Variable correspondence of female host preference and larval performance in a phytophagous ladybird beetle Epilachna pustulosa (Coleoptera: Coccinellidae)

Genetic and environmental factors causing intraspecific variation of the thistle Cirsium kamtschaticum Ledeb. as a host plant of the phytophagous ladybird Epilachna pustulosa Kôno were investigated through simple food-choice tests and rearing of larvae. Two thistle clones (T1U₁ and T4H₂) were used, originally growing approximately 12 km apart. A previous study showed that adult female ladybirds preferred T1U₁ to T4H₂, and that larval performance was better on T1U₁, when leaves from the clones in situ were examined. The two clones retained their characteristics with respect to beetle preference after transplantation into a common garden. However, the difference between T1U₁ and T4H₂ with respect to larval performance was reduced after the transplantation. When leaves from shoots of T1U₁ exposed to different sunlight intensities were offered, adult female ladybirds did not show obvious preferences. Larval eclosion rates increased significantly with the increase in leaf sunlight intensity exposure. These results suggest strongly that both genetic and environmental factors are involved in interclonal variation of thistle quality in beetle preference and/or performance. It is suggested that the quality of thistle leaves for larval performance is largely affected by environmental factors, while leaf quality for beetle preference may be determined strictly by genetic factors. Under certain conditions, E. pustulosa females may behave maladaptively, preferring plants not appropriate for larval growth, or not choosing plants appropriate for the larval growth.     

Spatio-temporal dynamics of willow beetle (Phratora vulgatissima) in short-rotation coppice willows grown as monocultures or a genetically diverse mixture

1 Phratora vulgatissima (Chrysomelidae) is the major pest of short-rotation coppice willows in the U.K., capable of causing severe defoliation in monoculture plantations. As this beetle shows feeding preferences between willow clones, knowledge of the spatio-temporal dynamics of P. vulgatissima is needed in order to assess the effects of mixed clonal plantings on the management of this pest. 2 Distribution patterns of adult P. vulgatissima were monitored over a season at a spatial scale of every tree or every other tree in single clone plantings (monocultures) of three willow clones and in a regular row mixture of these clones. 3 Distribution indices (deviation from Complete Spatial Randomness) demonstrated that P. vulgatissima adults were spatially aggregated on favourable clones in the monocultures and the mixture. The degree of aggregation (measured by the k-parameter of the negative binomial) differed between willow clones. Beetles were highly aggregated at the start of the season, but less so later. 4 Dispersal studies of P. vulgatissima adults over time showed a pattern of beetle infestation along rows of the preferred clones in a regularly structured mixture. In the monocultures, movement was less directional. 5 Plantation design offers some potential for more effective non-chemical pest management for P. vulgatissima. Monitoring of the pest must take account of our findings that adults occur in mobile aggregations.     

Host plant suitability of a recently naturalized thistle Cirsium vulgare (Asteraceae) for a phytophagous ladybird beetle,Epilachna pustulosa (Coleoptera: Coccinellidae)

The phytophagous ladybird beetle, Epilachna pustulosa Kôno occurs mainly on the thistle Cirsium kamtschaticum Ledeb. In recent years, we have confirmed feeding and oviposition by E.pustulosa, and some degree of larval development of this beetle, on a naturalized congeneric thistle, Cirsium vulgare (Savi) Tenore. In this study, we assessed the host suitability of C.vulgare for E.pustulosa, using larval rearing and simple food choice tests, by comparing the performance on, and preference for, their legitimate host C.kamtschaticum. All experiments were initiated in July, when the utilization of C.vulgare by E.pustulosa became obvious. For larval performance, E.pustulosa showed a lower eclosion rate, shorter developmental duration and smaller body size when reared on C.vulgare than on C.kamtschaticum. Cirsium vulgare appeared to be a less adequate larval food than C.kamtschaticum. Adult feeding preference was altered seasonally; females preferred C.kamtschaticum to C.vulgare in a food choice test conducted in early July, whereas they showed the opposite preference in tests conducted in mid and late July. The utilization of C.vulgare by E.pustulosa appeared to be induced by qualitative deterioration of C.kamtschaticum leaves in July, which resulted in increased dispersion of adult beetles.     

Regeneration and transformation of Eucalyptus camaldulensis

Reliable regeneration protocols for Eucalyptus camaldulensis using leaf explants from in vitro-grown plants have been developed. Out of the 24 clones tested 13 were regenerated and of these, 6 showed regeneration from more than 60% of the explants. Identical protocols were also successful in the regeneration of some clones of E. microtheca, E. ochrophloia, E. grandis and E. marginata, but at lower frequencies. Co-cultivation of E. camaldulensis leaf explants with Agrobacterium tumefaciens strains carrying a kanamycin resistance gene and the reporter gene β-glucuronidase (GUS), followed by selection on kanamycin at 9 mg l^–1, allowed the selection of transformed shoots that could be rooted on selective media. Transformation of the plants was verified by staining for the GUS enzyme in various plant tissues, NptII assays and by Southern blotting on isolated DNA using specific probes for both the GUS and selectable marker genes. Transformed tissue was obtained with 5 clones of E. camaldulensis tested and a number of A. tumefaciens strains. However, only 1 clone regenerated transformed whole plants reliably. Received: 14 October 1996 / Revision received: 18 February 1997 / Accepted: 1 April 1997     

Preference-performance linkage in a herbivorous lady beetle: consequences of variability of natural enemies

We investigated the relationship between oviposition preference and offspring performance in a herbivorous lady beetle Epilachna pustulosa on two co-occurring plant species, thistle Cirsium kamtschaticum and blue cohosh Caulophyllum robustum, in 1994 and 1995. The relative importance of bottom-up effects by host plants and top-down effects by natural enemies on offspring performance were determined using field and laboratory experiments. In both years, egg density on blue cohosh was significantly higher than on thistle. A laboratory experiment demonstrated that larval survival from hatching to adult emergence was significantly higher, and developmental period shorter when larvae were reared on blue cohosh compared to thistle. The positive preference-performance linkage varied between years in the field. Top-down effects had a different impact on larval survival on the two host plant species. Arthropod predators, a lady beetle Harmonia axyridis and an earwig Forficula mikado, considerably depressed immature survival on thistle, while they were negligible on blue cohosh. Although the lack of effective predation increased larval survival on blue cohosh, it led to defoliation due to increased larval feeding late in the season. Because of severe intraspecific competition, old larvae had significantly lower survival on blue cohosh than on thistle. In 1994, as larval survival decreased due to defoliation on blue cohosh, the overall survival rate was significantly higher on thistle than on blue cohosh. This survival pattern was opposite to that found in the laboratory experiment. In contrast, in 1995, the increase in predatory lady beetles on thistle caused greater larval mortality. Thus, the overall survival was significantly lower on thistle than on blue cohosh, although severe intraspecific competition occurred on blue cohosh as it had in 1994. Consequently, the offspring performance on the two host plants is largely determined by the relative importance of arthropod predation determining larval survival on thistle and host plant defoliation reducing late larval survival on blue cohosh. These results indicate the important role of spatial and temporal variability of natural enemies on the preference-performance linkage of herbivorous insects. Received: 19 August 1998 / Accepted: 11 January 1999     

Spatial and temporal patterns of abundance of the potato leafhopper among red maples

A study was conducted in the 2001 growing season to examine the relative abundance, dispersion and distribution of the potato leafhopper (Empoasca fabae) among trees of 24 red maple (Acer rubrum) clonal selections. Yellow sticky traps hung from the trees were used to estimate the relative number of aerial potato leafhoppers among trees, starting on 1 May 2001 and stopping on 4 September 2001. More insects were collected from the east side of the plot than from the other sides in May, but no differences were significant late in the season. Few leafhoppers were collected during May, but the number of leafhoppers collected on traps rapidly increased and peaked during June. Afterwards, populations decreased and just a few individuals were trapped late in the season. Overall, significantly more leafhoppers were caught on traps hanging from trees of clone 56026 and 55410 than on trees of the other clones. Dispersion of the potato leafhopper changed from being aggregated in May, to being random in August, and also differed among the different clones. Populations tended to aggregate with increases in population numbers. Positive significant autocorrelations were detected at short distances in May, suggesting an aggregated spatial pattern early in the season. Yet, different autocorrelation patterns were detected within each block later in the season. Our results indicate that management practices could be developed to minimise pesticide use by targeting smaller areas with the highest populations at the right time of the season.     

Individual variation in two host plants of the ladybird beetle,Epilachna pustulosa (Coleoptera: Coccinellidae)

Individual variation in two species of host plants (thistle,Cirsium kamtschaticum, and blue cohosh,Caulophyllum robustum) of the herbivorous ladybird beetleEpilachna pustulosa was examined under laboratory conditions for their acceptability to adult beetles as a food resource, for adult preference and for larval performance. When clones of these plants were subjected to non-choice tests using posthibernating female beetles, there was found to be significant intraspecific variation among clones in terms of their acceptability, but interspecific variation was not detected. Significant intraspecific as well as interspecific variation were frequently detected in the two host plants when clones of these plants were subjected to choice tests using posthibernating female beetles; the magnitude of interspecific plant variation for beetle preference is not necessarily larger than that of intraspecific plant variation. Individual variation across plant species with respect to beetle larval performance was also significant. A positive correlation between adult preference and larval performance is suggested across the two taxonomically remote host plant species, thistle and blue cohosh, although this needs further investigation.     

Successful gonadal development and maturation of tench (Tinca tinca L.) in small concrete ponds

The experiments were performed in a tench farm from autumn until the spawning season (June–July). Tench broodstocks from natural habitats were maintained in 25 × 6 × 1 m concrete ponds and fed on commercial trout pellets. Females and males were separated and maintained under natural photoperiod and temperature conditions at densities around 2 kg m^?2. Water flow throughout was supplied at the rate of 15 L s^?1. When females showed external signs of advanced gonadal development, induction of spawning was made by luteinizing hormone releasing factor (LH‐RH) synthetic analogue administration at three different periods of the reproductive season (June–July). A single intramuscular injection (20 μg kg^?1 body weight) was administered to 110 mature females selected from a total of 150. The females were stripped 42 h (22°C) after hormone administration. The mean rate of stripped females to the number injected was 77%. Mean relative egg weight in relation to the weight of the stripped females was 5.61%. More than 90% of the males provided semen without hormonal induction. Differences in egg production and external egg quality were observed at different times of the spawning period. It was proven that tench maintained in small concrete tanks and fed on artificial diets were able to reach gonadal maturation.     

Year-round local movements of the japanese fluvial sculpin,Cottus pollux (large egg type), with special reference to the distribution of spawning nests

Year-round local movements of adult Japanese fluvial sculpin.Cottus pollux (large egg type), were investigated by a capture-mark-and-recapture method from July 1989 to July 1990 in the upper reaches of the Inabe River, central Japan. In the pre-breeding (July to January) and post-breeding seasons (June to July), the mean distance of movements in males and females was less than 20 m, and there was no apparent tendency to move into a particular channel-unit habitat, suggesting residential tendency in both sexes. In the breeding season (February to May), males tended to move into the raceways where most of the spawning nests were found, but females did not do likewise, indicating a sexual difference in movements in that season. Such sexual difference in movements was also confirmed by the records of individuals captured and recaptured more than three times: six (42.9%) of 14 males moved into the raceways in the breeding season, whereas only one (7.1%) of 14 females did so in that season. The reason behind this sexual difference in movements observed in the breeding season is discussed from the viewpoint of the patterns of spatial distribution between sexes during the pre-breeding and breeding seasons.     

Persistence of bacterial and archaeal communities in sea ice through an Arctic winter

The structure of bacterial communities in first‐year spring and summer sea ice differs from that in source seawaters, suggesting selection during ice formation in autumn or taxon‐specific mortality in the ice during winter. We tested these hypotheses by weekly sampling (January–March 2004) of first‐year winter sea ice (Franklin Bay, Western Arctic) that experienced temperatures from ?9°C to ?26°C, generating community fingerprints and clone libraries for Bacteria and Archaea. Despite severe conditions and significant decreases in microbial abundance, no significant changes in richness or community structure were detected in the ice. Communities of Bacteria and Archaea in the ice, as in under‐ice seawater, were dominated by SAR11 clade Alphaproteobacteria and Marine Group I Crenarchaeota, neither of which is known from later season sea ice. The bacterial ice library contained clones of Gammaproteobacteria from oligotrophic seawater clades (e.g. OM60, OM182) but no clones from gammaproteobacterial genera commonly detected in later season sea ice by similar methods (e.g. Colwellia, Psychrobacter). The only common sea ice bacterial genus detected in winter ice was Polaribacter. Overall, selection during ice formation and mortality during winter appear to play minor roles in the process of microbial succession that leads to distinctive spring and summer sea ice communities.     

Entamoeba Clone‐Recognition Experiments: Morphometrics,Aggregative Behavior,and Cell‐Signaling Characterization

Studies on clone‐ and kin‐discrimination in protists have proliferated during the past decade. We report clone‐recognition experiments in seven Entamoeba lineages (E. invadens IP‐1, E. invadens VK‐1:NS, E. terrapinae, E. moshkovskii Laredo, E. moshkovskii Snake, E. histolytica HM‐1:IMSS and E. dispar). First, we characterized morphometrically each clone (length, width, and cell‐surface area) and documented how they differed statistically from one another (as per single‐variable or canonical‐discriminant analyses). Second, we demonstrated that amebas themselves could discriminate self (clone) from different (themselves vs. other clones). In mix‐cell‐line cultures between closely‐related (E. invadens IP‐1 vs. E. invadens VK‐1:NS) or distant‐phylogenetic clones (E. terrapinae vs. E. moshkovskii Laredo), amebas consistently aggregated with same‐clone members. Third, we identified six putative cell‐signals secreted by the amebas (RasGap/Ankyrin, coronin‐WD40, actin, protein kinases, heat shock 70, and ubiquitin) and which known functions in Entamoeba spp. included: cell proliferation, cell adhesion, cell movement, and stress‐induced encystation. To our knowledge, this is the first multi‐clone characterization of Entamoeba spp. morphometrics, aggregative behavior, and cell‐signaling secretion in the context of clone‐recognition. Protists allow us to study cell–cell recognition from ecological and evolutionary perspectives. Modern protistan lineages can be central to studies about the origins and evolution of multicellularity.     

Serum and Egg Antibody Responses in Chickens to Escherichia coli

The effects of Freund’s adjuvants on antibody production in chickens against E. coli whole cells were examined. The levels of anti-E. coli IgG antibodies in serum were higher when Freund’s complete (FCA) or incomplete adjuvant (FIA) was administered than that without adjuvant. Production of antibodies recognizing E. coli cells and their lipopolysaccharide was enhanced by FIA, while both FIA and FCA enhanced production of antibodies recognizing outer membrane components. In contrast, serum IgM antibody levels were higher when no adjuvant was used. Anti-E. coli IgG antibodies in serum were efficiently transferred to egg yolk, giving antibody activity in egg yolk similar to that in serum. However, anti-E. coli IgM antibodies were not detected in the egg, suggesting that egg (white) IgM was not influenced by antigenic stimulation of the humoral immune system. Antimicrobial activity of the egg yolk IgG was highest when the bacteria antigen was injected with FIA.     

Indirect interaction between two native thistles mediated by an invasive exotic floral herbivore

Spatial and temporal variation in insect floral herbivory is common and often important. Yet, the determinants of such variation remain incompletely understood. Using 12 years of flowering data and 4 years of biweekly insect counts, we evaluated four hypotheses to explain variation in damage by the Eurasian flower head weevil, Rhinocyllus conicus, to the native North American wavyleaf thistle, Cirsium undulatum. The four factors hypothesized to influence weevil impact were variations in climate, weevil abundance, phenological synchrony, and number of flower heads available, either on wavyleaf thistle or on the other cooccurring, acquired native host plant (Platte thistle, Cirsium canescens), or on both. Climate did not contribute significantly to an explanation of variation in R. conicus damage to wavyleaf thistle. However, climate did influence weevil synchrony with wavyleaf flower head initiation, and phenological synchrony was important in determining R. conicus oviposition levels on wavyleaf thistle. The earlier R. conicus was active, the less it oviposited on wavyleaf thistle, even when weevils were abundant. Neither weevil abundance nor availability of wavyleaf flower heads predicted R. conicus egg load. Instead, the strongest predictor of R. conicus egg load on wavyleaf thistle was the availability of flower heads on Platte thistle, the more common, earlier flowering native thistle in the sand prairie. Egg load on wavyleaf thistle decreased as the number of Platte thistle flower heads at a site increased. Thus, wavyleaf thistle experienced associational defense in the presence of flowering by its now declining native congener, Platte thistle. These results demonstrate that prediction of damage to a native plant by an exotic insect may require knowledge of both likely phenological synchrony and total resource availability to the herbivore, including resources provided by other nontarget native species.     

Spatial distribution of colonizing Listronotus maculicollis populations: implications for targeted management and host preference

We characterized the spatio‐temporal distribution of emerging overwintered adult Listronotus maculicollis populations colonizing golf course fairways with Spatial Analyses by Distance IndicEs (SADIE) with the goal to better target management tactics and to test assumptions for weevil preference for annual bluegrass (Poa annua). Adults randomly colonized and moved throughout fairways. However, cumulative captures were significantly aggregated along fairway edges closest to overwintering sites demonstrating progressive movement through the edges. Spatial association analyses suggest that the spatial patterns of cumulative captures of adults rather than weekly patterns were strongly associated with larvae, indicating that the adults enter fairways from the edges and deposit eggs over the course of several weeks. We did not observe an effect of host species on the distribution of either adult or larval L. maculicollis which leads us to question traditional assumptions of host preference for short mown P. annua. Instead, we propose that the aggregated distribution of larvae is generated by a low encounter rate of short mown hosts, rather than preference for species or cultivar. This study indicates that caution need to be applied when using preference‐performance criteria in host preference studies. Future behavioural studies need to address the contributions of encounter rate and host species on L. maculicollis host selection and oviposition.     

Co-inoculation of Ceratocystis polonica and Heterobasidion annosum with callus of two Norway spruce clones with different in vivo susceptibility

We have studied how callus cultures from two clones of Norway spruce influence the growth of two pathogens, Ceratocystis polonica and Heterobasidion annosum, when co-cultivated in vitro. In field experiments, trees of clone 409 were susceptible to both fungi, whereas clone 589 was less affected. Callus was cultured on medium containing cytokinins (benzylaminopurine, kinetin) and with or without auxin (2,4-dichlorophenoxyacetic acid). For co-cultivation with fungus, one piece of callus was placed towards the edge of each Petri dish. One and 14 days after inoculation with callus the dishes were co-inoculated with the fungus. Both clones strongly stimulated the initial growth of both fungi. Clone 589 inhibited the growth of both fungi when the fungi were inoculated one day after the callus. When the callus was cultured on medium without auxin for 14 days before co-inoculation clone 589 strongly inhibited the growth of both fungi, whereas clone 409 inhibited H. annosum only. This revised version was published online in June 2006 with corrections to the Cover Date.     

Life tables for worker honeybees

Life tables for worker honeybees covering all life span, and those for adults, were prepared for three seasonal cohorts, June bees, July bees and wintering bees. Survivorship curves for June and July bees show a convex type being exceptional for insects, with relatively high mortality at egg and feeding larval stages and at later adult stage after most bees became potential foragers. Adult longevity greatly lengthens in Winteriing bees and survivorship curve drops approximately with the same rate. A remarkable similarity of survivorship curves for men and honeybees was demonstrated, apparently due to highly developed social care in both. Some comments were given on mortality factors. The importance of life tables for population researches was shown by applying our result to the population growth curve made byBodenheimer , based upon the data byNolan . At the asymptote of the uncorrected curve, the ratio of total population estimated by uncorrected curve to that by corrected curve reaches about 3∶2.     

Leaf nutrient levels and the spatio-temporal distributions of <Emphasis Type="Italic">Plutella xylostella</Emphasis> and its larval parasitoids <Emphasis Type="Italic">Diadegma insulare</Emphasis> and <Emphasis Type="Italic">Microplitis plutellae</Emphasis> in canola

Seasonal distribution patterns of the diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), and its principal parasitoids Diadegma insulare (Cresson) (Hymenoptera: Ichneumonidae) and Microplitis plutellae (Muesebeck) (Hymenoptera: Braconidae) were investigated over three site-years in commercial fields of canola (Brassica napus L.) in southern Alberta, Canada. The sampling of P. xylostella, D. insulare, and M. plutellae from points arranged in grid patterns, together with the mapping and analysis of their spatial distributions over time, generated a detailed picture of the pattern of crop infestation by the herbivore and its parasitoids. Plutella xylostella exhibited significant aggregations on different scales most often when its host plants were in early flowering. Diadegma insulare adults exhibited significant aggregated distributions during early flowering and distributions subsequently became more uniform as the wasps moved into the crop later in the season. However, M. plutellae distributions were aggregated in mid flowering in only one site-year. The close spatial associations between densities of D. insulare and P. xylostella indicated that host abundance was the main determinant of parasitoid distribution patterns. Spatial distributions of nutrient contents in leaf tissue and their spatial associations with the herbivore and parasitoids were also investigated. Significant spatial associations existed between certain nutrients (e.g. nitrogen, sulfur, and potassium) and P. xylostella distributions. Sulfur exhibited a positive effect on the distributions of D. insulare but not of M. plutellae. We observed similar relationships between nutrients and the distribution of P. xylostella parasitoids as for nutrients and P. xylostella, but these relationships lacked consistency and may be the results of the spatial associations between the parasitoids and their hosts. Aggregated distributions of adults and larvae of P. xylostella hold promise for spatially targeted insecticidal applications as a means for reducing the environmental impact of insecticides on nontarget and beneficial species in canola agroecosystems.     

Testing a new hypothesis: plant vigor and phylloxera distribution on wild grape in Arizona

Summary Longer, meaning more vigorous, shoots of a wild grape clone (Vitis arizonica) were more susceptible to attack by second and third generations of leaf-galling grape phylloxera,Daktulopsphaira vitifoliae, as the growing season progressed. Although there was no significant difference in mean shoot length between attacked and unattacked shoots within a clone at the beginning of shoot elongation, attacked shoots were significantly longer than unattacked shoots when elongation had ceased (P<0.01). Also, long attacked shoots had a significantly greater population of phylloxera galls than short attacked shoots (P<0.01) as the season progressed. The phylloxera population on long shoots increased rapidly while the population on short shoots remained the same. Longer shoots also produced significantly more axillary shoots than shorter shoots as the season progressed (P<0.001), and the number of axillary shoots accounted for 66 percent of the variance in number of attacked leaves on a shoot. Experimental evidence showed that there was a significantly greater percentage of available leaves attacked on long shoots than on short shoots (P<0.05) and the leaves on long shoots generally had a greater number of galls per leaf. The relationship between shoot length and probability of attack was also tested by comparing shoots lengths of 10 attacked clones and 10 unattacked clones at a second location. Mean shoot lengths of attacked clones were significantly longer than mean shoot lengths of unattacked clones (P<0.05), and mean shoot lengths of attacked shoots within a clone were significantly longer than unattacked shoots (P<0.001). Longer shoot length accounted for 81 percent of the variance in probability of attack. The reason for this pattern of attack was that long shoots produced newly expanding leaves over a longer time during the growing season and multivoltine phylloxera require undifferentiated tissue to initiate gall formation. Patterns of attack within a shoot were characterized by an uneven distribution of galls among leaves. This was due to development time between generations and the current availability of undifferentiated tissue at times of colonization. This study supports the hypothesis that some herbivore species are favored more by vigorous plants than by stressed plants.     

Relative preference of Lygus hesperus (Hemiptera: Miridae) to selected host plants in the field

Abstract Information on host plant preference of agriculturally important insect pests, such as Lygus hesperus (Knight), can be helpful in predicting its occurrence and future movement among crop and non-crop host plants. A field study was conducted during 2005 and 2006 to evaluate the host preference of Lygus to cotton and four other host plants in the Texas High Plains, including alfalfa (Medicago sativa L.), wild sunflower (Helianthus annuus L.), Russian thistle (Salsola iberica L.) and pigweed (Amaranthus palmeri L.). Sampling for both nymphs and adults during 2005 (July to November) and 2006 (June to November) showed that alfalfa and Russian thistle were the two most preferred hosts out of the five hosts evaluated. Abundance of nymphs (numbers per 50 sweeps per host plant) during the sampling period also indicated the superior reproductive suitability of alfalfa and Russian thistle. Cotton appeared to be the least attractive host plant for Lygus when Russian thistle and alfalfa were available in the host mosaic. Seasonal abundance of Lygus was found to be lower during 2006 compared to 2005, which may be explained by the difference in rainfall patterns during these two years. In terms of species dominance, L. hesperus was the most dominant species in the sampled population followed by an inconsiderable fraction of L. elisus and L. lineolaris.     

Cloning and Expression of Chlorobium vibrioforme Uroporphyrinogen Decarboxylase Gene in a Salmonella Heme Auxotroph

To study the post-uroporphyrin steps in heme and chlorophyll biosynthesis in Chlorobium, we attempted to clone the uroporphyrinogen decarboxylase (hemE) gene. A Chlorobium genomic library was used to transform a restriction-minus Salmonella typhimurium strain. The recombinant DNA molecules were transduced into an auxotrophic Salmonella double mutant (hemA⁻ hemE⁻) by phage P22. Faster-growing colonies indicated complementation of the hemE mutation. Each clone was tested by backcross transduction of the mutant. Growth rates of the confirmed clones in LB medium were comparable to wild-type Salmonella. HPLC analysis of the substrate (uroporphyrinogen) and the product (coproporphyrinogen) of the decarboxylase activity was performed in one such clone. This clone showed an active hemE gene within a 4-kb insert. Received: 21 February 2002 / Accepted: 8 May 2002