Formation of Mycorrhiza-like Structures in Cultured Root/Callus of Cathaya argyrophylla Chun et Kuang Infected with the Ectomycorrhizal Fungus Cenococcum geophilum Fr.

An in vitro system was used for ectomycorrhizal synthesis of Cenococcum geophilum Fr. with Cathaya argyrophylla Chun et Kuang, an endangered species. Calli initiated from stem segments and adventitious roots differentiated from young seedlings were removed and cocultured with Cenococcum geophllum on a modified Murashlge-Skoog medium. Fungal hyphae were visible within intercellular spaces of the callus 4 weeks after inoculation, but definite and well-developed Hartig net structures did not form in the calU 8 weeks after inoculation. The typical ectomycorrhizal structures （i.e. hyphal mantle and Intracortical Hartig net） were observed in root segments 8 weeks after inoculation. This is the first report of aseptic ectomycorrhlzal-like formation/infection between root organ/callus of Cathaya argyrophylla and the ectomycorrhizal fungus Cenococcum geophflum. This culture system is useful for further investigation of mycorrhizal synthesis in Cathaya trees.     

In situ and in vitro colonization of Cathaya argyrophylla (Pinaceae) by ectomycorrhizal fungi

Cathaya argyrophylla, a critically endangered conifer, is found to grow at four isolated areas located in subtropical mountains of China. To examine the involvement and usefulness of mycorrhizas for sustaining the population of this tree, we compared the root system, morphology, and structure of mycorrhizal roots of C. argyrophylla, which were collected from a natural stand and an artificial stand, each grown at a different location. More mycorrhizal roots were found for trees from an artificial stand. The presence of extramatrical mycelium, mantle, and Hartig net revealed that C. argyrophylla formed an ectomycorrhizal association in both sampling sites. Starch granules were found in mycorrhizal roots collected only from a natural stand. The aseptic synthesis of C. argyrophylla and Cenococcum geophilum was established for the first time in vitro. Typical ectomycorrhizas formed on seedlings on RM medium containing 0.1 g/l glucose, 5 weeks after inoculation. By light microscopy, the synthesized mycorrhizas showed a thin mantle from which emanated extramatrical hyphae and highly branched Hartig net. A simple, rapid, and convenient mycorrhiza synthesis system was developed, which facilitates further studies on ectomycorrhizal development of C. argyrophylla.     

The Effect of Various Culture Media on the Formation of Embryo-like Structures in Cultures Derived from Explants Taken from Mature Larix decidua

The effect of various collection dates and nine different culture media on the formation of ‘embryo-like structures’ (ELS) in cultures derived from explants taken from a 42-year-old Larix decidua tree was studied. The best medium was AFC, a medium low in NH₄⁻, NO₃ ⁺, Mg²⁺ and SO₄²⁻ but high in PO₄³⁻ compared with the concentration of these elements in the other media. On AFC, ELS production reached a peak with collections made in late summer during the period when needle primordia are being initiated. For the other media, collection date had a lesser effect on ELS initiation.     

Fasciola hepatica: Morphological changes in vitelline cells following treatment in vitro with the deacetylated (amine) metabolite of diamphenethide (DAMD)

Fasciola hepatica: morphological changes in vitelline cells following treatment in vitro with the deacetylated (amine) metabolite of diamphenethide (DAMD). International Journal for Parasitology 18: 1061–1069. The effect of the deacetylated (amine) metabolite of diamphenethide (DAMD, 10 μg ml⁻¹) on the vitelline cells of Fasciola hepatica over an 18 h period in vitro has been determined by transmission electron microscopy. DAMD acts preferentially on the undifferentiated stem cells and the intermediate cells in the early stages of protein synthesis; it appears to prevent their continued development. In the stem cell the nucleolus is absent after 6 h. During the rest of the drug treatment period considerable clumping of heterochromatin takes place, the cells round up and become electron-dense. Signs of autophagy are also evident, and the mitochondria become swollen and disorganized. From 6 h onwards there are progressive changes in the It1 (intermediate type 1) cells, including clumping of the heterochromatin in the nucleus, a decrease in the number of egg-shell globules (and consequently a reduction in the number and size of the shell globule clusters), and a decrease in the number of ribosomes on the GER cisternae, although the GER system remains well-developed. By 18 h the nucleolus is absent, and the cells are very rounded and electron-dense; the mitochondria are swollen and disorganized. Similar changes are evident in the It2 (intermediate type 2) cells, so that by 18 h it is difficult to distinguish between the It1 and It2 cells. In the mature cells there is a progressive decrease in the number and size of the shell globule clusters from 9 h onwards. Glycogen synthesis and ‘yolk’ formation may also be impaired and lipid droplets are present. Spaces begin to appear between the nurse cell cytoplasm and the vitelline cells after 9 h, and disruption of the nurse cell cytoplasm is evident after 12 h, becoming very severe by 18 h. By this time the follicle is very disorganized and empty-looking. In more severely affected follicles the mature cells are seen to be breaking down. Over the 18 h drug treatment period, a change in the cell population of the follicle takes place, with relatively more stem, early It1 and mature cells being present, whilst few if any characteristic It1 and It2 cells remain. The results are interpreted as being due to an inhibition of protein synthesis in the vitelline cells by DAMD.     

Associations of variants in MTHFR and MTRR genes with male infertility in the Jordanian population

Folate pathway is expected to play an important role in spermatogenesis since it is involved in DNA synthesis, repair and methylation. The purpose of this study was to examine the association between male infertility and the MTHFR (C677T and A1298C) and MTRR (A66G) polymorphisms. A group of 300 males was recruited in this study from different Jordanian infertility clinics. Of these, 150 cases of infertile men that included oligozoospermia cases (n = 45), severe oligozoospermia (n = 71) and azoospermia (n = 34) were studied. The other 150 males were age matched fertile controls. Genotyping of MTHFR and MTRR polymorphisms was performed using PCR-RFLP technique. The results showed an association between MTHFR 677TT genotype and male infertility (P < 0.05). However, the distribution of MTHFR A1298C and MTRR A66G genotypes were not different between the fertile and infertile groups (P > 0.05). In addition, none of the examined polymorphisms was related to any of the semen parameters in the infertile group. In conclusion, this study showed that MTHFR C677T polymorphism is associated with male infertility in Jordanians.