Man-made Substrates Research Articles

Antifouling effect of two saturated copper coatings applied on carbon steel structures

Biofouling is the colonization of man-made substrata by sessile organisms. The aim of this paper is to evaluate the performance of two antifouling saturated copper coating. Bioassays were carried out at a harbor in Argentine (38°02’S- 57°32’W). During six months, one series of pipes and panels were removed monthly to estimate the recruitment of macro and microfouling species and immediately replaced by clean ones. Another series was removed from the beginning of exposure to monitor the development of the established community (accumulative pipes and panels along six months). Data obtained from control (without a saturated copper coating) and saturated-copper coated pipes and panels were compared in order to estimate performance of the coating. One of two saturated copper coating demonstrated a good effect antifouling.

Settlement of Crassostrea ariakensis Larvae: Effects of Substrate, Biofilms, Sediment and Adult Chemical Cues

The Suminoe oyster (Crassostrea ariakensis) is being considered for introduction into the Chesapeake Bay. However, our current understanding of the biology and ecology of C. ariakensis is insufficient to predict whether an introduction will be successful, provide desired benefits, or have adverse impacts. Behavior of native Eastern oyster (C. virginica) pediveligers has been studied for many years and it is well established that they use a variety of habitat characteristics when selecting a site for colonization. Perhaps the most important of these are chemical cues emitted by adult conspecifics, which can lead to gregarious larval settlement and dense, persistent reef communities. Conversely, almost nothing is known about the mechanisms that regulate larval settlement and metamorphosis for C. ariakensis or how pediveligers might respond to conditions found in Chesapeake Bay. In a comparative study with C. virginica, we examined how environmental factors such as substrate type, natural biofilms, sediment and waterborne chemical cues influence larval settlement for two C. ariakensis strains (''south China'' and ''west coast''). Our results demonstrate many similarities but also potentially important differences. Both species and strains of larvae greatly prefer natural substrates (e.g., shell) covered with biofilms for colonization but the west coast strain of C. ariakensis exhibited greater attachment onto manmade substrates (e.g., fiberglass) than C. virginica. Waterborne chemical cues emitted by adult oysters were also found to enhance substrate attachment for all larval forms but cues do not appear to be species specific. These results provide critical insight to the ability of C. ariakensis larvae to identify and colonize suitable substrates in the Chesapeake Bay, which will have a large impact on recruitment success and their ability to establish self-sustaining populations.

Pattern of settlement and natural chimerism in the colonial urochordate Botryllus schlosseri

Colonies of the cosmopolitan urochordate Botryllus schlosseri that share one or both alleles at a single allorecognition locus (Fu/HC) and come into tissue contacts, may fuse and form a mixed entity, a chimera. Botryllus populations worldwide exhibit unprecedented extensive polymorphism at this locus, a result that restricts fusions to kin encounters. This study aims to compare spatiotemporal configurations in source and introduced B. schlosseri populations, residing on natural and man-made substrata, respectively. By using four microsatellite loci, we tested genetic consanguinity of colonies settled naturally along spatial vectors on both, natural (native populations) and man-made (introduced) substrates. Four populations were studied. Results revealed that B. schlosseri colonies, on both substrate types, assemble in groups of relatives that share similar microsatellite profiles. We suggest that this pattern of settlement promotes the formation of chimeras, which evoke conflicting interactions: cooperation between different somatic cell lines that constitute the colonial soma and competition between germ cells that inhabit the chimera gonads. Under natural conditions, the chimera may allow genetic flexibility that depends on joint genomic fitness of its partners. This is probably one of the life history characteristics that led to the worldwide distribution success of this species.

The Distribution and Conservation Status of the Gull-billed Tern (Gelochelidon nilotica) in North America

The Gull-billed Tern (Gelochelidon nilotica) has until recently received little conservation and management attention within North America despite a relatively low overall population size and significant declines in parts of the breeding range. This lack of attention may stem in part from the wide distribution of the species, encompassing parts of six continents, and from its tendency to nest in relatively small, scattered and often ephemeral colonies. Populations of North American subspecies are alarmingly small. The current population of the eastern subspecies aranea in the U.S. is unlikely to exceed 3,600 pairs, with over 60% of these birds occurring in Texas. The Texas population has remained generally stable, but declines of populations in Maryland (where probably extirpated), Virginia, North Carolina, Florida, and possibly Georgia give cause for concern for this subspecies. For the western subspecies vanrossemi, as few as 250 pairs nest at only two locations in the U.S., both in California. When populations in western Mexico are considered, the entire vanrossemi population numbers only 600-800 pairs. Currently the Gull-billed Tern is listed as “endangered” or “threatened” in four states, and is considered to be of management concern in five others. The breeding range of the species has contracted and shifted slightly from its known historic range in the middle Atlantic states, but otherwise occupies its historic range in the United States and has expanded slightly to coastal southern California. Some range contraction in Mexico (e.g., in Sonora) may have occurred. In eastern Mexico, historical information is almost non-existent and knowledge of current distribution and abundance is incomplete. Main threats to populations in North America include loss of natural nesting islands through beach erosion or perturbations to estuarine functions, development or modification of upland habitats near breeding areas that may be important for foraging, and disturbances to colonies by humans and feral or human-subsidized predators. This species often nests on man-made substrates suggesting it could be responsive to management of breeding sites. Key research needs include more frequent and refined population monitoring, a better understanding of demographics, metapopulation dynamics and factors limiting populations as well as refinement of subspecies’ breeding distributions and wintering ranges.

Aeroterrestrial Microalgae Growing in Biofilms on Facades—Response to Temperature and Water Stress

The photosynthetic performance of a microalgal biofilm colonizing a building facade was investigated between February and July 2004, with an emphasis on changing water availability and air humidity. The fluorimetric measurements of the quantum efficiency (F (v)/F (m)) indicated diurnal activity patterns. At most sampling dates the algal biofilm photosynthesized particularly in the morning and substantially less in the afternoon. As long as liquid water was present, the microalgae exhibited at least some degree of photosynthesis. However, F (v)/F (m) values never exceeded 0.4, pointing to slight photoinhibition or damage of the cells. Dried cells without photosynthesis could recover within minutes after artificial moistening. Three microalgal strains were isolated from aeroterrestrial biofilms and established as unialgal cultures. Their photosynthesis and growth were characterized under different air humidities and temperatures. Photosynthesis and growth of strain ROS 55/3 (Stichococcus sp.) showed similar patterns with decreasing relative air humidity. Positive growth and optimum photosynthesis were recorded at 100% relative air humidity. At air humidities below 93%, both processes were strongly inhibited. All studied strains grew between 1 and 30 degrees Celsius with optimum rates at 20-23 degrees Celsius, indicating eurythermal features. The data indicate that liquid water or 100% air humidity are the prerequisite for optimum photosynthesis and growth of aeroterrestrial microalgae. However, when dried and consequently inactive, these microorganisms can recover quickly if water is suddenly available, e.g., after rain events. These physiological capabilities explain well the ecological success of aeroterrestrial microalgae in occupying many man-made substrata such as building facades and roof tiles in urban areas.

Antifouling diketopiperazines produced by a deep-sea bacterium, Streptomyces fungicidicus

Modern antifouling coatings use heavy metals and toxic organic molecules to prevent biofouling, the undesirable growth of marine organisms on man-made substrata. In an ongoing survey of deep-sea microorganisms aimed at finding low toxic antifouling metabolites, an actinomycete bacterium was isolated from the Pacific sediment at the depth of about 5000 m. The bacterium was closely related to Streptomyces fungicidicus (99% similarity) according to 16S ribosomal RNA sequence information. The spent culture medium of this bacterium inhibited barnacle larval attachment. Bioassay-guided fractionation was employed to isolate antifouling compounds. The ethyl acetate extract was fractionated by using an open silica gel column. Active fractions were further purified on a HPLC C18 column. Five diketopiperazines, cyclo-(L-Leu-L-Pro), cyclo-(L-Phe-L-Pro), cyclo-(L-Val-L-Pro), cyclo-(L-Trp-L-Pro), and cyclo-(L-Leu-L-Val) were isolated for the first time from a deep sea bacterium, and the structures of the compounds were elucidated by nuclear magnetic resonance spectroscopy and mass spectrometry. The pure diketopiperazines were tested for antilarval activity using the barnacle Balanus amphitrite. Effective concentrations that inhibited 50% larval attachment (EC50) after 24 h ranged from 0.10 – 0.27 mM. The data suggest that diketopiperazines and other compounds from deep-sea bacteria may be used as novel antifoulants.

Antifouling potential of cyanobacteria: a mini-review

Cyanobacteria produce a variety of bioactive metabolites that may have allelochemical functions in the natural environment, such as in the prevention of fouling by colonising organisms. Chemical compounds from cyanobacteria are also of biotechnological interest, especially for clinical applications, because of their antibiotic, algicidal, cytotoxic, immunosupressive and enzyme inhibiting activities. Cyanobacterial metabolites have the potential for use in antifouling technology, since they show antibacterial, antialgal, antifungal and antimacrofouling properties which could be expoited in the prevention of biofouling on man-made substrata in the aquatic environment. Molecules with antifouling activity represent a number of types including fatty acids, lipopeptides, amides, alkaloids, terpenoids, lactones, pyrroles and steroids. The isolation of biogenic compounds and the determination of their structure may provide leads for future development of, for example, environmentally friendly antifouling paints. An advantage of exploring the efficacy of cyanobacterial products is that the organisms can be grown in mass culture, which can be manipulated to achieve optimal production of bioactive substances. Phycotoxins and related products from cyanobacteria may serve as materials for antimicro- and antimacrofouling applications. A survey of antibiotic compounds with antifouling potential revealed more than 21 different antifouling substances from 27 strains of cyanobacteria.

Effects of PVC, cast iron and concrete conduit on concentrations of copper in stormwater

Architectural applications of copper on roofs, gutters and facades exposes it to weathering forces, such as wind and all forms of precipitation. This leads to the dissolution of copper from these surfaces and its introduction into local watersheds. The potential for exposure in local watersheds is a function of the amount of copper entering the watershed and the assimilation capacity of a wide variety of natural and manmade substrates that transform and sequester copper, thus reducing exposure of sensitive organisms to bioavailable copper. This study investigates one type of manmade substrate, stormwater conduit, commonly used to transport stormwater away from buildings. Conduits made of PVC and cast iron did not significantly remove copper from a synthetic stormwater spiked with copper averaging 2391 μg Cu/L. Concrete conduit significantly removed copper from the synthetic water and at high rates (12 – 18%) over a short distance (610 cm). A high percentage (81 – 100%) of the copper removed from the water was from the dissolved copper fraction. Once adsorbed to the concrete, copper did not readily leach back into copper-free water subsequently pumped into the conduit. These characteristics have significant implications for watershed management and best management practices for reducing concentrations of copper in stormwater runoff.

HORNET CLEARWING MOTH (SESIA APIFORMIS [CLERCK]) AND DIEBACK OF POPLARS IN EASTERN ENGLAND

Summary Poplar trees showing severe dieback, or defoliation and death within a growing season, have been reported during recent years from the region around Bedford, Milton Keynes and Northampton. Many of the affected trees have also been found to be attacked by the hornet clearwing moth, Sesia apiformis (Cl.). Larvae of this moth tunnel beneath the bark of the lower trunk and roots, and it has been suggested that the moth might be the cause of the decline in tree health. However, a survey of 801 poplar trees in 1999 confirmed that, although there was a correlation between the amount of dieback and the number of moth exit holes visible in the trunk (r2 = 0.38), there was considerable variation in the relationship and a significant number of trees showed severe dieback but no evidence of infestation. The data demonstrate that dieback in many trees cannot be attributed to S. apiformis and that the moth most likely acts as a secondary agent in tree decline. Up to 53 per cent of trees with severe dieback occurred in shelterbelts, and many of these sites were situated on reclaimed land or other man-made substrates. The area where dieback has been most prevalent is one of the hottest and driest parts of the UK, and the reports of dieback in poplar and the apparent increase in S. apiformis activity, follow particular dry springs and early summers in 1995 and 1996. The available evidence suggests, therefore, that the deterioration in the health of poplars in this region is not caused directly by S. apiformis, but by a combination of climate and perhaps human influences.

Properties of Urbic Anthrosols from an abandoned shunting yard in the Ruhr area, Germany

The soil conditions of an abandoned shunting yard in the Ruhr area were studied to find the most important factors influencing plant growth and nutrient cycling. The chemical and physical conditions of five Urbic Anthrosols with different development histories were compared with those of a nearby Calcic Cambisol under agricultural use. In the second half of the 19th century, the ground level of the shunting yard site was raised about 2 m. The soils are now mainly Regosols, mostly well drained in the topsoil. The coarse material contents (>2 mm), which are up to 100% in base layers of railway tracks, and the quality of the man-made substrates (crushed rock, slags, ashes, coke) are the most important factors influencing plant growth. Poor physical structure and low contents of fine fraction (<2 mm) lead to low water storage. Together with a low available nutrient stock, these properties lead to poor establishment of vegetation. The burning of coal by steam engines produced highly polluted ashes that fill pore spaces in the railway ballast layers. All the investigated fine substrata of the shunting yard layers exhibit a higher magnetic susceptibility than the rural soil. The ashes still contain unburned coal so that, although the total organic C content of the man-made soils reaches 37%, the wide C:N ratio does not indicate the actual quality of humic substances. The acid neutralization potential of the topsoil layers of the shunting yard will be consumed by acidic atmospheric deposition within a few decades, whereas in the surrounding Calcic Cambisols, it will take more than a thousand years. This may cause further problems of groundwater quality, as the heavy metals will become more soluble. The low soil quality makes such sites suitable for vegetation species that cannot compete in the surrounding very eutrophic agricultural landscape.

DISPERSAL PATTERNS OF TERRICOLOUS LICHENS BY THALLUS FRAGMENTS

Dispersal patterns from seven terricolous lichen species, with a high capacity for asexual reproduction by fragmentation (Cetraria muricata, Cladonia species), were studied in differing vegetation types in north-eastern Germany. Marked lichen thalli were fragmented by trampling. After 15 days the spread of thallus fragments was monitored. Whereas most of the fragments that were dispersed by wind remained within a 20-cm radius from the source, the maximal dispersal distance was 57cm in a dry sand grassland and 68cm in an open pioneer pine forest. Dispersal was negligible in a closed old-growth pine forest. Several fragmented lichen cushions were disturbed and removed by animals, and led to a maximal dispersal distance of 9·70m. These results suggest that: (a) thallus fragments provide good short-distance dispersal in open vegetation, but are inefficient for long-distance dispersal, and (b) wind and animals are important factors for the dispersal of thallus fragments. For restoration management of man-made substrata, artificial introduction of lichen thalli is proposed.

NOTES ONXANTHORIATH. FR. III. TWO NEW SPECIES OF THEXANTHORIA CANDELARIAGROUP

Two new species ofXanthoriaare described and illustrated, and comments about their differences from related taxa are provided. These are:X. ascendensfrom bark and shrub remains in sea-shore sand dunes and hillsides in Argentina, Chile and Peru, andX. ucrainica, from bark, stones, and some man-made substrata in the Ukraine, Sweden, United Kingdom, Germany, Austria and Asian Russia.

Notes on Xanthoria Th. Fr. III. Two New Species of the Xanthoria Candelaria Group

Two new species of Xanthoria are described and illustrated, and comments about their differences from related taxa are provided. These are: X. ascendens from bark and shrub remains in sea-shore sand dunes and hillsides in Argentina, Chile and Peru, and X. ucrainica, from bark, stones, and some man-made substrata in the Ukraine, Sweden, United Kingdom, Germany, Austria and Asian Russia.

Calcium oxalate in lichen biodeterioration studied using FT-Raman spectroscopy

The biodeterioration of diverse natural and man-made substrata by Caloplaca aurantia, Lecanora muralis and Acarospora oxytoma has been studied using FT-Raman spectroscopy with 1064 nm laser excitation. Each of the three lichen species produce relatively large amounts of calcium oxalate in encrustations at the thallus—substratum interface during the biodeterioration process; the Raman spectroscopic technique is capable of identifying non-destructively the monohydrate, with ν(CO) stretching bands at 1463 and 1496 cm −1 and the dihydrate, with a ν(CO) stretching band at about 1475 cm(su−1). In this work, the presence of calcium oxalate monohydrate and dihydrate in the lichen encrustations is identified for these high-oxalate producing biodeteriorative lichen systems. The results indicate that the lichens adopt different methods for the production and removal of the hydrated calcium oxalates in the encrustations.

Retention of the Gram-negative bacterium SW8 on surfaces under conditions relevant to the subsurface environment: Effects of conditioning films and substratum nature

Shallow (5–13 m) and deep (35–65 m) groundwaters were evaluated for their ability to generate conditioning films which affect bacterial adhesion to natural (sandstone, shale, andesite) and man-made substrata (polypropylene, stainless steel). Water contact angles indicated that all water samples produced conditioning films. Most films modified retention of the nonmotile Gram-negative bacterium SW8, but attachment of the organism did not correlate with water contact angles. Each borewater produced conditioning films with a characteristic attachment profile of SW8. Films adsorbed from standing borewaters often retained SW8 in different numbers than coatings derived from pumped bores. Groundwater chemistry was very heterogeneous and microbiological data indicated the presence of a diverse aerobic and anaerobic microbial community. These results indicate that conditioning films derived from dissolved compounds may play a significant role in controlling the interaction of bacteria with substrata in the subsurface.

Dispersal and Recruitment of Zebra Mussel (Dreissena polymorpha) in a Nearshore Area in West-central Lake Erie: The Significance of Postmetamorphic Drifting

Quantitative evidence that early postmetamorphic stages of zebra mussel (Dreissena polymorpha) can disperse in the water column and colonize substrates has been obtained using off-bottom scouring pad collectors and plankton sampling in a nearshore, shallow-water area (2–7 m) near Wheatley, Lake Erie, during August 1991. Collectors were deployed for 24–72 h, thus minimizing growth of recruits during deployments. Size criteria determined through an in situ growth experiment were used to discriminate between individuals arriving at a collector as (1) settling larvae or (2) postmetamorphic stages. During certain periods, 20–80% of individuals settling on collectors were postmetamorphic stages. Plankton samples taken near collectors also confirmed the presence of postmetamorphic stages in the water column. Most drifting juveniles ranged from about 300 μm shell length to 800 μm (some up to 1–2 mm). Although settlement by postmetamorphic stages occurred during various periods, they drifted and settled on collectors in much higher numbers during periods of strong wave action generated during storms. Colonization of natural and man-made substrates by postmetamorphic stages may be significant in exposed or turbulent areas and may impact on population dynamics. Whether postmetamorphic drifting is adaptive and which mechanisms are involved are unknown.

Epilithic lichen vegetation on man-made, calcareous substrates in The Netherlands

This is the first survey of calcareous, saxicolous lichen vegetation on man-made substrates in The Netherlands. The fieldwork was carried out in the 1970's using the Braun-Blanquet approach. All communities are classified in the Verrucarietalia nigrescentis Klem. 50, Verrucarietea nigrescentis Wirth 80. The Caloplacetum citrinae, Lecanora albescens-Caloplaca citrina coenon, Caloplacetum saxicolae, Physcio nigricantis-Candelarielletum mediantis and Caloplacetum teicholytae belong to the Caloplacion decipientis Klem 50 (...)

Barnacle Adhesion: Reattachment of the Adult Barnacle Chthamalus Fragilis Darwin to Polystyrene Surfaces Followed by Centrifugational Shearing

Adult specimens of Chthamalusfragilis Darwin were removed carefully from the leaves and stalks of marsh grass and placed with their bases on the bottoms of polystyrene dishes containing sea water in order to allow reattachment of the animals. During overnight culture, animals secreted small volumes of a sea water-insoluble material which reattached the animals to the surface of the polystyrene dishes. The secreted material made a contact angle with the polystyrene surface of ~ 15?, indicating that, before setting, the liquid precursor of the set material has considerable wetting properties. Dishes with reattached barnacles were mounted in a centrifuge to load the animals in shear and were subjected to increasing speeds of rotation until each of the reattached barnacles became detached. Reattached barnacles sustained relative centrifugal forces of up to 6,800 x g before detachment, which usually occurred at the barnacle basis/ cement interface. The strength of adhesion of individual barnacles reattached to polystyrene ranged from 0.0014-0.3589 Newtons. Using the area of attachment between cement and polystyrene dish, the strength of adhesion averaged 1.05 x 105 Nm-2, with a range of 0.14 x 105 to 2.79 x 105 Nm-2. These data are comparable to forces of adhesion reported for the temporary attachment of cyprid larvae and the normal attachment of adult barnacles to plastics and other materials measured by techniques which load the animals in tension. Our observations and measurements indicate that the material secreted by adult barnacles under in vitro culture conditions exhibits significant adhesive properties. It is unlikely that adult barnacles detached from marsh grass reattach to polystyrene by Stefan effects. It is well known that barnacles secrete, under water, adhesive materials which facilitate and maintain their attachment to a variety of natural and man-made substrates including the coverings of other marine organisms such as other barnacles, mussels, oysters, crabs, turtles, fishes, and whales, bare and painted metals and woods, concrete, plastics, and various other synthetic materials. Attachment is initiated during a short period of the barnacle life cycle, the cyprid larval stage, towards the end of which exploring cyprids settle on a suitable substrate and secrete an adhesive which firmly binds the larvae to the surface (Nott and Foster, 1969). An extensive literature relates light (Visscher and Luce, 1928; Pomerat and Reiner, 1942), fluid flow rates (Smith, 1946; Crisp, 1955; Crisp and Stubbings, 1957; Rittschof et al., 1984), surface texture (Pomerat and Weiss, 1946; Gregg, 1948; Wethey, 1986), and other environmental factors to cyprid settlement (Barnes, 1970). Attached larvae quickly metamorphose to young adult forms and begin the sedentary phase of their life cycle (Doochin, 1951; Bernard and Lane, 1962; Walley, 1969). The strength of attachment of both cyprid larvae and postmetamorphic adult barnacles to a variety of substrates has been measured by several investigators. By attaching cyprids to an electrobalance and loading them in tension, Yule and Crisp (1983) determined that the strength of temporary attachment of exploring cyprid larvae of Semibalanus balanoides (L.) to slate panels exhibited forces in the range of 1.47 x 105 to 3.81 x 105 Newtons per square meter (Nm-2). The strength of temporary adhesion of exploring cyprids of S. balanoides to slate, glass, and several plastics was reported by Yule and Walker (1984b, 1987) to be in the range of 0.6-3 x 105 Nm-2, with larvae attaching more avidly to surfaces with inherently higher surface energies such as slate and glass. It has been argued that these ranges of forces do not support the view that suction is the underlying mechanism holding barnacle larvae in place (Yule and Crisp, 1983). The strength of adhesion of adult sessile

Sem observations on the interfacial surface of the cement of the adult barnacle, attached to natural and synthetic adherends

The barnacle, Chthamalus fragilis Darwin, is found attached to a variety of natural and man-made substrates in the marine environment. Bases of specimens of this species attached to the leaves and stalks of marsh grass, Spartina alterniflora, and to polyethylene films placed in the high intertidal zone of the marsh were studied by scanning electron microscopy in order to determine the interfacial features of the cement holding these animals in place. When removed carefully, the interfacial surfaces of the barnacle cement appeared as finely detailed replicas of the leaf surfaces and of the polyethylene film surfaces, mimicking the highly contoured surface features and peripheral interdigitations of the adjacent epithelial cells which compose the surfaces of the leaves and ‘crow's foot’ and linear perturbations in the relatively smooth polyethylene films. These observations indicate that the bioadhesive secretion of this barnacle species loses its liquidity and becomes a hardened adhesive cement. It is probable that this mold-like replication of adherend surface details contributes to barnacle adhesion by mechanical interaction between the barnacle cement and the adherend surfaces.

Genetic aspects of biodegradation by pseudomonads.

Microorganisms that are isolated on the basis of their ability to degrade natural or man-made organic substrates very often belong to the genus Pseudomonas. This may be because the pseudomonads have an extraordinary range of catabolic pathways, are easy to handle under laboratory conditions and generally grow on defined synthetic media. A single species such as P. cepacia utilizes more than 100 different substrates (out of 146 compounds tested) as the only C, N, or S source 22,98. Many recalcitrant chemicals (e.g., aromatics, chlorinated aliphatic compounds, pesticides) can be degraded by different Pseudomonas species or related soil bacteria isolated from nature. It is therefore pertinent to discuss the genetic organization of biodegradation functions in those Pseudomonas species that have been examined in some detail by genetic techniques. In addition, numerous methods that are potentially applicable to the study of biodegradation have been developed for Agrobacterium and Rhizobium genetics and some of these will be mentioned. Several reviews have been published describing the catabolic potential 22, the degradative plasmids 14,36 and chromosome maps 48 of Pseudomonas as well as the manipulation of soil bacteria by genetic engineering techniques 2, 36, 91. Microbial communities and cometabolism of organic substrates undoubtedly play an important role in nature 97. The geneticists, however, tend to shy away from such complex systems. They prefer pure cultures of distinct species and well-defined medium conditions allowing the selection of particular phenotypes. In the case of biodegradation pathways this means that a growth substrate should be utilized as the only C, N, S or P source. Occasionally, the disappearance of a colored substrate or the formation of a colored product may be helpful to monitor catabolic functions. Bacteria should grow on agar plates some strains with interesting biodegradation properties refuse to do that and allow at least one type of genetic transfer: conjugation, transduction, or transformation. The genetic analysis of bacteria used to be confined to members of the same or closely related species. In recent years, however, the scope of genetic research has greatly widened due to 3 major developments: a) Broad-host-range plasmids have been found that can replicate in virtually any gram-negative bacterium and enable the transfer of genetic information across the species barriers. b) The properties of transposons and other insertion elements may be used to produce insertion mutations and deletions, and mutated genes carrying a transposon insert may subsequently be isolated by recombinant DNA techniques. c) The gene cloning methods are universal in that the DNA from practically any organism can be inserted into cloning vectors. Several plasmid vectors that can be used in Pseudomonas species have been derived from broad-host-range plasmids.