High-Throughput Sequencing Reveals a Dynamic Bacterial Linkage between the Captive White Rhinoceros and Its Environment.

Soil is an essential part of the animal habitat and has a large diversity of microbiota, while the animal body was colonized by a complex bacterial community; so far, the relationship between the animal host microbial community and the soil microbial ecosystem remains largely unknown. In this study, 15 white rhinoceros from three different captive grounds were selected and the bacterial community of the gut, skin, and environment of these rhinoceros were analyzed by 16S rRNA sequencing technology. Our results showed that Firmicutes and Bacteroidota were the predominant phyla in the gut microbiome, whereas skin and environment samples share similar microbiome profiles and are dominated by the phyla of Actinobacteriota, Chloroflexi, and Proteobacteria. Although the bacterial composition of the gut differs from that of the skin and environment, the Venn diagrams showed that there were 22 phyla and 186 genera shared by all the gut, skin, and environmental microbes in white rhinoceroses. Further cooccurrence network analysis indicated a bacterial linkage based on a complex interaction was established by the bacterial communities from the three different niches. In addition, beta diversity and bacterial composition analysis showed that both the captive ground and host ages induced shifts in the microbial composition of white rhinoceroses, which suggested that the bacterial linkage between the captive white rhinoceros and its environment is dynamic. Overall, our data contribute to a better understanding of the bacterial community of the captive white rhinoceros, especially for the relationship between the environment and animal bacterial communities. IMPORTANCE The white rhinoceros is one of the world's most endangered mammals. The microbial population plays a key role in animal health and welfare; however, studies regarding the microbial communities of the white rhinoceros are relatively limited. As the white rhinoceros has a common behavior of mud baths and thus is in direct contact with the environment, a relationship between the animal microbial community and the soil microbial ecosystem appears possible, but it remains unclear. Here, we described the characteristics and interaction of bacterial communities of the white rhinoceros in three different niches, including gut, skin, and environment. We also analyzed the effects of captive ground and age on the composition of the bacterial community. Our findings highlighted the relationship among the three niches and may have important implications for the conservation and management of this threatened species.

[siRNA-mediated silencing of ClC-2 gene inhibits proliferation of human U-87 glioma cells].

BACKGROUND & OBJECTIVE: Small interfering RNA (siRNA), which has been used to inhibit mammalian gene expression, is demonstrated to be an effective tool for gene function investigation. The aim of the present study was to observe the effect of siRNA, which was designed to target volume-regulated chloride channel ClC-2 gene, on the proliferation of a human glioma cell line U-87. METHODS: Two recombinant eukaryotic CIC-2 siRNA expression vectors were designed and constructed. Sequences were identified and confirmed by restrictive endonuclease digestion and DNA sequencing. The empty vector, pSUPER. puro, and two recombinant plasmids, pSUPER. puro-shRNA-ClC-21 and pSUPER. puro-shRNA-ClC-22, were transfected into U-87 cells using Lipofectamine2000 (Groups: PP0, PP1 and PP2, respectively). ClC-2 mRNA expression was detected by reverse transcription polymerase chain reaction (RT-PCR); the cellular proliferation rate was determined by MTT assay; the cell cycle was measured by flow cytometry (FCM); and the cell colony formation rate was measured by plate colony formation assay. RESULTS: The DNA fragments encoding siRNA targeting ClC-2-gene were successfully connected onto pSUPER. puro vector. ClC-2 mRNA expression and the cell growth rate in PP1 and PP2 groups were significantly inhibited compared to those in PP0 and control groups. Meanwhile, cell cycle was arrested in G1 phase and the percentage of G1 phase cells were increased by about 30.24% in PP1 and 18.04% in PP2 vs. in control and PP0 groups, P < 0.05. Moreover, the cell colony formation rates were statistically decreased in siRNA treated groups, which were (11.0+/-1.0)% in PP1 and (20+/-3.1)% in PP2 vs. (46.5+/-1.6)% in control and (47.5+/-2.8)% in PP0 groups (P<0.01). CONCLUSION: These results demonstrate that CIC-2 siRNA could inhibit the cell proliferation of a human glioma cell line U-87, thus ClC-2 gene may be used as a novel target for the suppression of the growth of human malignant glioma cells.

Involvement of anion channel(s) in the modulation of the transient outward K(+) channel in rat ventricular myocytes.

The cardiac Ca(2+)-independent transient outward K(+) current (I(to)), a major repolarizing ionic current, is markedly affected by Cl(-) substitution and anion channel blockers. We reexplored the mechanism of the action of anions on I(to) by using whole cell patch-clamp in single isolated rat cardiac ventricular myocytes. The transient outward current was sensitive to blockade by 4-aminopyridine (4-AP) and was abolished by Cs(+) substitution for intracellular K(+). Replacement of most of the extracellular Cl(-) with less permeant anions, aspartate (Asp(-)) and glutamate (Glu(-)), markedly suppressed the current. Removal of external Na(+) or stabilization of F-actin with phalloidin did not significantly affect the inhibitory action of less permeant anions on I(to). In contrast, the permeant Cl(-) substitute Br(-) did not markedly affect the current, whereas F(-) substitution for Cl(-) induced a slight inhibition. The I(to) elicited during Br(-) substitution for Cl(-) was also sensitive to blockade by 4-AP. The ability of Cl(-) substitutes to induce rightward shifts of the steady-state inactivation curve of I(to) was in the following sequence: NO(3)(-) > Cl(-) approximately Br(-) > gluconate(-) > Glu(-) > Asp(-). Depolymerization of actin filaments with cytochalasin D (CytD) induced an effect on the steady-state inactivation of I(to) similar to that of less permeant anions. Fluorescent phalloidin staining experiments revealed that CytD-pretreatment significantly decreased the intensity of FITC-phalloidin staining of F-actin, whereas Asp(-) substitution for Cl(-) was without significant effect on the intensity. These results suggest that the I(to) channel is modulated by anion channel(s), in which the actin cytoskeleton may be implicated.