Iophenoxic acid as a bait marker for wild mammals: efficacy and safety considerations

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Crystallographic analysis reveals the structural basis of the high-affinity binding of iophenoxic acid to human serum albumin

Background  

Iophenoxic acid is an iodinated radiocontrast agent that was withdrawn from clinical use because of its exceptionally long half-life in the body, which was due in part to its high-affinity binding to human serum albumin (HSA). It was replaced by Iopanoic acid, which has an amino rather than a hydroxyl group at position 3 on the iodinated benzyl ring and, as a result, binds to albumin with lower affinity and is excreted more rapidly from the body. To understand how iophenoxic acid binds so tightly to albumin, we wanted to examine the structural basis of its interaction with HSA.     

Specificity and success of oral-bait delivery to Eurasian wild boar in Mediterranean woodland habitats

Eurasian wild boar (Sus scrofa) is an important reservoir host for pathogens affecting humans and domestic animals. The eradication of these diseases may require the development of control strategies that reduce pathogen transmission between wildlife and domestic animals. Baiting for oral vaccine delivery is often considered for wildlife disease control. The effective and efficient field vaccination of wildlife requires species-specific baits as delivery vehicles for oral vaccines and designing appropriate baiting strategies. The objective of this study was to determine the proportion of young and adult wild boar and non-target animals that consumed baits containing a chemical marker, iophenoxic acid (IPA), in delivery trials conducted in summer in four different sites in the Mediterranean region of Spain where wild boar are abundant. The proportion of wild boar showing IPA markers in serum in autumn ranged from 11.5% to 56.4%. When attending to age classes, 12.6% to 72.7% of young individuals presented IPA. The results evidenced that the percent of wild boar that ingested the baits varied among study sites and age classes. Placing baits inside selective cages (for juveniles) and under heavy pavel stones (for adults) contributed to improve age specificity in bait consumption. We suggest ways for improving the age specificity of bait delivery systems used for young and adult wild boar.     

Field trials of slow‐release synthetic lures for stoats (Mustela erminea) and ferrets (M. furo)

Abstract

Experimental field trials were used to compare the attractiveness to stoats (Mustela erminea) and ferrets (M. furo) of traps set with either a synthetic scent lure or with fresh food bait. One lure, containing 2‐n‐propylthietane, effectively attracted as many ferrets as did rabbit meat bait. Fewer stoats were caught with this lure than with egg bait. Made up in a slow‐release formulation in “plastic rope”, the lure remained attractive to ferrets for at least 2 weeks under sunny field conditions. Traps containing a second lure, 3‐n‐propyl‐l,2‐dithiolane, were less successful at catching both mustelid species than those containing fresh food baits. The scent lures were not attractive to non‐target species, but neither did they deter some species.     

Did ‘precautionary’ 1080 baiting have a realistic potential to eradicate Red Fox (Vulpes vulpes) in Tasmania without in situ monitoring data?

Anecdotal reports in 2001 suggested that the European Red Fox (Vulpes vulpes) had been deliberately released in Tasmania and thereafter an eradication programme using buried fluoroacetic acid (1080) baits was believed to be a necessary precautionary action until mid‐2013. Prerequisites for the successful eradication of foxes relate to the scale of the undertaking and the ability to collect in situ data such as the distribution and abundance of the target population and measures of the efficacy of the control technique. Previously, 1080 baiting has demonstrated only limited potential as a fox eradication technique on islands when used on a scale between 685 and 2141 times smaller than Tasmania. In the absence of empirical monitoring data confirming the distribution or abundance of extant foxes, buried baiting was targeted to specific landscapes believed to be preferred by foxes. No empirical data was collected concerning the in situ effectiveness of baiting in Tasmania, yet an a priori assumption of lethal efficacy was extrapolated from four heterogeneous mainland studies to suggest that foxes would have only a 0.23 probability of surviving each bait treatment. We show that these studies were unrepresentative of Tasmanian baiting methods used and influenced by imprecise fox population surveys and misreported data. Overall, in the absence of key population monitoring and efficacy data, the ‘precautionary’ baiting strategy adopted did not have a realistic potential to eradicate fox incursions in Tasmania, nor is it an appropriate risk management strategy for other large offshore Australian islands. Contingency plans to counter fox incursions on offshore islands must address the currently inadequate technical capacity to reliably detect and monitor low‐density fox populations, which is an essential component of successful fox eradication.     

Brugia pahangi and Dirofilaria immitis: experimental infections in the ferret, Mustela putorius furo.

Ferrets were inoculated with 160 third-stage larvae of the filarial nematode Brugia pahangi, followed 23 days later by 15 larvae of another filarial nematode, Dirofilaria immitis. Other ferrets received only one of these species. Microfilaremia developed in some ferrets with single infections of each species and in some ferrets with dual infections. The nature of the experiment did not permit a thorough study of microfilaremia, but B. pahangi microfilariae were found in numbers as high as 15,650/ml. At necropsy, approximately 8 months after inoculation, adult B. pahangi were recovered from the lymphatic vessels of all 8 ferrets inoculated only with that species, the recovery rate (based on 6 animals only) varying from 2 to 50% of the inoculum (mean 25%). Adult D. immitis were recovered from the heart of all three ferrets inoculated only with that species, the recovery rate being 7, 47, and 60% (mean 38%) of the inoculum. All 5 ferrets inoculated with both species yielded both adult B. pahangi (6 to 23%, mean 16% of inoculum) and adult D. immitis (13 to 67%, mean 37% of inoculum). It is concluded that the ferret is highly susceptible to both species and that concurrent infections with both species may readily be established.     

Intertribal somatic hybrids between <Emphasis Type="Italic">Brassica napus</Emphasis> and <Emphasis Type="Italic">Camelina sativa</Emphasis> with high linolenic acid content

Intertribal somatic hybrids of Brassica napus and Camelina sativa were developed by protoplast electrofusion. Hybrid identity of the regenerants was determined using flow cytometric analysis of nuclear DNA content and simple sequence repeat (SSR) marker analysis. Three hybrids exhibited specific bands for B. napus and C. sativa. These hybrids showed intermediate leaf, flower and seed morphology compared with the two parental species. The seeds of these three hybrids had a modified fatty acid profile, indicating higher level of linolenic and eicosanoic acids than those of B. napus. Our results suggest that somatic hybridization offers opportunities for transferring entire genomes between B. napus and C. sativa in improving rapeseed breeding.     

Presence of Bacillus coagulans spores and vegetative cells in rat intestine and feces and their physiological effects

ABSTRACT

This study was aimed to investigate the presence of Bacillus coagulans vegetative cells in the intestine and fecal samples in rats fed B. coagulans spores as well as to estimate the ratios of spores and vegetative cells in these samples. A two-step process has been developed to enumerate B. coagulans in different mixed bacterial samples, specifically (1) observation of yellow ring formation on modified GYEA medium upon incubation at 55°C, (2) microscopic examination of spore formation after 7 d of incubation. Our results have demonstrated the presence of vegetative cells in the intestinal and fecal samples in rats fed B. coagulans spores. The ratios of B. coagulans spores and vegetative cells in cecal fluid, colonic content, and feces were approximately 2:8, 2:8, and 4:6, respectively. The existence of B. coagulans vegetative cells improved the intestinal milieu through an elevated short-chain fatty acid concentrations, higher fecal moisture, and lower fecal pH.     

Fox‐baiting in agricultural landscapes in south‐eastern Australia: a case‐study appraisal and suggestions for improvement

Summary In south‐eastern Australia, the introduced Red Fox (Vulpes vulpes) is a major predator of native wildlife and livestock. Fox control in agricultural landscapes is heavily reliant on the laying of poisoned baits by private landholders, yet there have been few assessments of the application or success of landholder‐baiting practices. We evaluated a community‐based fox‐baiting campaign, typical of programs employed throughout the agricultural regions of south‐eastern Australia to control foxes. We recorded the spatial coverage of 1080 baits deployed by landholders, assessed baiting procedures, monitored the survival of six radio‐collared foxes during and after baiting, and compared the spatial coverage and likely effectiveness of the baiting program with two alternative (theoretical) baiting strategies. Relative to other baiting programs, coordination among neighbours was reasonably high, with 37.5% of baited properties (n = 40) adjoining ≥3 neighbouring properties that also contained baits. Nevertheless, the maximum distance from the centre of a baited property to the nearest edge of an unbaited property was <750 m (mean = 380 m ± 147 m SD). On average, 33% (±17% SD) of each fox’s home range overlapped with baited properties, but only two foxes died during the baiting program. The remaining four foxes were still alive 10 weeks after baiting ceased. Modelling of simulated fox home ranges showed that 13.5% contained no bait stations based on the community baiting program, whereas alternative roadside‐ and grid‐baiting strategies (theoretically) delivered baits to all simulated home ranges. Some landholders employed practices that could reduce the effectiveness of baiting programs such as not removing decayed baits before deploying new ones or placing bait stations too close together. Our research illustrates the difficulties of managing a coordinated baiting program on private land that effectively controls foxes. Alternative baiting strategies such as roadside baiting need to be considered to improve fox control in agricultural landscapes.     

Demographic and functional responses of wild dogs to poison baiting

Lethal control of wild dogs – that is Dingo (Canis lupus dingo) and Dingo/Dog (Canis lupus familiaris) hybrids – to reduce livestock predation in Australian rangelands is claimed to cause continental‐scale impacts on biodiversity. Although top predator populations may recover numerically after baiting, they are predicted to be functionally different and incapable of fulfilling critical ecological roles. This study reports the impact of baiting programmes on wild dog abundance, age structures and the prey of wild dogs during large‐scale manipulative experiments. Wild dog relative abundance almost always decreased after baiting, but reductions were variable and short‐lived unless the prior baiting programme was particularly effective or there were follow‐up baiting programmes within a few months. However, age structures of wild dogs in baited and nil‐treatment areas were demonstrably different, and prey populations did diverge relative to nil‐treatment areas. Re‐analysed observations of wild dogs preying on kangaroos from a separate study show that successful chases that result in attacks of kangaroos by wild dogs occurred when mean wild dog ages were higher and mean group size was larger. It is likely that the impact of lethal control on wild dog numbers, group sizes and age structures compromise their ability to handle large difficult‐to‐catch prey. Under certain circumstances, these changes sometimes lead to increased calf loss (Bos indicus/B. taurus genotypes) and kangaroo numbers. Rangeland beef producers could consider controlling wild dogs in high‐risk periods when predation is more likely and avoid baiting at other times.     

碱性条件下苏云金芽胞杆菌基础代谢分析

【目的】探索苏云金芽胞杆菌(Bacillus thuringiensis)形成转录差异的碱性条件,明确B.thuringiensis在该条件下的基础代谢途径变化。【方法】采用半定量RT-PCR技术及实时荧光定量PCR技术,确定碱刺激下参考基因psp A存在表达差异的碱性处理条件。在该条件下提取RNA进行Agilent定制B.thuringiensis表达谱芯片杂交,对芯片数据进行差异表达分析、GO富集分析及生物途径富集分析等。【结果】通过检测psp A表达变化,将对数生长中期的菌体加入终浓度为28 mmol/L的Na OH并诱导培养10min,作为B.thuringiensis响应碱刺激的研究条件。富集分析表明碳代谢、脂肪酸合成代谢、氨基酸合成代谢途径变化明显。细胞糖酵解途径至少19个酶促基因上调表达,三羧酸循环中催化α-酮戊二酸转化为苹果酸的大部分酶蛋白编码基因上调2倍以上。【结论】本研究发现在碱性条件下B.thuringiensis基础代谢明显增强,细胞可能通过大量合成酸性物质如乳酸、苹果酸等来提高细胞对于碱性环境的适应能力。     

液体植物保护膜防治瓜实蝇的效果评价

【背景】瓜实蝇是一种重要的世界性检疫害虫,广泛分布在我国的热带、亚热带地区。在室内和田间测定了液体植物保护膜对瓜实蝇的控制效能,并初步探索了该保护膜的作用方式,以期为瓜实蝇的绿色防控提供一定的技术支撑。【方法】测定了应用保护膜后瓜实蝇的产卵、存活以及对苦瓜的为害率。【结果】保护膜显著影响了瓜实蝇产卵、存活以及选择性。随着保护膜浓度的升高,瓜实蝇的存活率和在供试苦瓜段上的产卵量逐渐下降,但300和400倍保护膜之间的差异不显著。当浓度达到400倍时产卵量最低,为0.33粒,死亡率最高,约为50%;室内研究发现保护膜可持续影响瓜实蝇产卵5 d;此外,通过将保护膜应用在损伤的苦瓜上,发现保护膜并没有抑制瓜实蝇产卵。在田间,研究发现苦瓜的果长影响了保护膜的应用效能,2种果长的苦瓜应用保护膜后12~14 cm苦瓜的被害率显著低于6~8 cm苦瓜的被害率。【结论与意义】400倍的植物液体保护膜是防治瓜实蝇的最佳浓度,应用时间建议每4 d一次。保护膜最合适的应用时期为果长发育至12~14 cm时,即生长后期。施用保护膜时一定均匀,且在寄主果实受到损伤时不宜喷施保护膜。     

Field evaluation of alpha-chlorohydrin against the Indian mole rat: studies on toxic and antifertility effects

Field acceptance and efficacy of a toxicant-sterilant, alpha-chlorohydrin (α-CH), at its 0.5% concentration in bait were evaluated against rodents in sugarcane fields which harboured high populations of the Indian mole rat Bandicota bengalensis (Trap Index (TI) = 53.6 rats/100 traps/24 h) followed by that of the Indian bush rat Golunda ellioti (TI = 28.6) and soft-furred field rat Rattus meltada (TI = 1.8). Acceptance of the α-CH bait by rodents in the fields was evident from complete consumption of the offered bait at most of the baiting points. The treatment (72 h exposure to the poison bait) resulted in 63.7% to 82.9% rodent mortality. Survey of the mature sugarcane crop revealed that the percentage of rodent cut canes in the treated fields (7.6% to 16.2%) was significantly less than that of the reference fields (26.3%). Most of the surviving male B. bengalensis, captured after 15 days of the treatment, had developed sterility as revealed by the functional abnormalities in their testes and epid-idymides. They showed decreased thickness of the seminiferous tubules, lower population of spermatogenic cells, cauda epididymal sperm concentration, live sperm and sperm mortality. A nonsignificant positive correlation between the testicular weight and sperm motility in males from treated fields indicated the effect of α-CH at maturation level in the cauda epididymides. Overall, it was evident that a significant proportion of the surviving B. bengalensis had become permanently sterile as a consequence to the development of spermatocoeles in the caput epididymides and the remaining had significantly low numbers (< 30%) of motile sperm in their cauda epididymides which was, obviously, a handicap for successful fertilisation.     

Microbial control of Agrilus planipennis (Coleoptera: Buprestidae) with Beauveria bassiana strain GHA: field applications

The effects of Beauveria bassiana strain GHA, applied as BotaniGard ES, on newly colonised and well-established populations of emerald ash borer, Agrilus planipennis (Coleoptera: Buprestidae) were evaluated in the field using foliar and trunk sprays in Michigan in 2004–2005. Results from field trials at a newly colonised white ash site showed a 41% reduction in A. planipennis population in fungal-treated trees compared with that of untreated controls. In addition, fungal infection was also found in 20% of the larval population within 14 days of incubation under laboratory conditions. At a site with a well-established Agrilus planipennis population in green ash trees, larval density was reduced by 47% for trees treated with the fungus compared with that of the controls; 21% of larvae from the current generation were found infected after 14 days of laboratory incubation. Fungal-treated green ash trees also produced fewer adults emerging in the next generation, with a 63% reduction in adult density observed in treated trees compared to that of controls. As a result, fungal-treated trees sustained 42% less crown dieback than did controls. A. planipennis larval density was negatively correlated with trunk height above the ground, and positively correlated with log diameter. Results of laboratory leaf bioassays on A. planipennis adults showed that fungal conidia persisted well under field conditions, with mortality of 78–100% at 7 – days post – exposure for leaves collected between 2 and 264 h after application. Potential strategies for using B. bassiana strain GHA for managing A. planipennis are discussed.     

Baiting with Sclerotium rolfsii for selective isolation of Gliocladium virens from natural soil

A baiting technique for selective isolation of Gliocladium virens from natural soil was developed by mixing Sclerotium rolfsii —colonized sorghum grains with moist natural soil and incubating at 30 ± 5°C for 6–10 days. G. virens developed large, distinct colonies on the soil surface by colonizing S. rolfsii and was then easily isolated for use in biocontrol programmes. Trichoderma spp. were present in the soil but never developed conspicuous colonies on the soil surface, even when the soil was supplemented with large numbers of conidia.     

Simulated aerial sprays for field cage evaluation of Beauveria bassiana and Metarhizium brunneum (Ascomycetes: Hypocreales) against Anabrus simplex (Orthoptera: Tettigoniidae) in Montana

Field efficacy of the entomopathogenic Ascomycetes Beauveria bassiana strain GHA and Metarhizium brunneum strain F52 was evaluated against nymphs of the Mormon cricket, Anabrus simplex. Fungi were applied with a new apparatus that allows simulated aerial sprays to 0.1-m² areas in the field. The Mormon crickets were then individually housed in cylindrical, metal hardware cloth cages on treated grass. Both fungi demonstrated only marginal success in reducing immature Mormon cricket survival in the field cages. After 28 days, the field mortality of insects, corrected for control deaths, was 28 and 45% for B. bassiana and M. brunneum, respectively. Field-exposed but laboratory-incubated insects, however, suffered 90–100% mortality within 12 days with initial deaths occurring on Day 6 (Beauveria) or Day 5 (Metarhizium), indicating a lethal rate was applied and acquired by the crickets in the field. Potential daily body temperatures were determined for the entire post-treatment observation period using special thermal surrogates. High ambient temperatures and/or thermoregulation by Mormon crickets, in excess of the upper thermal limits of both fungi, prevented higher mortality from being expressed in the field. Thermal surrogates were used to develop models for predicting onset of mortality from infections. The surrogate data indicated mortality should begin between 8 and 26 days after treatment with M. brunneum and 11 and 33 days after treatment with B. bassiana. The timing of mortality in field cages was consistent with the upper boundaries of the temporal models developed from the thermal surrogates, i.e., at or after 28 days after treatment.     

An optimized microsatellite marker set for detection of Metarhizium anisopliae genotype diversity on field and regional scales

Thirty-three Metarhizium anisopliae isolates sampled across Switzerland as well as 35 and 36 M. anisopliae isolates sampled from two field sites were assembled in three isolate collections. All isolates were analyzed using 27 newly developed and 14 previously published microsatellite markers. The 41 markers allowed for detection of 25 genotypes in the Swiss collection while 30 and 11 genotypes were detected in the two field collections. This indicated high genetic diversity on a regional as well as on a field scale. In order to improve genotyping efficiency, an optimized marker set, which allows discrimination of a large number of genotypes with as few markers as possible was developed. The optimized marker set consisted of 16 common markers, which provided resolution close to maximal resolution in all three collections (91–93 %). The results demonstrated that optimized marker sets have to be validated before large scale application to previously unassessed collections in order to avoid suboptimal resolution. This genetic tool will be valuable for analyses of genetic population structure of M. anisopliae in different habitats on a regional as well as on a field scale.     

Structural improvement of LidNA: delta-type LidNA is a potent miRNA inhibitor constructed with unmodified DNA

ABSTRACT

Many miRNA inhibitors have been developed, including chemically modified oligonucleotides, such as 2′-O-methylated RNA and locked nucleic acid (LNA). Unmodified DNA has not yet been reported as a miRNA inhibitor due to relatively low DNA/miRNA binding affinity. We designed a structured DNA, LidNA, which was constructed with unmodified DNA, consisting of a complementary sequence to the target miRNA flanked by two structured DNA regions, such as double-stranded DNA. LidNA inhibited miRNA activity more potently than 2′-O-methylated RNA or LNA. To optimize LidNA, two double-stranded regions were joined, causing the molecule to assume a delta-like shape, which we termed delta-type LidNA. Delta-type LidNAs were developed to target endogenous and exogenous miRNAs, and exhibited potent miRNA inhibitory effects with a duration of at least 10 days. Delta-type LidNA-21, which targeted miR-21, inhibited the growth of cancer cell lines. This newly developed LidNA could contribute to miRNA studies across multiple fields.

Abbreviations: LidNA: DNA that puts a lid on miRNA function; LNA: locked nucleic acid; 3′-UTR: 3′-untranslated regions; RISC: RNA-induced silencing complex; MBL: Molecular beacon-like LidNA; YMBL: Y-type molecular beacon-like LidNA; TDMD: target-directed microRNA degradation.     

Efficient assembly of a large fragment of monkeypox virus genome as a qPCR template using dual-selection based transformation-associated recombination

Transformation-associated recombination (TAR) has been widely used to assemble large DNA constructs. One of the significant obstacles hindering assembly efficiency is the presence of error-prone DNA repair pathways in yeast, which results in vector backbone recircularization or illegitimate recombination products. To increase TAR assembly efficiency, we prepared a dual-selective TAR vector, pGFCS, by adding a P_ADH1-URA3 cassette to a previously described yeast-bacteria shuttle vector, pGF, harboring a P_HIS3-HIS3 cassette as a positive selection marker. This new cassette works as a negative selection marker to ensure that yeast harboring a recircularized vector cannot propagate in the presence of 5-fluoroorotic acid. To prevent pGFCS bearing ura3 from recombining with endogenous ura3-52 in the yeast genome, a highly transformable Saccharomyces cerevisiae strain, VL6-48B, was prepared by chromosomal substitution of ura3-52 with a transgene conferring resistance to blasticidin. A 55-kb genomic fragment of monkeypox virus encompassing primary detection targets for quantitative PCR was assembled by TAR using pGFCS in VL6-48B. The pGFCS-mediated TAR assembly showed a zero rate of vector recircularization and an average correct assembly yield of 79% indicating that the dual-selection strategy provides an efficient approach to optimizing TAR assembly.     

Baiting improves CPUE in nine‐spined stickleback (Pungitius pungitius) minnow trap fishery

Whether or not baiting influences stickleback catch per unit effort (CPUE) remains a matter of debate among stickleback researchers: While the opinions about the impact of baiting on CPUE differ, supporting quantitative data are scarce. The effect of baiting and trap type on nine‐spined stickleback (Pungitius pungitius) CPUE was studied in a field experiment conducted over four consecutive days in a small pond in northeastern Finland. The results show that baited traps yielded better (mean CPUE = 1.24 fish/trap/d) catches than unbaited traps (mean CPUE = 0.66); however, there were also differences in CPUE depending on the type of collapsible trap that was used. The trap type effect on CPUE seemed to differ among age classes – the finer meshed trap caught more young‐of‐the‐year fish than the coarse‐meshed one, whereas the opposite was true for the older and larger individuals. The results agree with those of an earlier more restricted study conducted in the same locality: Together, these results provide strong evidence for the positive impact of baiting on nine‐spined stickleback CPUE.