Institutes: Ukrainian Medical and Dental Academy, Regional HIV/AIDS Prevention and Control Center, Poltava Ukraine Toll-like receptor 4 polymorphism influence on the development of opportunistic diseases in HIV-infected patients The presenting author is Tetiana Kyrychenko Tetiana Kyrychenko Authors: G. Dubynska, T.Koval, I.Kaidashev, V.Korshenko

Ukraine has one of the fastest growing HIV/AIDS epidemics in the world and the highest in all of Europe. Late 2011 Ukraine numbered 210,000 HIV- positive persons. Dynamics of officially registered new cases of HIV- infection in Ukraine (on 100 thousand population)

Toll-like receptors (TLRs) have a key role in innate immunity and mutations in the genes encoding these receptors have been associated with increased or decreased susceptibility to infections.

 Objectives:  to study the prevalence of TLR4 Asp299Gly SNP among the healthy and HIV-positive persons in Ukraine ;  to find out HIV clinical manifestations depending on TLR4 genotype variants. Asp299Gly, A>G chromosome 9q32–33

Methods Patients. The study included 180 antiretroviral treatment-naive HIV-1 infected patients. Patients. The study included 180 antiretroviral treatment-naive HIV-1 infected patients. The stage of the disease was recorded in accordance with the expanded European AIDS case definition. Laboratory procedures. Laboratory procedures. Routine CD-4 cell count was determined using conventional flow cytometry with fresh blood samples from all patients. Plasma HIV-RNA was also routinely measured by the polymerase chain reaction (PCR) (Real time HIV-1, Abbout). TLR4 genotyping was performed by real- time PCR using oligonucleotide primers Tercik PCR system, DNA-technology, Moscow) TLR4 genotyping was performed by real- time PCR using oligonucleotide primers (Tercik PCR system, DNA-technology, Moscow)

Characteristic of a Cohort of 180 patients infected with HIV-1 and studied in Relation to SNP of TLR4 Characteristic Wild type (AA, n=165) Asp299Gly heterozygous (AG, n=15) Male-to-female ratio 98:67 10:5 Ageª, mean years ± standard deviation 35,21±0,5135,13±1,33 Nadir CD-4 cell countª, mean cell/mm³ ± standard deviation 259,52±16,2233,6±30,38 Maximum viral loadª, median copies/mL Clinical stage(%) ABC 11 (6,6) 72 (43,7) 82 (49,7) 2(13,3)2(13,3)11(73,4)

Historyof opportunistic infections in 180 HIV-infected patients History of opportunistic infections in 180 HIV-infected patients

Historyof opportunistic infections in 180 HIV-infected patients in relation to SNP of the Toll-like receptor 4 gene (No. (%) of patients) History of opportunistic infections in 180 HIV-infected patients in relation to SNP of the Toll-like receptor 4 gene (No. (%) of patients) Variable Wild type (AA, n=165) Asp299Gly heterozygous (AG, n=15) p Mycobacterium tuberculosis, pulmonary or extrapulm. 59 (35.7) 11 (73.3) 0.007* Bacterial pneumonia, recurrent 42 (25.5) 8 (53.3) 0.021* Hairy leukoplakia, oral 51 (30.9) 9 (60) 0.022* Herpes zoster (shingles), involving two or more episodes or at least one dermatome 29 (17.6) 7 (46.7) 0.007* AG compared to AA, Mann-Whitney U-test *AG compared to AA, Mann-Whitney U-test

Historyof opportunistic infections in 180 HIV-infected patients in relation to SNP of the Toll-like receptor 4 gene (No. (%) of patients) History of opportunistic infections in 180 HIV-infected patients in relation to SNP of the Toll-like receptor 4 gene (No. (%) of patients) Variable Wild type (AA, n=165) Asp299Gly heterozygous (AG, n=15) p Chronic hepatitis C 80 (48.5) 12 (80) 0.018* Candidiasis, oropharyngeal or esophageal 78 (47.3) 8 (53.3) >0.05 Wasting syndrome caused by HIV (involuntary weight loss >10% of baseline body weight) associated with either chronic diarrhea or chronic weakness and documented fever for ≥1 month) 42 (25.5) 6 (40) >0.05 Herpes simplex: chronic ulcers (>1 month in duration) 28 (17) 5 (33.3) >0.05 Any 19 (11.5) 3 (20) >0.05 AG compared to AA, Mann-Whitney U-test *AG compared to AA, Mann-Whitney U-test

Distribution of the 299 alleles of the patients when they were grouped by CD-4 T-cell counts equal to or above and below 350, cells/mm³, respectively Only patients with CD4 T-cell counts above 200cells/mm³ had the prevalence of the Asp299Gly polymorphism (12%) *Only patients with CD4 T-cell counts above 200cells/mm³ had the prevalence of the Asp299Gly polymorphism (12%) significantly higher than patients with CD4 T-cell counts below than 200 cells/mm³ (3.7%), p<0.05

Detection of opportunistic infections in HIV- infected patients with different levels of CD4 (No. (%) of patients) CD-4 T-cell counts /mm³ < >350 TLR-4 genotype AA,n=31AG,n=3AA,n=102AG,n=9AA,n=32AG,n=3 Tuberculosis20(64.5)3(100)34(33.3)6(66.7)*5(15.6)2(66.7)* Toxoplasmosis12(38.8)3(100)*0(0)1(11.1)*00 Herpes zoster 2(6.5)1(33.3)24(23.6)5(55.6)*3(9.3)1(33.3) Pneumonia10(32.3)1(33.3)22(21.6)5(55.6)*10(31)2(66.7) AG compared to AA, X²-test, p<0.05 * AG compared to AA, X²-test, p<0.05

Conclusions  This study suggests a grater risk for development of active tuberculosis, bacterial pneumonia, herpes zoster, hairy leukoplakia, toxoplasmosis of brain, chronic hepatitis C in patients with the Asp299Gly TLR4 polymorphism  our results demonstrate the impact of a TLR4 genetic polymorphism on development of opportunistic infections for CD4 levels more than 100.

Acknowledgments  G.Dubynska, prof., department of infectious diseases, Ukrainian Medical and Dental Academy, Poltava, Ukraine  T.Koval, prof., department of infectious diseases, Ukrainian Medical and Dental Academy, Poltava, Ukraine  I. Kajdashev, prof., chair of research department, Ukrainian Medical and Dental Academy, Poltava, Ukraine  V.Korshenko, chief doctor of Regional centre of HIV/AIDS Poltava, Ukraine