Novel Papillomaviruses in Free-Ranging Iberian Bats: No Virus-Host Co-evolution, No Strict Host Specificity, and Hints for Recombination

Papillomaviruses (PVs) are widespread pathogens. However, the extent of PV infections in bats remains largely unknown. This work represents the first comprehensive study of PVs in Iberian bats. We identified four novel PVs in the mucosa of free-ranging Eptesicus serotinus (EserPV1, EserPV2, and EserPV3) and Rhinolophus ferrumequinum (RferPV1) individuals and analyzed their phylogenetic relationships within the viral family. We further assessed their prevalence in different populations of E. serotinus and its close relative E. isabellinus. Although it is frequent to read that PVs co-evolve with their host, that PVs are highly species-specific, and that PVs do not usually recombine, our results suggest otherwise. First, strict virus-host co-evolution is rejected by the existence of five, distantly related bat PV lineages and by the lack of congruence between bats and bat PVs phylogenies. Second, the ability of EserPV2 and EserPV3 to infect two different bat species (E. serotinus and E. isabellinus) argues against strict host specificity. Finally, the description of a second noncoding region in the RferPV1 genome reinforces the view of an increased susceptibility to recombination in the E2-L2 genomic region. These findings prompt the question of whether the prevailing paradigms regarding PVs evolution should be reconsidered.

Novel Papillomaviruses in Free-Ranging Iberian Bats: No Virus-Host Co-evolution, No Strict Host Specificity, and Hints for Recombination

Papillomaviruses (PVs) are widespread pathogens. However, the extent of PV infections in bats remains largely unknown. This work represents the first comprehensive study of PVs in Iberian bats. We identified four novel PVs in the mucosa of free-ranging Eptesicus serotinus (EserPV1, EserPV2, and EserPV3) and Rhinolophus ferrumequinum (RferPV1) individuals and analyzed their phylogenetic relationships within the viral family. We further assessed their prevalence in different populations of E. serotinus and its close relative E. isabellinus. Although it is frequent to read that PVs co-evolve with their host, that PVs are highly species-specific, and that PVs do not usually recombine, our results suggest otherwise. First, strict virus-host co-evolution is rejected by the existence of five, distantly related bat PV lineages and by the lack of congruence between bats and bat PVs phylogenies. Second, the ability of EserPV2 and EserPV3 to infect two different bat species (E. serotinus and E. isabellinus) argues against strict host specificity. Finally, the description of a second noncoding region in the RferPV1 genome reinforces the view of an increased susceptibility to recombination in the E2-L2 genomic region. These findings prompt the question of whether the prevailing paradigms regarding PVs evolution should be reconsidered.

Development, validation, and utilization of a competitive enzyme-linked immunosorbent assay for the detection of antibodies against Brucella species in marine mammals

A competitive enzyme-linked immunosorbent assay (cELISA) was developed by using a whole-cell antigen from a marine Brucella sp. isolated from a harbor seal (Phoca vitulina). The assay was designed to screen sera from multiple marine mammal species for the presence of antibodies against marine-origin Brucella. Based on comparisons with culture-confirmed cases, specificity and sensitivity for cetacean samples tested were 73% and 100%, respectively. For pinniped samples, specificity and sensitivity values were 77% and 67%, respectively. Hawaiian monk seal (Monachus schauinslandi; n = 28) and bottlenose dolphin (Tursiops truncatus; n = 48) serum samples were tested, and the results were compared with several other assays designed to detect Brucella abortus antibodies. The comparison testing revealed the marine-origin cELISA to be more sensitive than the B. abortus tests by the detection of additional positive serum samples. The newly developed cELISA is an effective serologic method for detection of the presence of antibodies against marine-origin Brucella sp. in marine mammals.

Response Specificity to Advertisement Vocalization in the Chinese Alligator (Alligator sinensis)

Stringency in the identification of conspecific call properties is essential among sympatric species to ensure conspecific mating, as the risk of improper recognition and heterospecific mating is high. In this study we investigated the basic signal structure required for intraspecies communication in the Chinese alligator (Alligator sinensis), a species that has no relatives living in sympatry, by playback of signals modified in the temporal (truncating original bellows with first 1/4, 1/2, 3/4 or last 1/4, 1/2, 3/4 portion) or frequency domain (with low- or high-pass filters at frequencies 100, 250, 500 and 1000 Hz), or by reversal of natural bellows. The playback experiments revealed that relatively large modifications in bellow temporal or frequency structure failed to impair Chinese alligators' bellowing behavior; even reversed bellows effectively evoked a positive response. In general, the first half of the bellow in temporal domain and frequencies below 500 Hz were critical for behavioral induction, while the last half of the bellow in temporal domain and frequencies above 500 Hz failed to produce a single positive response, implying a potential functional signal redundancy. The observed high tolerance to bellow variations in Chinese alligators may be an evolutionary adaptation to (1) the acoustic constraints of propagation imposed by dense vegetative habitats; or (2) a lack of selection pressure due to the low risk of incorrect conspecific recognition and heterospecific mating.

Morphological and genetic differences of Trypanosoma in some Chinese freshwater fishes: difficulties of species identification

Blood smears and purified trypanosome from freshwater fishes yellow catfish (Pseudobagras fulvidraco) and common carp (Cyprinus carpio) captured from Niushan Lake, Hubei Province were examined to determine whether all of their trypanosomes were Trypanosoma pseudobagri, a species of supposed host specificity and widespread existence across China. Trypanosomes occurred in 16/16 blood smears, and morphometric character analysis of trypanosomes from these smears showed that there were three morphospecies, Trypanosoma sp Carpio, T. sp Pseudobagri, and T. sp. 18S rDNA sequences of trypanosomes from 16 samples revealed three genetic groups among these fish trypanosomes. Group 1 was from C. carpio containing T. sp Carpio; groups 2 and 3 were from P. fulvidraco containing T. sp Pseudobagri and T. sp, respectively. The high similarity of morphometric characters and 18S rDNA sequences showed that T. sp Carpio and T. siniperca probably were the same species. T. sp Pseudobagri was the first occurrence in China. Sequence comparison showed that T. sp Pseudobagri sequence was most similar to that of clone Marv, whereas T. sp sequence differ from those of T. sp Carpio and T. sp Pseudobagri by 5.4 and 5.8%, respectively, and tentatively identified as T. pseudobagri. It was concluded that three species of trypanosomes, at least three genotypes occur in Niushan Lake fishes, and P. fulvidraco in this region appear to contain both types, although the identification of T. pseudobagri remains a problem.

Evidence of host specificity and congruence between phylogenies of bitterling and freshwater mussels

Evidence of host specificity and congruence between phylogenies of bitterling and freshwater mussels. Zoological Studies 45(3): 428-434. Bitterling (Cyprinidae: Acheilognathinae) are freshwater fishes with a unique spawning relationship with freshwater mussels on whose gills they lay their eggs. During the breeding season of bitterling fishes, we collected 843 mussels belonging to 16 species from Lake Qinglan, central China and examined their gill chambers for the presence of bitterling larvae. Three species of bitterling larvae were identified; Acheilognathus tonkinensis, Ach. cf. meridianus, and Ach. barbatulus, in 3 species of mussel: Unio douglasiae, Lamprotula caveata, and L. tortuosa, suggesting host specialization. Using our own and other published data, we compared the respective phylogenies of bitterling and mussels, but failed to show clear congruence. However, broad specializations are evident, with Acheilognathus and Tanakia showing preferences for mussels with a relatively simple gill structure (Ableminae), and Rhodeus spp. showing preferences for mussels of the Anodontinae and Unioninae, which have more-complex gill structures.

Scanning electron microscopy of Aspidogaster ijimai Kawamura, 1913 and A-conchicola Baer, 1827 (Aspidogastrea, Aspidogastridae) with reference to their fish definitive-host specificity

Two species of aspidogastreans, namely Aspidogaster ijimai and A. conchicola, were studied by scanning electron microscopy. In nine lakes and an old river course, the Tian'ezhou oxbow, investigated in the flood plain of the Yangtze River, A. ijimai was obtained from the common carp (Cyprinus carpio) in three lakes, and A. conchicola from the black carp Mylopharyngodon piceus in three lakes and the oxbow. In none of the localities, however, were the two species found together. It is suggested that A. ijimai may be considered as a specialist parasite for the common carp, at least in the flood-plain lakes of the Yangtze River. The two parasites were similar in many aspects of their morphology. Their bodies can both be separated into a dorsal part and a ventral disc, with the body surface of the dorsal part elevated by transverse folds, and the disc subdivided into alveoli by transverse and longitudinal septa, although the number of alveoli was different in the two species. The depression on the ventral surface of the neck region was prominent for both species, and their ventral disc was covered densely with non-ciliated bulbous papillae. The position of mouth, osmo-regulatory pore and marginal organ was also similar for A. ijimai and A. conchicola. However, microridges in the trough of the folds in the neck region and numerous small pits on the upper part of the septa were found exclusively in A. ijimai, but uniciliated sensory papillae in A. conchicola.

Molecular variation of Bothriocephalus acheilognathi Yamaguti, 1934 (Cestoda : Pseudophyllidea) in different fish host species based on ITS rDNA sequences

The molecular variation in Bothriocephalus acheilognathi Yamaguti, 1934 from 11 species of freshwater fish collected in Australia, China, the Czech Republic, England and Hawaii was investigated by determining the nucleotide sequences of the internal transcribed spacer region. The length of the first and second internal transcribed spacer sequences of multiple individuals ranged from 553 to 571 bp and 553 to 615 bp, and the G + C content from 53.1 to 53.5%. The percentage sequence divergence varied between 0 and 0.9% in the ITS1 and 0 and 6.6% in the ITS2, respectively, indicating the occurrence of intraspecific variation. It is demonstrated that the fragment length variation resulted primarily from microsatellite polymorphisms present in the ITS region, especially in the ITS2 region. Phylogenetic analyses revealed that B. acheilognathi examined in this study consisted of three closely related genotypes with certain degrees of host-specificity, and the genotype representing isolates from Cyprinus carpio L. was the most common and diverse form within the species B. acheilognathi.

A prochelator activated by beta-secretase inhibits Abeta aggregation and suppresses copper-induced reactive oxygen species formation.

The intersection of the amyloid cascade hypothesis and the implication of metal ions in Alzheimer's disease progression has sparked an interest in using metal-binding compounds as potential therapeutic agents. In the present work, we describe a prochelator SWH that is enzymatically activated by beta-secretase to produce a high affinity copper chelator CP. Because beta-secretase is responsible for the amyloidogenic processing of the amyloid precursor protein, this prochelator strategy imparts disease specificity toward copper chelation not possible with general metal chelators. Furthermore, once activated, CP efficiently sequesters copper from amyloid-beta, prevents and disassembles copper-induced amyloid-beta aggregation, and diminishes copper-promoted reactive oxygen species formation.

Assessing the sensitivity and specificity of fish community indicators to management action

We assessed ten trophodynamic indicators of ecosystem status for their sensitivity and specificity to fishing management using a size-resolved multispecies fish community model. The responses of indicators to fishing depended on effort and the size selectivity (sigmoid or Gaussian) of fishing mortality. The highest specificity against sigmoid (trawl-like) size selection was seen from inverse fishing pressure and the large fish indicator, but for Gaussian size selection, the large species indicator was most specific. Biomass, mean trophic level of the community and of the catch, and fishing in balance had the lowest specificity against both size selectivities. Length-based indicators weighted by biomass, rather than abundance, were more sensitive and specific to fishing pressure. Most indicators showed a greater response to sigmoid than Gaussian size selection. Indicators were generally more sensitive at low levels of effort because of nonlinear sensitivity in trophic cascades to fishing mortality. No single indicator emerged as superior in all respects, so given available data, multiple complementary indicators are recommended for community monitoring in the ecosystem approach to fisheries management.

Phage Display-Derived Binders Able to Distinguish Listeria monocytogenes from Other Listeria Species

The objective of this study was to produce phage display-derived binders with the ability to distinguish Listeria monocytogenes from other Listeria spp., which may have potential utility to enhance detection of Listeria monocytogenes. To obtain binders with the desired binding specificity a series of surface and solution phage-display biopannings were performed. Initially, three rounds of surface biopanning against gamma-irradiated L. monocytogenes serovar 4b cells were performed followed by an additional surface biopanning round against L. monocytogenes 4b which included prior subtraction biopanning against gamma-irradiated L. innocua cells. In an attempt to further enhance binder specificity for L. monocytogenes 4b two rounds of solution biopanning were performed, both rounds included initial subtraction solution biopanning against L. innocua. Subsequent evaluations were performed on the phage clones by phage binding ELISA. All phage clones tested from the second round of solution biopanning had higher specificity for L. monocytogenes 4b than for L. innocua and three other foodborne pathogens (Salmonella spp., Escherichia coli and Campylobacter jejuni). Further evaluation with five other Listeria spp. revealed that one phage clone in particular, expressing peptide GRIADLPPLKPN, was highly specific for L. monocytogenes with at least 43-fold more binding capability to L. monocytogenes 4b than to any other Listeria sp. This proof-of-principle study demonstrates how a combination of surface, solution and subtractive biopanning was used to maximise binder specificity. L. monocytogenes-specific binders were obtained which could have potential application in novel detection tests for L. monocytogenes, benefiting both the food and medical industries. © 2013 Morton et al.

The specificity of TRIM5 alpha-mediated restriction is influenced by its coiled-coil domain.

Retroviruses are both powerful evolutionary forces and dangerous threats to genome integrity. As such, they have imposed strong selective pressure on their hosts, notably triggering the emergence of restriction factors, such as TRIM5 alpha, that act as potent barriers to their cross-species transmission. TRIM5 alpha orthologues from different primates have distinct retroviral restriction patterns, largely dictated by the sequence of their C-terminal PRYSPRY domain, which binds the capsid protein of incoming virions. Here, by combining genetic and functional analyses of human and squirrel monkey TRIM5 alpha, we demonstrate that the coiled-coil domain of this protein, thus far essentially known for mediating oligomerization, also conditions the spectrum of antiretroviral activity. Furthermore, we identify three coiled-coil residues responsible for this effect, one of which has been under positive selection during primate evolution, notably in New World monkeys. These results indicate that the PRYSPRY and coiled-coil domains cooperate to determine the specificity of TRIM5 alpha-mediated capture of retroviral capsids, shedding new light on this complex event.

Molecular detection and identification of zygomycetes species from paraffin-embedded tissues in a murine model of disseminated zygomycosis: a collaborative European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Fungal Infection Study Group (EFISG) evaluation.

The present study was performed to assess the interlaboratory reproducibility of the molecular detection and identification of species of Zygomycetes from formalin-fixed paraffin-embedded kidney and brain tissues obtained from experimentally infected mice. Animals were infected with one of five species (Rhizopus oryzae, Rhizopus microsporus, Lichtheimia corymbifera, Rhizomucor pusillus, and Mucor circinelloides). Samples with 1, 10, or 30 slide cuts of the tissues were prepared from each paraffin block, the sample identities were blinded for analysis, and the samples were mailed to each of seven laboratories for the assessment of sensitivity. A protocol describing the extraction method and the PCR amplification procedure was provided. The internal transcribed spacer 1 (ITS1) region was amplified by PCR with the fungal universal primers ITS1 and ITS2 and sequenced. As negative results were obtained for 93% of the tissue specimens infected by M. circinelloides, the data for this species were excluded from the analysis. Positive PCR results were obtained for 93% (52/56), 89% (50/56), and 27% (15/56) of the samples with 30, 10, and 1 slide cuts, respectively. There were minor differences, depending on the organ tissue, fungal species, and laboratory. Correct species identification was possible for 100% (30 cuts), 98% (10 cuts), and 93% (1 cut) of the cases. With the protocol used in the present study, the interlaboratory reproducibility of ITS sequencing for the identification of major Zygomycetes species from formalin-fixed paraffin-embedded tissues can reach 100%, when enough material is available.

Plant rhizosphere specificity and variability in the insect and plant adhesins, Madl and Mad2, within the genus Metarhizium suggest plant adaptation as an evolutionary force

Metarhizium is a soil-inhabiting fungus currently used as a biological control agent against various insect species, and research efforts are typically focused on its ability to kill insects. In section 1, we tested the hypothesis that species of Metarhizium are not randomly distributed in soils but show plant rhizosphere-specific associations. Results indicated an association of three Metarhizium species (Metarhizium robertsii, M. brunneum and M. guizhouense) with the rhizosphere of certain types of plant species. M. robertsii was the only species that was found associated with grass roots, suggesting a possible exclusion of M. brunneum and M. guizhouense, which was supported by in vitro experiments with grass root exudate. M. guizhouense and M. brunneum only associated with wildflower rhizosphere when co-occurring with M. robertsii. With the exception of these co-occurrences, M. guizhouense was found to associate exclusively with the rhizosphere of tree species, while M. brunneum was found to associate exclusively with the rhizosphere of shrubs and trees. These associations demonstrate that different species of Metarhizium associate with specific plant types. In section 2, we explored the variation in the insect adhesin, Madl, and the plant adhesin, Mad2, in fourteen isolates of Metarhizium representing seven different species. Analysis of the transcriptional elements within the Mad2 promoter region revealed variable STRE, PDS, degenerative TATA box, and TATA box-like regions. Phylogenetic analysis of 5' EF-Ia, which is used for species identification, as well as Madl and Mad2 sequences demonstrated that the Mad2 phylogeny is more congruent with 5' EF-1a than Madl. This suggests Mad2 has diverged among Metarhizium lineages, contributing to clade- and species-specific variation. While other abiotic and biotic factors cannot be excluded in contributing to divergence, it appears that plant associations have been the driving factor causing divergence among Metarhizium species.

Pathogens occuring in the winter pea-maize-winter wheat rotation, their host specificity and the potential of compost in suppressing foot and root disease of peas

The overall aim of the work presented was to evaluate soil health management with a specific focus on soil borne diseases of peas. For that purpose field experiments were carried out from 2009 until 2013 to assess crop performance and pathogen occurrence in the rotation winter pea-maize-winter wheat and if the application of composts can improve system performance. The winter peas were left untreated or inoculated with Phoma medicaginis, in the presence or absence of yard waste compost at rate of 5 t dry matter ha-1. A second application of compost was made to the winter wheat. Fusarium ssp. were isolated and identified from the roots of all three crops and the Ascochyta complex pathogens on peas. Bioassays were conducted under controlled conditions to assess susceptibility of two peas to Fusarium avenaceum, F. solani, P. medicaginis and Didymella pinodes and of nine plant species to F. avenaceum. Also, effects of compost applications and temperature on pea diseases were assessed. Application of composts overall stabilized crop performance but it did not lead to significant yield increases nor did it affect pathogen composition and occurrence. Phoma medicaginis was dominating the pathogen complex on peas. F. graminearum, F. culmorum, F. proliferatum, Microdochium nivale, F. crookwellense, F. sambucinum, F. oxysporum, F. avenaceum and F. equiseti were frequently isolated species from maize and winter wheat with no obvious influence of the pre-crop on the Fusarium species composition. The spring pea Santana was considerably more susceptible to the pathogens tested than the winter pea EFB33 in both sterile sand and non-sterilized field soil. F. avenaceum was the most aggressive pathogen, followed by P. medicaginis, D. pinodes, and F. solani. Aggressiveness of all pathogens was greatly reduced in non-sterile field soil. F. avenaceum caused severe symptoms on roots of all nine plant species tested. Especially susceptible were Trifolium repens, T. subterraneum, Brassica juncea and Sinapis alba in addition to peas. Reduction of growing temperatures from 19/16°C day/night to 16/12°C and 13/10°C did not affect the efficacy of compost. It reduced plant growth and slightly increased disease on EFB33 whereas the highest disease severity on Santana was observed at the highest temperature, 19/16°C. Application of 20% v/v of compost reduced disease on peas due to all four pathogens depending on pea variety, pathogen and growing media used. Suppression was also achieved with lower application rate of 3.5% v/v. Tests with γ sterilized compost suggest that the suppression of disease caused by Fusarium spp. is biological in origin, whereas chemical and physical properties of compost are playing an additional role in the suppression of disease caused by D. pinodes and P. medicaginis.