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Abstract: HIV-1 is the major cause of the worldwide pandemic. Therefore, Determination of the HIV-1 genetic subtypes has been shown to be useful for understanding the origin and global spread of the virus. HIV-1 subtyping may also be helpful for evaluation of the pathogenicity, transmissibility,andcandidateHIVvaccines. ThisstudyaimedtodeterminetheHIV-1genotypesintheIraqipopulationfromdifferentcitiesof Baghdad province –Iraq. This study included 65 HIV infected patients; they were referred for diagnosis and treatment to the Ibn Zuhur Hospital and AL- Karama Hospital in Baghdad during the period from October 2019 to May 2020. Data were collected for each patient using a special form. Following amplification gp41 region of env gene for HIV-1 with nested -RT- PCR, the sequences of this region was determined, HIV-1 subtype was reported and phylogenetically analysed. The age of HIV infected patients in this study ranged between 20 - 76 years. Most of them were males (83%). After amplified 628bp of env gene coding region by nested PCR, Nucleic acid sequencing was conducted to emphasize their specificity and introduce the ultimate means to detect the virus subtype. Therefore, in Iraqi patients from Baghdad -Iraq, HIV-1 genotype was belonging to group M subtypes A sub- subtypes A1 was detected.
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Human Immunodeficiency Virus Genotyping in Baghdad, Iraq
Manuscript Number: S-957
NAAS Rating: 5.79
Abstract: HIV-1 is the major cause of the worldwide pandemic. Therefore, Determination of the HIV-1 genetic subtypes has been shown to be
useful for understanding the origin and global spread of the virus. HIV-1 subtyping may also be helpful for evaluation of the pathogenicity,
transmissibility, and candidate HIV vaccines. This study aimed to determine the HIV-1 genotypes in the Iraqi population from different cities of
Baghdad province –Iraq. This study included 65 HIV infected patients; they were referred for diagnosis and treatment to the Ibn Zuhur Hospital
and AL- Karama Hospital in Baghdad during the period from October 2019 to May 20 . Data were collected for each patient using a special 20
form. Following amplification gp41 region of gene for HIV-1 with nested -RT- PCR, the sequences of this region was determined, HIV-1 env
subtype was reported and phylogenetically analysed. The age of HIV infected patients in this study ranged between 20 - 76 years. Most of
them were males (83%). After amplified 628bp of gene coding region by nested PCR, Nucleic acid sequencing was conducted to env
emphasize their specificity and introduce the ultimate means to detect the virus subtype. Therefore, in Iraqi patients from Baghdad -Iraq, HIV-1
genotype was belonging to group M subtypes A sub- subtypes A1 was detected.
Keywords: HIV-1, Genotyping, Nested-PCR, Baghdad, Phylogenetic analysis
Hayder Saad Al- Qassab and Nawal Mohammed Utba*
Department of Biology, College of Science, University of Baghdad, Al-Jaderia, Baghdad-10071, Iraq
Indian Journal of Ecology (2022) 49 Special Issue (19): 318-323
Acquired immunodeficiency syndrome (AIDS) is
associated with high morbidity and mortality due to the
increased risk of opportunistic infections and malignancy that
caused by global pandemic virus which is called the Human
immunodeficiency virus (HIV). The WHO reported that HIV
prevalence in the Eastern Mediterranean Region is low
epidemic (0.1%), since Iraq is one of the countries in this
region was characterized as a low-prevalence HIV epidemic.
However, the number of HIV infected cases registered in the
Iraqi Ministry of Health after 2008 have increased more than
previous years. The new cases in Iraq were increased each
year from 5 cases in 2008 to 124 cases in 2018 (MOH 2019).
The human immunodeficiency viruses have a high degree of
genetic variation; which regards one of an important
advantage to confer survival against negative selection
pressures such as host antiretroviral agents and immune
response. The high genetic diversity of HIV results from a
high replication rate of the virus, error in replication, and
recombination-prone reverse transcription (Sherpa et al
2020). HIV has two types HIV-1 results from chimpanzees
and gorillas living in Western Africa, and consists of four
groups (M, N, O, and P) ( HIV-2 Nikolopoulos et al 2016).
result from endangered West African primate sooty
mangabey ( The HIV-1 differing genetically D'arc et al 2015).
by 55% and it is more widespread than from HIV-2 (Beloukas
et al 2016). The group M from HIV-1 causing up to 85% of
global HIV infections and can be subdivided into 9 subtypes
(A, B, C, D, F, G, H, J, and K), at least six sub-subtypes of A
and F (A1–A6, F1–F2), circulating recombinant forms
(CRFs), and numerous unique recombinant forms (URFs),
104 CRFs have been registered in April 2020 by the Los
Alamos National Laboratory. The A subtype is common in
Eastern Africa The B (Abecasis and Vandamme 2015).
subtype is the dominant form in Europe, the Americas,
Japan, and Australia. The subtype B is the most common
form in the Middle East and North Africa. The C subtype is the
main in Southern and Eastern Africa, India, Nepal, and parts
of China. The D subtype is seen only in Eastern and Central
Africa. The E subtype in Southeast Asia is the dominant form
for heterosexuals as transmission rate is much higher than
most other subtypes The F Subtype has (Sallam et al 2017).
been found in central Africa, South America, and Eastern
Europe. The G and the CRF02-AG Subtypes have been
found in Africa and Central Europe, The H subtype is limited
to central Africa. The K Subtype is limited to the Democratic
Republic of Congo (DRC) and Cameroon (Fox et al 2010) .
The J Subtype found in North, Central and West Africa, and
the Caribbean ( Determination of the genetic Cohen 2014).
subtypes of human immunodeficiency virus type 1 (HIV-1)
has been shown to be useful for understanding the origin and
global spread of the virus. Subtyping may also be helpful for
evaluation of pathogenicity, transmissibility, and candidate
HIV vaccines ( Depending on our best Kessler et al 2001).
knowledge, no study has been performed in Iraq for
identifying HIV genotypes and its distribution among Iraqi
patients. This study aimed to determination HIV genotypes
and its distribution among Iraqi patients in Baghdad province
and compares the result with the neighbor countries. In
addition, identification of genetic relatedness among local
HIV isolates.
Study subjects: A total of 65 patients infected with HIV-1
were included in this study and were referred for the
diagnosis and treatment to IbnZuhur Hospital and AL-
Karama Hospital in Baghdad during October 201 to May 9
2020. Anti - HIV antibodies were detected in their sera by
ELISA technique and confirmed by Western blot test and real
time PCR analysis to expose the viral genetic material. This
investigation with the diagnosis was done by the advisor
medical staff in the previous hospital. Data were collected for
each patient using a special form; all participants gave
informed consent before enrolling in the research, which was
approved by the Biology Department, College of Science,
Baghdad University ethics committee.
Sample collection: Initially, 5 mL blood samples were
collected in ethylene diaminetetra acetic acid blood collection
tubes. The plasma was separated by centrifugation at 3000
r/min and frozen at −20°C until use for RNA extraction. The
HIV gp 41 regain (628bp), were amplified and sequenced env
to determine the subtype .HIV RNA extraction and cDNA
synthesis: μ HIV RNA was extracted from 200 L of plasma
using the column purification method (Viral Nucleic Acid
Extraction Kit II (Geneaid, Taiwan) according to manufacturer
instructions. PCR reaction for amplification of gp41env
regains was done according to Badreddine et al (2007). With
some adjustment GoTaq® 1-Step RT-qPCR System kit
(Promega/ USA) contains all the components necessary for
sequential cDNA synthesis and amplification by PCR in one
tube (one-step RT-PCR).
PCR mixture was prepared by using an Eppendorf
gradient PCR system thermal cycler in two runs In the first .
run conventional PCR was achieved with 10 l volume (2 l of μ μ
extracted RNA + 8 l of Master Mix, RT-Mix, MgCl2 primers μ ,
(Macrogen, Korea), and Nuclease free water (Table 1). In
reaction conditions as follows; enzyme activation at 37 C for ˚
15 min , initial denaturation at 95 C for 5 min for 1 for 1 cycle ˚
cycle, after those 40 cycles of PCR were performed. Each
Id Primer name Thermo cycler type Sequence (5ˊ-3ˊ)
Table 1. Sequence of primers
cycle consisted of denaturation at 95 C for 30 Sec., ˚
a at 50 for 30 Sec. at72 for 1 min. ˚ ˚ nnealing C and extension C
then, at for 7 min. for 1 cycleand˚ final extension 72 C hold at
10˚for 10min.
In the second run, Nested PCR with 25 l volume was μ
achieved. An aliquot (3 L) of the primary PCR product and μ
primers (Table 1) were used as follows: initial denaturation at
95°C for 5 min. for 1 cycle, then 30 cycles consist
denaturation for 0.30 min. at 95°C, annealing for 0.30 min. at
50°C and extension for 1 min. at 72 °C. Finally, 1 cycle of final
elongation at 72°C for 7 min. and hold 10 °C for 10 min. was
used for all PCR reactions. The results were checked by
electrophoresis of the nested PCR products on 1.5%
agarose gel and visualized with ethidium bromide under UV
light. The PCR products (628 pb) were confirmed by
comparing its molecular weight with 100bp DNA Ladder.
However, PCR product was sequenced using ABI3730XL,
automated DNA sequencer by Macrogen Corporation
Statistical analysis: Sequences were then compared with
the HIV sequences present in Los Alamos National
Laboratory on line at (
/sequence/BASIC_BLAST/basic_blast.html), Geneious
program version 2021.01 and GenBank using the National
Center Biotechnology Information (NCBI) online at
( the NCBI program and aligned
with the corresponding sequence fragments of the database
using the Basic Local Alignment Search Tool (BLAST)
BLASTX program ( Phylogenetic trees Morgulis et al 2008).
were created by the neighbor-joining method using the
MEGA_ X program version 10.1.8.
The plasma samples were collected from 65 Iraqi HIV
infected patients. Of these 83% were males and rest were
females The primary routes of with age 20-76 years.
HIV/AIDS transmission were sexual contact (72.3%),
homosexual routes (12.3%) and other routs (15.4%). To
understand the genetic variation and characterization of HIV-
1 in the current study, eleven positive samples were
sequenced after amplified 628bp of gene coding region env
by nested PCR. Nucleic acid sequencing was conducted to
319Human Immunodeficiency Virus Genotyping in Baghdad, Iraq
emphasize their specificity and introduce the ultimate means
to detect the virus subtype. Therefore, amplicons have HIV-1
isolated sequences of conserved regions that had not been
previously sequencing recorded in Iraq and showed that the
HIV-1 genotype was belonging to group M subtypes A sub-
subtypes A1. env Ten sequences isolates of the gene region
from HIV-1 was recorded in Genebank (NCBI) under
accession numbers [LC594633, LC594634.1, LC594635.1,
LC594636.1, LC594637.1, LC594638.1, LC594639.1,
LC594640.1, LC594641.1, and LC594642.1].
The results of this study with studies conducted in
neighbouring countries such as Iran, Turkey, Kuwait and
Saudi Arabia, and there are differences in the distribution and
prevailing HIV-1 subtypes in each country Although, HIV-1 .
Subtype B was predominated in most neighbouring
countries, Subtype A rather than B was reported in Baghdad
–Iraq and this different results evaluated may be due to the
main source of infection is different from other countries and
to the small samples size in this study. In Iran Sarrami-
Forooshani et al (2006) reported that the first molecular study
of HIV-1 genotypes suggests two parallel outbreaks in
distinct high-risk populations and may offer clues to the origin
and spread of infection. HIV-1 subtyping and phylogenetic
analysis were done to HIV-1 seropositives belonging to the
two groups (hemophiliacs and intravenous drug users) and
all hemophiliacs are infected with HIV-1 subtype B and all
intravenous drug users are infected with HIV-1 subtype A.
Since intravenous drug abuse is the major transmission route
in Iran, HIV-1 subtype A is likely to be the dominant viral
subtype circulating in the country The analysis of genetic .
distances showed subtype B viruses in Iran to be twice as
heterogeneous as the subtype A viruses. phylogenetic
analysis showed subtypes A1 and B in 58 and 42% patients,
respectively (Baesi et al 2012). Baesi et al (2016) the
recombinant CRF 35_AD (88%) be the predominant HIV
subtype, followed by subtypes CRF 28_BF in 8% and CRF
29_BF in 4% of patients failing treatment in Tehran, which is
consistent with other reports of HIV in Iran, and in addition.
Mozhgani et al (2017) reported that CRF35-AD was specified
as the predominant circulating subtype among different high-
risk groups. Most of the HIV-positive patients (70%) were
intravenous drug users (IDUs) after the p17 region of the gag
and the c2-v5 region of the genes were sequenced and env
then clustered by phylogenetic analyses. In Turkey, Ziyade et
al (2019) indicated that HIV-1 subtype B was most frequent
type (39.3%). Subtype A (17.9%;), CRF02_AG (14.3%),
subtype C (10.7%), B+CRF02_AG recombinant (3.6%),
CRF01_AE (3.6%), subtype D (3.6%), as well as subtype F
(3.6%) and subtype G (3.6%) strains were also detected in
the circulation. Biçeroğlu et al (2014) reported that the
subtypes of the isolates according to phylogenetic analysis
were as 44.2% subtype B, 34.2% CRF42_BF, 8.5%
B/CRF02_AG recombinants, 7.1% sub-subtype A1, 1.4%
sub-subtype F1, %1.4 CRF 25_cpx, 1.4% CRF02_AG and
1.4% CRF01_AE In Kuwait, Chehadeh et al (2015) .
observed that a total of ten different HIV-1 subtypes and
recombinant forms were in Kuwait with predominance of
subtypes B, C, and CRF01_AE. However, in Saudi Arabia,
Zaki et al (2020) reported that a total of 57 samples of
treatment-naive patients were tested for HIV-1 genotypes
through direct sequencing. The HIV-1 Genotypes were as
follow; C: 66.6%, D: 10.5%, G: 8.8%, B: 7.0%, CRF01_AE:
3.5%, A and CRF02_AG: 1.8% Molecular epidemiological .
studies showed that, with the exception of Sub-Saharan
Africa, where almost all subtypes, CRFs, and several URFs
have been detected, there is a specific geographic
distribution pattern for HIV-1 subtypes (Hemelaar et al2006).
This distribution pattern seems to be the consequence of
either accidental trafficking (viral migration), with a resulting
“founder effect,” or a prevalent route of transmission, which
results in a strong advantage for and local predominance of
the prevalent subtype transmitted in that population
(Buonaguro et al 2007). HIV-1 sequences from the gene env
region studied in all 11 HIV-1 isolates are displayed in the
phylogenetic tree constructed by the neighbour-joining
method. The optimal tree with the sum of branch length =
0.36477717. The tree is drawn to scale, with branch lengths
(above the branches) in the same units as those of the
evolutionary distances used to infer the phylogenetic tree.
The evolutionary distances were computed using the
maximum composite likelihood method, and are in the units
of the number of base substitutions per site. This analysis
involved 17 nucleotide sequences. There were a total of 523
positions in the final dataset (Fig. 1). Separate clusters of
related HIV-1 strains can be observed among isolates from
Iraqi HIV-1 infected patients in Baghdad province of
genotypes M Subtype A sub- subtypes A1. However, distinct
clusters of highly related sequences were observed in A1
sub-sub genotypes, suggesting highly prevalence and
endemic circulation of HIV-1 strains belonged to M subtype in
this area.
In phylogenetic tree analysis the isolate showed nearly to
the Uganda, Tanzania, Cameroon, and Rwanda isolates.
Greater percent near to Uganda isolate under accession
number [ID: KU865746.1] and less percent with another
countries Rwanda accession numbers [ID: KX983501.1, and
ID: JX508883.1], Tanzania accession numbers [ID:
AY7 75603.1 ], Came roon ac c ession number [ID:
MK086122.1], and Kenya accession number [KT022382.1].
The genotype results of HIV in this study were comparable
320 Hayder Saad Al- Qassab and Nawal Mohammed Utba
Fig. 1. Phylogenetic tree of 11 HIV-1 isolates sequences in the region, clustering based on neighbor-joining method by env
using MEGA-X program version 10.1.8
with the results of many neighbor countries. In Iran a study
conducted at 2013 reported that all HIV -1 genotype M
subtype A1 related to Ugandan , Kenyan and Sub-Saharan
isolates detected by used gene and gene (env gag Baesi et al
2014). In other study the viral sequences of the Iranian IDUs
are strongly related and form a single cluster within the A
subtype related to African Ugandan/ Kenyan Sub-Saharan
isolate ( pointed Tagliamonte et al 2007). Leung et al (2019)
that in Hong Kong, most of the A1 isolates were strongly
clustered with the reference sequences collected in African
countries such as Cameroon, Kenya, and Uganda.
The origin of HIV-1 has been documented around
Kinshasa in the present day Democratic Republic of Congo
around the 1920s from where it spread along a transport
network to other areas in Sub-Saharan Africa, West Africa,
Europe and the rest of the world ( This global Faria et al 2014).
spread was marked by a geographically defined distribution
of several genetically distinct viruses. Subtype A remains the
most prevalent strain in parts of East Africa, Russia and
former Soviet Union countries; subtype B in Europe,
Americas and Oceania; subtype C in Southern Africa and
India; CRF01_AE in Asia and CRF02_AG in Western Africa.
Recent studies based on near full-length genome
sequencing highlighted the growing importance of
recombinant variants and subtype C viruses (Bbosa et al
2019). There was significant increase in the prevalence of
recombinants in some countries (Lee et al (2017) and several
newly discovered CRFs/URFs in parts of Asia and Africa
( The co-infection or super-infection by Miao et al 2018).
divergent HIV-1 strains has become more common in regions
where multiple subtypes co-circulate (Li et al 2017) Viral .
sequences from recombinant forms affect the accuracy of
phylogenetic reconstructions. This is due to the different
regions of the aligned recombinant genome having distinct
evolutionary relationships that affect tree topology and
branch lengths ( Furthermore, such Paraskevis et al 2009).
sequences introduce errors in phylodynamic inferences by
biasing ancestral state reconstruction and estimations for the
most recent common ancestor (Frost et al 2015) As the .
prevalence of recombinant forms increases, these
challenges are expected to become more important form
molecular epidemiological-based investigations. Recovery
321Human Immunodeficiency Virus Genotyping in Baghdad, Iraq
of near full-length HIV-1 genomes from a plasma source
coupled with advances in next generation sequencing (NGS)
technologies and development of universal methods for
amplifying whole genomes of HIV-1 circulating in a target
geography or population provides the opportunity for a
detailed analysis of HIV-1 strain identification, evolution and
dynamics ( The scale up of cheaper Alampalli et al 2017).
near full-length genome sequencing approaches is critical in
improving the accuracy of viral classification. The high
mutation rate of HIV is one of the major factors that have
hindered development of an HIV vaccine; this problem is
compounded by the enhanced diversity of the virus resulting
from its high recombination rate ( Thus, Su et al 2000).
understanding the molecular epidemiology of HIV/AIDS is
increasingly important as it will shed light on the origin and
global distribution of AIDS, and will facilitate.
Although, HIV infection prevalence in Iraq was low, the
number of new cases of HIV infected patients increased in
the past recent years accompanied with changing in the
transmission mode of HIV from the contaminated blood and
blood products to the sexual contact. The determination of
HIV genetic subtypes has been shown to be useful for
understand the origin and global spread of the virus. On the
other hand, HIV subtyping may also be helpful for
development of candidate HIV vaccines, evaluation of
pathogenicity and transmissibility.
Ethics Approval and Consent to Participate: Ethical
clearance was obtained from the Biology Department,
College of Science, University of Baghdad ethics committee
in accordance with the Helsinki Declaration.
Human and Animal Rights: All human research procedures
followed were in accordance with the ethical standards of the
committee responsible for human experimentation
(institutional and national), and with the Helsinki Declaration,
2013.No Animals were used in this research.
Consent for publication: Written and informed consent was
obtained from the participants prior to the study.
Conflict of interest: There are no conflicts of interest.
Acknowledgements: Authors would like to express their
sincere thanks to the all HIV patients participated in this
study. We are grateful to staff of IbnZuhur Hospital, AL-
Karama Hospital and CDC for their support for this project.
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Received 11 January, 2022; Accepted 05 May, 2022
323Human Immunodeficiency Virus Genotyping in Baghdad, Iraq
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Determination of human immunodeficiency virus-1 (HIV-1) genotypes and identification of antiretroviral drug-resistant mutations. Among treatment naïve HIV patients in Jazan, Saudi Arabia. HIV is a major public health problem. HIV genotyping and antiretroviral resistance testing is an important guide for better management of treatment-naive. Antiretroviral resistance testing before starting of treatment regimen leads to a better virological response. A total of 57 samples of treatment-naive patients were collected from King Fahd Central Hospital in Jazan, Saudi Arabia. Samples were tested for HIV-1 antibodies, western blot, viral load, HIV-1 genotypes through direct sequencing, and antiretroviral resistance testing. The HIV-1 Genotypes were as follow; C: 66.6%, D: 10.5%, G: 8.8%, B: 7.0%, CRF01_AE: 3.5%, A and CRF02_AG: 1.8% each. 77.2% of cases showed susceptibility to the 3 major classes of antiretroviral drugs; Protease inhibitor (PI), Nucleoside reverse transcriptase inhibitor (NRTI), and non-nucleoside reverse transcriptase inhibitors (NNRTI); while 8.8% had mutations conferring resistance to NRTI. Mutations conferring resistance to PI were detected in 7.0% of cases, and 1.8% of cases had mutations conferring resistance to both NRTI and PI. Mutations conferring resistance to NNRTI were detected in 5.3% of cases. Mutations associated with antiretroviral drugs include (V82A+I84IV), (L10F+Q58E), (L10F+V82Y), L10FV, L33LF, L89LMV, M184V, E138A, V106I, and V179VD. The prevalence of HIV-1 antiretroviral resistance mutations is 22.8% in the studied population, which may warrant antiretroviral drug resistance testing as a pretreatment to help and guide physicians for the proper HIV treatment.
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Human immunodeficiency virus (HIV) comprises two genotypes, namely HIV-1 (group M, N, O and P) and HIV-2 (group A to H), which differ in their envelope glycoproteins and other antigenic epitopes despite their morphological and biological resemblance.Group M of HIV-1 responsible for 95% of HIV infections worldwide is composed of nine subgroups. In addition to subgroups, group M contains also two recombinant forms, known as circulating recombinant form (CRF) and unique recombinant form (URF). The first case of HIV/acquired immun deficiency virus (AIDS) in Turkey was reported in 1985 and the current number of cases reached a total of 18.557 including 1736 with AIDS based upon the surveillance data of Ministry of Health between October 1985 and November 2018. The aim of this study was to determine the prevalence of HIV-1 strains isolated from HIV positive autopsy cases detected by HIV polymerase chain reaction (PCR) and determine drug resistance. Twenty eight cases [17 males, 11 female: age ranged between 3 months and 66 years (median: 35 years)] found to be HIV positive among the autopsy cases sent for HIV1 PCR study and serological screening between 2011-2017 were recruited in the study. For identification of subtypes in HIV-1 isolates, most-preferred analysis tool was used [HIVdb Stanford University Genotypic Resistance Interpretation Algorithm (]. Phylogenetic tree was made according to direct sequencing of HIV-1 reverse transcriptase (pol) region and phylogenetic analysis was evaluated in 23 cases. Los Alamos National Laboratory were trimmed from full-length genomes. Phylogenetic analysis of the 870 base pair of the pol gene region was performed using CLC Sequence Viewer v8.0 (Qiagen Aarhus A/S, software. The phylogenetic tree was obtained according to the neighbor-joining method and the Jukes-Cantor nucleotide distance scale and bootstrap value was set at 1000. In our study, subtype B was found to be most frequent type (39.3%; 11/28). Subtype A (17.9%; 5/28), CRF02_AG (14.3%; 4/28), subtype C (10.7%; 3/28), B+CRF02_AG recombinant (3.6%; 1/28), CRF01_AE (3.6%; 1/28), subtype D (3.6%; 1/28), as well as subtype F (3.6%; 1/28) and subtype G (3.6%; 1/28) strains were also detected in the circulation. Analysis of our results showed that 32.1% (9/28) of the samples exhibited resistance mutations. Detected mutations were as follows: M41L, T215C, K65R, M184V, responsible for nucleoside reverse transcriptase inhibitor (NRTI) resistance; K103N, Y181C, G190A, responsible for non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance; D30N, M46I, responsible for protease inhibitor (PI) resistance. NRTI, NNRTI and PI mutation rates in the samples were found as 21.4%, 7.1% and 3.6%, respectively. Although number of samples analyzed in our study is low, we can propose that they resemble the strains circulating in Turkey. The results of our study; although the subtype B is still dominant in our country, it supports other studies reporting that there are non-B subtypes and an increase in CRF rates in recent years. Phylogenetic analysis is widely regarded as the gold standard technique to determine the subtypes of HIV-1. Molecular epidemiologic studies related to HIV may be important in monitoring HIV subtype patterns and spreading pathways in that country. As a result; the opportunity to collect postmortem HIV sequences in a database appears to have occurred, and as this database expands, its usability is available. Therefore, it is thought that HIV subtypes and mutation information may be useful.
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Background The HIV-1 spread in the Middle East and North Africa (MENA) has not been previously characterised using the phylogenetic approach. The aim of the current study was to investigate the genetic diversity and domestic transmission of HIV-1 in the MENA. Methods A total of 2036 HIV-1 sequences available in Genbank and collected in the MENA during 1988–2016 were used together with 715 HIV-1 reference sequences that were retrieved from Genbank based on genetic similarity with the MENA sequences. The REGA and COMET tools were used to determine HIV-1 subtypes and circulating recombinant forms. Maximum Likelihood and Bayesian phylogenetic analyses were used to identify and date HIV-1 transmission clusters. Results At least 21 HIV-1 subtypes and recombinant forms were prevalent in the MENA. Subtype B was the most common variant (39%), followed by CRF35_AD (19%) and CRF02_AG (14%). The most common genetic region was pol, and 675 partial pol sequences (average of 1005 bp) were eligible for detailed phylogenetic analysis. Fifty-four percent of the MENA sequences formed HIV-1 transmission clusters. Whereas numerous clusters were country-specific, some clusters indicated transmission links between countries for subtypes B, C and CRF02_AG. This was more common in North Africa compared with the Middle East (p < 0.001). Recombinant forms had a larger proportion of clustering compared to pure subtypes (p < 0.001). The largest MENA clusters dated back to 1991 (an Algerian CRF06_cpx cluster of 43 sequences) and 2002 (a Tunisian CRF02_AG cluster of 48 sequences). Conclusions We found an extensive HIV-1 diversity in the MENA and a high proportion of sequences in transmission clusters. This study highlights the need for preventive measures in the MENA to limit HIV-1 spread in this region.
Purpose of review: To provide a summary of the current data on the global HIV subtype diversity and distribution by region. HIV is one of the most genetically diverse pathogens due to its high-mutation and recombination rates, large population size and rapid replication rate. This rapid evolutionary process has resulted in several HIV subtypes that are heterogeneously globally distributed. Recent findings: Subtype A remains the most prevalent strain in parts of East Africa, Russia and former Soviet Union countries; subtype B in Europe, Americas and Oceania; subtype C in Southern Africa and India; CRF01_AE in Asia and CRF02_AG in Western Africa. Recent studies based on near full-length genome sequencing highlighted the growing importance of recombinant variants and subtype C viruses. Summary: The dynamic change in HIV subtype distribution presents future challenges for diagnosis, treatment and vaccine design and development. An increase in recombinant viruses suggests that coinfection and superinfection by divergent HIV strains has become more common necessitating continuous surveillance to keep track of the viral diversity. Cheaper near full-length genome sequencing approaches are critical in improving HIV subtype estimations. However, missing subtype data and low sequence sampling levels are still a challenge in some geographical regions. VIDEO ABSTRACT:
Currently, complex HIV-1 recombination among the B', C, and CRF01_AE genotypes have frequently arisen in Yunnan, China. A novel HIV-1 complex circulating recombinant form (CRF) consisting of B', C, and CRF01_AE (CRF96_cpx) was recently characterized from three epidemiologically unlinked individuals. Two strains of them were isolated from the injecting drug users in this study, the remaining one strain (JL. RL01) obtained from a previous report in Jilin province. Phylogenetic analysis based on near full-length genome (NFLG) revealed that CRF96_cpx formed a distinct monophyletic cluster supported by a high bootstrap value of 100%, distantly related to all known HIV-1 subtypes/CRFs. CRF96_cpx had a CRF01_AE backbone with three subtype B' and C segments inserted, respectively, in the gag and pol region. Further, subregion tree analysis showed that CRF01_AE backbone and subtype B segment inserted originated from a Thai-CRF01_AE lineage, whereas subtype C fragment inserted was from an India C lineage. They are different from previously documented CRF01_AE/B/C forms in its distinct backbone, inserted fragment size, and breakpoints. This highlighted the importance of continual monitoring of genetic diversity and complexity of HIV-1 strains in Yunnan, China.
Finding the predominant circulating subtype of human immunodeficiency virus type 1 (HIV-1) and surveying co-infection with other infectious viruses are crucial to making preventive decisions. To this end, 50 Iranian HIV-positive patients made up of 37 men and 13 women were selected. Most of the HIV-positive patients (70%) were intravenous drug users (IDUs), and 48 and 32% of patients were co-infected with HCV and HBV, respectively. The rate of simultaneous infection with HIV, HCV, and HBV was found to be 6%. The p17 region of the gag and the c2-v5 region of the env genes were sequenced and then clustered by phylogenetic analyses. CRF35-AD was specified as the predominant circulating subtype among different high-risk groups. In our survey, most of the patients in the IDU group had co-infections with HCV and HBV. Some possible reasons for the increased transmission risk of HIV in IDUs could be low levels of education, poor hygiene and housing conditions, and limited access to health services.
Objectives: To estimate prevalence, examine time trends, and test for clinical correlates and outcomes associated with HIV-1 intersubtype recombination under a full-genome sequencing context in a rural community in Mbarara, Uganda, where HIV-1 subtypes A1 and D co-circulate. Methods: Near-full-genome HIV-1 Sanger sequence data was collected from plasma samples of 504 treatment-naïve individuals, who then received PI or NNRTI-containing regimens and were monitored for up to 7.5 years. Subtypes were inferred by Los Alamos RIP 3.0 and compared with Sanger/REGA and MiSeq/RIP. "Non-recombinants" and "recombinants" infections were compared in terms of pre-therapy viral load, CD4 count, post-therapy time to virologic suppression, virologic rebound, first CD4 rise above baseline and sustained CD4 recovery. Results: Prevalence of intersubtype recombinants varied depending on the genomic region examined: gag (15%), prrt (11%), int (8%), vif (10%), vpr (2%), vpu (9%), GP120 (8%), GP41 (18%), and nef (4%). Of the 200 patients with near-full-genome data, prevalence of intersubtype recombination was 46%; the most frequently observed recombinant was A1-D (25%). Sanger/REGA and MiSeq/RIP yielded generally consistent results. Phylogenetic tree revealed most recombinants did not share common ancestors. No temporal trend was observed (all p > 0.1). Subsequent subtype switches were detected in 27 of 143 (19%) subjects with follow-up sequences. Non-recombinant versus recombinants infections were not significantly different in any pre- nor post-therapy clinical correlates examined (all p > 0.2). Conclusion: Intersubtype recombination was highly prevalent (46%) in Uganda if the entire HIV genome was considered, but was not associated with clinical correlates nor therapy outcomes.
Five near full-length genome of novel second-generation HIV-1 recombinant virus (JS150021, JS150029, JS150129, JS150132 and AH150183) were identified from five HIV-positive people in Jiangsu and Anhui province, east China. Phylogenic analyses showed that these five sequences are all composed of two well-established circulating recombinant forms CRF07_BC and CRF01_AE, grouped into four new discovered recombinant forms ,which show several very similar but not identical recombinant breakpoints. The four recombinant forms are also identified to be a sort of family or related viruses, seems to be the results of different recombination events. The emergence of a serious new closed related CRF07_BC/CRF01_AE recombinant strain indicate the increasing complexity sexual transmission of the HIV-1 epidemic in China.
Human Immunodeficiency Virus type 1 (HIV-1) is characterised by vast genetic diversity. Globally circulating HIV-1 viruses are classified into distinct phylogenetic strains (subtypes, sub-subtypes) and several recombinant forms. Here we describe the characteristics and evolution of European HIV-1 epidemic over time through a review of published literature and updated queries of existing HIV-1 sequence databases. HIV-1 in Western and Central Europe was introduced in the early-1980s in the form of subtype B, which is still the predominant clade. However, in Eastern Europe (Former Soviet Union (FSU) countries and Russia) the predominant strain, introduced into Ukraine in the mid-1990s, is subtype A (AFSU) with transmission mostly occurring in people who inject drugs (PWID). In recent years, the epidemic is evolving towards a complex tapestry with an increase in the prevalence of non-B subtypes and recombinants in Western and Central Europe. Non-B epidemics are mainly associated with immigrants, heterosexuals and females but more recently, non-B clades have also spread amongst groups where non-B strains were previously absent - non-immigrant European populations and amongst men having sex with men (MSM). In some countries, non-B clades have spread amongst the native population, for example subtype G in Portugal and subtype A in Greece, Albania and Cyprus. Romania provides a unique case where sub-subtype F1 has predominated throughout the epidemic. In contrast, HIV-1 epidemic in FSU countries remains more homogeneous with AFSU clade predominating in all countries. The differences between the evolution of the Western epidemic and the Eastern epidemic may be attributable to differences in transmission risk behaviours, lifestyle and the patterns of human mobility. The study of HIV-1 epidemic diversity provides a useful tool by which we can understand the history of the pandemic in addition to allowing us to monitor the spread and growth of the epidemic over time.