Claudio Casoli

Antiretroviral Activity of Thiosemicarbazone Metal Complexes

Thiosemicarbazones display a wide antimicrobial activity by targeting bacteria, fungi, and viruses. Here, we report our studies on the antiviral activity of two thiosemicarbazone metal complexes, [bis(citronellalthiosemicarbazonato)nickel(II)] and [aqua(pyridoxalthiosemicarbazonato)copper(II)] chloride monohydrate, against the retroviruses HIV-1 and HTLV-1/-2. Both compounds exhibit antiviral properties against HIV but not against HTLVs . In particular, the copper complex shows the most potent anti-HIV activity by acting at the post-entry steps of the viral cycle.

Evolutionary rate and genetic heterogeneity of human T-cell lymphotropic virus type II (HTLV-II) using isolates from European injecting drug users

Abstract. Seven new Italian and two new British HTLV-II isolates were obtained from injecting drug users and the entire long terminal repeat (LTR) region was sequenced. Restriction analysis showed that all the Italian isolates are of the IIb subtype, whereas the British isolates are of the IIa subtype. To understand whether the further differentiation of each two principal HTLV-II subtypes in several subgroups could be statistically supported by phylogenetic analysis, the neighbor-joining, parsimony, and maximum likelihood methods were used. The separation between IIa and IIb is very well supported by all three methods. At least two phylogenetic subgroups exist within the HTLV-IIa and at least three within the HTLV-IIb subtype. In the present analysis, no statistical support was obtained for additional phylogroups. Two particular subgroups seem interesting because they include all European and North American injecting drug user strains within the IIa and IIb subtypes, respectively. These data confirm that European HTLV-II infection among drug users is probably derived from North America. They also suggest that though a certain differentiation by restriction analysis in different subgroups is possible, carefully interpreted phylogenetic analyses remain necessary. Using the likelihood ratio test, a molecular clock for the drug user strains was calibrated. A fixation rate between 1.08 × 10−4 and 2.7 × 10−5 nucleotide substitutions per site per year was calculated for the IIa and IIb injecting drug user strains. This is the lowest fixation rate so far reported for RNA viruses, including for HIV, which typically range between 10−2 and 10−4.

Coinfection with HIV-1 and human T-Cell lymphotropic virus type II in intravenous drug users is associated with delayed progression to AIDS

Human T-cell lymphotropic virus (HTLV) type II has spread among intravenous drug users (IDUs), many of whom are coinfected with HIV-1. We have investigated the rate of HTLV-II infection in 3574 Italian IDUs screened for HIV-1, HTLV-I, and HTLV-II from 1986 to the present. HTLV-II proviral load was determined by a real-time polymerase chain reaction specifically designed for tax amplification. The frequency of HTLV-II infection was 6.7% among HIV-1-positive subjects and 1.1% among HIV-1-negative subjects (P < 0.0001). For examination of AIDS progression, a group of 437 HIV-1-monoinfected subjects and another group of 96 HIV-1/HTLV-II-coinfected subjects were monitored. Enrollees were matched at entry by CD4 cell counts and followed for an average of 13 years. HIV-1/HTLV-II coinfection was associated with older age (P < 0.0001) and higher CD4 (P < 0.0001) and CD8 (P < 0.001) cell counts compared with monoinfected IDUs. The number of long-term nonprogressors for AIDS was significantly higher (P < 0.0001) among coinfected patients (13 [13.5%] of 96 patients) than HIV monoinfected patients (5 [1.1%] of 437 patients), showing that HTLV-II exerts a protective role. An increased incidence of liver disease and hepatitis C virus positivity among coinfected IDUs was observed. Five coinfected subjects undergoing antiretroviral therapy showed a significant (P < 0.05) increase in HTLV-II proviral load concomitant to a decrease in HIV-1 viremia, suggesting that the treatment is ineffective against HTLV-II infection.

Molecular characterization of two isolates of human T cell leukaemia virus type II from Italian drug abusers and comparison of genome structure with other isolates

The human T cell leukaemia virus type II (HTLV-II), whose pathogenicity is as yet unclear, was recently found to be associated with intravenous drug abuse in North America and Europe. HTLV-II was isolated from two Italian drug abusers belonging to the same cohort and coinfected with human immunodeficiency virus type 1. Two new isolates, HTLV-II Gu and Va, were established in a culture of BJAB cells, a continuous B cell line (Epstein-Barr virus-negative), and characterized by nucleotide sequence analysis of the long terminal repeat (LTR) and portions of the gag, env and X regions. These sequences were compared to those of the HTLV-II Mo isolate reported in the literature. No major variations were observed in important regulatory elements of LTR nor in the stem-bulge-loop configuration known to be essential for binding of rex protein. The results obtained from the sequence of the 1988 nucleotides examined indicated a 1.6% variability between the Gu and Va isolates and about 6% with respect to Mo. Notable differences were found in the structure of putative open reading frames of the X region when compared to those reported for the Mo isolate. Restriction analysis of proviral DNA of two isolates and comparison with the physical map of the Mo isolate confirmed the existence of genetic heterogeneity in the HTLV-II group and demonstrated that the new isolates Gu and Va belong to the HTLV-IIb subtype. The results of this study show that the new isolates have distinct features with respect to the Mo isolate though all important regulatory elements of the LTR appear to be well conserved.

Complete nucleotide sequence of the Italian human T-cell lymphotropic virus type II isolate Gu and phylogenetic identification of a possible origin of South European epidemics

The complete nucleotide sequence of a human T-cell lymphotropic virus type II isolate (HTLV-II-Gu) from an Italian injecting drug user was obtained, representing the first entire sequence of a European HTLV-II isolate. The HTLV-II-Gu genome was more similar to the HTLV-IIb-NRA isolate (98.4%) and HTLV-IIb-G12 (98.2%) than to HTLV-IIa-Mo (95.2%). The classification of HTLV-II-Gu as subtype IIb was confirmed by restriction analysis. Just as for HTLV-IIa strain Mo, HTLV-IIb-Gu cultured lymphocytes produce two additional mRNAs generated through alternative splicing in the pX region. A phylogenetic analysis was performed by using the methods of neighbour-joining and parsimony with bootstrapping, and maximum likelihood. The different gene regions were analysed separately, comparing Gu with all other HTLV-II strains presently available. In the LTR, as well as in other genome regions, a clear separation between IIa and IIb was evident, and within the IIb subtype three clusters were present of which two were well supported; one contained exclusively Amerindian strains and the other included all Italian and Spanish strains together with two strains obtained from New York drug users. All data clearly showed that HTLV-IIa and IIb subtypes are closely related and are equidistant from HTLV-I, suggesting that both groups evolved simultaneously. The results suggest that HTLV-II-Gu and other IIb South European isolates were probably derived from North American IIb isolates. The data also indicate that sequence analysis is necessary to further classify IIa and IIb subtypes.

Vaccination of Lactating Dairy Cows for the Prevention of Aflatoxin B1 Carry Over in Milk

The potential of anaflatoxin B1 (AnAFB1) conjugated to keyhole limpet hemocyanin (KLH) as a vaccine (AnAFB1-KLH) in controlling the carry over of the aflatoxin B1 (AFB1) metabolite aflatoxin M1 (AFM1) in cow milk is reported. AFB1 is the most carcinogenic compound in food and foodstuffs amongst aflatoxins (AFs). AnAFB1 is AFB1 chemically modified as AFB1-1(O-carboxymethyl) oxime. In comparison to AFB1, AnAFB1 has proven to be non-toxic in vitro to human hepatocarcinoma cells and non mutagenic to Salmonella typhimurium strains. AnAFB1-KLH was used for immunization of cows proving to induce a long lasting titer of anti-AFB1 IgG antibodies (Abs) which were cross reactive with AFB1, AFG1, and AFG2. The elicited anti-AFB1 Abs were able to hinder the secretion of AFM1 into the milk of cows continuously fed with AFB1. Vaccination of lactating animals with conjugated AnAFB1 may represent a solution to the public hazard constituted by milk and cheese contaminated with AFs.

The origin and evolution of human T-cell lymphotropic virus type II (HTLV-II ) and the relationship with its replication strategy

In this review, the origin and evolution of the human T-cell lymphotropic virus type II (HTLV-II) are discussed, with particular emphasis on its high genomic stability. In particular, it appears that the virus originated in the African continent and has been infecting human populations for several thousands of years. The very low divergence accumulated on average between different viral strains during such a long period could be explained by considering that in infected individuals the viral amplification could be due mainly to the clonal expansion of the infected cells, via cellular mitosis, rather than to reverse transcription. HTLV-II was introduced into the American continent during one or more migrations of HTLV-II-infected Asian populations over the Bering land bridge, some 15,000-35,000 years ago. Finally, during the last few decades, HTLV-II has been transmitted from native Amerindians to injecting drug users (IDUs). It might be speculated that at least two separate introductions of HTLV-II in European IDUs from US IDUs have occurred, due to the practice of needle-sharing among IDUs.

The MHC class II transactivator: prey and hunter in infectious diseases

The MHC class II transcriptional activator (CIITA) is the major regulator of expression of MHC class II genes. Thus, CIITA plays a fundamental role in the regulation of the immune response. Here, we discuss our findings on the dual role of CIITA during infections, as the target (prey) for certain pathogens but the host effector (hunter) against other pathogens, including HIV-1. This dual role is placed in an evolutionary context as a rather peculiar example of a strategy used by pathogens to evade host defenses and a counteraction of the host to minimize the survival and spread of the pathogen.

Rifampicin-resistant RNA polymerase and NAD transferase activities in coliphage N4 virions

SEVERAL animal viruses carry within the infectious particle a number of enzymatic activities1,2. Prominent among these are the RNA synthetases of orthomixo, paramixo, rhabdo-and diplomaviruses3–6, the DNA-dependent RNA polymerase of poxviruses7 and the reverse transcriptases of oncornaviruses8,9, all of which play important roles in the virus life cycle. Exclusion of vital enzymes from tine virion of all complex bacterial viruses seems, on the other hand, to have been the result of the evolutionary trend followed by these agents. A notable exception to this rule seems to be represented by coliphage N4 (ref. 10). It has been reported that transcription of N4 early genes11,12 and incorporation of 32P-orthophosphate into several host proteins13 occur in Escherichia coli cells treated before infection with rifaimpicin and chloramphenicol. A possible explanation for these findings was that N4 injects into the cell the relevant enzymes along with its DNA. We report here the detection of a rifampicin-resistant RNA polymerase and a nicotinamide adenine dinucleotide (NAD) transferase in purified N4 particles.

Highlights on distinctive structural and functional properties of HTLV Tax proteins

Human T cell leukemia viruses (HTLVs) are complex human retroviruses of the Deltaretrovirus genus. Four types have been identified thus far, with HTLV-1 and HTLV-2 much more prevalent than HTLV-3 or HTLV-4. HTLV-1 and HTLV-2 possess strictly related genomic structures, but differ significantly in pathogenicity, as HTLV-1 is the causative agent of adult T cell leukemia and of HTLV-associated myelopathy/tropical spastic paraparesis, whereas HTLV-2 is not associated with neoplasia. HTLVs code for a protein named Tax that is responsible for enhancing viral expression and drives cell transformation. Much effort has been invested to dissect the impact of Tax on signal transduction pathways and to identify functional differences between the HTLV Tax proteins that may explain the distinct oncogenic potential of HTLV-1 and HTLV-2. This review summarizes our current knowledge of Tax-1 and Tax-2 with emphasis on their structure, role in activation of the NF-κB (nuclear factor kappa-B) pathway, and interactions with host factors.