Influence of phosphine resistance genes on flight propensity and resource location in Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae): the landscape for selection

Phosphine resistance in Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) has evolved through changes to enzymes involved in basic metabolic pathways. These changes impose metabolic stress and could affect energy-demanding behaviours. We therefore tested whether phosphine resistance alleles impact the movement of these insects in their quest for new resources. We measured walking and flight parameters of four T. castaneum genotypes: (1) a field-derived population, (2) a laboratory cultured, phosphine-susceptible reference strain, (3) a laboratory cultured, phosphine-resistant reference strain, and (4) a resistant introgressed strain that is almost identical genetically to the susceptible population. The temporal pattern of flight was identical across all populations, but resistant beetles took flight significantly less, walked more slowly, and located resources less successfully than did susceptible beetles. Also, the field-derived beetles (proved not to be carrying resistance genes) walked significantly faster and more directly towards food resources, and had a higher propensity for flight when compared to the susceptible laboratory beetles. These negative effects suggest survival of beetles with the resistance alleles will be compromised should they leave phosphine application sites. The field for selection therefore extends beyond the site at which phosphine fumigant imposed its effect, and other mutations are also likely to be affected in this way.

Selection and characterisation of two attenuated vaccine lines of Eimeria tenella in Australia

Objective To attenuate two strains of Eimeria tenella by selecting for precocious development and evaluate the strains in characterisation trials and by field evaluation, to choose one precocious line for incorporation into an Australian live coccidiosis vaccine for poultry. Design Two strains from non-commercial flocks were passaged through chickens while selecting for precocious development. Each strain was characterised for drug sensitivity, pathogenicity, protection against homologous and heterologous challenge, and oocyst output in replicated experiments in which the experimental unit was a cage of three birds. Oocyst output and/or body weight gain data collected over a 10 to 12 day period following final inoculation were measured. Feed conversion ratios were also calculated where possible. Results Fifteen passages resulted in prepatent periods reduced by 24 h for the Redlands strain (from 144 h to 120 h)and 23 h for the Darryl strain (from 139 h to 116 h). Characterisation trials demonstrated that each precocious line was significantly less pathogenic than its parent strain and each effectively induced immunity that protected chickens against challenge with both the parent strain and other virulent field strains. Both lines had oocyst outputs that, although significantly reduced relative to the parent strains, remained sufficiently high for commercial vaccine production, and both showed susceptibility to coccidiostats. Conclusion Two attenuated lines have been produced that exhibit the appropriate characteristics for use in an Australian live coccidiosis vaccine.

Genetic diversity, aggressiveness and metalaxyl sensitivity of Pythium aphanidermatum populations infecting cucumber in Oman

Seventy three isolates of Pythium aphanidermatum obtained from cucumber from four different regions of Oman and 16 isolates of muskmelon from the Batinah region in Oman were characterized for aggressiveness, sensitivity to metalaxyl and genetic diversity using AFLP fingerprinting. Twenty isolates of P. aphanidermatum from diverse hosts from different countries were also included in the study. Most isolates from Oman were found to be aggressive on cucumber seedlings and all were highly sensitive to metalaxyl (EC50 < 0•80 µg mL−1). Isolates from cucumber and muskmelon were as aggressive as each other on both hosts (P > 0.05), which implies a lack of host specialization in P. aphanidermatum on these two hosts in Oman. AFLP analysis of all isolates using four primer-pair combinations resolved 152 bands, of which 61 (~40%) were polymorphic. Isolates of P. aphanidermatum from Oman and other countries exhibited high genetic similarity (mean = 94.1%) and produced 59 different AFLP profiles. Analysis of molecular variance indicated that most AFLP variation among populations of P. aphanidermatum in Oman was associated with geographical regions (FST = 0.118; P < 0.0001), not hosts (FST = -0.004; P = 0.4323). These data were supported by the high rate of recovery (24%) of identical phenotypes between cucumber and muskmelon fields in the same region as compared to the low recovery (10%) across regions in Oman, which suggests more frequent movement of Pythium inoculum among muskmelon and cucumber fields in the same region compared to movement across geographically separated regions. However, recovering clones among regions and different countries may imply circulation of Pythium inoculum via common sources in Oman and also intercontinental spread of isolates.

Survival and reproduction of Tribolium castaneum (Herbst), Rhyzopertha dominica (F.) and Sitophilus oryzae (L.) following periods of starvation

This study determined the starvation tolerance of Tribolium castaneum (Herbst), Rhyzopertha dominica (F.) and Sitophilus oryzae (L.) in terms of both adult survival and reproduction, the impact of starvation on reproduction not having been studied before. Experiments were conducted at 30°C and 55% or 70% r.h. using a laboratory strain and a field strain of each species. The number of progeny was a better indicator of the impact of starvation on a species than adult survival. Tribolium castaneum was the most tolerant species, requiring up to 35 d starvation before no progeny were produced. Rhyzopertha dominica and S. oryzae required up to 8 d starvation before no progeny were produced. The results suggest that hygiene will have a greater impact on populations of S. oryzae and R. dominica than T. castaneum.

Rickettsia-like-organisms and phytoplasmas associated with diseases in Australian strawberries

Strawberry lethal yellows (SLY) disease in Australia is associated with the phytoplasmas Candidatus Phytoplasma australiense and tomato big bud, and a rickettsia-like-organism (RLO). Ca. P. australiense is also associated with strawberry green petal (SGP) disease. This study investigated the strength of the association of the different agents with SLY disease. We also documented the location of SLY or SGP plants, and measured whether they were RLO or phytoplasma positive. Symptomatic strawberry plants collected from south-east Queensland (Australia) between January 2000 and October 2002 were screened by PCR for both phytoplasmas and the RLO. Two previously unreported disease symptoms termed severe fruit distortion (SFD) and strawberry leaves from fruit (SLF) were observed during this study but there was no clear association between these symptoms and phytoplasmas or the RLO. Only two SGP diseased plants were observed and collected, compared with 363 plants with SLY disease symptoms. Of the 363 SLY samples, 117 tested positive for the RLO, 67 tested positive for Ca. P. australiense AGY strain and 11 plants tested positive for Ca. P. australiense PYL variant strain. On runner production farms at Stanthorpe, Queensland the RLO was detected in SLY diseased plants more frequently than for the phytoplasmas. On fruit production farms on the Sunshine Coast, Queensland, Ca. P. australiense was detected in SLY disease plants more frequently than the RLO.

Mortality and blood loss by blue swimmer crabs (Portunus pelagicus) after simulated capture and discarding from gillnets

Two laboratory experiments were carried out to quantify the mortality and physiological responses of juvenile blue swimmer crabs (Portunus pelagicus) after simulated gillnet entanglement, air exposure, disentanglement, and discarding. In both experiments, all but control blue swimmer crabs were entangled in 1-m(2) gillnet panels for 1 h, exposed to air for 2 min, subjected to various treatments of disentanglement ranging between the forceful removal of none, one, two, and four appendages, then "discarded" into individual experimental tanks and monitored for 10 d. In Experiment 1, mortalities were associated with the number of appendages removed and the occurrence of unsealed wounds. In Experiment 2, live blue swimmer crabs were sampled for blood at 2 min and 6, 24, and 72 h post-discarding to test for the effects of disentanglement and appendage removal on total haemocyte counts, clotting times, protein levels (by refractive index), and blood ion concentrations. Compared with blue swimmer crabs that had sealed or no wounds, those with unsealed wounds had lower total haemocyte counts, protein, and calcium concentrations and increased clotting ties and magnesium and sodium levels. Induced autotomy, as opposed to the arbitrary, forceful removal of a appendages has the potential to minimize the mortality and stress of discarded, juvenile blue swimmer crabs.

Effect of timing of a deficit-irrigation allocation on corn evapotranspiration, yield, water use efficiency and dry mass

Water regulations have decreased irrigation water supplies in Nebraska and some other areas of the USA Great Plains. When available water is not enough to meet crop water requirements during the entire growing cycle, it becomes critical to know the proper irrigation timing that would maximize yields and profits. This study evaluated the effect of timing of a deficit-irrigation allocation (150 mm) on crop evapotranspiration (ETc), yield, water use efficiency (WUE = yield/ETc), irrigation water use efficiency (IWUE = yield/irrigation), and dry mass (DM) of corn (Zea mays L.) irrigated with subsurface drip irrigation in the semiarid climate of North Platte, NE. During 2005 and 2006, a total of sixteen irrigation treatments (eight each year) were evaluated, which received different percentages of the water allocation during July, August, and September. During both years, all treatments resulted in no crop stress during the vegetative period and stress during the reproductive stages, which affected ETc, DM, yield, WUE and IWUE. Among treatments, ETc varied by 7.2 and 18.8%; yield by 17 and 33%; WUE by 12 and 22%, and IWUE by 18 and 33% in 2005 and 2006, respectively. Yield and WUE both increased linearly with ETc and with ETc/ETp (ETp = seasonal ETc with no water stress), and WUE increased linearly with yield. The yield response factor (ky) averaged 1.50 over the two seasons. Irrigation timing affected the DM of the plant, grain, and cob, but not that of the stover. It also affected the percent of DM partitioned to the grain (harvest index), which increased linearly with ETc and averaged 56.2% over the two seasons, but did not affect the percent allocated to the cob or stover. Irrigation applied in July had the highest positive coefficient of determination (R2) with yield. This high positive correlation decreased considerably for irrigation applied in August, and became negative for irrigation applied in September. The best positive correlation between the soil water deficit factor (Ks) and yield occurred during weeks 12-14 from crop emergence, during the "milk" and "dough" growth stages. Yield was poorly correlated to stress during weeks 15 and 16, and the correlation became negative after week 17. Dividing the 150 mm allocation about evenly among July, August and September was a good strategy resulting in the highest yields in 2005, but not in 2006. Applying a larger proportion of the allocation in July was a good strategy during both years, and the opposite resulted when applying a large proportion of the allocation in September. The different results obtained between years indicate that flexible irrigation scheduling techniques should be adopted, rather than relying on fixed timing strategies.

Identification of putative virulence-associated genes of Haemophilus parasuis through suppression subtractive hybridization.

Haemophilus parasuis is the causative agent of Glässer's disease. Up to now 15 serovars of H. parasuis have been identified, with significant differences existing in virulence between serovars. In this study, suppression subtractive hybridization (SSH) was used to identify the genetic difference between Nagasaki (H. parasuis serovar 5 reference strain, highly virulent) and SW114 (H. parasuis serovar 3 reference strain, non-virulent). A total of 191 clones were obtained from the SSH library. Using dot hybridization and PCR, 15 clones were identified containing fragments that were present in the Nagasaki genome while absent in the SW114 genome. Among these 15 fragments, three fragments (ssh1, ssh13, ssh15) encode cell surface-associated components; three fragments (ssh2, ssh5, ssh9) are associated with metabolism and stress response; one fragment (ssh8) is involved in assembly of fimbria and one fragment (ssh6) is a phage phi-105 ORF25-like protein. The remaining seven fragments are hypothetical proteins or unknown. Based on PCR analysis of the 15 serovar reference strains, eight fragments (ssh1, ssh2, ssh3, ssh6, ssh8, ssh10, ssh11 and ssh12) were found in three to five of most virulent serovars (1, 5, 10, 12, 13 and 14), zero to two in three moderately virulent serovars (2, 4 and 15), but absent in the low virulent serovar (8) and non-virulent serovars (3, 6, 7, 9 and 11). In vivo transcription fragments ssh1, ssh2, ssh8 and ssh12 were identified in total RNA samples extracted from experimental infected pig lung by RT-PCR. This study has provided some evidence of genetic differences between H. parasuis strains of different virulence.

The spatiotemporal dynamics of Tribolium castaneum (Herbst): adult flight and gene flow.

Tribolium castaneum (Herbst) has been used as a model organism to develop and test important ecological and evolutionary concepts and is also a major pest of grain and grain products globally. This beetle species is assumed to be a good colonizer of grain storages through anthropogenic movement of grain, and active dispersal by flight is considered unlikely. Studies using T. castaneum have therefore used confined or walking insects. We combine an ecological study of dispersal with an analysis of gene flow using microsatellites to investigate the spatiotemporal dynamics and adult flight of T. castaneum in an ecological landscape in eastern Australia. Flying beetles were caught in traps at grain storages and in fields at least 1 km from the nearest stored grain at regular intervals for an entire year. Significantly more beetles were trapped at storages than in fields, and almost no beetles were caught in native vegetation reserves many kilometres from the nearest stored grain. Genetic differentiation between beetles caught at storages and in fields was low, indicating that although T. castaneum is predominantly aggregated around grain storages, active dispersal takes place to the extent that significant gene flow occurs between them, mitigating founder effects and genetic drift. By combining ecological and molecular techniques, we reveal much higher levels of active dispersal through adult flight in T. castaneum than previously thought. We conclude that the implications of adult flight to previous and future studies on this model organism warrant consideration.

Evaluation of the impacts of spaying by either the dropped ovary technique or ovariectomy via flank laparotomy on the welfare of Bos indicus beef heifers and cows

The welfare outcomes for Bos indicus cattle (100 heifers and 50 cows) spayed by either the dropped ovary technique (DOT) or ovariectomy via flank laparotomy (FL) were compared with cattle subjected to physical restraint (PR), restraint by electroimmobilization in conjunction with PR (EIM), and PR and mock AI (MAI). Welfare assessment used measures of morbidity, mortality, BW change, and behavior and physiology indicative of pain and stress. One FL heifer died at d 5 from peritonitis. In the 8-h period postprocedures, plasma bound cortisol concentrations of FL, DOT, and EIM cows were not different and were greater (P < 0.05) than PR and MAI. Similarly, FL and DOT heifers had greater (P < 0.05) concentrations than PR and MAI, with EIM intermediate. Creatine kinase and aspartate aminotransferase concentrations were greater (P < 0.05) in FL and EIM heifers compared with the other treatments, with a similar pattern seen in the cows. Haptoglobin concentrations were significantly (P < 0.05) increased in the FL heifers compared with other treatments in the 8- to 24-h and 24- to 96-h periods postprocedures, and in cows were significantly (P < 0.05) increased in the FL and DOT compared with PR in the 24- to 96-h period. Behavioral responses complemented the physiological responses; standing head down was shown by more (P < 0.05) FL cows and heifers to 3 d postprocedures compared with other treatments, although there was no difference between FL and DOT heifers at the end of the day of procedures. At this same time, fewer (P < 0.05) FL and DOT heifers and cows were observed feeding compared with other treatments, although in cows there was no difference between FL, DOT, and EIM. There were no significant differences (P > 0.05) between treatments in BW changes. For both heifers and cows, FL and DOT spaying caused similar levels of acute pain, but FL had longer-lasting adverse impacts on welfare. Electroimmobilization during FL contributed to the pain and stress of the procedure. We conclude that: i) FL and DOT spaying should not be conducted without measures to manage the associated pain and stress; ii) DOT spaying is preferable to FL spaying; iii) spaying heifers is preferable to spaying cows; and iv) electroimmobilization causes pain and stress and should not be routinely used as a method of restraint.

Inheritance and relative dominance, expressed as toxicity response and delayed development, of phosphine resistance in immature stages of Rhyzopertha dominica (F.) (Coleoptera: Bostrichidae)

Fumigation of stored grain with phosphine (PH 3) is used widely to control the lesser grain borer Rhyzopertha dominica. However, development of high level resistance to phosphine in this species threatens control. Effective resistance management relies on knowledge of the expression of resistance in relation to dosage at all life stages. Therefore, we determined the mode of inheritance of phosphine resistance and strength of the resistance phenotype at each developmental stage. We achieved this by comparing mortality and developmental delay between a strongly resistant strain (R-strain), a susceptible strain (S-strain) and their F 1 progenies. Resistance was a maternally inherited, semi-dominant trait in the egg stage but was inherited as an autosomal, incompletely recessive trait in larvae and pupae. The rank order of developmental tolerance in both the sensitive and resistant strains was eggs > pupae > larvae. Comparison of published values for the response of adult R. dominica relative to our results from immature stages reveals that the adult stage of the S-strain is more sensitive to phosphine than are larvae. This situation is reversed in the R-strain as the adult stage is much more resistant to phosphine than even the most tolerant immature stage. Phosphine resistance factors at LC 50 were eggs 400×, larvae 87× and pupae 181× with respect to reference susceptible strain (S-strain) adults indicating that tolerance conferred by a particular immature stage neither strongly nor reliably interacts with the genetic resistance element. Developmental delay relative to unfumigated control insects was observed in 93% of resistant pupae, 86% of resistant larvae and 41% of resistant eggs. Increased delay in development and the toxicity response to phosphine exposure were both incompletely recessive. We show that resistance to phosphine has pleiotropic effects and that the expression of these effects varies with genotype and throughout the life history of the insect. © 2012.

Water-use efficiency and productivity trends in Australian irrigated cotton: a review

The aim of this review is to report changes in irrigated cotton water use from research projects and on-farm practice-change programs in Australia, in relation to both plant-based and irrigation engineering disciplines. At least 80% of the Australian cotton-growing area is irrigated using gravity surface-irrigation systems. This review found that, over 23 years, cotton crops utilise 6-7ML/ha of irrigation water, depending on the amount of seasonal rain received. The seasonal evapotranspiration of surface-irrigated crops averaged 729mm over this period. Over the past decade, water-use productivity by Australian cotton growers has improved by 40%. This has been achieved by both yield increases and more efficient water-management systems. The whole-farm irrigation efficiency index improved from 57% to 70%, and the crop water use index is >3kg/mm.ha, high by international standards. Yield increases over the last decade can be attributed to plant-breeding advances, the adoption of genetically modified varieties, and improved crop management. Also, there has been increased use of irrigation scheduling tools and furrow-irrigation system optimisation evaluations. This has reduced in-field deep-drainage losses. The largest loss component of the farm water balance on cotton farms is evaporation from on-farm water storages. Some farmers are changing to alternative systems such as centre pivots and lateral-move machines, and increasing numbers of these alternatives are expected. These systems can achieve considerable labour and water savings, but have significantly higher energy costs associated with water pumping and machine operation. The optimisation of interactions between water, soils, labour, carbon emissions and energy efficiency requires more research and on-farm evaluations. Standardisation of water-use efficiency measures and improved water measurement techniques for surface irrigation are important research outcomes to enable valid irrigation benchmarks to be established and compared. Water-use performance is highly variable between cotton farmers and farming fields and across regions. Therefore, site-specific measurement is important. The range in the presented datasets indicates potential for further improvement in water-use efficiency and productivity on Australian cotton farms.

Biological management of the invasive Vachellia nilotica ssp. indica (Benth.) Kyal. & Boatwr. (Fabales: Mimosoideae) in tropical Australia: stress-inducing potential of Anomalococcus indicus Ramakrishna (Insecta: Hemiptera: Coccoidea: Lecanodiaspididae), an agent of promise

Vachellia nilotica ssp. indica (hereafter, V. n. indica) is an important tree weed in Australia. Its dense populations induce undesirable changes in the vast areas of northern Australia. Because chemical and mechanical management options appear unviable for various reasons, biological management of this tree is considered a better option. Among the many trialled arthropods in Australian context, Anomalococcus indicus, a lecanodiaspid native to India, has been identified as a potent-candidate, since in India, its native terrain, it is the most widespread and occurs throughout the year. Severe infestations of A. indicus cause defoliation, wilting and death of branches, and occasionally the tree. Populations of A. indicus have been brought into Australia and are being tested for its host specificity under quarantine conditions. This article reports the physiological damage and stress it inflicts in the shoots of V. n. indica. Younger-nymphal instars of A. indicus feed on cortical-parenchyma cells of young stems, whereas the older instars and adults feed from the phloem of old stems. Two conspicuous responses of V. n. indica arising in response to the feeding action of A. indicus are changes in the cell-wall dynamics and irregular cell divisions. The feeding action of A. indicus elicits a sequence of reactions in the stem tissues of V. n. indica such as differentiation of thick-walled elements in the outer cortical parenchyma, differential thickening of cells with supernumerary layers of either suberin or lignin, proliferations of parenchyma and phloem, wall thickening and obliteration of inner lumen of phloem cells, and the sieve plates plugged with callosic deposits. The responses are the culminations of interaction between the virulence factor (one or more of the salivary proteins?) from A. indicus and the resistance factor in V. n. indica. We have analysed structural changes in the context of their functions, by comparing the feeding action of A. indicus with that of other hemipteroids. From the level of stress it induces, this study confirms that A. indicus has the potential to be an effective biological management of V. n. indica in Australia. © 2014 © 2014 Taylor & Francis and Aboricultural Association.

Prevalence and potential fitness cost of weak phosphine resistance in Tribolium castaneum (Herbst) in eastern Australia

The prevalence of resistance to phosphine in the rust-red flour beetle, Tribolium castaneum, from eastern Australia was investigated, as well as the potential fitness cost of this type of resistance. Discriminating dose tests on 115 population samples collected from farms from 2006 to 2010 showed that populations containing insects with the weakly resistant phenotype are common in eastern Australia (65.2 of samples), although the frequency of resistant phenotypes within samples was typically low (median of 2.3). The population cage approach was used to investigate the possibility that carrying the alleles for weak resistance incurs a fitness cost. Hybridized populations were initiated using a resistant strain and either of two different susceptible strains. There was no evidence of a fitness cost based on the frequency of susceptible phenotypes in hybridized populations that were reared for seven generations without exposure to phosphine. This suggests that resistant alleles will tend to persist in field populations that have undergone selection even if selection pressure is removed. The prevalence of resistance is a warning that this species has been subject to considerable selection pressure and that effective resistance management practices are needed to address this problem. The resistance prevalence data also provide a basis against which to measure management success.