A NEW SPECIES OF PRASADISEIUS WAINSTEIN, 1970 (ACARI: OTOPHEIDOMENIDAE) FROM HAWK MOTHS (LEPIDOPTERA: SPHINGIDAE) IN PERU

During an expedition to Peru in August 2010, a new species of otopheidomenid mite, Prasadiseius incanus Prasad and Guanilo, n. sp. (Acari: Otopheidomenidae), was collected from Xylophanes fusimaculata (Felder, 1874) and Xylophanes undata Rothschild and Jordan, 1903 (Sphinghidae). The female and male of the new species are described using current nomenclature for the idiosomal chaetotaxy and illustrated in a series of photographs. A key for the identification of all species of the genus is provided.


INTRODUCTION
The species of the genus Prasadiseius Wainstein, 1972 (Mesostigmata: Otopheidomenidae) are only known to parasitize hawk moths (Lepidoptera: Sphingidae) and have been found primarily in the neotropical region with two species from other tropical regions of the world (Prasad 1976(Prasad , 2011d. So far, the following seven 7 species of the genus Prasadiseius have been described: P. achlora (Prasad 1972) and P. aporodes (Prasad 1972) from Uganda, P. cocytes (Prasad 1970a) and P. donahuei (Prasad 1970a) from Peru, P. indicus (Prasad 1973) from India, P. kayosiekeri (Prasad 1970b) from Honduras, Panama Canal Zone and Peru, and P. pholusis (Prasad 1970b) from Bolivia and Honduras (Prasad 1970a(Prasad , b, 1972(Prasad , 1973. Recently, P. cocytes was found to be widely distributed in several neotropical countries: Brazil, Ecuador, Guatemala, and Peru (Prasad 2011d). All stages of the mite are found dorsally mainly in the rectangular area of the tympanum, the metathorax, and first abdominal tergite of the moths; occasionally, they are found on the proboscis, around eyes, and the dorsum or venter of the head, thorax, abdomen, and wings. All the species were described from dead specimens collected from museum preserved dead hawk moths that, in some cases, were collected over 50 years before the mites were discovered by the first author. Although the eggs, larvae, protonymphs, deutonymphs, females and males of these mites are often found on these moths when they are heavily infested, the biological parameters of these mites have not been studied yet.
Species of Otopheidomenidae are known to parasitize insects of Hemiptera, Isoptera, Lepidoptera and Orthoptera (Treat 1975;Fain and Lukoschus 1983;Syed and Goff 1983;Halliday 1994;Zhang 1995;Mo 1996). Presently, the family is subdivided into the following 3 subfamilies: Katydiseiinae Fain and Lukoschus, 1983 which infests grasshoppers (Orthoptera) and termites (Isoptera); Otopheidomeninae Treat, 1955 which infests noctuid and sphingid moths (Lepidoptera); and Treatiinae Wainstein, 1972 which infests various true bugs (Hemiptera). The subfamily Otopheidomeninae is comprised of 3 genera: Noctuiseius Prasad, 1968and Otopheidomenis Treat, 1955which infest noctuid moths, and Prasadiseius Wainstein, 1972 which infests sphingid moths. The family Otopheidomenidae is considered to be closely related to the Phytoseiidae (Krantz and Khot 1962;Evans 1963;Chant 1965;Chant and Yoshida-Shaul 1992;Chant and McMurtry 2007) but have adapted to parasitizing various insects. Unlike the chelicerae of phytoseiids which have a fixed digit and movable digit, the otopheidomenids only have the movable digit. In addition, there is a reduction in the number of setae in Otopheidomenidae. The spermatheca of the species in which it is known, are cup-shaped or tubular-shape similar to those found in some of the phytoseiid species. Because of these unique features and their close relation to phytoseiids, otopheidomenid mites have received more attention in recent years, including DNA studies, which are still in progress (pers. comm., 2010, Kreiter -France, Ragusa di Chiara -Italy). Detailed studies of the morphology of these mites using scanning electron micrographs are also being conducted (Prasad and Walker 2011).
Since live otopheidomenid mites on live hawk moths had never been observed before and consequently their biology never studied, the first author organized an international expedition in August 2010 involving Alberto D. Guanilo, an acarologist from Peru; Juan Grados, a lepidopterist from Peru; and Indira Prasad, expedition supporter and lifelong companion of the first author. During this expedition, not only were live otopheidomenid mites observed for the first time on live sphingid moths captured in the Amazonian Forest of Cusco Department, but an interesting new species of Prasadiseius was found which is described herein, thus, making a total of 8 species of Prasadiseius known worldwide.

MATERIALS AND METHODS
After obtaining permit from the Government of Peru months ahead of our visit for collecting the moths and mites, the live sphingid moths were collected at night in transparent plastic jars with screw caps (100 ml, 7.5 x 4.5 cm, NCS Diagnostics Inc., Mississauga, Canada) during August 10 -18, 2010 in different localities of Cusco Department from 7 -10 pm using a mercury vapor light. A Honda generator was used to produce the electricity for the light bulb as no electricity was available in the jungle. Each live moth was photographed before collection using a Ricoh (R8) camera, labeled with a corresponding collection number, and examined next day by first two authors under a Bausch and Lomb stereo binocular microscope having total magnification of 25x for the presence of mites. When mites were observed, the moths with the mites were photographed and a video was taken using a High Definition Kodak EasyShare Z812 IS camera and/or Ricoh camera. After observation on live moths, the mites were collected in 70% ethanol, stored in small glass vials (4.5 x 1.5 cm) having screw caps, and brought to USA for preparation and identification by the first author. The moths were euthanized with 0.2 ml of wind shield washing liquid which was injected into the thorax and then taken to Natural History Museum, Lima, Peru for the identification.
The mites were extracted from the 70% ethanol, and mounted in Hoyer's medium, one specimen per glass microscope slide (3 x 1 inch; 75 x 25 mm), and covered with a #18 mm, zero-thickness round cover slip. Each slide was dried for a week between 45 -47°C on a slide-warming hot plate, rung with Glyptal, and labeled. The mites were identified by the first author using a Accu-Scope 3000 phase-contrast microscope (Acc-Scope, New York, USA) under 400x. Occasionally, mag-nification of 1000x was needed to see the structural and setal details under the oil immersion lens. Many photographs of the mites were taken using the mounted Micrometrics TM camera on the microscope and saved in Photoshop CS2. Measurements were taken directly from the slide mounted specimens using the Micrometrics system.
The hawk moth hosts on which this paper is based were identified by Mr. Juan Grados, lepidopterist and a team member of the expedition. The species of the mite was identified and description details were prepared by the first author. The second author made significant contribution not only by coming from Australia to join the expedition but took hundreds of photos of live sphingid moths at night before the collection from UV light screen and helped in observation and collection of the mites on which this paper is based. The fourth author not only helped in tagging of moths with the collection data at night in the Amazon forest but provided hand-held torch light daily for hours as no electricity was available in the room and provided every supply during examination of the moths. Prasad (2011a) recently provided idiosomal chaetotaxy of all known species of Prasadiseius which is followed in this paper. All measurements were taken from the slide mounted specimens and given in micrometers (µm) with the range and average given in brackets. These were taken, unless otherwise mentioned, at the longest and widest part of the structure. Length of the setae were measured from base to the tip and the distance between the two setae were measured, excluding setae, from inner base of one seta to the inner base of the other seta. Measurements of the length and width of idiosoma and the length of legs (taken in center of each leg from base of coxa to tip of pretarsus) were taken at 100x, the length and width of idiosomal shield and length of palps was taken in 200x, and all other measurements, including the length of setae and distance between the bases of the setae were taken in 400x. Occasionally, some details of setae and structures were observed in 1000x. As most dorsal idiosomal setae were minute and could not be seen clearly at low magnification at the same time (up to 200x), several photos were taken from different locations (up to 400x) to show all the setae. Each photo shows the magnification in which it was taken (200x vs 400x), followed by mite collection number (VP10-36 or VP10-38) and the photo number. These are taken from holotype and paratype mites and are to be used as 'type photos' for the new species described in this paper.
Different standardized abbreviations used in the description of mites in the present study are as given below. It should be noted that if the idiosoma (= ID) is covered dorsally by one shield as in female and male of most Otopheidomeninae, it is called idiosomal shield (= IDS) or dorsal shield (DS) which is divided anteriorly in podosoma or podosomal region (= PO) having podonotal shield (POS) and posteriorly in opisthosoma or opisthosomal region (= OP) having opisthonotal shield (OPS). Chant and McMurtry (2007) used term 'podonotum' for podonotal shield and 'opisthonotum' for opisthonotal shield (OPS). Zhang (1995) used terms podonotal shield and opisthonotal shield for above and referred podonotal area and opisthonotal area when giving the number of setae. The dorsal shield in the adults of Otopheidomenidae is either entire (Eickwortius termes, Zhang 1995), partially incised laterally (Prasadiseius spp.), or separated into 2 shields (Noctuiseius spp. or male of P. pholusis). When together as DS, these are called podonotal shield (podonotum) and opisthonotal shield (opisthonotum) in the description. When both shields are separate from each other, the former is still called podonotal shield and latter is called as opisthonotal shield which has also been called as pygidial shield to indicate its separation from the former. Both above shields in larvae, protonymphs and deutonymphs could be separate (N. treati), together as 1 dorsal shield in female but 2 separate shields in male (P. pholusis) (Prasad, 2011c), or separate in larvae and protonymphs but fused in deutonymphs (some species of Prasadiseius, unpublished data).
Diagnosis -Dorsal shield entire (O. ascalaphae) or incised laterally (females of all remaining species except P. pholusis in which the dorsal shield of the male is divided into 2 separate shields). Dorsal shield with 14 or fewer pairs of setae. Tritosternum absent (Noctuiseius spp. and Prasadiseius spp.) or vestigial (Otopheidomenis spp.). Metasternal setae (ST4) absent. Genital (= epigynial) shield rounded posteriorly and with or without genital setae. Ventrianal shield present by anal shield having only 3 anal setae (PA, PST) surrounding the anus. One or 3 pairs of setae present in between genital shield and anal shield or lateral to anal shield. Metapodal plates absent (in all species except O. zalelestes). Movable digit of chelicera with 3 -13 teeth. Palp trochanter without seta (except in O. ascalaphae with 1 seta). Spermatheca tubular (except in N. batoridgi which is cup-shaped). Males with specialized seta on venter of femur II in several species (e.g. Prasadiseius). No macrosetae on tibia or tarsus of legs I -IV (except O. ascalaphae with 4 large setae on the dorsum of tarsus IV). Parasites of noctuid and sphingid moths.

Prasadiseius incanus Prasad and Guanilo, n. sp. (Figs. 1 -26)
Description Diagnosis -Dorsal idiosoma with 12 pairs of setae of which 11 pairs (j3, j4, j5, j6, J2, J5, z2, z3, z5, Z5, s4) on the large mediolaterally incised dorsal shield and 1 pair (r3) on the anterolateral integument. Setae j4 present, z2 located posterolateral to j3, and peritremes long in female. Sternal (female) or sternogenital (male) shield with 3 pairs of setae; setae ST1 in both female and male not extending beyond the base of ST2, setae ST2 reaching or extending beyond the base of ST3 in same vertical row in female but not in male. Genital shield without genital setae. Male with a very large and thick spinose spur on ventral basifemur of leg II.
Differential -The new species P. incanus Prasad and Guanilo is close to P. indicus but can easily be separated from the latter by having short peritreme, absent S2 and by having a very large and thick spinose spur on ventral basifemur of leg II in the male. None of the species of Prasadiseius or other genera in the subfamily Otopheidomeninae have such a spur.
Female (n = 8, Figures. 1-14, 21, 22A (Figures 1,2), having a well pronounced lateral invagination on each side between setae j6 and J2 (Figure 3) which partially divides the dorsal shield into 2 parts -the podonotal shield comprising approximately the anterior two-thirds, and the opisthonotal shield comprising the posterior one-third of the shield ( Figure  3). Dorsal shield may also have shallow concavities or convexities on the margin of the shield near setae z3, r3 ( Figure 1) and in between Z5 (Figures 1, 4).
Gnathosoma -Similar to that of female, tectum triangular, rounded and smooth anteriorly ( Figure  19). Movable digit of chelicera bearing a pointed spermatodactyl. Hypostome with 3 setae set in a triangle on each side. Posterior medial seta set slightly posterior to posterolateral seta. Corniculi with pointed to blunt apices. Palp length (trochanter to tarsus) = 91 -197 (125). Number of setae on palp segments (palptrochanter to tibia) as in female.

Types
Holotype: Female, with above collection details, deposited in the Natural History Museum, Lima, Peru. Paratypes: 12 females, 4 males, 2 larvae, 5 protonymphs, and 3 deutonymphs on glass slides, and several others in 70% and 100% ethanol for further studies. Slide mounted paratypes deposited as follows: Two females and 1 male in the Natural History Museum, Lima, Peru; 1 female and 1 male in the Museum of Biodiversity, The Ohio State University, 1315 Kinnear Road, Columbus, OH 43212, USA; 1 female and 1 male in the US National Museum, Washington, DC, USA; and others in author's collection.

Etymology
The new species name "incanus" is provided to honor famous Inca tribe and Civilization of Peru that once stretched north to south along the high mountainous Andean range from Colombia to Chile and reached west to east from coastal desert of Atacama to rain forest of Amazon.

Remarks
The location of the z2 pair of setae in relation to the j3 setae seems to be a good species specific characteristic; however, a study of a large series of specimens is necessary before reaching this conclusion.
The length of the peritreme and the number of denticles on movable digit of the chelicerae are also significant in defining a species. The number of denticles in different specimens of P. incanus varies by 1 -2 in the material studied so far.  The position of setae j4 in relation to j5 is variable. Occasionally, an additional j4 or j5 seta may be present (Figure 14) or only 1 of the j4 pair of setae may be present. Seta J2 on one side in female may be absent (Figures 23, 24). Although present in several females, pair of seta J2 and one of J5 on one side was found absent in male (Figures 25, 26). The position of z3 in relation to j4 or to j6 may also vary within the same species. Setae z3 may be located in same or transverse line with j4 -j5 (Fig. 2) or may be slightly anterior to these ( Figure 14). The position of z2 in relation to j3 (anterior or posterior) appears to be of significance in defining a species.
Of all morphological characters studied, the presence of the specific type of spur on venter of femur of leg II in the male is very species specific. Males are often present among the females in populations of 8 or more mites on the host; without the males, it may be difficult to identify a species. Differences in the shape and size of spermatodactyl of different species of Prasadiseius spp. may prove to be useful in distinguishing the different species. It appears to be a complex structure and needs to be studied in higher magnification, along with other gnathosomal structures, including scanning electron micrographs (SEM) of all of the species.
Several questions on the development and bioecology of these mites remain to be answered and need to be investigated in future. Do females infest new moth host first or males, or vice versa, or both together? Are females gravid when they infest a new moth host? In case of heavy infestations, do more than one species infest the same moth? Is there any migration of these mites involved in the case of a heavy infestation? Do any species develop to a tritonymphal stage? The first author has found eggs, protonymphs, deutonymphs, and adult females and males of these mites together on same host but has not observed presence of tritonymphs. As we have seen these mites alive for the first time in Peru, it would be interesting to study these aspects in future in this and other countries where these mites have been reported infesting various sphingid moths by the first author.

CONCLUSION
It was 1970, over 40 years ago, when two new species of the otopheidomenid mites from museum preserved sphingid moths were described by the first author from Peru. Later, after survey of preserved moths in different museums of the world in a brief period of about 3 years, he added 5 more new species from various countries to this list. These mites had never been seen alive before until August 2010 in Peru when collections were made of the new species described in this paper, which brought a total number of eight species of the genus Prasadiseius known from sphingid moths. Research has proved that these ectoparasitic mites are present on many sphingid hosts in various neotropical and tropical countries of the world. This work indicates the likelihood that many new species are yet to be discovered in the future and that surveys of moths housed in museums and those collected in the native countries must be conducted to learn more about these mites. It is unfortunate that the lack of research funding for such work is hampering these discoveries worldwide. As financial resources are limited due to economic crisis in many countries, authors feel that global efforts are necessary to work as team members, as done in the present work, in obtaining the necessary funding and conducting the expedition and exploration for the mite fauna in a few selected countries at a time in future.