Factors affecting the accuracy of pregnancy scanning in ewes.

The objective was to evaluate the accuracy of pregnancy scanning by transabdominal ultrasonography and affecting factors. Altogether 44,783 ewes were registered (2008-2010), 39,724 diagnosed as pregnant and subsequently lambed. The ewes, 0.5-14 years old, were divided into 4 age and 6 breed groups and analysed. The accuracy (per cent; number of scanned fetuses/number of born lambs) decreased with increasing number of fetuses (P < 0.001). Overall accuracy was 90.3 per cent, highest (93.7 per cent) in ewes carrying one fetus, 91.9 per cent, 82.4 per cent, and 74.9 per cent in ewes with 2, 3 and ≥4 fetuses at scanning, respectively. Finnsheep ewes with highest number of lambs showed lowest accuracy (P < 0.001). Analyses of a more complete sub set of data (n = 23,396), showed that number of fetuses diagnosed, breed, age of ewe, operator and time in gestation, significantly affected the accuracy. Accuracy decreased with age of ewe (P < 0.001) and was 71.8 per cent, 91.6 per cent and 89.3 per cent for scanning at <40, 40-80 and 81-100 days of gestation, respectively (P < 0.001). In general, the numbers of fetuses were overestimated at scanning and increased gradually with number of fetuses diagnosed. In conclusion, the accuracy was affected by several factors, which should be considered when interpreting/implementing the results, especially in breeds with high fecundity.

Predominance of HIV-1 subtype A and D infections in Uganda.

To better characterize the virus isolates associated with the HIV-1 epidemic in Uganda, 100 specimens from HIV-1-infected persons were randomly selected from each of two periods from late 1994 to late 1997. The 200 specimens were classified into HIV-1 subtypes by sequence- based phylogenetic analysis of the envelope (env) gp41 region; 98 (49%) were classified as env subtype A, 96 (48%) as D, 5 (2.5%) as C, and 1 was not classified as a known env subtype. Demographic characteristics of persons infected with the two principal HIV-1 subtypes, A and D, were very similar, and the proportion of either subtype did not differ significantly between early and later periods. Our systematic characterization of the HIV-1 epidemic in Uganda over an almost 3-year period documented that the distribution and degree of genetic diversity of the HIV subtypes A and D are very similar and did not change appreciably over that time.

Genetic characterization and phylogenetic analysis of HIV-1 subtype C from Uganda.

To better understand the emergence of subtype C and its potential impact on vaccine efforts in Uganda, we have characterized subtype C sequences from Uganda (n = 13), Zimbabwe (n = 11), Mozambique (n = 5), South Africa (n = 4), and India (n = 3). Phylogenetic analysis of subtype C sequences in the env gp41 gene region revealed multiple subclusters within subtype C. Further, while most Ugandan specimen subclustered together, other subclusters did not reflect a clear geographic location. The nucleotide divergence within the Ugandan subset was 8.2% (6.1-9.8%) compared with 9.5% (2.5-15%) for the other subtype C gp41 sequences. The protein sequence alignment revealed marked sequence conservation of major immunodominant epitopes within the gp41 region.

Identification of a window period for susceptibility to dual infection with two distinct human immunodeficiency virus type 2 isolates in a Macaca nemestrina (pig-tailed macaque) model.

The potential to establish dual retroviral infections was investigated in this study. Groups of macaques infected with human immunodeficiency virus type 2 (HIV-2) isolate (either GB122 or CDC77618) were exposed to the other virus at 2, 4, 8, 12, 14, or 72 weeks after primary inoculation. Dual infections were established in macaques simultaneously exposed to both viruses. In other groups, secondary infections were observed only if challenge occurred at early intervals after primary infection but before a full seroconversion. Polymerase chain reaction and virus-isolation data demonstrated that challenges at 8, 12, 14, or 72 weeks after infection with the initial isolate failed to result in a dual infection. Anti-HIV-2 serologic titers, CD4 levels, virus burden, and the ability to superinfect peripheral blood mononuclear cells in vitro were not correlated with susceptibility to or protection from secondary challenges in this investigation. These findings demonstrate a window period for susceptibility to dual infection and indicate that protection from retroviral infection may be achievable.

Detection of phylogenetically diverse human immunodeficiency virus type 1 groups M and O from plasma by using highly sensitive and specific generic primers.

The high degree of genetic diversity within human immunodeficiency virus type 1 (HIV-1), which includes two major groups, M (major) and O (outlier), and various env subtypes within group M (subtypes A to J), has made designing assays that will detect all known HIV-1 strains difficult. We have developed a generic primer set based on the conserved immunodominant region of transmembrane protein gp41 that can reliably amplify as few as 10 copies/PCR of viral DNA from near-full-length clones representing group M subtypes A to H (subtypes I and J were not available). The assay is highly sensitive in detecting plasma viral RNA from HIV-1 strains of diverse geographic origins representing different subtypes of HIV-1 group M as well as HIV-1 group O. Of the 253 group M plasma specimens (subtypes A, 68 specimens; B, 71; C, 19; D, 27; E, 23; F, 33; and G, 12), 250 (98.8%) were amplified by using the gp41 M/O primer set. More importantly, all 32 (100%) group O plasma samples were also amplified with these primers. In vitro spiking experiments further revealed that the assay could reliably detect as few as 25 copies/ml of viral RNA and gave positive signals in HIV-1-seropositive specimens with plasma copy numbers below the limits of detection by all commercially available viral load assays. In addition, analysis of five seroconversion panels indicated that the assay is highly sensitive for early detection of plasma viremia during the "window period." Thus, the highly sensitive assay will be useful for early detection of HIV-1 in clinical specimens from all known HIV-1 infections, regardless of their genotypes and geographic origins.

Genetic analysis of human immunodeficiency virus in Abidjan, Ivory Coast reveals predominance of HIV type 1 subtype A and introduction of subtype G.

To better understand the molecular epidemiology of HIV genetic diversity in Abidjan, Ivory Coast, we performed a genetic analysis of 170 HIV-1-seropositive specimens representing newly diagnosed tuberculosis patients (n = 143) and women monitored in a mother-to-child transmission cohort study (n = 27). Preliminary screening with RFLP presumptively classified 162 (95.3%) of these as subtype A. The envelope region of 108 specimens was subtyped by sequence analysis: 102 (94.4%) were subtype A, 2 (1.9%) were subtype D, and 4 (3.7%) were subtype G. Subtyping gag and env regions of the genome suggested that five of the six nonsubtype A isolates exhibited a potentially mosaic structure. A comparative phylogenetic analysis of HIV-1 subtype A C2V3 from 27 Ivory Coast and 21 Ugandan sequences revealed a striking clustering among Ivory Coast variants, and an independent segregation from Ugandan subtype A. Despite independent clustering with other subtype A specimens, limited variability of the V3 loop apex was observed; the globally predominant V3 motif, GPGQ, represented 90.1% of the HIV-1 strains. This study demonstrates that clade A is the predominant HIV-1 subtype in HIV-seropositive individuals in Abidjan, Ivory Coast and that these strains are phylogenetically distinct from other subtype A strains observed in East Africa.

Introduction of HIV-2 and multiple HIV-1 subtypes to Lebanon.

HIV genetic variability, phylogenetic relationships, and transmission dynamics were analyzed in 26 HIV-infected patients from Lebanon. Twenty-five specimens were identified as HIV-1 and one as HIV-2 subtype B. The 25 strains were classified into six env-C2-V3 HIV-1 subtypes: B (n = 10), A (n = 11), C (n = 1), D (n = 1), G (n = 1), and unclassifiable. Potential recombinants combining parts of viral regions from different subtypes Aenv/Dpol/Agag, Genv/Apol, and the unclassifiable-subtype(env)/unclassifiable-subtype(pol)/Agag were found in three patients. Epidemiologic analysis of travel histories and behavioral risks indicated that HIV-1 and HIV-2 subtypes reflected HIV strains prevalent in countries visited by patients or their sex partners. Spread of complex HIV-subtype distribution patterns to regions where HIV is not endemic may be more common than previously thought. Blood screening for both HIV-1 and HIV-2 in Lebanon is recommended to protect the blood supply. HIV subtype data provide information for vaccine development.

Optimizing the delivery of HIV counseling and testing services: the Uganda experience using rapid HIV antibody test algorithms.

The AIDS Information Center (AIC) was established in Kampala, Uganda in 1990 in response to increasing interest by members of the general public who wished to know their HIV serostatus. By 1996, >300,000 clients had been seen. HIV serologic testing was performed at a central laboratory and results reported back to AIC after 2 weeks. Approximately 25% of clients failed to learn their HIV serostatus as a result of failure to return or late arrival of results. To address these issues, AIC carried out an evaluation of 3 rapid HIV assays, Sero-Strip, SeroCard, and Capillus, against a standard criterion to identify a testing algorithm that could be used as an on-site confirmatory testing strategy. The study was carried out over a period of 5 working days and 325 clients were seen. An algorithm was identified, which gave no indeterminate results with unambiguously positive or negative specimens, which was 100% sensitive and specific, and which could be integrated with minimal disruption into existing counseling procedures. All clients left AIC knowing their HIV serostatus and having spent <2 hours at the Center. The results of this evaluation demonstrate that "same-day" results can be provided in counseling and testing settings without compromising the quality of counseling or the accuracy of HIV testing.

PIP: An evaluation conducted at the AIDS Information Center in Kampala, Uganda, demonstrated that same-day HIV results can be provided in counseling and testing centers without compromising service quality. The Center was established in 1990 in response to widespread public interest in HIV serodiagnosis. By 1996, more than 300,000 clients had been tested. However, 25% of these clients never received their result because of failure to report back to the Center after 2 weeks (the time required for results to be returned from an off-site laboratory) or late arrival of results. To address this problem, the Center evaluated three rapid HIV assays (Sero-Strip, SeroCard, and Capillus) against a standard criterion to identify a testing algorithm that could be used as an on-site confirmatory testing strategy. 325 clients were enrolled in the 5-day evaluation. Individually, all three rapid tests performed well when compared with the standard criterion result. The resulting algorithm (a combination of Capillus as the screening assay and SeroCard as a supplementary assay for initially positive specimens and Multispot as a tie-breaker assay) gave no indeterminate results, was 100% sensitive and specific, and could be integrated easily into existing counseling protocols. The entire process (registration, test decision counseling, phlebotomy, laboratory testing, prevention counseling, and post-test counseling) took an average of 2 hours to complete.

Detection of genetically diverse human immunodeficiency virus type 1 group M and O isolates by PCR.

A panel of 136 genetically diverse group M and 5 group O adult isolates from outside the United States and Europe were evaluated by PCR with the Roche AMPLICOR HIV-1 test, a modified version of the AMPLICOR HIV-1 test, and a new primer pair/probe system. Detection of some of these isolates was less efficient with the AMPLICOR HIV-1 test; however, the assay was significantly improved by reducing the sample input and lowering the annealing temperature. The new primer pair/probe set detected 140 of 141 isolates, including the 5 group O isolates that were not detected with either of the AMPLICOR HIV-1 test formats.