Shlomo Maayan

Mutation D30N Is Not Preferentially Selected by Human Immunodeficiency Virus Type 1 Subtype C in the Development of Resistance to Nelfinavir

ABSTRACT Differences in baseline polymorphisms between subtypes may result in development of diverse mutational pathways during antiretroviral treatment. We compared drug resistance in patients with human immunodeficiency virus subtype C (referred to herein as “subtype-C-infected patients”) versus subtype-B-infected patients following protease inhibitor (PI) therapy. Genotype, phenotype, and replication capacity (Phenosense; Virologic) were determined. We evaluated 159 subtype-C- and 65 subtype-B-infected patients failing first PI treatment. Following nelfinavir treatment, the unique nelfinavir mutation D30N was substantially less frequent in C (7%) than in B (23%; P = 0.03) while L90M was similar (P < 0.5). Significant differences were found in the rates of M36I (98 and 36%), L63P (35 and 59%), A71V (3 and 32%), V77I (0 and 36%), and I93L (91 and 32%) (0.0001 < P < 0.05) in C and B, respectively. Other mutations were L10I/V, K20R, M46I, V82A/I, I84V, N88D, and N88S. Subtype C samples with mutation D30N showed a 50% inhibitory concentration (IC50) change in susceptibility to nelfinavir only. Other mutations increased IC50 correlates to all PIs. Following accumulation of mutations, replication capacity of the C virus was reduced from 43% ± 22% to 22% ± 15% (P = 0.04). We confirmed the selective nature of the D30N mutation in C, and the broader cross-resistance of other common protease inhibitor mutations. The rates at which these mutational pathways develop differ in C and subtype-B-infected patients failing therapy, possibly due to the differential impact of baseline polymorphisms. Because mutation D30N is not preferentially selected in nelfinavir-treated subtype-C-infected patients, as it is in those infected with subtype B, the consideration of using this drug initially to preserve future protease inhibitor options is less relevant for subtype-C-infected patients.

Genotypic variation of HIV-1 reverse transcriptase and protease: comparative analysis of clade C and clade B.

Objective To compare drug-resistant variants from untreated (naive) and treated patients infected with clade B or C virus. Methods Consecutive samples (165) from patients throughout Israel were analyzed. All those in the treated group were failing highly active antiretroviral therapy. Results There were 78 clade C (20 naive) and 87 clade B (14 naive) with significant differences in the prevalence of known drug-resistance mutations between the clades: in naive patients in the protease region M36I 7% and 95% (P < 0.0001), K20R 0% and 27% (P = 0.063), A71V 18% and 0% (P = 0.063), M46I 0% and 13%, and V77I 18% and 0% (P = 0.063), respectively, and in the reverse transcriptase region A98G/S 0% and 20% (P = 0.12), respectively. Most clade C viruses also showed significant differences from clade B consensus sequence at additional protease sites: R41K 100%, H69K/Q 85%, L89M 95% and I93L 80% (P < 0.0001). There were also significant differences (P < 0.03 to < 0.0001) in treated patients in clades B and C: in the protease region L10I 40% and 12%, M36I 26% and 95%, L63P 67% and 40%, A71I 38% and 7%, G73I and V77I 18% and 0%, I84V 16% and 3%, and L90M 40% and 12%, respectively; in the reverse transcriptase M41L 41% and 17%, D67N 41% and12%, K70R 30% and 7%, T215Y 48% and 29%, K219Q 21% and 7%, and A98G/S 3% and 24%, respectively. Conclusion Significantly differences between clade B and C viruses may be associated with development of differing resistance patterns during therapy and may affect drug utility in patients infected with clade C.

Nucleotide and Amino Acid Polymorphisms at Drug Resistance Sites in Non-B-Subtype Variants of Human Immunodeficiency Virus Type 1

ABSTRACT We have compared nucleotide substitutions and polymorphisms at codons known to confer drug resistance in subtype B strains of human immunodeficiency virus type 1 (HIV-1) with similar substitutions in viruses of other subtypes. Genotypic analysis was performed on viruses from untreated individuals. Nucleotide and amino acid diversity at resistance sites was compared with a consensus subtype B reference virus. Among patients with non-subtype B infections, polymorphisms relative to subtype B were observed at codon 10 in protease (PR). These included silent substitutions (CTC→CTT, CTA, TTA) and an amino acid mutation, L10I. Subtype A viruses possessed a V179I substitution in reverse transcriptase (RT). Subtype G viruses were identified by silent substitutions at codon 181 in RT (TAT→TAC). Similarly, subtype A/G viruses were identified by a substitution at position 67 in RT (GAC→GAT). Subtype C was distinguished by silent substitutions at codons 106 (GTA→GTG) and 219 (AAA→AAG) in RT and codon 48 (GGG→GGA) in PR. Variations relative to subtype B were seen at RT position 215 (ACC→ACT) for subtypes A and A/E. These substitutions and polymorphisms reflect different patterns of codon usage among viruses of different subtypes. However, the existence of different subtypes may only rarely affect patterns of drug resistance-associated mutations.

The impact of syndromic treatment of sexually transmitted diseases on genital shedding of HIV-1.

Objectives: To examine the impact of sexually transmitted diseases (STD) syndromic treatment on genital shedding of HIV and the impact among women in whom STD treatment was not successful. Design: Seventy-one HIV-infected women were included; 60 had symptomatic STD [72% with genital discharge syndrome (GDS) and 28% with genital ulcer syndrome (GUS)] and 11 controls did not have symptomatic STD. Cervical HIV load in 94% women was measured at baseline and after STD treatment. Results: Cervical HIV load at entry was significantly higher in women with symptomatic STD than in controls [median, 3.15; interquartile range (IQR), 1.90–3.34 versus median, 1.90; IQR, 1.90–2.19 log10 RNA copies/swab, respectively; P = 0.024]. Women with STD were also more likely to have detectable cervical HIV RNA (68% versus 27%; P = 0.016). Cervical HIV load was significantly higher in women with GUS than in those with GDS (median 3.46; IQR, 2.84–4.18 versus median, 2.83; IQR, 1.90–3.31 log10 copies/swab; P = 0.019). There was no significant reduction in genital HIV shedding after syndromic treatment of GDS or GUS. However, significant decreases were limited to only those with clinical improvement (median, 2.91; IQR, 1.90–3.45 versus median, 2.25; IQR, 1.90–3.08 log10 RNA copies/swab, respectively; P = 0.006). GUS was significantly associated with treatment failure, independent of plasma HIV RNA load and CD4 T-cell count (odds ratio, 4.79; 95% confidence interval, 1.32–17.46). Conclusions: The fact that STD syndromic treatment impacts very little in reducing genital HIV shedding underscores the need for appropriate validation of STD syndromic diagnosis and management to control heterosexual transmission of HIV.

Meningococcal meningitis among Rwandan refugees: Diagnosis, management, and outcome in a field hospital

OBJECTIVEnTo study the diagnostic process, clinical course, and outcome of Rwandan refugees with meningococcal meningitis, treated in an Israeli field hospital in Goma, Zaire, in the summer of 1994.nnnMETHODSnPatient hospital charts and laboratory records were reviewed with critical evaluation of clinical presentation and diagnostic tests. Patients were treated as part of a disaster relief effort in a refugee camp experiencing several coexisting lethal epidemics.nnnRESULTSnA total of 65 patients were identified as having group A meningococcal meningitis. Latex agglutination test for Neisseria meningitidis soluble antigen in the cerebrospinal fluid was found to be a superior diagnostic tool, as compared to Gram stain, and at least as effective as culture. The mortality rate was 14%; mortality was markedly affected by co-morbidity (e.g., dysentery, pneumonia, and malnutrition).nnnCONCLUSIONSnThe outcome of patients with meningococcal meningitis, treated in referral centers within a disaster area may be favorable, despite overwhelming coexisting epidemics, and may be comparable to that achieved in advanced medical facilities. Encephalopathy may be a diagnostic pitfall in the perspective of coexisting epidemics, requiring a high index of suspicion and routine lumbar puncture. The latex agglutination test is highly useful in achieving prompt diagnosis of meningococcal meningitis, in particular when sample handling for culture and microscopy is suboptimal.

The expression of CD8 on B lymphocytes in HIV-infected individuals

In HIV-1 infected individuals the CD8 + T lymphocyte population is markedly activated as reflected by increased expression of the CD38 and CD45RO activation markers and elevated serum levels of soluble CD8 antigen. We have previously shown that in vitro activation of peripheral blood lymphocytes results in the appearance of T cell markers on B cells. In the present study B lymphocytes from HIV-1-infected individuals were tested for the expression of the CD8 T cell antigen, using F(ab)2 fragments of antibodies against the CD8 and CD19 antigens. The proportion of CD19 + B cells which co-expressed CD8 was significantly elevated among 55 HIV-infected individuals (7.20 +/- 1.24%, mean +/- S.E.) as compared with among 22 normal controls (3.32 +/- 0.70%). The proportion of CD4 + cells decreased significantly in HIV-infected individuals in accordance with the progression of the infection, but no significant change in the level of CD8 + CD19 + B cells was seen in different stages of the disease. In contrast, the proportion of CD8 + B cells showed a significant correlation with the proportion of CD8 + cells.

A Population-Structured HIV Epidemic in Israel: Roles of Risk and Ethnicity.

Background HIV in Israel started with a subtype-B epidemic among men who have sex with men, followed in the 1980s and 1990s by introductions of subtype C from Ethiopia (predominantly acquired by heterosexual transmission) and subtype A from the former Soviet Union (FSU, most often acquired by intravenous drug use). The epidemic matured over the last 15 years without additional large influx of exogenous infections. Between 2005 and 2013 the number of infected men who have sex with men (MSM) increased 2.9-fold, compared to 1.6-fold and 1.3-fold for intravenous drug users (IVDU) and Ethiopian-origin residents. Understanding contemporary spread is essential for effective public health planning. Methods We analyzed demographic and virologic data from 1,427 HIV-infected individuals diagnosed with HIV-I during 1998–2012. HIV phylogenies were reconstructed with maximum-likelihood and Bayesian methods. Results Subtype-B viruses, but not A or C, demonstrated a striking number of large clusters with common ancestors having posterior probability ≥0.95, including some suggesting presence of transmission networks. Transmitted drug resistance was highest in subtype B (13%). MSM represented a frequent risk factor in cross-ethnic transmission, demonstrated by the presence of Israeli-born with non-B virus infections and FSU immigrants with non-A subtypes. Conclusions Reconstructed phylogenetic trees demonstrated substantial grouping in subtype B, but not in non-MSM subtype-A or in subtype-C, reflecting differences in transmission dynamics linked to HIV transmission categories. Cross-ethnic spread occurred through multiple independent introductions, with MSM playing a prevalent role in the transmission of the virus. Such data provide a baseline to track epidemic trends and will be useful in informing and quantifying efforts to reduce HIV transmission.

Training Laboratory Technicians from the Ethiopian Periphery in the MODS Technique Enables Rapid and Low-Cost Diagnosis of Mycobacterium tuberculosis Infection

Tuberculosis (TB) is a leading cause of morbidity and mortality and is frequently complicated by emergence of drug-resistant strains. Diagnosis of TB in developing countries is often based on the relatively insensitive acid-fast staining that does not enable susceptibility profiling. Microscopic observation drug susceptibility assay (MODS) is an inexpensive, simple method that enables rapid TB culture coupled with susceptibility testing. A 3-week MODS training of three Ethiopian laboratory technicians was conducted at Hadassah-Hebrew University Medical Center, Israel. Results of the trainee readings were blindly assessed by an experienced instructor. Two hundred fifty-five (255) trainee culture readings were evaluated throughout the course. The sensitivity and specificity were 75-100% and 31.5-100%, respectively. Multivariate analysis revealed that sensitivity and duration of incubation were positively correlated, although specificity was positively correlated with the length of training. MODS can be reliably performed by laboratory technicians inexperienced in culture techniques in developing countries, with high sensitivity and specificity reached after a brief learning period.

HIV/AIDS among Palestinians: detection, clinical presentation, prognosis and HIV testing patterns, 1994–2010

OBJECTIVESnTo describe the detection, clinical presentation, and prognosis of West Bank and East Jerusalem Palestinians infected with HIV/AIDS, and HIV testing patterns of Palestinians in the Jerusalem area.nnnDESIGN AND METHODSnThis was a case-control analysis comparing all 33 Palestinian HIV/AIDS patients who were referred to the Hadassah AIDS Center (HAC) over 17 years (1994-2010) with 77 non-Palestinian patients seen over the same period. The systematic sampling method was used to select the control group. Patterns of HIV testing were observed for the years 2002 and 2007.nnnRESULTSnMany Palestinian patients (36%) were diagnosed during their initial hospitalization, while 47.1% of non-Palestinians were diagnosed as outpatients. Significantly more opportunistic infections were detected during diagnosis among Palestinians (48.5%) than among non-Palestinians (9.1%, p<0.001). Overall mortality among Palestinian patients was 36.4% (12/33) vs. 6.5% (5/77) among non-Palestinians (p<0.001). No significant differences in the initial CD4 counts and viral load levels were noted between Palestinians and non-Palestinians (256/mm(3) and log 4.58 copies/ml vs. 271/mm(3) and log 4.49 copies/ml, respectively). Follow-up visits were more infrequent among Palestinians than among non-Palestinians: 9.8 (± 1.0) compared with 23.4 (± 12.9), respectively (p<0.001), over a median follow-up of 2.7 years for Palestinians and 8.1 years for non-Palestinians (p<0.001). With regard to HIV testing, 7.3% (72/989) of individuals tested in 2002 and 10.9% (202/1851) in 2007 were Palestinians. The most frequent reason for being tested among Palestinians was medical (e.g., before in vitro fertilization, 69.4% in 2007); among non-Palestinians it was intimate relationships (31% in 2007).nnnCONCLUSIONnThese results show that despite an overall small number of Palestinian HIV/AIDS patients, late diagnosis and high mortality are very much in evidence.

HIV infection and susceptibility to epidemic bacterial infections among Rwandan refugees

Abstract Objective: To study HIV prevalence and susceptibility to epidemic bacterial infections among Rwandan refugees treated at the Israeli military field hospital in Goma, Zaire, during a 2-week period of massive outbreaks of cholera, shigellosis, and meningitis in the summer of 1994. Methods: Anonymous testing was performed on serum samples obtained for laboratory analysis from patients who had had an intravenous line inserted at the hospitals emergency facility. The prevalence of HIV was compared among patients who presented at the emergency facility because of watery or bloody diarrhea, pneumonia, meningitis, or trauma. Results: Of the 1350 patients who were seen during the period, 127 were tested: 35 of 127 (27.5%) were HIV seropositive by enzyme-linked immunosorbent assay (ELISA) and Western blot. No statistical difference in the prevalence of HIV was found among the four categories. Conclusion: A high prevalence of HIV infection was observed among Rwandan refugees treated for severe cholera, shigellosis, pneumonia, meningitis, or trauma in a field hospital in Goma, Zaire in the summer of 1994. This rate was similar to that reported among the healthy Rwandan population. Although based on only a small sample of the sick refugees in the camps, this study suggests that HIV infection did not cause increased susceptibility to the epidemic bacterial infections seen during the 1994 refugee crisis in Rwanda.