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The Frankia sp. strain R82 genome is described as representative of a novel candidate species within Frankia cluster 1, as indicated by average nucleotide identity (ANI) analyses, with its closest relatives being Frankia nodulisporulans AgTrs and strains Ag45/Mut15 and AgPM24 (86% identity).

The genomes of two nitrogen-fixing

In this study, we describe the genomes of two novel candidate species of non-nitrogen fixing Frankia that were isolated from the root nodules of Coriaria nepalensis and Alnus glutinosa, genospecies CN and Ag, respectively. Comparative genomic analyses revealed that both genospecies lack genes essential for nitrogen-fixation and possess genes involved in the degradation of plant cell walls. Additionally, we found distinct biosynthetic gene clusters in each genospecies. The availability of these genomes will contribute to the study of the taxonomy and evolution of actinorhizal symbioses.

The effect of host plants on the abundance and distribution of introduced and indigenous Frankia populations was assessed in soils and root nodules of four alder species, Alnus glutinosa,Alnus cordata,Alnus rubra and Alnus viridis. Plants were grown in microcosms with either a sandy soil without detectable frankiae, with or without inoculation of a mixture of Frankia isolates, or a silty clay loam soil with indigenous Frankia. The presence of frankiae in soils increased plant height and root nodule formation, with significant increases in the presence of indigenous frankiae. Abundance in soils increased significantly for both introduced and indigenous Frankia populations independent of alder species, with generally largest increases in cluster 1b frankiae. Root nodules formed by introduced frankiae did not reflect the diversity of strains inoculated, with nodules generally only formed by strain ArI3 representing cluster 1a/d. All indigenous Frankia populations detected in soil were also found in A. glutinosa nodules, while A. cordata or A. rubra nodules contained different subsets of frankiae with unique abundances dependent on plant species. These results demonstrate the intrageneric differences of host plants in the selection of specific Frankia populations in soils for root nodule formation.

Head-starting of the federally endangered Houston toad (Anaxyrus houstonensis), that is, the release of egg strands, tadpoles, and metamorphic juveniles produced in captivity into the original breeding ponds, requires assessment of potential threats for the transmission of pathogens from captive to free-ranging toads. We used Illumina-based 16S rRNA V3 amplicon sequencing to investigate the community structure of bacteria from skin lesions of captive Houston toad and habitat (pond) samples. Proteobacteria, alone or together with Actinobacteria and, in some samples, Cyanobacteria represented virtually all reads in tissue lesion samples, whereas pond samples were much more diverse, with Acidobacteria, Actinobacteria, Bacteriodetes, Chloroflexi, Cyanobacteria, Firmicutes, Planctomycetes, Proteobacteria, and Verrucomicrobia present with little variation between samples. If present in lesions, Actinobacteria were largely represented by Mycobacteriaceae, and here mainly by one sequence identical to sequences of members of the Mycobacterium chelonae-abscessus complex. In pond samples, mycobacteria represented only a small portion of the actinobacteria, although at higher diversity with six distinct reads. Sequences for reads obtained from pond samples were identical to those representing the M. chelonae-abscessus complex, a group with Mycobacterium marinum, Mycobacterium kansasii, Mycobacterium avium, a group with Mycobacterium vaccae, Mycobacterium fortuitum, Mycobacterium poriferae, and a group with Mycobacterium elephantis and Mycobacterium celeriflavum, whereas sequences of high similarity were detected for reads related to those of Mycobacterium holsaticum, Mycobacterium pallens, and Mycobacterium obuense, and Mycobacterium goodii. Our results indicated that lesions observed on the Houston toad in captivity are not the result of mycobacteria in every case, and that the presence of mycobacteria in the captive colony does not represent a novel pathogen threat to the wild populations because such bacteria are also seen in the natural pond habitats for the Houston toad.

Illumina-based 16S rRNA V3 amplicon sequencing of total DNA obtained from soft tissue lesions (joint granulomas) of the endangered Houston toad (Anaxyrus houstonensis) demonstrated that many reads represented members of the actinobacterial Mycobacterium chelonae-abscessus complex. In order to quantify members of this complex in those lesions, we designed three complex-specific primer set/probe combinations (sets I, II and III) targeting variable regions on the 23S rRNA gene for SybrGreen- and Taqman-based quantitative polymerase chain reaction (qPCR). Both SybrGreen- and Taqman-based analyses specifically detected members of the M. chelonae-abscessus complex in lesion samples, with numbers between 10

Members of the nitrogen-fixing actinobacterial genus Frankia are typically isolated from root nodules and generally infective on the same plant species. Several Frankia strains originally isolated from Casuarina species, however, have been found to be non-infective on Casuarina species. The goal of this study was to investigate the potential role of infective isolates from Casuarina species on the potential establishment of these non-infective Frankia strains in root nodule formation on Casuarina equisetifolia. Soil microcosms were established with plants of C. equisetifolia and inoculated with Frankia casuarinae strain CcI3 or cluster 3 strain R43, or combinations of both at different densities. Basic plant growth characteristics, root nodule formation and localization of both Frankia strains in nodule periderm and cortex, as well as population development in soils were monitored. The presence of strain R43 did not affect plant growth performance nor root nodule formation, while inoculation with strain CcI3 enhanced plant growth and resulted in root nodule formation. qPCR analyses on selected nodule lobes revealed the presence of strain CcI3 in cortex samples in all treatments, while strain R43 was not detected in any cortex samples but in 40% of the periderm samples from lobes from treatments with highest inoculation values. In situ hybridization detected cells of strain R43 on the outside of the nodules, i.e. on the periderm only. These results demonstrate that the cluster 3 Frankia strain R43 is not co-infecting root nodules formed by the F. casuarinae strain CcI3 on C. equisetifolia, but has likely been isolated as a surface contaminant from Casuarina nodules.

Rodent species were assessed as potential hosts of Trypanosoma cruzi, the etiologic agent of Chagas disease, from five sites throughout Texas in sylvan and disturbed habitats. A total of 592 rodents were captured, resulting in a wide taxonomic representation of 11 genera and 15 species. Heart samples of 543 individuals were successfully analyzed by SybrGreen-based quantitative PCR (qPCR) targeting a 166 bp fragment of satellite DNA of T. cruzi. Eight rodents representing six species from six genera and two families were infected with T. cruzi. This is the first report of T. cruzi in the pygmy mouse (Baiomys taylori) and the white-footed mouse (Peromyscus leucopus) for the USA. All infected rodents were from the southernmost site (Las Palomas Wildlife Management Area). No differences in pathogen prevalence existed between disturbed habitats (5 of 131 tested; 3.8%) and sylvan habitats (3 of 40 tested; 7.5%). Most positives (n = 6, 16% prevalence) were detected in late winter with single positives in both spring (3% prevalence) and fall (1% prevalence). Additionally, 30 Triatoma insects were collected opportunistically from sites in central Texas. Fifty percent of these insects, i.e., 13 T. gerstaeckeri (68%), and two T. lecticularia (100%) were positive for T. cruzi. Comparative sequence analyses of 18S rRNA of samples provided identical results with respect to detection of the presence or absence of T. cruzi and assigned T. cruzi from rodents collected in late winter to lineage TcI. T. cruzi from Triatoma sp. and rodents from subsequent collections in spring and fall were different, however, and could not be assigned to other lineages with certainty.

Targeted Illumina sequencing of nitrogenase reductase (nifH) gene fragments and analyses of pair-end reads through a modified QIIME pipeline were used to assess the diversity of the actinomyceteous genus Frankia in three soils. Soils were vegetated with host or non-host plants, and included locations in Illinois (ABA, host), Colorado (CoMt, non-host), and Wisconsin (FMWI, non-host). After filtering, seven unique sequences were recovered for soil ABA, six for CoMt, and four sequences for FMWI. These sequences were included in a Bayesian topology anchored by published sequence data from pure cultures of Frankia. Sequences from all three soils showed affinities to Frankia strains from both the Alnus and Elaeagnus host infection groups. Reads representing Casuarina-infective strains were not detected. Four sequences from soil CoMt and five sequences from soil ABA did not cluster, at 97% similarity, into a shared OTU that contained a cultured relative. These results demonstrate that targeted Illumina sequencing provides an efficient and economical method for assessing haplotype diversity of ecofunctional genes (e.g. nifH) at the genus level in microorganisms that perform important ecosystem functions.

Freshwater turtles from 28 species representing 6 families, categorized as “wild,” “captive,” or “wild-caught, captive-raised,” were analyzed for the presence of intraerythrocytic, parasitic protozoa. Microscopic analyses of blood smears from 327 individual turtles revealed parasites in the blood of 29% of these individuals (n = 96), with levels of parasitemia ranging from 0.003% to 2.1%. SYBR ® Green–based quantitative polymerase chain reaction confirmed these results with 29% overall prevalence (94 of 324), with 66% prevalence in the Geoemydidae, 26% prevalence in the Emydidae, and 23% prevalence in the Kinosternidae. No infections were detected in members of the Chelidae, Pelomedusidae, and Trionychidae. Prevalence was highest in wild turtles with 67%–100% from 7 locations, followed by wild-caught, captive-raised turtles with 4%–26% from 2 locations, while detections in captive turtles were zero in 2 locations and 3% in the remaining third location. Comparative sequence analyses of 583-bp amplicons of 18S rRNA gene fragments allowed us to identify 80 Haemogregarina infections and one Hemolivia infection. Parasites representing the genus Hepatozoon were not detected. Because parasites were generally detected in wild turtles and only rarely in captive turtles, captive breeding with subsequent release of turtles would seem to pose little risk to free-ranging turtles with regard to the spread of these parasites. On the contrary, wild turtles would be more likely sources of parasite transmission to captive populations if used in breeding programs.

Chytridiomycosis, an emerging infectious disease caused by the fungal pathogenBatrachochytrium dendrobatidis(Bd), has caused amphibian population declines worldwide.Bdwas first described in the 1990s and there are still geographic gaps in the genetic analysis of this globally distributed pathogen. Relatively few genetic studies have focused on regions whereBdexhibits low virulence, potentially creating a bias in our current knowledge of the pathogen’s genetic diversity. Disease-associated declines have not been recorded in Texas (USA), yetBdhas been detected on amphibians in the state. These strains have not been isolated and characterized genetically; therefore, we isolated, cultured, and genotypedBdfrom central Texas and compared isolates to a panel of previously genotyped strains distributed across the Western Hemisphere. We also isolated other chytrids not known to infect amphibians from east Texas. To identify larval amphibian hosts, we sequenced part of the COI gene. Among 37Bdisolates from Texas, we detected 19 unique multi-locus genotypes, but found no genetic structure associated with host species, Texas localities, or across North America. Isolates from central Texas exhibit high diversity and genetically cluster withBd-GPL isolates from the western U.S. that have caused amphibian population declines. This study genetically characterizes isolates ofBdfrom the south central U.S. and adds to the global knowledge ofBdgenotypes.

The prevalence of salmonellae in the intestines of the invasive suckermouth catfish Hypostomus plecostomus was assessed in the San Marcos River, just down-stream of its spring-fed headwaters. In 2014, H. plecostomus, sediment, and water samples were collected during 15 sampling events. A combination of semi-selective enrichment and quantitative polymerase chain reaction (qPCR) revealed the presence of salmonellae in 45% of the fish intestines across the entire year, with a prevalence range of 13–100% per sampling event. Repetitive element sequence-based PCR (rep-PCR) and multi-locus sequence typing (MLST) revealed a high diversity of salmonellae from fish intestine samples at individual sampling times, single or multiple presence of rep-PCR patterns and serotypes within individual fish, and identical rep-PCR patterns and serotypes for different fish within and across sampling events. Overall, 15 serotypes were identified by MLST, with a diversity range between one and seven serotypes per sampling event. Some serotypes were retrieved only once, while others were detected more frequently. A few serotypes were retrieved at several sampling times, nearly evenly distributed over the entire sampling period. Prevalence and diversity were independent of precipitation events, indicating the potential presence of environmental strains that are capable of long-term persistence in the environment.

Actinorhizal plants form nitrogen-fixing root nodules in symbiosis with soil-dwelling actinobacteria within the genus Frankia , and specific Frankia taxonomic clusters nodulate plants in corresponding host infection groups. In same-soil microcosms, we observed that some host species were nodulated ( Alnus glutinosa , Alnus cordata , Shepherdia argentea , Casuarina equisetifolia ) while others were not ( Alnus viridis , Hippophaë rhamnoides ). Nodule populations were represented by eight different sequences of nifH gene fragments. Two of these sequences characterized frankiae in S. argentea nodules, and three others characterized frankiae in A. glutinosa nodules. Frankiae in A. cordata nodules were represented by five sequences, one of which was also found in nodules from A. glutinosa and C. equisetifolia , while another was detected in nodules from A. glutinosa . Quantitative PCR assays showed that vegetation generally increased the abundance of frankiae in soil, independently of the target gene (i.e., nifH or the 23S rRNA gene). Targeted Illumina sequencing of Frankia -specific nifH gene fragments detected 24 unique sequences from rhizosphere soils, 4 of which were also found in nodules, while the remaining 4 sequences in nodules were not found in soils. Seven of the 24 sequences from soils represented >90% of the reads obtained in most samples; the 2 most abundant sequences from soils were not found in root nodules, and only 2 of the sequences from soils were detected in nodules. These results demonstrate large differences between detectable Frankia populations in soil and those in root nodules, suggesting that root nodule formation is not a function of the abundance or relative diversity of specific Frankia populations in soils. IMPORTANCE The nitrogen-fixing actinobacterium Frankia forms root nodules on actinorhizal plants, with members of specific Frankia taxonomic clusters nodulating plants in corresponding host infection groups. We assessed Frankia diversity in root nodules of different host plant species, and we related specific populations to the abundance and relative distribution of indigenous frankiae in rhizosphere soils. Large differences were observed between detectable Frankia populations in soil and those in root nodules, suggesting that root nodule formation is not a function of the abundance or relative diversity of specific Frankia populations in soils but rather results from plants potentially selecting frankiae from the soil for root nodule formation. These data also highlight the necessity of using a combination of different assessment tools so as to adequately address methodological constraints that could produce contradictory data sets.

A considerable amount of information is available on root nodule formation of the nitrogen fixing actinomycete Frankia and its host plants; however, studies on Frankia populations in soil, its second ecological niche, are generally hampered by the complexity of soil systems and the lack of adequate quantification methods for frankiae. The aim of this study was, using recently developed quantitative PCR methods, to follow population dynamics of introduced Frankia strains in soil microcosms under different edaphic conditions. Populations dynamics of four Frankia strains (ArI3, Ag45/Mut15, CcI3 and EAN1pec) representing frankiae of the Alnus host infection group (i.e. subgroups I and II infective on Alnus species, Casuarina-infective frankiae) and frankiae of the Elaeagnus host infection group, respectively, were analyzed by quantitative PCR (qPCR) in a factorial design of microcosms with two plant species (A. glutinosa, C. equisetifolia), two potential carbon resources (rhizosphere, leaf litter), and two matric potentials (wet, dry). Introduction of frankiae into soil resulted in about 10-fold declines in abundance within the first 6 weeks of incubation in both bulk and rhizosphere soils, independently of plant species, leaf litter amendment and matric potential. While this decline continued in bulk soil during an additional 6 week period, numbers in the rhizosphere recovered and increased to values close to- or exceeding those of introduced frankiae. Independently of plant species, carbon resource or matric potential, cell numbers of strain EAN1pec recovered to inoculum values in nearly all treatment combinations, while those of strain ArI3 remained low. Strain Ag45/Mut15 recovered most prominently in the rhizosphere in leaf-litter-amended microcosms, regardless of plant species and matric potential, while cell numbers of Casuarina-infective Frankia strain CcI3 recovered only in the presence of C. equisetifolia, with higher numbers in the presence of leaf litter and under dry conditions. These results demonstrate differential effects of environmental conditions including plant species, carbon resources, and matric potentials on the fate of specific Frankia strains in soil, and identifies factors potentially affecting subpopulations of indigenous frankiae under natural conditions.

Recent models suggest a relationship exists between community diversity and pathogen prevalence, the proportion of individuals in a population that are infected by a pathogen, with most inferences tied to assemblage structure. Two contrasting outcomes of this relationship have been proposed: the “dilution effect” and the “amplification effect.” Small mammal assemblage structure in disturbed habitats often differs from assemblages in sylvan environments, and hantavirus prevalence is often negatively correlated with habitats containing high species diversity via dilution effect dynamics. As species richness increases, prevalence of infection often is decreased. However, anthropogenic changes to sylvan landscapes have been shown to decrease species richness and/or increase phylogenetic similarities within assemblages. Between January 2011 and January 2016, we captured and tested 2406 individual small mammals for hantavirus antibodies at 20 sites across Texas and México and compared differences in hantavirus seroprevalence, species composition, and assemblage structure between sylvan and disturbed habitats. We found 313 small mammals positive for antibodies against hantaviruses, evincing an overall prevalence of 9.7% across all sites. In total, 40 species of small mammals were identified comprising 2 taxonomic orders (Rodentia and Eulipotyphla). By sampling both habitat types concurrently, we were able to make real-world inferences into the efficacy of dilution effect theory in terms of hantavirus ecology. Our hypothesis predicting greater species richness higher in sylvan habitats compared to disturbed areas was not supported, suggesting the characteristics of assemblage structure do not adhere to current conceptions of species richness negatively influencing prevalence via a dilution effect.

The nodule-forming actinobacterial genus Frankia can generally be divided into 4 taxonomic clusters, with clusters 1, 2, and 3 representing nitrogen-fixing strains of different host infection groups and cluster 4 representing atypical, generally non-nitrogen-fixing strains. Recently, quantitative PCR (qPCR)-based quantification methods have been developed for frankiae of clusters 1 and 3; however, similar approaches for clusters 2 and 4 were missing. We amended a database of partial 23S rRNA gene sequences of Frankia strains belonging to clusters 1 and 3 with sequences of frankiae representing clusters 2 and 4. The alignment allowed us to design primers and probes for the specific detection and quantification of these Frankia clusters by either Sybr Green- or TaqMan-based qPCR. Analyses of frankiae in different soils, all obtained from the same region in Illinois, USA, provided similar results, independent of the qPCR method applied, with abundance estimates of 10 × 10 5 to 15 × 10 5 cells (g soil) −1 depending on the soil. Diversity was higher in prairie soils (native, restored, and cultivated), with frankiae of all 4 clusters detected and those of cluster 4 dominating, while diversity in soils under Alnus glutinosa , a host plant for cluster 1 frankiae, or Betula nigra , a related nonhost plant, was restricted to cluster 1 and 3 frankiae and generally members of subgroup 1b were dominating. These results indicate that vegetation affects the basic composition of frankiae in soils, with higher diversity in prairie soils compared to much more restricted diversity under some host and nonhost trees. IMPORTANCE Root nodule formation by the actinobacterium Frankia is host plant specific and largely, but not exclusively, correlates with assignments of strains to specific clusters within the genus. Due to the lack of adequate detection and quantification tools, studies on Frankia have been limited to clusters 1 and 3 and generally excluded clusters 2 and 4. We have developed tools for the detection and quantification of clusters 2 and 4, which can now be used in combination with those developed for clusters 1 and 3 to retrieve information on the ecology of all clusters delineated within the genus Frankia . Our initial results indicate that vegetation affects the basic composition of frankiae in soils, with higher diversity in prairie soils compared to much more restricted diversity under some host and nonhost trees.

The 23S rRNA gene was evaluated as target for the development of Sybr Green-based quantitative PCR (qPCR) for the analysis of nitrogen-fixing members of the genus Frankia or subgroups of these in soil. A qPCR with a primer combination targeting all nitrogen-fixing frankiae (clusters 1, 2 and 3) resulted in numbers similar to those obtained with a previously developed qPCR using nifH gene sequences, both with respect to introduced and indigenous Frankia populations. Primer combinations more specifically targeting three subgroups of the Alnus host infection group (cluster 1) or members of the Elaeagnus host infection group (cluster 3) were specific for introduced strains of the target group, with numbers corresponding to those obtained by quantification of nitrogen-fixing frankiae with both the 23S rRNA and nifH genes as target. Method verification on indigenous Frankia populations in soils, i.e. in depth profiles from four sites at an Alnus glutinosa stand, revealed declining numbers in the depth profiles, with similar abundance of all nitrogen-fixing frankiae independent of 23S rRNA or nifH gene targets, and corresponding numbers of one group of frankiae of the Alnus host infection only, with no detections of frankiae representing the Elaeagnus, Casuarina, or a second subgroup of the Alnus host infection groups.

We investigated the effect of different nitrogen (N) sources on exopolysaccharide (EPS) production and composition by Frankia strain CcI3, a N2-fixing actinomycete that forms root nodules with Casuarina species. Frankia cells grown in the absence of NH4Cl (i.e., under N2-fixing conditions) produced 1.7-fold more EPS, with lower galactose (45.1 vs. 54.7 mol%) and higher mannose (17.3 vs. 9.7 mol%) contents than those grown in the presence of NH4Cl as a combined N-source. In the absence of the combined N-source, terminally linked and branched residue contents were nearly twice as high with 32.8 vs. 15.1 mol% and 15.1 vs. 8.7 mol%, respectively, than in its presence, while the content of linearly linked residues was lower with 52.1 mol% compared to 76.2 mol%. To find out clues for the altered EPS production at the transcriptional level, we performed whole-gene expression profiling using quantitative reverse transcription PCR and microarray technology. The transcription profiles of Frankia strain CcI3 grown in the absence of NH4Cl revealed up to 2 orders of magnitude higher transcription of nitrogen fixation-related genes compared to those of CcI3 cells grown in the presence of NH4Cl. Unexpectedly, microarray data did not provide evidence for transcriptional regulation as a mechanism for differences in EPS production. These findings indicate effects of nitrogen fixation on the production and composition of EPS in Frankia strain CcI3 and suggest posttranscriptional regulation of enhanced EPS production in the absence of the combined N-source.

The diversity of Salmonella was analysed in biofilm and water samples from the spring and slough arms of Spring Lake, the headwaters of the San Marcos River, Texas, during the drought of 2011, with only one potential run-off event at the beginning of the study. Salmonellae were detected in semiselective enrichment cultures by end-point PCR during the entire sampling period of (11 sampling events during 2 months). From the spring arm site, 73% of the biofilms and 41% of the water samples were positive for salmonellae, while only 9% of the biofilms and 23% of the water samples were positive from the slough arm site. Salmonellae could be isolated from all positive samples, with higher diversity in biofilms compared with water samples and more strains obtained from the spring arm than from the slough arm. Differences between sites were generally caused by less frequently detected isolates, while the majority of isolates that were present in both biofilms and water from both sites was represented by three strains only. Quantification attempts by quantitative PCR directly in samples without prior enrichment did not result in a reliable detection of salmonellae, suggesting that numbers in all samples were below the detection limit.

Patterns of spatial variation of mercury and methylmercury (MeHg) were examined in sediments and muscle tissue of largemouth bass (Micropterus salmoides) from Amistad International Reservoir, a large and hydrologically complex subtropical water body in the Rio Grande drainage. The distributions of both Hg and MeHg were compared with environmental and biological factors known to influence production of MeHg. The highest concentrations of total Hg (THg) in sediment were found in the Rio Grande arm of the reservoir, whereas MeHg was highest at sites in the Devils River arm and inundated Pecos River (often more than 3.0 ng/g). Conditions in the sediments of the Devils River arm and Pecos River channel were likely more favorable to the production of MeHg, with higher sediment porewater dissolved organic carbon, and porewater sulfate levels in the optimal range for methylation. Although the detection of different groups of sulfate-reducing bacteria by polymerase chain reaction (PCR) was generally correlated with MeHg concentrations, bacterial counts via fluorescent in situ hybridization (FISH) did not correlate with MeHg. A sample of 156 largemouth bass (30 cm) showed a spatial pattern similar to that of MeHg in sediments, where fish from the Devils River arm of the reservoir had higher muscle Hg concentrations than those collected in the Rio Grande arm. In 88 bass of legal sport fishing size (35 cm), 77% exceeded the 0.3 mg/kg U.S. Environmental Protection Agency screening value. This study shows that significant variation in sediment MeHg and biotic Hg concentration can exist within lakes and reservoirs and that it can correspond to variation in environmental conditions and Hg methylation.

The diversity and distribution of salmonellae in freshwater biofilms were analyzed at a fine scale (i.e. in 20 locations from a 324 cm(2) area) for two sites in San Marcos, TX. A concrete storm water overflow channel (City Park) was sampled 4 times and a concrete surface in the spring-fed headwaters of the San Marcos River (Spring Lake) 5 times between April and September 2009, and each biofilm sample analyzed by a combination of traditional enrichment methods and molecular techniques. PCR detection of the invA gene, that encodes a protein of a type III secretion system present in salmonellae, after semi-selective enrichment of salmonellae was achieved in biofilms from all 20 locations at the City Park site, with locations generally being positive 2-3 times out of 4 sampling times for a total of 59% positive samples. InvA gene fragment detection in biofilms was less frequent for the 5 sampling times and 20 locations from the Spring Lake site (18% of all samples), with 1 sampling time being entirely negative and 8 locations remaining negative throughout the study. Rep-PCR fingerprinting of 491 Salmonella isolates obtained from both sites resulted in 30 distinct profiles, with 26 and 7 profiles retrieved from City Park and Spring Lake samples, respectively, and thus with 3 profiles present at both sites, and multiple strains frequently obtained from single locations at both sites. The composition of Salmonella strains in the area analyzed changed in time with large differences between early (April, June) and late sampling times (September) within and among sites, except for one strain (S12) that was present at almost all sampling times at both sites, though often at different locations within the area analyzed. These results demonstrate the presence of salmonellae in natural biofilms and a significant micro-heterogeneity with differences in diversity and persistence of salmonellae.

Two isolates, designated CadH11T and Cad448T, representing uncultured purple sulfur bacterial populations H and 448, respectively, in the chemocline of Lake Cadagno, a crenogenic meromictic lake in Switzerland, were obtained using enrichment and isolation conditions that resembled those used for cultured members of the genus Thiocystis. Phenotypic, genotypic and phylogenetic analyses of these isolates confirmed their assignment to the genus Thiocystis. However, 16S rRNA gene sequence similarities of 98.2 % between CadH11T and Cad448T, and similarities of 97.7 and 98.5 %, respectively, with their closest cultured relative Thiocystis gelatinosa DSM 215T, as well as differences in DNA G+C content and carbon source utilization suggested that the isolates belonged to two distinct species. DNA–DNA hybridization of CadH11T and Cad448T with T. gelatinosa DSM 215T showed relatedness values of 46.4 and 60.8 %, respectively; the relatedness value between CadH11T and Cad448T was 59.2 %. Based on this evidence, strains CadH11T and Cad448T represent two novel species within the genus Thiocystis, for which the names Thiocystis chemoclinalis sp. nov. and Thiocystis cadagnonensis sp. nov. are proposed, respectively. The type strains of T. chemoclinalis sp. nov. and T. cadagnonensis sp. nov. are CadH11T ( = JCM 15112T = KCTC 5954T) and Cad448T ( = JCM 15111T = KCTC 15001T), respectively.

Salmonellae are pathogenic bacteria often detected in waters impacted by human or animal wastes. In order to assess the fate of salmonellae in supposedly pristine environments, water and natural biofilm samples along with snails (Tarebia granifera) and crayfish (Procambarus clarkia) were collected before and up to 7 days following four precipitation events from sites within the headwater springs of Spring Lake, San Marcos, TX. The samples were analyzed for the presence of salmonellae by polymerase chain reaction (PCR) after semi-selective enrichment. Salmonellae were detected in one water sample directly after precipitation only, while detection in ten biofilm and two crayfish samples was not related to precipitation. Salmonellae were not detected in snails. Characterization of isolates by rep-PCR revealed shared profiles in water and biofilm samples, biofilm and crayfish samples, and biofilm samples collected 23 days apart. These results suggest that salmonellae are infrequently washed into this aquatic ecosystem during precipitation runoff and can potentially take up residency in biofilms which can help facilitate subsequent long-term persistence and eventual transfer through the food chain.

Matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry (MS) was evaluated as a technique to characterize strains of the nitrogen-fixing actinomycete Frankia. MALDI-TOF MS reliably distinguished 37 isolates within the genus Frankia and assigned them to their respective host infection groups, i.e., the Alnus/Casuarina and the Elaeagnus host infection groups. The assignment of individual strains to sub-groups within the respective host infection groups was consistent with classification based on comparative sequence analysis of nifH gene fragments, confirming the usefulness of MALDI-TOF MS as a rapid and reliable tool for the characterization of Frankia strains.

The genetic diversity of Frankia populations in soil and in root nodules of sympatrically grown Alnus taxa was evaluated by rep-polymerase chain reaction (PCR) and nifH gene sequence analyses. Rep-PCR analyses of uncultured Frankia populations in root nodules of 12 Alnus taxa (n = 10 nodules each) growing sympatrically in the Morton Arboretum near Chicago revealed identical patterns for nodules from each Alnus taxon, including replicate trees of the same host taxon, and low diversity overall with only three profiles retrieved. One profile was retrieved from all nodules of nine taxa (Alnus incana subsp. incana, Alnus japonica, Alnus glutinosa, Alnus incana subsp. tenuifolia, Alnus incana subsp. rugosa, Alnus rhombifolia, Alnus mandshurica, Alnus maritima, and Alnus serrulata), the second was found in all nodules of two plant taxa (A. incana subsp. hirsuta and A. glutinosa var. pyramidalis), and the third was unique for all Frankia populations in nodules of A. incana subsp. rugosa var. americana. Comparative sequence analyses of nifH gene fragments in nodules representing these three profiles assigned these frankiae to different subgroups within the Alnus host infection group. None of these sequences, however, represented frankiae detectable in soil as determined by sequence analysis of 73 clones from a Frankia-specific nifH gene clone library. Additional analyses of nodule populations from selected alders growing on different soils demonstrated the presence of different Frankia populations in nodules for each soil, with populations showing identical sequences in nodules from the same soil, but differences between plant taxa. These results suggest that soil environmental conditions and host plant genotype both have a role in the selection of Frankia strains by a host plant for root nodule formation, and that this selection is not merely a function of the abundance of a Frankia strain in soil.

Clone libraries of nifH gene fragments specific for the nitrogen-fixing actinomycete Frankia were generated from six soils obtained from five continents using a nested PCR. Comparative sequence analyses of all libraries (n=247 clones) using 96 to 97% similarity thresholds revealed the presence of three and four clusters of frankiae representing the Elaeagnus and the Alnus host infection groups, respectively. Diversity of frankiae was represented by fewer clusters (i.e., up to four in total) within individual libraries, with one cluster generally harboring the vast majority of sequences. Meta-analysis including sequences previously published for cultures (n=48) and for uncultured frankiae in root nodules of Morella pensylvanica formed in bioassays with the respective soils (n=121) revealed a higher overall diversity with four and six clusters of frankiae representing the Elaeagnus and the Alnus host infection groups, respectively, and displayed large differences in cluster assignments between sequences retrieved from clone libraries and those obtained from nodules, with assignments to the same cluster only rarely encountered for individual soils. These results demonstrate large differences between detectable Frankia populations in soil and those in root nodules indicating the inadequacy of bioassays for the analysis of frankiae in soil and the role of plants in the selection of frankiae from soil for root nodule formation.

Seasonal variation of arbuscular mycorrhizal fungi (AMF) in roots of the high salt marsh plant Spartina patens, the diversity of nitrogen-fixing bacteria in the rhizosphere and plant growth performance was studied at key stages of the growing season coinciding with major plant phenological stages, i.e., vegetative growth, reproduction and senescence. AMF colonization was highest during vegetative growth, with values declining during the growing season to the same level seen at plant dormancy. AMF colonization was reduced at lower depths in the sediments where anoxic conditions were observed and in plants treated with the systemic fungicide Benomyl. Only small changes in diversity of nitrogen-fixing bacteria in general and more specifically of those belonging to the e-subdivision of Proteobacteria were detected during the season or between treatments by PCR-RFLP of nifH gene fragments with DNA as template for amplification; however, greater seasonal changes were displayed when cDNA was used as template for amplification as a proxy for gene expression and thus active bacteria. DGGE analyses of nifH gene fragments representing nitrogen-fixing bacteria of the e-subdivision of Proteobacteria using both using DNA and cDNA as template showed highly diverse profiles that changed during the season and in response to treatment. Seasonal changes were observed for a suite of plant growth attributes and differences were observed between treatments, with higher values generally obtained on non-treated plants compared to Benomyl-treated plants. These differences were most pronounced during vegetative growth; however, differences between non-treated and Benomyl-treated plants were reduced seasonally and disappeared by the onset of senescence. This study demonstrates seasonal changes in AMF colonization on S. patens and in the community structure of nitrogen-fixing members of the e-subdivision of Proteobacteria in the plant root zone. Plant growth performance changed seasonally with some effects of Benomyl-treatment.

The ability of Frankia strains to grow in the rhizosphere of a nonactinorhizal plant, Betula pendula, in surrounding bulk soil and in soil amended with leaf litter was analyzed 6 weeks after inoculation of pure cultures by in situ hybridization. Growth responses were related to taxonomic position as determined by comparative sequence analysis of nifH gene fragments and of an actinomycetes-specific insertion in Domain III of the 23S rRNA gene. Phylogenetic analyses confirmed the basic classification of Frankia strains by host infection groups, and allowed a further differentiation of Frankia clusters within the Alnus host infection group. Except for Casuarina-infective Frankia strains, all other strains of the Alnus and the Elaeagnus host infection groups displayed growth in the rhizosphere of B. pendula, and none of them grew in the surrounding bulk soil that was characterized by a very low organic matter content. Only a small number of strains that all belonged to a distinct phylogenetic cluster within the Alnus host infection group grew in soil amended with ground leaf litter from B. pendula. These results demonstrate that saprotrophic growth of frankiae is a common trait for most members of the genus, and the supporting factors for growth (i.e. carbon utilization capabilities) varied with the host infection group and the phylogenetic affiliation of the strains.

Bioassays with Morella pensylvanica as capture plant and comparative sequence analyses of nifH gene fragments of Frankia populations in nodules formed were used to investigate the diversity of Frankia in soils over a broad geographic range, i.e., from sites in five continents (Africa, Europe, Asia, North America, and South America). Phylogenetic analyses of 522-bp nifH gene fragments of 100 uncultured frankiae from root nodules of M. pensylvanica and of 58 Frankia strains resulted in a clear differentiation between frankiae of the Elaeagnus and the Alnus host infection groups, with sequences from each group found in all soils and the assignment of all sequences to four and five clusters within these groups, respectively. All clusters were formed or dominated by frankiae obtained from one or two soils with single sequences occasionally present from frankiae of other soils. Variation within a cluster was generally low for sequences representing frankiae in nodules induced by the same soil, but large between sequences of frankiae originating from different soils. Three clusters, one within the Elaeagnus and two within the Alnus host infection groups, were represented entirely by uncultured frankiae with no sequences from cultured relatives available. These results demonstrate large differences in nodule-forming frankiae in five soils from a broad geographic range, but low diversity of nodule-forming Frankia populations within any of these soils.

The potential role of host plant species in the selection of symbiotic, nitrogen-fixing Frankia strains belonging to the Elaeagnus host infection group was assessed in bioassays with two Morella, three Elaeagnus, and one Shepherdia species as capture plants, inoculated with soil slurries made with soil collected from a mixed pine/grassland area in central Wisconsin, USA. Comparative sequence analysis of nifH gene fragments amplified from homogenates of at least 20 individual lobes of root nodules harvested from capture plants of each species confirmed the more promiscuous character of Morella cerifera and Morella pensylvanica that formed nodules with frankiae of the Alnus and the Elaeagnus host infection groups, while frankiae in nodules formed on Elaeagnus umbellata, Elaeagnus angustifolia, Elaeagnus commutata, and Shepherdia argentea generally belonged to the Elaeagnus host infection group. Diversity of frankiae of the Elaeagnus host infection groups was larger in nodules on both Morella species than in nodules formed on the other plant species. None of the plants, however, captured the entire diversity of nodule-forming frankiae. The distribution of clusters of Frankia populations and their abundance in nodules was unique for each of the plant species, with only one cluster being ubiquitous and most abundant while the remaining clusters were only present in nodules of one (six clusters) or two (two clusters) host plant species. These results demonstrate large effects of the host plant species in the selection of Frankia strains from soil for potential nodule formation and thus the significant effect of the choice of capture plant species in bioassays on diversity estimates in soil.

Sediment, water, and fish gut samples taken at three sites near the headwaters of the San Marcos River, Texas, were analyzed for salmonellae Salmonella spp. by culture and molecular techniques. While enrichment cultures from sediment and water samples from the two uppermost sites were negative for salmonellae in polymerase chain reaction analyses, both sediment and water samples were positive at the downstream site. At all sites, salmonellae were present in the guts of different fishes (e.g., largemouth bass Micropterus salmoides, channel catfish Ictalurus punctatus, common carp Cyprinus carpio, and suckermouth catfish Hypostomus plecostomus). The highest percentage of detection (33% of analyzed fish) occurred at the downstream site, whereas detection percentages at the upper two sites were 18% and 17%. Detection of salmonellae was usually limited to one segment of the gut (i.e., upper or lower part). Serovars were highly variable among individuals and differed between the upper and lower gut in the only individual (a common carp) that had salmonellae in both gut segments. In situ hybridization demonstrated that salmonellae were normally associated with particulate material in the gut and occurred in highly variable numbers ranging from an occasional organism to a majority of the gut microbe population. These results demonstrate the presence of different serovars of potentially human pathogenic salmonellae among four ecologically distinct fishes within natural environments. They also suggest that salmonellae are not components of the indigenous microbial community in fish intestines but rather are ingested with particulate material.

Information on the importance of captive turtles as sources of human Salmonella infections is well established; however, data on the potential of free-ranging turtles as carriers of salmonellae are scarce and contradictory. We combined traditional culture techniques and molecular tools to analyze swabs obtained from the cloacae and from different body parts of captive and free-ranging turtles for salmonellae. Salmonellae were detected in 50% of captive turtles (N = 10). A similar percentage of detection (51%) was obtained for salmonellae in free-ranging turtles from the Rio Grande (N = 80) with six sites at Big Bend National Park, Texas, and one site at Elephant Butte Reservoir, New Mexico, analyzed. Here, 46% of Trachemys gaigeae (N = 36), 56% of Apalone spinifera (N = 43), and the only individual of Chrysemys picta were positive for salmonellae. These percentages of detection of salmonellae in turtles were independent of the location of the sampling in the Rio Grande, the species and the gender of the turtles. Although individuals of captive turtles testing positive for salmonellae were generally positive for all body parts tested (i.e., the cloacae, the carapace, the ventral base of the left rear leg, underneath one or more of the claws on the front feet, and the ventral base of the tail), individuals of free-ranging turtles testing positive for salmonellae were often positive for only one or two body parts. Our results demonstrate that salmonellae are prevalent in high rates in both captive and free-ranging turtles.

Based on phylogenetic analysis of clones retrieved from two nifH gene clone libraries that were created using cDNA from suboxic sediment samples obtained from areas densely vegetated with the high-salt marsh plant Spartina patens , a primer set was designed to target nitrogen-fixing bacteria with sequence similarities to members of the ε subclass of Proteobacteria . Nested PCR, denaturing gel electrophoresis, and subsequent sequence analysis of reamplified fragments confirmed the specificity of the primer set by retrieving nifH sequences of only putative members of the ε subclass of Proteobacteria , all of which were characterized by a highly divergent 27- or 36-bp insertion in both DNA and cDNA.

The fate and effects of selected heavy metals were examined in sediment from a restored salt marsh. Sediment cores densely covered with Spartina patens were collected and kept either un-amended or artificially amended with nickel (Ni) under standardized greenhouse conditions. Ni-amendment had no significant effect on the fate of other metals in sediments, however, it increased root uptake of the metals. Metal translocation into the shoots was small for all metals. Higher Ni concentrations in plants from amended cores were accompanied by seasonal reductions in plant biomass, photosynthetic capacity and transfer efficiency of open photosystem II reaction centers; these effects, however, were no longer significant at the end of the growing season. Root colonization by arbuscular mycorrhizal fungi (AMF) resembled that of natural salt marshes with up to 20% root length colonized. Although Ni-amendment increased AMF colonization, especially during vegetative growth, in general AMF were largely unaffected.

Sediments and water from the pristine headwaters of the San Marcos River, Texas, USA, as well as swabs from biofilms on the carapace and from the cloacae of 17 musk turtles (Sternotherus odoratus) and one snapping turtle (Chelydra serpentina serpentina) caught at the same site, were analysed for salmonellae by culture and molecular techniques. Whereas enrichment cultures from sediment and water samples were negative for salmonellae in PCR- and in situ hybridization-based analyses, both techniques detected salmonellae after enrichments from both carapace and cloacae of nine (i.e. of 53%) musk turtles. Further characterization of 10 isolates obtained from the enrichment cultures of four selected individuals and confirmed as salmonellae by PCR analysis was achieved by fingerprinting techniques (rep-PCR). The results show differences between individuals and, in one case, variation among isolates from a single individual. All isolates from two individuals displayed identical profiles. These profiles were different from those obtained from the isolates of the third individual, which were, themselves, also identical for all isolates. Salmonellae were much more diverse in samples from the carapace of the last individual with five different rep-PCR profiles retrieved. Serotyping of seven isolates representative for each rep-PCR profile identified all isolates as representing Salmonella enterica subspecies enterica serotype Rubislaw, which demonstrates the presence of different strains of potentially human pathogenic salmonellae naturally occurring on turtles even within pristine environments. The frequent detection of these organisms in biofilms on the carapace opens the door for speculations on the role of this habitat as a reservoir for salmonellae, and on potential implications for turtles acting as a dispersal vector.

The effect of a synthetic iron(III)-porphyrin meso-tetra(2,6-dichloro-3-sulfonatophenyl)porphyrinate as a biomimetic catalyst in the oxidative treatment of 2,4-dichlorophenol (2,4-DCP) with humic substances and H2O2 was evaluated in factorial design experiments conducted at different concentrations of 2,4-DCP (0–25 ppm) and different incubation treatment times (0, 24, 96, or 120 h). In the absence of this treatment, bioassays with the bacterium Pseudomonas putida (ATCC11250) showed decreasing specific growth rates μ (used here to quantify 2,4-DCP toxicity) with increasing concentrations of 2,4-DCP. However, when 2,4-DCP was treated as mentioned above the toxicity of the resulting 2,4-DCP solution was reduced significantly. At low 2,4-DCP concentrations (up to 5 ppm) and long incubation periods (as long as 120 h), the specific growth rate μ was comparable to that of cultures grown in the absence of 2,4-DCP. The reduction in toxicity was directly correlated to a decrease in the concentration of 2,4-DCP in the treated solutions, as measured by high-performance liquid chromatography. The reduced concentrations of 2,4-DCP in the treated solutions could be correctly predicted based on the relationship between the specific growth rates and the 2,4-DCP concentrations in untreated solutions. These results indicate that the oxidative coupling of 2,4-DCP to humic substances catalyzed by the synthetic iron(III)-porphyrin catalyst in the presence of H2O2 is responsible for the removal of 2,4-DCP from solutions. This approach appears to be a promising alternative treatment to reduce 2,4-DCP bioavailability and thus toxicity in the environment.

Population analyses in water samples obtained from the chemocline of crenogenic, meromictic Lake Cadagno, Switzerland, in October for the years 1994 to 2003 were studied using in situ hybridization with specific probes. During this 10-year period, large shifts in abundance between purple and green sulfur bacteria and among different populations were obtained. Purple sulfur bacteria were the numerically most prominent phototrophic sulfur bacteria in samples obtained from 1994 to 2001, when they represented between 70 and 95% of the phototrophic sulfur bacteria. All populations of purple sulfur bacteria showed large fluctuations in time with populations belonging to the genus Lamprocystis being numerically much more important than those of the genera Chromatium and Thiocystis . Green sulfur bacteria were initially represented by Chlorobium phaeobacteroides but were replaced by Chlorobium clathratiforme by the end of the study. C. clathratiforme was the only green sulfur bacterium detected during the last 2 years of the analysis, when a shift in dominance from purple sulfur bacteria to green sulfur bacteria was observed in the chemocline. At this time, numbers of purple sulfur bacteria had decreased and those of green sulfur bacteria increased by about 1 order of magnitude and C. clathratiforme represented about 95% of the phototrophic sulfur bacteria. This major change in community structure in the chemocline was accompanied by changes in profiles of turbidity and photosynthetically available radiation, as well as for sulfide concentrations and light intensity. Overall, these findings suggest that a disruption of the chemocline in 2000 may have altered environmental niches and populations in subsequent years.

Lake Cadagno, a crenogenic meromictic lake located in the catchment area of a dolomite vein rich in gypsum in the Piora Valley in the southern Alps of Switzerland, is characterized by a compact chemocline with high concentrations of sulfate, steep gradients of oxygen, sulfide and light and a turbidity maximum that correlates to large numbers of bacteria (up to 107 cells ml-1). The most abundant taxa in the chemocline are large- and small-celled purple sulfur bacteria, which account for up to 35% of all bacteria, and sulfate- reducing bacteria that represent up to 23% of all bacteria. Depending on the season, as much as 45% of all bacteria in the chemocline are associated in aggregates consisting of different populations of small-celled purple sulfur bacteria of the genus Lamprocystis (up to 35% of all bacteria) and sulfate-reducing bacteria of the family Desulfobulbaceae (up to 12% of all bacteria) that are almost completely represented by bacteria closely related to Desulfocapsa thiozymogenes. Their association in aggregates is restricted to small-celled purple sulfur bacteria of the genus Lamprocystis, but not obligate since non-associated cells of bacteria related to D. thiozymogenes are frequently found, especially under limited light conditions in winter and early summer. Aggregate formation and concomitant growth enhancement of isolates of both partners of this association suggests synergistic interactions that might resemble a sulfide-based source-sink relationship between the sulfate-reducing bacterium that is able to sustain growth by a disproportionation of inorganic sulfur compounds (sulfur, thiosulfate, sulfite), with the purple sulfur bacteria acting as a biotic scavenger. The availability of these isolates opens up the door for future studies considering other facets of potential interactions in aggregates since both types of organisms are metabolically highly versatile and interactions may not be limited to sulfur compounds only.

The interaction between the salt marsh plant Spartina patens , arbuscular mycorrhizal fungi (AMF) and bacteria in salt marsh sediment was examined in a long-term arbuscular mycorrhizas (AM) suppression study by applying the systemic fungicide benomyl to field-collected sediment cores with and without S. patens plants. Microbial populations were sampled four times corresponding to major plant phenological stages (dormancy, vegetative growth, reproduction, and senescence) previously linked to changes in microbial populations under field conditions. Benomyl-treatment of soil cores significantly suppressed AM colonization on S. patens , keeping values relatively consistent throughout the growing season (11.5%) whereas plants in non-treated cores experienced seasonal increases and declines in AM colonization (26.6% during vegetative growth to 11.5% during dormancy). Soil physicochemical parameters were not affected by benomyl application. In unvegetated cores, no benomyl- or seasonal effects were displayed by cell numbers and specific biomass of DAPI-stained organisms, members of the domain bacteria and here especially members of the α-, β-, γ- and δ-subdivisions of proteobacteria that were the most abundant bacterial groups. In vegetated cores, the microbial community as well as specific bacterial populations were at least twice as large in terms of number and biomass than in samples from unvegetated cores with significant seasonal changes for DAPI-stained cells, for members of the domain bacteria and for members of the α- and γ-subdivisions of proteobacteria. In benomyl-treated cores, the population of γ-subdivision of proteobacteria was significantly smaller than in non-treated cores, and a positive association was found between this bacterial group and root length colonized by AM suggesting that AM-suppression can affect populations of specific soil bacterial populations in salt marsh sediment. Benomyl-treatment had no effect on the diversity of N-fixing bacteria as evidenced by PCR-RFLP analysis, but seasonal changes were noted in vegetated cores with populations during active plant growth substantially different from populations during dormancy and senescence.

Sulfate-reducing bacteria were analyzed in the chemocline of meromictic Lake Cadagno, Switzerland, in March, June, August and October using in situ hybridization. Numbers of sulfate-reducing bacteria determined as the sum of cells hybridizing to Probes SRB385Db targeting Desul- fobacteriaceae and SRB385 targeting Desulfovibrionaceae were similar at all samplings and accounted for 13 to 18% of the total microbial community, respectively. Abundance of cells detected with either probe, however, changed during the year, with cell numbers detected with SRB385Db being larger in early summer (June) and those detected with SRB385 being larger in late summer (October). Increasing cell numbers detected with SRB385Db were mainly caused by a yet uncultured and phylogenetically unidentified bacterium with a peculiar morphology ('Morphotype R') that followed the sulfide profile in June as well as in August with increasing numbers at increasing concentrations with depth. From the fraction of cells detected with SRB385, only a minor part rep- resented a yet uncultured population without close cultured relatives. At all samplings, the majority of cells detected with SRB385 (93 to 99%) represented populations phylogeneti- cally related to Desulfocapsa thiozymogenes DSM7269. Their cells were generally found in association with aggregates of small-celled phototrophic sulfur bacteria. This association was not specific for 1 of the 4 populations representing all small-celled phototrophic sulfur bacteria in Lake Cadagno. The association was also not obligate since non-associated cells were frequently found, especially in winter and early summer when limited light conditions caused by snow and ice cover reduced the abundance of small-celled phototrophic sulfur bacteria to about 50% of the values found in late summer. Nonetheless, the association between populations related to D. thiozymogenes and small-celled phototrophic sulfur bacteria suggests an ecological advantage to both organisms under appropriate environmental conditions.