Antifungal activity of essential oils from leaves and flowers of Inula viscosa (Asteraceae) by Apulian region.

Some essential oils from several plants (Artemisia verlotorum, Lavandula augustifolia, Ocimum gratissimum) have proved to have acaricidal, antifungal and antibacterial activity. Inula viscosa Ait. (Asteraceae), a plant growing spontaneously in the Mediterranean area, is currently used by popular medicine for its therapeutic effects. Flavonoids, azulenes, sesquiterpenes, and essential oils have been isolated and identified from its leaves. This paper reports the results of the composition and antifungal activity in vitro against dermatophytes and Candida spp. of the four essential oils obtained by steam distillation of the leaves, flowers, whole plant and whole plants without flower extracts of I. viscosa. All the extracts proved to have a significant antifungal activity against dermatophytes even at low concentrations (0.01 mg/ml). The leaf extracts exhibited the greatest antifungal efficacy. The high concentration of the sesquiterpene (carboxyeudesmadiene), occurring in the leaf extracts, may explain its greater antifungal activity.

Differentiation among three species of bovine Thelazia (Nematoda: Thelaziidae) by polymerase chain reaction-restriction fragment length polymorphism of the first internal transcribed spacer ITS-1 (rDNA).

Thelazia gulosa, Thelazia rhodesi and Thelazia skrjabini are nematodes transmitted by some species of Musca (Diptera: Muscidae) which cause ocular infestations in bovines. Differences in the rDNA of these species were determined by a PCR using different sets of relatively conserved oligonucleotide primers. PCR on the first internal transcribed spacer (ITS-1) revealed differences in size in Thelazia species (437 bp for T. gulosa, 370 bp for T. rhodesi and 506 bp for T. skrjabini) while the DNA control of Musca spp. was not amplified. The ITS-1 amplicons of the three species were sequenced and then analysed. The GC contents ranged from 26 to 36% and the level of differences in the nucleotide sequences of ITS-1 was lower between T. skrjabini and T. gulosa (39%) than the latter and T. rhodesi (49-56%). Restriction fragment length polymorphism (RFLP) of ITS-1 amplicons was also carried out and the restriction profiles compared. Clear genetic differences among the three Thelazia examined were demonstrated by using the enzymes HpaII, CpoI and SspI. This PCR-RFLP for the delineation of T. gulosa, T. rhodesi and T. skrjabini offers prospects as a molecular epidemiological tool to study parasite transmission patterns and prevalence.

Screening of commercial milk samples using ELISA for immuno-epidemiological evidence of infection by the cattle grub (Diptera: Oestridae).

Pasteurised and ultra high temperature (UHT)-treated milk were tested over the period November 1997-February 1998 in order to evaluate the applicability of an hypodermosis ELISA test on commercial milk samples. Milk samples from six dairy companies were analysed and the development of anti-Hypoderma antibodies recorded for the period from September 1998 to June 1999. For one dairy (no. 3) bulk milk samples were also obtained from the tankers, transporting milk away from the farms, and analysed; the results were compared to the parasitological status of the cows whose milk had been collected on the farms. Out of 32 pasteurised samples tested, 23 were positive, exceeding the cut-off (22%), while 9 samples were negative. UHT milk was always negative. The antibody levels in milk samples from four of the six companies were highest during January and decreased gradually during February to May 1999. The bulk milk samples also had high antibody levels and 47.2% of milk producing cows had lesions of hypodermosis. It was concluded that testing commercial milk for antibodies is an efficient way of detecting the presence of hypodermosis in cattle, especially in those countries for which no data on this disease are available.

Comparative study of anti-Hypoderma antibody kinetics in sera, single and bulk milk samples of naturally infested cattle.

The investigation was carried out in Basilicata region (Southern Italy) from October 1997 to June 1998. Fifteen dairy cows bred in semiconfined conditions on a farm with a history of hypodermosis were sampled once a month for sera and milk; bulk milk from these animals was also collected monthly from the farm's tanker. Samples were tested for anti-Hypoderma spp. antibodies (Abs) with an ELISA technique and clinical parasitological examination was carried out monthly from January to July on all the animals in order to detect grubs. Blood and single and bulk milk samples yielded similar antibody kinetics and patterns in accordance with results obtained in previous immunological surveys in Italy. All animals were warbled in the spring time. November-January was confirmed to be the most suitable period for seroepidemiological survey for weather conditions in Southern Italy. The ELISA test proved once again to be very useful because it is simple to perform and cost effective. Either blood or milk samples may be used for epidemiological surveys; bulk milk may be very useful for the preliminary detection of hypodermosis on farms or in areas where there is no data available on the diffusion of the disease.

[Fungistatic activity of a sesquiterpene lactone (tomentosin) isolated from fresh Inula viscosa (Asteraceae) flowers from the Puglia region]. / Attività fungistatica di un sesquiterpene lattone (tomentosina) isolato dai fiori freschi di Inula viscosa (Asteraceae) della regione Puglia.

A sesquiterpene lactone, tomentosin, has been isolated and identified form Inula viscosá Ait (Asteraceae) flowers. The molecule, at 1 mg/ml concentration, showed to be active in vitro against Microsporum canis, Microsporum gypseum and Trichophyton mentagrophytes.

Goat warble fly infestation by Przhevalskiana silenus (Diptera: Oestridae): immunoepidemiologic survey in the Basilicata region (southern Italy).

The demonstration of serological cross-reactivity between the Hypoderma lineatum antigen and anti-Przhevalskiana silenus antibodies led us to prepare an immunological test (ELISA) for an early diagnosis of goat warble fly infestation. Using the Hypodermosis ELISA-Kit (Vétoquinol Diagnostic, France) produced for the immunodiagnosis of bovine hypodermosis, an epidemiological assay was carried out in Basilicata region where goat breeding is very common and no data are reported with regards to the distribution of goat warble fly infestation. Out of a total of 1,100 flocks and 41,200 goats, 105 randomly extracted flocks proved to be infected and 262 sera out of 1,316 were positive; goat warble fly infestation proved to be present in Basilicata with values similar to those recorded in the surrounding regions (Apulia and Calabria). Statistical evaluation showed highly significant differences between the number of infected flocks in the mountainous areas and hills and those of the mountainous areas and the plain, but no differences between hills and plains. The higher number of positive sera and antibody titres in November-December confirmed that these months are the optimal period for sampling sera in order to perform an early diagnosis. The ELISA test was confirmed to be an easy and economic tool especially when sera sampled within a brucellosis eradication program are used.

Differentiation by polymerase chain reaction - restriction fragment length polymorphism of some Oestridae larvae causing myiasis.

The cytochrome oxidase I (COI) gene of the most wide-spread Italian species of Oestridae larvae causing myiasis (Gasterophilus spp., Hypoderma bovis, Hypoderma lineatum, Oestrus ovis and Przhevalskiana silenus) was amplified by polymerase chain reaction (PCR) using conserved primers. Restriction fragment length polymorphism (RFLP) of amplicons was also carried out and their restriction profiles compared. A clear genetic difference between the Oestridae larvae examined was demonstrated by using Taq(alpha) I, Hinf I, Rsa I and Hpa II enzymes. No intra-specific variation in RFLPs was detected between the two species of Hypoderma. The results highlight the taxonomic and phylogenetic relationships among larvae belonging to the different subfamilies, and thus offer additional diagnostic and epidemiological instruments.

Further evidence on the internal life cycle of Przhevalskiana silenus (Diptera, Oestridae).

Knowledge of the internal life cycle of goat warble fly infestation is scarce despite ample data available on the aetiology, epidemiology, immunodiagnosis and treatment of such infestations. This study was carried out at the slaughterhouse of Rossano Calabro (Cosenza, southern Italy) on 154 animals from 10 months to 6 years of age from May 1997 to June 1998. 1206 Przhevalskiana silenus larvae were collected during the trial period from the subcutaneous tissue of the slaughtered animals. The larval stage average size ranged from 4.7 mm, for first instar larvae (May), to 16.6 mm, for third instar larvae (February), in the first cycle of infestation. No larvae were found in March-April, coinciding with the pupation period. Small first instar larvae were found at the beginning of the second cycle of infestation (May-June). Necroscopic examinations were also carried out on internal organs and no larvae were found. The results pointed out that the internal life cycle of P. silenus is exclusively subcutaneous and there is no internal migration of the larvae.

Thelazia gulosa Railliet & Henry, 1910 and T. skrjabini Erschow, 1928 infection in southern Europe (Italy).

The occurrence of Thelazia gulosa and T. skrjabini in cattle in Italy, together with an update of the prevalence of Thelazia spp. in the southern Italy, is reported. On 764 slaughtered native cattle, fifty-five (7.2%) bovines were infected by Thelazia spp: T. rhodesi was found in 44 (80%) animals, T. gulosa was found in 19 (34.5%) cattle and only one (1.8%) cattle harboured T. skrjabini. This is the first report of T. gulosa and T. skrjabini in Southern Europe.

Serodiagnosis of goat warble fly infestation by Przhevalskiana silenus with a commercial ELISA kit.

Cross-reactivity between Hypoderma lineatum antigen and anti-Przhevalskiana silenus antibodies has been demonstrated by an ELISA technique. To evaluate the applicability of a commercial ELISA kit for the immunodiagnosis of goat warble fly infestation, different dilutions of serum and conjugate were tested, the development of antibody to P silenus in naturally infested goats was studied, and the results were compared with an ELISA technique using an antigen extracted from the first instar larvae of H lineatum. The best results were obtained with a serum dilution of 1:50; with both techniques the highest antibody concentration were recorded in October, November and December. In view of the confirmed cross-reactivity between H lineatum antigen and anti-P silenus antibodies, and the simplicity and rapidity of the assay, the commercial ELISA kit can be considered as a useful tool for the diagnosis of goat warble fly infestation.

[Research on antifungal activity of flowers and leaves of Inula viscosa (Asteraceae)]. / Ricerche sull'attività fungistatica di fiori e foglie di Inula viscosa (Asteraceae).

The authors carried out a preliminary screening about the in vitro antifungal activity of some extracts of flowers and leaves of Inula viscosa obtained with different solvents. All extracts showed antifungal activity against dermatophytes and Candida species. The best results were obtained with Inula viscosa flowers extracts. These results may be ascribed to the different flavonoids and different flavonoid concentrations in our samples.

Antifungal activity of Apulia region propolis.

A study was carried out to assess the in vitro antifungal activity of some natural Apulian propolis extracts of different origin. Their antifungal activity was compared to the antifungal activity of conifers and commercial propolis extracts. All extracts revealed antifungal activity against dermatophytes and Candida species. The antifungal activity differences found depended on the origin of the propolis and the solvent used for extraction. The best antifungal activity was given by the 'Orimini' propolis. The antifungal activity may have been influenced by the presence of different cinnamic and flavonoid components and their different concentration in the extracts. Further investigations are needed to validate this hypothesis.

Hypoderma lineatum antigen and anti-Przhevalskiana silenus antibodies: cross-reactivity and antibody kinetics in naturally infested goats.

To evaluate the cross-reactivity between Hypoderma lineatum antigen and anti-Przhevalskiana silenus antibodies six protocols with different concentrations of antigen and different dilutions of sera and conjugate were applied. The highest cross-reaction between the H. lineatum antigen and the anti-P. silenus antibodies is given by 2 micrograms/ml of antigen concentration, 1:400 of serum and 1:10,000 conjugate dilution. The study on the kinetic development of antibodies in goats naturally infested by P. silenus and the natural course of infestation pointed out the existence of a good correlation between individual antibody kinetics and the natural evolution of the cycle of infestation. The highest antibody concentration may be registered from October through November, coinciding with the end of the migration fo the larvae inside the animal's body. In our condition, this period can be considered as a favorable sampling period for immunodiagnosis and immunoepidemiological studies of goat warble fly infestation.

Evaluation of polymorphonuclear cell and monocyte functions in Leishmania infantum-infected dogs.

Leishmania infantum is the causative agent of canine leishmaniasis in the Mediterranean area. The aspects of the canine immune response which may explain the chronic severe disease in these animals have not been well investigated. Therefore, in this work we have evaluated the parasite killing ability by circulating polymorphonuclear (PMN) cells and monocytes in dogs with spontaneous leishmaniasis and in healthy dogs. Moreover, in order to analyse the mechanisms responsible for killing activity, we evaluated the oxidative burst of circulating PMN cells, in terms of O2- and H2O2 production. The killing ability of infected dog PMN cells and monocytes was significantly lower than in a group of healthy dogs and a group of infected dogs treated with meglumine antimoniate and without symptoms of leishmaniasis. Superoxide anion and hydrogen peroxide production by PMN cells was significantly lower in a group of Leishmania-infected dogs than in healthy dogs, suggesting that these mediators may be involved in the reduced killing activity of infected dogs.

Leishmania donovani lipophosphoglycan (LPG) inhibits respiratory burst and chemotaxis of dog phagocytes.

Lipophosphoglycan (LPG) is the major glycoconjugate of Leishmania promastigote surface membrane. Previous studies on human and murine models have demonstrated that this molecule is involved in the attachment and survival of Leishmania in the host cells. Dog is the main reservoir of Leishmania strains responsible for human leishmaniasis in Italy. Since no studies have been performed on the LPG-canine phagocyte interactions, we investigated the LPG effects on dog phagocyte functions by evaluating: 1) the chemotactic activity of peripheral monocytes and polymorphonuclear (PMN) cells, in terms of cellular polarization; 2) the PMN cell respiratory burst, by measuring superoxide anion and hydrogen peroxide production. Results demonstrated a significant reduction of metabolic and chemotactic activity in LPG-preincubated cells, thus emphasizing the ability of this molecule to impair also the canine phagocyte responses.

Canine leishmaniasis in the Gargano promontory (Apulia, South Italy).

We examined 444 dogs by visiting farms and peripheral districts on the Gargano promontory (Province of Foggia, Apulia, South Italy). Clinical examination and immunofluorescence antibody test for leishmaniasis were performed. Bone marrow and lymph node samples obtained from 25 dogs with positive serological test were cultured on Tobie-Evans medium. The results obtained show both a higher seropositivity rate for canine leishmaniasis (14.4%) and a higher percentage of asymptomatic dogs with positive serological test (53.1%) compared to previous research in the same area. Seven strains isolated from infected dogs belonged to Leishmania infantum species zymodeme Montpellier 1. None of 82 sera of humans living in close contact with infected dogs were positive for leishmaniasis.

Infection by the warble fly Przhevalskiana silenus Brauer, 1858, in the Italian goats. An update up to 1988.

This paper is a review of the work done at the University of Bari on the goat warble fly Przhevalskiana silenus Brauer, 1858. It reports data concerning taxonomy, biology, and epidemiology of the goat warble fly, as well as therapy against the infection. Grunin in 1962, and later Madel in 1969, were the first researchers to reach the conclusion that the three species of Przhevalskiana up to then recognized--namely P. silenus, P. aegagri, and P. crossii--should be unified in just one--Przhevalskiana silenus. The genetical studies conducted by us on the third instar larvae with the use of 9 gene-enzyme systems confirmed the assumption that P. silenus is the only species which parasitizes goats. Studies were conducted on the external life-cycle of the parasite with the purpose of elucidating the relationship existing among time of dropping, length of time needed for adults to emerge from pupae, and temperature of the environment. Both total sum of temperature and length of time seem to play an important role in determining the duration of the pupal stage. Epidemiological data derived from a two-year survey carried out in six infected groups of goats are also reported. The interrelationship existing among prevalence, mean intensity, variance, variance/mean rate, and parameter K of the negative binomial are presented. It appears that age, parameter K, and variance of infection intensity are closely related. A trial experiment was also carried out to evaluate the efficacy of ivermectin against a natural infection with P. silenus larvae. A single subcutaneous injection of 1% ivermectin was given, at doses of 50, 100 or 200 micrograms/kg b.w. The drug was found to be fully effective at all three doses.

Acute and chronic hemodynamic effects of propranolol in unselected cirrhotic patients.

Different and contradictory results concerning the use of propranolol in the treatment of portal hypertension have been reported. This study was designed to investigate the hemodynamic effects of short- and long-term administration of propranolol in portal hypertensive patients. Portal pressure, cardiac index, heart rate and blood pressure were obtained in 18 unselected alcoholic cirrhotic patients with esophageal varices before and 60 min after the oral administration of 40 mg propranolol and again after 106 +/- 35 days of continuous oral administration (mean dose = 158 +/- 63 mg per day). Baseline portal pressure was 21.7 +/- 7.2 mm Hg. It decreased after 60 min to 17.2 +/- 5.5 mm Hg (p less than 0.01) and after long-term administration of propranolol to 16.1 +/- 5.7 mm Hg (p less than 0.01). No decrease in portal pressure was noted in 9 of 18 (50%) patients after acute administration and 5 of 17 (30%) patients after long-term administration. Baseline cardiac index was 5.1 +/- 1.2 liters X min-1 X m-2. It decreased after 60 min to 3.9 +/- 1.4 liters X min-1 X m-2 (p less than 0.01) and to 3.6 +/- 1.0 liters X min-1 X m-2 after long-term administration (p less than 0.001). Baseline heart rate was 85 +/- 11 beats per min. It decreased after 60 min to 75 +/- 9 (p less than 0.001) and after long-term administration to 62 +/- 6 (p less than 0.001) beats per min. Baseline mean arterial pressure was 108 +/- 11 Hg. It decreased after 60 min to 97 +/- 14 mm Hg (p less than 0.01) and after long-term administration to 103 +/- 14 mm Hg (not statistically significant).(ABSTRACT TRUNCATED AT 250 WORDS)