Characterization of LORF11, a unique gene common to the three Marek's disease virus serotypes

The unique open reading frame 11 (LORF11) of Marek's disease virus (MDV) is present in all three serotypes of MDV and is located in the unique long region of the MDV genome. In the serotype 1 Md5 genome, LORF11 comprises 2711 nucleotides and encodes a predicted protein of 903 amino acids. In order to study the biological function of LORF11 we deleted it from the MDV cosmid A6 by using the RecA-assisted restriction endonuclease cleavage method. The recombinant cosmid, A6DeltaLORF11, was transfected into duck embryo fibroblasts (DEF) in conjunction with parental SN5, P89, SN16, and B40 cosmid clones. Recombinant rMd5DeltaLORF11 plaques were evident at 12-13 days after transfection. Polymerase chain reaction amplification of DEF cells infected with rMd5DeltaLORF11 viruses confirmed the deletion of a 2.57-kb fragment resulting in a 296-bp fragment. Three rMd5DeltaLORF11 mutants were generated and their biological functions were studied in vitro and in vivo. In vitro growth characteristics of rMd5DeltaLORF11 viruses were similar to those of parental rMd5, indicating that LORF11 is not essential for replication in vitro. In vivo studies of rMd5DeltaLORF11 mutants showed that they were impaired in viral replication in the lymphoid organs and had 100x lower viremia than chickens infected with the parental rMd5 virus. Furthermore, rMd5-infected chickens horizontally transmitted the virus to contact controls whereas no horizontal transmission occurred in rMd5DeltaLORF11-infected chickens. Three independent deletion mutants were tested and showed the same phenotypes, so it is unlikely that the observed phenotype is because of any random mutation in the genome. Therefore the LORF11 gene of MDV is essential for normal virus replication in chickens and deletion of LORF11 renders an attenuated virus.     

Hematologic characteristics of captive western barred bandicoots (Perameles bougainville) from Western Australia

BACKGROUND: The western barred bandicoot (Perameles bougainville) is an Australian marsupial species now considered endangered as a consequence of habitat destruction and predation. A recently discovered papillomatosis syndrome is hindering efforts to repopulate this species. Hematology reference intervals have been lacking for P bougainville, preventing optimal interpretation of hematology results from wart-affected and clinically normal animals. OBJECTIVES: The purpose of this study was to establish hematology reference values and describe morphologic characteristics of blood cells of healthy western barred bandicoots. METHODS: Fifty-nine whole blood samples were collected by jugular venipuncture into EDTA from 47 clinically healthy captive western barred bandicoots at 3 locations on the Western Australian mainland. A CBC was performed using an ADVIA-120 analyzer. Data were compared on the basis of geographic location, sex, age, and lactation status, and reference intervals were calculated. Blood cell morphology was evaluated using light microscopy, and transmission and scanning electron microscopy. RESULTS: Significant differences were found based on sex (RBC indices, fibrinogen), age (% polychromatophilic RBCs), and geographic location (RBC, neutrophil, and lymphocyte counts, MCHC, % polychromatophilic RBCs, fibrinogen). Combined reference intervals were calculated for hemoglobin concentration (122-165 g/L), HCT (0.36-0.49 L/L), and total WBC (2.9-14.9 x 10(9)/L), monocyte (0-0.6 x 10(9)/L), eosinophil (0-0.9 x 10(9)/L), and total plasma protein (47-63 g/L) concentrations. Leukocyte, erythrocyte, and platelet morphology were similar to those of other marsupial peramelid species. Nuclei in neutrophils, monocytes, and eosinophils occasionally had an annular configuration. CONCLUSIONS: Reference intervals and blood cell morphology obtained in this study will be useful for the evaluation of laboratory data from ill animals and assist with population health monitoring of western barred bandicoots.     

Occurrence of Giardia and Cryptosporidium in Norwegian red foxes (Vulpes vulpes)

Faecal samples from 269 Norwegian wild red foxes (Vulpes vulpes) shot during the hunting season (October-April) in 2002-2004 were examined for the presence of Giardia and Cryptosporidium. Cryptosporidium oocysts were detected in samples from 6 (2.2%) of the foxes, and Giardia cysts in 13 (4.8%) of the foxes. The prevalence of Giardia infection was significantly higher in juvenile male foxes than in adult male foxes, but no other significant differences between age and sex were found. No significant differences in prevalence related to geographical origin of animals were found. Insufficient nucleated Cryptosporidium oocysts were isolated for successful PCR, but genotyping of Giardia duodenalis isolates from seven foxes demonstrated a high degree of heterogeneity amongst them, with all isolates belonging to the zoonotic Assemblages A and B.     

Comparison of progestin-based protocols to synchronize estrus and ovulation before fixed-time artificial insemination in postpartum beef cows

This experiment was designed to compare pregnancy rates in postpartum beef cows resulting from fixed-time AI (FTAI) after treatment with 1 of 2 protocols to synchronize estrus and ovulation. Cross-bred, suckled beef cows (n = 650) at 4 locations (n = 210; n = 158; n = 88; and n = 194) were assigned within a location to 1 of 2 protocols within age group by days postpartum and BCS. Cows assigned to the melengestrol acetate (MGA) Select treatment (MGA Select; n = 327) were fed MGA (0.5 mg x head(-1) x d(-1)) for 14 d, GnRH (100 microg of Cystorelin i.m.) was injected on d 26, and prostaglandin F2alpha (PG; 25 mg of Lutalyse i.m.) was injected on d 33. Cows assigned to the CO-Synch + controlled internal drug release (CIDR) protocol (CO-Synch + CIDR; n = 323) were fed a carrier for 14 d, were injected with GnRH and equipped with an EAZI-BREED CIDR insert (1.38 g of progesterone, Pfizer Animal Health, New York, NY) 12 d after carrier removal, and PG (25 mg of Lutalyse i.m.) was injected and the CIDR were removed on d 33. Fixed-time AI was performed at 72 or 66 h after PG for the MGA Select or CO-Synch + CIDR groups, respectively. All cows were injected with GnRH (100 microg of Cystorelin i.m.) at the time of insemination. Blood samples were collected 8 and 1 d before the beginning of MGA or carrier to determine estrous cyclicity status of the cows (estrous cycling vs. anestrus) before treatment [progesterone > or = 0.5 ng/mL (MGA Select, 185/327, 57%; CO-Synch + CIDR, 177/323, 55%); P = 0.65]. There was no difference (P = 0.20) in pregnancy rate to FTAI between treatments (MGA Select, 201/327, 61%; CO-Synch + CIDR, 214/323, 66%). There was also no difference (P = 0.25) between treatments in final pregnancy rate at the end of the breeding period (MGA Select, 305/327, 93%; CO-Synch + CIDR, 308/323, 95%). These data indicate that pregnancy rates to FTAI were comparable after administration of the MGA Select or CO-Synch + CIDR protocols. Both protocols provide opportunities for beef producers to utilize AI and potentially eliminate the need to detect estrus.     

Generating and exploiting polarity in bacteria

Bacteria are often highly polarized, exhibiting specialized structures at or near the ends of the cell. Among such structures are actin-organizing centers, which mediate the movement of certain pathogenic bacteria within the cytoplasm of an animal host cell; organized arrays of membrane receptors, which govern chemosensory behavior in swimming bacteria; and asymmetrically positioned septa, which generate specialized progeny in differentiating bacteria. This polarization is orchestrated by complex and dynamic changes in the subcellular localization of signal transduction and cytoskeleton proteins as well as of specific regions of the chromosome. Recent work has provided information on how dynamic subcellular localization occurs and how it is exploited by the bacterial cell. The main task of a bacterial cell is to survive and duplicate itself. The bacterium must replicate its genetic material and divide at the correct site in the cell and at the correct time in the cell cycle with high precision. Each kind of bacterium also executes its own strategy to find nutrients in its habitat and to cope with conditions of stress from its environment. This involves moving toward food, adapting to environmental extremes, and, in many cases, entering and exploiting a eukaryotic host. These activities often involve processes that take place at or near the poles of the cell. Here we explore some of the schemes bacteria use to orchestrate dynamic changes at their poles and how these polar events execute cellular functions. In spite of their small size, bacteria have a remarkably complex internal organization and external architecture. Bacterial cells are inherently asymmetric, some more obviously so than others. The most easily recognized asymmetries involve surface structures, e.g., flagella, pili, and stalks that are preferentially assembled at one pole by many bacteria. "New" poles generated at the cell division plane differ from old poles from the previous round of cell division. Even in Escherichia coli, which is generally thought to be symmetrical, old poles are more static than new poles with respect to cell wall assembly (1), and they differ in the deposition of phospholipid domains (2). There are many instances of differential polar functions; among these is the preferential use of old poles when attaching to host cells as in the interaction of Bradyrhizobium with plant root hairs (3) or the polar pili-mediated attachment of the Pseudomonas aeruginosa pathogen to tracheal epithelia (4). An unusual polar organelle that mediates directed motility on solid surfaces is found in the nonpathogenic bacterium Myxococcus xanthus. The gliding motility of this bacterium is propelled by a nozzle-like structure that squirts a polysaccharide-containing slime from the pole of the cell (5). Interestingly, M. xanthus, which has nozzles at both poles, can reverse direction by closing one nozzle and opening the other in response to end-to-end interactions between cells.     

Park user preferences for establishing a sustainable forest park in Taipei,Taiwan

In Taiwan, urban parks have mainly cared for demands associated with recreational use and landscape aesthetics, while ecological functions have often been neglected. The purpose of this study was to examine park users’ awareness of the functions of urban parks, their attitudes toward Amsterdam Bos Park as well as their preferences and support for a sustainable forest park in Taipei metropolitan area. Moreover, the study intended to inform the decision makers in Taiwan about people's perception of urban parks. The findings suggested that participants were moderately aware of the economic benefit of urban parks, and highly aware of urban parks’ benefits regarding landscape beauty, social values, health and safety, and ecological conservation. Participants moderately agreed with the financial demand, and highly agreed with the development strategy, managerial operation, and design approach of the Bos Park. In addition, participants liked the concept of the Bos Park very much, and strongly supported the establishment of a sustainable forest park in Taipei metropolitan area. Based on the findings, recommendations were made to the Taipei City government for the decision-making in the development of a metropolitan park, as well as to landscape architects in the planning and design of urban parks.     

Phenotypic and Genotypic Heterogeneity among Streptococcus iniae Isolates Recovered from Cultured and Wild Fish in North America,Central America and the Caribbean Islands

Abstract

Streptococcus iniae, the etiological agent of streptococcosis in fish, is an important pathogen of cultured and wild fish worldwide. During the last decade outbreaks of streptococcosis have occurred in a wide range of cultured and wild fish in the Americas and Caribbean islands. To gain a better understanding of the epizootiology of S. iniae in the western hemisphere, over 30 S. iniae isolates recovered from different fish species and geographic locations were characterized phenotypically and genetically. Species identities were determined biochemically and confirmed by amplification and sequencing of the 16S rRNA gene. Repetitive-element palindromic PCR fingerprinting as well as biochemical and antimicrobial susceptibility profiles suggest that a single strain of S. iniae was responsible for two different disease outbreaks among reef fishes in the Caribbean, one in 1999 and another in 2008. Interestingly, a majority of the isolates recovered from cultured fish in the Americas were genetically distinct from the Caribbean isolates and exhibited a trend toward higher minimal inhibitory concentration with respect to several antibiotics as well as greater genetic variability. The biological significance of this genetic variability is unclear, but it could have implications for future vaccine development and treatment.

Received April 20, 2014; accepted July 7, 2014