Comparison of Worm Control Strategies in Grazing Sheep in Denmark

Control of nematode parasites with reduced reliance on the use of anthelmintics was studied in 16 ewes with suckling twin lambs on contaminated pasture in Denmark. Ewes and lambs were treated with albendazole at turn-out 3 May. Ewes were removed from the groups on 26 July, and lambs were slaughtered on 11 October. The animals were allocated to 4 groups of 8 lambs and their 4 ewes. Group TS was treated with albendazole at weeks 3, 6 and 8 after turnout and set-stocked; group TM was similarly treated but moved to clean pasture in conjunction with the last drenching; group US was untreated and set-stocked, and group UM was left untreated but moved to clean pasture week 8 after turn-out. Supplementary feed was offered in June and August due to scarcity of pasture. Strategic treatments of ewes and lambs weeks 3, 6 and 8 after turn-out, with or without a move to clean pasture, were highly effective in controlling nematode infections for most of the season. This was reflected in better weight gains and carcass characteristics in the treated compared to untreated lambs, resulting in an average increase in the value of the product by 36%. The effect of moving without treatment (UM) on faecal egg counts was limited but peak pasture infectivity was reduced to less than 10% compared to the set-stocked group and weight gains of lambs were significantly better despite poor feed availability in late season. The study showed that under set-stocked conditions repeated anthelmintic treatments of both ewes and lambs in early season may ensure sufficient nematode control whereas moving animals to clean pasture without dosing was less efficient. The latter may, however, still be a viable option in organic and other production systems where routine use of anthelmintics is banned, particularly if weaning and moving are combined or a second move is performed.     

Investigation of apoptosis in cultured cells infected with equine herpesvirus 1

Many viruses alter different stages of apoptosis of infected cells as a strategy for successful infection. Few studies have addressed mechanisms of equine herpesvirus 1 (EHV-1) strain-induced cell death. We investigated the effect of an abortigenic strain (AR8 strain) on heterologous Madin–Darby bovine kidney cells and homologous equine dermis (ED) cells cell lines. We compared morphologic and biochemical features of early and late apoptosis at different postinfection times. We investigated translocation of phosphatidylserine to the cell surface, nuclear fragmentation and changes in the cytoskeleton using flow cytometry and annexin V/propidium iodide staining, DNA laddering, terminal deoxynucleotidyl transferase UTP nick-end labeling assay and immunofluorescence staining of cytokeratin 18 cleavage. AR8 EVH-1 strain interfered with apoptosis in both cell lines, particularly during the middle stage of the replication cycle; this was more evident in ED cells. Although this antiapoptotic effect has been reported for other alpha herpesviruses, our findings may help elucidate how EHV-1 improves its infectivity during its cycle.     

在中国应用垂直波束雷达监测褐飞虱和其他水稻害虫迁飞的可行性

近年来,可用于昆虫迁飞研究且可自动运行的垂直波束雷达(vertical-looking radar,VLR)的发展使得对迁飞性害虫的周年长期自动监测成为可能。本文提供了我们对能否将这种雷达应用于中国的褐飞虱和其他水稻害虫的监测与预测体系以改善其综合治理的可行性研究结果。以往的研究已经表明,这些害虫一般在300—2000m高度迁飞;而我们根据褐飞虱的雷达和射有效截面的计算结果表明,目前使用的3．2cm波长的VLR对褐飞虱个体目标的最大可检测高度仅约240m;虽然建造一部8．8mm波长的VLR即可覆盖褐飞虱迁飞高度的绝大部分,但其造价和维护费用均过于昂贵。为此,一个更可行的解决方案是,以3．2cm波长的VLR作为包括大多数水稻害虫在内的个体较大的迁飞性害虫的监测工具。     

Functional changes in human leukemic cell line HL-60. A model for myeloid differentiation

Polar solvents induce terminal differentiation in the human promyelocytic leukemia cell line HL-60. The present studies describe the functional changes that accompany the morphologic progression from promyelocytes to bands and poly-morphonuclear leukocytes (PMN) over 9 d of culture in 1.3 percent dimethylsulfoxide (DMSO). As the HL-60 cells mature, the rate of O(2-) production increase 18-fold, with a progressive shortening of the lag time required for activation. Hexosemonophosphate shunt activity rises concomitantly. Ingestin of paraffin oil droplets opsonized with complement or Ig increases 10-fold over 9 d in DMSO. Latex ingestion per cell by each morphologic type does not change significantly, but total latex ingestion by groups of cells increases with the rise in the proportion of mature cells with greater ingestion capacities. Degranulation, as measured by release of β-glucuronidase, lysozyme, and peroxidase, reaches maximum after 3-6 d in DMSO, then declines. HL-60 cells contain no detectable lactoferrin, suggesting that their secondary granules are absent or defective. However, they kill staphylococci by day 6 in DMSO. Morphologically immature cells (days 1-3 in DMSO) are capable of O(2-) generation, hexosemonophosphate shunt activity, ingestion, degranulation, and bacterial killing. Maximal performance of each function by cells incubated in DMSO for longer periods of time is 50-100 percent that of normal PMN. DMSO- induced differentiation of HL-60 cells is a promising model for myeloid development.     

Assessing the reliability of eBURST using simulated populations with known ancestry

Background  

The program eBURST uses multilocus sequence typing data to divide bacterial populations into groups of closely related strains (clonal complexes), predicts the founding genotype of each group, and displays the patterns of recent evolutionary descent of all other strains in the group from the founder. The reliability of eBURST was evaluated using populations simulated with different levels of recombination in which the ancestry of all strains was known.     

Coronavirus Genetically Redirected to the Epidermal Growth Factor Receptor Exhibits Effective Antitumor Activity against a Malignant Glioblastoma

Coronaviruses are positive-strand RNA viruses with features attractive for oncolytic therapy. To investigate this potential, we redirected the coronavirus murine hepatitis virus (MHV), which is normally unable to infect human cells, to human tumor cells by using a soluble receptor (soR)-based expression construct fused to an epidermal growth factor (EGF) receptor targeting moiety. Addition of this adapter protein to MHV allowed infection of otherwise nonsusceptible, EGF receptor (EGFR)-expressing cell cultures. We introduced the sequence encoding the adaptor protein soR-EGF into the MHV genome to generate a self-targeted virus capable of multiround infection. The resulting recombinant MHV was viable and had indeed acquired the ability to infect all glioblastoma cell lines tested in vitro. Infection of malignant human glioblastoma U87ΔEGFR cells gave rise to release of progeny virus and efficient cell killing in vitro. To investigate the oncolytic capacity of the virus in vivo, we used an orthotopic U87ΔEGFR xenograft mouse model. Treatment of mice bearing a lethal intracranial U87ΔEGFR tumor by injection with MHVsoR-EGF significantly prolonged survival compared to phosphate-buffered saline-treated (P = 0.001) and control virus-treated (P = 0.004) animals, and no recurrent tumor load was observed. However, some adverse effects were seen in normal mouse brain tissues that were likely caused by the natural murine tropism of MHV. This is the first demonstration of oncolytic activity of a coronavirus in vivo. It suggests that nonhuman coronaviruses may be attractive new therapeutic agents against human tumors.Already for quite some years oncolytic viruses are being investigated for use in human tumor therapy (for recent reviews, see references 3, 16, 22, 37, and 44). Their success in destroying human cancer cells depends on their ability to selectively infect and kill these cells. Although some oncolytic viruses appear to have a natural tropism for tumor cells, most viruses need to be modified in some way to achieve infection and/or lytic activity in these cells. One of the ways to accomplish specific infection of tumor cells is by redirecting the virus to epitopes expressed on such cells. Thus, different targeting approaches have been explored for a variety of viruses. These include pseudotyping, modification of viral surface proteins, and the use of bispecific adapters (14, 35, 45). All of these approaches require that the viability of the virus is not hampered and that the targeting moiety is properly exposed to allow directed infection. The ability to genetically modify a particular virus combined with the availability of an appropriate targeting epitope determines the success of the approaches.Coronaviruses are enveloped, positive-strand RNA viruses belonging to the order Nidovirales. The nonhuman coronavirus murine hepatitis virus (MHV) is the best-studied coronavirus and, more importantly, convenient reverse genetics systems are available to modify its genome (19, 50). MHV has several appealing characteristics that might make it suitable as an oncolytic virus. First, it has a narrow host range, determined by the interaction of its spike (S) glycoprotein with the cellular receptor mCEACAM1a. Since mCEACAM1a is not expressed on human cells, MHV cannot establish an infection in either normal or cancerous human cells. Second, infection by MHV induces the formation of large multinucleated syncytia, to which also surrounding uninfected cells are recruited (42). Hence, given also its relatively short replication cycle (6 to 9 h), MHV destroys populations of cells rapidly once they have become infected. Third, the tropism of MHV can be modified either by substitution of the viral spike ectodomain (19) or by the use of adapter proteins (43, 47, 48). These adapter proteins, composed of a virus-binding moiety coupled to a target cell-binding device, can redirect the virus to a specific receptor on the target cell (43, 47, 48). The studies revealed that, once the host cell tropism barrier is alleviated, MHV is capable of establishing infection in nonmurine cells.The use of adapter proteins to target therapeutic viruses to tumor cells (modeled in Fig. ​Fig.1A)1A) is limited by the necessity to externally provide and replenish the adapter protein. To overcome this obstacle, the genetic information for the targeting device can be introduced into the viral genome to allow the virus to produce the adaptor itself in infected cells, thereby creating a self-targeted virus. Indeed, we demonstrated the feasibility of this concept by expressing an adapter protein—composed of the relevant (soluble) domain of the mCEACAM1a receptor linked to a His tag—as an additional protein from the MHV genome (43). We extend these investigations here by coupling the epidermal growth factor (EGF) protein to the mCEACAM1a fragment. Introduction of this expression cassette encoding the adapter protein allowed multiround infection in EGF receptor (EGFR)-expressing human cells, resulting in extensive cell-cell fusion and efficient killing of target human glioblastoma cells. Using an orthotopic intracranial tumor model of aggressive U87ΔEGFR glioblastomas in nude mice, we show for the first time that redirected coronavirus has oncolytic potential.Open in a separate windowFIG. 1.Redirection of MHV to the EGFR. (A) Rationale for using adapter proteins to target viruses to a specific receptor present on target cells. Adapters, either added or expressed from the viral genome when incorporated, enable targeted infection of otherwise nonsusceptible cells. (B) Schematic diagram of the expression constructs pSTsoR-h-His and pSTsoR-EGF. The Igκ signal sequence directs adapter protein secretion, the N-terminal D1 domain of mCEACAM1a (soR) provides for binding to and induction of conformational changes in the MHV spike protein, and the six-amino-acid His tag (His) provides for binding to the artificial His receptor. The hinge (h) linker region present in soR-h-His allows formation of disulfide-linked dimers of the resulting adapter protein (43) (T7, T7 promoter; myc, myc tag; ala3, alanine tripeptide). (C) Targeting of MHV-EFLM to the His receptor and to the EGFR on CHO cells using soR-h-His and soR-EGF, respectively. CHO, CHO-His.scFv, and CHO-EGFR cells were inoculated with MHV-EFLM preincubated with soR-h-His or soR-EGF, and the luciferase activities (expressed as relative light units [RLU]) were measured. The values depicted are the means of an experiment performed in triplicate. Error indicators show the standard deviations.