Assessment of seawater bacterial infection in rabbit tibia by Illumina MiSeq sequencing and bacterial culture

Abstract

Abstract Objectives We aimed to explore the bacterial community composition following ocean bacterial infection using an animal model. Methods This animal-based experiment was conducted from September 2019 to November 2019. Eighteen seawater filter membranes were collected from Changle City, Fujiian Province, China, on September 8, 2019. Ten filter membranes were used for implantation. Eight filter membranes that were used in the bacterial culture for the exploration of seawater bacteria were assigned to the seawater group (SG). Fourteen healthy adult New Zealand rabbits were randomly divided into the experimental group (EG) and control group (CG). Seawater filter membranes and asepsis membranes were implanted into the tibia in the EG and CG, respectively. One week after surgery, tibial bone pathology tissues were collected and assessed using light microscopy and scanning electron microscopy (SEM). Medullary cavity tissues were collected for the performance of Illumina MiSeq sequencing and bacterial culture. The differences between EG and CG were assessed by pathological observation under light microscopy and SEM, high-throughput bacterial sequencing, and bacterial culture. Results Compared with the CG, the infection rate was 100%, and the mortality value was 20% after the implantation of the filter membranes in the EG. Both light microscopy and SEM showed that a large number of bacteria were distributed in the bone marrow cavity after ocean bacterial infection. No bacterial growth was found in the CG. Illumina MiSeq sequencing found that Firmicutes, Proteobacteria, Thermotogae, Fusobacteria, Bacteroidetes, and Actinobacteria were the dominant bacteria at the phylum level and Clostridium_sensu_stricto_7, Haloimpatiens, Clostridium_sensu_stricto_15, Clostridiaceae_1, Clostridium_sensu_stricto_18, and Oceanotoga were the dominant bacteria in genus level among the EG. In the bacterial culture of the medullary cavity tissues, Klebsiella pneumoniae, Shewanella algae, Staphylococcus aureus, Escherichia coli, Enterobacter cloacae, and Vibrio vulnificus were the predominant infective species. Moreover, compared with the SG, the EG showed a higher detection rate of E. coli and S. aureus (P = 0.008 and P = 0.001, respectively). The detection rates of V. alginolyticus, V. parahaemolyticus, and V. fluvialis were higher in the SG than the EG (P = 0.007, P = 0.03, and P = 0.03, respectively). Conclusions Our model, which was comprehensively evaluated using four techniques: histopathology and SEM observation, gene detection, and bacteria culture, provides a scientific basis for the clinical diagnosis and treatment of patients in such settings.

Keywords

• Scanning electron microscopy
• Sequencing
• Marine
• New Zealand rabbit
• Bacterial infection