J. Neil Simonsen

Female to male transmission of human immunodeficiency virus type 1: risk factors for seroconversion in men

To determine the frequency and risk factors for female to male sexual transmission of human immunodeficiency virus type 1 (HIV-1), a prospective study was carried out in 422 men who had acquired a sexually transmitted disease (STD) from a group of prostitutes with a prevalence of HIV-1 infection of 85%. The initial seroprevalence of HIV among the men was 12%. 24 of 293 (8.2%) initially seronegative men seroconverted to HIV-1. Newly acquired infection was independently associated with frequent prostitute contact (risk ratio 3.2, 95% confidence interval 1.2-8.1), with the acquisition of genital ulcer disease (risk ratio 4.7, 95% confidence interval 1.3-17.0), and with being uncircumcised (risk ratio 8.2, 95% confidence interval 3.0-23.0). 96% of documented seroconversions occurred in men with one or both of the latter two risk factors. In a subgroup of 73 seronegative men who reported a single prostitute sexual contact, the frequency of HIV-1 infection was 8.2% during 12 weeks of observation. No man without a genital ulcer seroconverted. A cumulative 43% of uncircumcised men who acquired an ulcer seroconverted to HIV-1 after a single sexual exposure. These data indicate an extremely high rate of female to male transmission of HIV-1 in the presence of STD and confirm a causal relation between lack of male circumcision, genital ulcer disease, and susceptibility to HIV-1 infection.

Mass Spectrometric Characterization of Proteins from the SARS Virus A Preliminary Report

A new coronavirus has been implicated as the causative agent of severe acute respiratory syndrome (SARS). We have used convalescent sera from several SARS patients to detect proteins in the culture supernatants from cells exposed to lavage another SARS patient. The most prominent protein in the supernatant was identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as a ∼46-kDa species. This was found to be a novel nucleocapsid protein that matched almost exactly one predicted by an open reading frame in the recently published nucleotide sequence of the same virus isolate (>96% coverage). A second viral protein corresponding to the predicted ∼139-kDa spike glycoprotein has also been examined by MALDI-TOF MS (42% coverage). After peptide N-glycosidase F digestion, 12 glycosylation sites in this protein were confirmed. The sugars attached to four of the sites were also identified. These results suggest that the nucleocapsid protein is a major immunogen that may be useful for early diagnostics, and that the spike glycoprotein may present a particularly attractive target for prophylactic intervention in combating SARS.

HIV-1-specific cellular immune responses among HIV-1-resistant sex workers.

The goal of the present study was to determine whether there were HIV‐1 specific cellular immune responses among a subgroup of women within a cohort of Nairobi prostitutes (n = 1800) who, despite their intense sexual exposure to HIV‐1, are epidemiologically resistant to HIV‐1 infection. Of the 80 women defined to be resistant, 24 were recruited for immunological evaluation. The HIV‐1‐specific T‐helper responses were determined by IL‐2 production following stimulation with HIV‐1 envelope peptides and soluble gp120. Cytotoxic T lymphocyte responses were determined by lysis of autologous EBV‐transformed B cell lines infected with control vaccinia virus or recombinant vaccinia viruses containing the HIV‐1 structural genes env, gag and pol. Resistant women had significantly increased HIV‐1 specific T‐helper responses, as determined by in vitro IL‐2 production to HIV‐1 envelope peptides and soluble glycoprotein 120, compared with low‐risk seronegative and HIV‐1‐infected controls (P ≤ 0.01, Students t‐test). Seven of the 17 (41%) resistant women showed IL‐2 stimulation indices ≥ 2.0. HIV‐1‐specific CTL responses were detected among 15/22 (68.2%) resistant women compared with 0/12 low‐risk controls (Chi‐squared test, P < 0.001). In the two resistant individuals tested, the CTL activity was mediated by CD8+ effectors. Many HIV‐1‐resistant women show evidence of HIV‐1‐specific T‐helper and cytotoxic responses. These data support the suggestion that HIV‐1‐specific T‐cell responses contribute to protection against HIV‐1 infection.

Oral Antimicrobial Therapy for Diabetic Foot Osteomyelitis

Background: Osteomyelitis in the foot of a diabetic individual is a common complication of peripheral neuropathy, peripheral vascular disease, and infection. Operative facilities and home intravenous antibiotic therapy programs may not be available in remote or rural communities. Limited data are available regarding the treatment results of oral antimicrobial therapy, with or without limited office debridement for diabetic foot osteomyelitis. Methods: This retrospective medical record review of 325 consecutive diabetic patients who were evaluated at a multidisciplinary foot clinic identified 94 (29%) patients with 117 episodes of osteomyelitis. The most common group of organisms isolated were aerobic gram-positive cocci, and the single most frequent organism was Staphylococcus aureus. A mean of 1.6 ± 0.8 (range 1 to 4) pathogens were recovered per episode of osteomyelitis. Therapy was guided by culture results. There were 93 episodes of osteomyelitis (79 patients) that were treated with a mean of 3 ± 1 oral antimicrobial agents (with or without an initial short course of intravenous antimicrobial agents) and had adequate followup to evaluate outcome of treatment; office treatment included bone debridement in 26 (28%) and toe amputation in nine (10%) of the 93 episodes (79 patients). Results: Of the 93 episodes treated with oral antimicrobial agents (with or without an initial short course of intravenous antimicrobial agents), 75 (80.5%) episodes were put into remission. Mean duration of oral antimicrobial therapy was 40 ± 30 weeks. Mean relapse-free followup duration was 50 ± 50 weeks. Conclusions: Diabetic foot osteomyelitis was effectively managed with oral antimicrobial therapy with or without limited office debridement in most patients. This regimen may be especially useful in communities where infectious disease specialists and operative resources are limited.

Genetic analysis of human DNA recovered from minute amounts of serum or plasma

A rapid and inexpensive method is described where a small amount of serum or plasma was used as the source of DNA for genetic analysis. Using a silica gel matrix DNA was isolated from 50 microliters of archived serum or plasma. The specimens were collected from 13 individuals at two separate time points 3-6 years apart. The polymorphic region of second exon of the MHC class II gene HLA DQA1 was amplified using the polymerase chain reaction (PCR) to sufficient quantities to permit genetic analysis using allele-specific oligonucleotides (ASO). Allelic typing of each specimen was performed and the reproducibility of the method was demonstrated in that in all 13 cases the two independently isolated specimens produced the identical ASO binding patterns. No qualitative difference was noted in the amplified product generated from plasma or serum. This study demonstrates (a) that minute amounts of serum or plasma are able to provide sufficient quantity and quality of DNA to permit genetic analyses (b) and that the source of serum can be archived for many years.

Gonococcal Cervicitis Is Associated with Reduced Systemic CD8+ T Cell Responses in Human Immunodeficiency Virus Type 1—Infected and Exposed, Uninfected Sex Workers

Neisseria gonorrhoeae cervicitis and human immunodeficiency virus (HIV) type 1 frequently coinfect core transmitter populations, such as female sex workers. Gonococcal cervicitis is associated with increased viral shedding and plasma viremia in HIV-1-infected women and increased HIV-1 susceptibility in uninfected women. We studied the influence of gonococcal cervicitis on CD8(+) interferon (IFN)-gamma responses to HIV-1 and cytomegalovirus (CMV) epitopes in HIV-1-infected and in highly-exposed, persistently seronegative (HEPS) female sex workers. In HIV-1-infected women, gonococcal cervicitis was associated with reduced IFN-gamma responses in bulk CD8(+) lymphocyte populations, and intracellular cytokine staining, combined with class I major histocompatibility complex (MHC)-peptide tetramer studies, demonstrated reduced IFN-gamma production by HIV-1 epitope-specific CD8(+) lymphocytes. In HEPS sex workers, cervicitis was associated with the transient loss of systemic HIV-1-specific CD8(+) responses and with reduced function of CMV-specific CD8(+) lymphocytes. Impaired function of virus-specific CD8(+) lymphocytes may partly explain the deleterious effects of gonococcal cervicitis on HIV-1 immune control and susceptibility.

Enrichment Of Variations In KIR3DL1/S1 And KIR2DL2/L3 Among H1n1/09 Icu Patients: An Exploratory Study

Background Infection by the pandemic influenza A (H1N1/09) virus resulted in significant pathology among specific ethnic groups worldwide. Natural Killer (NK) cells are important in early innate immune responses to viral infections. Activation of NK cells, in part, depend on killer-cell immunoglobulin-like receptors (KIR) and HLA class I ligand interactions. To study factors involved in NK cell dysfunction in overactive immune responses to H1N1 infection, KIR3DL1/S1 and KIR2DL2/L3 allotypes and cognate HLA ligands of H1N1/09 intensive-care unit (ICU) patients were determined. Methodology and Findings KIR3DL1/S1, KIR2DL2/L3, and HLA -B and -C of 51 H1N1/09 ICU patients and 105 H1N1-negative subjects (St. Theresa Point, Manitoba) were characterized. We detected an increase of 3DL1 ligand-negative pairs (3DL1/S1+ Bw6+ Bw4−), and a lack of 2DL1 HLA-C2 ligands, among ICU patients. They were also significantly enriched for 2DL2/L3 ligand-positive pairs (P<0.001, Pc<0.001; Odds Ratio:6.3158, CI95%:2.481–16.078). Relative to St. Theresa aboriginals (STh) and Venezuelan Amerindians (VA), allotypes enriched among aboriginal ICU patients (Ab) were: 2DL3 (Ab>VA, P = 0.024, Pc = 0.047; Odds Ratio:2.563, CI95%:1.109–5.923), 3DL1*00101 (Ab>VA, P<0.001, Pc<0.001), 3DL1*01502 (Ab>STh, P = 0.034, Pc = 0.268), and 3DL1*029 (Ab>STh, P  = 0.039, Pc = 0.301). Aboriginal patients ligand-positive for 3DL1/S1 and 2DL1 had the lowest probabilities of death (Rd) (Rd = 28%), compared to patients that were 3DL1/S1 ligand-negative (Rd = 52%) or carried 3DL1*029 (Rd = 52%). Relative to Caucasoids (CA), two allotypes were enriched among non-aboriginal ICU patients (NAb): 3DL1*00401 (NAb>CA, P<0.001, Pc<0.001) and 3DL1*01502 (CA<NAb, P = 0.012, Pc = 0.156). Non-aboriginal patients with ligands for all three KIRs (3DL1/S1, 2DL2/L3, and 2DL1) had the lowest probabilities of death (Rd = 36%), compared to subjects with 3DL1*01502 (Rd = 48%) and/or 3DL1*00401 (Rd = 58%). Conclusions Specific KIR3DL1/S1 allotypes, 3DL1/S1 and 2DL1 ligand-negative pairs, and 2DL2/L3 ligand-positive pairs were enriched among ICU patients. This suggests a possible association with NK cell dysfunction in patients with overactive immune responses to H1N1/09, leading to severe disease.

Diabetic Foot Complications in a Northern Canadian Aboriginal Community

Background: Limited access to basic foot care and protective footwear may contribute to diabetic foot complications. The purpose of this study was to determine the prevalence of foot complications, ongoing foot care, and footwear use in diabetic subjects in a remote northern Canadian Aboriginal community. Methods: This was a cross-sectional cohort study of 169 diabetic people, including interview, physical examination, and retrospective chart review. Results: The mean age of the 169 diabetic individuals in the study was 56 ± 12 years and their duration of diabetes 10 ± 7 years. There were 139 (82%) individuals who had 418 diabetic foot complications (average, 3.0 complications per subject with complications), including toenail pathology, foot and ankle deformities, calluses, impaired pulses, neuropathy, past or present ulcer, amputation, and Charcot arthropathy. Risk classification showed that 69 (41%) individuals were at risk for future ulceration. Fifty-five (33%) individuals had inadequate footwear for their foot risk category, and only 11 (17%) of 66 individuals in the higher risk categories (categories 2 and 3) had suitable footwear. In a 7-year period, only 0.7 screening foot examinations per diabetic subject per year were documented. However, during this period, foot problems accounted for 498 (18%) local emergency room visits, 359 (16%) hospitalization days, 109 (11%) nonemergency transfers, and 4 (6%) emergency transfers to a tertiary care hospital. Conclusions: Foot and ankle complications of diabetes in this remote Aboriginal community were common and associated with substantial morbidity. Preventive diabetic foot screening examinations and footwear were inadequate. The results suggest that programs for prevention and early detection of complications are needed, including foot screening, provision of appropriate footwear, and foot care.

A Common CD4 Gene Variant Is Associated with an Increased Risk of HIV-1 Infection in Kenyan Female Commercial Sex Workers

BACKGROUND It has been predicted that CD4 C868T, a novel CD4 single-nucleotide polymorphism (SNP) that has been found to be highly prevalent among Africans, changes the tertiary structure of CD4, which may alter susceptibility to human immunodeficiency virus (HIV) infection. METHODS Participants were from a Kenyan cohort and included 87 uninfected and 277 HIV-1-infected individuals. DNA sequencing was used to determine CD4 genotype. A2.01 cells expressing similar levels of either wild-type CD4 or CD4-Trp240 as well as peripheral blood mononuclear cells from uninfected donors were infected with HIV-1(IIIB) or a Kenyan primary HIV-1 isolate. HIV-1 p24 enzyme-linked immunosorbent assay was used to determine the outcome of infection. RESULTS CD4 C868T was found to be significantly more prevalent among HIV-1-infected participants than among HIV-1-uninfected participants (P = .002), and C868T was associated with an increased incidence of HIV-1 infection as well (P = .005, log-rank test; P = .009, Wilcoxon test), with an odds ratio of 2.49 (P = .009). Both in vitro and ex vivo models demonstrated a significant association between CD4 C868T and susceptibility to HIV-1 infection (P < .001 and P = .003, respectively). CONCLUSION Overall, the present study found a strong correlation between CD4 C868T and increased susceptibility to HIV-1 infection. Given the high prevalence of both HIV infection and CD4 C868T in African populations, the effect of this SNP on the epidemic in Africa could be dramatic.

Multidisciplinary treatment of diabetic foot ulcers in Canadian Aboriginal and non-Aboriginal people

BACKGROUND Diabetic foot ulcers are a major cause of morbidity and mortality. This study evaluated the clinical outcomes in Canadian non-Aboriginal and Aboriginal diabetic patients with foot ulcers managed at a multidisciplinary, tertiary care diabetic foot clinic. METHODS A retrospective review of medical records was done for 325 patients receiving care during a 2-year period. All patients were followed at least 1 year after the initial visit. RESULTS There were 224 (69%) non-Aboriginal and 101 (31%) Aboriginal patients with 697 foot ulcers. At the initial office visit, 204 (63%) patients had lesions in Wagner grades 2-4. At the most recent evaluation (average, 79+/-73 weeks after initial clinic visit), 190 (58%) patients were rated as having a good outcome (either healed or healing), but a poor outcome (static, progression, amputation, or death) was noted in 135 (42%) patients. At the most recent evaluation, the majority of the 697 ulcers that were noted at the initial or subsequent clinic visits were healed. Aboriginal patients had a shorter average time from initial clinic visit to major lower extremity amputation (Aboriginal, 50+/-64 weeks; non-Aboriginal, 62+/-56 weeks; P<0.01). Residence in a rural or reserve community also correlated with shorter average time from initial clinic visit to major lower extremity amputation (rural or reserve, 45+/-56 weeks; urban, 66+/-61 weeks; P<0.002). When controlled for non-urban residence, Aboriginal ethnicity was not associated with poorer clinical outcome. Earlier major lower extremity amputation was significantly associated with non-urban residence, Aboriginal ethnicity, and arterial insufficiency. Poor clinical outcome was significantly associated with being referred with a lesion present, age greater than 60 years, prior lower extremity amputation or revascularization, arterial insufficiency, more than one lesion on initial presentation, longer duration of type 2 diabetes, and a higher initial Wagner grade for the most advanced lesion. CONCLUSIONS A multidisciplinary diabetic foot clinic may be successful in treating diabetic foot ulcers in Aboriginal and non-Aboriginal people. However, the frequency of poor outcome is high, consistent with the high prevalence of associated significant risk factors in this population.