The expression and activation of Toll-like receptors (TLRs) was investigated in leprosy, a spectral disease in which clinical manifestations correlate with the type of immune response mounted toward Mycobacterium leprae. TLR2-TLR1 heterodimers mediated cell activation by killed M. leprae, indicating the presence of triacylated lipoproteins. A genome-wide scan of M. leprae detected 31 putative lipoproteins. Synthetic lipopeptides representing the 19-kD and 33-kD lipoproteins activated both monocytes and dendritic cells. Activation was enhanced by type-1 cytokines and inhibited by type-2 cytokines. In addition, interferon (IFN)-gamma and granulocyte-macrophage colony-stimulating factor (GM-CSF) enhanced TLR1 expression in monocytes and dendritic cells, respectively, whereas IL-4 downregulated TLR2 expression. TLR2 and TLR1 were more strongly expressed in lesions from the localized tuberculoid form (T-lep) as compared with the disseminated lepromatous form (L-lep) of the disease. These data provide evidence that regulated expression and activation of TLRs at the site of disease contribute to the host defense against microbial pathogens.

Leprosy is a chronic and debilitating human disease caused by infection with the Mycobacterium leprae bacillus. Despite the marked reduction in the number of registered worldwide leprosy cases as a result of the widespread use of multidrug therapy, the number of new cases detected each year remains relatively stable. This indicates that M. leprae is still being transmitted and that, without earlier diagnosis, M. leprae infection will continue to pose a health problem. Current diagnostic techniques, based on the appearance of clinical symptoms or of immunoglobulin M (IgM) antibodies that recognize the bacterial phenolic glycolipid I, are unable to reliably identify early-stage leprosy. In this study we examine the ability of IgG within leprosy patient sera to bind several M. leprae protein antigens. As expected, multibacillary leprosy patients provided stronger responses than paucibacillary leprosy patients. We demonstrate that the geographic locations of the patients can influence the antigens they recognize but that ML0405 and ML2331 are recognized by sera from diverse regions (the Philippines, coastal and central Brazil, and Japan). A fusion construct of these two proteins (designated leprosy IDRI diagnostic 1 [LID-1]) retained the diagnostic activity of the component antigens. Upon testing against a panel of prospective sera from individuals who developed leprosy, we determined that LID-1 was capable of diagnosing leprosy 6 to 8 months before the onset of clinical symptoms. A serological diagnostic test capable of identifying and allowing treatment of early-stage leprosy could reduce transmission, prevent functional disabilities and stigmatizing deformities, and facilitate leprosy eradication.Cases in which Mycobacterium leprae infection manifests to cause leprosy present as a bacteriologic, clinical, immunologic, and pathological spectrum ranging from the extremes observed in paucibacillary (PB) and multibacillary (MB) patients (21,24). PB patients have one or a few skin lesions and a low or absent bacterial index (BI; a measure of the number of acidfast bacilli in the dermis, expressed on a logarithmic scale) and demonstrate specific cell-mediated immunity against M. leprae, but they have low or absent titers of M. leprae-specific antibodies and a granulomatous dermatopathology. In marked contrast, MB patients have multiple symmetric skin lesions and a high BI and demonstrate high titers of anti-M. leprae antibodies but an absence of specific cell-mediated immunity and a dermatopathology largely devoid of functional lymphocytes (21). Despite the implementation of a WHO-directed eradication program over the last 20 years, the worldwide annual rate of new case detection for leprosy remains stable at approxi-

Human immunodeficiency virus type 1 integrase (HIV-1 IN) is thought to have several putative roles at steps prior to integration, such as reverse transcription and nuclear transport of the preintegration complex (PIC).Here, we investigated new functional aspects of HIV-1 IN in the context of the viral replication cycle through point mutagenesis of Ser, Thr, Tyr, Lys, and Arg residues conserved in IN, some of which are located at possible phosphorylation sites. Our results showed that mutations of these Ser or Thr residues had no effect on reverse transcription and nuclear transport of PIC but had a slight effect on integration. Of note, mutations in the conserved KRK motif (amino acids 186 to 189), proposed previously as a putative nuclear localization signal Retroviruses establish a proviral state in which a doublestranded DNA copy of the viral genomic RNA is integrated into the host genome in a stable manner, through several steps following binding and entry into the target cell. These early events include uncoating, reverse transcription, nuclear transport of the viral genome, and integration. The viral enzyme integrase (IN) is encoded by the pol gene and the attachment (att) site located at the U3 and U5 termini of the viral DNA and is required for integration, which is the last event (6,14,43,47,53,56,57,61,66,74). The detailed mechanism of retroviral integration has been elucidated from in vitro studies using recombinant IN protein and a synthetic DNA substrate mimicking the viral att sites. These studies, using in vitro assays, have contributed much information toward the currently accepted mechanism of retroviral integration (reviewed in references 34, 42, and 75). Mutational and structural studies of human immunodeficiency virus type 1 (HIV-1) IN have identified three functional domains: a central catalytic core domain, an N-terminal zinc binding domain, and a C-terminal nonspecific DNA binding domain. The core domain contains the highly conserved D,D35E motif, which is directly involved in the catalytic activities of IN (7,23,46,48). The N-terminal domain contains a highly conserved HHCC motif, which binds to zinc. Through a tetrahedral attachment to the HHCC motif, zinc enhances both multimerization and enzymatic activities of 8,21,79). The C terminus, consisting of a structure that closely resembles Src homology 3 domains, possesses sequence-and metal ion-independent DNA binding activity (20,51). Each domain has been demonstrated to form a dimer or higher multimerization state of IN (8,19,20), which might be required for its full activity (13,21,22,66,70,73).Genetic analysis of HIV-1 IN has demonstrated multiple effects of mutations at steps distinct from integration. These steps include correct viral particle formation (24, 59), uncoating (54, 59), and reverse transcription (49,54). During the early events of the infection cycle, prior to integration, around 50-100 protomers of IN exist as one of the major components of the preintegration complex (PIC). This is composed of the viral genome, the matr...

Sequences of the folP1, rpoB, and gyrA genes were analyzed for 88 isolates of Mycobacterium leprae from leprosy patients in Japan, Haiti, Indonesia, Pakistan, and the Philippines. Thirteen isolates (14.8%) showed representative mutations in more than two genes, suggesting the emergence of multidrug-resistant M. leprae.

This multicenter prospective study showed favorable short-term outcomes for gastric ESD.

Investigations were done to determine whether the replication kinetics of human immunodeficiency virus (HIV)-1 were altered when the virus acquired a set or subsets of five mutations (A62V, V75I, F77L, F116Y, and Q151M) in the pol gene conferring resistance to multiple dideoxynucleosides. In the absence of drugs, the replication rate of all infectious clones generated was comparable to that of wild type HIV-1. However, in the presence of zidovudine or didanosine, the comparative order for replication was HIV-1(62/75/77/116/151) > HIV-1(77/116/151) > HIV-1(75/77/116/151) approximately HIV-1(151), whereas that for drug resistance was HIV-1(75/77/116/151) > HIV-1(62/75/77/116/151) > or = HIV-1(77/116/151) > HIV-1(151). The virologic features of these infectious mutants suggest that HIV-1 develops drug resistance through one or more mutations, which, however, sacrifice replicative capability; then it finally acquires optimal replication competence by additional mutations when the multi-dideoxynucleoside-resistant mutant emerges.

Host defense against Mycobacterium leprae infection is chiefly mediated by gamma interferon (IFN-␥؉ -and CD8 ؉ -T-cell subsets. M. leprae is a unique pathogen which remains resistant to DC-mediated T-cell immunity, at least in the early stages of infection.

Although the role of the APOBEC3-dependent retroelement restriction system as an intrinsic immune defense against human immunodeficiency virus type1 (HIV-1) infection is becoming clear, only the rat ortholog of mammalian APOBEC1s (A1) thus far has been shown to possess antiviral activity. Here, we cloned A1 cDNAs from small animal species, and showed that similar to rat A1, both wild-type and Δvif HIV-1 infection was inhibited by mouse and hamster A1 (4- to 10-fold), whereas human A1 had negligible effects. Moreover, rabbit A1 significantly reduced the infectivity of both HIV-1 virions (>300-fold), as well as that of SIVmac, SIVagm, FIV and murine leukemia virus. Immunoblot analysis showed that A1s were efficiently incorporated into the HIV-1 virion, and their packaging is mediated through an interaction with the nucleocapsid Gag domain. Interestingly, there was a clear accumulation of particular C-T changes in the genomic RNAs of HIV-1 produced in their presence, with few G-A changes in the proviral DNA. Together, these data reveal that A1 may function as a defense mechanism, regulating retroelements in a wide range of mammalian species.