RESUMEN
Rhodiola sachalinensis (family Crassulaceae), a perennial herbaceous plant with adaptogenic properties, cardiopulmonary protective effects, and central nervous system activities, is widely used as a traditional Chinese medicine (3). R. sachalinensis naturally exists only above 1,500 m elevation in the mountain area of Changbai Mountain (average July temperature ≤10°C), China. Introduction and cultivation of R. sachalinensis has been carried out in several low-altitude districts of northeast China. From 2007 to 2010, severe root rot disease was observed on R. sachalinensis in Siping districts, Jilin, China. Approximately 75 to 95% of the fields were affected with root rot disease incidence ranging from 85 to 100% under conditions of high temperatures (24 to 30°C) and high humidity. As the disease progressed, brown lesions expanded on lateral and main roots, and aboveground tissues shriveled and died. Over the 4- to 5-year period from culture to harvest, root rot became more serious. Symptomatic plants were collected from Siping districts. Samples were rinsed in tap water, necrotic areas were excised and cut into 2-mm pieces, surface sterilized with 5% NaOCl for 30 s, and rinsed in four successive changes of sterile distilled water. A single fungus was isolated on potato dextrose agar (PDA). The fungus was white, then pink and cottony, with nearly round margins after 8 days (27°C). Hyphae were separate and hyaline but macroconidia were sparse and occurred in a false head. Conidiogenous cells were monophialides. Microconidia in chains were abundant and mostly nonseptate, oval to clavate, and measured 8.1 to 14.5 × 2.0 to 3.4 µm. Morphological characteristics suggested the isolate was Fusarium verticillioides (Gibberella fujikuroi), which differed from the reported root rot pathogen of R. sachalinensis, F. oxysporum by forming microconidia in chains (1). The sexual stage of F. verticillioides was not observed. The internal transcribed spacer (ITS) fragments were amplified using ITS1 and ITS4 as primers and the 351-bp sequence was deposited in GenBank (Accession No. HQ025928). The ITS sequence showed 100% nucleotide sequence identity with F. verticillioides (GenBank Accession No. AY188916.1.). For pathogenicity tests, the isolate was cultured on PDA for 8 days. Inoculations were performed on 15 healthy R. sachalinensis plants by spraying a conidial suspension (2.0 × 105 microconidia ml-1) on roots wounded with a metal needle (6 wounds cm-2) (2). Ten plants were mock inoculated with water. Plants were incubated in a growth chamber (25°C, 70 to 80% relative humidity, 300 µmol·m-2·s-1 light intensity, and a 12-h photoperiod). After 15 days, defoliation and root rot symptoms were similar to the original symptoms observed under field conditions. F. verticillioides was reisolated from the roots of infected plants. Control plants remained asymptomatic. To our knowledge, this is the first report of F. verticillioides on R. sachalinensis in China. References: (1) X. Y. Li et al. J. Northeast For. Univ. 34:12, 2003. (2) M. Ma. Syahit et al. Am. J. Appl. Sci. 6:902, 2009. (3) T. F. Yan et al. Conserv. Genet. 4:213, 2003.
RESUMEN
The filamentous fungus Cochliobolus carbonum produces endo-alpha 1,4-polygalacturonase (endoPG), exo-alpha 1,4-polygalacturonase (exoPG), and pectin methylesterase when grown in culture on pectin. Residual activity in a pgn1 mutant (lacking endoPG) was due to exoPG activity, and the responsible protein has now been purified. After chemical deglycosylation, the molecular mass of the purified protein decreased from greater than 60 to 45 kDa. The gene that encodes exoPG, PGX1, was isolated with PCR primers based on peptide sequences from the protein. The product of PGX1, Pgx1p, has a predicted molecular mass of 48 kDa, 12 potential N-glycosylation sites, and 61% amino acid identity to an exoPG from the saprophytic fungus Aspergillus tubingensis. Strains of C. carbonum mutated in PGX1 were constructed by targeted gene disruption and by gene replacement. Growth of pgx1 mutant strains on pectin was reduced by ca. 20%, and they were still pathogenic on maize. A double pgn1/pgx1 mutant strain was constructed by crossing. The double mutant grew as well as the pgx1 single mutant on pectin and was still pathogenic despite having less than 1% of total wild-type PG activity. Double mutants retained a small amount of PG activity with the same cation-exchange retention time as Pgn1p and also pectin methylesterase and a PG activity associated with the mycelium. Continued growth of the pgn1/pgx1 mutant on pectin could be due to one or more of these residual activities.