High Host Specificity Research Articles

Ecology of bat flies in Singapore: A study on the diversity, infestation bias and host specificity (Diptera: Nycteribiidae).

Bat flies are highly-specialized, hematophagous arthropods that are globally ubiquitous. There is little published research on bat flies (Diptera: Nycteribiidae) in Singapore and understanding the diversity of nycteribiids, host association and infestation rates can provide insight into this host-ectoparasite relationship. Nycteribiids were collected from bats trapped in Singapore (2011–2016) and identified using morphological keys. Host-ectoparasite relationships were investigated with logistic regression and Bayesian poisson regression. Nycteribiids were found to be monoxenously associated with their host bat species and host age, sex, species, and BBCI appear to contribute to differences in prevalence and intensity. Differences in host specificity between bat fly species in Singapore and their conspecifics in less disturbed habitats with higher bat biodiversity, such as Malaysia, Philippines and Thailand, suggest that the high host specificity in Singapore derives from the paucity of suitable hosts and abundance of single species roosts and not from their coevolved restrictions to them.

D-mannose-specific Immunoglobulin M in Grass Puffer (Takifugu niphobles), a Nonhost Fish of a Monogenean Ectoparasite Heterobothrium okamotoi, Can Act as a Trigger for its Parasitism.

Heterobothrium okamotoi, a monogenean gill parasite, exhibits high host specificity for the tiger puffer, Takifugu rubripes, and it has been experimentally verified that the parasite cannot colonize either closely related species such as the grass puffer Takifugu niphobles or distantly related fish such as the red seabream Pagrus major. Previously, we demonstrated in T. rubripes that immunoglobulin M (IgM) with d-mannose affinity induced deciliation of the oncomiracidia, the first step of parasitism, indicating that the parasite utilizes the molecule as a receptor for infection. In the present study, we purified mannose-specific IgM from 2 nonhost species, T. niphobles and P. major, by affinity and gel-filtration chromatography techniques and compared their deciliation-inducing activity against H. okamotoi oncomiracidia. The IgM of the former showed activity, whereas the latter had no effect, suggesting that in addition to d-mannose-binding ability, the crystallizable fragment domain of IgM, which is not part of the antigen-binding domain, plays an important role in host recognition by the oncomiracidia, such as direct binding to the parasites. It also suggests that the host specificity of H. okamotoi is relatively low upon initial recognition, and the specificity is established by exclusion in nonhosts during a later stage.

Identification, pathogenicity and histopathology of Colletotrichum sansevieriae causing anthracnose of Sansevieria trifasciata in Malaysia.

To characterize causal pathogen of Sansevieria trifasciata anthracnose through morphology and molecular analysis; to evaluate the host range of the pathogen; and to explicate the infection process by the pathogen histopathologically. Symptomatic leaves of S. trifasciata were collected from five states in Malaysia. The causal pathogen was isolated and identified for the first time in Malaysia as C. sansevieriae based on morphological and multi-gene phylogenetic analyses using ITS, TUB2 and GAPDH sequences. Pathogenicity tests were conducted on different hosts. Colletotrichum sansevieriae was not pathogenic towards S. cylindrica, S. masoniana, Furcraea foetida, Chlorophytum comosum, Aloe vera and Gasteria carinata, confirming the exceptionally high host specificity for a species of Colletotrichum. Histopathology was performed using light microscope and scanning electron microscopy to study the infection process of C. sansevieriae on S. trifasciata. Colonization of host leaves by the pathogen was observed 2days after inoculation. Colletotrichum sansevieriae caused anthracnose of S. trifasciata in Malaysia. It is a host-specific pathogen and colonized the host intracellularly. This is the first report of C. sansevieriae causing anthracnose of S. trifasciata in Malaysia. The host range test and understanding of the infection process will provide better understanding of the host-pathogen relationship and beneficial for effective disease management.

Native parasite affecting an introduced host in aquaculture: cardiac henneguyosis in the red seabream Pagrus major Temminck & Schlegel (Perciformes: Sparidae) caused by Henneguya aegea n. sp. (Myxosporea: Myxobolidae)

BackgroundHenneguya Thélohan, 1892 (Myxobolidae) is one of the most species-rich genera of myxosporean parasites infecting fish. Although common in nature, there are few reports of these parasites causing important disease in aquaculture. In this paper, we describe a new species of Henneguya infecting Pagrus major (Temminck & Schlegel), a fish host introduced to the Mediterranean Sea from Japan in the late 1980s.ResultsLarge plasmodia of the parasite were found in the bulbus arteriosus and in the ventricle of the infected fish. Spores were found mainly in the kidney and heart and were accompanied by melanized macrophages or vascular intimal proliferation mixed with a mild non-suppurative response, respectively. Comparisons of morphometric data for spore and polar capsule length and width, suggest a unique combination of features in the newly described species. Molecular analysis, based on 18S rDNA sequence of the parasite, followed by phylogenetic analysis, indicated that the parasite described here is a novel species of Henneguya, clustered with the marine congeneric species.ConclusionsHenneguya aegea n. sp. infects in aquaculture P. major, a host introduced as eggs to the Mediterranean from Japan. Despite the high host specificity of the myxobolid parasites, H. aegea n. sp. seems to be able to use P. major as a host and propagate successfully, causing morbidity and mortality. This could result in spillback of the new species from high density cultured non-native P. major to native fish hosts.

Isolation of Four Lytic Phages Infecting Klebsiella pneumoniae K22 Clinical Isolates from Spain.

The emergence of multi-drug-resistant bacteria represents a major public-health threat. Phages constitute a promising alternative to chemical antibiotics due to their high host specificity, abundance in nature, and evolvability. However, phage host specificity means that highly diverse bacterial species are particularly difficult to target for phage therapy. This is the case of Klebsiella pneumoniae, which presents a hypervariable extracellular matrix capsule exhibiting dozens of variants. Here, we report four novel phages infecting K. pneumoniae capsular type K22 which were isolated from environmental samples in Valencia, Spain. Full genome sequencing showed that these phages belong to the Podoviridae family and encode putative depolymerases that allow digestion of specific K22 K. pneumoniae capsules. Our results confirm the capsular type-specificity of K. pneumoniae phages, as indicated by their narrow infectivity in a panel of K. pneumoniae clinical isolates. Nonetheless, this work represents a step forward in the characterization of phage diversity, which may culminate in the future use of large panels of phages for typing and/or for combating multi-drug-resistant K. pneumoniae.

Genetic variability on worldwide populations of the scale insect Pulvinariella mesembryanthemi

The South African scale insect Pulvinariella mesembryanthemi was introduced worldwide in several coastal areas with Mediterranean climate, probably through infested plants of Carpobrotus sp. Its high host specificity and its capacity to produce severe damages in the invasive Carpobrotus sp. plants makes this insect a potential biocontrol agent. To test the efficiency and host range of insects used for biocontrol, population genetic studies can help to unravel cryptic complexes and intraspecific diversity. In this study we performed a genetic analysis including native and exotic populations of P. mesembryanthemi, through Sanger sequencing of mitochondrial (cytochrome c oxidase I, COI) and ribosomal (D2–D3 expansion segments of the large subunit ribosomal RNA gene 28S) gene fragments. Accidentally, an endosymbiont was sequenced with one of the pair of primers used. The exotic populations of the insect did not show any variability among populations for both studied genes, which suggest a common origin of all studied introduced populations. Contrastingly, native populations showed high variability and seemed to be a cryptic species complex. Moreover, the Gauteng populations (from NE South Africa) were phylogenetically the closest to the exotic ones, suggesting that the exotic populations could be original from somewhere near this area. An endosymbiont of P. mesembryanthemi was detected, and the sequenced coxA gene was similar to that of the Rickettsiaceae family from the α-Proteobacteria, and close to other insect endosymbionts. To the best of our knowledge, this was the first mention of this endosymbiont in P. mesembryanthemi, although α-Proteobacteria endosymbionts have been reported for other sap-sucking insects.

Diversity of Symbiodiniaceae in 15 Coral Species From the Southern South China Sea: Potential Relationship With Coral Thermal Adaptability.

It is well-known that the adaptability of coral-Symbiodiniaceae symbiosis to thermal stress varies among coral species, but the cause and/or mechanism behind it are not well-understood. In this study, we aimed to explore this issue based on zooxanthellae density (ZD) and Symbiodiniaceae genus/subclade. Hemocytometry and next-generation sequencing of the internal transcribed spacer region 2 (ITS2) marker gene were used to observe ZDs and Symbiodiniaceae genera/subclades associated with 15 typical coral species in the southern South China Sea (SCS). Average ZDs of all corals were in low levels, ranging from 0.84 to 1.22 × 106 cells cm−2, with a total of five Symbiodiniaceae genera, Symbiodinium, Cladocopium, Durusdinium, Fugacium, and Gerakladium, as well as 24 dominant subclades, were detected and varied among these coral species. Pocillopora verrucosa was dominated by Durusdinium (subclade D1/D1a), and other colonial corals were dominated by Cladocopium, but the subclades were varied among these species. Porites lutea and Montipora efflorescens were dominated by C15, and Echinopora lamellosa, Hydnophora exesa, and Coscinaraea exesa were dominated by C40. Acropora corymbosa, Merulina ampliata, and five species of Faviidae were mainly associated with Cladocopium types of C3u and Cspc. In contrast to other colonial corals, the dominant subclade of solitary Fungia fungites was C27, with high host specificity. Our study indicates that coral thermal stress adaptability is mainly affected by dominant Symbiodiniaceae type instead of ZD in the southern SCS. Some heat-sensitive corals, such as P. verrucosa corals, have acquired a high abundance of heat-tolerant Durusdinium to adapt to thermal stress. This could be the main reason for these corals becoming the dominant corals in this reef region. Background subclades analyses showed significant differences among coral species in subclade quantity and diversity. These suggest that numbers of coral species may have adapted to high environmental temperature by adopting various symbionts and/or associating with heat-tolerant Symbiodiniaceae.

Structural Basis for Cell-Wall Recognition by Bacteriophage PBC5 Endolysin

Phage endolysins are hydrolytic enzymes that cleave the bacterial cell wall during the lytic cycle. We isolated the bacteriophage PBC5 against Bacillus cereus, a major foodborne pathogen, and describe the molecular interaction between endolysin LysPBC5 and the host peptidoglycan structure. LysPBC5 has an N-terminal glycoside hydrolase 25 domain, and a C-terminal cell-wall binding domain (CBD) that is critical for specific cell-wall recognition and lysis. The crystal and solution structures of CBDs reveal tandem SH3b domains that are tightly engaged with each other. The CBD binds to the peptidoglycan in a bidentate manner via distal β sheet motifs with pseudo 2-fold symmetry, which can explain its high affinity and host specificity. The CBD primarily interacts with the glycan strand of the peptidoglycan layer instead of the peptide crosslink, implicating the tertiary structure of peptidoglycan as the recognition motif of endolysins.

Monogenoidea (Dactylogyridae, Gyrodactylidae) from Red River Pupfish, Cyprinodon rubrofluviatilis (Cyprinodontiformes: Cyprinodontidae), from Kansas and Texas, USA.

The gills of Red River Pupfish (Cyprinodon rubrofluviatilis) collected from Kansas and Texas, U.S.A., were found to be infected with three monogenoideans, Fundulotrema prolongis, Gyrodactylus rubrofluviatilaen. sp., and a species of Salsuginus. Gyrodactylus rubrofluviatilae n. sp. appears to be a member of a group of six closely related species possessing hamuli with mesially folded roots and a linguiform ventral bar shield, and parasitizing closely related species of Cyprinodon of the North American Gulf Coast and inland waters of the Southwestern United States and northern México. Gyrodactylus rubrofluviatilae n. sp. differs from its close congeners in the morphology of the hamuli, ventral bar, ventral bar shield, and marginal hooks. Because the morphology of the male copulatory complex was not determined of what we believe will eventually be a new species of Salsuginus, a species description is deferred. This is the first report of any parasite from this host. ManyNorth American species currently recognized within the Cyprinodontiformes have not yet been surveyed for species of Fundulotrema, Gyrodactylus, or Salsuginus. This, coupled with the high host specificity generally recognized for these monogeneans, portends there are likely additional new species yet to be discovered.When possible, to help augment morphological data, further studies should employ comprehensiveintra- and interspecies molecular analyses to help resolve host-parasite phylogenies.

Abstract 4772: Evaluation of immune response following ultrasound targeted gene therapy in a murine model of prostate cancer

Abstract Human Adenoviruses (hAd) have been broadly used as gene delivery tool in preclinical and clinical studies of cancer gene therapy. The main challenges associated with systemic delivery of hAds are their high immunogenicity and host-specificity. Adenoviruses can elicit strong innate and acquired immune responses that reduce therapeutic gene transfer and expression in malignant sites. Murine cells generally lack some of the receptors necessary for hAd adhesion and internalization making them not permissive to infection and replication and limiting translational studies using murine models. We have developed a gene transfer method, which uses lipid-encapsulated perfluorocarbon microbubbles (MBs) and ultrasound (US) to protect the hAds from the immune system and to deliver them to a targeted tissue, bypassing the necessity for specific receptors. The expression of the Ad-receptors (CAR, Integrins αvβ3 and αvβ5), transduction efficiency of Ad-GFP, and GFP transgene expression were measured in vitro in murine (TRAMP-C2) and human (DU145) prostate cancer cells lines by flow cytometry and immunofluorescence. The activation of the innate and acquired immune system following the injection of Ad-GFP/MBs complexes was evaluated in vivo in C57BL/6 mice healthy or bearing a TRAMP-C2 syngeneic tumor graft. ELISA was used to quantify the inflammatory cytokines TNF-α, IL-6 and measure Neutralizing Antibodies. ELISpot assay was performed to measure the frequency of DBP-specific CD8+ T cells secreting INF-γ. We showed in vitro that murine TRAMP-C2 cells display an expression pattern of Ad-receptors comparable to human DU145 prostate cancer cells. We also demonstrated that both murine and human cells showed a dose dependent increase in the percentage of cells transduced by Ad-GFP at 24 hours. Additionally, we showed that TRAMP-C2 cells efficiently express the GFP transgene at 48 and 72 hours post transduction. To assess in vivo if our gene transfer method could effectively protect the Ads form both the innate and adaptive immunity, we injected C57BL/6 mice with the hAds-GFP/MBs complexes +/-US. Notably we did not observe activation of either innate (secretion of TNF- and IL-6 cytokines) or acquired immunity (neutralizing antibodies and presence of adenovirus-specific CD8+ T cells producing INF-γ). Our data provides evidence that the TRAMP-C2 syngeneic model of prostate cancer is a suitable system to study in immunocompetent mice the capability of the MBs/US to protect the Adenoviruses from the immune system while delivering them to a targeted site bypassing the requirement of specific receptors. Citation Format: Flavia De Carlo, Elliot T. Varney, Pier Paolo Claudio, Candace M. Howard. Evaluation of immune response following ultrasound targeted gene therapy in a murine model of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4772.

Biological and molecular characterization of Rachiplusia nu single nucleopolyhedrovirus, a promising biocontrol agent against the South American soybean pest Rachiplusia nu

Rachiplusia nu (Lepidoptera: Noctuidae) is a key soybean pest in Argentina. Current management of this moth relies mainly on the use of synthetic insecticides and transgenic plants. In search of biological control-based alternatives, a baculovirus from R. nu (hereafter RanuNPV) was characterized and its insecticidal properties tested under laboratory conditions. RanuNPV occlusion bodies (OBs) were nearly tetrahedral, averaging 1.0 ± 0.2 µm in their longest edge and containing singly enveloped nucleocapsids. Histopathology of infected late-instar larvae revealed broad tissue tropism, where fat bodies and epidermis were the most affected organs. Phylogenetic analysis of concatenated polh, lef-8 and lef-9 partial sequences classified RanuNPV as a new species that clusters with other group II alphabaculoviruses infecting larvae of Plusiinae. Bioassays performed with R. nu neonates determined the median lethal dosage to be approximately 2.5 OBs/larva; most insects died within 4–5 days post inoculation showing typical baculovirus-induced liquefaction. No effects were observed in other lepidopteran species assayed, including Spodoptera frugiperda, Cydia pomonella and Diatraea saccharalis. High pathogenicity and host specificity make RanuNPV a good candidate for controlling R. nu.

Host range testing of Tamarixia dryi (Hymenoptera: Eulophidae) sourced from South Africa for classical biological control of Trioza erytreae (Hemiptera: Psyllidae) in Europe

The African citrus psyllid, Trioza erytreae, vectors citrus greening or huanglongbing (HLB) disease. The psyllid has been reported from mainland Europe, where it is rapidly spreading from the northwest to the southwest of the Iberian Peninsula. In order to reduce its spread and population levels, a classical biological control program with the parasitoid Tamarixia dryi is under development in Spain. We evaluated the host specificity of T. dryi using 11 non-target psyllid (NTP) species, including five species of the genus Trioza. The psyllids were selected based on phylogenetic and ecological criteria. Tamarixia dryi exhibited a high host specificity. Females did not parasitize any of the 11 NTPs tested, except for one nymph of a gall-forming Trioza species closely related to Trioza montanetana. Tamarixia dryi only laid one egg on a nymph when it was removed from the gall on Convolvulus canariensis and exposed directly to the parasitoid. However, the immature parasitoid died before emerging. We further confirmed that T. dryi did not parasitize a representative triozid species, Trioza laurisilvae, of the endemic Canarian fauna after long time exposure. Our results demonstrate that T. dryi is a highly specific parasitoid and its introduction, release and establishment in Europe within the classical biological control program of T. erytreae should not affect other psyllid species. Therefore, no significant environmental impact is expected.

Structure and composition of Nycteribiidae and Streblidae flies on bats along an environmental gradient in northeastern Brazil

Bats can be parasitized by several arthropod groups, including ectoparasitic flies. The high host specificity is a common phenomenon between flies and bats. In recent years, more efforts have been employed to understand how environmental variables can influence richness and parasitic load (PL). However, many gaps still need to be filled to better understand this issue. We analyzed the PL of flies on bats sampled in three environments with different rain volume and vegetation types to verify if PL is correlated with rainfall and if there are differences in the PL on bats within and between environments. Overall, there was no correlation between rainfall and PL in the same environment, nor a difference between the three environments. When tested separately, Seba’s short-tailed bats (Carollia perspicillata (Linnaeus, 1758)) had a difference in prevalence of flies between environments and flat-faced fruit-eating bats (Artibeus planirostris (Spix, 1823)) had a greater abundance of flies in the rainy season in a semiarid area. There was no difference in PL between male and female bats. Our results suggest that bat–fly interactions are driven by several factors, not only by the amount of rainfall or vegetation, and that different host species may respond differently with no obvious general pattern.

A paradise for parasites? Seven new haemogregarine species infecting lizards from the Canary Islands.

Oceanic islands are hotspots of biodiversity due to their high levels of endemism, with the Canary Islands being a notable example. A previous molecular study on the biogeography and host associations of haemogregarines (Apicomplexa: Adeleorina) infecting lizards from this archipelago detected seven parasite haplogroups. These haplogroups exhibited high host-specificity and geographical structure, suggesting that they might correspond to distinct biological identities. In this study, along with sequencing a longer fragment of the 18S rRNA, we further explore the distinctiveness of these parasites by analysing their morphology, effects on host erythrocytes and parasitaemia levels. These lines of evidence together with their genetics, host associations, frequency of occurrence and geographical distribution support them as different biological entities. As such, we describe seven new species: Karyolysus canariensis sp. nov., Karyolysus galloti sp. nov., Karyolysus stehlini sp. nov., Karyolysus gomerensis sp. nov., Karyolysus atlanticus sp. nov., Karyolysus tinerfensis sp. nov. and Karyolysus makariogeckonis sp. nov. These new taxa are further examples of endemic diversity in the Canarian archipelago. They also contribute to clarify the taxonomy within the Apicomplexa, a phylum estimated to have one of the lowest percentages of described species.

Lactarius guangdongensis sp. nov. (Russulaceae, Russulales), a species of Lactarius sect. Deliciosi growing with a vulnerable five-needle pine, Pinus kwangtungensis

Lactarius sect. Deliciosi is a group of ectomycorrhizal mushrooms showing high host specificity with trees of Pinaceae and more rarely Fagales. A new species of this section, L. guangdongensis, is described from South China. This new species has the smallest basidiocarps (pileus 1–3 cm) in the section. The pure orange basidiocarps with faint greenish discoloration are similar to those of Abies-associated L. abieticola and L. laeticolor and Pinus-associated L. vividus. The subdistant lamellae and absence of pleuromacrocystidia can be used as valid characters for this species. It grows with Chinese national protected plant Pinus kwangtungensis and might be one of the very few species that form symbiosis with five-needle pines. Phylogenetic analyses of the ITS region and glyceraldehyde-3-phosphate dehydrogenase gene (gpd) show that L. guangdongensis clearly diversified from its relatives and look-alikes and probably represents an early diverging lineage in the section. Molecular and morphological characters provided in this study will be useful for in situ and ex situ conservation of P. kwangtungensis in the future.

Bacteriophages applications in agriculture

The bacteriophages life cycle has two stages: a lytic stage where the phages reproduce inside the bacteria and lyse bacteria and a lysogenic stage where the phage is in a stationary stage where do not exist phage reproduction. The understanding of the life cycle of phages is fundamental to understand the advantages of phage offers as biological control applications and how engineered phages work. The bacteriophages are an alternative to fight against the antimicrobial or pesticides because phages offer advantages such as high host specificity, the ability of long term effect, are active against dividing or not dividing bacterial cells, effective elimination of biofilms and are capable vehicles for nucleic acids delivery. Phages have been isolated from water or soil samples in different parts of the world and for specific bacterial pathogens. In the following review, in the main topics in bacteriophages and its applications in agriculture: the bacteriophages life cycle, advantages of phages in biological control applications, the last isolated phages and described for different pathogens and the last advances in phage engineering applications for biological control.

A novel approach using microsporidia to estimate the flight route of the common cutworm, Spodoptera litura (Lepidoptera: Noctuidae)

Microsporidia are unique fungi that exist as obligate intracellular parasites. Approximately 40% of the known microsporidian strains infect various insects. Due to their high host specificity, microsporidia have potential use as powerful biological pesticides. Here, we analyzed microsporidian strains isolated from the common cutworm, Spodoptera litura Fabricius (Lepidoptera: Noctuidae) of Japan and Taiwan, which is considered a pest insect. Two experiments were performed: a comparison of spore size and a phylogenetic analysis using small subunit ribosomal RNA gene sequencing. We then estimated the flight routes of S. litura by using the phylogenetic relationships of the isolated microsporidian strains as a marker. An analysis of spore size indicated that the test strains comprised three groups: Pleistophora, Nosema or Vairimorpha, and Trachipleistophora or Vavraia. The results of the phylogenetic analysis suggested a classification into five genera, including Vavraia. The genus Vavraia was first detected from lepidopteran insects in Japan. We estimated the sources and flight routes of S. litura using phylogenetic data for the genera Nosema and Trachipleistophora. In this study, we used microsporidia as a novel marker to estimate sources and flight routes of S. litura from Ogasawara and Southern China to mainland Japan, demonstrating the usefulness of this approach.

Bacteriophages Against Pathogenic Vibrios in Delaware Bay Oysters (Crassostrea virginica) During a Period of High Levels of Pathogenic Vibrio parahaemolyticus

Eastern oysters (Crassostrea virginica) from three locations along the Delaware Bay were surveyed monthly from May to October 2017 for levels of total Vibrio parahaemolyticus, pathogenic strains of V. parahaemolyticus and Vibrio vulnificus, and for strain-specific bacteriophages against vibrios (vibriophages). The objectives were to determine (a) whether vibriophages against known strains or serotypes of clinical and environmental vibrios were detectable in oysters from the Delaware Bay and (b) whether vibriophage presence or absence corresponded with Vibrio abundances in oysters. Host cells for phage assays included pathogenic V. parahaemolyticus serotypes O3:K6, O1:KUT (untypable) and O1:K1, as well as clinical and environmental strains of V. vulnificus. Vibriophages against some, but not all, pathogenic V. parahaemolyticus serotypes were readily detected in Delaware Bay oysters. In July, abundances of total and pathogenic V. parahaemolyticus at one site spiked to levels exceeding regulatory guidelines. Phages against three V. parahaemolyticus host serotypes were detected in these same oysters, but also in oysters with low V. parahaemolyticus levels. Serotype-specific vibriophage presence or absence did not correspond with abundances of total or pathogenic V. parahaemolyticus. Vibriophages were not detected against three V. vulnificus host strains, even though V. vulnificus were readily detectable in oyster tissues. Selected phage isolates against V. parahaemolyticus showed high host specificity. Transmission electron micrographs revealed that most isolates were ~ 60-nm diameter, non-tailed phages. In conclusion, vibriophages were detected against pandemic V. parahaemolyticus O3:K6 and O1:KUT, suggesting that phage monitoring in specific host cells may be a useful technique to assess public health risks from oyster consumption.

Structural Basis for Cell Wall Recognition by Bacteriophage PBC5 Endolysin

Phage endolysins are hydrolytic enzymes that cleave the bacterial cell wall during the lytic cycle. We isolated the bacteriophage PBC5 against Bacillus cereus, a major foodborne pathogen, and describe the molecular interaction between endolysin LysPBC5 and the host peptidoglycan structure. LysPBC5 has an N-terminal glycoside hydrolase 25 domain, and a C-terminal cell-wall binding domain (CBD) that is critical for specific cell-wall recognition and lysis. The crystal and solution structure of CBD reveals tandem SH3b domains that are tightly engaged with each other. CBD binds to peptidoglycan in a bidentate manner via distal β-sheet motifs with pseudo two-fold symmetry, which can explain its high affinity and host specificity. CBD primarily interacts with the glycan strand of the peptidoglycan layer instead of the peptide crosslink, implicating the tertiary structure of peptidoglycan as the recognition motif of endolysins.

Identification and genomic sequence analysis of a new Spodoptera exigua multiple nucleopolyhedrovirus, SeMNPV-QD, isolated from Qingdao, China.

A new beet armyworm (Spodoptera exigua) multiple nucleopolyhedrovirus SeMNPV-QD was isolated from dead larvae in the field in Qingdao, Shandong, China. The virus has a polyhedron size of 1.39 ± 0.28 μm, and typical virions contain one to seven nucleocapsids per envelope. SeMNPV-QD only infects the larvae of S. exigua; it does not infect larvae of S. litura, Agrotis ipsilon, A. segetum, Bombyx mori, Hyphantria cunea, or Stilpnotia salicis, and thus, has higher host specificity. SeMNPV-QD has a circular double-stranded DNA genome of 128,525 bp with a GC content of 37.41% and 127 putative open reading frames (ORF), each of which encodes more than 50 amino acids. These were identified and annotated in the SeMNPV-QD genome, accounting for 87.53% of the whole genome. Phylogenetic analysis of 38 core genes of the baculovirus confirmed that SeMNPV-QD is a Group II Alphabaculovirus and is most closely related to SeMNPV American isolate (SeMNPV-US1), S. litura nucleopolyhedrovirus II (SpliNPV-II), S. frugiperda multiple nucleopolyhedrovirus (SfMNPV), A. segetum nucleopolyhedrovirus (AgseNPV), A. segetum nucleopolyhedrovirus B (AgseNPV-B) and A. ipsilon multiple nucleopolyhedrovirus (AgipNPV). The pairwise distance of the nucleotide sequences of lef-8, lef-9, polh and concatenated lef-8/lef-9/polh fragments between SeMNPV-QD and several sister viruses mentioned above were all above 0.05 substitutions/site. SeMNPV-QD has 123 ORFs similar to those of SeMNPV-US1, and the genomic similarity was 45.8%. Compared to SeMNPV-US1, SeMNPV-QD has four additional ORFs such as two baculovirus repeat ORF (bro) genes, bro-1, bro-2, orf39 and orf95, but lacks 17 ORFs that have no effect on viral transcription and replication. The above results indicate that SeMNPV-QD is a new species of S. exigua multiple nucleopolyhedrovirus.