Intestinal parasites in immigrants in the city of Naples (southern Italy).

The present study was aimed at carrying out a cross-sectional copromicroscopic survey of helminths and intestinal protozoa in immigrants in Naples (southern Italy). Between October 2008 and November 2009, a total of 514 immigrants were tested comparing the FLOTAC dual technique and the ethyl acetate concentration technique. Combined results of the two techniques served as a diagnostic 'gold' standard and revealed an overall prevalence of parasitic infections of 61.9% (318/514). The ethyl acetate concentration technique detected a low number of positive results (49.0%) and this was confirmed for each helminth/protozoa species detected. Among helminths, Trichuris trichiura (3.9%), hookworms (3.7%) and Ascaris lumbricoides (1.4%) were the most prevalent. Strongyloides stercoralis (0.4%), Enterobius vermicularis (0.4%), Schistosoma mansoni (1.0%), Hymenolepis nana (1.6%) and Taenia spp. (0.2%) were also found, as well as zoonotic helminths, as Trichostrongylus spp. (0.8%) and Dicrocoelium dendriticum (0.8%). As regard to pathogenic protozoa, Blastocystis hominis was the most commonly detected (52.7%), followed by Entamoeba histolytica/Entamoeba dispar/Entamoeba moshkovskii (11.9%) and Giardia duodenalis (4.5%). Several issues concerning diagnosis, epidemiology and public health impact of parasitic infections in immigrants are offered for discussion. In conclusion, the present paper pointed out the need of better diagnosis and cure of the immigrant population in order to improve access to health care of this neglected and marginalised population group, for its own protection and care.

Processing of DNA base damage by DNA polymerases. Dihydrothymine and beta-ureidoisobutyric acid as models for instructive and noninstructive lesions.

The processing of unrepaired DNA lesions is a key to understanding and predicting the biological end points of particular DNA damages. In this study, we prepared single-stranded f1 phage (f1-K12) DNA containing dihydrothymine or beta-ureidoisobutyric acid as models for instructive or noninstructive base lesions and assessed the potential biological consequences of these lesions in vitro and in vivo. To determine the effect of the two lesions on in vitro DNA synthesis, the extent of DNA synthesis was measured by 3H-labeled nucleotide incorporation, and the newly synthesized DNA was analyzed by DNA sequencing gels. The results showed that dihydrothymine in the template was at most a weak block to in vitro DNA synthesis catalyzed by Escherichia coli DNA polymerase I Klenow fragment (Pol I) and T4 DNA polymerase. In contrast, beta-ureidoisobutyric acid constituted a very strong (probably absolute) replicative block in vitro. With Pol I, termination bands were observed either opposite or one base prior to (3' to) the putative beta-ureidoisobutyric acid depending on its position in the template. However, when DNA synthesis was catalyzed by Pol I lacking a 3'----5' exonuclease activity, termination bands were only observed opposite beta-ureidoisobutyric acid, with purine nucleotides being incorporated preferentially opposite the lesion. With T4 DNA polymerase that contains a very active 3'----5' exonuclease activity, DNA synthesis was arrested almost exclusively one base prior to (3' to) the putative beta-ureidoisobutyric acid site in the template. We also measured survival of transfecting DNA containing dihydrothymine or beta-ureidoisobutyric acid in an attempt to correlate the in vitro data with in vivo processing. In keeping with the results obtained in vitro, dihydrothymine present in transfecting f1-K12 DNA did not constitute an inactivating lesion. On the other hand, it took 0.9 beta-ureidoisobutyric acid residues per molecule to inactivate transfecting f1-K12 DNA, indicating that the lesion was an absolute replicative block in vivo. When host cells were ultraviolet-irradiated to induce the SOS response, a slight increase (about 2-fold) in survival of transfecting f1-K12 DNA containing beta-ureidoisobutyric acid was observed. The potential effects of the structures of base lesions on lesion-polymerase interactions are discussed.

Oxidative damage in DNA. Lack of mutagenicity by thymine glycol lesions.

Thymine glycol (5,6-dihydroxy-5,6-dihydrothymine) is a base damage common to oxidative mutagens and the major stable radiolysis product of thymine in DNA. We assessed the mutagenic potential of thymine glycols in single-stranded bacteriophage DNA during transfection of Escherichia coli wild-type and umuC strains. cis-Thymine glycols were induced in DNA by reaction with the chemical oxidant, osmium tetroxide (OsO4); modification of thymines was quantitated by using anti-thymine glycol antibody. Inactivation of transfecting molecules showed that one lethal hit corresponded to 1.5 to 2.1 thymine glycols per phage DNA in normal cells, whereas conditions of W-reactivation (SOS induction) reversed 60 to 80% of inactivating events. Forward mutations in the lacI and lacZ' (alpha) genes of f1 and M13 hybrid phage DNAs were induced in OsO4-treated DNA in a dose-dependent manner, in both wild-type and umuC cells. Sequence analysis of hybrid phage mutants revealed that mutations occurred preferentially at cytosine sites rather than thymine sites, indicating that thymine glycols were not the principal pre-mutagenic lesions in the single-stranded DNA. A mutagenic specificity for C----T transitions was confirmed by OsO4-induced reversion of mutant lac phage. Pathways for mutagenesis at derivatives of oxidized cytosine are discussed.

Effect of a 2-methylthio-N6-isopentenyladenosine deficiency on peptidyl-tRNA release in Escherichia coli.

We examined the effect of miaA, a mutation conferring a deficiency in 2-methylthio-N6-isopentenyladenosine in tRNA, on patterns of peptidyl-tRNA accumulation in Escherichia coli strains deficient in peptidyl-tRNA hydrolase activity. A specific reduction in peptidyl-tRNA accumulation was seen for tRNAs which normally contain the 2-methylthio-N6-isopentenyladenosine modification. These results provide new evidence in support of the ribosome editor model, which links peptidyl-tRNA release to mistranslation events.

Novel E. coli mutants deficient in biosynthesis of 5-methylaminomethyl-2-thiouridine.

Novel E. coli mutants deficient in biosynthesis of 5- methylaminomethyl -2-thiouridine were isolated based on a phenotype of reduced readthrough at UAG codons. They define 2 new loci trmE and trmF , near 83' on the E. coli map. These mutants are different from strains carrying trmC mutations, which are known to confer a methylation deficiency in biosynthesis of 5- methylaminomethyl -2-thiouridine. tRNA from mutants carrying trmE or trmF mutations was shown to carry 2-thiouridine instead of 5- methylaminomethyl -2-thiouridine. This deficiency affects the triplet binding properties of the mutant tRNA. Our results suggest that the 5- methylaminomethyl group stabilizes the basepairing of this modified nucleotide with G, most likely through direct interaction with the ribosomal binding site(s).

The role of 2-methylthio-N6-isopentenyladenosine in readthrough and suppression of nonsense codons in Escherichia coli.

Readthrough and suppression of nonsense codons was compared in Escherichia coli strains with and without a miaA mutation, which confers a loss of the isopentenyladenosine modification in transfer RNA. Generally speaking, our results conform to predictions based on previous literature. In addition, we showed that the miaA mutation in strain TRPX is itself a UAA mutation. An antagonism between miaA and rpsL mutations, which confer streptomycin resistance, was also discovered. Our data further suggest that slight alterations of the translation apparatus are easily detectable by monitoring readthrough and suppression of nonsense codons. Our findings are discussed in the context of old and recent reports.

Frameshift mutations: relative roles of simple intercalation and of adduct formation.

The contribution of "simple" intercalation and of adduct formation on the expression of frameshift mutations in Salmonella typhimurium was investigated using 9-aminoacridine and derivatives capable of either only intercalating between DNA base pairs or of forming adducts with DNA as well. For a chemical capable of intercalating as well as of forming an adduct, only a small portion of the frameshift mutagenicity is due to "simple" intercalation. Analogs only able to induce frameshift mutations as a result of intercalation generally display only a fraction (approx. 1%) of the frameshift activity of the analog capable of forming DNA adducts.

4-Nitroquinoline-1-oxide: factors determining its mutagenicity in bacteria.

In bacteria, 4-nitroquinoline-1-oxide (NQO) causes primarily mutations of the base-substitution type although frameshift mutations are also induced. The adducts formed are presumably recognized by error-prone DNA repair enzymes as evidenced by the much greater activity in plasmid pKM101-bearing tester strains. Although reduction of the nitro group appears to be required for mutagenic activity, this reduction is not catalyzed by the nitroreductase required for the demonstration of the mutagenicity in bacteria of other nitro-containing mutagens (nitrofurans, 2-nitronaphthalene, nitrofluorenes). The reduction of the nitro group appears to be catalyzed by a different nitroreductase. The mutagenicity of the non-carcinogenic 3-methyl-4-nitroquinoline-1-oxide (meNQO) may be related to this newly recognized nitroreductase. It is proposed, further, that the ultimate mutagenic intermediates derived from NQO and MeNQO differ.

Structural basis of the mutagenicity in bacteria of nitrated naphthalene and derivatives.

While 2-nitronaphthalene was a weak direct-acting base-substitution mutagen (1.4 revertants/nanomole) for Salmonella typhimurium, the analogous nitronaphthalic acid anhydride and imides were moderate frameshift mutagens (approximately 20 rev/nanomole in strain TA98). Although imide derivatives are efficient DNA intercalators, mutagenicity data indicate that the bulk of the frameshift activity is derived from adduct formation between hydroxylamine intermediates and DNA. The low level of frameshift activity (approximately 8% of total) resulting from simple intercalation (measured in strain TA 1537) is not dependent upon reduction of the nitro function. Evidence is presented that suggests that the reduction of the nitro function to the corresponding hydroxylamine might not involve a free nitroso intermediate. The introduction of a second nitrofunction into nitronaphthalenes results in great positional effects of the various isomers on mutagenic activity and specificity.

Non-mutagenic genotoxicants: novobiocin and nalidixic acid, 2 inhibitors of DNA gyrase.

The antimicrobial agents nalidixic acid and novobiocin, both inhibitors of DNA gyrase, preferentially inhibit the growth of DNA repair-deficient Escherichia coli, Salmonella typhimurium and Bacillus subtilis. On the other hand, these agents exhibit no demonstrable mutagenicity for S. typhimurium and E. coli even when tested under conditions designed to take the metabolic properties of these agents into consideration. Novobiocin and nalidixic acid are widely used in animal and human medicine and further studies to assess their genetic and carcinogenic potentials appear to be in order.