Evaluation of chemical treatments to mitigate or eradicate Gnathia maxillaris infestations

The eradication of the haematophagous isopod ectoparasite Gnathia maxillaris was not achieved after several years of physical elimination of larvae by filtration and chemical treatment with trichlorfon in a captive marine fish population in exhibition aquaria. In this study, different in vitro laboratory assays were performed to find effective alternative anti‐parasitic treatments to trichlorfon (lufenuron, emamectin benzoate, cypermethrin and abamectin). The lethal concentration 50 (LC50) values at 96 hr were calculated for each compound using both larval stages (zuphea and praniza); morphological deformities generated throughout the life cycle, reduction in egg laying and survival of adult forms were also determined. Abamectin, cypermethrin and emamectin benzoate proved effective at limiting further Gnathia maxillaris development, but their efficacy at eradicating an infestation in large tanks was not supported by laboratory assays. However, the use of lufenuron proved to be a good substitute for trichlorfon when treating this type of infestation in large‐volume tanks (LC50 55.8 mg m^?3 for praniza). Routine lufenuron treatment once a month via an oral dose at 10 mg kg^?1 body weight eradicated G. maxillaris from the exhibition aquaria.     

Notes on the long‐term transport of the scalloped hammerhead shark (Sphyrna lewini)

The capture and transport of scalloped hammerhead sharks (Sphyrna lewini Griffith and Smith, 1834) historically has represented a difficult, expensive, and uncertain undertaking for the public aquarium community. In this study, techniques were developed to improve the successful long‐term transport of S. lewini by mitigating some of the deleterious effects associated with hyperactivity and impaired swimming patterns. The relationship between the transport vessel size and shark sizes, numbers, and swimming behavior was considered when formulating the transport regime. By balancing these factors and adopting a comprehensive water treatment method, it was possible to extend the duration of a successful transport by up to 60 hr. Implications for the future transport of S. lewini and other free‐swimming sharks are discussed. Zoo Biol 21:243–251, 2002. © 2002 Wiley‐Liss, Inc.     

Environmental DNA analysis shows high potential as a tool for estimating intraspecific genetic diversity in a wild fish population

Environmental DNA (eDNA) analysis has recently been used as a new tool for estimating intraspecific diversity. However, whether known haplotypes contained in a sample can be detected correctly using eDNA‐based methods has been examined only by an aquarium experiment. Here, we tested whether the haplotypes of Ayu fish (Plecoglossus altivelis altivelis) detected in a capture survey could also be detected from an eDNA sample derived from the field that contained various haplotypes with low concentrations and foreign substances. A water sample and Ayu specimens collected from a river on the same day were analysed by eDNA analysis and Sanger sequencing, respectively. The 10 L water sample was divided into 20 filters for each of which 15 PCR replications were performed. After high‐throughput sequencing, denoising was performed using two of the most widely used denoising packages, unoise3 and dada2 . Of the 42 haplotypes obtained from the Sanger sequencing of 96 specimens, 38 (unoise3 ) and 41 (dada2 ) haplotypes were detected by eDNA analysis. When dada2 was used, except for one haplotype, haplotypes owned by at least two specimens were detected from all the filter replications. Accordingly, although it is important to note that eDNA‐based method has some limitations and some risk of false positive and false negative, this study showed that the eDNA analysis for evaluating intraspecific genetic diversity provides comparable results for large‐scale capture‐based conventional methods. Our results suggest that eDNA‐based methods could become a more efficient survey method for investigating intraspecific genetic diversity in the field.     

On an outbreak of the citrus flat-headed borer,Agrilus auriventris E. Saunders in Nagasaki Prefecture

The process of an outbreak of the citrus flat-headed borer, Agrilus auriventris in Nagasaki Prefecture was described with some ecological considerations on causative factors. This outbreak which showed a typical “Gradation”, initiated around 1958, and terminated in 1963. The peak year was seen in 1961. At first attack by the borer was restricted to old and weak trees, but it was also found in young and healthy trees in the peak year. Some environmental conditions in orchards were examined in relation to the intensity of infestation by the borer. Soil management and drought seemed to have remarkable correlations with the borer's infestations. Probably, this outbreak was caused by the weakening of citrus tree by gummosis and abnormal defoliation, aging of orchard trees, and drought in the summer of 1959∼1960. Interrelation between these causes and processes are shown schematically in Fig. 12.     

INTRA‐CELLULAR AND EXTRA‐CELLULAR TOXINS IN THE RED TIDE DINOFLAGELLATE,GYMNODINIUM BREVE

The marine dinoflagellate, Gymnodinium breve (Davis), produces several neurotoxins that cause neurotoxic shellfish poisoning (nsp), massive fish kills and respiratory irritation in marine mammals and humans. The common method for discerning toxic levels of G. breve for public health advisories is enumeration of live cells in a given water mass. In this study, laboratory cultures, as well as natural blooms, were added to a stirred ultra‐filtration cell concentrator to separate viable cells containing intra‐cell toxins from ambient water containing extra‐cell toxins. Methods were validated using various mixtures of lysed and whole G. breve laboratory culture. Extractions and recovery of brevetoxins were done using a C‐18 bonded‐phase glass fiber extraction disc eluted with methanol. Total PbTx toxin concentrations were quantified by HPLC/UV using a C‐18 column and an 85:15 methanol:water (1 ml min^?1) isocratic elution at 215 nm. This method of separation and extraction was subsequently applied to water samples collected during natural blooms along two different areas of the Florida Gulf coast. The results indicated that early stages of G. breve blooms contained primarily intra‐cell toxins with extra‐cell toxins increasing as the bloom progressed, even though very few viable G. breve cells were present. This suggests that enumeration of cells alone may be insufficient and additional toxin quantitation is necessary.     

Spatio‐temporal dynamics of an Ips acuminatus outbreak and implications for management

• 1 Understanding spatio‐temporal processes of bark beetle infestations is crucial for predicting beetle behaviour and aiding management decisions aiming to prevent or mitigate tree mortality. We recorded the spatial and temporal distribution of killed trees during the 5‐year period of an Ips acuminatus outbreak.
• 2 Killed trees were always grouped in well‐defined patches (infestation spots). In years of high population density, infestation spots were large and aggregated, whereas, in years of low density, infestation spots were small and weakly aggregated or randomly distributed within the study area.
• 3 Most trees were killed in the spring by beetles that had hibernated but, in some years, trees were also killed in the summer by new‐generation beetles originating from spring attacks. Spring‐killed trees always formed new infestation spots at new locations (i.e. spot proliferation). By contrast, summer‐killed trees always occurred at the edge of active spots established in the spring, thus resulting in spot growth.
• 4 With regard to management strategies, the results obtained in the present study suggest that areas located in close proximity to infestations of the previous year should be prioritized for risk assessment. Because large spots account for most of the observed tree mortality, the cut‐and‐remove method should be focused on these spots as soon as crown discoloration appears in the summer. If applied timely, this strategy will remove the new‐generation beetles originating from the spring attacks before they emerge and also reduce the risk of spot growth.

     

What explains landscape patterns of tree mortality caused by bark beetle outbreaks in Greater Yellowstone?

Aim Bark beetle outbreaks have recently affected extensive areas of western North American forests, and factors explaining landscape patterns of tree mortality are poorly understood. The objective of this study was to determine the relative importance of stand structure, topography, soil characteristics, landscape context (the characteristics of the landscape surrounding the focal stand) and beetle pressure (the abundance of local beetle population eruptions around the focal stand a few years before the outbreak) to explain landscape patterns of tree mortality during outbreaks of three species: the mountain pine beetle, which attacks lodgepole pine and whitebark pine; the spruce beetle, which feeds on Engelmann spruce; and the Douglas‐fir beetle, which attacks Douglas‐fir. A second objective was to identify common variables that explain tree mortality among beetle–tree host pairings during outbreaks. Location Greater Yellowstone ecosystem, Wyoming, USA. Methods We used field surveys to quantify stand structure, soil characteristics and topography at the plot level in susceptible stands of each forest type showing different severities of infestation (0–98% mortality; n= 129 plots). We then used forest cover and beetle infestation maps derived from remote sensing to develop landscape context and beetle pressure metrics at different spatial scales. Plot‐level and landscape‐level variables were used to explain outbreak severity. Results Engelmann spruce and Douglas‐fir mortality were best predicted using landscape‐level variables alone. Lodgepole pine mortality was best predicted by both landscape‐level and plot‐level variables. Whitebark pine mortality was best – although poorly – predicted by plot‐level variables. Models including landscape context and beetle pressure were much better at predicting outbreak severity than models that only included plot‐level measures, except for whitebark pine. Main conclusions Landscape‐level variables, particularly beetle pressure, were the most consistent predictors of subsequent outbreak severity within susceptible stands of all four host species. These results may help forest managers identify vulnerable locations during ongoing outbreaks.     

Snail responses to cues produced by an invasive decapod predator

Abstract. Cues released by predators and injured prey often induce shifts in prey behavior that allow prey to evade predators, but also affect prey resource use. I investigated the effects of chemical and mechanical signals produced by injured snails (Physella gyrina) and predatory crayfish (Procambarus clarkii) on microdistributions of P. gyrina. In an initial experiment, I observed snail responses to the presence of a caged crayfish predator, to injured conspecifics, or to both. There were significant effects of time and the treatment × time interaction on the proportion of snails moving above the water line, with greater proportions of snails above the water line at night than during the day and with weak snail crawl‐out behavior being elicited by caged crayfish at night, but not during the day. In a second experiment, I examined snail microdistributions when exposed to crayfish confined to a small cage within each aquarium, crayfish confined to half of each aquarium, and crayfish ranging freely throughout each aquarium. Snails responded most strongly to free‐ranging crayfish by moving above the water line, but also demonstrated significant, but reduced, crawl‐out responses to crayfish confined to half of each aquarium; however, snails did not respond behaviorally to crayfish confined to small cages. In both experiments, there were marginally significant effects of unfed caged crayfish on the proportions of snail populations hiding under benthic shelters, with this response being the strongest at the start of the experiments but weak overall (with only 4–5% of P. gyrina responding in each experiment). These results indicate that cues (e.g., chemical or mechanical) produced by predators altered prey microdistributions, but that the exact prey responses (e.g., moving above the water line or into horizontal or benthic refugia) depended on the intensity and nature of cues.     

The Effect of Larch Casebearer on Larch Growth and Reproductive Organ Formation

Dependencies of larch (Larix sibirica Ledeb.) growth and reproductive organ formation on its infestation by a phyllophagous miner, larch casebearer (Coleophora sibiricella Flkv., Lepidoptera, Coleophoridae) were studied. The work was performed in the sustained outbreak of the larch casebearer in the forest-steppe-larch forest at Kuznetsk Alatau. For the representative tree groups, we determined shoot linear growth, the number of axillary buds, trunk radial growth, the number of micro- and macrostrobili produced, and the contents of water, monoses, starch, and nitrogenous compounds in needles as dependent on the infestation density. We found that infestation exceeding 0.5 larvae per brachyblast significantly suppressed tree growth and reproductive organ formation. At this threshold value of infestation, a significant inhibition of shoot linear growth can serve a marker for suppressed trunk radial growth and reproductive organ formation. The lower degree of infestation did not suppress larch growth and even stimulated it. This observation, along with our data on the changes in the composition of nitrogenous and carbohydrate compounds in damaged needles, demonstrates the active character of interaction between the host tree and the phyllophagous insect, a transit of the source function from damaged needles to healthy ones, and the stimulated reparation of the assimilation apparatus.     

Habitat preferences of the burbot (Lota lota) from the River Elbe: an experimental approach

An experimental approach was used to analyze preferences of the burbot (Lota lota) with regard to water depth, substrate type and flow velocity. In total, 30 burbots used in the experiments were caught in the middle reaches of the River Elbe by electro‐fishing. Immediately after capture, they were placed in a 180 L transportation box with aerated freshwater and taken to the research aquarium of the Zoological Museum Hamburg where they were divided into two groups according to total length: group I, 10.0–16.5 cm; group II, 20.0–30.0 cm. Prior to the experiments the fish were adapted to laboratory conditions for four weeks at 20°C in four tanks, each containing 720 L water. For each experiment, three L. lota were transferred to an additional 1350 L experimental tank. After a one‐day adaptation phase each experiment ran for three continuous day and night phases. Burbot preferences were tested for (i) water depth, (ii) substrate size, (iii) water depth and substrate size combined, and (iv) water depth, substrate and flow velocity in combination. Diurnal and nocturnal activities were recorded and analyzed throughout all experiments. Burbots of both length groups showed a significant preference for deeper water depths and large cobble substrate. Substrate type was found to be the dominant factor determining habitat preferences of L. lota when investigated in combination with water depth. In experiments combined with flow velocity, substrate and water depth were the most important factors determining burbot habitat preferences, and flow velocity of minor importance. In general, burbots were most active at night, while daytime activity was much lower. However, in all experimental approaches in the smaller length group the burbots displayed much higher day‐ and night‐time activity than did the individuals in the larger length group.     

Classification of Prosopis juliflora invasion in the Lake Baringo basin and environmental correlations

The spread of Prosopis juliflora in the Baringo basin, Kenya, has led to severe changes in the ecosystem with negative socio‐economic impacts. The drivers that foster the invasiveness of Prosopis are not fully understood. Thus, a method to quantify the degree of infestation will support the determination of environmental preferences and the risk assessment of future Prosopis invasion. We developed a methodology for characterising and classifying degrees of Prosopis infestation in vegetation stands and propose its application in environmental correlation models. The relative cover was identified as the most suited attribute for assessing and monitoring the invasion of Prosopis. The distance of invaded stands from original plantations and environmental attributes related to water availability (ground water table, rainfall and soil water–holding capacity) have potential to predict potential or future invasion risks.     

Rainbow trout resistance to bacterial cold water disease: two new quantitative trait loci identified after a natural disease outbreak on a French farm

In rainbow trout farming, Flavobacterium psychrophilum, the causative agent of bacterial cold water disease, is responsible for important economic losses. Resistance to F. psychrophilum is heritable, and several quantitative trait loci (QTL) with moderate effects have been detected, opening up promising perspectives for the genetic improvement of resistance. In most studies however, resistance to F. psychrophilum was assessed in experimental infectious challenges using injection as the infection route, which is not representative of natural infection. Indeed, injection bypasses external barriers, such as mucus and skin, that likely play a protective role against the infection. In this study, we aimed at describing the genetic architecture of the resistance to F. psychrophilum after a natural disease outbreak. In a 2000‐fish cohort, reared on a French farm, 720 fish were sampled and genotyped using the medium‐throughput Axiom? Trout Genotyping Array. Overall mortality at the end of the outbreak was 25%. Genome‐wide association studies were performed under two different models for time to death measured on 706 fish with validated genotypes for 30 060 SNPs. This study confirms the polygenic inheritance of resistance to F. psychrophilum with a few QTL with moderate effects and a large polygenic background, the heritability of the trait being estimated at 0.34. Two new chromosome‐wide significant QTL and three suggestive QTL were detected, each of them explaining between 1% and 4% of genetic variance.     

Bioturbation and the role of microniches for sulfate reduction in coastal marine sediments

The effects of bioturbation in marine sediments are mainly associated with an increase in oxic and oxidized zones through an influx of oxygen‐rich water deeper into the sediment and the rapid transport of particles between oxic and anoxic conditions. However, macrofaunal activity also can increase the occurrence of reduced microniches and anaerobic processes, such as sulfate reduction. Our goal was to determine the two‐dimensional distribution of microniches associated with burrows of a ghost shrimp (Neotrypaea californiensis) and to determine microbial activities. In laboratory experiments, detailed measurements of sulfate reduction rates (SRR) were measured by injecting, in a 1 cm grid, radiolabelled sulfate directly into a narrow aquarium (40 cm × 30 cm × 3 cm) containing the complex burrow of an actively burrowing shrimp. Light‐coloured oxidized burrow walls, along with black reduced microniches, were clearly visible through the aquarium walls. Direct injection of radiotracers allowed for whole‐aquarium incubation to obtain two‐dimensional documentation of sulfate reduction. Results indicated SRR were up to three orders of magnitude higher (140–790 nmol SO₄^2? cm^?3 day^?1) in reduced microniches associated with burrows when compared with the surrounding sediment. Additionally, some of the subsurface sulfate‐reducing microniches associated with the burrow system appeared to be zones of dinitrogen fixation. Bioturbation may also lead to decreased sulfate reduction in other microniches and the sum of the activity in all microniches might not result in a total increase of sulfate reduction compared with non‐bioturbated control sediments.     

Rapid detection of <Emphasis Type="Italic">Salmonella typhi</Emphasis> in foods by combination of immunomagnetic separation and polymerase chain reaction

Summary A combination of immunomagnetic separation and polymerase chain reaction (IMS-PCR) was used to detect Salmonella typhi in food and water samples. IMS was found to be an effective method for specific capture of S. typhi from artificially inoculated meat rinse samples. The bacteria could be detected within 6 h by IMS-PCR with a sensitivity of 10⁵ cells. However, when tested in milk samples, the method was less effective. In comparison to conventional culture method, IMS-PCR is a rapid and specific method for detection of S. typhi and could be useful in outbreak situations for tracing the source of infection.     

Effect of black bean aphid, Aphis fabae, on transpiration, stomatal conductance and crude protein content of faba bean

This study was initiated to investigate effects of damage by 0, 5 and 10 aphids/plant on the physiology of faba bean plants throughout different feeding periods and at two plant development stages. Immediately following removal of Aphis fabae, measurements showed 84–229% increase in transpiration rate. These changes were proportional to the number of aphids and infestation duration. Injury by A. fabae caused the stomatal conductance to be much higher in the leaves of infested plants. Leaf stomatal conductance of the infested plants increased significantly by 51–224% depending on initial aphid densities and feeding intervals. This increase was proportional to the infestation level for each date. Length of infestation period and plant growth stage seemed to have no clear effect on stomatal apertures. Aphid feeding caused a damage of about 7–33% of crude protein levels in the leaf tissue. This reduction increased with increasing infestation levels and time, except for 28‐day‐old plants on 28 days. The physiological effects of aphid feeding on water vapour and chemical composition of damaged leaves are particularly serious when the population is high.     

Method for estimating density of Cryphalus piceae (Ratz.) brood galleries using a regression model

Estimation of the severity of infestation by the bark beetle (Cryphalus piceae Ratz.) on silver fir (Abies alba Mill.) is commonly done using methods that lack satisfactory statistical foundations. The object of this study was to develop a statistical method for estimating stem infestation density which permits calculation of the estimation error and which also does not require debarking of the entire stem. The study was carried out in the near‐natural forests of the ?wi?tokrzyski National Park in central Poland. Two general patterns (A and B) of distribution of C. piceae brood galleries on stems of A. alba were observed. In pattern A, brood galleries occurred on the 1st 1‐m length of stem (measured from the base), and the probability of infestation on successive 1‐m lengths further up the tree was also very high (above 90%). In pattern B, brood galleries were absent from the 1st 1‐m length but the probability of infestation in successive 1‐m lengths increased as the distance from the base increased. In a statistical analysis of linear correlations between the numbers of C. piceae brood galleries in selected 1‐m‐long stem lengths and the total density of infestation, the most significant correlations for pattern A were found for the 1st, 2nd, 6th and 7th 1‐m lengths (counting from the base). Here, mean relative errors of estimation did not exceed 35%. For pattern B, the most significant correlations were found for the 8th, 10th, 11th, 12th, and 13th 1‐m lengths, where mean relative errors of estimation did not exceed 42%. The method described is only mildly invasive and therefore may be used in strictly protected areas of reserves and national parks.     

Comparison of enumeration and Jolly-Seber estimation of population size in the common voleMicrotus arvalis

Capture-recapture data on common volesMicrotus arvalis (Pallas, 1779) in central Europe have been almost exclusively analysed by means of the enumeration technique (minimum number alive or calendar of catches). Here we compare enumeration and Jolly-Seber (JS) estimation of population size in the common vole using live-trapping data from an alfalfa field-population in southern Moravia, Czech Republic. Over the entire study the enumeration estimate of the population size was smaller by an average of 28% than the JS estimate. The negative bias increased with density, decreased with both capture probability and the survival rate, and was more pronounced in males at high density. We conclude that the method of direct enumeration is not reliable for estimating population size in the common vole.     

Comparison of culture-based methods to enumerate Escherichia coli in tropical and temperate freshwaters

Aims: The specificity of a method for the enumeration of Escherichia coli (chromocult agar, CC) was tested using freshwater samples from a tropical area (Cuba Island) by isolating colonies and identifying them with API (Appareillage et procédé d’identification) strips. Enumerations of E. coli by the most probable number (MPN) microplate method were compared with counts on chromogenic and fluorogenic agar media [CC, rapid E. coli (REC), fluorocult] in tropical and temperate freshwater samples. Methods and Results: A high percentage of specificity (95·7%) for the CC agar enumeration of E. coli was observed. High regression coefficients (log‐log linear regressions) were found between E. coli counts on agar media and by the MPN method. In the tropical environment, counts with REC medium were significantly different from those obtained with the other methods. MPN counts were found to be significantly higher than those obtained using the plate counts methods in the temperate environment. Conclusions: Escherichia coli enumeration methods based on glucuronidase activity appear to be suitable for the evaluation of microbiological quality in the tropical environment featured in this study. Significance and Impact of the Study: The methods for the enumeration of E. coli tested in this study should help improve the evaluation of microbiological contamination of Cuban freshwaters.     

Farm‐scale dispersal of Bactericera cockerelli in potato crops measured using Bt mark‐capture techniques

Natural populations of Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), also known as tomato/potato psyllid, were marked in potato [Solanum tuberosum L. (Solanaceae)] crops using Bacillus thuringiensis Berliner (Bt) to investigate the impact of dispersal on crop infestation and management of potential insecticide resistance in New Zealand. The technique was adapted from previous studies that used conventional spray applications of Bt to mark Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae), and identified marked individuals with selective microbiological assays and identification of characteristic crystal inclusions. Initially, marking rates of B. cockerelli were improved by using ultra‐low volume applications of undiluted Bt, but this result was not consistent. Several other pests and natural enemies were also marked. In mark‐capture studies, marked B. cockerelli were captured over 3 days on yellow sticky traps in small trap plots of potatoes at 60, 120, 180, 250, and 350 m from the sprayed crop. Bactericera cockerelli flight activity occurred throughout daylight hours with evidence of bimodal diurnal peaks. Significantly greater numbers of B. cockerelli were captured in downwind traps. The combined dispersal curve derived from two mark‐capture experiments estimated a mean dispersal distance for B. cockerelli of 100 m in 3 days and indicated that 10% of the population dispersed further than ca. 250 m. Over the period of a growing season, this level of dispersal suggests that B. cockerelli can disperse throughout a vegetable‐growing region, with implications for crop infestation and management of potential insecticide resistance.     

Inter‐laboratory validation of a rapid assay for the detection and quantification of Legionella spp. in water samples

Aims: To compare the standard culture method with a new, rapid test (ScanVIT‐Legionella?) using fluorescently labelled gene probes for the detection and enumeration of Legionella spp. The new technique was validated through experiments conducted on both artificially and naturally contaminated water and through an inter‐laboratory comparison. Methods and Results: All samples were processed by the ScanVIT test according to the manufacturer’s instructions and by a culture method (ISO 11731). ScanVIT detected significantly more positive samples, although concentrations were similar and a strong positive correlation between the two methods was observed (r = 0·888, P < 0·001). The new test was more accurate in identifying the co‐presence of Legionella pneumophila and Leg. non‐pneumophila. ScanVIT showed a slightly higher Legionella recovery from water samples artificially contaminated with Leg. pneumophila alone or together with Pseudomonas aeruginosa. Lastly, the inter‐laboratory comparison revealed that the ScanVIT test exhibits a lower variability than the traditional culture test (mean coefficient of variation 8·7 vs 16·1%). Conclusions: The results confirmed that the ScanVIT largely overlaps the reference method and offers advantages in terms of sensitivity, quantitative reliability and reduced assay time. Significance and Impact of the Study: The proposed method may represent a useful validated alternative to traditional culture for the rapid detection and quantification of Legionella spp. in water.