Eustachio Tarasco

Occurrence of entomopathogenic fungi in soils in Southern Italy

The natural occurrence of entomopathogenic fungi was investigated in Southern Italian soils using larvae of Galleria mellonella L. (Lepidoptera: Galleriidae) as “bait insect”. Since September (1996) to March (1997) 188 samples of soil were collected from different habitats (woodland, orchard, field, seacoast, grassland, uncultivated land and salt pan). Parasitic fungi were obtained from 14.9% of the soil samples. There were 3 entomopathogenic species: Metarhizium anisopliae (Metchnikoff) Sorokin, Paecilomyces lilacinus (Thom) Samson and Beauveria bassiana (Balsamo) Vuillemin. The most common fungal pathogen was B. bassiana. Both M. anisopliae and P. lilaci- nus were isolated only once. The occurrence of B. bassiana seems to be affected by the soil type and the habitat. Key words: survey, Beauveria bassiana, Metarhizium anisopliae, Paecilomyces lilacinus. FUNGHI ENTOMOPATOGENI NEI TERRENI DELL’ITALIA MERIDIONALE E stata effettuata un’indagine per valutare la presenza e la distribuzione di funghi entomopatogeni nei terreni dell’Italia meridionale utilizzando larve di Galleria mellonella L. (Lepidoptera: Galleriidae) come insetto esca. Da settembre 1996 a marzo 1997, sono stati esaminati 188 cam- pioni di terreno provenienti da diversi biotopi. Sono state rinvenute larve infettate da funghi in 28 campioni (14,9%) e sono state identificate 10 specie di funghi parassiti di cui 3 propriamente entomopatogeni: Metarhizium anisopliae (Metchnikoff) Sorokin, Paecilomyces lilacinus (Thom) Samson e Beauveria bassiana (Balsamo) Vuillemin. Il fungo patogeno rinvenuto con maggior frequenza e risultato essere B. bassiana. La sua presenza sembra correlata alle caratteristiche del suolo e dell’habitat. Parole chiave: indagine, Beauveria bassiana, Metarhizium anisopliae, Paecilomyces lilacinus.

Efficacy and Persistence of Entomopathogenic Nematodes in Controlling Larval Populations of Thaumetopoea pityocampa (Lepidoptera: Thaumetopoeidae)

A 3-year study was conducted in a Pinus halepensis reforestation of Apulia Region (Southern Italy) injecting IJs (infective juveniles) of Steinernema feltiae , S. carpocapsae and Heterorhabditis bacteriophora in aqueous and gel suspensions (Idrosorb SR 2002 [Nigem ® ], and Compex) into the nests of Thaumetopoea pityocampa caterpillar. This study showed that the gel suspensions do not percolate and that slow release of water from the gels allowed nematodes to survive and complete their life cycle in the host. Results demonstrate the feasibility of reducing overwintering larval populations by injecting gel suspension of S. feltiae . We found no negative effects on the endoparasite Phryxe caudata .

Laboratory studies on the competition for insect haemocoel between Beauveria bassiana and Steinernema ichnusae recovered in the same ecological niche

Abstract Laboratory assays evaluated the combined action of the entomopathogenic nematode Steinernema ichnusae and the entomopathogenic fungus Beauveria bassiana isolated from the same ecological niche, an oak forest in Sardinia (Italy). Galleria mellonella larvae were used as the test insect with the aim of understanding what happens in host haemocoel during a simultaneous infection with two different entomopathogens. Larval mortality assays were performed using nematodes and fungi both alone and together, at the same and different times, and in different concentrations. No additive or synergistic effects were observed, but there was a clear antagonism and competition for survival space in the haemocoel. Moreover, mutual effects between the symbiotic bacteria of the entomopathogenic nematode Xenorhabdus bovienii and entomopathogenic fungi were investigated. In laboratory experiments, X. bovienii crude extracts were tested for their activity against fungal growth. Compounds produced by B. bassiana were tested for their activity against the growth of bacteria, revealing that X. bovienii and B. bassiana are antagonistic to each other.

Steinernema ichnusae sp. n. (Nematoda: Steinernematidae) a new entomopathogenic nematode from Sardinia Island (Italy).

A new Steinernema species was isolated from three different sandy soil samples along the Platamona Beach, in the north-west coast of Sardinia Island (Italy). This new species is characterized by the following morphological characters: infective third-stage juvenile with a body length of 866+/-61 (767-969)microm, distance from head to excretory pore of 63+/-2.7 (59-68)microm, tail length of 81+/-3.2 (76-89)microm, ratio E (%) 77+/-3.4 (68-83); male tail with a mucron only in the second generation, spicule length of 66+/-1.4 (64-67)microm and gubernaculum length of 44+/-1.4 (43-46)microm in the first generation male; female of first generation with a slight vulval protrusion and ratio D (%) of 53+/-4.0 (47-63). The new species differs distinctly from the related species (S. feltiae, S. kraussei, S. litorale, S. oregonense and S. cholashanense) in some morphometric values such as percentage of hyaline portion, ratios of gubernaculum/spicule length, spicule head length/width. The DNA analyses of the internal transcribed spacers and D2D3 regions show that the studied nematode isolates are a new species. Cross hybridisation tests with S. feltiae, S. kraussei, S. litorale, S. weiseri and S. oregonense showed that these species were reproductively isolated.

Blowing a nose black and blue

In June, 2006, a 32-year-old woman, working as an architect at the archaeological site of Pompeii, noticed that her nose was blocked, especially the right nostril. When she blew her nose, the mucus was black. A few days later, she developed a dry cough and intermittent fever. Her general practitioner referred her to an otolaryngologist. Nasal examination showed no abnormality, other than mild septal deviation. Skin-prick tests for allergy gave negative results, as did nasal smears for bacteria and fungi; CT of the sinuses showed nothing of note. Nonetheless, the patient’s nasal blockage and rhinorrhoea worsened; she had occasional fever spikes, and was referred to an infectious-diseases specialist. Chest radiography and pulmonary function tests gave normal results; blood tests showed a high concentration of C-reactive protein, mild neutrophilia, and, although no fungal hyphae had been found in nasal smears, antibodies to aspergillus. Oral itraconazole (100 mg daily) was therefore prescribed; 2 months later, no benefi t had been observed. We saw the patient in September, 2007. Nasal endoscopy, CT of the sinuses, and allergy tests again gave negative results. Nasal smears showed no fungi, but we observed black, amorphous bodies, which we interpreted as cellular debris (fi gure). We suggested that the patient clean her nose and sinuses with a douche, which allowed a continuous fl ow of saline through the nose. During the fi rst washing, a small insect was expelled from the nostril; immediate symptomatic improvement ensued. The patient informed us, and the insect was collected and identifi ed as Clogmia albipunctata [=Telmatoscopus albipunctatus]. We deduced that the amorphous black objects were insect faeces. The fever, cough, and nasal obstruction completely resolved within a few days—however, the black discharge continued, although, given the mucociliary clearance rate, we had expected it to disappear in about 2 days. The patient became depressed and frustrated, and feared that she had other insects in her nose, although this possibility was excluded by CT and MRI. During a consultation, we noticed that she wore eyeliner. She confi rmed that she sometimes used a kohl pencil. We asked her to stop. The black secretion disappeared within 2 days. We realised that kohl, mixed with tears, had been passing though an abnormally patent nasolacrimal duct, and produced the blackpigmented bodies previously identifi ed as insect faeces. When the patient wore blue eyeliner, blue nasal secretions appeared. Our lessons from this event? When usual diagnoses for common illnesses such as rhinitis are excluded, refl ect on underlying mechanisms, and consider the (nearly) impossible. In this case, the nasolacrimal duct was, unusually, wide enough to host an insect, and to allow kohl fragments to enter the nose. What caused the systemic infl ammation? The patient did not have conjunctivitis, although the conjunctiva was in direct contact with her eyeliner. Moreover, the infl ammatory symptoms stopped after the fl y was ejected. We therefore blame the fl y. We note, in passing, that commercial kohl can contain more than 25 diff erent substances, some of which can provoke allergy; health problems have been caused by kohl containing lead. But we do not wish to issue a blanket warning against kohl. Ingredients of kohl vary widely. In south Asia, and across much of the Muslim world, traditional recipes are regarded as medicinal. Much-feared Pashtun warriors historically applied kohl before surging into battle.

Oscheius onirici sp. n. (Nematoda: Rhabditidae): a new entomopathogenic nematode from an Italian cave

Oscheius onirici sp. n. (Nematoda: Rhabditidae) was isolated from a karst cave soil of Central Italy. Molecular and morphological analyses were performed. Total DNA was extracted from individual nematodes and the mitochondrial COI, the ITS containing region, the D2-D3 expansion domains of the 28S rRNA gene and the 18S rRNA gene were amplified and sequenced. BLAST search at NCBI by using all molecular markers revealed that this taxon is similar to Oscheius species. Phylogenetic trees of ITS, 28S and 18S rDNA revealed that O. onirici sp. n. belongs to Dolichura-group. Oscheius onirici sp. n. is characterized by small body size and stoma rhabditoid type. Female reproductive system is amphidelphic. Males are rare with peloderan bursa, spicules slender and small, nine pairs of papillae of different lengths, arranged in a 1+1+1/3+3 pattern. Entomopathogenicity bioassay revealed that this nematode is capable of infecting larvae of Galleria mellonella and Tenebrio molitor.

Steinernema vulcanicum n. sp. (Rhabditida: Steinernematidae), a new entomopathogenic nematode species from Sicily (Italy)

A new species of entomopathogenic nematode, Steinernema vulcanicum n. sp., is described from the slopes of Mount Etna (Sicily, Italy). It was recovered from soil in a chestnut wood. Molecular and morphological data indicate that the new species belongs to the glaseri group of Steinernema spp. The new species can be distinguished from the other described species of the group by morphological characteristics, crossbreeding tests and by the DNA sequences of ITS1 rDNA, 12S and cox1 mitochondrial loci. Steinernema vulcanicum n. sp. is characterised by the long and slender male spicules, with manubrium wider than long, velum prominent, rounded tip with ventral concavity and tail without mucro. The female has double epiptygmata in both generations. The third-stage infective juveniles have an average body length of 1200 μm, excretory pore generally located level with posterior part of the pharyngeal corpus (D% = 47-68), lateral field pattern of variable formula and hyaline part of tail forming ca 50% of its length. The DNA sequences, life cycle and the endosymbiotic bacterium of S. vulcanicum n. sp., Xenorhabdus kozodoii, indicate the similarity of the new species with S. arenarium, S. apuliae and S. boemarei, from which it can be distinguished by both morphological and molecular characters.

TIPOLOGIE STRUTTURALI E CARATTERISTICHE FUNZIONALI DELLE AREE VERDI PERIURBANE: IL CASO DI STUDIO DELLA CITTÀ DI BARI

Si riportano i risultati di uno studio multidisciplinare finalizzato alla comprensione dei rapporti struttura-processo tra aree verdi periurbane e specie dell’entomofauna (coleotteri terricoli). Lo studio ha interessato una vasta superficie verde posta ai margini dell’agglomerato urbano di Bari denominata «Lama Balice» (Parco Naturale Regionale). Per tale area sono stati effettuati rilievi in campo al fine di determinare gli habitat vegetazionali prevalenti e le principali specie di coleotteri terricoli presenti in ciascun habitat. Complessivamente sono stati identificati 1779 coleotteri, di cui 1209 carabidi, 99 silfidi, 240 stafilinidi, 231 tenebrionidi. L’analisi statistica dei dati raccolti ha evidenziato una relazione significativa tra habitat e coleotteri carabidi e silfidi, con prevalenza dei coleotteri nell’habitat macchia-gariga. Di contro, nell’analisi non sono state riscontrate relazioni significative tra stafilinidi e tenebrionidi e i diversi habitat vegetazionali. Lo studio si conclude con alcune considerazioni circa il ruolo delle aree verdi in ambito urbano e periurbano in ragione della specifica tipologia (componente strutturale) e della presenza di specie vegetali ed animali (componente funzionale).

Survey of Steinernema and Heterorhabditis (Rhabditida: Nematoda) in Southern Italian soils

The occurrence of Steinernematid and Heterorhabditid entomopathogenic nematodes was pointed out in samples of soil collected from cultivated and uncultivated habitats in Southern Italy. Nematodes were isolated from samples of soil by “baiting” with larvae of Galleria mellonella L. (Lepidoptera: Galleriidae). Entomopathogenic nematodes were obtained from 26 (13.82%) of 188 soil samples collected: Steinernema was located in 16 sites (8.51%), Heterorhabditis in 10 sites (5.32%). The Steinernema strains were identified as S. feltiae (Filipjev, 1934) (50%), S. anomali (Kozodoi, 1984) (25%) and as S. affinis (Bovien, 1937) (25%). The Heterorhabditis strains were identified as H. bacteriophora Poinar, 1976. This is the first report of S. anomali and S. affinis in Italy; the presence of S. anomali in the soil of salt pan borders with 3,550 mg of Cl- /100 g of soil is extremely interesting. Key words: Entomopathogenic nematodes, occurrence, Steinernema feltiae, S. anomali, S. affinis, Heterorhabditis bacteriophora. INDAGINI SU STEINERNEMA E HETERORHABDITIS (RHABDITIDA: NEMATODA) NEI TERRENI DELL’ITALIA MERIDIONALE E stata effettuata una indagine per valutare la presenza e la distribuzione di nematodi entomopatogeni (Steinernema ed Heterorhabditis) in campioni di terreno provenienti da diversi habitat dell’Italia meridionale. I nematodi sono stati estratti dai campioni di terreno utilizzando larve di Galleria mellonella L. (Lepidoptera: Galleriidae), come insetto esca. La presenza di nematodi entomopatogeni e stata accertata in 26 campioni (13,82%) su 188 esaminati: gli Steinernema sono stati rinvenuti in 16 campioni (8,51%), Heterorhabditis in 10 (5,32%). Tra gli isolati di Steinernema sono state identificate 3 specie: S. feltiae Filipjev, 1934 (=S. bibionis) (Bovien, 1937) (50%), S. anomali (Kozodoi, 1984) (25%) e S. affinis (Bovien, 1937) (25%). I ceppi di Heterorhabditis sono risultati appartenere alla specie H. bacteriophora Poinar, 1976. La presenza di S. anomali e S. affinis viene segnalata per la prima volta in Italia. Di particolare interesse e il ritrovamento di un isolato di S. anomali in un terreno con Cl- /100 g di 3.550 mg. Parole chiave: nematodi entomopatogeni, distribuzione, Steinernema feltiae, S. anomali, S. affi- nis, Heterorhabditis bacteriophora.

Mosquitoes LTR retrotransposons: a deeper view into the genomic sequence of Culex quinquefasciatus.

A set of 67 novel LTR-retrotransposon has been identified by in silico analyses of the Culex quinquefasciatus genome using the LTR_STRUC program. The phylogenetic analysis shows that 29 novel and putatively functional LTR-retrotransposons detected belong to the Ty3/gypsy group. Our results demonstrate that, by considering only families containing potentially autonomous LTR-retrotransposons, they account for about 1% of the genome of C. quinquefasciatus. In previous studies it has been estimated that 29% of the genome of C. quinquefasciatus is occupied by mobile genetic elements. The potential role of retrotransposon insertions strictly associated with host genes is described and discussed along with the possible origin of a retrotransposon with peculiar Primer Binding Site region. Finally, we report the presence of a group of 38 retrotransposons, carrying tandem repeated sequences but lacking coding potential, and apparently lacking “master copy” elements from which they could have originated. The features of the repetitive sequences found in these non-autonomous LTR retrotransposons are described, and their possible role discussed. These results integrate the existing data on the genomics of an important virus-borne disease vector.