Bacterial Diversity and Functions in Aerobic Phenoldegrading Granules

Aerobic granules can be viewed as a special form of biofilm, but without carriers for biofilm attachment. Growth environments for biofilm communities are different from planktonic communities, and micro-bial communities in attached biofilms have been shown to be highly distinct from the suspended biomass, even within a single reactor system. Recognizing the diversity and the linkages among the key functional groups in any given biological system can lead to better ways to model and understand diversity and function as well as to improve process stability. In a recent study, culture-independent and culture-dependent methods were used in combination to study the microbial community of aerobic phenol-degrading granules and to isolate, characterize, and identify ecologically relevant microorganisms (Jiang et al., 2004b).

The direct isolation technique was used to obtain bacterial colonies by incubating biomass from aerobic phenol-degrading granules on MP medium agar plates supplemented with 500 mg phenol l-1. A final set of ten strains, designated PG-01 to PG-10, was assembled after screening of 16S rRNA genes with REP-PCR (Table 9.1). Seven of the ten isolates belonged to the fi- or y-Proteobacteria group. These culture-based data are consistent with previous studies which demonstrated that fi- and y-Proteobacteria constitute a large fraction of the bacteria in wastewater treatment plants (Bond et al., 1995; Snaidr et al., 1997) or in glucose-fed aerobic granules (Tay et al., 2002). Members of fi-Proteobacteria have also been implicated in phenol degradation in activated sludge, as demonstrated in isolation experiments (Watanabe et al., 1998).

Another interesting observation was the prevalence of gram-positive high G+C bacteria in the phenol-degrading aerobic granules. In contrast, gram-positive high G+C bacteria were not dominant members in phenol-degrading activated sludge systems (Watanabe et al., 1998, 1999; Whiteley et al., 2001). These observations could probably be explained by the fact that high G+C bacteria preferred to grow in attached biofilms than to remain in a planktonic state (Lehman et al., 2001; Tresse et al., 2002). These microorganisms are also known to be resilient to external stresses, because of the presence of a strong cell envelope (Zhuang et al., 2003). In addition, several gram-positive high G+C strains are known to consume soluble COD (chemical oxygen demand) rapidly and store them as storage polymers to survive low nutrient conditions (Maszenan et al., 2000; Liu et al., 2001). These competitive traits can allow the gram-positive high G+C bacteria to thrive in the highly variable feast-famine situations encountered in the granulation systems, where phenol can be completely consumed within the first 30min of each 4 h cycle (Jiang et al., 2004a).

DGGE analysis of amplified 16S rRNA gene fragments from activated sludge, granules, and isolates showed that the dominant DGGE bands

Table 9.1. Characteristics of ten strains isolated from phenol-degrading granules

Isolates

Minimum cell density

Closest relative

Taxon. affiliation

16S rRNA

Number

(CFU g VSS-1

)

gene sequence identity (%)

of bases analyzed

PG-01

5.64 ± 0.87 x

1010

Pandoraea apista strain LMG 16407

$-Proteobacteria

98.7

1326

PG-02

1.01 ± 0.92 x

108

Propioniferax innocua ATCC 49929

Actinobacteria, HGC Gram positive bacteria,

93.5

1315

106

Propionibacteriacaea

PG-03

5.49 ± 1.80 x

Rhodococcus erythropolis strain HV1 00/50/6670

Actinobacteria, HGC Gram positive bacteria, Nocardioidaceae

99.8

1433

PG-04

3.05 ± 1.42 x

106

Propionibacterium cyclohexanicum strain IAM 14535

Actinobacteria, HGC Gram positive bacteria, Propionibacteriacaea

87.7

1370

PG-05

1.53 ± 1.37 x

107

Xenophilus azovorans KF46FT

$-Proteobacteria

98.8

1437

PG-06

2.55 ± 1.32 x

106

Acidovorax avenae ATCC 29625

$-Proteobacteria

97.9

1437

PG-07

1.93 ± 0.72 x

105

Xanthomonas axonopodis strain s53

y-Proteobacteria

98.1

1409

PG-08

3.56 ± 1.52 x

106

Comamonas sp. D22

$-Proteobacteria

97.0

1408

PG-09

7.62 ± 2.80 x

106

Pigmentiphaga

$-Proteobacteria

99.6

1432

PG-10

4.56 ± 1.72 x

106

Hydrogenophaga palleronii DSM 63

$-Proteobacteria

98.5

TO Ci

TO S

associated with the activated sludge did not co-migrate with the dominant bands from the granules (Fig. 9.2). However, DGGE bands associated with strains PG-01, PG-02, and PG-08 co-migrated with bands from the aerobic granules, which were found to have partial sequences that were identical to the sequences of the corresponding isolates. These three strains therefore represented dominant populations of $-Proteobacteria and gram-positive high G+C group within the granule community.

Additional experimental results provided independent evidence to support the contention that PG-01 was a numerically important microorganism in the aerobic granules (Jiang et al., 2004b). Fluorescent in situ hybridization (FISH) with confocal laser scanning microscopy (CLSM) was used to elucidate the abundance and spatial distribution of strain PG-01 in the aerobic granules (Fig. 9.3). The granules consisted of a dense layer of bacterial cells, surrounding a less dense central region. This structural pattern was repeatedly observed in all sections analyzed. Most PG-01 cells were distributed in clusters in the outer layers of the granules. Direct counting of probe-hybridized cells after disruption of granules revealed that PG-01 cells were numerically abundant in the granules, accounting for 4.1 ± 3.2% of all bacterial cells. Furthermore, PG-01 had a high specific growth rate and high specific phenol degradation rate and these attributes might have contributed significantly to PG-01's dominant role in phenol degradation in the granules.

Was this article helpful?

0 0

Post a comment