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The foot-and-mouth disease virus (FMDV) nonstructural protein 3A plays an important role in viral replication, virulence, and host range. It has been shown that deletions of 10 or 19-20 amino acids in the C-terminal half of 3A attenuate serotype O and C FMDVs, which replicate poorly in bovine cells but normally in porcine-derived cells, and the C-terminal half of 3A is not essential for serotype Asia1 FMDV replication in BHK-21 cells. In this study, we constructed a 3A deletion FMDV mutant based on a serotype O FMDV, the wild-type virus O/YS/CHA/05, with a 60-amino acid deletion in the 3A protein sequence, between residues 84 and 143. The rescued virus O/YS/CHA/05-Δ3A exhibited slower growth kinetics and formed smaller plaques compared to O/YS/CHA/05 in both BHK-21 and IBRS-2 cells, indicating that the 60-amino acid deletion in the 3A protein impaired FMDV replication. After 14 passages in BHK-21 cells, the replication capacity of the passaged virus O/YS/CHA/05-Δ3A-P14 returned to a level similar to the wild-type virus, suggesting that amino acid substitutions responsible for the enhanced replication capacity occurred in the genome of O/YS/CHA/05-Δ3A-P14. By sequence analysis, two amino acid substitutions, P153L in VP1 and T135I in 2C, were found in the O/YS/CHA/05-Δ3A-P14 genome compared to the O/YS/CHA/05-Δ3A genome. Subsequently, the amino acid substitutions VP1 P153L and 2C T135I were separately introduced into O/YS/CHA/05-Δ3A to rescue mutant viruses for examining their growth kinetics. Results showed that the 2C T135I instead of the VP1 P153L enhanced the virus replication capacity. The 2C T135I substitution also improved the replication of the wild-type virus, indicating that the effect of 2C T135I substitution on FMDV replication is not associated with the 3A deletion. Furthermore, our results showed that the T135I substitution in the nonstructural protein 2C enhanced O/YS/CHA/05 replication through promoting viral RNA synthesis.

Begomoviruses have increased pathogenicity because of their adaptation to a wide host range; consequently, these viruses cause a major loss to agroeconomic crops worldwide. In this study, we designed a gene construct representing an antisense coat protein gene. We also analyzed the efficacy of the induced resistance against Croton yellow vein mosaic virus (CrYVMV) affecting papaya in Nicotiana tabacum plants. Positive control plants developed typical leaf curl symptoms, whereas transgenic plants were symptomless. Moreover, the key component (i.e., short interfering RNA) of the antisense pathway was upregulated in transgenic plants. This finding demonstrates the activation of the gene silencing mechanism in transgenic plants. Thus, these results confirm that our construct is functional and effectively induces transient resistance against CrYVMV infections.

Tomato mild yellow leaf curl Aragua virus (ToMYLCV) is a begomovirus first reported infecting tomato (Solanum lycopersicum) and milkweed (Euphorbia heterophylla) in Venezuela. In this study, a ToMYLCV isolate (Zulia-219) was completely sequenced and its host range was evaluated. The DNA-A and DNA-B components of isolate Zulia-219 showed 93 and 85% nucleotide sequence identity with the respective counterparts of the ToMYLCV type strain. According to current demarcation criteria for begomovirus species, Zulia-219 is a new strain of ToMYLCV. Interestingly, tomato plants inoculated with ToMYLCV Zulia-219 displayed severe symptoms, including severe chlorotic leaf curling, in contrast to mild symptoms associated with the type strain of this begomovirus. These results indicate potential risks associated with this new ToMYLCV strain for tomato production in Venezuela.

Recognition of specific cell surface glycans, mediated by the VP8* domain of the spike protein VP4, is the essential first step in rotavirus (RV) infection. Due to lack of direct structural information of virus-ligand interactions, the molecular basis of ligand-controlled host ranges of the major human RVs (P[8] and P[4]) in P[II] genogroup remains unknown. Here, through characterization of a minor P[II] RV (P[19]) that can infect both animals (pigs) and humans, we made an important advance to fill this knowledge gap by solving the crystal structures of the P[19] VP8* in complex with its ligands. Our data showed that P[19] RVs use a novel binding site that differs from the known ones of other genotypes/genogroups. This binding site is capable of interacting with two types of glycans, the mucin core and type 1 histo-blood group antigens (HBGAs) with a common GlcNAc as the central binding saccharide. The binding site is apparently shared by other P[II] RVs and possibly two genotypes (P[10] and P[12]) in P[I] as shown by their highly conserved GlcNAc-interacting residues. These data provide strong evidence of evolutionary connections among these human and animal RVs, pointing to a common ancestor in P[I] with a possible animal host origin. While the binding properties to GlcNAc-containing saccharides are maintained, changes in binding to additional residues, such as those in the polymorphic type 1 HBGAs may occur in the course of RV evolution, explaining the complex P[II] genogroup that mainly causes diseases in humans but also in some animals.

The Poxviridae family is comprised of double-stranded DNA viruses belonging to nucleocytoplasmic large DNA viruses (NCLDV). Among the NCLDV, poxviruses exhibit the widest known host range, which is likely observed because this viral family has been more heavily investigated. However, relative to each member of the Poxviridae family, the spectrum of the host is variable, where certain viruses can infect a large range of hosts, while others are restricted to only one host species. It has been suggested that the variability in host spectrum among poxviruses is linked with the presence or absence of some host range genes. Would it be possible to extrapolate the restriction of viral replication in a specific cell lineage to an animal, a far more complex organism? In this study, we compare and discuss the relationship between the host range of poxvirus species and the abundance/diversity of host range genes. We analyzed the sequences of 38 previously identified and putative homologs of poxvirus host range genes, and updated these data with deposited sequences of new poxvirus genomes. Overall, the term host range genes might not be the most appropriate for these genes, since no correlation between them and the viruses' host spectrum was observed, and a change in nomenclature should be considered. Finally, we analyzed the evolutionary history of these genes, and reaffirmed the occurrence of horizontal gene transfer (HGT) for certain elements, as previously suggested. Considering the data presented in this study, it is not possible to associate the diversity of host range factors with the amount of hosts of known poxviruses, and this traditional nomenclature creates misunderstandings., Competing Interests: The authors declare no conflict of interest.

Background: Our main purpose was to evaluate the expression of plastic and evolved genes involved in ecological speciation in the noctuid moth Spodoptera frugiperda, the fall armyworm (FAW); and to demonstrate how host plants might influence lineage differentiation in this polyphagous insect. FAW is an important pest of several crops worldwide, and it is differentiated into host plant-related strains, corn (CS) and rice strains (RS). RNA-Seq and transcriptome characterization were applied to evaluate unbiased genetic expression differences in larvae from the two strains, fed on primary (corn) and alternative (rice) host plants. We consider that genes that are differently regulated by the same FAW strain, as a response to different hosts, are "plastic". Otherwise, differences in gene expression between the two strains fed on the same host are considered constitutive differences., Results: Individual performance parameters (larval and pupal weight) varied among conditions (strains vs. hosts). A total of 3657 contigs was related to plastic response, and 2395 contigs were differentially regulated in the two strains feeding on preferential and alternative hosts (constitutive contigs). Three molecular functions were present in all comparisons, both down- and up-regulated: oxidoreductase activity, metal-ion binding, and hydrolase activity., Conclusions: Metabolization of foreign chemicals is among the key functions involved in the phenotypic variation of FAW strains. From an agricultural perspective, high plasticity in families of detoxifying genes indicates the capacity for a rapid response to control compounds such as insecticides.

Ecotropic, xenotropic, and polytropic mouse leukemia viruses (E-, X-, and P-MLVs) exist in mice as infectious viruses and endogenous retroviruses (ERVs) inserted into mouse chromosomes. All three MLV subgroups are linked to leukemogenesis, which involves generation of recombinants with polytropic host range. Although P-MLVs are deemed to be the proximal agents of disease induction, few biologically characterized infectious P-MLVs have been sequenced for comparative analysis. We analyzed the complete genomes of 16 naturally occurring infectious P-MLVs, 12 of which were typed for pathogenic potential. We sought to identify ERV progenitors, recombinational hot spots, and segments that are always replaced, never replaced, or linked to pathogenesis or host range. Each P-MLV has an E-MLV backbone with P- or X-ERV replacements that together cover 100% of the recombinant genomes, with different substitution patterns for X- and P-ERVs. Two segments are always replaced, both coding for envelope (Env) protein segments: the N terminus of the surface subunit and the cytoplasmic tail R peptide. Viral gag gene replacements are influenced by host restriction genes Fv1 and Apobec3 Pathogenic potential maps to the env transmembrane subunit segment encoding the N-heptad repeat (HR1). Molecular dynamics simulations identified three novel interdomain salt bridges in the lymphomagenic virus HR1 that could affect structural stability, entry or sensitivity to host immune responses. The long terminal repeats of lymphomagenic P-MLVs are differentially altered by recombinations, duplications, or mutations. This analysis of the naturally occurring, sometimes pathogenic P-MLV recombinants defines the limits and extent of intersubgroup recombination and identifies specific sequence changes linked to pathogenesis and host interactions. IMPORTANCE During virus-induced leukemogenesis, ecotropic mouse leukemia viruses (MLVs) recombine with nonecotropic endogenous retroviruses (ERVs) to produce polytropic MLVs (P-MLVs). Analysis of 16 P-MLV genomes identified two segments consistently replaced: one at the envelope N terminus that alters receptor choice and one in the R peptide at the envelope C terminus, which is removed during virus assembly. Genome-wide analysis shows that nonecotropic replacements in the progenitor ecotropic MLV genome are more extensive than previously appreciated, covering 100% of the genome; contributions from xenotropic and polytropic ERVs differentially alter the regions responsible for receptor determination or subject to APOBEC3 and Fv1 restriction. All pathogenic viruses had modifications in the regulatory elements in their long terminal repeats and differed in a helical segment of envelope involved in entry and targeted by the host immune system. Virus-induced leukemogenesis thus involves generation of complex recombinants, and specific replacements are linked to pathogenesis and host restrictions., (Copyright © 2017 American Society for Microbiology.)

Recombinant vectors based on the human adenovirus type 5 (HAdV5) have been developed and extensively used in preclinical and clinical studies for over 30 years. However, certain restrictions of HAdV5-based vectors have limited their clinical applications because they are rather inefficient in specifically transducing cells of therapeutic interest that lack the coxsackievirus and adenovirus receptor (CAR). Moreover, enhanced vector-associated toxicity and widespread preexisting immunity have been shown to significantly hamper the effectiveness of HAdV-5-mediated gene transfer. However, evolution of adenoviruses in the natural host is driving the generation of novel types with altered virulence, enhanced transmission, and altered tissue tropism. As a consequence, an increasing number of alternative adenovirus types were identified, which may represent a valuable resource for the development of novel vector types. Thus, researchers are focusing on the other naturally occurring adenovirus types, which are structurally similar but functionally different from HAdV5. To this end, several strategies have been devised for getting genetic access to adenovirus genomes, resulting in a new panel of adenoviral vectors. Importantly, these vectors were shown to have a host range different from HAdV5 and to escape the anti-HAdV5 immune response, thus underlining the great potential of this approach. In summary, this review provides a state-of-the-art overview of one essential step in adenoviral vector development.

Bacterial etiolation and decline (BED), caused by Acidovorax avenae, is an emerging disease of creeping bentgrass on golf courses in the United States. We performed the first comprehensive analysis of A. avenae on a nationwide collection of turfgrass- and maize-pathogenic A. avenae. Surprisingly, our results reveal that the turfgrass-pathogenic A. avenae in North America are not only highly divergent but also belong to two distinct phylogroups. Both phylogroups specifically infect turfgrass but are more closely related to maize pathogens than to each other. This suggests that, although the disease is only recently reported, it has likely been infecting turfgrass for a long time. To identify a genetic basis for the host specificity, we searched for genes closely related among turfgrass strains but distantly related to their homologs from maize strains. We found a cluster of 11 such genes generated by three ancient recombination events within the type III secretion system (T3SS) pathogenicity island. Ever since the recombination, the cluster has been conserved by strong purifying selection, hinting at its selective importance. Together our analyses suggest that BED is an ancient disease that may owe its host specificity to a highly conserved cluster of 11 T3SS genes.

The "Papaya ringspot virus (PRSV) cluster" of cucurbit-infecting potyviruses contains five acknowledged species that have similar biological, serological and molecular properties. Additional data suggest there are other uncharacterized species from various locations in the world that likely belong to the PRSV cluster including a new PRSV-like virus reported from Sudan in 2003. Molecular and biological data indicated that the virus from Sudan belongs to a new species, tentatively named wild melon vein banding virus (WMVBV). The complete nucleotide sequence of a second virus from Sudan revealed it was a divergent relative of Moroccan watermelon mosaic virus (MWMV). Based on sequence similarity this virus was determined to be a distinct species and tentatively named Sudan watermelon mosaic virus (SuWMV). Molecular analyses indicate that SuWMV is a recombinant between WMVBV- and MWMV-related viruses. Based on surveys performed in Sudan between 1992 and 2012, SuWMV appeared 10 times more frequent than WMVBV in that country (14.6% vs. 1.5% of the samples tested). The geographic structure and molecular diversity patterns of the putative and acknowledged species suggest that the PRSV-like cluster originated in the Old World about 3600 years ago, with an important diversification in Africa., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Cucurbit yellow stunting disorder virus (CYSDV), emerged in the Sonoran Desert region of the southwestern USA in 2006 and has become well established. Symptoms induced by CYSDV infection include a striking interveinal chlorosis or yellowing and reduced yield and quality. The virus is transmitted by Bemisia tabaci, and the cryptic species MEAM1 has been present in the region since the early 1990s. CYSDV has now become the most economically important of the viruses affecting cucurbit production in the southwestern US. Here, we present a review of recent studies on CYSDV in the southwestern US, with implications for management of this virus throughout the world. Field surveys have established that CYSDV results in late-season infection of spring melon crops with limited economic impact; however, all summer and fall cucurbits become infected shortly after emergence due to high B. tabaci populations and abundant sources of inoculum. Studies have also demonstrated that CYSDV has an extensive host range among crops and weeds prevalent in the region. Recent studies demonstrated considerable variation in virus accumulation and transmission rates among the host plants evaluated as potential reservoirs. Cucurbit hosts had the highest CYSDV titers, were efficient sources for virus acquisition, and showed a positive correlation between titer in source plants and transmission to cucurbit plants. Non-cucurbit hosts had significantly lower CYSDV titers and varied in their capacity to serve as sources for transmission. Experiments demonstrated that multiple factors influence the efficiency with which a host plant species will be a reservoir for vector transmission of CYSDV to crops. Melon PI 313970 was identified as a new source of host plant resistance to CYSDV, in addition to the previously identified TGR 1551 (=PI 482420) and TGR 1937 (=PI 482431). Potential new sources of CYSDV resistance were identified by field screening of ca. 500 melon accessions with naturally occurring inoculum from 2007 through 2012. Host plant resistance to B. tabaci has also been identified in melon germplasm resistant to CYSDV and could be an important factor in reducing losses to CYSDV. Resistance to CYSDV is being transferred to US western shipping type cantaloupe and honeydew., (Published by Elsevier B.V.)

Tomato leaf curl New Delhi virus (ToLCNDV) is an exceptional Old World bipartite begomovirus. On the Indian subcontinent, a region in which monopartite DNA satellite-associated begomoviruses with mostly narrow geographical ranges predominate, it is widespread, with a geographical range also including the Far East, Middle East, North Africa and Europe. The success of ToLCNDV probably derives from its broad host range and highly flexible genomic configuration: its DNA-A component is capable of productively interacting with, and trans-replicating, diverse DNA-B components and betasatellites. An understanding of the capacity of ToLCNDV to infect a variety of hosts and spread across a broad and ecologically variable geographical range could illuminate the potential economic threats associated with similar begomoviral invasions. Towards this end, we used available ToLCNDV sequences to reconstruct the history of ToLCNDV spread., Taxonomy: Family Geminiviridae, Genus Begomovirus. ToLCNDV is a bipartite begomovirus. Following the revised begomovirus taxonomic criteria of 91% and 94% nucleotide identity for species and strain demarcation, respectively, ToLCNDV is a distinct species with two strains: ToLCNDV and ToLCNDV-Spain., Host Range: The primary cultivated host of ToLCNDV is tomato (Solanum lycopersicum), but the virus is also known to infect 43 other plant species from a range of families, including Cucurbitaceae, Euphorbiaceae, Solanaceae, Malvaceae and Fabaceae., Disease Symptoms: Typical symptoms of ToLCNDV infection in its various hosts include leaf curling, vein thickening, puckering, purpling/darkening of leaf margins, leaf area reduction, internode shortening and severe stunting., (© 2016 BSPP AND JOHN WILEY & SONS LTD.)

Wheat blast first emerged in Brazil in the mid-1980s and has recently caused heavy crop losses in Asia. Here we show how this devastating pathogen evolved in Brazil. Genetic analysis of host species determinants in the blast fungus resulted in the cloning of avirulence genes PWT3 and PWT4 , whose gene products elicit defense in wheat cultivars containing the corresponding resistance genes Rwt3 and Rwt4 Studies on avirulence and resistance gene distributions, together with historical data on wheat cultivation in Brazil, suggest that wheat blast emerged due to widespread deployment of rwt3 wheat (susceptible to Lolium isolates), followed by the loss of function of PWT3 This implies that the rwt3 wheat served as a springboard for the host jump to common wheat., (Copyright © 2017, American Association for the Advancement of Science.)

Symbiosis is a process that can generate evolutionary novelties and can extend the phenotypic niche space of organisms. Symbionts can act together with their hosts to co-construct host organs, within which symbionts are housed. Once established within hosts, symbionts can also influence various aspects of host phenotype, such as resource acquisition, protection from predation by acquisition of toxicity, as well as behaviour. Once symbiosis is established, its fidelity between generations must be ensured. Hosts evolve various mechanisms to screen unwanted symbionts and to facilitate faithful transmission of mutualistic partners between generations. Microbes are the most important symbionts that have influenced plant and animal phenotypes; multicellular organisms engage in developmental symbioses with microbes at many stages in ontogeny. The co-construction of niches may result in composite organisms that are physically nested within each other. While it has been advocated that these composite organisms need new evolutionary theories and perspectives to describe their properties and evolutionary trajectories, it appears that standard evolutionary theories are adequate to explore selection pressures on their composite or individual traits. Recent advances in our understanding of composite organisms open up many important questions regarding the stability and transmission of these units.

Tick-borne flaviviruses (TBFVs), including Powassan virus and tick-borne encephalitis virus cause encephalitis or hemorrhagic fevers in humans with case-fatality rates ranging from 1-30%. Despite severe disease in humans, TBFV infection of natural rodent hosts has little noticeable effect. Currently, the basis for resistance to disease is not known. We hypothesize that the coevolution of flaviviruses with their respective hosts has shaped the evolution of potent antiviral factors that suppress virus replication and protect the host from lethal infection. In the current study, we compared virus infection between reservoir host cells and related susceptible species. Infection of primary fibroblasts from the white-footed mouse (Peromyscus leucopus, a representative host) with a panel of vector-borne flaviviruses showed up to a 10,000-fold reduction in virus titer compared to control Mus musculus cells. Replication of vesicular stomatitis virus was equivalent in P. leucopus and M. musculus cells suggesting that restriction was flavivirus-specific. Step-wise comparison of the virus infection cycle revealed a significant block to viral RNA replication, but not virus entry, in P. leucopus cells. To understand the role of the type I interferon (IFN) response in virus restriction, we knocked down signal transducer and activator of transcription 1 (STAT1) or the type I IFN receptor (IFNAR1) by RNA interference. Loss of IFNAR1 or STAT1 significantly relieved the block in virus replication in P. leucopus cells. The major IFN antagonist encoded by TBFV, nonstructural protein 5, was functional in P. leucopus cells, thus ruling out ineffective viral antagonism of the host IFN response. Collectively, this work demonstrates that the IFN response of P. leucopus imparts a strong and virus-specific barrier to flavivirus replication. Future identification of the IFN-stimulated genes responsible for virus restriction specifically in P. leucopus will yield mechanistic insight into efficient control of virus replication and may inform the development of antiviral therapeutics.

Although introduced hemosporidian (malaria) parasites (Apicomplexa: Haemosporida) have hastened the extinction of endemic bird species in the Hawaiian Islands and perhaps elsewhere, little is known about the temporal dynamics of endemic malaria parasite populations. Haemosporidian parasites do not leave informative fossils, and records of population change are lacking beyond a few decades. Here, we take advantage of the isolation of West Indian land-bridge islands by rising postglacial sea levels to estimate rates of change in hemosporidian parasite assemblages over a millennial time frame. Several pairs of West Indian islands have been connected and separated by falling and rising sea levels associated with the advance and retreat of Pleistocene continental glaciers. We use island isolation following postglacial sea-level rise, ca. 2.5 ka, to characterize long-term change in insular assemblages of hemosporidian parasites. We find that assemblages on formerly connected islands are as differentiated as assemblages on islands that have never been connected, and both are more differentiated than local assemblages sampled up to two decades apart. Differentiation of parasite assemblages between formerly connected islands reflects variation in the prevalence of shared hemosporidian lineages, whereas differentiation between islands isolated by millions of years reflects replacement of hemosporidian lineages infecting similar assemblages of avian host species., Competing Interests: The authors declare no conflict of interest.

Sodium taurocholate cotransporting polypeptide (NTCP) is the functional receptor for human hepatitis B virus (HBV) and its satellite hepatitis D virus (HDV). Species barriers to HBV/HDV infection are mainly determined at entry level by variations in the sequences of particular NTCP orthologs. In this study, we sought to determine whether the NTCP ortholog in woodchuck (Marmota monax), woodchuck NTCP (wNTCP) supports viral infection. We found that wNTCP is capable of supporting HBV/HDV infection in HepG2 cells, but to much lower extent than human NTCP (hNTCP), which is about 90% reduction of hNTCP. Comprehensive site-directed mutagenesis mapping of hNTCP and wNTCP revealed that the residue at position 263 is a novel site crucial for viral entry. The important role of site 263 in infection is conserved among NTCP orthologs and may therefore be a potential target for blocking the viral entry., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Plant phylogeny constrains orchid mycorrhizal (OrM) fungal community composition in some orchids. Here, we investigated the structures of the OrM fungal communities of eight Dendrobium species in one niche to determine whether similarities in the OrM fungal communities correlated with the phylogeny of the host plants and whether the Dendrobium-OrM fungal interactions are phylogenetically conserved. A phylogeny based on DNA data was constructed for the eight coexisting Dendrobium species, and the OrM fungal communities were characterized by their roots. There were 31 different fungal lineages associated with the eight Dendrobium species. In total, 82.98% of the identified associations belonging to Tulasnellaceae, and a smaller proportion involved members of the unknown Basidiomycota (9.67%). Community analyses revealed that phylogenetically related Dendrobium tended to interact with a similar set of Tulasnellaceae fungi. The interactions between Dendrobium and Tulasnellaceae fungi were significantly influenced by the phylogenetic relationships among the Dendrobium species. Our results provide evidence that the mycorrhizal specificity in the eight coexisting Dendrobium species was phylogenetically conserved.

Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis globally. HEV comprises four genotypes with different geographic distributions and host ranges. We utilize this natural case-control study for investigating the evolution of zoonotic viruses compared to single-host viruses, using 244 near-full-length HEV genomes. Genome-wide estimates of the ratio of nonsynonymous to synonymous evolutionary changes ( dN / dS ratio) located a region of overlapping reading frames, which is subject to positive selection in genotypes 3 and 4. The open reading frames (ORFs) involved have functions related to host-pathogen interaction, so genotype-specific evolution of these regions may reflect their fitness. Bayesian inference of evolutionary rates shows that genotypes 3 and 4 have significantly higher rates than genotype 1 across all ORFs. Reconstruction of the phylogenies of zoonotic genotypes demonstrates significant intermingling of isolates between hosts. We speculate that the genotype-specific differences may result from cyclical adaptation to different hosts in genotypes 3 and 4. IMPORTANCE Hepatitis E virus (HEV) is increasingly recognized as a pathogen that affects both the developing and the developed world. While most often clinically mild, HEV can be severe or fatal in certain demographics, such as expectant mothers. Like many other viral pathogens, HEV has been classified into several distinct genotypes. We show that most of the HEV genome is evolutionarily constrained. One locus of positive selection is unusual in that it encodes two distinct protein products. We are the first to detect positive selection in this overlap region. Genotype 1, which infects humans only, appears to be evolving differently from genotypes 3 and 4, which infect multiple species, possibly because genotypes 3 and 4 are unable to achieve the same fitness due to repeated host jumps., (Copyright © 2017 American Society for Microbiology.)

Species of Proctoeces Odhner, 1911 (Trematoda: Fellodistomidae) have been reported from a wide range of marine animals globally. Members of the genus tend to lack strongly distinguishing morphological features for diagnosis, making identification difficult and the true number of species in the genus contentious. Combined morphological and molecular analyses were used to characterise three species of Proctoeces from Moreton Bay and the southern Great Barrier Reef. Data for two ribosomal regions and one mitochondrial region were generated for specimens collected from Australia. Three unique 18S-genotypes were identified which corresponded to subtle, but reliable, morphological differences. Two species of Proctoeces were identified from fishes of Moreton Bay, Proctoeces insolitus (Nicoll, 1915) Yamaguti, 1953 and P. major Yamaguti, 1934, and a third, P. choerodoni n. sp. from off Heron Island on the southern Great Barrier Reef. Phylogenetic analyses of partial 18S and partial 28S rDNA indicated that these three species differ from the four species reported outside of Australia for which sequence data are available. Phylogenetically, Proctoeces proved to be a reliable concept, with all species of Proctoeces that have been characterised genetically forming a well-supported clade in all analyses. Dramatically different patterns of host-specificity were identified for each of the three Australian species; P. insolitus apparently infects a single species of fish, P. choerodoni n. sp. infects multiple species of a single genus of fish, and P. major infects multiple species of two teleost orders., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

The fungus Parastagonospora nodorum is the causal agent of Septoria nodorum blotch of wheat (Triticum aestivum). The interaction is mediated by multiple fungal necrotrophic effector-dominant host sensitivity gene interactions. The three best-characterized effector-sensitivity gene systems are SnToxA-Tsn1, SnTox1-Snn1 and SnTox3-Snn3. These effector genes are highly expressed during early infection, but expression decreases as the infection progresses to tissue necrosis and sporulation. However, the mechanism of regulation is unknown. We have identified and functionally characterized a gene, referred to as PnPf2, which encodes a putative zinc finger transcription factor. PnPf2 deletion resulted in the down-regulation of SnToxA and SnTox3 expression. Virulence on Tsn1 and Snn3 wheat cultivars was strongly reduced. The SnTox1-Snn1 interaction remained unaffected. Furthermore, we have also identified and deleted an orthologous PtrPf2 from the tan spot fungus Pyrenophora tritici-repentis which possesses a near-identical ToxA that was acquired from P. nodorum via horizontal gene transfer. PtrPf2 deletion also resulted in the down-regulation of PtrToxA expression and a near-complete loss of virulence on Tsn1 wheat. We have demonstrated, for the first time, evidence for a functionally conserved signalling component that plays a role in the regulation of a common/horizontally transferred effector found in two major fungal pathogens of wheat., (© 2016 THE AUTHORS. MOLECULAR PLANT PATHOLOGY PUBLISHED BY BRITISH SOCIETY FOR PLANT PATHOLOGY AND JOHN WILEY & SONS LTD.)

Escherichia coli producing Shiga toxins (Stx) and the attaching-effacing (AE) lesion (AE-STEC) are responsible for (bloody) diarrhoea in humans and calves while the enteropathogenic E. coli (EPEC) producing the AE lesion only cause non-bloody diarrhoea in all mammals. The purpose of this study was (i) to identify the pathotypes of enterohaemolysin-producing E. coli isolated between 2009 and 2013 on EHLY agar from less than 2 month-old diarrhoeic calves with a triplex PCR targeting the stx1, stx2, eae virulence genes; (ii) to serotype the positive isolates with PCR targeting the genes coding for ten most frequent and pathogenic human and calf STEC O serogroups; and (iii) to compare the MLSTypes and virulotypes of calf and human O5 AE-STEC after Whole Genome Sequencing using two server databases (www.genomicepidemiology.org). Of 233 isolates, 206 were triplex PCR-positive: 119 AE-STEC (58%), 78 EPEC (38%) and 9 STEC (4%); and the stx1+eae+ AE-STEC (49.5%) were the most frequent. Of them, 120 isolates (84% of AE-STEC, 23% of EPEC, 22% of STEC) tested positive with one O serogroup PCR: 57 for O26 (47.5%), 36 for O111 (30%), 10 for O103 (8%) and 8 for O5 (7%) serogroups. The analysis of the draft sequences of 15 O5 AE-STEC could not identify any difference correlated to the host. As a conclusion, (i) the AE-STEC associated with diarrhoea in young calves still belong to the same serogroups as previously (O5, O26, O111) but the O103 serogroup may be emerging, (ii) the O5 AE-STEC from calves and humans are genetically similar., (Copyright © 2016. Published by Elsevier B.V.)

It is well documented that a variety of viral quasispecies are found in the patients with chronic infection of hepatitis C virus (HCV). However, the significance of quasispecies in the specific infectivity to individual cell types remains unknown. In the present study, we analyzed the role of quasispecies of the genotype 2a clone, JFH1 (HCVcc), in specific infectivity to the hepatic cell lines, Huh7.5.1 and Hep3B. HCV RNA was electroporated into Huh7.5.1 cells and Hep3B/miR-122 cells expressing miR-122 at a high level. Then, we adapted the viruses to Huh7 and Hep3B/miR-122 cells by serial passages and termed the resulting viruses HCVcc/Huh7 and HCVcc/Hep3B, respectively. Interestingly, a higher viral load was obtained in the homologous combination of HCVcc/Huh7 in Huh7.5.1 cells or HCVcc/Hep3B in Hep3B/miR-122 cells compared with the heterologous combination. By using a reverse genetics system and deep sequence analysis, we identified several adaptive mutations involved in the high affinity for each cell line, suggesting that quasispecies of HCV participate in cell-specific infectivity.

The soybean cyst nematode (SCN) Heterodera glycines is a major threat to soybean production, made more challenging by the current limitations of natural resistance for managing this pathogen. The use of resistant host cultivars is effective, but, over time, results in the generation of virulent nematode populations able to robustly parasitize the resistant host. In order to understand how virulence develops in SCN, we utilized a single backcross BC 1 F 2 strategy to mate a highly virulent inbred population (TN20), capable of reproducing on all current sources of resistance, with an avirulent one (PA3), unable to reproduce on any of the resistant soybean lines. The offspring were then investigated to determine how virulence is inherited on the main sources of SCN resistance, derived from soybean lines Peking, PI 88788, PI 90763, and the broad spectrum resistance source PI 437654. Significantly, our results suggest virulence on PI 437654 is a multigenic recessive trait that allows the nematode to reproduce on all current sources of resistance. In addition, we examined how virulence on different sources of resistance interact by placing virulent SCN populations under secondary selection, and identified a strong counter-selection between virulence on PI 88788- and PI 90763-derived resistances, while no such counter-selection existed between virulence on Peking and PI 88788 resistance sources. Our results suggest that the genes responsible for virulence on PI 88788 and PI 90763 may be different alleles at a common locus. If so, rotation of cultivars with resistance from these two sources may be an effective management protocol., (Copyright © 2017 Gardner et al.)

Background: Despite continuous research efforts, bacterio(phages) infecting Lactococcus lactis starter strains persist as a major threat to dairy fermentations. The lactococcal P335 phages, which are currently classified into four sub-groups (I-IV), are the second most frequently isolated phage group in an industrial dairy context., Results: The current work describes the isolation and comparative genomic analysis of 17 novel P335 group phages. Detailed analysis of the genomic region of P335 phages encoding the so-called "baseplate", which includes the receptor binding protein (RBP) was combined with a functional characterization of the RBP of sub-group III and IV phages. Additionally, calcium-dependence assays revealed a specific requirement for calcium by sub-group IV phages while host range analysis highlighted a higher number of strains with CWPS type A (11 of 39 strains) are infected by the P335 phages assessed in this study than those with a C (five strains), B (three of 39 strains) or unknown (one of 39 strains) CWPS type., Conclusions: These analyses revealed significant divergence among RBP sequences, apparently reflecting their unique interactions with the host and particularly for strains with a type A CWPS. The implications of the genomic architecture of lactococcal P335 phages on serving as a general model for Siphoviridae phages are discussed.

Enterocytozoon bieneusi is an obligate eukaryotic intracellular parasite that infects a wide variety of vertebrate and invertebrate hosts. Although considerable research has been conducted on this organism, relatively little information is available on the occurrence of E. bieneusi in captive Asiatic black bears. The present study was performed to determine the prevalence, genetic diversity, and zoonotic potential of E. bieneusi in captive Asiatic black bears in zoos in southwestern China. Fecal specimens from Asiatic black bears in four zoos, located in four different cities, were collected and analyzed for the prevalence of E. bieneusi. The average prevalence of E. bieneusi was 27.4% (29/106), with the highest prevalence in Guiyang Zoo (36.4%, 16/44). Altogether, five genotypes of E. bieneusi were identified among the 29 E. bieneusi-positive samples, including three known genotypes (CHB1, SC02, and horse2) and two novel genotypes named ABB1 and ABB2. Multi-locus sequence typing using three microsatellites (MS1, MS3, and MS7) and one minisatellite (MS4) revealed V, III, V, and IV genotypes at these four loci, respectively. Phylogenetic analysis showed that the genotypes SC02 and ABB2 were clustered into group 1 of zoonotic potential, the genotypes CHB1 and ABB1 were clustered into a new group, and the genotype horse2 was clustered into group 6 of unclear zoonotic potential. In conclusion, this study identified two novel E. bieneusi genotypes in captive Asiatic black bears, and used microsatellite and minisatellite markers to reveal E. bieneusi genetic diversity. Moreover, our findings show that genotypes SC02 (identified in humans) and ABB2 belong to group 1 with zoonotic potential, suggesting the risk of transmission of E. bieneusi from Asiatic black bears to humans and other animals., Competing Interests: The authors have declared that no competing interests exist.

Schizaphis graminum (green bug; GB) and Sipha flava (yellow sugarcane aphid; YSA) are two cereal aphid species with broad host ranges capable of establishing on sorghum (Sorghum bicolor) and several switchgrass (Panicum virgatum) cultivars. Switchgrass and sorghum are staple renewable bioenergy crops that are vulnerable to damage by aphids, therefore, identifying novel targets to control aphids has the potential to drastically improve yields and reduce losses in these bioenergy crops. Despite the wealth of genomic and transcriptomic information available from a closely related model aphid species, the pea aphid (Acyrthosiphon pisum), similar genomic information, including the identification of small RNAs, is still limited for GB and YSA. Deep sequencing of miRNAs expressed in GB and YSA was conducted and 72 and 56 miRNA candidates (including 14 and eight novel) were identified, respectively. Of the identified miRNAs, 45 were commonly expressed in both aphid species. Further, plant derived miRNAs were also detected in both aphid samples, including 13 (eight known and five novel) sorghum miRNAs and three (novel) barley miRNAs. In addition, potential aphid gene targets for the host plant-derived miRNAs were predicted. The establishment of miRNA repertoires in these two aphid species and the detection of plant-derived miRNA in aphids will ultimately lead to a better understanding of the role of miRNAs in regulating gene expression networks in these two aphids and the potential roles of plant miRNAs in mediating plant-insect interactions., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Colletotrichum graminicola and C. sublineola cause anthracnose leaf and stalk diseases of maize and sorghum, respectively. In spite of their close evolutionary relationship, the two species are completely host-specific. Host specificity is often attributed to pathogen virulence factors, including specialized secondary metabolites (SSM), and small-secreted protein (SSP) effectors. Genes relevant to these categories were manually annotated in two co-occurring, contemporaneous strains of C. graminicola and C. sublineola. A comparative genomic and phylogenetic analysis was performed to address the evolutionary relationships among these and other divergent gene families in the two strains., Results: Inoculation of maize with C. sublineola, or of sorghum with C. graminicola, resulted in rapid plant cell death at, or just after, the point of penetration. The two fungal genomes were very similar. More than 50% of the assemblies could be directly aligned, and more than 80% of the gene models were syntenous. More than 90% of the predicted proteins had orthologs in both species. Genes lacking orthologs in the other species (non-conserved genes) included many predicted to encode SSM-associated proteins and SSPs. Other common groups of non-conserved proteins included transporters, transcription factors, and CAZymes. Only 32 SSP genes appeared to be specific to C. graminicola, and 21 to C. sublineola. None of the SSM-associated genes were lineage-specific. Two different strains of C. graminicola, and three strains of C. sublineola, differed in no more than 1% percent of gene sequences from one another., Conclusions: Efficient non-host recognition of C. sublineola by maize, and of C. graminicola by sorghum, was observed in epidermal cells as a rapid deployment of visible resistance responses and plant cell death. Numerous non-conserved SSP and SSM-associated predicted proteins that could play a role in this non-host recognition were identified. Additional categories of genes that were also highly divergent suggested an important role for co-evolutionary adaptation to specific host environmental factors, in addition to aspects of initial recognition, in host specificity. This work provides a foundation for future functional studies aimed at clarifying the roles of these proteins, and the possibility of manipulating them to improve management of these two economically important diseases.

Human coronaviruses (HCoVs), including SARS-CoV and MERS-CoV, are zoonotic pathogens that originated in wild animals. HCoVs have large genomes that encode a fixed array of structural and nonstructural components, as well as a variety of accessory proteins that differ in number and sequence even among closely related CoVs. Thus, in addition to recombination and mutation, HCoV genomes evolve through gene gains and losses. In this review we summarize recent findings on the molecular evolution of HCoV genomes, with special attention to recombination and adaptive events that generated new viral species and contributed to host shifts and to HCoV emergence. VIDEO ABSTRACT., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Although genetic transfer between viruses and vertebrate hosts occurs less frequently than gene flow between bacteriophages and prokaryotes, it is extensive and has affected the evolution of both parties. With retroviruses, the integration of proviral DNA into chromosomal DNA can result in the activation of adjacent host gene expression and in the transduction of host transcripts into retroviral genomes as oncogenes. Yet in contrast to lysogenic phage, there is little evidence that viral oncogenes persist in a chain of natural transmission or that retroviral transduction is a significant driver of the horizontal spread of host genes. Conversely, integration of proviruses into the host germ line has generated endogenous retroviral genomes (ERV) in all vertebrate genomes sequenced to date. Some of these genomes retain potential infectivity and upon reactivation may transmit to other host species. During mammalian evolution, sequences of retroviral origin have been repurposed to serve host functions, such as the viral envelope glycoproteins crucial to the development of the placenta. Beyond retroviruses, DNA viruses with complex genomes have acquired numerous genes of host origin which influence replication, pathogenesis and immune evasion, while host species have accumulated germline sequences of both DNA and RNA viruses. A codicil is added on lateral transmission of cancer cells between hosts and on migration of host mitochondria into cancer cells.

Here we describe an initial characterization of Campylobacter jejuni bacteriophages by host range analysis, genome size determination by pulsed-field gel electrophoresis, and receptor-type identification by screening mutants for phage sensitivity.

Species of the Fusarium fujikuroi species complex (FFC) cause a wide spectrum of often devastating diseases on diverse agricultural crops, including coffee, fig, mango, maize, rice, and sugarcane. Although species within the FFC are difficult to distinguish by morphology, and their genes often share 90% sequence similarity, they can differ in host plant specificity and life style. FFC species can also produce structurally diverse secondary metabolites (SMs), including the mycotoxins fumonisins, fusarins, fusaric acid, and beauvericin, and the phytohormones gibberellins, auxins, and cytokinins. The spectrum of SMs produced can differ among closely related species, suggesting that SMs might be determinants of host specificity. To date, genomes of only a limited number of FFC species have been sequenced. Here, we provide draft genome sequences of three more members of the FFC: a single isolate of F. mangiferae, the cause of mango malformation, and two isolates of F. proliferatum, one a pathogen of maize and the other an orchid endophyte. We compared these genomes to publicly available genome sequences of three other FFC species. The comparisons revealed species-specific and isolate-specific differences in the composition and expression (in vitro and in planta) of genes involved in SM production including those for phytohormome biosynthesis. Such differences have the potential to impact host specificity and, as in the case of F. proliferatum, the pathogenic versus endophytic life style., (© The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Plasmodium reichenowi, an ape malaria parasite is morphologically identical and genetically similar to Plasmodium falciparum, infects chimpanzees but not humans. Genomic studies revealed that all primate malaria parasites belong to Laverania subgenus. Laverania parasites exhibit strict host specificity, but the molecular mechanisms underlying these host restrictions remain unexplained. Plasmodium merozoites express multiple binding ligands that recognize specific receptors on erythrocytes, including micronemal proteins belonging to P. falciparum EBL family. It was shown that erythrocyte binding antigen-175 (EBA-175), erythrocyte binding ligand-1 (EBL-1), erythrocyte binding antigen-140 (EBA-140) recognize erythrocyte surface sialoglycoproteins - glycophorins A, B, C, respectively. EBA-140 merozoite ligand hijacks glycophorin C (GPC), a minor erythrocyte sialoglycoprotein, to invade the erythrocyte through an alternative invasion pathway. A homolog of P. falciparum EBA-140 protein was identified in P. reichenowi. The amino acid sequences of both EBA-140 ligands are very similar, especially in the conservative erythrocyte binding region (Region II). It has been suggested that evolutionary changes in the sequence of EBL proteins may be associated with Plasmodium host restriction. In this study we obtained, for the first time, the recombinant P. reichenowi EBA-140 ligand Region II using baculovirus expression vector system. We show that the ape EBA-140 Region II is host specific and binds to chimpanzee erythrocytes in the dose and sialic acid dependent manner. Further identification of the erythrocyte receptor for this ape ligand is of great interests, since it may reveal the molecular basis of host restriction of both P. reichenowi and its deadliest human counterpart, P. falciparum., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Animal guts are often colonized by host-specialized bacterial species to the exclusion of other transient microorganisms, but the genetic basis of colonization ability is largely unknown. The bacterium Snodgrassella alvi is a dominant gut symbiont in honey bees, specialized in colonizing the hindgut epithelium. We developed methods for transposon-based mutagenesis in S. alvi and, using high-throughput DNA sequencing, screened genome-wide transposon insertion (Tn-seq) and transcriptome (RNA-seq) libraries to characterize both the essential genome and the genes facilitating host colonization. Comparison of Tn-seq results from laboratory cultures and from monoinoculated worker bees reveal that 519 of 2,226 protein-coding genes in S. alvi are essential in culture, whereas 399 are not essential but are beneficial for gut colonization. Genes facilitating colonization fall into three broad functional categories: extracellular interactions, metabolism, and stress responses. Extracellular components with strong fitness benefits in vivo include trimeric autotransporter adhesins, O antigens, and type IV pili (T4P). Experiments with T4P mutants establish that T4P in S. alvi likely function in attachment and biofilm formation, with knockouts experiencing a competitive disadvantage in vivo. Metabolic processes promoting colonization include essential amino acid biosynthesis and iron acquisition pathways, implying nutrient scarcity within the hindgut environment. Mechanisms to deal with various stressors, such as for the repair of double-stranded DNA breaks and protein quality control, are also critical in vivo. This genome-wide study identifies numerous genetic networks underlying colonization by a gut commensal in its native host environment, including some known from more targeted studies in other host-microbe symbioses., Competing Interests: The authors declare no conflict of interest.

Chronic wasting disease (CWD) in cervids and bovine spongiform encephalopathy (BSE) in cattle are prion diseases that are caused by the same protein-misfolding mechanism, but they appear to pose different risks to humans. We are interested in understanding the differences between the species barriers of CWD and BSE. We used real-time, quaking-induced conversion (RT-QuIC) to model the central molecular event in prion disease, the templated misfolding of the normal prion protein, PrP c , to a pathogenic, amyloid isoform, scrapie prion protein, PrP Sc We examined the role of the PrP c amino-terminal domain (N-terminal domain [NTD], amino acids [aa] 23 to 90) in cross-species conversion by comparing the conversion efficiency of various prion seeds in either full-length (aa 23 to 231) or truncated (aa 90 to 231) PrP c We demonstrate that the presence of white-tailed deer and bovine NTDs hindered seeded conversion of PrP c , but human and bank vole NTDs did the opposite. Additionally, full-length human and bank vole PrP c s were more likely to be converted to amyloid by CWD prions than were their truncated forms. A chimera with replacement of the human NTD by the bovine NTD resembled human PrP c The requirement for an NTD, but not for the specific human sequence, suggests that the NTD interacts with other regions of the human PrP c to increase promiscuity. These data contribute to the evidence that, in addition to primary sequence, prion species barriers are controlled by interactions of the substrate NTD with the rest of the substrate PrP c molecule., Importance: We demonstrate that the amino-terminal domain of the normal prion protein, PrP c , hinders seeded conversion of bovine and white-tailed deer PrP c s to the prion forms, but it facilitates conversion of the human and bank vole PrP c s to the prion forms. Additionally, we demonstrate that the amino-terminal domain of human and bank vole PrP c s requires interaction with the rest of the molecule to facilitate conversion by CWD prions. These data suggest that interactions of the amino-terminal domain with the rest of the PrP c molecule play an important role in the susceptibility of humans to CWD prions., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

The P[19] genotype belongs to the P[II] genogroup of group A rotaviruses (RVs). However, unlike the other P[II] RVs, which mainly infect humans, P[19] RVs commonly infect animals (pigs), making P[19] unique for the study of RV diversity and host ranges. Through in vitro binding assays and saturation transfer difference (STD) nuclear magnetic resonance (NMR), we found that P[19] could bind mucin cores 2, 4, and 6, as well as type 1 histo-blood group antigens (HBGAs). The common sequences of these glycans serve as minimal binding units, while additional residues, such as the A, B, H, and Lewis epitopes of the type 1 HBGAs, can further define the binding outcomes and therefore likely the host ranges for P[19] RVs. This complex binding property of P[19] is shared with the other three P[II] RVs (P[4], P[6], and P[8]) in that all of them recognized the type 1 HBGA precursor, although P[4] and P[8], but not P[6], also bind to mucin cores. Moreover, while essential for P[4] and P[8] binding, the addition of the Lewis epitope blocked P[6] and P[19] binding to type 1 HBGAs. Chemical-shift NMR of P[19] VP8* identified a ligand binding interface that has shifted away from the known RV P-genotype binding sites but is conserved among all P[II] RVs and two P[I] RVs (P[10] and P[12]), suggesting an evolutionary connection among these human and animal RVs. Taken together, these data are important for hypotheses on potential mechanisms for RV diversity, host ranges, and cross-species transmission., Importance: In this study, we found that our P[19] strain and other P[II] RVs recognize mucin cores and the type 1 HBGA precursors as the minimal functional units and that additional saccharides adjacent to these units can alter binding outcomes and thereby possibly host ranges. These data may help to explain why some P[II] RVs, such as P[6] and P[19], commonly infect animals but rarely humans, while others, such as the P[4] and P[8] RVs, mainly infect humans and are predominant over other P genotypes. Elucidation of the molecular bases for strain-specific host ranges and cross-species transmission of these human and animal RVs is important to understand RV epidemiology and disease burden, which may impact development of control and prevention strategies against RV gastroenteritis., (Copyright © 2016 Liu et al.)

Several studies have demonstrated that E. coli appears to display some level of host adaptation and specificity. Recent studies in our laboratory support these findings as determined by logic regression modeling of single nucleotide polymorphisms (SNP) in intergenic regions (ITGRs). We sought to determine the degree of host-specific information encoded in various ITGRs across a library of animal E. coli isolates using both whole genome analysis and a targeted ITGR sequencing approach. Our findings demonstrated that ITGRs across the genome encode various degrees of host-specific information. Incorporating multiple ITGRs (i.e., concatenation) into logic regression model building resulted in greater host-specificity and sensitivity outcomes in biomarkers, but the overall level of polymorphism in an ITGR did not correlate with the degree of host-specificity encoded in the ITGR. This suggests that distinct SNPs in ITGRs may be more important in defining host-specificity than overall sequence variation, explaining why traditional unsupervised learning phylogenetic approaches may be less informative in terms of revealing host-specific information encoded in DNA sequence. In silico analysis of 80 candidate ITGRs from publically available E. coli genomes was performed as a tool for discovering highly host-specific ITGRs. In one ITGR (ydeR-yedS) we identified a SNP biomarker that was 98% specific for cattle and for which 92% of all E. coli isolates originating from cattle carried this unique biomarker. In the case of humans, a host-specific biomarker (98% specificity) was identified in the concatenated ITGR sequences of rcsD-ompC, ydeR-yedS, and rclR-ykgE, and for which 78% of E. coli originating from humans carried this biomarker. Interestingly, human-specific biomarkers were dominant in ITGRs regulating antibiotic resistance, whereas in cattle host-specific biomarkers were found in ITGRs involved in stress regulation. These data suggest that evolution towards host specificity may be driven by different natural selection pressures on the regulome of E. coli among different animal hosts., (Copyright © 2016 Elsevier Inc. All rights reserved.)

A new dactylogyrid monogenean Dactylogyrus bicorniculus sp. nov. is described from the gills of the kazetoge bitterling, Rhodeus atremius atremius (Jordan and Thompson, 1914), an endemic species in Japan, from Saga Prefecture, northern Kyūshū. D. bicorniculus sp. nov. resembles Dactylogyrus bicornis Malevitskaja, 1941 and Dactylogyrus lophogonus Zhang and Ji, 1980 because they have two common features, a large V-shaped ventral bar and well-developed second marginal hooks. However, the new species is distinguished from these congeners by a shorter penis and an accessory piece. A phylogenetic analysis of 28S rDNA shows that D. bicorniculus sp. nov. is a basal species with the T-shaped ventral bar in the genus. The new species has strict host-specificity to R. a. atremius, one of the endangered freshwater fishes in Japan, and may face the danger of co-extinction with its host., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

The objectives of this study were to examine the genetic variation in fruit flies (Diptera: Tephritidae) in Thailand and to test the efficiency of the mitochondrial cytochrome c oxidase subunit I (COI) barcoding region for species-level identification. Twelve fruit fly species were collected from 24 host plant species of 13 families. The number of host plant species for each fruit fly species ranged between 1 and 11, with Bactrocera correcta found in the most diverse host plants. A total of 123 COI sequences were obtained from these fruit fly species. Sequences from the NCBI database were also included, for a total of 17 species analyzed. DNA barcoding identification analysis based on the best close match method revealed a good performance, with 94.4% of specimens correctly identified. However, many specimens (3.6%) had ambiguous identification, mostly due to intra- and interspecific overlap between members of the B. dorsalis complex. A phylogenetic tree based on the mitochondrial barcode sequences indicated that all species, except for the members of the B. dorsalis complex, were monophyletic with strong support. Our work supports recent calls for synonymization of these species. Divergent lineages were observed within B. correcta and B. tuberculata, and this suggested that these species need further taxonomic reexamination.

Recent studies have revealed a large diversity of Plasmodium spp. among African great apes. Some of these species are related to Plasmodium falciparum, the most virulent agent of human malaria (subgenus Laverania), and others to Plasmodium ovale, Plasmodium malariae and Plasmodium vivax (subgenus Plasmodium), three other human malaria agents. Laverania parasites exhibit strict host specificity in their natural environment. Plasmodium reichenowi, Plasmodium billcollinsi, Plasmodium billbrayi and Plasmodium gaboni infect only chimpanzees, while Plasmodium praefalciparum, Plasmodium blacklocki and Plasmodium adleri are restricted to gorillas and Plasmodium falciparum is pandemic in humans. This host specificity may be due to genetic and/or environmental factors. Infrastructures hosting captive primates, such as sanctuaries and health centres, usually concentrate different primate species, thus favouring pathogen exchanges. Using molecular tools, we analysed blood samples from captive non-human primates living in Gabon to evaluate the risk of Plasmodium spp. transfers between host species. We also included blood samples from workers taking care of primates to assess whether primate-human parasite transfers occurred. We detected four transfers of Plasmodium from gorillas towards chimpanzees, one from chimpanzees to gorillas, three from humans towards chimpanzees and one from humans to mandrills. No simian Plasmodium was found in the blood samples from humans working with primates. These findings demonstrate that the genetic barrier that determines the apparent host specificity of Laverania is not completely impermeable and that parasite exchanges between gorillas and chimpanzees are possible in confined environments., (Copyright © 2016 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Unlabelled: Two alleles of segment 8 (NS) circulate in nonchiropteran influenza A viruses. The A allele is found in avian and mammalian viruses, but the B allele is viewed as being almost exclusively found in avian viruses. This might reflect the fact that one or both of its encoded proteins (NS1 and NEP) are maladapted for replication in mammalian hosts. To test this, a number of clade A and B avian virus-derived NS segments were introduced into human H1N1 and H3N2 viruses. In no case was the peak virus titer substantially reduced following infection of various mammalian cell types. Exemplar reassortant viruses also replicated to similar titers in mice, although mice infected with viruses with the avian virus-derived segment 8s had reduced weight loss compared to that achieved in mice infected with the A/Puerto Rico/8/1934 (H1N1) parent. In vitro, the viruses coped similarly with type I interferons. Temporal proteomics analysis of cellular responses to infection showed that the avian virus-derived NS segments provoked lower levels of expression of interferon-stimulated genes in cells than wild type-derived NS segments. Thus, neither the A nor the B allele of avian virus-derived NS segments necessarily attenuates virus replication in a mammalian host, although the alleles can attenuate disease. Phylogenetic analyses identified 32 independent incursions of an avian virus-derived A allele into mammals, whereas 6 introductions of a B allele were identified. However, A-allele isolates from birds outnumbered B-allele isolates, and the relative rates of Aves-to-Mammalia transmission were not significantly different. We conclude that while the introduction of an avian virus segment 8 into mammals is a relatively rare event, the dogma of the B allele being especially restricted is misleading, with implications in the assessment of the pandemic potential of avian influenza viruses., Importance: Influenza A virus (IAV) can adapt to poultry and mammalian species, inflicting a great socioeconomic burden on farming and health care sectors. Host adaptation likely involves multiple viral factors. Here, we investigated the role of IAV segment 8. Segment 8 has evolved into two distinct clades: the A and B alleles. The B-allele genes have previously been suggested to be restricted to avian virus species. We introduced a selection of avian virus A- and B-allele segment 8s into human H1N1 and H3N2 virus backgrounds and found that these reassortant viruses were fully competent in mammalian host systems. We also analyzed the currently available public data on the segment 8 gene distribution and found surprisingly little evidence for specific avian host restriction of the B-clade segment. We conclude that B-allele segment 8 genes are, in fact, capable of supporting infection in mammals and that they should be considered during the assessment of the pandemic risk of zoonotic influenza A viruses., (Copyright © 2016 Turnbull et al.)

Aeromonas hydrophila is an opportunistic pathogenic bacterium causing diseases in human and fish. The emergence of multidrug-resistant A. hydrophila isolates has been increasing in recent years. In this study, we have isolated a novel virulent podophage of A. hydrophila, designated as Ahp1, from waste water. Ahp1 has a rapid adsorption (96% adsorbed in 2 min), a latent period of 15 min, and a burst size of 112 PFU per infected cell. At least eighteen Ahp1 virion proteins were visualized in SDS-polyacrylamide gel electrophoresis, with a 36-kDa protein being the predicted major capsid protein. Genome analysis of Ahp1 revealed a linear doubled-stranded DNA genome of 42,167 bp with a G + C content of 58.8%. The genome encodes 46 putative open reading frames, 5 putative phage promoters, and 3 transcriptional terminators. Based on high degrees of similarity in overall genome organization and among most of the corresponding ORFs, as well as phylogenetic relatedness among their DNAP, RNAP and major capsid proteins, we propose a new subgroup, designated Ahp1-like subgroup. This subgroup contains Ahp1 and members previously belonging to phiKMV-like subgroup, phiAS7, phi80-18, GAP227, phiR8-01, and ISAO8. Since Ahp1 has a narrow host range, for effective phage therapy, different phages are needed for preparation of cocktails that are capable of killing the heterogeneous A. hydrophila strains., Competing Interests: The authors have declared that no competing interests exist.

Cultivated common beans are the primary protein source for millions of people around the world who subsist on low-input agriculture, enabled by the symbiotic N2 -fixation these legumes perform in association with rhizobia. Within a single agricultural plot, multiple Rhizobium species can nodulate bean roots, but it is unclear how genetically isolated these species remain in sympatry. To better understand this issue, we sequenced and compared the genomes of 33 strains isolated from the rhizosphere and root nodules of a particular bean variety grown in the same agricultural plot. We found that the Rhizobium species we observed coexist with low genetic recombination across their core genomes. Accessory plasmids thought to be necessary for the saprophytic lifestyle in soil show similar levels of genetic isolation, but with higher rates of recombination than the chromosomes. However, the symbiotic plasmids are extremely similar, with high rates of recombination and do not appear to have co-evolved with the chromosome or accessory plasmids. Therefore, while Rhizobium species are genetically isolated units within the microbial community, a common symbiotic plasmid allows all Rhizobium species to engage in symbiosis with the same host in a single agricultural plot., (© 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Leucocytozoon, a widespread hemosporidian blood parasite that infects a broad group of avian families, has been studied in corvids (family: Corvidae) for over a century. Current taxonomic classification indicates that Leucocytozoon sakharoffi infects crows and related Corvus spp., while Leucocytozoon berestneffi infects magpies (Pica spp.) and blue jays (Cyanocitta sp.). This intrafamily host specificity was based on the experimental transmissibility of the parasites, as well as slight differences in their morphology and life cycle development. Genetic sequence data from Leucocytozoon spp. infecting corvids is scarce, and until the present study, sequence data has not been analyzed to confirm the current taxonomic distinctions. Here, we predict the phylogenetic relationships of Leucocytozoon cytochrome b lineages recovered from infected American Crows (Corvus brachyrhynchos), yellow-billed magpies (Pica nuttalli), and Steller's jays (Cyanocitta stelleri) to explore the host specificity pattern of L. sakharoffi and L. berestneffi. Phylogenetic reconstruction revealed a single large clade containing nearly every lineage recovered from the three host species, while showing no evidence of the expected distinction between L. sakharoffi and L. berestneffi. In addition, five of the detected lineages were recovered from both crows and magpies. This absence of the previously described host specificity in corvid Leucocytozoon spp. suggests that L. sakharoffi and L. berestneffi be reexamined from a taxonomic perspective.

Background: Many species belonging to the genus Colletotrichum cause anthracnose disease on a wide range of plant species. In addition to their economic impact, the genus Colletotrichum is a useful model for the study of the evolution of host specificity, speciation and reproductive behaviors. Genome projects of Colletotrichum species have already opened a new era for studying the evolution of pathogenesis in fungi., Results: We sequenced and annotated the genomes of four strains in the Colletotrichum acutatum species complex (CAsc), a clade of broad host range pathogens within the genus. The four CAsc proteomes and secretomes along with those representing an additional 13 species (six Colletotrichum spp. and seven other Sordariomycetes) were classified into protein families using a variety of tools. Hierarchical clustering of gene family and functional domain assignments, and phylogenetic analyses revealed lineage specific losses of carbohydrate-active enzymes (CAZymes) and proteases encoding genes in Colletotrichum species that have narrow host range as well as duplications of these families in the CAsc. We also found a lineage specific expansion of necrosis and ethylene-inducing peptide 1 (Nep1)-like protein (NLPs) families within the CAsc., Conclusions: This study illustrates the plasticity of Colletotrichum genomes, and shows that major changes in host range are associated with relatively recent changes in gene content.

Background: The swine-origin influenza A (H1N1) virus (S-OIV) has come to the forefront since 2009 and was identified as a new reassortant strain. The hemagglutinin (HA) glycoprotein mediates virus binding, contains antigenic regions recognized by neutralizing antibodies, and is associated with viral cross-species infection and adaption. The comparison study of codon usage preferences in influenza viral genomes was less extensive. In this study, we used codon usage pattern analyses to validate the adaption and origins of S-OIV., Methods: Codon usage pattern was used to estimate the host adaption of S-OIVs. Phylogenetic analysis of the HA gene was conducted to understand the phylogeny of H1N1 viruses isolated from different hosts. Amino acid signature pattern on antigenic sites of HA was analyzed to understand the antigenic characteristics., Results: Results of phylogenetic analyses of HA gene indicate that S-OIVs group in identical clusters. The synonymous codon usage pattern analyses indicate that the effective number of codons versus GC content at the third codon position in the HA1 gene slightly differ from those in swine H1N1 and gradually adapted to human. Our data indicate that S-OIV evolution occurred according to positive selection within these antigenic regions. A comparison of antigenic site amino acids reveals similar signature patterns between S-OIV and 1918 human influenza strains., Conclusion: This study proposes a new and effective way to gain a better understanding of the features of the S-OIV genome and evolutionary processes based on the codon usage pattern. It is useful to trace influenza viral origins and cross-species virus transmission., (Copyright © 2014. Published by Elsevier B.V.)

Interspecific hybridization is widely observed within diverse eukaryotic taxa, and is considered an important driver for genome evolution. As hybridization fuels genomic and transcriptional alterations, hybrids are adept to respond to environmental changes or to invade novel niches. This may be particularly relevant for organisms that establish symbiotic relationships with host organisms, such as mutualistic symbionts, endophytes and pathogens. The latter group is especially well-known for engaging in everlasting arms races with their hosts. Illustrated by the increased identification of hybrid pathogens with altered virulence or host ranges when compared with their parental lineages, it appears that hybridization is a strong driver for pathogen evolution, and may thus significantly impact agriculture and natural ecosystems., (Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The aim of this study was to compare the dynamics of three previously isolated bacteriophages (or phages) individually (phSE-1, phSE-2 and phSE-5) or combined in cocktails of two or three phages (phSE-1/phSE-2, phSE-1/phSE-5, phSE-2/phSE-5 and phSE-1/phSE-2/phSE-5) to control Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) in order to evaluate their potential application during depuration. Phages were assigned to the family Siphoviridae and revealed identical restriction digest profiles, although they showed a different phage adsorption, host range, burst size, explosion time and survival in seawater. The three phages were effective against S. Typhimurium (reduction of ∼2.0 log CFU/mL after 4h treatment). The use of cocktails was not significantly more effective than the use of single phages. A big fraction of the remained bacteria are phage-resistant mutants (frequency of phage-resistant mutants 9.19×10(-5)-5.11×10(-4)) but phage- resistant bacterial mutants was lower for the cocktail phages than for the single phage suspensions and the phage phSE-1 presented the highest rate of resistance and phage phSE-5 the lowest one. The spectral changes of S. Typhimurium resistant and phage-sensitive cells were compared and revealed relevant differences for peaks associated to amide I (1620cm(-1)) and amide II (1515cm(-1)) from proteins and from carbohydrates and phosphates region (1080-1000cm(-1)). Despite the similar efficiency of individual phages, the development of lower resistance indicates that phage cocktails might be the most promising choice to be used during the bivalve depuration to control the transmission of salmonellosis., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Wolbachia pipientis are obligate intracellular bacteria commonly found in many arthropods. They can induce various reproductive alterations in hosts, including cytoplasmic incompatibility, male-killing, feminization, and parthenogenetic development, and can provide host protection against some viruses and other pathogens. Wolbachia differ from many other primary endosymbionts in arthropods because they undergo frequent horizontal transmission between hosts and are well known for an abundance of mobile elements and relatively high recombination rates. Here, we compare the genomes of two closely related Wolbachia (with 0.57% genome-wide synonymous divergence) that differ in their reproductive effects on hosts. wVitA induces a sperm-egg incompatibility (also known as cytoplasmic incompatibility) in the parasitoid insect Nasonia vitripennis, whereas wUni causes parthenogenetic development in a different parasitoid, Muscidifurax uniraptor Although these bacteria are closely related, the genomic comparison reveals rampant rearrangements, protein truncations (particularly in proteins predicted to be secreted), and elevated substitution rates. These changes occur predominantly in the wUni lineage, and may be due in part to adaptations by wUni to a new host environment, or its phenotypic shift to parthenogenesis induction. However, we conclude that the approximately 8-fold elevated synonymous substitution rate in wUni is due to a either an elevated mutation rate or a greater number of generations per year in wUni, which occurs in semitropical host species. We identify a set of genes whose loss or pseudogenization in the wUni lineage implicates them in the phenotypic shift from cytoplasmic incompatibility to parthenogenesis induction. Finally, comparison of these closely related strains allows us to determine the fine-scale mutation patterns in Wolbachia Although Wolbachia are AT rich, mutation probabilities estimated from 4-fold degenerate sites are not AT biased, and predict an equilibrium AT content much less biased than observed (57-50% AT predicted vs. 76% current content at degenerate sites genome wide). The contrast suggests selection for increased AT content within Wolbachia genomes., (© The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Avian influenza viruses (AIVs) are known to cross species barriers, and emergent highly pathogenic H5N6 AIVs pose a serious threat to human health and the poultry industry. Here, we serially passaged an H5N6 virus 10 times in BALB/c mice. The pathogenicity of the wild-type 6D2 (WT-6D2) and mammal-adapted 6D2 strain (MA-6D2) were compared. The viral titer in multiple organs and the death rate for MA-6D2 were significantly higher than for WT-6D2. We provide evidence that the mutations HA A150V, NA R143K and G147E, PB2 E627K, and PA A343T may be important for adaptation of H5N6 AIVs to mammals.