From laboratory to point of entry: development and implementation of a loop‐mediated isothermal amplification (LAMP)‐based genetic identification system to prevent introduction of quarantine insect species

BACKGROUND

Rapid genetic on‐site identification methods at points of entry, such as seaports and airports, have the potential to become important tools to prevent the introduction and spread of economically harmful pest species that are unintentionally transported by the global trade of plant commodities. This paper reports the development and evaluation of a loop‐mediated isothermal amplification (LAMP)‐based identification system to prevent introduction of the three most frequently encountered regulated quarantine insect species groups at Swiss borders, Bemisia tabaci, Thrips palmi and several regulated fruit flies of the genera Bactrocera and Zeugodacus.

RESULTS

The LAMP primers were designed to target a fragment of the mitochondrial cytochrome c oxidase subunit I gene and were generated based on publicly available DNA sequences. Laboratory evaluations analysing 282 insect specimens suspected to be quarantine organisms revealed an overall test efficiency of 99%. Additional on‐site evaluation at a point of entry using 37 specimens performed by plant health inspectors with minimal laboratory training resulted in an overall test efficiency of 95%. During both evaluation rounds, there were no false‐positives and the observed false‐negatives were attributable to human‐induced manipulation errors. To overcome the possibility of accidental introduction of pests as a result of rare false‐negative results, samples yielding negative results in the LAMP method were also subjected to DNA barcoding.

CONCLUSION

Our LAMP assays reliably differentiated between the tested regulated and non‐regulated insect species within <1 h. Hence, LAMP assays represent suitable tools for rapid on‐site identification of harmful pests, which might facilitate an accelerated import control process for plant commodities. © 2018 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Effect of glutamic acid content in the diet on the catabolic rate of the isotope-labeled glutamic acid in rats. 4. Measurement of 14C-glutamic acid oxidation to 14CO2 with various energy sources in the diet

48 male rats (body weight 80-100 g) were fed with 2 diets different in the glutamic acid content (diet I 2.42 and diet II 6.24% glutamic acid in the dry matter). The mixture of the other synthetic L-amino acids was adapted to the egg protein pattern corresponding 10% crude protein in the diet. Each diet was fed either on 73% or 98 to 104% of the energy maintenance requirement. After 7 days of experimental feeding 14C-U-L-glutamic acid was given to each group by intragastric infusion (i.g.), intraperitoneal (i.p.) or subcutaneous injection (s.c.), respectively, followed by a measurement of the CO2-and 14CO2-excretion during two subsequent periods of 3 hours. The CO2-excretion was lower in animals with restricted energy intake especially during the first 3 hour-period, which was started 2 hours after feed intake. The relative 14CO2-excretion (percent of the dose) was neither significantly influenced by the level of energy intake nor by the amount of dietary glutamic acid. The highest degradation rates of 14C-glutamic acid to 14CO2 were measured after i.g. application (more than 50%), followed by the i.p. injection (nearly 50%) and the lowest values were observed after s.c. injection (about 40%). These differences were only evident during the first CO2-absorption period. Furthermore the s.c. injection caused a lower specific radioactivity of CO2 compared with the data after i.g. and i.p. application. The results suggest the high metabolic activity of the intestinal tissue for glutamic acid.     

Utilization of Animal-Protein and Plant-Protein Supplements by Red Swamp Crayfish Procambarus clarkii Fed Formulated Diets1

A study was conducted to evaluate alternative protein supplements that could be used to reduce the cost of formulated crayfish diets. Red swamp crayfish Procambarus clarkii were held in the laboratory for 9 wk and fed 30% crude protein diets containing isonitrogenous mixtures of plant and animal protein in a 65:35 ratio. Combinations tested were: soybean meal/menhaden fish meal (SOY/FSH); cottonseed meal/menhaden fish meal (COT/FSH); soybean meal/meat and bone meal (SOY/MB); cottonseed mealheat and bone meal (COT/MB); soybean meal/meat and bone meal/ blood meal (SOYIMB-B); and cottonseed meal/meat and bone meal/blood meal (COTIMB-B). Comparison of crayfish weight gain, feed efficiency ratio, apparent net protein and energy retention, and body composition indicated that SOY/FSH was the best protein combination tested. Weight gain was reduced when cottonseed meal replaced soybean meal in diets that contained either fish meal or meat and bone meal. Feed consumption of crayfish fed SOY/MB-B and COT/MB-B was lower than that of crayfish fed other diets containing the same plant-protein supplement and weight gain was lower in crayfish fed blood meal in all but one case. Differences in amino acid composition and amino acid availability of protein supplements, inhibitory effects of gossypol in cottonseed meal, and reduced consumption of diets containing blood meal could have contributed to diet effects. Results suggest that soybean meal is a better dietary protein source for crayfish than cottonseed meal, menhaden fish meal is better than meat and bone meal, and both fish meal and meat and bone meal are superior to a 60:40 meat and bone meal/blood meal mixture. However, cottonseed meal, meat and bone meal, and meat and bone meal/blood meal mixtures could be useful as lower-cost alternatives to soybean meal and fish meal in diets for pond-raised crayfish, since the apparent lower protein (amino acid) quality of these ingredients would be unlikely to depress growth of crayfish that have access to natural food in ponds.     

Toxicity of the chitin synthesis inhibitors,diflubenzuron and its dichloro-analogue,to Spodoptera littoralis larvae

The activities of the chitin synthesis inhibitors, diflubenzuron and PH 60–38, against Spodoptera littoralis larvae were assayed by feeding treated alfalfa or poisoned wheat bran baits, by allowing the larvae to imbibe sucrose-containing aqueous dispersions of the compounds, and by injection into larvae. PH 60–38 was less active than diflubenzuron. On alfalfa, diflubenzuron had to be fed for at least 2 days to prevent formation of normal pupae and emergence of adults. For very big (480–540 mg) larvae, feeding diflubenzuron at concentrations of 50 mg/litre for 2 days or 2.5 mg/litre for 3 days prevented adult emergence. For 200–250 mg larvae, this was achieved by feeding concentrations of 100 mg/litre for 2 days, 5 mg/litre for 3 days or 3.5 mg/litre for 4 days. In all larvae > 150 mg, mortality in feeding experiments occurred in the prepupal or the pupal stage. Only with 30–50 mg and 100–150 mg larvae was there considerable mortality during moults between larval instars, the larvae being unable to liberate themselves from the old larval skins and head capsules. Diflubenzuron incorporated into wheat bran baits at concentrations of from 2.5 to 10 000 μg/g killed approximately 70–90% of the insects. When imbibed, diflubenzuron was much less toxic as a wettable powder than as a liquid formulation but the two formulations were equitoxic when injected into the larvae.     

Conservation of animal genetic resources — A reviewLa conservation des resources génétiques animalesErhaltunf genetischer tierressourcen

The success in breeding highly productive farm animals has resulted in a situation where animal breeders and other people consider it necessary to preserve less productive breeds. Arguments for conservation cover emotional, cultural and scientific reasons, as well as the concern of losing genetic material which could be valuable for future animal production. Costs and time require priorities for objectives, species and forms of conservation. International co-operation in documentation, evaluation, and conservation of genetic material seems necessary to ensure efficiency of actions.     

Effect of bone diameter and eccentric loading on fatigue life of cortical screws used with interlocking nails

OBJECTIVE: To test the effects of bone diameter and eccentric loading on fatigue life of 2.7-mm-diameter cortical bone screws used for locking a 6-mm-diameter interlocking nail. SAMPLE POPULATION: Eighteen 2.7-mm-diameter cortical bone screws. PROCEDURE: A simulated bone model with aluminum tubing and a 6-mm-diameter interlocking nail was used to load screws in cyclic 3-point bending. Group 1 included 6 screws that were centrally loaded within 19-mm-diameter aluminum tubing. Group 2 included 6 screws that were centrally loaded within 31.8-mm-diameter aluminum tubing. Group 3 included 6 screws that were eccentrically loaded (5.5 mm from center) within 31.8-mm-diameter aluminum tubing. The number of cycles until screw failure and the mode of failure were recorded. RESULTS: An increase in the diameter of the aluminum tubing from 19 to 31.8 mm resulted in a significant decrease in the number of cycles to failure (mean +/- SD, 761,215 +/- 239,853 to 16,941 +/- 2,829 cycles, respectively). Within 31.8-mm tubing, the number of cycles of failure of eccentrically loaded screws (43,068 +/- 14,073 cycles) was significantly greater than that of centrally loaded screws (16,941 +/- 2,829 cycles). CONCLUSIONS AND CLINICAL RELEVANCE: Within a bone, locking screws are subjected to different loading conditions depending on location (diaphyseal vs metaphyseal). The fatigue life of a locking screw centrally loaded in the metaphyseal region of bone may be shorter than in the diaphysis. Eccentric loading of the locking screw in the metaphysis may help to improve its fatigue life.