and Nematodes (Nematoda) in Bark Beetles

ACTA ZOOLOGICA BULGARICA Acta zool. bulg., 60 (3), 2008: 227-232

Prevalence of Gregarina typographi Fuchs (Apicomplexa: Gregarinidae) and Nematodes (Nematoda) in Bark Beetles (Coleoptera: Scolytinae) from Bulgaria Depending on the Host Gender Danail Takov, Daniela Pilarska Institute of Zoology, Bulgarian Academy of Sciences, 1, Tzar Osvoboditel Blvd, Sofia 1000, Bulgaria; e-mails: [email protected], [email protected]

Abstract:

The prevalence of the protozoan Gregarina typographi and nematodes in males and females of 12 bark beetle species attacking spruce, white and black pine from 10 localities in Bulgaria was investigated. Gr. typographi was detected in 2 bark beetle species and showed higher prevalence in males (37%) and females (31.7%) of Ips sexdentatus compared with I. acuminatus. The highest prevalence of nematodes was in I. sexdentatus. 87.9% of the males and 91% of the females were infected. The lowest percentage was recorded in the males (8.3%) and in the females (16.7%) of Trypodendron lineatum. There were no statistical significant differences in the prevalence of Gr. typographi and nematodes in both genders of all investigated bark beetle species.

Key words: Bulgaria, bark beetles, gender, Gregarina typographi, nematodes, prevalence

Introduction Bark beetles attack coniferous trees every year and cause enormous damages of forest economy, especially during intensive outbreaks (TZANKOV, MIRCHEV 1985). Protozoans and nematodes are one of the most important pathogens infecting bark beetles. In Europe pathogens of Ips typographus have been intensively studied (WEISER 1954, 1955, 1977, WEGENSTEINER 1994; WEGENSTEINER, WEISER 1995, 2004; ZIZKA et al.1997). However there are few publications concerning pathogens of other bark beetle species attacking coniferous trees and their prevalence in both genders (PURRINI 1980, PURRINI, WEISER 1985, KOHLMAYER et al. 2003, HÄNDEL et al. 2003). In Bulgaria TAKOV et al. (2006, 2007) started the first investigations of the distribution and biodiversity of

bark beetle pathogens. They studied 20 bark beetle species including Ips typographus. The aim of this study is to indicate the prevalence of Gregarina typographi and nematodes in male and female bark beetles attacking spruce, white and black pine.

Material and Methods Twelve bark beetle species from ten localities in the mountains Vitosha, Pirin, the Rodopes, Maleshevska and Stara Planina were investigated from March 2003 to October 2006 (Таble 1). The material was collected and prepared according to the method of WEGENSTEINER et al. (1996). 227

Takov D., D. Pilarska Таble 1. Investigated bark beetles, localities and number of male and female specimens Bark beetle species

Number of the investigated specimens

Locality

Male

Female

Ips sexdentatus

Zheleznitsa (Vitosha mountain); Tsalim (Pirin mountain); Tsaparevo (Maleshevska mountain)

108

145

Ips acuminatus

Tsalim, Sandanski (Pirin mountain); Tsaparevo (Maleshevska mountain); Garmen, Borino (the Rodopes); Tremoshtnitsa (Pirin mountain)

134

330

Tomicus piniperda

Borino, Garmen (the Rodopes); Veliushtets (Maleshevska mountain)

80

78

Pityogenes bistridentatus

Veliushtets (Maleshevska mountain)

6

6

Hylastes ater

Tsaparevo (Maleshevska mountain)

2

2

H.attennuatus


12

12

H. cunicularius

Garmen (the Rodopes )

18

20

Polygraphus poligraphus

Artista (Vitosha mountain)

30

31

Orthotomicus erosus

Gorni Lom (Stara Planina mountain)

37

34

O.laricis


3

8

Trypodendron lineatum


12

7

Hylurgus ligniperda


5

6

After revealing of attacked spruce and pine trees, the bark was eliminated. The specimens were collected from the trees bark. After determination of the host species the bark beetles were dissected in vivo under a stereoscope. Fresh smears from the internal organs of the hosts were made in a drop of physiological solution. For discovering of pathogens the preparations were examined with a light microscope Amplival, at magnification 100х, 400х. The gender of the studied host species was determined according to the preserved gonads which are the most precise criterion for the sex determination especially for species with small size (Polygraphus poligraphus, Pityogenes bistridentatus, Hylastes sp.). The χ2 test was used for statistical calculation of differences in infection prevalences (ESSI, 1987).

Results Gregarina typographi Fuchs, 1915 The protozoan Gregarina typographi was identified on the basis of its morphological and morphometrical characteristics. It was found in males and 228

females of Ips sexdentatus and I. acuminatus. The gregarine was localized in the lumen of bark beetle midgut, where stages of the life cycle of the parasite – trophozoits, mature gamonts and sizigia – were observed. The prevalence of Gr. typographi depending on the gender of I. sexdentatus was 37% in the males and 31.7% in females (p>0,05) (Fig.1). For I. acuminatus this proportion was 14.2% for the males and 8.2% for the females (p>0.05) (Fig.1). There was no statistical significant difference in the prevalence of gregarine in males and females of both bark beetle species. Nematoda Nematodes were found in males and females of 12 bark beetle species. They were localized in the haemolymph and digestive system of the bark beetles. According RÜHM (1956) one beetle host can be infected by several nematode species. It was difficult to identify the species of nematodes we have observed in each infected beetle. In many cases we have found only larvae without developed reproductive system. Therefore we have presented the pre-

Prevalence of Gregarina typographi Fuchs... 40

the prevalence of nematodes for males and females showed the following proportions: Tomicus piniperda – 81.2% and 83.3% (p>0.05) (Fig. 2), Pityogenes bistridenatus – 33.3% and 33.3% (p>0.05) (Fig.2), H. attennuatus – 83,3% and 66,7% (p>0.05) (Fig. 3), Polygraphus poligraphus – 60% and 41.2% (p>0,05) (Fig. 3), Hylastes cunicularius – 33.3% and 40% (p>0,05) (Fig. 3), H. ater – 50.0% and 50.0% (Fig. 3), Orthotomicus erosus – 83.8% and 67.6% (p>0.05) (Fig. 4), Orthotomicus laricis – 66.7% and 50% (p>0.05) (Fig. 4), Trypodendron lineatum – 8.3% and 16.7% (p>0.05) (Fig. 4), Hylurgus ligniperda 40% and 33.3% (p>0.05) (Fig. 4). As it is seen from the values of p (statistical significance coefficient) no statistical significant differences in the prevalence of nematodes in males and females of the investigated beetles were found.

37 male female

35 31,7

Prevalence, %

30 25 20 14,2

15 10

8,2

5 0

I. sexdentatus

I. acuminatus Bark beetle

Fig. 1. Prevalence of Gregarina typographi in males and females of Ips sexdentatus and Ips acuminatus.

Discussion

valence of the observed nematodes as total. For Ips sexdentatus the prevalence of nematodes was 87.9% in the male and 91.0% in the female bark beetles (p>0.05) (Fig. 2). 73.1% of the males and 66.9% of the females of I. acuminatus were infected with nematodes (p>0.05) (Fig. 2). For the other bark beetles

The protozoan Gregarina typographi was found in males and females in two out of twelve investigated beetles. Its prevalence was higher in males and females of Ips sexdentatus compared with I. acuminatus. TAKOV et al. (2006) studied the prevalence of this protozoan in males and females of I. typo-

100 90

87,9

91

80

73,1 66,9

70 Prevalence,%

81,2 83,3

male female

60 50 40

33,3 33,3

30 20 10 0

I. sexdentatus

I. acuminatus T. piniperda Bark beetle

P. bistridentatus

Fig. 2. Prevalence of nematodes in males and females of Ips sexdentatus, I. acuminatus, Tomicus piniperda and Pityogenes bistridentatus.

229

Takov D., D. Pilarska 90

83,3

male female

80 70

66,7 60

Prevalence, %

60 50

50

50 41,2

40

40 33,3

30 20 10 0

H. attennuatus

H. ater

P. poligraphus

H. cunicularius

Bark beetle

Fig. 3. Prevalence of nematodes in males and females of Hylastes attennuatus, H. ater, Polygraphus poligraphus and H. cunicularius. 90

83,8

male female

80 70

67,6

66,7

Prevalence, %

60 50

50

40

40

33,3

30 20

16,7 8,3

10 0

O. erosus

O. laricis T. lineatum Bark beetle

H. ligniperda

Fig. 4. Prevalence of nematodes in males and females of Orthotomicus erosus, O. laricis, Trypodendron lineatum and Hylurgus ligniperda.

graphus and recorded high prevalence of the gregarine in both genders (up to 56.1% in males and up to 47.1% in the females) for one of the studied sites – Bistrishko Branishte. In single samples collected from 2 locations these authors found significantly higher infection rates of Gr. typographi in male beetles than in the females. However they stated that the overall prevalence of the gregarine in both genders 230

did not differ significantly. Our results are also similar to the results of ZITTERER (2002) who investigated male and female specimens of I. acuminatus and did not find statistically significant differences in the prevalence of the gregarine in this host. The nematodes showed higher prevalence almost in all studied males and females compared with the gregarine. One of the reasons for these results

Prevalence of Gregarina typographi Fuchs... could be the synchronization of bark beetle life cycle with that of the nematodes. In more cases the infection of bark beetles with nematodes begins in larval galleries where the new generation of the host develops and favourable conditions for infection of the bark beetles occur. The highest prevalence of nematodes was 87.9% in the males and 91% in the females of I. sexdentatus. The lowest percentage was detected in the males (8.3%) and in the females (16.7%) of Tripodendron lineatum. The nematode prevalence in both genders was higher than the prevalence of the gregarine. Similarly to Gr. typographi no gender dependant differences in the infection rates were observed for nematodes. This finding is in accordance with the investigations of ZITTERER (2002). The author also did not find statistically significant differ-

ences in the prevalence of nematodes in males and females of Ips acuminatus. We did not find information for the prevalence of nematodes in males and females of other bark beetle species except for I. acuminatus and therefore we were not able to compare our results with these of other authors. The study reported here gives original information about the prevalence values of Gregarina typographi and nematodes in different bark beetle species depending on their gender and suggests that the infection rates values in male and female individuals of all studied species were similar, without statistical significant differences. Acknowledgements: We are especially indebted to eng. Danail Doychev and Elena Tzvetanska for their contributions to this work. We are also grateful to the funding provided by the National Science Fund of Bulgaria, grant B 1312/2003.

References ESSI A. 1987. Statistishe Methoden in der Tierproduction. Wien. Verlagsunion Agrar, 316 p. H ÄNDEL U., R W EGENSTEINER , J. W EISER , Z. Z IZKA 2003. Occurrence of pathogens in associated living bark beetles (Col., Scolytidae) from different spruce stands in Austria. – Anzeiger für Schädlingskunde, 76: 22-32. KOHLMAYER B. J. WEISER, R. WEGENSTEINER , U. HÄNDEL, Z. Z IZKA . 2003. Infection of Tomicus piniperda (Col.; Scolytidae) with Canningia tomici sp.n. (Microsporidia, Unikaryonidae). – J. of Pest Science, 76: 65-73. P URRINI K.1980. Malamoeba scolyti sp.n. (Amoebidae, Rhizopoda, Protozoa) parasitizing the bark beetles Dryocetes autographus Ratz. and Hylurgops palliatus Gyll. (Scolytidae, Col.). – Archiv für Schädlingskunde, Pflanzenschutz, Umweltschutz. 52: 167-183. PURRINI K., J. WEISER 1985. Ultrastructuralstudy of the microsporidian Chytridiopsis typographi (Chytridiopsida: Microspora) infecting the bark beetle Ips typographus (Scolytidae: Coleoptera), with new data on spore dimorphism. – J. of Invertebrate Pathology, 45: 66-74. RÜHM W. 1956. Die Nematoden der Ipiden. – Parasitologische Schriftenreihe, 6: 1-435. TAKOV D., D. DOYCHEV., WEGENSTEINER, R., D. PILARSKA 2007. Study on the pathogens of bark beetles (Coleoptera, Scolytidae) from different coniferous stands in Bulgaria. – Acta zool. bulg., 59 (1): 87-96. TAKOV D., D. PILARSKA, R. WEGENSTEINER 2006. Оccurrence of pathogens in Ips typographus (Coleoptera, Scolytidae) from several Picea abies (L.) (Karst.) stands in Bulgaria. – Acta zool. bulg., 58 (3): 409-420. T ZANKOV G. , P. M IRCHEV 1985. A study with aggregation pheromones on some peculiarities in the development of the spruce bark beetle (Ips typographus L.) and the lineate bark beetle, (Trypodendron lineatum Oliv.) in the biosphere

reserve Parangalica. – Internat. Symp. Conservation of Natural areas and of genetic material they contain, MAB of UNESCO, 23-28.09. 1985. Blagoevgrad, 132-139. WEGENSTEINER R. 1994. Chytridiopsis typographi (Protozoa, Microsporidia) and other pathogens in Ips typographus (Coleoptera, Scolytidae). – IOBC/ wprs Bulletin, 17 (3): 39-42. WEGENSTEINER R. , J. WEISER 1995. A new Entomopoxvirus in the bark beetle Ips typographus (Coleoptera:Scolytidae). – J. of Invertebrate Pathology, 65 (2): 203-205. WEGENSTEINER R., J. WEISER 2004. Annual variation of pathogen occurrence and pathogen prevalence in Ips typographus L. (Col., Scolytidae) from the BOKU University Forest Demonstration Centre. – J. of Pest Science, 77: 221-228. WEGENSTEINER R., J. WEISER, E. FUHRER 1996. Observations on the occurrence of pathogens in the bark beetle Ips typographus L. (Coleoptera, Scolytidae). – J. of Appl. Entomol., 120: 199-204. WEISER J. 1954 a. Prispevek k znalosti cizopasniku kurovce Ips typographus L. I. (Contributions to the knowledge of Ips typographus parasites, II). – Vestnik Ceskoslovenske Zoologicke Spolecnosti, 18: 217-224. WEISER J. 1955. Prispevek k znalosti cizopasniku kurovce Ips typographus L. II. (Contributions to the knowledge of Ips typographus L. parasites II.). – Acta societatis zoologicae bohemoslovenicae, 9: 374-380. WEISER J. 1977. An Atlas of Insect Diseases. Academia, Prague, 240 p. ZIZKA Z., J. WEISER, R. WEGENSTEINER 1997. Ultrstructure of Oocysts of Mattesia sp. in Ips typographus. – J. of Eukaryotic Microbiology, 44: 25A, 98. ZITTERER P. M. 2002. Antagonists of Ips acuminatus (Gyllenhall) with special consideration of pathogens. – Wien. Diploma thesis. Universität für Bodenkultur Vienna. 57 p. Received: 05.08.2008 Accepted: 27.10.2008

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Takov D., D. Pilarska

Екстензивност на заразяване на корояди (Coleoptera: Scolytinae) с Gregarina typographi Fuchs (Apicomplexa: Gregarinidae) и нематоди (Nematoda) от България в зависимост от пола на гостоприемника Д. Таков, Д. Пиларска

(Резюме) Изследвана е екстензивността на заразяване с Gregarina typographi и нематоди при двата пола на 12 вида корояди, нападащи иглолистни видове дървета (смърч, бял и черен бор) от 10 находища в България. Gr. typographi е установена в 2 вида корояди и показва по-висока екстензивност на заразяване при мъжки (37%) и женски (31,7%) на Ips sexdentatus в сравнение с I. acuminatus. Екстензивността на заразяване с нематоди е най-висока при мъжките – 87,9% и женските – 91% на I. sexdentatus. Най-нисък е процентът на нематодите, установен в мъжки (8,3%) и женски (16,7%) на Trypodendron lineatum. Няма статистически достоверни разлики в заразеността при двата пола с Gr. typographi и нематоди при всички изследвани видове корояди.

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