Prosthecomicrobium and Ancalomicrobium: New Prosthecate Freshwater Bacteria

Direct microscopic examination of natural freshwater samples reveals a variety of small microorganisms having elaborate cellular appendages. Several strains have been isolated from crude cultures containing low concentrations of organic nutrients. All of the isolates are procaryotic. They are aerobic chemoorganotrophs that require vitamins for growth. Because they cannot be assigned to any of the existing bacterial genera, two new genera are proposed: Ancalomicrobium for organisms which have several long appendages and which reproduce by budding; Prosthecomicrobium for organisms which have many short appendages tapering toward a blunt tip and which reproduce by binary fission. Gas vacuoles have been found in strains of each genus. The term prostheca is proposed for the rigid appendages of procaryotic cells bounded by the cell wall, and is defined to include the structures on these new bacteria, as well as the stalks of the caulobacters and the hyphae of the hyphomicrobia.     

High Numbers of Prosthecate Bacteria in Pulp Mill Waste Aeration Lagoons

Prosthecate bacteria comprised 0.6 to 10.5% of the bacterial community in samples from 11 pulp mill waste aeration lagoons. Because of their distinct morphology, the genera Ancalomicrobium, Caulobacter, Prosthecobacter, Prosthecomicrobium, Stella, and Hyphomicrobium or Hyphomonas could be identified and enumerated by direct microscopic examination. Monthly samples from one lagoon showed that several genera varied from undetectable to predominant among the appendaged organisms. Temperature (season), type of wood pulped, and pulping process did not significantly affect the density of prosthecate bacteria.     

Deoxyribonucleic Acid Base Sequence Homologies of Some Budding and Prosthecate Bacteria

The genetic relatedness of a number of budding and prosthecate bacteria was determined by deoxyribonucleic acid (DNA) homology experiments of the direct binding type. Strains of Hyphomicrobium sp. isolated from aquatic habitats were found to have relatedness values ranging from 9 to 70% with strain "EA-617," a subculture of the Hyphomicrobium isolated by Mevius from river water. Strains obtained from soil enrichments had lower values with EA-617, ranging from 3 to 5%. Very little or no homology was detected between the amino acid-utilizing strain Hyphomicrobium neptunium and other Hyphomicrobium strains, although significant homology was observed with the two Hyphomonas strains examined. No homology could be detected between prosthecate bacteria of the genera Rhodomicrobium, Prosthecomicrobium, Ancalomicrobium, or Caulobacter, and Hyphomicrobium strain EA-617 or H. neptunium LE-670. The grouping of Hyphomicrobium strains by their relatedness values agrees well with a grouping according to the base composition of their DNA species. It is concluded that bacteria possessing cellular extensions represent a widely diverse group of organisms.     

Environmental Factors Correlated with Size of Bacterial Populations in a Polluted Stream

Samples of water were taken from a polluted zone of the Gallinas River and analyzed as to numbers of total bacteria, coliforms, and fecal streptococci. Environmental factors measured were temperature, pH and concentrations of detergent, nitrate plus nitrite nitrogen, sulfate, chloride, bicarbonate, and phosphate. Thirty-two observations were made from 12 March through 22 July 1971. Stepwise multiple linear regression analyses of the data were carried out by computer to determine which of the environmental factors were significantly correlated with numbers of bacteria present. A multiple linear regression equation was constructed for each bacteriological parameter as a function of significant variables only. Log total bacteria was correlated positively with bicarbonate, phosphate, and detergent concentrations. Log coliforms was correlated positively with phosphate and sulfate concentrations and negatively with chloride concentration. Log fecal streptococci was correlated positively with bicarbonate and chloride concentrations.     

A Study of Ciliate Protozoa from a Small Polluted Stream in East-Central Illinois

The role of species of ciliate Protozoa in trickling filterwaste treatment processes awaits redefinition. Using the smallfresh-water Salt Fork stream for our study, a stream which receivesall of the discharge from the Champaign-Urbana Waste TreatmentPlant in east-central Illinois, analyses of samples of carefullyidentified ciliates taken from selected sites over a three-yearperiod yielded some interesting results. Four specific situationswere noted: (1) free-swimming microphagous forms feeding selectivelyon primarily attached bacteria, and, in turn, being fed on bycarnivorous ciliates; (2) sessile ciliates (peritrichs) feedingon non-attached bacterial species, but with high selectivity;(3) free-swimming carnivorous gymnostomes feeding on the sessileperitrichs but, again in turn, serving as prey themselves tostalked suctorian species attached to the peduncles of the peritrichs;(4) attached peritrichs and suctorians both serving as foodfor several groups of small invertebrates frequenting the stream.With respect to numbers and diversity of ciliates found, some155 different species, belonging to 93 genera, were identified(including several to be described elsewhere as new species)in the stream and/or the trickling filter settling basin. Thehighest populations and greatest diversity were found at sitesclosest to the waste water influx; such protozoan populationsmight thus be considered as a form of tertiary treatment inthe drainage basin.     

Screening and Isolation of PHB-Producing Bacteria in a Polluted Marine Microbial Mat

The characteristics of microbial mats within the waste stream from a seafood cannery were compared to a microbial community at a pristine site near a sandy beach at Puerto San Carlos, Baja California Sur, Mexico. Isolation of poly-beta-hydroxybutyrate (PHB)-producing bacteria, recognition of brightly refractile cytoplasmatic inclusions, lipophilic stains with Sudan Black and Nile Red, and chemical extraction of PHB were used as a culture-dependent strategy for the detection of PHB-producing bacteria. The culture-independent approach included denaturing gradient gel electrophoresis of phylotypes of 16S rRNA of microbial communities from environmental samples. Significant differences in community structure were found among the polluted and pristine sites. These differences were correlated with the physicochemical characteristics of the seawater column. At the polluted site, the seawater was rich in nutrients (ammonia, phosphates, and organic matter), compared to the pristine location. Partial sequencing of 16S rDNA of cultures of bacteria producing PHB included Bacillus and Staphylococcus at both sites; Paracoccus and Micrococcus were found only at the polluted site and Rhodococcus and Methylobacterium were found only at the pristine site. Bands of the sequences of 16S rDNA from both field samples in the denaturing gradient gel electrophoresis (DGGE) analyses affiliated closely only with bacterial sequences of cultures of Bacillus and Staphylococcus. High concentrations of organic and inorganic nutrients at the polluted site had a clear effect on the composition and diversity of the microbial community compared to the unpolluted site.     

Contribution of Fungi and Bacteria to Leaf Litter Decomposition in a Polluted River

The contribution of fungi and bacteria to the decomposition of alder leaves was examined at two reference and two polluted sites in the Ave River (northwestern Portugal). Leaf mass loss, microbial production from incorporation rates of radiolabeled compounds into biomolecules, fungal biomass from ergosterol concentration, sporulation rates, and diversity of aquatic hyphomycetes associated with decomposing leaves were determined. The concentrations of organic nutrients and of inorganic nitrogen and phosphorus in the stream water was elevated and increased at downstream sites. Leaf decomposition rates were high (0.013 day⁻¹ < k < 0.042 day⁻¹), and the highest value was estimated at the most downstream polluted site, where maximum values of microbial production and fungal biomass and sporulation were found. The slowest decomposition occurred at the other polluted site, where, along with the nutrient enrichment, the lowest current velocity and dissolved-oxygen concentration in water were observed. At this site, fungal production, biomass, and sporulation were depressed, suggesting that stimulation of fungal activity by increased nutrient concentrations might be offset by other factors. Although bacterial production was higher at polluted sites, fungi accounted for more than 94% of the total microbial net production. Fungal yield coefficients varied from 10.2 to 13.6%, while those of bacteria were less than 1%. The contribution of fungi to overall leaf carbon loss (29.0 to 38.8%) greatly exceeded that of bacteria (4.2 to 13.9%).     

Diversity of Fungi, Bacteria, and Actinomycetes on Leaves Decomposing in a Stream

Although fungi, bacteria, and specific bacterial taxa, such as the actinomycetes, have been studied extensively in various habitats, few studies have examined them simultaneously, especially on decomposing leaves in streams. In this study, sugar maple and white oak leaves were incubated in a stream in northeastern Ohio for 181 days during which samples were collected at regular intervals. Following DNA extraction, PCR-denaturing gradient gel electrophoresis (DGGE) was performed using fungus-, bacterium-, and actinomycete-specific primers. In addition, fungal and bacterial biomass was estimated. Fungal biomass differed on different days but not between leaves of the two species and was always greater than bacterial biomass. There were significant differences in bacterial biomass through time and between leaf types on some days. Generally, on the basis of DGGE, few differences in community structure were found for different leaf types. However, the ribotype richness of fungi was significantly greater than those of the bacteria and actinomycetes, which were similar to each other. Ribotype richness decreased toward the end of the study for each group except bacteria. Lack of differences between the two leaf types suggests that the microorganisms colonizing the leaf biofilm were primarily generalists that could exploit the resources of the leaves of either species equally well. Thus, we conclude that factors, such as the ecological role of the taxa (generalists versus specialists), stage of decay, and time of exposure, appeared to be more important determinants of microbial community structure than leaf quality.     

Recovery in Diversity of Fish and Invertebrate Communities Following Remediation of a Polluted Stream: Investigating Causal Relationships

Spatial and temporal responses of biota to anthropogenic disturbance were measured over a 15 year period in a contaminated stream undergoing remediation and recovery. Along the spatial gradient of the stream, levels of contaminants decreased downstream along with improved responses of instream biota at several levels of biological organization. Recovery of the biota in this stream over the 15 year study period is demonstrated by the temporal relationships between levels of decreasing contaminants and the concomitant responses of the periphyton, macroinvertebrate, and fish communities and changes in the various bioindicators of individual fish health. Decreases in contaminants over a temporal scale were followed closely by an improvement in physiological and organismal-level indicators, increases in the diversity of macroinvertebrate and fish communities, and rapid increases in the chlorophyll a biomass and photosynthesis rate of the periphyton community. These results emphasize that field studies designed to assess and evaluate the effectiveness of restoration activities on stream recovery should incorporate a variety of response endpoints ranging from sensitive and short-term responses to long-term but ecological relevant indicators of change. The close spatial and temporal relationships observed between changes in physicochemical factors and positive responses in various components of the stream biota over the 15-year study period suggest a strong cause and effect relationship between remediation activities and stream recovery. Understanding causal relationships and the mechanistic processes between environmental stressors, stress responses of biota, and the recovery process is important in the effective management and restoration of aquatic ecosystems. An erratum to this article is available at .     

Plasmid Incidence in Bacteria from Deep Subsurface Sediments

Bacteria were isolated from deep terrestrial subsurface sediments underlying the coastal plain of South Carolina. A total of 163 isolates from deep sediments, surface soil, and return drill muds were examined for plasmid DNA content and resistance to the antibiotics penicillin, ampicillin, carbenicillin, streptomycin, kanamycin, and tetracycline. MICs of Cu²⁺, Cr³⁺, and Hg²⁺ for each isolate were also determined. The overall frequency of plasmid occurrence in the subsurface bacteria was 33%. Resistance was most frequent to penicillin (70% of all isolates), ampicillin (49%), and carbenicillin (32%) and was concluded to be related to the concentrations of the individual antibiotics in the disks used for assaying resistance and to the production of low levels of β-lactamase. The frequencies of resistance to penicillin and ampicillin were significantly greater for isolates bearing plasmids than for plasmidless isolates; however, resistance was not transferable to penicillin-sensitive Escherichia coli. Hybridization of subsurface bacterial plasmids and chromosomal DNA with a whole-TOL-plasmid (pWWO) probe revealed some homology of subsurface bacterial plasmid and chromosomal DNAs, indicating a potential for those bacteria to harbor catabolic genes on plasmids or chromosomes. The incidences of antibiotic resistance and MICs of metals for subsurface bacteria were significantly different from those for drill mud bacteria, ruling out the possibility that bacteria from sediments were derived from drill muds.     

Nitrogen Transformations and Diversity of Ammonia-Oxidizing Bacteria in a Desert Ephemeral Stream Receiving Untreated Wastewater

Levels of inorganic nitrogen species (ammonia, nitrite, and nitrate), ammonia oxidation potential (AOP), and diversity of ammonia-oxidizing bacteria (AOB) were studied in the sediments of a 50-km-long segment of an ephemeral stream in the Negev desert, receiving untreated wastewater. Water analysis in downstream sampling points showed reductions of 91.7% in biological oxygen demand, 87.7% in chemical oxygen demand, 73.9% in total nitrogen, and 72.8% in total ammonia nitrogen. Significant AOP levels in the sediment were detected mainly in the fall and spring seasons. Denaturing gradient gel electrophoresis of AOB 16S rRNA gene fragments showed that in most sampling points, the streambed was dominated by Nitrosospira cluster 3 strains similar to those dominating the stream bank’s soils and sediments in nearby springs. Nitrosomonas strains introduced by discharged wastewater and others dominated some sections of the stream characterized by high organic carbon levels. The results suggest that climatic conditions in the Negev desert select for AOB belonging to Nitrosospira cluster 3, and these conditions dominate the aquatic environment effect along most of the stream sections. In addition, the nitrification–denitrification processes were not sufficient to reduce nitrogen levels in the sediment and prevent the eutrophication of some sections of the stream ecosystem. Thus, the discharge of high nitrogen wastewater into desert streams should be done carefully as it may endanger the already fragile ecosystem.     

Proliferation of Ciprofloxacin Resistant Bacteria in Polluted Sediments of Musi River,India

Improper disposal of domestic sewage and effluents from drug manufacturing units for several years has resulted in the accumulation of pollutants in Musi river sediments. There were no studies carried out before to quantify the antibiotic resistance in this river, despite the fact that its sediments are loaded with antibiotics. The present study investigated the relationship between the proliferation of ciprofloxacin resistant culture with the occurrence of fluoroquinolone and heavy metals in sediments of Musi River. The fluoroquinolones concentration in river sediments were observed in high concentration (13336.4 ng/g) and were found to be positively correlated with the occurrence of ciprofloxacin resistant bacteria (r = 0.386 to 0.675, p < 0.05). The occurrence of heavy metals was also in positive correlation with the distribution of antibiotic resistant bacteria in the river (r = 0.454 to 0.881, p < 0.05). This study indicated the spread of antibiotic resistance in polluted river sediments that might pose a serious threat to public health as the river water is used for irrigation, drinking, and recreational purposes; and needs an immediate risk assessment and mitigation strategies.     

Q-RT-PCR for Assessing Archaea, Bacteria, and Fungi During Leaf Decomposition in a Stream

Leaf disks of Tilia cordata were exposed for up to 5 weeks in a first-order stream in Nova Scotia, Canada. The exponential decay rate k was 0.008 day(-1). Ergosterol levels increased linearly to a maximum of 134 mug g(-1) dry leaf mass. Release of conidia peaked at 700 day(-1) mg(-1) on leaves that had been exposed for 3 weeks; after 5 weeks, it declined to 15 mg(-1). In total, 23 taxa of aquatic hyphomycetes were distinguished. Anguillospora filiformis contributed over 76% of the conidia during weeks 1, 2, and 3, and 16.5% in week 5. Three sets of primers specific for Bacteria, Archaea, and Fungi were applied in quantitative real-time polymerase chain reaction (Q-RT-PCR) to estimate relative DNA amounts. Archaeal DNA was consistently present at low levels. Bacterial and fungal DNA peaked between weeks 2 and 3, and declined in week 5. With the exception of week 1, fungal DNA exceeded bacterial DNA by between 12 and 110%.     

Culturability of Stream Bacteria Assessed at the Assemblage and Population Levels

Lotic bacterial communities can be examined at multiple levels: from the assemblage level to populations of individual species. In stream environments, as in many other systems, the percentage of bacteria that are culturable is quite low. In this study, the culturability of the overall bacterial assemblage, as well as the culturability of three common species (Acinetobacter calcoaceticus, Burkholderia cepacia, and Pseudomonas putida), was determined in samples collected from four streams on three dates. Colony hybridization (colonies were grown on modified nutrient agar) and fluorescent in situ hybridization were used to calculate the percentage of cells of a given species that were culturable. Approximately half of the overall assemblage was estimated to be viable but nonculturable cells (VBNC). The culturability of two of the species was low (0.29% for A. calcoaceticus and 0.46% for P. putida), whereas the value for B. cepacia (2.48%) exceeded the overall assemblage level culturability (0.90%). Overall, both bacterial assemblages and populations were dominated by VBNC. These results show quantitatively that not all members of a species that has culturable representatives are culturable when retrieved from natural populations, likely because of interspecific phenotypic and genotypic variability. Thus, the large pool of nonculturable cells includes representatives of species that are, under some circumstances, culturable.