Fergusobia tumifaciens ( Currie 1937 ) Wachek 1955

Davies, Kerrie A., Ye, Weimin, Giblin-Davis, Robin M., Taylor, Gary S., Hodda, Mike & Thomas, W. Kelley, 2014, Nematodes from galls on Myrtaceae. VII. Fergusobia from ‘ leafy’ leaf bud galls in Australia, with re-description of Fergusobia tumifaciens (Currie 1937) Wachek 1955 and descriptions of Fergusobia planchonianae n. sp. and Fergusobia viminalisae n. sp., Zootaxa 3856 (4), pp. 529-554 : 533-539

Re-description of Fergusobia tumifaciens ( Currie 1937) Wachek 1955

( Figs 2 , 3 , 7 A, 8A)

Measurements. Table 2 .

Material examined. The description presented here is based on measurements of 19 parthenogenetic ♀s, 19 infective ♀s and 21 ♂ s; from roadside vegetation on Jindera Street, Jindera, near Albury in NSW, Australia (36°04.32´S 146°92.90´E). Taken from ‘leafy’ leaf bud galls on E. bridgesiana R. Baker 1898 . Coll. K. A. Davies, 18.iii.2011.

Neotype. One parthenogenetic female, on a slide deposited at the Australian National Insect Collection ( ANIC), Canberra, ACT, Australia, collection data as above; with a paraneotype infective ♀ and a ♂.

Paraneotypes. Vouchers (collection data as above) deposited at the Waite Insect and Nematode Collection ( WINC), The University of Adelaide, SA, Australia, 4 parthenogenetic females, 4 infective females and 5 males ( WNC 2546); the National Museum of Australia, Sydney, NSW, Australia, 6 parthenogenetic females, 6 infective females and 11 males on slides; and at the USDA Nematode Collection, Beltsville, MD, USA 2 parthenogenetic females, 2 infective females and 3 males on slides.

Description. Parthenogenetic female. Body open C-shape, rarely arcuate, dorsally curved with ventral side convex; relatively small and broad; smaller than amphimictic pre-parasitic female and male; body narrows behind vulva to form short, arcuate, conoid tail with bluntly rounded tip. Cuticle appears smooth under light microscope, sub-cuticle with strong longitudinal striae. Lateral fields not seen.

Cephalic region about 50% diameter of body at anterior end, off-set, 1–2 µm long, unstriated; rounded outline in lateral view, circum-oral area raised <1 µm. Amphids not seen. Stylet small, with conus about 40% of length, rounded basal knobs just higher than wide, ca 2 µm diameter at base.

Orifice of dorsal pharyngeal gland about 1 µm posterior to stylet knobs. Anterior fusiform part of digestive tract occupying 54–65% (mean 60%, n = 20) of body diameter, length 0.7–1.9 times diameter; usually with two large nuclei, lumen of tract broadening at distal end of dorsal pharyngeal gland. Pharyngeal glands extending over intestine, large, diameter about 60–85% of body diameter, distance from head to end of glands being 48 (42–55)% of total body length. Gland nucleus large, with prominent nucleolus.

Neotype Partheno-genetic Parthenogenetic females Males Infective

female females Secretory/excretory pore opens at level in front of gland nucleus; duct obscure; secretory/excretory cell ovoid, about 7–11 µm long. Hemizonid extending over two or three annules, one or two annules in front of secretory/ excretory pore.

Reproductive tract variable in length, extending part-way along pharyngeal gland or to nerve ring; with 0–5 flexures; oviduct usually with two oocytes in a row; quadricolumella smooth, uterus relatively short and nonextensile, usually without egg; vulva a simple transverse slit with protruding lips in some specimens, no vulval plate. Anus pore-like. Tail conoid, relatively short (c = 15–22), convex on ventral side; length 1–2 times anal body diameter; tip bluntly rounded.

Infective pre-parasitic female. Infecting mature larval stage of Fn. carteri Tonnoir 1937 or pupa. J-shape when heat-relaxed; maximum body diameter at mid-body length; body tapers gradually posterior to vulva. Cuticle obscurely annulated, <1 µm long; longitudinal striae apparent with light microscope; lateral fields not seen.

Cephalic region slightly offset; circum-oral area rounded and barely raised; stylet slender, short, weakly sclerotised with basal knobs higher than wide, rounded, ca 2µm wide; conus about 50% of length.

Orifice of dorsal pharyngeal gland often obscure, about 1µm posterior to stylet knobs. Anterior fusiform part of digestive tract diameter 42–60% of body diameter, length 1.2–1.6 times diameter. Pharyngeal glands diameter about 25–70% of body diameter, distance from head to end of glands 24 (19–27)% of body length.

Secretory/excretory pore opens at level of nucleus of pharyngeal glands; duct obscure; ellipsoid secretory/ excretory cell about 3 µm long. Hemizonid not seen.

Uterus 70–80% of total gonad length in uninseminated females, packed with sperm in inseminated females; vagina at 90 to body axis; reproductive tract extending to nerve ring; usually with at least one flexure. Vulva a transverse slit, vulval lips raised about 1 µm, no vulval plate present. Anus an obscure pore. Tail relatively short (c = 14.5–29.5), sub-cylindroid; length about diameter at anus, tip almost hemispherical.

Male. Body almost straight or barely J-shaped when heat-relaxed, tail region more or less curved ventrally. Cuticle weakly annulated, annules ca 1 µm long; strong longitudinal striae apparent with light microscope; lateral fields not seen.

Cephalic region occupying about 80% anterior body diameter, offset, ca 2 µm long; circum-oral area flat; stylet short, with conus about 50% of length, stylet knobs rounded, slightly higher than wide, 2–3 µm wide at base.

Anterior fusiform part of digestive tract diameter about 65–75% of body diameter, length 1.1–1.9 times diameter. Orifice of dorsal pharyngeal gland 1–2 µm behind knobs. Pharyngeal glands diameter 60–90% of body diameter, distance from head to end of glands 20–34% of total body length.

Secretory/excretory pore opens opposite nucleus of pharyngeal gland; duct obscure; secretory/excretory cell not seen. Hemizonid extending over two annules, one or two annules in front of secretory/excretory pore.

Testis single, extending to nerve ring; may be reflexed; testis, seminal vesicle and vas deferens not clearly differentiated in specimens that have more sperm, i.e., appear to be more mature. Bursa almost peloderan but ends just in front of tail tip, smooth; obscure; arises 9–16% of length of body anterior to tail tip. Spicules paired, angular at about 45% of length, with manubrium and shaft slightly longer than blade; moderately sclerotised; manubrium wider than shaft, not offset; blade narrows gradually to bluntly rounded tip; opening terminal. Inconspicuous muscles associated with cloaca. Tail arcuate, ventrally concave; length 1–1.7 times diameter at cloaca; bluntly rounded tip.

Diagnosis (Emended) and Relationships. Fergusobia tumifaciens is morphologically characterized by the combination of a parthenogenetic female with a relatively small C-shaped body, a small head capsule, vulva at ca 85% of body length, and an arcuate, short, broadly conoid tail with a bluntly rounded tip; a J-shaped infective female with a relatively large body and a sub-cylindroid tail with an almost hemispherical tail tip and a relatively posterior vulva; and an arcuate to barely J-shaped male with a relatively large body, angular (not heavily sclerotised) spicules, short arcuate tail and short to mid-length leptoderan bursa. Morphologically, it is closest to F. viminalisae n. sp. From phylogenetic analyses based on sequence of 28S rDNA D2/D3, F. tumifaciens is closest to F. viminalisae n. sp., with 3-bp differences between them in a 852-bp DNA fragment. The blastn search of the 18S rDNA revealed that F. t u m i f a ci e ns is closest to F. morrisae (V330) and F. brittenae (V’s 56 and 201) with 6-bp differences and 2 gaps in the 1561-bp sequenced DNA fragment. The blastn search of the mtCOI revealed that F. tumifaciens is closest to vouchers 65, 1 and 2, but with 33, 41 and 46-bp nucleotide differences respectively among 578-bps of the sequenced DNA fragment. Thus, sequencing corroborates the species status of F. tumifaciens .

Currie (1937) described F. tumifaciens from parthenogenetic females and males only from ‘leafy’ leaf bud galls on Eucalyptus bridgesiana from Canberra, ACT. He published a photograph of the galls (Plate VI, Fig. 5 , in Currie 1937), and the galls collected from Jindera near Albury were similar. ‘Leafy’ leaf bud galls are a type of shoot bud gall, formed from fusion of up to five leaf blades. Leaf tissue commonly extends past the gall. The associated fly larvae collected from Jindera have dorsal shields similar to those of Fn. carteri (G.S. Taylor, unpub. data), the type fly associate. Parthenogenetic females have similar body shapes; tail shapes (short, arcuate, conoid, tapering to a bluntly rounded tip); V (77–89 vs 80%); ratio a (6–9 vs 7–8); and the head end is similarly small relative to anterior body width. The males have similar body shape and length (314–395 µm vs Currie’s 415 µm); tail shape (short, conoid, tapering to a bluntly rounded tip); ratio a (10–12 vs 8–9); and spicule shape and length (angular, 18–22 vs 21 µm).

Although similar in many ways, there are five important morphological differences between the F. tumifaciens specimens depicted by Currie and those collected by K.A. Davies from Jindera. Currie (1937) provided no scales for his drawings, and did not give the number of nematodes measured. 1) Stylet length of the parthenogenetic females differs (19 µm in Currie vs 9–12 µm). No described Fergusobia sp. since Currie has a stylet of 19 µm, and the longest observed was about 15 µm (K.A. Davies unpub. data). Currie’s stylet measurement may be an error due to the poorer quality microscopes available in the 1930’s, or it could be a typographical error. The male of F. tumifaciens has a stylet about 12 µm long both in Currie and here. 2) The parthenogenetic females and males are of similar size (both 415 µm long) in Currie’s description, but in the specimens from Jindera the parthenogenetic females are shorter (average 280 µm vs 359 µm in the males). Currie did not describe the pre-parasitic infective female, which in the Jindera specimens is a similar size to the males, and may have been a source of confusion for him. It is unusual for both males and parthenogenetic females of Fergusobia to have exactly the same length (K.A. Davies, unpub. data), and this may be an error in his paper. In addition, as drawn, his parthenogenetic female appears to be smaller than the male. 3) In Currie’s drawings, the dorsal pharyngeal gland appears to abut the intestine, and to have dorsal and ventral sectors of similar size. In all specimens of Fergusobia examined, the gland has a definite dorsal overlap of the intestine, and the ventral sector is small or non-existent (K.A. Davies, unpub. data). Currie’s drawings appear to be idealized. 4) Tail length is shorter in the parthenogenetic females (12–21 µm vs 39 µm) and males (18–22 µm vs 40 µm) collected from Jindera than in those measured by Currie. This is also reflected in the different ratio c values (15–22 vs 10–11 and 14–20 vs 10–12). This may be associated with uncertainty in the location of the anus in the parthenogenetic female when viewed with the earlier optics, but seems unlikely for the more prominent cloaca of males. 5) The bursa is shorter in the specimens from Jindera than recorded by Currie (arising at 9–16 vs 35% of body length).

The Jindera specimens were collected from the same host as F. tumifaciens , from a similar gall form, and associated with fly larvae having similar morphology, but the nematodes have some morphological differences. These differences may be related to geographic separation with the earlier collections from near Canberra and the later from near Albury, i.e., both are from the climatically similar southern tablelands region of NSW but about 220 kilometers apart. Albury is at a generally lower altitude than Canberra. The Jindera specimens may represent a new species. It is also possible that a species complex is associated with ‘leafy’ leaf bud galls on E. bridgesiana . Because F. tumifaciens is the type species for the genus Fergusobia , it is important that this be resolved. It is not possible to carry out cross-breeding experiments with Fergusobia , but the two collections are similar in many ways and seem to occupy a similar ecological niche. Extensive searches and queries of former staff of the Australian Museum, Sydney, and ANIC, Canberra have failed to locate the holotype or any paratypes for the species and hence the genus, so it is considered lost. Currie, the original author, is now deceased, and left Australia soon after the collections were made. If a new species is proposed for these nematodes from ‘leafy’ leaf bud galls on E. bridgesiana , there would be strong arguments that F. tumifaciens would become species inquirendum. In this case, F. curriei Fisher & Nickle 1968 —the next species described in the genus—would have priority and become the type. However, the status of F. curriei is also controversial, having been synonymised with F. tumifaciens by Siddiqi (1986) but restored by Davies et al. (2010a). In the interests of promoting stability within the genus Fergusobia , we decided to re-describe F. tumifaciens from the material collected from Jindera, i.e., to establish a neotype for the genus.

This decision is supported by the one collection of nematodes made by K.A. Davies from ‘leafy’ leaf bud galls near Canberra in January 2004, closer to Currie’s collection site. He gives the type location as ‘Canberra’. In the collection made by Davies, the galls were heavily parasitized by wasps, and only 9 Fergusobia nematodes were collected (WINC 2351, WNC 004790). They comprised 8 small parthenogenetic females with short tails like those collected from Jindera, and one male collected in poor condition. The spicule of this male is similar to that of the males from Jindera. These specimens are probably F. tumifaciens , but are not in good enough condition for neotype selection.

Therefore, because the parthenogenetic and male nematodes described here from E. bridgesiana were collected from an area not unlike that of the type locality, from a similar gall form on the type host and associated with Fergusonina flies (probably Fn. carteri ) having larvae with a similar body shape and shield morphology, and are in many respects similar to the description and diagnosis given by Currie (1937), these nematodes are considered to be F. tumifaciens .

Morphologically, Fergusobia tumifaciens is most similar to F. viminalisae n. sp., F. fisheri Davies & Lloyd 1996 , F. microcarpae Davies 2013 (in Davies et al. 2013a), and F. p oro s a e Davies 2013 (in Davies et al. 2013a).

The parthenogenetic female of F. tumifaciens (usually C-shaped) differs from that of F. i nd i c a ( Jairajpuri 1962) Siddiqi 1986 (tight C-shape); and from F. rileyi Davies 2012 (in Davies et al. 2012a) (almost straight to arcuate). In length (224–307 µm), it is shorter than the parthenogenetic female of F. brittenae (329–461 µm), F. cosmophyllae (354–406 µm); F. delegatensae Davies 2012 (in Davies et al. 2012d) (345–527 µm), F. floribundae (352–466 µm), F. magna Siddiqi 1986 sensu Davies 2010 (in Davies et al. 2010b) (500–780 µm), F. pimpamensis Davies 2012 (in Davies et al. 2012b) (313–429 µm), and F. ptychocarpae Davies 2008 (in Taylor & Davies 2008) (320–461 µm). The stylet (9–11 µm) is longer than in F. c ur r i e i (5–8 µm), F. juliae Davies 2012 (in Davies et al. 2012b) (5–7 µm), F. minimus Lisnawita 2013 (in Davies et al. 2013b) (4–8 µm), and F. morrisae (7–8 µm); and tends to be shorter than in F. camaldulensae (11–13 µm). In having large pharyngeal glands (b’ 1.8–2.4), it differs from F. brevicauda Siddiqi 1994 which has smaller glands, and from F. quinquenerviae Davies and Giblin-Davis 2004 which has larger glands with an extra lobe or flexure. It lacks the swollen cuticle of F. jambophila Siddiqi 1986 and F. pohutukawa Davies 2007 (in Taylor et al. 2007). In having a body posterior to the vulva that narrows gradually, is arcuate and conoid in shape, with a bluntly rounded tail tip, the parthenogenetic female of F. tumifaciens differs from that of F. camaldulensae , F. fasciculosae , F. brevicauda , F. cajuputiae Davies and Giblin-Davis 2004, F. leucadendrae Davies and Giblin-Davis 2004 , and F. viridiflorae Davies and Giblin-Davis 2004 (with broadly rounded tips); F. dealbatae Davies and Giblin-Davis 2004 , F. eugenioidae Davies 2012 (in Davies et al. 2012b), F. morrisae , F. philippinensis Siddiqi 1994 (more slender, arcuate to straight); and from F. diversifoliae Davies 2013 (in Davies et al. 2013b) and F. planchonianae n. sp. (body narrows rapidly behind the vulva). In length (12–21 µm), the tail of the parthenogenetic female of F. tumifaciens is shorter than in F. indica (50–65 µm), and tends to be shorter than in F. nervosae Davies and Giblin-Davis 2004 (20–30 µm). The ratio c’ (1–1.7) is smaller than in F. philippinensis Siddiqi 1994 (1.9–2.3). The parthenogenetic female of F. tumifaciens is similar to but can be separated from that of F. fisheri on the position of the hemizonid (respectively, 1–2 and 7–8 annules in front of the excretory/secretory pore). The parthenogenetic female can be separated from that of F. p oro s a e only by tending to have the conoid tail with a broader base, giving it an apparently larger volume. The parthenogenetic female of F. tumifaciens is morphologically similar to that of F. microcarpae and F. colbrani Davies 2014 (in Davies et al. 2014). It is separated from the latter by tending to have a longer stylet (respectively, 9–11 vs 7–9 µm long) and by having a narrower tail tip (bluntly vs broadly rounded). It is morphologically close to F. viminalisae n. sp., but the head capsule of F. tumifaciens is smaller compared to the anterior body diameter than in F. viminalisae n. sp.

The infective female of F. tumifaciens (J-shaped) differs in shape from F. brittenae , F. cajuputiae , F. camaldulensae , F. colbrani , F. cosmophyllae , F. dealbatae , F. delegatensae , F. diversifoliae , F. fasciculosae , F. leucadendrae , F. nervosae , F. microcarpae , F. philippinensis , F. planchonianae n. sp., and F. viridiflorae (arcuate to open C-shape); and from F. po ro s ae and F. r i l e yi (almost straight). In length (354–445 µm), it is smaller than the female of F. magna (537–633 µm); and larger than those of F. quinquenerviae (259–325 µm). The ratio a (9.3–12.3, mean 10.9) tends to be smaller than in F. floribundae (10.7–13.6, mean 12.0). Stylet length (7–10 µm) is longer than in F. minimus (4–6 µm). The infective female can be separated from the female of F. eugenioidae in having a flat rather than a raised circum-oral area. It lacks the cuticular plate around the vulva that is found in F. juliae . The pharyngeal gland in F. tumifaciens is larger than in F. morrisae (respectively, b’ ratio is 3.7–5.2 vs 4.4–6.9). Morphologically, it is difficult to separate the infective female of F. tumifaciens from F. ptychocarpae , but the tail of F. tumifaciens tends to be longer relative to the diameter at the anus (c’ ratios of 0.7–1.2 (mean 1.0) compared to 0.6–0.9 (mean 0.8) in F. ptychocarpae ). The infective female of F. pimpamensis , with a c ’ ratio of 1.5–2.4 (mean 2.1), has a tail that is longer relative to diameter at the anus than in F. tumifaciens . The infective female of F. tumifaciens mostly has a shorter tail than in F. viminalisae n. sp. (12–29 (mean 20µm) vs 21–55 (mean 31µm)). It is not possible morphologically to separate the infective female of F. tumifaciens from that of F. brevicauda , F. curriei , and F. fisheri .

The male of F. tumifaciens , almost straight to barely J-shaped, differs from F. diversifoliae , F. floribundae , F. magna , F. pimpamensis , F. ptychocarpae and F. viridiflorae (C-shaped); and F. juliae (strongly J-shaped). It is longer (314–395 µm) than the male of F. nervosae (277–312 µm); and smaller than that of F. pohutukawa (398–469 µm). The ratio a (9.9–11.7) is smaller than in the male of F. pohutukawa (12.2–15.5). The stylet (8–10 µm) is shorter than in F. rileyi (11–13 µm); and is longer than in F. minimus (4–7 µm). The shape of the tail (arcuate with a bluntly rounded tip) differs from that of F. philippinensis (truncate tip). The male tail is shorter than in F. curriei (18–25 vs 36–38 µm). The ratio c (14–20) of the male of F. tumifaciens is greater than in F. cajuputiae (6.8–9.6), F. dealbatae (8.3–10), F. leucadendrae (7.1–10.9), F. microcarpae (6.7–11.9), and F. ptychocarpae (7.6–8.6). The ratio c’ (1.0–1.7) is smaller than in F. jambophila (1.7–2.5). In having an angular spicule, F. tumifaciens differs from F. jambophila and F. pimpamensis , in which it is arcuate. The bursa is short (9–16% of body length) and almost peloderan, differing from F. diversifoliae , F. morrisae and F. planchonianae in which it arises near the secretory/excretory pore, from F. delegatensae in which it ends just posterior to the cloaca, and from F. brevicauda , F. camaldulensae , F. colbrani and F. viminalisae n. sp. (respectively, arising at about 33, 50–70, 30–65, 30–65 and 40% of body length). In having a flat ‘head’ with a small opening for the stylet, the male is separated from F. eugenioidae , in which it is peaked. The male lacks the large secretory/excretory cell of F. brittenae . The male of F. fasciculosae is mostly smaller than that of F. tumifaciens (length 274–336 vs 314–395 µm). The hemizonid is 1–2 annules anterior to the secretory/excretory pore in F. tumifaciens , compared to 10 in F. cosmophyllae and 4–5 in F. porosae . Morphologically, males of F. tumifaciens can be separated from F. fisheri only on the shape of the spicule, which has a longer manubrium and shaft compared to the blade than in F. fisheri . The male has a smaller pharyngeal gland than that of F. quinquenerviae (b’ 2.9–5.1 vs 1.9–2.6).

Figures

FIGURE 2. Fergusobia tumifaciens from E. bridgesiana (all in lateral view): A, Entire parthenogenetic female; B, Head of parthenogenetic female; C, Habitus; D, Tails of parthenogenetic females; E, Entire infective female; F, Head of infective female; G; Habitus; H, Tails of infective females; I, Entire male; J, Head of male; K, Habitus; L, Tails of males; M, Spicules. Scale bars: A, E, I = 50 Μm. B, F, J, M = 5 Μm. Tails not drawn to scale.

FIGURE 3. Fergusobia tumifaciens from E. bridgesiana (in lateral view), re-drawn from Currie (1937): A, entire parthenogenetic female; B, male. No scales given on original drawing.

FIGURE 5. Fergusobia viminalisae n. sp. from E. viminalis (A-L lateral view, M ventral view): A, Entire parthenogenetic female; B, Head of parthenogenetic female; C, Habitus; D, Tails of parthenogenetic females; E, Entire infective female; F, Head of infective female; G; Habitus; H, Tails of infective females; I, Entire male; J, Head of male; K, Habitus; L, Tails of males; M, Ventral view of male tail. Scale bars: A, E, I = 50 Μm. B, F, J = 5 Μm. Tails not drawn to scale.

FIGURE 7. Habitus drawings of Fergusobia spp. collected from ‘ leafy’ leaf bud galls: A, F. tumifaciens from E. bridgesiana; B, F. viminalisae n. sp. from E. viminalis; C, MSp 76 from E. siderophloia; D, F. planchonianae n. sp. from E. planchoniana; E, MSp 68 from E. aromaphloia. Scale bars = 50 Μm.

Tables

TABLE 2. Measurements (µm) of Fergusobia tumifaciens from E. bridgesiana (mean ± standard deviation (range )); including measurements from Currie (1937) for comparison.

	This work Currie (1937)	This work Currie (1937)
n	19	20 19
Length 270	280±20.5 415 (224–307)	359±22 415 397±28 (314–395) (354–445)
a 7.4	7.3±0.7 7–8 (6.2–8.9)	11.1±0.6 8–9 10.9±0.8 (9.9–11.7) (9.3–12.3)
b’ 2.5	2.1±0.2 3–4 (1.8–2.5)	4.1±0.5 3–4 4.2±0.5 (2.9–5.1) (3.7–5.2)
c 15.2	18.2±2.2 10–11 (15.2–22.1)	16.1±1.6 10–12 20.6±4.4 (13.6–20.0) (14.5–29.5)
c’ 1.6	1.3±0.2 (1.0–1.7)	1.3±0.2 1.0±0.2 (1.0–1.7) (0.7–1.2)
V% 85.5	85.9±2.4 79 (76.9–89.4)	81.0±1.4 (79.3–84.2)
T%		76.6±3.6 (65.7–80.9)
Body 36 diameter	40±5 56 (34–52)	32±2 49–50 36±1 (28–36) (34–38)
Stylet 9 length	10.2±0.7 19 (9–11)	9.5±0.5 12 8.4±0.7 (8.5–10) (6.5–9.4)
Ant. end to 77 S/E pore	74±8 90 (52–87)	92±13 89 97±20 (78–123) (77–147)
Spicule length		20.0±1.2 21 (18–22)
Tail length 18	16±2 39 (12–21)	22±2 40 20±4 (18–25) (12–29)