Neutralising antibodies for Mayaro virus in Pantanal, Brazil

The Pantanal hosts diverse wildlife species and therefore is a hotspot for arbovirus studies in South America. A serosurvey for Mayaro virus (MAYV), eastern (EEEV), western (WEEV) and Venezuelan (VEEV) equine encephalitis viruses was conducted with 237 sheep, 87 free-ranging caimans and 748 equids, including 37 collected from a ranch where a neurologic disorder outbreak had been recently reported. Sera were tested for specific viral antibodies using plaque-reduction neutralisation test. From a total of 748 equids, of which 264 were immunised with vaccine composed of EEEV and WEEV and 484 had no history of immunisation, 10 (1.3%) were seropositive for MAYV and two (0.3%) for VEEV using criteria of a ≥ 4-fold antibody titre difference. Among the 484 equids without history of immunisation, 48 (9.9%) were seropositive for EEEV and four (0.8%) for WEEV using the same criteria. Among the sheep, five were sero- positive for equine encephalitis alphaviruses, with one (0.4%) for EEEV, one (0.4%) for WEEV and three (1.3%) for VEEV. Regarding free-ranging caimans, one (1.1%) and three (3.4%), respectively, had low titres for neutralising antibodies to VEEV and undetermined alphaviruses. The neurological disorder outbreak could not be linked to the alphaviruses tested. Our findings represent strong evidence that MAYV and all equine encephalitis alphaviruses circulated in the Pantanal.     

Establishment of hepatitis E virus infection‐permissive and ‐non‐permissive human hepatoma PLC/PRF/5 subclones

PLC/PRF/5 cells show limited permissiveness, meaning that almost all subclones are permissive; however, some subclones do not exhibit permissiveness for hepatitis E virus (HEV) infection. In this study, the single‐cell cloning of PLC/PRF/5 was performed and heterogeneous subclones characterized. Notably, the efficiency of intracellular virus replication did not correlate with the permissiveness for HEV infection. However, as well as binding permissive subclones, virus‐like particles bound non‐permissive subclones on various levels, suggesting that these subclones have some deficiencies in the attachment and entry steps of infection. Our data would be useful for investigating the HEV life cycle.     

Effects of interaction between genetic variants in human leukocyte antigen DQ and granulysin genes in Chinese Han subjects infected with hepatitis B virus

Single nucleotide polymorphisms (SNPs) of HLA‐DQ and granulysin (GNLY) are reportedly associated with HBV infection. The aim of this study was to investigate the effects of interactions between SNPs in HLA‐DQ and GNLY on the outcome of hepatitis B virus (HBV) infection in Chinese Han subjects. HLA‐DQ (rs9275572) and GNLY (rs1866139 and rs11127) were genotyped in 310 subjects with HBV‐related chronic liver disease, 295 in whom spontaneous clearance of HBV had occurred and 316 who had not been exposed to HBV. HLA‐DQ rs9275572 was significantly correlated with HBV clearance (dominant genetic model: OR, 1.84; 95% CI, 1.30–2.61; adjusted P = 0.001). There was no statistical association of GNLY rs1866139 and rs11127with HBV infection outcomes. However, significant sex‐specific associations with HBV susceptibility were observed in men who carried rs1866139 CG or rs11127 TC and in women who carried rs1866139 GG or rs11127 CC. The findings were the same in the validation cohort, which was composed of 829 subjects. Based on a multifactor dimensionality reduction test with permutation correction, a three‐way interaction between SNPs in HLA‐DQ and GNLY was identified in terms of HBV clearance. In conclusion, additional evidence for an association of HLA‐DQ and GNLY SNPs with HBV infection outcomes has been identified and a SNP‐SNP interaction between HLA‐DQ and GNLY on HBV clearance observed.     

Exploring alternate states and oligomerization preferences of coiled‐coils by de novo structure modeling

Homomeric coiled‐coils can self‐assemble into a wide range of structural states with different helix topologies and oligomeric states. In this study, we have combined de novo structure modeling with stability calculations to simultaneously predict structure and oligomeric states of homomeric coiled‐coils. For dimers an asymmetric modeling protocol was developed. Modeling without symmetry constraints showed that backbone asymmetry is important for the formation of parallel dimeric coiled‐coils. Collectively, our results demonstrate that high‐resolution structure of coiled‐coils, as well as parallel and antiparallel orientations of dimers and tetramers, can be accurately predicted from sequence. De novo modeling was also used to generate models of competing oligomeric states, which were used to compare stabilities and thus predict the native stoichiometry from sequence. In a benchmark set of 33 coiled‐coil sequences, forming dimers to pentamers, up to 70% of the oligomeric states could be correctly predicted. The calculations demonstrated that the free energy of helix folding could be an important factor for determining stability and oligomeric state of homomeric coiled‐coils. The computational methods developed here should be broadly applicable to studies of sequence‐structure relationships in coiled‐coils and the design of higher order assemblies with improved oligomerization specificity. Proteins 2015; 83:235–247. © 2014 Wiley Periodicals, Inc.     

Molecular Evidence for the Association of Tomato leaf curl Gujarat virus with a Leaf Curl Disease of Phaseolus vulgaris L.

A leaf curl disease with symptoms typical of begomoviruses was observed in bean (Phaseolus vulgaris) at the Main Research Farm of the Indian Institute of Pulses Research, Kanpur, India. Infected plants had severe distortion of leaves and the plants were unproductive. PCR indicated the involvement of French bean leaf curl virus (JQ866297), a recently described Begomovirus, and Tomato leaf curl Gujarat virus (ToLCGV). The full‐length genome of ToLCGV associated with leaf curl disease of bean was 2757 nucleotides long and had maximum identity (97–98%) with seven isolates of ToLCGV (AY234383, AF449999, EU573714, GQ994098, AY190290, FR819708, AF413671) and is designated as Tomato leaf curl Gujarat virus‐(IN:Knp:Bean:2013) (KF440686). To the best of our knowledge, this is the first record of ToLCGV infecting a leguminous host, P. vulgaris.     

Molecular and Biological Analysis of Potato virus M (PVM) Isolates from the Czech Republic

The sequences of the 3′‐terminal region of four Czech Potato virus M isolates VIRUBRA 4/007, VIRUBRA 4/009, VIRUBRA 4/016 and VIRUBRA 4/035 were determined and compared with sequences of PVM isolates available in GenBank. Among the Czech isolates, VIRUBRA 4/007 and 4/016 as well as VIRUBRA 4/016 and 4/035 showed the highest nucleotide identity (93%). Isolates VIRUBRA 4/007, 4/016 and 4/035 were most similar to the PV0273 isolate from Germany and to the wild isolate from Russia. Interestingly, isolate VIRUBRA 4/009 significantly differed from the other three Czech isolates and was the only European isolate that showed the highest nucleotide identity with American isolates. Moreover, the PVM isolates from the Czech Republic and Germany differed in their host range. Phylogenetic analysis based on ORF5 coding for coat protein showed that the Czech isolates could be classified in two of the three groupings of the phylogenetic tree obtained. This is the first report on molecular and biological analysis of the genome sequences of PVM isolates from the Czech Republic.     

Molecular Detection and Characterization of Chinese Yam Mild Mosaic Virus Isolates

An improved RT‐PCR was developed and validated for the detection of Yam mild mosaic virus (YMMV). Sequences of the coat protein core region of 19 Chinese isolates were obtained, and analysis indicated the presence of different genetic variants. Phylogenetic analyses showed that the Chinese isolates were divided into two distinct clusters. Complete genomic sequences of two distinct Chinese variants were determined to be 9527 and 9529 nucleotides long, excluding the 3′ poly (A) tail. Their genomic structure and organization were virtually identical to that of a Brazilian isolate. The two variants shared identity of 87.3% to one another and 83.9–84.6% to the Brazilian variant at the genomic sequence level. Phylogenetic analyses supported that they represented two distinct YMMV lineages.     

Comparative Epidemiology of Three Virus Diseases on Zucchini Squash

Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus – type W (PRSV‐W) and Zucchini lethal chlorosis virus (ZLCV) cause important diseases on zucchini squash crops in Brazil. ZYMV and PRSV‐W belong to the genus Potyvirus and are transmitted by aphids, whereas ZLCV belongs to Tospovirus and is transmitted by the thrips Frankliniella zucchini. These three viruses may occur simultaneously in the field, and the epidemiology of the corresponding diseases may be determined by interactions among viruses, hosts and vectors. In this work, the progress of the diseases caused by these viruses was studied over a temporal and geographic range for three planting seasons (PS). For the lethal chlorosis (ZLCV), a monomolecular model was found to be the best fit for the data, though only during the third PS. For data collected during the first two PS, the Gompertz model was found to fit the data best. The spatial distribution of disease indicated disease aggregation at the end of the crop cycle. For the yellow mosaic (ZYMV), the model that best fit in the 1st PS was the logistic and in the 2nd and 3rd PS was monomolecular. The spatial pattern of the disease was random when the disease incidence was low but aggregated when the disease incidence was high. The common mosaic (PRSV‐W) showed the lowest incidence in all three PS. An exponential model was the best fit for data collected during all PS, and the spatial pattern of the disease was random. Interactions among the three viruses apparently did not result in changes in the epidemiology of the diseases. Removal of sources of inoculum and planting at an unfavourable time for reproduction of virus vectors are the two main measures recommended for the control of these diseases. The use of insecticide is indicated only for the control of the F. zucchini.