidney.
Disassociation of insulin action and Akt/FOXO signaling in skeletal muscle of older Akt-deficient mice
Authors: Reynolds Thomas H. , Merrell Erin , Cinquino Nicholas , Gaugler Megan , Ng Lily
Published: October 2012

The purpose of the present study was to determine the effect of Akt gene ablation on Akt/Forkhead Box O (FOXO) signaling and atrogene expression. This was accomplished by studying wild-type (WT) and isoform-specific Akt knockout (Akt1−/− and Akt2−/−) mice. The ability of insulin to promote Akt phosphorylation on Ser473 was significantly lower in extensor digitorum longus (EDL) and soleus muscles from Akt1−/− and Akt2−/− mice compared with WT mice. Total Akt1 protein levels were significantly lower in EDL muscles of Akt2−/− mice compared with WT mice, a process that appears to be posttranscriptionally regulated as Akt1 mRNA levels were unchanged. The loss of Akt1 protein in EDL muscles of Akt2−/− mice does not appear to be due to insulin resistance because 4 mo of a high-fat diet failed to reduce Akt1 protein levels in muscles of WT mice. Although FOXO3a phosphorylation and atrogin-1 expression were unaltered in muscles of Akt1−/− and Akt2−/− mice, the expression of the atrogenes Bnip3 and gabarapl were significantly elevated in muscles of both Akt1 and Akt2 knockout mice. Finally, the expression of striated activator of Rho signaling was significantly increased in muscles of Akt2−/− mice compared with Akt1−/− and WT mice. Our results demonstrate that the ablation of Akt isoforms disassociates insulin action and Akt/FOXO signaling to atrogenes.
Renal dopamine and angiotensin II receptor signaling in age-related hypertension
Authors: Chugh Gaurav , Pokkunuri Indira , Asghar Mohammad
Published: October 2012

Kidneys play a vital role in long-term regulation of blood pressure. This is achieved by actions of many renal and nonrenal factors acting on the kidney that help maintain the body's water and electrolyte balance and thus control blood pressure. Several endogenously formed or circulating hormones/peptides, by acting within the kidney, regulate fluid and water homeostasis and blood pressure. Dopamine and angiotensin II are the two key renal factors that, via acting on their receptors and counterregulating each other's function, maintain water and sodium balance. In this review, we provide recent advances in the signaling cascades of these renal receptors, especially at the level of their cross talk, and discuss their roles in blood pressure regulation in the aging process.
Immune reactivity to heat shock protein 70 expressed in the kidney is cause of salt-sensitive hypertension
Authors: Pons Héctor , Ferrebuz Atilio , Quiroz Yasmir , Romero-Vasquez Freddy , Parra Gustavo , Johnson Richard J. , Rodriguez-Iturbe Bernardo
Published: October 2012

Hypertension affects one-third of the adult population of the world. The causes of hypertension are incompletely understood, but relative impairment of sodium excretion is central to its pathogenesis. Immune cell infiltration in the kidney is a constant finding in hypertension that in association with local angiotensin and oxidants causes a defect in sodium excretion. However, it is unclear if the T cell influx into the kidney responds to nonspecific chemokine cues or is due to antigen-driven immune attraction. We found that T cells in experimentally induced salt-driven hypertension present a CD4 clonal response to heat shock protein 70 (HSP70) that is overexpressed in the kidney. We used a highly preserved amino acid sequence within the HSP molecule to induce immune tolerance associated with the generation of IL-10 producing regulatory T cells. Immune tolerant rats to HSP70 developed minimal renal inflammation and were protected from the development of salt-sensitive hypertension. Adoptive transfer of T lymphocytes isolated from spleen of tolerized rats also reversed hypertension. HSP70 gene delivery to the renal vein of the kidneys of rats sensitized to HSP70 caused an increment in blood pressure in response to a high-salt diet. The HSP70 peptide used in this work induces a strong proliferative response in peripheral blood lymphocytes of patients with essential hypertension. These studies provide evidence that autoimmunity plays a role in salt-sensitive hypertension and identifies HSP70 expressed in the kidney as one key antigen. These findings raise the possibility of novel approaches to the treatment of this condition.
On the origin of event-related potentials indexing covert attentional selection during visual search: timing of selection by macaque frontal eye field and event-related potentials during pop-out search
Authors: Purcell Braden A. , Schall Jeffrey D. , Woodman Geoffrey F.
Published: October 2012

Event-related potentials (ERPs) have provided crucial data concerning the time course of psychological processes, but the neural mechanisms producing ERP components remain poorly understood. This study continues a program of research in which we investigated the neural basis of attention-related ERP components by simultaneously recording intracranially and extracranially from macaque monkeys. Here, we compare the timing of attentional selection by the macaque homologue of the human N2pc component (m-N2pc) with the timing of selection in the frontal eye field (FEF), an attentional-control structure believed to influence posterior visual areas thought to generate the N2pc. We recorded FEF single-unit spiking and local field potentials (LFPs) simultaneously with the m-N2pc in monkeys performing an efficient pop-out search task. We assessed how the timing of attentional selection depends on task demands by direct comparison with a previous study of inefficient search in the same monkeys (e.g., finding a T among Ls). Target selection by FEF spikes, LFPs, and the m-N2pc was earlier during efficient pop-out search rather than during inefficient search. The timing and magnitude of selection in all three signals varied with set size during inefficient but not efficient search. During pop-out search, attentional selection was evident in FEF spiking and LFP before the m-N2pc, following the same sequence observed during inefficient search. These observations are consistent with the hypothesis that feedback from FEF modulates neural activity in posterior regions that appear to generate the m-N2pc even when competition for attention among items in a visual scene is minimal.