Isolation and characterization of anaerobic cellulose-degrading bacteria from East African porcupine (Hysterix cristata)

Two strains of obligately anaerobic cellulolytic bacteria designated as PS7 and PS8 (PS for porcupine species) were isolated from hindgut fluid of a crested porcupine (Hystrix cristata). The rates of cellulose degradation, total volatile fatty acids, and gas production from cellulose by the isolates were determined in comparison with Ruminococcus flavefaciens FD-1. Ruminococcus flavefaciens FD-1 degraded acid swollen cellulose and produced total volatile fatty acid at a faster rate (0.03145 mg/d;3.8350 mumol/mL) than PS7 (0.03113 mg/d' 2.5278 mumol/mL) and PS (0.0125 mg/d; 2.1080 mum/mL). However, PS7 degraded cellulose strips (untreated) faster (1.5 weeks) than R. flavefaciens FD-1 (2 weeks). Furthermore, PS7 produced gas at a higher rate (0.1055 ml/d) than R. flavefaciens FD-1 (0.03145 ml/d) more produced butyric, isovaleric acids and almost twice the amounts of total volatile fatty acids from acid swollen cellulose. Both PS7 and PS8 were Gram variable, rod shaped and motile. On cellobiose medium, PS7 grew at temperature ranges from 25 to 45 degree C while PS8 did not grow at 25 and 45 degree C. Both isolates grew at pH levels between 6.2 and 11. Characteristation based on carbohydrate fermentation and morphology indicated that these two isolates were similar. Characterisation by RAPD-PCR suggested that PS7 and PS8 were genotypically similar but distinct. Phylogenetic analysis using the nucleotide sequence (1450 bp) of the 16S rRNA gene suggested that PS7 clustered with Clostridium sub-phylum and exhibited the highest similarity (95 percent) with Clostridium lentocellum. The phylogenetic results suggest that PS7 might represent a new taxon

Characterization of tannin-tolerant bacterial isolates from East African ruminants

Several tannin-tolerant bacteria were isolated from enrichment cultures of rumen microflora of bush duiker, giraffe, Grant's gazelle, sheep, and goat, and established in medium containing crude tannin extracts or tannic acid. The isolates were characterized by classical and molecular methods. The isolates were also tested for the presence of tannin acylhydrolase. Characterization by restriction fragment length polymorphism of the 16S rRNA-PCR product was performed with Alu1, Dde1, Msp1, and Taq1. Amplified fragment length polymorphism analysis was performed only on the isolates that were curved rods. The nucleotide sequence of PCR products derived from the 16S rRNA genes of isolates was determined. The classical characterization suggested that, with one exception all the curved rods isolates were Selenomonas and the coccus was a Streptococcus. Only Selenomonas-like isolates had tannin acylhydrolase activities. One isolate lost the ability to completely hydrolyze tannins after prolonged storage at –70 degree C. The restriction fragment length polymorphism profiles suggested that the Selenomonas-like isolates exhibited heterogeneity in the ribosomal RNA locus. The coccus had the same profiles as Streptococcus caprinus, while the straight rods appeared to be similar to each other. Amplified fragment length polymorphism analysis suggested that the Selenomonas-like isolates clustered into two major groups. The 16S rRNA sequences of the coccus clustered with that of Streptococcus species and the Selenomonas-like isolates exhibited a high level of similarity with Selenomonas ruminantium, while the straight rods clustered with Klebsiella species accessions in the databases. A partial 16S sequence strongly indicated that one of the isolates was Butyrivibrio fibrisolvens