Isolation of Main Pathogens Causing Postharvest Disease in Fresh Codonopsis pilosula during Different Storage Stages and Ozone Control against Disease and Mycotoxin Accumulation

Codonopsis pilosula is an important Chinese herbal medicine. However, fresh C. pilosula is prone to decay during storage due to microorganism infections, seriously affecting the medicinal value and even causing mycotoxin accumulation. Therefore, it is necessary to study the pathogens present and develop efficient control strategies to mitigate their detrimental effects on the herbs during storage. In this study, fresh C. pilosula was collected from Min County in Gansu Province, China. The natural disease symptoms were observed during different storage stages, and the pathogens causing C. pilosula postharvest decay were isolated from the infected fresh C. pilosula. Morphological and molecular identification were performed, and pathogenicity was tested using Koch’s postulates. In addition, the control of ozone was examined against the isolates and mycotoxin accumulation. The results indicated that the naturally occurring symptom increased progressively with the extension of storage time. The mucor rot caused by Mucor was first observed on day 7, followed by root rot caused by Fusarium on day 14. Blue mold disease caused by Penicillum expansum was detected as the most serious postharvest disease on day 28. Pink rot disease caused by Trichothecium roseum was observed on day 56. Moreover, ozone treatment significantly decreased the development of postharvest disease and inhibited the accumulations of patulin, deoxynivalenol, 15-Acetyl-deoxynivalenol, and HT-2 toxin

Mutational spectrum of Chinese LGMD patients by targeted next-generation sequencing

<div><p>This study aimed to study the diagnostic value of targeted next-generation sequencing (NGS) in limb-girdle muscular dystrophies (LGMDs), and investigate the mutational spectrum of Chinese LGMD patients. We performed targeted NGS covering 420 genes in 180 patients who were consecutively suspected of LGMDs and underwent muscle biopsies from January 2013 to May 2015. The association between genotype and myopathological profiles was analyzed in the genetically confirmed LGMD patients. With targeted NGS, one or more rare variants were detected in 138 patients, of whom 113 had causative mutations, 10 sporadic patients had one pathogenic heterozygous mutation related to a recessive pattern of LGMDs, and 15 had variants of uncertain significance. No disease-causing mutation was found in the remaining 42 patients. Combined with the myopathological findings, we achieved a positive genetic diagnostic rate as 68.3% (123/180). Totally 105 patients were diagnosed as LGMDs with genetic basis. Among these 105 patients, the most common subtypes were LGMD2B in 52 (49.5%), LGMD2A in 26 (24.8%) and LGMD 2D in eight (7.6%), followed by LGMD1B in seven (6.7%), LGMD1E in four (3.8%), LGMD2I in three (2.9%), and LGMD2E, 2F, 2H, 2K, 2L in one patient (1.0%), respectively. Although some characteristic pathological changes may suggest certain LGMD subtypes, both heterogeneous findings in a certain subtype and overlapping presentations among different subtypes were not uncommon. The application of NGS, together with thorough clinical and myopathological evaluation, can substantially improve the molecular diagnostic rate in LGMDs. Confirming the genetic diagnosis in LGMD patients can help improve our understanding of their myopathological changes.</p></div

Immunohistochemical findings in different LGMD subtypes^a.

<p>Immunohistochemical findings in different LGMD subtypes<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175343#t002fn001" target="_blank">^a</a>.</p

Histopathological abnormalities in different LGMD subtypes^a.

<p>Histopathological abnormalities in different LGMD subtypes<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175343#t001fn001" target="_blank">^a</a>.</p

Flowchart of patients’ enrollment.

<p>Flowchart of patients’ enrollment.</p

Myopathological changes of LGMD patients.

<p>A. NADH-TR staining shows lobulated fibers (arrow) in a LGMD2B patient. B. MGT staining shows red-ragged fibers (arrow) in a LGMD2B patient. C. MGT staining shows nemaline bodies (arrow) in a LGMD2A patient. D. NADH-TR staining shows multi-minicores (arrow) in a LGMD2A patient. E. Immunohistochemical labelling of N-terminal dystrophin shows decreased or absent expression (asterisk) in a LGMD2A patient. F. Immunohistochemical labelling of dysferlin shows decreased expression (asterisk) in a LGMD2A patient. Abbreviations: LGMD = limb-girdle muscular dystrophy; NADH-TR = nicotinamide adenine dinucleotide-tetrazolium reductase; MGT = modified Gomori trichrome.</p