Chi-Guang Wu and Suh-Jen Lin

Soil Microbiology Lab, Agricultural Chemistry Department, Taiwan Agricultural Research Institute, Taiwan, R.O.C.

Key words: monograph, Entrophospora, systematics

In this study, five species of Entrophospora were illustrated from Taiwan, i.e., E. infrequens, E. columbiana, E. kentinensis, E. microspora, and E. macrospora. Among them, E. microspora and E. macrospora are newly described species and characterized by their unique spore sizes. Most of the species of Entrophospora were recovered from acidic soil, except E. kentinensis which was found in soil with wide pH range. Wall structures of E. kentinensis was revised in this study. In the description of E. macrospora, a new term, “arenaceous ?was coined to describe a sandy and fragile wall structure.

An extensive investigation of Glomales in Taiwan has been conducted to collect various species from natural reservation areas with a wet-sieving and decanting method (Gerdemann and Nicolson, 1963). The sievings were further purified by sucrose centrifugation (Jenkins, 1964). Spores were propagated in the green house with bahia grass. For taxonomic study, spores were mounted in PVLG medium and made into semipermanent slides. Some collections were also preserved in 0.05% Sodium Azide as voucher specimen.

In this study, we illustrated a monograph of Entrophospora from Taiwan, including two new species. Type specimens are deposited in the Soil Microbiology Lab, Agricultural Chemistry Department, Taiwan Agricultural Research Institute, Wu-feng, Taichung, Taiwan, R.O.C. and Mycological Herbarium, Department of Plant Pathology, University of Florida, Gainesville, Florida, U. S. A.

Entrophospora columbiana spain et Schenck

Azygospores produced singly in soil; spores formed within a slightly tapering hypha, terminating in a globose, subglobose swllen tip, 125-175 mm diam; hyphal terminus white to subhyaline, becoming transparent as contents empty to form a spore. Azygospores yellow brown to brown at maturity, globose, subglobose, ellipsoidal, 100-145 x 100-190 mm.

Spore wall: Two layers (L1 and L2). The outer layer (L1), hyaline, confluent with the wall of saccule, ephemeral, 0.5 mm thick. L2, yellow-brown, laminated, 1-3 mm thick.

Flexible inner walls: Three walls are distinguished in all spores. Inner Wall 1: a single hyaline, membranous wall closely adherent to spore wall, <0.5 mm. Inner Wall 2: Usually appears as a single layer, but it consists of two thin-layers, each 0.3-0.5 mm thick. These two layers are closely connected. Inner Wall 3: Obviously, two flexible layers are tightly adherent. L1, membranous, hyaline, and in PVLG looks ¡§beaded¡¨. L2, amorphous, staining red-purple in Melzer¡¦s reagent.

Site of the connecting hyphal attachment from the hyphal terminus frequently visible on the mature azygospore, 20-35 mm diam; the site of connecting hyphal attachement from the hyphal stalk, visible at all stages, 10-15 mm diam. Azygospores often attached with empty hyphal teminus in soil, having a dumbbell shaped configuration. Forming typical vesicular-arbuscular mycorrhizae.

notes: Entrophospora columbiana is similar to Acaulospora mellea by its spore color, wall structure development and spore size. However, E. columbiana could be distinguished from A. mellea by the numbers of cicatrices. This species has been found associated with A. laevis, Glomus claroideum, Scutellospora pellucida, G. leptotichum, G. clarum, and Entrophospora infrequens..

Entrophospora infrequens (Hall) Ames et Schneider

Azygospores produced singly in soil; spores formed within a slightly tapering hypha, terminating in a globose, subglobose swllen tip, 150-175 mm diam; hyphal terminus white to subhyaline, becoming transparent as contents empty to form a spore. Azygospores reddish brown to brown at maturity, globose, subglobose, ovoid, 120-135 x 130-150 mm.

Spore wall: Two layers (L1 and L2) are distinguised. The outer layer (L1), hyaline, confluent with the wall of saccule, ephemeral, up to 11 mm thick. L1 usually covered by a thin membranous sublayer, 1 ƒÝm thick. Layer 2 (L2), yellow-brown or orange brown, laminated, 0.5-1 mm thick, covered by polygonal spins 3-5 mm high and 1.5-2.5 mm wide at base. Spins separated from each other by less than 1 mm.

Flexible inner walls: One inner wall with two layers is distinguished in all spores. Inner wall , hyaline, flexible, 3-4 mm thick. L1: membranous, hyaline, looks minutely beaded in PVLG. L2: hyaline, 3-3.5 mm thick. These two layers are closely adherent. Sometimes, a membranous inner most layer was visible in some crushed spores. Inner wall staining inert or pale pink in the Melzer¡¦s reagent.

Site of the connecting hyphal attachment from the hyphal terminus frequently visible on the mature azygospore, 17.5-20 mm diam; the site of connecting hyphal attachement from the hyphal stalk, visible at all stages, 15 mm diam. Azygospores often attached with empty hyphal teminus in soil, having a dumbbell shaped configuration.

notes: Entrophospora infrequens is similar to E. macrospora by its spore color, and wall structure. However, E. infrequens is very distinguished from other species by its spiny ornamentation. Spin structures could also be seen in the neck between spore and saccule. This species has been found associated with A. tuberculata, A. rhemii, Glomus claroideum, G. constrictum, and Entrophospora columbiana..

ENTROPHOSPORA KENTINENSIS Wu et Liu

Azygospores produced singly in soil; spores formed within a slightly tapering hypha, terminating in a globose, subglobose swllen tip, 100-150 mm diam; hyphal terminus white to subhyaline, becoming transparent as contents empty to form a spore. Azygospores at first white to subhyaline becoming pale yellow to yellow brown at maturity, globose, subglobose, ellipsoidal, irregular, 85-140 x 95-210 mm.

Spore wall: Two layers are visible. Layer 1 (L1), hyaline, confluent with the wall of saccule, ephemeral, 0.5 mm thick; Layer 2 (L2), yellow-brown, laminated, ornamented with pits, 1.5-3 ƒÝm thick; pits on L2, 1-3 mm diam, separated by ridges (2-)3-4(-6) mm.

Flexible inner wall: Three flexible inner walls are visible. Inner wall 1: A single hyaline, membranous layer, <0.3 mm thick, closely adherent to spore wall. Inner wall 2: A rigid wall with two tightly adherent layers. L1, hyaline, membranous, <0.3 mm thick. L2, hyaline, ca. 1 mm thick. Inner wall 3: A flexible wall consisting of two closely attached layers. L1, hyaline, membranous, beaded in PVLG. L2, hyaline, amorphous, (1.5-)2-7.5(-12.5) mm thick, staining red-purple in Merlzer's reagent.

Site of the connecting hyphal attachment from the hyphal terminus frequently visible on the mature azygospore, 20-32.5 mm diam; the site of connecting hyphal attachement from the hyphal stalk, visible only on the newly-formed azygospore, 5-6 mm diam. Azygospores often attached with empty hyphal teminus in soil, having a dumbbell shaped configuration. Forming typical vesicular-arbuscular mycorrhizae.

notes: Entrophospora kentinensis is similar to Acaulospora scrobiculata by its spore color, pitted outer wall, and spore size. However, E. kentinensis differs from A. scrobiculata by (i) spore wall structures, (ii) spore ontogeny types, and (iii) sizes and numbers of cicatrices. Murograph of E. kentinensis is the same as A. mellea, but they mainly differ in spore ontogeny and wall ornamentation. Most Entrophospora species are extracted from acid soil, nevertheless E. kentinensis could survive in wide pH range. This species in this survery was associated with Scutellospora fulgida, A. morrowiae, A. delicata, G. leptotichum, G. derserticola, G. tortuosum.

Entrophospora macrospora Wu et Lin

Azygospores produced singly in soil; spores formed within a slightly tapering hypha, terminating in a globose, subglobose swllen tip, up to 500 mm diam; hyphal terminus yellow brown to pale olive brown. Azygospores globose, subglobose, ellipsoidal, 300-350 x 300-400 mm.

Spore wall: Three layers (L1, L2, L3) are visible in all spores. L1, pale yellow to yellow brown, confluent with the wall of saccule, 2.5-5 mm thick and sometimes covered by a hyaline sublayer, <1 mm thick. L2, yellow-brown, arenaceus, fragile, 4-5 mm thick; L3, yellow-brown to brwon, 1-1.5 mm thick, closely adherent to L2.

Flexible inner wall: A flexible inner wall with three layers is visible in all spores. L1, a single hyaline layer, 1-3 mm thick, closely adherent to L 3 of spore wall. L2, a hyaline, rigid, laminated layer, 5-6 mm thick. L3, a flexible layer, hyaline, membranous, <0.3 mm thick. Spore wall staining inert or yellow in Merlzer's reagent

Site of the connecting hyphal attachment from the hyphal terminus frequently visible on the mature azygospore, 40-50 mm diam; the site of connecting hyphal attachement from the hyphal stalk, 30-35 mm diam. Attached hypha usually constricted at the attachment and a septum was seen in the lower position of attached hypha. Azygospores often attached with empty hyphal teminus in soil, having a dumbbell shaped configuration. Forming typical vesicular-arbuscular mycorrhizae.

notes: Wall structure of E. macrospora is quite similar to E. infrequens. However, the spore size and the arenaceus layer of spore wall make it distinguished from other species in Entrophospora. Arenaceus layer is coined for a unique wall structure which has sandy and fragile features. Under microscope, it shows very minutely granular appearance and could be crushed into powder or fragments easily when making a slide. Spore ontogeny of this species follows the same pattern as E. kentinensis. Septal structures are found both in the neck between spore and sporiferous saccule and in the hyphal stalk connecting to the spore. In the spores of E. infrequens, the L1 ~ L3 of spore wall are continuous with the saccule wall and the connecting channel was occluded by wall thickening or plug. Entrophospora macrospora was associated with G. leptotichum and G. derserticola in the rhizosphere of P. communis.

Entrophospora microspora wu et Lin

Azygospores produced singly in soil; spores formed within a slightly tapering hypha, terminating in a globose, subglobose swllen tip, 45-62.5 mm diam; hyphal terminus white to subhyaline, becoming transparent as contents empty to form a spore. Azygospores at first white to subhyaline becoming pale yellow to yellow brown at maturity, pyriform or ellipsoidal, not often globose or subglobose, 50-77.5 x 62.5-100 mm.

Spore wall: Two layers are visible in all spores. Layer 1 (L1), hyaline, confluent with the wall of saccule, ephemeral, <0.5 mm thick; Layer 2 (L2), yellow-brown, laminated, smooth, 1.5-2 mm thick.

Flexible inner wall: Three flexible inner walls are visible. Inner wall 1: A single hyaline layer, frequently adherent to spore wall, <0.3 mm thick. Inner wall 2: A single hyaline layer, 0.3-0.5 mm thick. Inner wall 3: A flexible wall consisting of two closely attached layers. L1, hyaline, membranous, beaded in PVLG. L2 hyaline, amorphous, 1.5-2 mm thick, staining pink or pinkish-purple in Merlzer's reagent

Site of the connecting hyphal attachment from the hyphal terminus frequently visible on the mature azygospore, 15-22.5 mm diam; the site of connecting hyphal attachement from the hyphal stalk, visible only on the newly-formed azygospore, 7.5-10 mm diam. Azygospores often attached with empty hyphal teminus in soil, having a dumbbell shaped configuration. Forming typical vesicular-arbuscular mycorrhizae.

notes: Entrophospora microspora is similar to Acaulospora morrowiae by its spore size and wall structure. However, E. microspora differs from A. morrowiae by its type of spore ontogey and numbers of cicatrices. The former has two cicatrices and inner most layer staining pinkish purple in Melzer’s reagent; however, the latter has only one cicatrix and inner most layer staining red-purple in Melzer’s reagent. Entrophospora microspora was found in soil with pH 4.3-4.9 and associated with A.mellea,G. clarum and G.gerdemannii.

Ames R N Schneider R W 1979 Entrophospora, a new genus in the Endogonaceae. Mycotaxon 8:347-352.

Gerdemann J W Nicolson T H 1963 Spores of mycorrhizal Endogone species extracted from soil by wet-sieving and decanting. Trans. Br. Mycol. Soc. 46:235-244.

Jenkins W R 1964 A rapid centrifugal-flotation technique for separating nematodes from soil. Plant Dis. Rep. 48:692.

Schenck N C Spain J L Sieverding E Howeler R H 1984 Several new and unreported vesicular-arbuscular mycorrhizal fungi (Endogonaceae) from Columbia. Mycologia 76:685-699.

Wu C G Liu Y S Hwuang Y L Wang Y P Chao C C 1995 Glomales of Taiwan: V. Glomus chimonobambusae and Entrophospora kentinensis, spp. nov. Mycotaxon 53:283-294.

¡@

Chi-Guang Wu and Suh-Jen Lin

Soil Microbiology Lab, Agricultural Chemistry Department, Taiwan Agricultural Research Institute, Taiwan, R.O.C.

Key words: monograph, Entrophospora, systematics

In this study, five species of Entrophospora were illustrated from Taiwan, i.e., E. infrequens, E. columbiana, E. kentinensis, E. microspora, and E. macrospora. Among them, E. microspora and E. macrospora are newly described species and characterized by their unique spore sizes. Most of the species of Entrophospora were recovered from acidic soil, except E. kentinensis which was found in soil with wide pH range. Wall structures of E. kentinensis was revised in this study. In the description of E. macrospora, a new term, “arenaceous ?was coined to describe a sandy and fragile wall structure.

»OÆW¬Ù¹A·~¸ÕÅç©Ò¹A¤Æ¨t¤gÄ[·L¥Íª«¬ã¨s«Ç

¥»¬ã¨s¤¤´y­z¨Ó¦Û»OÆWªº¤º¬`¥ÍÌU¤lÄݪºÅnÂOªK¤º¥Íµß®Úµß, ¥]¬A¤­ºØEntrophospora columbiana, E. infrequens, E. kentinensis, E. microspora, E. macrosporaµ¥¡C¨ä¤¤E. microspora, E. macrospora¨âºØ¬O¤µ¦~µo²{ªº¥@¬É·sºØ¡C¸Ó·sºØªº¥D­n¯S¼x¦b©ó¨ä¿W¯SªºÌU¤l¤j¤p¡C°£¤FE. kentinensis¥i¥Í¦s©ó¸û¼e¼s»ÄÆP­Èªº¤gÄ[¥~, ¤j¦h¼Æªº¤º¬`¥ÍÌU¤lÄݪºÌU¤l³£µo²{©ó»Ä©Ê¤gÄ[¤¤¡C¥»¤å¨Ã°w¹ïE. kentinensisªºÌU¾Àºc³y´£¥X­×¥¿¡C¦b´y­zE. macrospora®É, §Ú­Ì´£¥X¤@·sªº¦Wµü, ¡§¬â½èÌU¾À¡¨¥Î¥H§Î®e¨ã¦³¬â½è¤Î¯Ü®zªºÌU¾À¯S©Ê¡C