Species Tropism of Chimeric SHIV Clones Containing HIV-1 Subtype-A and Subtype-E Envelope Genes

AbstractTo analyze HIV-1 genes in a nonhuman primate model for lentivirus infection and AIDS, recombinant SIV/HIV-1 (SHIV) clones were constructed from two HIV-1 subtype-A isolates (HIV-1SF170 and HIV-1Q23–17 from individuals in Africa) and two HIV-1 subtype-E isolates (HIV-19466 and HIV-1CAR402 from AIDS patients in Thailand and Africa), respectively. These four SHIV clones, designated SHIV-A-170, SHIV-A-Q23, SHIV-9466.33, and SHIV-E-CAR, contain envelope (env) genes from the subtype-A or -E viruses. Interestingly, SHIV-A-170, SHIV-A-Q23, and SHIV-9466.33 were restricted for replication in cultures of macaque lymphoid cells, whereas SHIV-E-CAR replicated efficiently in these cells. Additional studies to define the block to replication in macaque cells were focused on the subtype-E clone SHIV-9466.33. A SHIV intragenic env clone, containing sequence-encompassing V1/V2 regions of HIV-1CAR402 and V3/V4/V5 regions of SHIV-9466.33, infected and replicated in macaque lymphoid cells. These results indicated that the sequence-encompassing V1/V2 region of HIV-19466 was responsible for the block of the SHIV-9466.33 replication in macaque cells. Analysis of viral DNA in acutely infected macaque cells revealed that SHIV-9466.33 was blocked at a step at/or before viral DNA synthesis, presumably during the process of virion entry into cells. In a fluorescence-based cell–cell fusion assay, fusion pore formation readily took place in cocultures of cells expressing the SHIV-9466.33 env glycoprotein with macaque T-lymphoid cells. Taken together, these results demonstrated that the block of SHIV-9466.33 replication in macaque cells is at an early step after fusion pore formation but before reverse transcription

Fatal Immunopathogenesis by SIV/HIV-1 (SHIV) Containing a Variant Form of the HIV-1sf33 env Gene in Juvenile and Newborn Rhesus Macaques

AbstractSIV/HIV-1 (SHIV) chimeric clones, constructed by substituting portions of the pathogenic molecular clone SIVmac239 with counterpart portions from HIV-1 clones, provide a means to analyze functions of selected HIV-1 genes in vivo in nonhuman primates. Our studies focused on SHIVsf33, which contains the vpu, tat, rev, and env genes of the cytopathic, T-cell line tropic clone HIV-1sf33 (subtype-B); this clone has a premature stop codon in the vpu gene. In three juvenile macaques inoculated intravenously with SHIVsf33, low-level persistent infection was established; no disease was observed for a period of >2 years. However, at ∼16 months p.i., one of four SHIVsf33-infected juvenile macaques exhibited an increase in virus load, depletion of CD4+ T cells in peripheral blood and lymph nodes, and other symptoms of simian AIDS (SAIDS). Virus recovered from this animal in the symptomatic stage was designated SHIVsf33a (A, adapted); this virus displayed multiple amino acid sequence changes throughout the HIV-1 env gene compared with the input SHIVsf33 clone. Additionally, a mutation in all clones from SHIVsf33a restored the open reading frame for the vpu gene. In vitro evaluations in tissue-culture systems revealed that SHIVsf33a replicated to higher levels and exhibited greater cytopathicity than SHIVsf33. Furthermore cloned env genes for SHIVsf33a were more fusogenic in a cell-fusion assay compared with the env gene of the SHIVsf33. Intravenous inoculation of SHIVsf33a into juvenile and newborn macaques resulted in a rapid decline in CD4+ T cells to very low levels and development of a fatal AIDS-like disease. A cell-free preparation of this pathogenic chimeric virus also established persistent infection when applied to oral mucosal membranes of juvenile macaques and produced a fatal AIDS-like disease. These studies on pathogenic SHIVsf33a establish the basis for further investigations on the role of the HIV-1 env gene in virus adaptation and in mechanism(s) of immunodeficiency in primates; moreover, the chimeric virus SHIVsf33a can play a role in elucidating mucosal membrane transmission and development of antiviral vaccines in newborns as well as juvenile and adult macaques

Escherichia coli O157:H7 in Feral Swine near Spinach Fields and Cattle, Central California Coast1

We investigated involvement of feral swine in contamination of agricultural fields and surface waterways with Escherichia coli O157:H7 after a nationwide outbreak traced to bagged spinach from California. Isolates from feral swine, cattle, surface water, sediment, and soil at 1 ranch were matched to the outbreak strain

Re-shaping models of E.coli population dynamics in livestock faeces: Increased bacterial risk to humans?

Dung-pats excreted directly on pasture from grazing animals can contribute a significant burden of faecal microbes to agricultural land. The aim of this study was to use a combined field and modelling approach to determine the importance of Escherichia coli growth in dung-pats when predicting faecal bacteria accumulation on grazed grassland. To do this an empirical model was developed to predict the dynamics of an E. coli reservoir within 1 ha plots each grazed by four beef steers for six months. Published first-order die-off coefficients were used within the model to describe the expected decline of E. coli in dung-pats. Modelled estimates using first-order kinetics led to an underestimation of the observed E. coli land reservoir, when using site-specific die-off coefficients. A simultaneous experiment determined the die-off profiles of E. coli within fresh faeces of beef cattle under field relevant conditions and suggested that faecal bacteria may experience growth and re-growth in the period post defecation when exposed to a complex interaction of environmental drivers such as variable temperature, UV radiation and moisture levels. This growth phase in dung-pats is not accounted for in models based on first-order die-off coefficients. When the model was amended to incorporate the growth of E. coli, equivalent to that observed in the field study, the prediction of the E. coli reservoir was improved with respect to the observed data and produced a previously unquantified step-change improvement in model predictions of the accumulation of these faecal bacteria on grasslands. Results from this study suggest that the use of first-order kinetic equations for determining land-based reservoirs of faecal bacteria should be approached with caution and greater emphasis placed on accounting for actual survival patterns observed under field relevant conditions

Different subcellular localisations of TRIM22 suggest species-specific function

The B30.2/SPRY domain is present in many proteins, including various members of the tripartite motif (TRIM) protein family such as TRIM5α, which mediates innate intracellular resistance to retroviruses in several primate species. This resistance is dependent on the integrity of the B30.2 domain that evolves rapidly in primates and exhibits species-specific anti-viral activity. TRIM22 is another positively selected TRIM gene. Particularly, the B30.2 domain shows rapid evolution in the primate lineage and recently published data indicate an anti-viral function of TRIM22. We show here that human and rhesus TRIM22 localise to different subcellular compartments and that this difference can be assigned to the positively selected B30.2 domain. Moreover, we could demonstrate that amino acid changes in two variable loops (VL1 and VL3) are responsible for the different subcellular localisations

Survival of Escherichia coli in the environment: fundamental and public health aspects

In this review, our current understanding of the species Escherichia coli and its persistence in the open environment is examined. E. coli consists of six different subgroups, which are separable by genomic analyses. Strains within each subgroup occupy various ecological niches, and can be broadly characterized by either commensalistic or different pathogenic behaviour. In relevant cases, genomic islands can be pinpointed that underpin the behaviour. Thus, genomic islands of, on the one hand, broad environmental significance, and, on the other hand, virulence, are highlighted in the context of E. coli survival in its niches. A focus is further placed on experimental studies on the survival of the different types of E. coli in soil, manure and water. Overall, the data suggest that E. coli can persist, for varying periods of time, in such terrestrial and aquatic habitats. In particular, the considerable persistence of the pathogenic E. coli O157:H7 is of importance, as its acid tolerance may be expected to confer a fitness asset in the more acidic environments. In this context, the extent to which E. coli interacts with its human/animal host and the organism's survivability in natural environments are compared. In addition, the effect of the diversity and community structure of the indigenous microbiota on the fate of invading E. coli populations in the open environment is discussed. Such a relationship is of importance to our knowledge of both public and environmental health. The ISME Journal (2011) 5, 173-183; doi:10.1038/ismej.2010.80; published online 24 June 2010NATO [ESP.EAP.CLG 981785]; The Soil Biotechnology Foundationinfo:eu-repo/semantics/publishedVersio

SUMO-Interacting Motifs of Human TRIM5α are Important for Antiviral Activity

Human TRIM5α potently restricts particular strains of murine leukemia viruses (the so-called N-tropic strains) but not others (the B- or NB-tropic strains) during early stages of infection. We show that overexpression of SUMO-1 in human 293T cells, but not in mouse MDTF cells, profoundly blocks N-MLV infection. This block is dependent on the tropism of the incoming virus, as neither B-, NB-, nor the mutant R110E of N-MLV CA (a B-tropic switch) are affected by SUMO-1 overexpression. The block occurred prior to reverse transcription and could be abrogated by large amounts of restricted virus. Knockdown of TRIM5α in 293T SUMO-1-overexpressing cells resulted in ablation of the SUMO-1 antiviral effects, and this loss of restriction could be restored by expression of a human TRIM5α shRNA-resistant plasmid. Amino acid sequence analysis of human TRIM5α revealed a consensus SUMO conjugation site at the N-terminus and three putative SUMO interacting motifs (SIMs) in the B30.2 domain. Mutations of the TRIM5α consensus SUMO conjugation site did not affect the antiviral activity of TRIM5α in any of the cell types tested. Mutation of the SIM consensus sequences, however, abolished TRIM5α antiviral activity against N-MLV. Mutation of lysines at a potential site of SUMOylation in the CA region of the Gag gene reduced the SUMO-1 block and the TRIM5α restriction of N-MLV. Our data suggest a novel aspect of TRIM5α-mediated restriction, in which the presence of intact SIMs in TRIM5α, and also the SUMO conjugation of CA, are required for restriction. We propose that at least a portion of the antiviral activity of TRIM5α is mediated through the binding of its SIMs to SUMO-conjugated CA