Aspergillus on tree nuts: incidence and associations

California exports tree nuts to countries where they face stringent standards for aflatoxin contamination. Trade concerns have stimulated efforts to eliminate aflatoxins and Aspergillus flavus from almonds, pistachios and walnuts. Incidence of fungi on tree nuts and associations among fungi on tree nuts were studied. Eleven hundred pistachios, almonds, walnuts and brazil nuts without visible insect damage were plated on salt agar and observed for growth of fungi. Samples came both from California nut orchards and from supermarkets. To distinguish internal fungal colonization of nuts from superficial colonization, half the nuts were surface-sterilized before plating. The most common genera found were Aspergillus , Rhizopus and Penicillium . Each species of nut had a distinct mycoflora. Populations of most fungi were reduced by surface sterilization in all except brazil nuts, suggesting that they were present as superficial inoculum on (rather than in) the nuts. In general, strongly positive associations were observed among species of Aspergillus ; nuts infected by one species were likely to be colonized by other species as well. Presence of Penicillium was negatively associated with A. niger and Rhizopus in some cases. Results suggest that harvest or postharvest handling has a major influence on nut mycoflora, and that nuts with fungi are usually colonized by several fungi rather than by single species. This revised version was published online in June 2006 with corrections to the Cover Date.     

Impact of processing conditions on the phytoprostanes profile of three types of nut kernels

The goal of this work was the identification and quantification of phytoprostanes (PhytoPs) in three types of nuts: “Walnut”, “Macadamia”, and “Pecan”. This study represents a first approach to the relationship between the quantitative and qualitative PhytoP profiles in the “Macadamia” and “Pecan” nuts subjected to fried salt or fried honey processing. The kernels were found to contain 9-F_1t-PhytoP, 9-epi-9-F_1t-PhytoP, 16-B₁-PhytoP, ent-16-B₁-PhytoP, 9-L₁-PhytoP, and ent-9-L₁-PhytoP. “Macadamia” fried salt nuts were the only ones that produced 9-epi-9-D₁-PhytoP and 9-D₁-PhytoP. The total PhytoP concentration in raw nuts was in the range of 5541–7830?ng kg^?1 fresh weight (FW); for most of the PhytoPs, the concentrations were lowest in raw walnuts, indicating that concentration of each PhytoP was influenced by the genotype. The frying process increased the total PhytoPs concentration to the range of 8903–33,727?ng kg^?1 FW. Therefore, this is the first work describing PhytoPs in nuts and reinforces the capacity of these compounds to act as biomarkers to monitor the processing treatments that influence the final quality of nuts.     

Toxic aspergilli from pistachio nuts

Pistachio nut samples taken during various stages of development from orchards in Iran, showed that contamination with fungi occurred mainly during the later stages of nut development. Members of the genera Aspergillus and Penicillium occurred most frequently. Of the Aspergilli, the species A. niger, A. flavus and A. fischeri var. spinosus occurred most frequently, followed by A. terreus, A. tamarii and A. nidulans. Twenty-two isolates comprising 13 species were tested for toxicity to ducklings. Isolates of known toxic fungi included A. flavus, A. niger, A. parasiticus, A. ochraceus, A. versicolor, A. nidulans and A. terreus. The toxicity of A. fischeri var. spinosus is reported. Chemical analysis showed that all isolates of A. flavus and A. parasiticus produced aflatoxin B₁, the isolates of A. versicolor and A. nidulans produced sterigmatocystin while the toxic isolate of A. ochraceus did not produce ochratoxins.Toxic fungi have been shown to occur in a variety of nuts (4), (5), (11), (12), (13), (15), (18), (20), (21). Aflatoxin contamination of pistachio nuts has been reported and has in the past led to the rejection of consignments of Iranian pistachio nuts (1). In Iran, pistachio nuts are produced mainly in the south eastern provinces (Kerman & Zahedan) and to a limited extent in the Northern part (Kazvin & Damghan). In 1975 it was estimated (7) that there were some 24 million pistachio trees in Iran, of which 60% were situated in Rafsanjan, Kerman (Table 1). Economic considerations as well as the potential health hazard posed by aflatoxin-contaminated nuts, prompted the University of Isfahan to initiate a study of various aspects of the mycotoxin problem in pistachio nuts.     

Dispersal of sweet pignut hickory in a year of low fruit production,and the influence of predation by a curculionid beetle

Summary The rate at which fallen hickory nuts are removed from beneath the parent tree, and the effect on this rate of the seed predatorConotrachelus affinis, was studied in an oak-hickory forest in southeastern Michigan, USA, during a year in which few nuts were produced. The trees responded toConotrachelus, which destroyed half the nut crop, by aborting inviable nuts during the summer. The seed dispersers, mostly gray squirrels, removed fallen nuts rapidly, showing the ability to distinguish viable nuts and remove them preferentially. The number of nuts removed in a week varies directly with the number available, and removal rate increases when many viable nuts are falling. The death of most seeds before dispersal, and the squirrels' efficiency at foraging on nuts and recovering them after burial, imply that successful hickory reproduction takes place only in years of heavy nut production.     

Sooty mangabeys scavenge on nuts cracked by chimpanzees and red river hogs—An investigation of inter‐specific interactions around tropical nut trees

Carrion scavenging is a well‐studied phenomenon, but virtually nothing is known about scavenging on plant material, especially on remnants of cracked nuts. Just like meat, the insides of hard‐shelled nuts are high in energetic value, and both foods are difficult to acquire. In the Taï forest, chimpanzees (Pan troglodytes) and red river hogs (Potamochoerus porcus) crack nuts by using tools or strong jaws, respectively. In this study, previously collected non‐invasive camera trap data were used to investigate scavenging by sooty mangabeys (Cercocebus atys), two species of Guinea fowl (Agelestres meleagrides; Guttera verreauxi), and squirrels (Scrunidae spp.) on the nut remnants cracked by chimpanzees and red river hogs. We investigated how scavengers located nut remnants, by analyzing their visiting behavior in relation to known nut‐cracking events. Furthermore, since mangabeys are infrequently preyed upon by chimpanzees, we investigated whether they perceive an increase in predation risk when approaching nut remnants. In total, 190 nut‐cracking events were observed in four different areas of Taï National Park, Ivory Coast. We could confirm that mangabeys scavenged on the nuts cracked by chimpanzees and hogs and that this enabled them to access food source that would not be accessible otherwise. We furthermore found that mangabeys, but not the other species, were more likely to visit nut‐cracking sites after nut‐cracking activities than before, and discuss the potential strategies that the monkeys could have used to locate nut remnants. In addition, mangabeys showed elevated levels of vigilance at the chimpanzee nut‐cracking sites compared with other foraging sites, suggesting that they perceived elevated danger at these sites. Scavenging on remnants of cracked nuts is a hitherto understudied type of foraging behavior that could be widespread in nature and increases the complexity of community ecology in tropical rainforests.     

Impact of site‐induced mouse caching and transport behaviour on regeneration in Castanea crenata

Abstract. Dispersal and retrieval site selection by mice, transport distance, cache depth, and emergence and survival of seedlings of Castanea crenata (Japanese chestnut) were investigated by a magnet‐locating experiment in two habitat conditions (gap vs. forest understorey). Magnets were inserted into nuts (n= 450) and the nuts placed in the edge of forest gaps. Although wood mice (Apodemus speciosus and A. argenteus) initially buried nuts singly in shallow surface caches near the nut source, by the following spring these cached nuts were retrieved and re‐cached in larger, deeper caches farther from the source, particularly in forest understories, probably to reduce the threat of pilferage. All the nuts cached in the forest understories were consumed, but 4 seedlings emerged in gaps, apparently because of lower foraging activity in the gaps by the mice. Seed size was not correlated with cache depth or cache site selection. With increasing seed size, transport distance increased, particularly in gaps, possibly due to a greater potential energy gain (relative to handling cost to the cacher), or to attempts to prevent density‐ or mass‐dependent loss of caches by other foragers. Variable seed dispersal behaviour based on variation in seed size may influence the chances of colonization and distribution of the light‐demanding Castanea trees in mosaic landscapes and may play an important role in community organization and dynamics.     

Ochratoxin Production by the Aspergillus ochraceus Group and Aspergillus alliaceus

Ochratoxin A is a toxic and carcinogenic fungal secondary metabolite; its presence in foods is increasingly regulated. Various fungi are known to produce ochratoxins, but it is not known which species produce ochratoxins consistently and which species cause ochratoxin contamination of various crops. We isolated fungi in the Aspergillus ochraceus group (section Circumdati) and Aspergillus alliaceus from tree nut orchards, nuts, and figs in California. A total of 72 isolates were grown in potato dextrose broth and yeast extract-sucrose broth for 10 days at 30°C and tested for production of ochratoxin A in vitro by high-pressure liquid chromatography. Among isolates from California figs, tree nuts, and orchards, A. ochraceus and Aspergillus melleus were the most common species. No field isolates of A. ochraceus or A. melleus produced ochratoxin A above the level of detection (0.01 μg/ml). All A. alliaceus isolates produced ochratoxin A, up to 30 μg/ml. We examined 50,000 figs for fungal infections and measured ochratoxin content in figs with visible fungal colonies. Pooled figs infected with A. alliaceus contained ochratoxin A, figs infected with the A. ochraceus group had little or none, and figs infected with Penicillium had none. These results suggest that the little-known species A. alliaceus is an important ochratoxin-producing fungus in California and that it may be responsible for the ochratoxin contamination occasionally observed in figs.     

Acorn dispersal by the blue jay (Cyanocitta cristata)

Summary Blue jays transported and cached 133,000 acorns from a stand of Quercus palustris trees in Blacksburg, Virginia, representing 54% of the total mast crop. A further 20% (49,000) of the mast crop was eaten by jays at the collecting site. A large proportion of the nuts remaining beneath the collecting trees was parasitized by curculionid larvae. The number of nuts transported per caching trip ranged from 1–5 with a mean of 2.2. Mean distance between seed trees and caches was 1.1 km (range: 100 m–1.9 km). Jays appeared to choose species with small- to medium-sized nuts (Quercus palustris, Q. phellos, Q. velutina, Fagus grandifolia) and avoided the larger nuts of Q. borealis and Q. alba.Nuts were cached singly within a few meters of each other and were always covered with debris. Covering may improve germination and early growth by protecting the nut and radicle from desiccation. The vegetation structure of most suburban caching sites was analogous to open, disturbed environments in more natural landscapes. The presence of numerous Quercus seedlings in jay caching sites and the tendency for jays to cache nuts in environments conducive to germination and early growth indicate that blue jays facilitate colonization of members of the Fagaceae.     

Five-year study on the autumn food habits of the Asiatic black bear in relation to nut production

We studied the autumn food habits of the Asiatic black bear (Ursus thibetanus) from 1993 to 1997, based on 202 fecal samples in the Chichibu Mountains, central Japan. Nuts occupied the highest proportions of autumn foods (59.9–85.8% important values). Although the proportion of nuts of Quercus crispula, Fagus crenata and Fagus japonica varied greatly between the years, acorns of Q.crispula were most prevalent in four of the five years. We also determined the relative nut production of these three species by counting the number of nuts or cupules on the ground. Black bears consumed the nuts according to their relative availabilities. Nuts of Q.crispula appeared to be the most important food because: (i) these nuts were eaten in the highest proportion in four of the five study years; and (ii) even in poor years the bears consumed acorns of Q.crispula, whereas nuts of Fagus spp. were not consumed. We discuss the significance of alternative foods for black bears in relation to food tree diversity in the forest and the necessity for long-term studies examining the food habits of Asiatic black bears.     

Molecular analysis of <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Flavi</Emphasis> isolated from Brazil nuts

Brazil nuts are an important export market in its main producing countries, including Brazil, Bolivia, and Peru. Approximately 30,000 tons of Brazil nuts are harvested each year. However, substantial nut contamination by Aspergillus section Flavi occurs with subsequent production of aflatoxins. In our study, Aspergillus section Flavi were isolated from Brazil nuts (Bertholletia excelsa), and identified by morphological and molecular means. We obtained 241 isolates from nut samples, 41% positive for aflatoxin production. Eighty-one isolates were selected for molecular investigation. Pairwise genetic distances among isolates and phylogenetic relationships were assessed. The following Aspergillus species were identified: A. flavus, A. caelatus, A. nomius, A. tamarii, A. bombycis, and A. arachidicola. Additionally, molecular profiles indicated a high level of nucleotide variation within β-tubulin and calmodulin gene sequences associated with high genetic divergence from RAPD data. Among the 81 isolates analyzed by molecular means, three of them were phylogenetically distinct from all other isolates representing the six species of section Flavi. A putative novel species was identified based on molecular profiles.     

Inoculated Host Range and Effect of Host on Morphology and Size of Macroconidia Produced by Claviceps africana and Claviceps sorghi

Twenty graminaceous plant species were evaluated for their susceptibility to the two sorghum ergot pathogens Claviceps sorghi and Claviceps africana. Five species viz., Sorghum arundinaceum, Sorghum halepense, Sorghum versicolor, Sorghum virgatum and Pennisetum glaucum were found to become infected by both pathogens via inoculation with 10⁶ conidia/ml. Species which did not become infected under these conditions included Pennisetum pedicellatum, Zea mays, and species of Panicum, Brachiaria, Cenchrus, Andropogon, Dichanthium, Chrysopogon, Iseilema, Bothriochloa and Chloris. Honeydew secretions were observed from infected flowers of susceptible plant species. There was marked variation in size of macroconidia of both C. sorghi and C. africana on different hosts on which the pathogens were able to establish symptoms. Dimorphism was observed for macroconidia produced on P. glaucum, as elliptical and spindle shaped macroconidia were observed. Based on inoculation under greenhouse conditions, we conclude that C. sorghi and C. africana may have similar host ranges.     

Mental map in wild chimpanzees: An analysis of hammer transports for nut cracking

The mental map of wild chimpanzees is analyzed in the context of their transports of clubs and stones used for cracking two species of nuts of different hardness,Coula edulis andPanda oleosa, in the Tai National Park (Ivory Coast). For the harderPanda nuts, they transport the harder hammers, i.e., almost exclusively stones, hammers of greater weight, and the transports are longer than forCoula nuts. The analysis made for the transports forPanda nuts shows that they seem to remember the location of stones and to choose the stones so as to keep the transport distance minimal. The chimpanzees seem to possess an Euclidian space, which allows them to somehow measure and remember distances; to compare several such distances so as to choose the stone with the shortest distance to a goal tree; to correctly locate a new stone location with reference to different trees; and to change their reference point so as to measure the distance to eachPanda tree from any stone location. They also combine the weight and the distance. The wild chimpanzees of the Tai National Park seem to possess concrete operation abilities in spatial representation.     

Detection of Aflatoxin in Aspergillus Species Isolated from Pistachio in Iran

To estimate the incidence contamination of fresh pistachio nuts by aflatoxigenic fungi in Iran, nut samples were collected from pistachio orchards in Kerman, Rafsanjan and Isfahan regions. Out of the 200 Aspergillus isolates obtained, 11 species were identified as A. alliaceous, A. candidus, A. flavus, A. niger, A. niveus, A. ochraceus, A. parasiticus, A. tamari, A. terreus, A. unguis and A. wentii. For detection of aflatoxin production ability of the isolates, three target genes, namely aflR, aflJ, and omtB, used in PCR amplification. In all the examined cases, the degenerate primer designed for amplification of omtB gene, named omtBII, was able to amplify an expected 611 bp fragment in aflatoxigenic isolates in this study and yielded the same result as those obtained from TLC analysis and fluorescence ability by application of methylated β‐cyclodextrin in culture media. Using this procedure the significant incidence of aflatoxin‐producing aspergilli was confirmed in pistachio nuts produced in different regions of Iran. The results indicated that PCR method described here, in combination with fluorescence assay, is a reliable and simple confirmatory test for monitoring pistachio nuts contaminated with aflatoxinogenic aspergilli.     

Nut weight and nutrient contents ofCola acuminata andC. nitida (Sterculiaceae)

Cola acuminata andC. nitida nuts were classified into nut weight classes, and each nut weight class was divided into groups according to the number of nut cotyledons. The nuts in each group were analyzed for inorganic and organic constituents. Statistical analyses showed the effect of nut weight to be significant on nutrient composition in the two species (P = 0.001) while cotyledon number effect was not significant inC. acuminata. Germination percentage and growth performance varied directly with nutrient contents which, parenthetically, were positively and significantly correlated with the nut weights (0.77 < r < 0.99 and 0.1% <P < 5%). Relationship between nut cotyledon number and the nutrient contents was not consistent. The preferential taste forC. nitida by people as compared toC. acuminata is probably because the former contains more free sugar and is more nutritious.     

Improved walnut-feeding skills with experience in wood mice, <Emphasis Type="Italic">Apodemus speciosus</Emphasis>

The wood mouse Apodemus speciosus can consume the hard-walled walnut species Juglans ailanthifolia. The mice gnaw holes on two sides of the shell and then pick the meat from the holes. However, not all mice are able to eat these walnuts, because the shells are extremely hard and the process is labour-intensive. To consume all of the meat it is more efficient to eat from holes on the raphe than to attempt to eat from other parts of the shell. We examined the effect of experience on the walnut-feeding skills of mice in the field. Feeding behaviours were compared among mice from habitats with and without walnut trees. Mice from habitats with walnut trees tended to consume nuts more efficiently than mice from habitats without walnut trees. We also observed the feeding behaviour of mice in an experimental area into which walnuts were artificially placed over a period of one year. This manipulation increased the proportion of mice that were able to eat nuts frequently and efficiently. Therefore, the walnut-feeding skills of mice improved with experience. Because individual mice acquired efficient feeding during the 14-day period of walnut conditioning, trial-and-error learning may be an important mechanism contributing to this behaviour.     

Histological, Physiological and Biochemical Interactions between Vesicular-Arbuscular Mycorrhizae and Thielaviopsis basicola in Tobacco Plants

Interactions between the mycorrhizal fungus Glomus monosporum and the root rot pathogen Thielaviopsis basicola and their effects on tobacco plants were investigated over a 4 week period. Mycorrhizal tobacco plants, obtained by preinoculation with G. monosporum, showed a better tolerance to T. basicola than non-mycorrhizal seedlings. Root and leaf dry weights of mycorrhizal plants were greater than those of controls. Mycorrhizal plants inoculated with T. basicola showed higher root and leaf dry weights than non-mycorrhizal infected plants, but lower values than mycorrhizal plants which were not infected. No appreciable differences in free aminoacid composition were observed among the different treatments with two exceptions: proline content was higher in infected and mycorrhizal infected plants compared to control and mycorrhizal plants; arginine content was higher in infected and mycorrhizal infected plants compared to control and mycorrhizal plants; arginine content was higher in mycorrhizal plants than in all the other treatments. The mechanisms by which (VAM) fungi can reduce disease incidence and pathogen development are discussed.     

Observations on the Behavior of Rain Forest Peccaries in Perú: Why do White-lipped Peccaries Form Herds?

The ecology and behavior of Tayassu tajacu and T. pecari were studied for a total period of 16 months in the years 1975–1978 in the Manú National Park in southeastern Perú. It appears from 132 sightings of T. tajacu at the study site, and from reports from other regions, that groups of this species in rain forest usually contain fewer than 12 individuals. The more observers were at the study site, the more frequently this species was encountered. T. tajacu also repeatedly used wallows in the forest. These observations suggested that individuals of this species were relatively sedentary. Herds of T. pecari were encountered on 60 occasions. Five counts indicated that there were over 100 individuals in the herds. This species was encountered at practically random intervals, independent of number of observers at the site, but more frequently in the dry season than in the rainy season. Adults of both species are prey primarily of large cats and humans. Both species feed on green plant parts, fruits, nuts and seeds, but T. pecari feeds on more resistant seeds and nuts than T. tajacu. The hardest palm nuts that only T. pecari can consume are distributed in a patchy manner. Cracking these nuts between the teeth causes the animals to be heard more than 50 m away. The patchy distribution of the hard nuts and seeds prevents T. pecari from being sedentary, and group formation likely has several individual benefits for foraging efficiency and defense against predators. These may include (1) avoidance of searching for food in places recently visited by others, (2) benefitting from the knowledge of experienced foragers, (3) reducing the per capita probability of detection by predators, (4) reducing the probability of being captured after group detection by predators, (5) increasing the ability to counterattack as a group, (6) increasing the probability of detecting the predator before it can attack, and (7) “confusing” the predator through escape behavior. T. tajacu seems to live in small groups because its typical foods are distributed more evenly and because consumption of these foods does not cause individuals to be so noticeable to predators.     

Predispersal predation by rodents to beechnuts (Fagus crenata Blume)

The rodents predation intensity and discrimination ability toward the predispersal beechnuts (Fagus crenata) were investigated using a tree tower in a beech forest, central Japan in 1999 and 2000. In this stand, using seed traps, the densities of fallen viable nuts were 35.1m^–2 in 1999 and 8.4m^–2 in 2000. The vertebrate-damaged nuts had fallen 5.6 and 2.2m^–2 in 1999 and 2000, respectively. Yet, the crop of viable nuts in 1999 was not so rich as that in a mast year. In 1999, predispersal predation by rodents was recognized at 16–19m above ground through the bagging experiment. In 2000, there were no predispersal predation and yet we captured Apodemus argenteus three times and Glirulus japonicus frequently on the tree. Judging from the facts of their feeding behaviors and the tooth scars left on the cupules and nuts, Apodemus argenteus might have been more responsible for predation to the predispersal beechnuts rather than Glirulus japonicus. Apodemus argenteus population seemed to be abundant on the ground in both years. If the main agent of predispersal predator were Apodemus argenteus, their number shifted to the canopy would be much larger in 1999 than in 2000 according as the crop of viable nuts. In an additional experiment, rodents preferred intact cupules to insect-damaged cupules on the tree, suggesting that they discriminated the quality of the predispersal nuts, even in the cupule stage, through olfactory and/or visual senses. Thus, predispersal nut predation by rodents was prevalent during the limited period in autumn.     

Cytogeographical study of four aneuploids ofCarex oxyandra Kudo in Japan

Chromosome numbers were determined for 342 clones ofCarex oxyandra collected from 35 localities in Hokkaido, Honshu, Shikoku and Kyushu, Japan. Four intraspecific aneuploids, 2n=18, 20, 24 and 26, were found. In meiotic division, only bivalent chromosomes were observed in all clones at metaphases I and II, suggesting that the aneuploids are established gamodemes. In the mitotic metaphase chromosomes, trimodal variation in chromosome length was observed. The 2n=26 clones found on Mt. Hiko had two particularly small chromosomes. The cytodemes with higher number of chromosomes are distributed in more southern areas of Japan.Carex oxyandra, therefore, accompanied with chromosome fragmentations, might spread the geographical distribution to the southern parts. The morphological characters of leaves, spikes, scales, perigynia and nuts were similar among the four cytodemes, except for the small leaves on plants from Yaku Island.     

Chimpanzees using stones to crack open oil palm nuts in Liberia

The use of stone-tools to open palm nuts (Elaeis guineensis) was studied in a group of 16 chimpanzees (Pan troglodytes) released from captivity to a natural island setting in Liberia. The behaviour was started by one female of the group; it then spread to 12 others over periods varying from a few seconds to a few weeks. Nut-cracking soon spread to three other sites, both spontaneously and with human encouragement. Both nuts and tools were carried distances of several hundred meters. Social interaction at cracking sites was rich and varied, ranging from fights over possession of tools to unsolicited sharing of nuts. Nut-crackers showed selectivity in schoosing “good” nuts and varied their methods according to the qualities of the nuts. Individual differences in technique emerged. These are the first detailed behavioural data on palm-nut-cracking, and they show many parallels with nut-cracking of other species by chimpanzees of the Tai Forest, Ivory Coast. This is another example of hammer-stone use from a limited region of West Africa: southeastern Guinea, eastern Liberia, and western Ivory Coast. This suggests limited cultural diffusion of the custom.