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A New Species of Procyrnea Chabaud 1975 (Nematoda: Habronematidae) and Redescription of Two Chewing Lice (Mallophaga) from the Palawan Hill , Gracula religiosa palawanensis (Passeriformes: Sturnidae), Philippines

Salcedo L. Eduardo1*, DVM, MS, PhD and Sr. Fidelia D. Villa, DVM, SPC2

1Department of Veterinary Paraclinical Sciences, College of Veterinary Medicine, University of the Philippines Los Baños, College 4031, Laguna, Philippines 2Our Lady of Chartres Convent, Antipolo City, Philippines

ABSTRACT

Thirty four live and 30 dead Palawan hill , Gracula religiosa palawanensis were examined for ecto- and endoparasites, respectively at different periods. Specimens of nematodes from the dead and chewing lice (Mallophaga) from the live birds were collected. Specimens were fixed and stored in 70% ethyl alcohol until further study. Nematode specimens were examined in lacto-phenol as temporary mounts. Lice were processed for examination by cleaning in 5% potassium hydroxide solution, washing in distilled water, dehydrating in increasing grades of ethyl alcohol, clearing in terpineol and mounting in Canada balsam. Representative specimens of lice were stained with Ziehl-Neelsen (carbol-fuchsin) in absolute ethyl alcohol before clearing in terpineol and mounting in Canada balsam. The nematode specimens consisted of three species. One of which proved to be new to science and is named, described and illustrated as Procyrnea graculae with prevalence rate of 20%. It is differentiated from closely related species of the genus. The other two were provisionally identified as Synhimantus (Dispharynx) sp. and Tetrameres (Tetrameres) sp. with prevalence rates of 10% and 3.3 % respectively, due to insufficient number of specimens for examination. The lice collection consisted of two species namely, Myrsidea invadens (Kellogg & Chapman, 1902) of the order Amblycera, family Menoponidae and Brueelia chayanh Ansari 1955 of the order Ischnocera of the family Philopteridae with prevalence rates of 11.76% and 23.52%, respectively. They are redescribed and illustrated based on the present materials, and reported for the first time in the Philippines constituting new locality records for the species.

Key words: Brueelia chayanh, Mallophaga, Myrsidea invadens, Palawan hill myna, Procyrnea graculae, nematodes, Gracula religiosa palawanensis

Philipp. J. Vet. Med., 48 (2): 77-85, 2011

INTRODUCTION Gracula religiosa palawanensis commonly known as Palawan hill myna or Palawan Mynas together with the belong talking myna, in recent years has been captured to the family Sturnidae of the avian order in large numbers for the pet trade and Passeriformes. Philippine mynas include four extirpated in some areas (Kennedy et al., 2000). species and their races as follows (Kennedy et Captive breeding would become necessary to al., 2000): Sarcops calvus (coleto) with three save this endemic species from possible races, calvus, melanotus & lowii; Acridotheres extinction. Knowledge of the health aspects of cristatellus (crested myna) with one race, this bird including parasitism would be useful cristatellus; miranda () to make the program successful. which is monotypic and Gracula religiosa (hill The latest summary of the parasites of myna) with one race, palawanensis. The last Philippine birds revealed that only very few two species are endemic to the Philippines. parasites of Philippine mynas have been reported (Price et al., 2003; Eduardo, 2007). *FOR CORRESPONDENCE Opportunities to examine Palawan hill myna 78 SL EDUARDO AND FD VILLA

arose when birds were confiscated from pet Measurements were made with the aid of a traders and kept at the Parks and Wildlife calibrated eyepiece micrometer. Photo- Bureau (PAWB) and a few additional birds kept micrographs were taken using an Olympus at the Laguna Wildlife Park & Rescue Center BX51 research microscope with digital camera (LWPRC). The authors were allowed by the attachment. Drawings were made with the aid respective authorities of these centers to of an Olympus drawing apparatus attached to examine these birds for parasites. This paper an Olympus CX31 research microscope. presents our observation on the parasites Voucher and type specimens were deposited collected from these birds. in the Parasite Collection Center of the College of Veterinary Medicine, UPLB. MATERIALS AND METHODS RESULTS Thirty four live and 30 dead Palawan hill mynas, Gracula religiosa palawanensis were Only nematodes and Mallophaga were found examined for ecto- and endoparasites, in and on the birds examined, respectively. The respectively at different periods. Birds were kept species of parasites, their organ location and at the Parks and Wildlife Bureau (PAWB) of their prevalence based on the 30 dead and 34 the DENR and at the Laguna Wildlife Park and live birds examined are given in the table below. Rescue Center (LWPRC). Most birds were confiscated from their owners by DENR Table. Species of Nematoda and Mallophaga found personnel. Dead birds were kept in deep freezer in 30 dead and 34 live Palawan Hill Myna until examined. (Gracula religiosa palawanensis), respectively Lice were picked out with thumb forceps and their prevalence rates. during careful examination of feathers of live Group: Species Organ location Prevalence birds. Nematodes were collected from autopsied (Family) (%) Nematoda: (n=30) birds following the sedimentation-decantation Procyrnea graculae n. Proventriculus 20 technique. Collected parasites were fixed and sp. and gizzard stored in 70% ethyl alcohol until further study. (Habronematidae) Nematode specimens were cleared and Synhimantus Proventriculus 10 examined as temporary mounts in lacto-phenol (Dispharynx) sp. (Acuariidae) solution or in glycerin after slow evaporation at Gizzard 3.3 room temperature of the alcohol in glycerin- Tetrameres (Tetrameres) sp. alcohol solution, after which, they were returned (Tetrameridae) to 70% ethyl alcohol for storage. Some Mallophaga: (n=34) proventricular and gizzard tissues were Myrsidea invadens Body feathers 11.76 sectioned following the standard paraffin (Menoponidae) method and stained in hematoxylin-eosin. Brueelia chayanh Wing feathers 23.52 (Philopteridae) Lice specimens were processed for

examination first by keeping them in 5% The nematode specimens consisted of three potassium hydroxide solution until the soft species. One is here identified as a new species tissues, mainly the digestive tract and its and is described and illustrated below as contents can be expelled easily from the body Procyrnea graculae (Figs. 1-6) of the family by teasing and pressing carefully with a fine Habronematidae. The other two however were needle under a stereomicroscope. These were insufficient in number to warrant identification then washed in distilled water, dehydrated in to species level and full description. Until increasing grades of ethyl alcohol, cleared in additional materials are made available, they terpineol and mounted in Canada balsam. are provisionally assigned as Synhimantus Representative specimens were stained in Ziehl- (Dispharynx) sp. (Fig.7) of the family Acuariidae Neelsen (carbol fuchsin) in absolute alcohol, also and Tetrameres (Tetrameres) sp. (Fig. 8) of the cleared in terpineol and mounted in Canada family Tetrameridae following the keys to the balsam. NEMATODA AND MALLOPHAGA FROM THE PALAWAN HILL MYNA 79

Figs. 1-5. Procyrnea graculae, new species. 1. Anterior end at region of the whole esophagus. Note position of cervical papillae and nerve ring, and the two parts of the esophagus, muscular and glandular. 2. Anterior end, en face view. 3. At the vulvar region. 4. Posterior end of female, lateral view. 5. Posterior end of male, ventral view. Note asymmetrical distribution of caudal papillae and unequal spicules. genera of the Order Spirurida by Chabaud Myrsidea invadens and Brueelia chayanh by (1975). Tetrameres (Tetrameres) sp. was Kellogg & Chapman (1902) and Ansari (1955), observed only in tissue sections of the gizzard. respectively were inadequate to conform to The lice specimens consisted of two species present day standard of mallophagan diagnosis. namely, Myrsidea invadens (Figs. 9-15) of the As far as the authors are aware, no other suborder Amblycera, family Menoponidae and subsequent descriptions have been made for Brueelia chayanh (Figs. 16-20) of the suborder these two species. They are therefore Ischnocera, family Philopteridae. Although redescribed and illustrated below based on the previously known, the original descriptions of present specimens. 80 SL EDUARDO AND FD VILLA

Nematoda anterior end. Vulva is 1.98-2.30 mm from anterior end. Vagina is tube-like and 0.23-0.28 Description: mm long. Anus is located 0.22-0.29 mm from Procyrnea graculae Eduardo & Villa, posterior extremity. Eggs in utero measure 39.5- new species 43.3 by 21.6-23.3 . (Figs. 1-6) Type host: Gracula religiosa palawanensis (Sharpe, 1890) (Passeriformes: Male (n=16): Body measures 7.6-8.00mm Sturnidae), Palawan hill myna. length and 0.31-0.37 mm in greatest width Organ location: Proventriculus and gizzard. attained at middle of body. Cuticle with Type locality: Palawan (Kept at PAWB and striations measuring 6.25-12 , wider at middle LWPRC). than in both anterior and posterior parts of the Type specimens: Holotype (male), paratypes. body, Mouth with two lips or pseudolabia Deposited in the Parasite Collection and surrounded by four pairs of papillae, two inner Reference Center, College of Veterinary and two outer pairs. Oral aperture is slit-like. Medicine, UPLB. Buccal capsule measures 0.028-0.030 mm in Etymology: Specific epithet is after the depth and 0.02-0.03 mm wide, with thick genus name of the host. sclerotized lining. Muscular and glandular parts of esophagus measure 0.45-0.46 mm and 2.56- Mallophaga (chewing lice) 2.69 mm, respectively. Cervical papillae are anterior to nerve ring and 0.173-0.176 mm from Redescription: anterior end. Nerve ring about 0.30-0.32 mm Myrsidea invadens (Kellogg & Chapman, from anterior extremity. Excretory pore is 9.25- 1902) 0.30 mm from anterior end. Spicules unequal, (Figs. 9-15) left (longer) measures 1.20-1.42 mm long and right (shorter) 0.35-0.37 mm long, the right Male (n=3): As in Fig. 9. Hypopharyngeal about 3.2-4.0 times longer than the right. sclerite as in Fig. 11. Gula as in Fig. 12. Margin Gubernaculum is absent. Papillae with short of pronotum with 3 short setae on each side. peduncles, round on top view and distributed Metanotum with 4 marginal corner setae on asymmetrically as follows: precloacal, 3 on the each side (3 short lateral and 1 very long left and 4 on the right side; postcloacal, 2 on median). Metasternal plate as in Fig. 13, large the left, 1 on the midline and 1 on the right and with 6 long setae. Abdomen with all tergites two minute papillae on each side anterior to the undivided and of equal length, without anterior mucron. Cloacal aperture is about 0.31-0.36 mm setae. Post spiracular setae extremely long on from posterior end. Posterior end is extended II, IV and VIII, very long on I and VII, and long into a mucron-like appendage measuring 0.017- on III, V and VI. Marginal tergal setae: I=12, 0.200 mm long. II=11-12, III-VI = 14-16, VII = 10-13, VIII = 8- Female (n=14): Body measures 12.03-14.45 10 without median tergal setae, VIII with very mm long and 0.42-0.47 mm in greatest width long seta flanked by shorter setae, none with attained at middle of body. Cuticle with anterior setae. Sternite I without setae. striations as in males, striations measure 6.25- Genitalia and genital sac sclerite as in Figs. 14 13.7 microns. Mouth and buccal capsule are as and 15, respectively. Dimensions: Temple in males. Buccal capsule measures 0.023-0.031 width (TW) 0.430-0.482 mm, head length (HL) mm in depth and 0.019-0.025 mm wide. 0.301-0.412 mm, prothorax width (PW) 0.234- Muscular and glandular parts of esophagus 0.311 mm, metathorax width (MW) 0.293-0.376 measure 0.48-0.53 mm and 0.28-0.33 mm mm, abdomen width (AW) 0.354-0.572 mm (at respectively. Cervical papillae are anterior to segment V), male genitalia length (GL) 0.330- nerve ring and 0.21-0.23 mm from anterior end. 0.387 mm. Total body length (TBL) 1.195-1.877 Nerve ring is 0.34-0.39 mm from anterior mm. extremity. Excretory pore is 0.39-0.43 mm from NEMATODA AND MALLOPHAGA FROM THE PALAWAN HILL MYNA 81

Figs. 6-8. Nematodes. 6. Procyrnea graculae, male, whole. 7. Synhimantus (Dispharynx) sp., anterior end. 8. Tetrameres (Tetrameres) sp. in section of proventricular tissue (stained with hematoxylin-eosin). Female (n=5): As in Fig. 10. Margin of compression made by VII which is expanded in pronotum with 1 short, 2 long setae each side. its antero-median margin. Post spiracular setae Hypopharyngeal sclerite, gula as in the male. extremely long on II, IV and VIII, very long on Metanotum with 4 marginal corner setae (3 I and VII, and long on III, V and VI. Marginal short lateral and 1 very long median). tergal setae: I=12, II=11-12, III-VI = 14-16, VII Metasternal plate as in the male, large with 6 = 10-13, VIII = 8-10 without median tergal long setae. Abdomen with all tergites undivided setae, VIII with very long seta flanked by and of equal length, without anterior setae. shorter setae, none with anterior setae. Sternite Tergite V-VI arching medially due to I without setae. Subgenital plate formed from 82 SL EDUARDO AND FD VILLA

Figs. 9-15. Myrsidea invadens. 9. Male, whole, ventral view. 10. Female, whole, ventral view. 11. Hypopharyngeal sclerite. 12. Gula. 13. Metasternal plate. 14. Male genitalia. 15. Male genital sac sclerite. (Figs. 9 & 10, stained with carbol fuchsin) fusion of sternite VIII-IX, its posterior margin with 15-18 setae in region of fused VIII-IX. Anal Brueelia chayanh Ansari 1955 fringe with 25-29 dorsal and 18-24 ventral (Figs. 16-20) setae. Dimensions: TW 0.450-0.575 mm, HL 0.323-0.3985 mm, PW 0.301-0.392 mm, MW Male (n=4): As in Fig. 16. Gula as in Fig. 0.436-0.497 mm, AW 0.593-0.658 mm (at 18. Chaetotaxy: prothorax with 1 long seta on segment III-IV), female anus width (ANW) each posterolateral corner; pterothorax with 4 0.132-0.187 mm across.TBL1.462-1.753 mm. setae (1 long, 1very long, 1 long, 1 very long) on NEMATODA AND MALLOPHAGA FROM THE PALAWAN HILL MYNA 83 each postero-lateral margin. Tergal setae on graculae in Gracula religiosa of the family each side of abdominal segment II-V = 0, VI- Sturnidae, order Passeriformes. The effects of VII = 1, VIII = 2, IX = 3. Pleural setae on each proventricular and gizzard worms on wild birds side of segment II-III = 0, IV-VII = 2, VIII-IX = have not been clearly elucidated. We have 3. Sternal seta on each side of segments II-III = observed destruction of the epithelium of the 0, IV=1, V-VIII=2. Genitalia as in Fig. 19, 0.157- proventriculus and gizzard in the Palawan hill 0.189 mm long. Dimensions: TW 0.312-0.432 myna infected with the nematodes mentioned mm; HL 0.395-0.423 mm; PW 0.232-0.316 mm, in this study. Foster et al. (2002) have found PL 0.087-0.099 mm, pterothorax width (PTW) gross changes in the texture, color and integrity 0.342-0.397 mm, pterothorax length (PTL) of the cornified epithelium of the gizzard of red- 0.109-0.194 mm; AW 0.423-0.687 mm (at bellied associated with infection segment V-VI), AL 0.780-0.842 mm. TBL 1.201- with Procyrnea pileata. 0.154 mm. The original descriptions of Myrsidea Female (n=5): As in Fig. 17. Gula as in the invadens and Brueelia chayanh by Kellogg & male. Prothorax with 1 medium seta on each Chapman (1902) and Ansari (1955), respectively side. Pterothorax with 6 long setae on each were wanting in details required in present day postero-lateral side. Tergal setae on each side mallophagan diagnosis. As far as the authors of abdominal segments II-IV = 1, V-VII = 2, are aware no other subsequent descriptions VII=0, IX= 3. Pleural setae on each side of were made for these species. They are here segments II-III = 0, IV-VII = 2, VIII = 3, IX = 1. redescribed and illustrated including the Sternal setae on each side of segments II-VI = characters of the genitalia, thoracic and 1. Genital plate as in Fig. 20. Dimensions: TW abdominal chaetotaxy and others. Although 0.401-0.456 mm, HL 0.402-0.486 mm, PW there were studies of the Mallophaga by genera 0.278-0.301 mm, PL 0.103-0.195 mm, PTW group of many of the families of the width 0.356-0.400 mm, PTL 0.125-0.186 mm, Passeriformes, there is none of the family AW 0.597-0.634 mm (at segment V-VI), AL Sturnidae. Based on the character of the genital 1.093-1.206 mm. TBL 1.789-2.040 mm. sac sclerite, Myrsidea invadens may be assigned to the plumosi species group of Myrsidea found DISCUSSION on the Pycnonotidae (Passeriformes) as defined by Hellenthal and Price (2003) on account of The present specimens of the new nematode the character of the male genital sac sclerite. species belong to Procyrnea as first erected and However, this structure in this species is bifid defined by Chabaud (1958) at subgeneric level in its posterior half (Fig. 15). Archawaranon & and later elevated to generic rank (Chabaud, Subinprasert (2005) found Myrsidea invadens 1975). This move has been accepted by and Brueelia chayanh infesting Gracula subsequent authors (Skrjabin & Sobolev, 1963; religiosa intermedia (hill myna northern race) Mawson, 1966; Pinto et al., 1996; Zhang et al., kept in five 33 aviaries in Thailand with 2004). According to Zhang et al. (2004) there prevalence rates of 64.5% and 34.5%, are more than 40 species described under the respectively. The two species were observed to genus, but many of which occur in the avian avoid interspecific competition, the former orders Falconiformes and Strigiformes. preferring the dorsal area and the latter, the Procyrnea graculae stands closest to Procyrnea ventral area of the bird. We obtained in this salomonseni and P. asymmetricum. It differs study much lower rates (11.76% and 23.52%, however from both species in the number and respectively) for the two chewing lice species in distribution of the caudal papillae and ratio of Palawan hill myna. The effects of these lice to the length of the spicules. It differs further from their host are not exactly known. More studies the latter in the number of papillae around the are needed on this aspect. Brueelia spp. have oral opening and body measurements. P. been incriminated as the probable cause of asymmetricum occurs in Circus spp. of the family feather holes in barn swallows and small Accipitridae, order Falconiformes while P. birds (Vas et al., 2008). Myrsidea 84 SL EDUARDO AND FD VILLA

Figs. 16-20. Brueelia chayanh. 16. Male, whole, ventral view. 17. Female, whole, ventral view. 18. Gula. 19. Male genitalia. 20. Female genital plate.

invadens and Brueelia chayanh are reported for Dr. Nielsen Donato of LWPRC for permission the first time in the Philippines constituting new to examine birds in their respective centers. locality records for the species. REFERENCES ACKNOWLEDGMENTS Archawaranon M and Subinprasert S. 2005. Bird- The authors are grateful to Mr. Fernando parasite relations: a hill mynah case study. P. Micosa for assistance in microphotography Journal of Entomology 2: 112–116. and Dr. Theresa Mundita S. Lim of PAWB and Ansari MAR. 1955. Studies on ischnoceran NEMATODA AND MALLOPHAGA FROM THE PALAWAN HILL MYNA 85

Mallophaga infesting birds in the Panjab. Indian Mawson PM. 1968. Habronematinae (Nematoda: Journal of Entomology 20: 46–62. Spiruridae) from Australian birds. Parasitology Chabaud AG. 1958. Essai de classification des 58: 745–767. nematodes Habronematinae. Annales de Skrjabin KI and Sobolev AA. 1963. Principles of Parasitologie Humaine et Comparee 33: 445–508. Nematology. 11. Spirurata of and Man Chabaud AG. 1975. Keys to the genera of the Order and the Diseases Caused by Them. Part 1. Spirurida Part 2. Spiruroidea, Habrone- Spiruroidea. Moscow: Izdatel’vo Akademii matoidea and Acuarioidea. (p. 29-58). In: CIH Nauka, SSSR (in Russian). Keys to the Nematode Parasites of Vertebrates Pinto RM, Vicente JJ and Noronha D. 1996. (Anderson RC, Chabaud A.G. & Willmott S., Nematode parasites of Brazilian Piciformes eds.), England: CAB. birds: a general survey with description of Eduardo SL. 2007. Synopsis of the Parasites of Birds Procyrnea anterovulvata n.sp. (Habro- in the Philippines. National Academy of Science nematodoidea, Habronematidae). Memorias do & Technology: Metro Manila. Instituto Oswaldo Cruz 91: 479–487. Foster GW, Kinsella JW, Walters EL, Schrader MS Price RD, Hellenthal RA, Palma RL, Johnson KP and Forrester DJ. 2002. Parasitic helminths of and Clayton DH. 2003. The Chewing Lice: World red-bellied woodpeckers (Melanerpes carolinus) Checklist and Biological Overview. Special from the Apalachicola National Forest in Publication 24. USA: Illinois Natural History Florida. Journal of Parasitology 88: 1140–1142. Survey. Hellenthal RA and Price RD. 3003. The genus Vas Z, Csörgö T, Møller AP and Rozsa L. 2008. The Myrsidea Waterston (Phthiraptera: Meno- feather holes on the barn swallow Hirundo ponidae) from bulbuls (Passeriformes: rustica and other small are probably Pycnonotidae), with description of 16 new caused by Brueelia spp. lice. Journal of species. Zootaxa 354: 1–20. Parasitology 94: 1438–1440. Kennedy RS, Gonzales PC, Dickinson EC, Miranda Zhang L, Brooks DR and Causey D. 2004. Procyrnea HC and Fisher TH. 2000. A Guide to the Birds Chabaud, 1958 (Nematoda: Habronematoidea: of the Philippines. United Kingdom: Oxford Habronematidae) in birds from the Area de University Press. Conservacion Guanacaste, Costa Rica including Kellogg VL and Chapman BL. 1902. Mallophaga description of 3 new species. Journal of from birds of the Hawaiian Islands. Journal of Parasitology 90: 364–373. the New York Entomological Society 10: 155–169.