Deep sequencing analysis of toad Rhinella schneideri skin glands and partial biochemical characterization of its cutaneous secretion

Animal poisons and venoms are sources of biomolecules naturally selected. Rhinella schneideri toads are widespread in the whole Brazilian territory and they have poison glands and mucous gland. Recently, protein from toads' secretion has gaining attention. Frog skin is widely known to present great number of host defense peptides and we hypothesize toads present them as well. In this study, we used a RNA-seq analysis from R. schneideri skin and biochemical tests with the gland secretion to unravel its protein molecules. Methods: Total RNA from the toad skin was extracted using TRizol reagent, sequenced in duplicate using Illumina Hiseq2500 in paired end analysis. The raw reads were trimmed and de novo assembled using Trinity. The resulting sequences were submitted to functional annotation against non-redundant NCBI database and Database of Anuran Defense Peptide. Furthermore, we performed caseinolytic activity test to assess the presence of serine and metalloproteases in skin secretion and it was fractionated by fast liquid protein chromatography using a reverse-phase column. The fractions were partially sequenced by Edman's degradation. Results: We were able to identify several classes of antimicrobial peptides, such as buforins, peroniins and brevinins, as well as PLA2, lectins and galectins, combining protein sequencing and RNA-seq analysis for the first time. In addition, we could isolate a PLA2 from the skin secretion and infer the presence of serine proteases in cutaneous secretion. Conclusions: We identified novel toxins and proteins from R. schneideri mucous glands. Besides, this is a pioneer study that presented the in depth characterization of protein molecules richness from this toad secretion. The results obtained herein showed evidence of novel AMP and enzymes that need to be further explored.(AU)

Results of Pulmonary Function Tests as Part of a Health Care Program for a Regional Community at a Kanagawa Hospital: A Review / 日本農村医学会雑誌

The results of pulmonary function tests (PFTs) given to those inhabitants in the western part of Kanagawa Prefecture who visited our hospital for medical examinations between April 1991 and March 1992 were studied comparatively, with their occupation, age, sex, environmental factors and smoking habits taken into account. For this purpose, the subjects totaling 1, 322 were classified into three groups-those who live in the hilly area, those who live in the suburban area and those who live in the coastal area. In the present study, the results of the four PFT items-FVC, FEV 1.0%, FVC and FEV 1.0%-were checked. Careful examination brought into relief the startling fact that many aged people, non-farmers and nonsmokers in the suburban subject group have impaired pulmonary function. It is said that because of the Tokyo-Nagoya expressway and many other motorways, the air in the suburban area is fouled up with exhaust gas to a greater extent than in the other two areas. Although our finding alone could not identify the cause of impaired pulmonary function definitely, air pollution was thought to be a culprit. Further investigation should be made into the living conditions of the examinees. Moreover, environmental monitoring and data analysis have to be carried out in the future.