Giardia species and other intestinal parasites in dogs in north-east and central Italy.
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Age Factors↗
Biomedical subjects
A Giangaspero
Publications and source records attributed to A Giangaspero.
First description of the endogenous life cycle of Hypoderma sinense affecting yaks and cattle in China.
Larvae belonging to five species of Hypoderma spp. (Diptera, Oestridae) cause myiasis in wild and domestic ruminants that is characterized by migrations within deep tissues. In China hypodermosis is one of the most important arthropod diseases affecting ruminants and, moreover, represents a significant zoonosis, with numerous reports of Hypoderma spp. affecting farmers. Recently, a sixth species, Hypoderma sinense Pleske, has been rediscovered but the endogenous migration pathway within the host body is completely unknown and it represents a major constraint for the control of larval infection. In December 2003 a total of 165 larval stages of Hypoderma spp. were collected from different anatomical sites of 40 yaks slaughtered at an abattoir in the province of Gansu, China. The morphological characters and size of the recovered larvae were used to infer migratory routes and 45 specimens were also subjected to a polymerase chain reaction (PCR) assay of cox1 mtDNA and amplicons sequenced. All the larvae molecularly processed were identified as H. sinense and sequence identity was confirmed by a PCR-restriction fragment length polymorphism (PCR-RFLP) tool carried out using BfaI and HinfI endonucleases. The finding of H. sinense larvae only in the oesophagus or both in oesophagus and subcutaneous tissue of 12 and 15 animals, respectively, indicates that H. sinense larvae migrate through the oesophagus similarly to Hypoderma lineatum (De Villers). The description of the endogenous life cycle of H. sinense will help to determine the timing of specific treatment programmes to guarantee the improvement of animal welfare and health, thus resulting in an increase in livestock production in China.
Animals↗
Giardia, Cryptosporidium and the spectre of zoonosis: the Italian experience from land to sea.
In the last decade, a major concern for the scientific community has been whether infected animals can serve as reservoirs of Giardia and Cryptosporidium infection for humans. Worldwide, prevalence studies and molecular tools have provided insights into the taxonomy and epidemiology of these protozoa in order to better understand such a relation. This paper presents data on the prevalence and molecular genotyping studies from several sample types from land to sea (humans, companion animals, sheep, cattle, goats, wastewaters, surface water, and shellfish) available in Italy. The contribution of Italian researchers to the international debate on the veterinary significance of these infections and their impact on public health is highlighted and the main objectives to be pursued in the future depicted.
Adult↗
A new tool for the diagnosis in vivo of habronemosis in horses.
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Animals↗
Biology of Thelazia callipaeda (Spirurida, Thelaziidae) eyeworms in naturally infected definitive hosts.
Thelazia callipaeda (Spirurida, Thelaziidae) eyeworm causes ocular infection in carnivores and humans in the Far East; this infection has been recently reported also in Europe--northern and southern Italy--in dogs, cats and foxes. The natural vector/s of T. callipaeda is/are unknown and the development of the nematode in its definitive hosts is limited to an experimental trial on dogs. To contribute new insights into the development of T. callipaeda in the definitive host in field conditions, eyeworms were collected from naturally infected dogs from an area with a high prevalence of infection (up to 60.14%) in the Basilicata region of southern Italy, from January 2002 to December 2003. Conjunctival secretions were also collected and examined for the presence of immature stages. The presence of blastomerized eggs throughout the period--except for the months from May to November--indicates a seasonality in the reproductive activity of T. callipaeda, coinciding with the presence/absence of the vector. In fact, 1st-stage larvae were found in the lachrymal secretions of dogs in summer (June--July 2002 and 2003), ready to be ingested by flies feeding about the eyes. The evidence of 4th-stage larvae in March 2002 and April, July and October 2003 may be accounted for by the presence of flies that act as intermediate hosts of T. callipaeda from early spring to early autumn. The presence of immature stages in October indicates an overlapping generation of nematodes and a 2nd cycle of vector infection. This basic knowledge of the development of T. callipaeda will hopefully help future epidemiological studies to identify the intermediate hosts and define the likely risk for vectors in field conditions.
Animals↗
Semi-nested PCR for the specific detection of Habronema microstoma or Habronema muscae DNA in horse faeces.
Habronema microstoma and Habronema muscae (Spirurida: Habronematidae) are parasitic nematodes which infect the stomach and/or skin of equids. The accurate diagnosis of gastric habronemosis is central to studying its epidemiology, but data on its distribution and prevalence are lacking, mainly due to the limitations of clinical and coprological diagnosis in live horses. To overcome this constraint, a two-step, semi-nested PCR-based assay was validated (utilizing genetic markers in the nuclear ribosomal DNA) for the specific amplification of H. microstoma or H. muscae DNA from the faeces from horses (n = 46) whose gastrointestinal parasite status had been determined at autopsy and whose faeces were examined previously using a conventional parasitological approach. Of these horses examined at autopsy, some harboured adults of either H. microstoma (n= 19) or H. muscae (n =4), and others (n = 7) harboured both species. Most of them were also infected with other parasites, including strongylid nematodes (subfamilies Cyathostominae and Strongylinae), bots and/or cestodes; there was no evidence of metazoan parasites in 2 horses. Larvated spirurid eggs were detected in the faeces of 1 of the 30 horses (3.3 %) shown to be infected with Habronema at autopsy. For this set of 46 samples, the PCR assay achieved a diagnostic specificity of 100 % and a sensitivity of approximately 97 % (being able to specifically detect as little as approximately 0.02 fg of Habronema DNA). The specificity of the assay was also tested using a panel of control DNA samples representing horse, the gastric spirurid Draschia megastoma and 26 other species of parasites from the alimentary tract of the horse. H. microstoma, H. muscae and D. megastoma could be readily differentiated from one another based on the sizes of their specific amplicons in the PCR. The results of this study showed that the performance of the PCR for the diagnosis of gastric habronemosis was similar to that of autopsy but substantially better than the traditional coprological examination procedure used. The ability to specifically diagnose gastric habronemosis in equids should have important implications for investigating the epidemiology and ecology of H. microstoma and H. muscae.
Animals↗
A third species of Hypoderma (Diptera: Oestridae) affecting cattle and yaks in China: molecular and morphological evidence.
Cattle and yak hypodermosis in China is caused by Hypoderma bovis and H. lineatum, with a prevalence reaching up to 98-100% of the animals and maximum intensities exceeding 400 warbles for each animal. A third species, H. sinense, is also considered by Chinese researchers to affect livestock. The molecular characterization of the most variable region of the mitochondrial cytochrome oxidase I gene and of the ribosomal 28S gene has been performed for the third-stage larvae collected from cattle and yaks in China and identified (on the basis of the spinulation on the ventral side of the 10th segment) as H. bovis, H. lineatum, and H. sinense. Amplicons were digested with the HinfI and BfaI restriction enzymes, which provided diagnostic profiles to simultaneously differentiate the 3 Hypoderma species. Third-stage larvae of H. sinense were also examined by scanning electron microscopy, which revealed proper morphological characteristics different from those of H. bovis and H. lineatum. The molecular and morphological evidence herein reported support the existence of a third species of Hypoderma affecting cattle and yaks in China, and the results provide new tools for unequivocal identification of this species and present key components for the evaluation of its endogenous cycle and pathogenicity in animals and humans.
Animals↗
[Animal echinococcosis in Italy: epidemiological update].
Cystic Echinococcosis (CE) is one of the most widespread parasitoses in the Mediterranean Region (MR). This is due to various factors, the most important being the close association between man, sheep and dogs in areas where open farming is practised. Although this disease has been known for several years and many studies have been carried out, nowadays in Italy there are no complete epidemiological data on its diffusion and distribution. The available data show that CE is mainly diffused in those districts where the sheep-dog cycle can be perpetuated, such as central and southern Italy, and the islands. Furthermore, no data are available on biomolecular characterisation of the strains of Echinococcus granulosus in Italy, apart form those in Sardinia, where the G1 (sheep-dog) and G7 (pig-dog) strains were recently isolated. One of the reasons why CE is a problem with no easy solution is undoubtedly the difficulty of making a certain diagnosis in the dog, the principal definitive host of E. granulosus.
Animals↗
[Myiasis caused by Oestridae: serological and molecular diagnosis].
Myiasis-causing Oestridae (bot flies) infect several animal species world-wide, from palaearctic to subtropical/tropical areas. Oestrids affect livestock production causing abortion, reduced milk production, losses in weight and fertility, poor hide quality and an impairment of the host's immune system. In the last few years much research has been carried out on the immunology of these infestations, in order to acquire efficient and reliable diagnostic serological tools; the genome of the different species of Oestridae has been studied to further their molecular identification, taxonomy and phylogenesis. The immunodiagnostic methods for many myiasis causing Oestrids have proven to be a viable alternative to the clinical parasitological examination or the post-mortem examination. Numerous serological tests have been developed for the diagnosis of bovine hypodermosis caused by Hypoderma bovis and Hypoderma lineatum, and ELISAs using larval hypodermin C as the antigen are currently used on serum, individual and pooled milk samples to detect the presence of circulating anti-Hypoderma antibodies. In Italy the best period to sample the animals is November-January, since it is in this period that the antibody kinetics of the animals reaches a peak. Recently the efficacy of the ELISA test on pasteurized milk samples has been demonstrated, allowing the diagnosis of bovine hypodermosis also in areas where there is no information on the presence of the disease and the sampling of the animals is laborious. The cross-reactivity between Przhevalskiana silenus antigens and anti-Hypoderma antibodies led to assessing the usefulness of a simple and cost-effective ELISA for the diagnosis of goat warble fly infection. In particular, it has been demonstrated that infected goats display an antibody peak in November-December in blood and milk, thus making this period suitable for sampling. Although no extensive data is available on the immunology of sheep and goat oestrosis caused by Oestrus ovis, the efficacy of ELISA has been demonstrated by correlating serological results with clinical post-mortem examinations. No immunological techniques are currently used to diagnose gasterophilosis of equids and only one study reports the efficacy of ELISA for detecting anti-Gasterophilus antibodies in infected equids. Several studies have been conducted into the molecular characterization of the mitochondrial DNA (mtDNA)--in particular of the gene encoding for the cytochrome oxidase I (COI)--for many free-living and parasitic arthropods for diagnostic, taxonomic and phylogenetic purposes. As regards Oestridae causing myiasis, the first study features a PCR-RFLP assay of the most common Italian species (i.e. H. bovis, H. lineatum, Gasterophilus intestinalis, P. silenus, O. ovis), which showed clear genetic differences among the genera examined, but no inter-specific variation between the two species of Hypoderma considered. The molecular characterization of the most variable region of the COI gene (encoding for the region from E4 to the terminal COOH) was able to clearly differentiate H. bovis and H. lineatum. The E4-COOH region of the COI gene has been characterized for 18 oestrid species and from a taxonomical point of view, molecular data confirm the morphological classification, with the examined species divided into four subfamilies. New insights have also been gained on the molecular differentiation of the most common species of Hypoderma (i.e. H. bovis, H. lineatum, Hypoderma actaeon, Hypoderma diana and Hypoderma tarandi) and, in particular, the restriction enzyme BfaI, provides a diagnostic profile that can be used to simultaneously differentiate all the species examined. The characterization of the E4-COOH COI gene and the hypervariable region of the gene encoding for the ribosomal Isu revealed the identity of Hypoderma sinense as a new species, infecting cattle and yaks in China. Finally, the molecular analysis of the same mitochondrial and ribosomal regions showed that P. silenus, Przhevalskiana aegagri and Przhevalskiana crossii are morphotypes of the same species.
Animals↗
[Ecology of Thelazia spp. in cattle and their vectors in Italy].
The genus Thelazia (Spirurida, Thelaziidae) includes a cosmopolitan group of eyeworm spirurids responsible for ocular infections in domestic and wild animals and transmitted by different species of muscids. Bovine thelaziosis is caused by Thelazia rhodesi Desmarest 1828, Thelazia gulosa Railliet & Henry 1910, and Thelazia skrjabini Erschow 1928, which occur in many countries; T. gulosa and T. skrjabini have been reported mainly in the New World, while T. rhodesi is particularly common in the Old World. In Italy, T. rhodesi was reported in southern regions a long time ago and, recently, T. gulosa and T. skrjabini have been identified in autochthonous cattle first in Apulia and then in Sardinia. Thirteen species of Musca are listed as intermediate hosts of eyeworms, but only Musca autumnalis and Musca larvipara have been demonstrated to act as vectors of Thelazia in the ex-URSS, North America, ex-Czechoslovakia and more recently in Sweden. In Italy, after the reports of T. gulosa and T. skrjabini in southern regions, the intermediate hosts of bovine eyeworms were initially only suspected as the predominant secretophagous Muscidae collected from the periocular region of cattle with thelaziosis were the face flies, M. autumnalis and M. larvipara, followed by Musca osiris, Musca tempestiva and Musca domestica. The well-known constraints in the identification of immature eyeworms to species by fly dissection and also the time-consuming techniques used constitute important obstacles to epidemiological field studies (i.e. vector identification and/or role, prevalence and pattern of infection in flies, etc.). Molecular studies have recently permitted to further investigations into this area. A PCR-RFLP analysis of the ribosomal ITS-1 sequence was developed to differentiate the 3 species of Thelazia (i.e. T. gulosa, T. rhodesi and T. skrjabini) found in Italy, then a molecular epidemiological survey has recently been carried out in field conditions throughout five seasons of fly activity and has identified the role of M. autumnalis, M. larvipara, M. osiris and M. domestica as vectors of T. gulosa and of M. autumnalis and M. larvipara of T. rhodesi. Moreover, M. osiris was described, for the first time, to act as a vector of T. gulosa and M. larvipara of T. gulosa and T. rhodesi. The mean prevalence in the fly population examined was found to be 2.86%. The molecular techniques have opened new perspectives for further research on the ecology and epidemiology not only of Thelazia in cattle but also of other autochthonous species of Thelazia which have been also recorded in Italy, such as Thelazia callipaeda, which is responsible for human and canid ocular infection and Thelazia lacrymalis, the horse eyeworm whose epidemiological molecular studies are in progress.
Animals↗
Prevalence of Giardia spp. in dogs and humans in northern and central Italy.
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Animals↗
Molecular epidemiological survey on the vectors of Thelazia gulosa, Thelazia rhodesi and Thelazia skrjabini (Spirurida: Thelaziidae).
A Polymerase Chain Reaction (PCR)- based assay developed for the specific identification of Thelazia gulosa, Thelazia rhodesi and Thelazia skrjabini (Nematoda, Spirurida), which cause bovine ocular thelaziosis, was evaluated for its usefulness in detecting the intermediate hosts and in estimating the infection prevalence of vectors in field conditions throughout 5 years (from 1997 to 2001). A total of 5190 flies were captured and identified as Musca larvipara, Musca osiris, Musca autumnalis, Musca tempestiva or Musca domestica. Genomic DNA was extracted from pools constituted by heads, thoraces, abdomens and wings of 10 flies of each species, and 2076 samples were subjected to a PCR assay to specifically detect the ribosomal ITS-1 sequence of bovine Thelazia. Amplicons were sequenced and subjected to digestion with CpoI restriction enzyme. M. autumnalis, M. larvipara, M. osiris and M. domestica species were shown to be PCR positive. T. gulosa was specifically detected by PCR in M. autumnalis, M. larvipara, M. osiris and M. domestica, whereas T. rhodesi is in M. autumnalis and M. larvipara. Of 27 positive samples, 23 were positive for T. gulosa and 4 for T. rhodesi, with a mean prevalence of 2.86% in the whole fly population collected. The highest mean prevalence values of infection were detected in M. autumnalis (4.46%) and M. larvipara (3.21%), and the former species was confirmed to be the vector of T. gulosa and T. rhodesi. This study is the first report of M. osiris as a vector of T. gulosa and M. larvipara as a vector of T. gulosa and T. rhodesi under natural conditions. The occurrence of Thelazia in fly populations in the Apulia region of Italy (in the 5 grazing seasons considered) indicates that cattle thelaziosis is enzootic in southern Italy. This molecular assay should be a useful epidemiological tool for assessing the role of different species of flies as intermediate hosts of thelaziae.
Animals↗
Efficacy of pyrantel embonate, febantel and praziquantel against Giardia species in naturally infected adult dogs.
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Animals↗
A novel [60]fullerene amino acid for use in solid-phase peptide synthesis.
[see structure]. A fullerene derivative containing a free amino group has been condensed with N-Fmoc-L-glutamic acid alpha-tert-butyl ester to give a C60-functionalized amino acid. The carboxylic end of this amino acid has been deprotected in acidic conditions, and the resulting acid has been used for solid-phase peptide synthesis. The final peptide, cleaved from the resin, was very soluble in water solutions and showed antimicrobial activity against two representative bacteria.
Amino Acids↗
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.
Amino Acid Sequence↗
Efficacy of 5 MU of interferon in combination with ribavirin for naïve patients with chronic hepatitis C virus: a randomized controlled trial.
BACKGROUND: In chronic hepatitis C the schedule of interferon (IFN), 3 MU thrice weekly (tiw) plus ribavirin (1000-1200 mg/daily) needs further evaluation, as IFN dosages >3 MU achieve better responses. AIMS: To compare the efficacy of 5 MU tiw of IFN with (96 patients) or without ribavirin (96 patients) for 12 months in naïve patients, to evaluate the effect of baseline features on the response to therapy, and to determine a reliable point in time during treatment to predict non-response. RESULTS: Sustained virologic response was 20.8% (95% CI 13-29) with IFN monotherapy and 54.2% (95% CI 44-64) with combination (P = 0.0001), the relapse rate 39.4% (95% CI 23-56) and 9% (95% CI 1-16) (P = 0.0007), and the combined rate of sustained biochemical and virologic response 22.7% (95% CI 14-31) and 60.5% (95% CI 50-71) (P = 0.0001), respectively. Patients given combination therapy were more likely to respond regardless of baseline features. Apart from genotype non-1, predictive factors for IFN monotherapy were ineffective in predicting response to combination therapy. Using logistic regression analysis, IFN-ribavirin was the strongest predictor of response (X2 = 21.3; P = 0.0001). Viral persistence at month 3 of therapy was a more accurate predictor than aminotransferase values for non-response to IFN monotherapy but not to combination therapy (positive predictive values of 98 and 82%, respectively). CONCLUSION: In this study, 5 MU of IFN combined with a standard dose of ribavirin has yielded the highest rate of sustained response reported to date. Further dose finding studies are warranted.
Adult↗
Amphipathic alpha helical antimicrobial peptides.
Antimicrobial peptides (AMPs) that assume an amphipathic alpha helical structure are widespread in nature. Their activity depends on several parameters including the sequence, size, degree of structure formation, cationicity, hydrophobicity and amphipathicity. The analysis of numerous natural AMPs provided representative values for these parameters and led to a sequence template with which to generate potent artificial lead AMPs. Sequences were then varied in a rational manner, using both natural and nonproteinogenic amino acids, to probe the individual roles of each parameter in modulating biological activity. A high cationicity combined with a stabilized amphipathic alpha helical structure conferred enhanced cidal activity towards all the cell types considered, and was a requirement for Gram-positive bacteria and fungi. An elevated helicity also correlated with increased hemolytic activity. The structural requirements for activity against several Gram-negative bacteria were instead considerably less stringent, so that it persisted in peptides in which formation of a helical structure and/or amphipathicity were impeded. Either a reduced charge or a reduced hydrophobicity resulted in generally inactive peptides. These observations, combined with the kinetics of bacterial membrane permeabilization and time-killing are discussed in terms of currently accepted models of action for this type of peptide. The simple guidelines obtained in this study allowed the design of highly active shortened AMPs and may be generally useful in the development of this type of peptides as anti-infective agents.
Amino Acid Sequence↗
A randomized trial of amantadine and interferon versus interferon alone as initial treatment for chronic hepatitis C.
The aim of this study was to compare, in an open-label study, the efficacy and safety of a combination of interferon (IFN) and amantadine (AMA) with that of IFN alone in previously untreated patients with chronic hepatitis C. A total of 200 patients were randomized to 6 MU of IFN-alpha2a 3 times per week, with 200 mg of AMA daily (n = 99) or to an identical dose of interferon alpha2a (n = 101). Patients were treated for 12 months and observed for 6 months' posttreatment. At the completion of treatment, 28.7% of patients in the monotherapy group and 45.5% in the combination group had a virologic response (P =.014). At 6 months' posttreatment, a sustained virologic response was observed in 16.8% (95% CI: 9-23) of patients with IFN alone versus 29.3% (95% CI: 19-37) of patients who were treated with combination therapy (P =.036). In each of the 2 treatments, genotype was the only predictive parameter for a sustained response. At the logistic regression analysis, therapy and genotype were the only 2 parameters with an independent predictive value. In the combination group, at examination of month 3, hepatitis C virus (HCV)-RNA status had a 97.6% (95% CI: 93-102) positive predictive value and a 50% (95% CI: 37-63) negative predictive value for a sustained virologic clearance. A substantial proportion of naïve patients with chronic hepatitis C have an end-of-treatment and end-of-follow-up virologic and biochemical response to a combination of IFN and AMA. This new treatment appears safe and well tolerated.
Adult↗