Novel acsF Gene Primers Revealed a Diverse Phototrophic Bacterial Population, Including Gemmatimonadetes, in Lake Taihu (China)

Abstract

Anoxygenic phototrophs represent an environmentally important and phylogenetically diverse group of organisms. They harvest light using bacteriochlorophyll-containing reaction centers. Recently, a novel phototrophic bacterium, Gemmatimonas phototrophica, belonging to a rarely studied phylum, Gemmatimonadetes, was isolated from a freshwater lake in the Gobi Desert. To obtain more information about the environmental distribution of phototrophic Gemmatimonadetes, we collected microbial samples from the water column, upper sediment, and deeper anoxic sediment of Lake Taihu, China. MiSeq sequencing of the 16S rRNA, pufM, and bchY genes was carried out to assess the diversity of local phototrophic communities. In addition, we designed new degenerate primers of aerobic cyclase gene acsF, which serves as a convenient marker for both phototrophic Gemmatimonadetes and phototrophic Proteobacteria Our results showed that most of the phototrophic species in Lake Taihu belong to Alpha- and Betaproteobacteria Sequences of green sulfur and green nonsulfur bacteria (phototrophic Chlorobi and Chloroflexi, respectively) were found in the sediment. Using the newly designed primers, we identified a diverse community of phototrophic Gemmatimonadetes forming 30 operational taxonomic units. These species represented 10.5 and 17.3% of the acsF reads in the upper semiaerobic sediment and anoxic sediment, whereas their abundance in the water column was <1%.

Importance: Photosynthesis is one of the most fundamental biological processes on Earth. Recently, the presence of photosynthetic reaction centers has been reported from a rarely studied bacterial phylum, Gemmatimonadetes, but almost nothing is known about the diversity and environmental distribution of these organisms. The newly designed acsF primers were used to identify phototrophic Gemmatimonadetes from planktonic and sediment samples collected in Lake Taihu, China. The Gemmatimonadetes sequences were found mostly in the upper sediments, documenting the preference of Gemmatimonadetes for semiaerobic conditions. Our results also show that the phototrophic Gemmatimonadetes present in Lake Taihu were relatively diverse, encompassing 30 operational taxonomic units.

FIG 1

(A) UPGMA clustering and relative abundance of top 10 phyla in three samples based on 16S rRNA sequences. “Others” refers to all phyla other than the top ten. (B) Clustering and heat map of the 35 most dominant genera based on 16S rRNA sequences. Phylum abbreviations: bac, Bacteroidetes; pro, Proteobacteria (with a-pro, b-pro, g-pro, and d-pro meaning Alpha-, Beta-, Gamma-, and Deltaproteobacteria, respectively); ver, Verrucomicrobia; fir, Firmicutes; aci, Acidobacteria; eur, Euryarchaeota; nit, Nitrospirae; spi, Spirochaetes. Samples: TH11, water; TH12, 3-mm sediment; TH13, 10-mm sediment.

FIG 2

(A) Clustering and heat map of 18 genera of anoxygenic phototrophs detected in Lake Taihu using the bchY gene. (B) Clustering and heat map of 32 genera of anoxygenic phototrophs detected in Lake Taihu using the pufM gene. TH11, water; TH12, 3-mm sediment; TH13, 10-mm sediment. High and low abundances are indicated by range of dark red and dark blue intensities, respectively. The units on the heat map scales were generated by the software HEML.

FIG 3

Polyacrylamide gel of acsF gene PCR products from Taihu samples. Lanes: M, DNA ladder; 1, water sample; 2, 3-mm sediment sample; 3, 10-mm sediment sample; 4, G. phototrophica AP64 (positive control).

FIG 4

Clustering and heat map of 31 genera of anoxygenic phototrophs detected in Lake Taihu by the newly designed acsF primers. TH11, water; TH12, 3-mm sediment; TH13, 10-mm sediment.

FIG 5

Abundance of Gemmatimonadetes in three samples detected by acsF and 16S rRNA gene, respectively. TH11, water; TH12, 3-mm sediment; TH13, 10-mm sediment.

FIG 6

Phylogenetic tree of 30 OTUs belonging to the phylum Gemmatimonadetes and the reference acsF genes from G. phototrophica AP64 and others. The phylogenetic tree was constructed using the ML algorithm with MEGA 6.1. Bootstrap values are based on 1,000 replicates, and only values of >50% are shown. The numbers in parentheses after the OTUs are the OTU reads in water column, upper- and deep-sediment samples, separated by slashes (“/”). OTUs without a number in parentheses means the overall reads for this OTU were <10.