Populations of Anaerobic Phototrophic Bacteria in a Spartina alterniflora Salt Marsh.

Habitat-simulating media were used with the Hungate anaerobic roll tube technique to enumerate culturable anaerobic photosynthetic bacteria in sediment, tidal waters, and Spartina alterniflora plant samples collected from the salt marsh at Sapelo Island, Ga. No phototrophs were detected in samples of creekside (low marsh) sediment or in tidal waters in creekside regions. In the high marsh region, 90% of anaerobic phototrophic bacteria occurred in the top 5 mm of sediment and none were detected below 6 mm. There was a seasonal variation, with maximal populations occurring in summer and fall (mean, 4.4 x 10 phototrophs g of dry sediment) and minimal numbers occurring in winter (mean, 3.9 x 10 phototrophs g of dry sediment). During winter and late spring, phototrophs had a patchy distribution over the high marsh sediment surface. In contrast, during late summer they had a random uniform distribution. Tidal water collected over high marsh sediment contained an average of 8.7 x 10 phototrophs ml, with no significant seasonal variation. Anaerobic phototrophic bacteria were also cultured from the lower stem tissue of S. alterniflora growing in both the high (4.3 x 10 phototrophs g of dry tissue) and creekside (4.9 x 10 phototrophs g of dry tissue) marsh regions. Chromatium buderi, Chromatium vinosum, Thiospirillum sanguineum, Rhodospirillum molischianum, and Chlorobium phaeobacteroides were the predominant anaerobic phototrophic species cultured from high marsh sediment. The two Chromatium species were dominant.

Degradation of selected phenylurea herbicides by anaerobic pond sediment.

Anaerobic degradation of diuron [3-(3,4-dichlorophenyl)-1,1-dimethylurea], monuron [3-(4-chlorophenyl)-1,1-dimethylurea], and fenuron [1,1-dimethyl-3-phenylurea] were studied. Herbicide containing media (reduced with cysteine-HCl and under 95% N2:5% CO2 gas phase) were inoculated with pond sediments. Sediment from a diuron-treated pond dehalongenated diuron to 3-(3-chlorophenyl)-1,1-dimethylurea (CPDU) in 17 to 25 days but sterile sediment from the pond did not. Sediments from non-diuron treated ponds were also ineffective. Particles from diuron-treated sediment were essential for dehalogenation and they could not be replaced with other solid surfaces such as clay, sand or cellulose. Diuron was degraded by sediment at 25 and 30 degrees C (maximal rate) but not at 5, 15 and 37 degrees C after 55 days incubation. The product CPDU produced in laboratory cultures was found in sediment of the diuron-treated pond, indicating in situ reductive dechlorination. Sediment-inoculated cultures containing monuron and fenuron showed no degradation after 74 days incubation.

Populations of methane-producing bacteria and in vitro methanogenesis in salt marsh and estuarine sediments.

Most probable numbers (MPNs) of methanogens in various salt marsh and estuarine sediments were determined with an anaerobic, habitat-simulating culture medium with 80% H(2) plus 20% CO(2) as substrate. Average MPNs for the short Spartina (SS) marsh sediments of Sapelo Island, Ga., were maximal at the 5- to 7-cm depth (1.2 x 10/g of dry sediment). Populations decreased to approximately 880/g of dry sediment at the 34- to 36-cm depth. There was no significant difference between summer and winter populations. In tall Spartina (TS) marsh sediments, average populations were maximal (1.2 x 10/g of dry sediment) in the upper 0- to 2-cm zone; populations from the 5- to 36-cm zones were similar (average of 9 x 10/g of dry sediment). Methanogenic populations for TS sediments of James Island Creek marsh, Charleston, S.C., were similar (average of 3 x 10/g of dry sediment) for all depths tested (0 to 22 cm), which was comparable to the trend observed for TS sediments at Sapelo Island, Ga. Sediment grab samples collected along a transect of James Island Creek and its adjacent Spartina marsh had MPNs that were approximately 20 times greater for the region of Spartina growth (average of 10/g of dry sediment) compared with the channel (approximately 5 x 10 methanogens per g of dry sediment). A similar trend was found at Pawley's Island marsh, S.C., but populations were approximately one order of magnitude lower. In vitro rates of methanogenesis with SS sediments incubated under 80% H(2)-20% CO(2) showed that the 5- to 7-cm region exhibited maximal activity (51 nmol of CH(4) g h), which was greater than rates for sediments above and below this depth. SS sediment samples (5 to 7 cm) incubated under 100% N(2) and supplemented with formate exhibited rates of methanogenesis similar to those generated by samples under 80% H(2)-20% CO(2). Replacing the N(2) atmosphere with H(2) resulted in an eightfold decrease in the rate of methanogenesis. In vitro methanogenic activity by TS salt marsh sediments, incubated under 80% H(2)-20% CO(2), was similar for all depths tested (0 to 22 cm). TS sediment samples (0 to 7 cm) supplemented with formate and incubated under 100% N(2) had greater rates of methanogenesis compared with unsupplemented samples.

Enumeration of bacteriophages and host bacteria in sewage and the activated-sludge treatment process.

Bacteriophage populations in an activated-sludge sewage treatment plant were enumerated. A newly developed assay for quantitation of total phages, employing direct electron microscopic counts, was used in conjunction with the plaque assay. The total concentration of phages was significantly higher in reactor mixed liquor and effluent than in influent sewage, indicating a net production of phages within the reactor. Maximum total phage concentrations in the fluid phase of sewage, activated-sludge mixed liquor, and reactor effluent were 2.2 x 10(7), 9.5 x 10(7), and 8.4 x 10(7)/ml, respectively. Conditions were optimized for isolation of predominant heterotrophic aerobic bacteria from sewage and mixed liquor. Blending at ice water temperatures was superior to ultrasound or enzyme treatments for maximum release of viable bacteria from microbial floc. A solidified extract of mixed liquor was superior to standard media for cultivating maximum numbers of heterotrophic bacteria. The highest culture counts for sewage and mixed liquor were 1.4 x 10(7) and 1.3 x 10(9)/ml, respectively, which represented only 3 and 6.8% of the total microscopic cell counts. Only 3 out of 48 dominant bacterial isolates from either mixed liquor or sewage were hosts for phages present in the system. The sum of phage populations infecting these three hosts accounted for, at best, 3.8% (sewage) and 0.2% (mixed liquor) of the total number of phages present. Generally, specific phage titers were lower in mixed liquor than in sewage, indicating that these hosts were not responsible for the net production of phages in the reactor. This study emphasizes the limitations of the plaque assay for ecological studies of phages, and it suggests that bacteria responsible for phage production in activated-sludge mixed liquor are either minor components of the heterotrophic population, floc-producing strains, or members of other physiological groups.

Technique for determining total bacterial virus counts in complex aqueous systems.

A direct method is described for measuring bacteriophage concentrations in complex aqueous systems. Conditions for sample clarification, phage recognition, and recovery were optimized. In contrast to the plaque assay, this procedure permits quantitation of total numbers of phages independent of bacterial host. Also, the modifications increase the sensitivity of the sedimentation assay, permitting detection of particles at a minimum concentration of 10(4) per ml. Maximal total phage concentrations in the aqueous phase of sewage and activated sludge mixed liquor were 1.3 x 10(6) and 4.3 x 10(7) per ml, respectively.

Some morphological types of bacteriophages in bovine rumen contents.

Six morphological types of bacteriophage were found in bovine rumen contents. Minimal total phage count was 5 x 10(7) per ml of rumen fluid.

Characterization of Methanobacterium mobilis, sp. n., isolated from the bovine rumen.

A methanogenic bacterium, present in bovine rumen contents at concentrations of approximately 2 x 10(8) cells per ml, has been isolated in pure culture. The organism is a strictly anaerobic, weakly motile, nonsporeforming, gram-negative rod (0.7 mu x 1.5 to 2.0 mu) with rounded ends. There is a single polar flagellum. The organism grows at temperatures between 30 and 45 C, with an optimum at 40 C, and at pH values between 5.9 and 7.7, with optimal growth between pH 6.1 and 6.9. Of the 17 substrates tested, only formate and H(2) plus CO(2) supported growth. An unidentified, heat-stable factor(s) was required by the organism. The factor, which was not one of the common ones, was present in rumen fluid, mixed rumen bacteria, and yeast extract. On the basis of colony morphology, Gram reaction, and motility, the organism is classified as a new species of methanogenic bacterium, and the name Methanobacterium mobilis sp. n. is proposed.