Prevalence of SARS-CoV-2 infection in neonates born to mothers or relatives with COVID-19.

BACKGROUND: In December 2019, in Wuhan, China, coronavirus disease 2019 (COVID-19) was emerged due to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). It seems that children and neonates, similar to adult and elderly individuals, are at risk of SARS-CoV-2 infection. However, adequate data are not available about neonates infected with SARS-CoV-2. METHODS: This study evaluated the presence of SARS-CoV-2 infection in neonates born to mothers or relatives with COVID-19. This cross-sectional study was performed on 25,044 consecutive Iranian participants in Tehran, Iran, from January 2020 to August 2020. Viral ribonucleic acid (RNA) was extracted from 500 µl of the oropharyngeal and nasopharyngeal specimens of the participants. The genomic RNA of SARS-CoV-2 was detected by real-time polymerase chain reaction (PCR) assay. RESULTS: Out of all participants, 98 (0.40%) cases were neonates born to mothers or relatives with SARS-CoV-2 infection. Therefore, the current study was performed on these neonates. Out of 98 studied neonates, 6 (6.1%) cases had positive PCR results for SARS-CoV-2 infection. Moreover, among 98 studied neonates' mothers, 25 (25.5%) cases had positive PCR results for SARS-CoV-2 infection. CONCLUSION: The findings of this study demonstrated that the rate of COVID-19 in neonates born to mothers or relatives with SARS-CoV-2 infection in the Iranian population is about 6.1%.

Comparison of real-time PCR and nested PCR for toxoplasmosis diagnosis in toxoplasmic retinochoroiditis patients.

BACKGROUNDS: PCR is a proper technique that significantly improves toxoplasmosis diagnosis. However, a more sensitive technique is required. This study compared real-time PCR with nested PCR using B1, SAG-4, and MAG-1 bradyzoite genes to diagnose toxoplasmosis in toxoplasmic retinochoroiditis patients. METHODS: Blood samples were collected from 10 patients with active toxoplasmic chorioretinal lesions and 10 healthy individuals. Blood samples including peripheral blood mononuclear cells (PBMCs), serum and whole blood samples were used for DNA extraction. Serum was also used to detect anti-toxoplasma IgG and IgM antibodies. Nested PCR and real-time PCR were performed using B1, SAG-4, and MAG-1 target genes. RESULTS: Five (50%) out of the 10 patients were tested positive for toxoplasmosis with nested PCR using the PBMC samples. All the five patients tested positive with nested PCR were also tested positive for toxoplasmosis with real-time PCR using the PBMC samples. The real-time PCR results demonstrated that 9(90%) out of the 10 patients were positive based on B1 and the remaining one (10%) was positive only based on MAG-1. In general, of the patients, five (50%) were positive using SAG-4 and three (30%) were positive in term of MAG-1 using PBMCs with real-time PCR. CONCLUSION: It appears that PBMC samples have the best performance as the PCR extraction method and are a good source for toxoplasmosis diagnosis. The use of B22 and B23 target genes due to their high sensitivity and specificity along with bradyzoite genes are recommended for toxoplasmosis diagnosis using PBMC samples with real-time PCR.

Occult HCV and occult HBV coinfection in Iranian human immunodeficiency virus-infected individuals.

The presence of hepatitis C virus (HCV) genome in liver biopsy or peripheral blood mononuclear cell (PBMC) specimens in the absence of detectable HCV-RNA in plasma of the people with or without anti-HCV antibodies has defined as occult HCV infection (OCI), whereas occult hepatitis B virus infection (OBI) is detection of hepatitis B virus (HBV) genome in the absence of traceable hepatitis B surface antigen in the plasma samples of patients. The purpose of this study is to determine the presence of OBI and OCI in human immunodeficiency virus (HIV)-infected individuals. In this cross-sectional research, 190 Iranian HIV-infected individuals were enrolled from September 2015 to February 2019. All participants were tested regarding various serological markers for HCV and HBV infections. Viral RNA and DNA were extracted from plasma and PBMC specimens, and the presence of HCV-RNA in plasma and PBMC samples was tested using reverse transcriptase-nested polymerase chain reaction (PCR), HBV viral load was determined in plasma samples using COBAS TaqMan 48 Kit, and also the presence of the HBV-DNA in PBMC samples was tested by real-time PCR. In this study, the prevalence of OBI and OCI in HIV-infected individuals was 3.1% and 11.4%, respectively. The genotypes of HCV in the patients with OCI were as follows: 57.1% were infected with subtype 3a, 35.7% were infected with subtype 1a, and 7.1% was infected with subtype 1b. It is noteworthy that in this study, two patients (1.1%) had OCI/OBI coinfections. The present study revealed that 1.1% of Iranian HIV-infected individuals had OBI and OCI at the same time. Therefore, it seems that designing prospective surveys to determine the presence of this coinfection in HIV-infected individuals is informative.

Evaluation of CCR5-Δ32 mutation among individuals with high risk behaviors, neonates born to HIV-1 infected mothers, HIV-1 infected individuals, and healthy people in an Iranian population.

One of the important genetic factors related to resistance to HIV-1 infection is the presence of the C-C chemokine receptor type 5 delta 32 (CCR5-Δ32) homozygous genotype (Δ32/Δ32). The aim of this study was to evaluate the CCR5-Δ32 mutation among individuals with high-risk behaviors, neonates born to HIV-1-infected mothers in the prevention of mother-to-child transmission (PMTCT) project, HIV-1-infected individuals, and healthy people. The frequency of the CCR5-Δ32 genotype was assessed in a cross-sectional survey carried out from March 2014 to March 2019 among four different groups of the Iranian population. Genomic DNA was extracted from peripheral blood mononuclear cells of 140 Iranian healthy people, 84 neonates born to HIV-1-infected mothers in the PMTCT project, 71 people with high-risk behaviors, and 76 HIV-1-infected individuals. The polymerase chain reaction method was used for the amplification of the CCR5 gene. The CCR5-Δ32 heterozygous deletion was detected in five (6.6%) HIV-1-infected individuals, four (4.7%) neonates born to HIV-1 positive mothers, two (1.4%) healthy people, and also three (4.2%) people with high-risk behaviors whereas the CCR5-Δ32 homozygous deletion was absent in all the groups (Fisher's exact test, P = .0242). The allele of CCR5-Δ32 homozygous was not detected in the four study groups, and no significant difference was seen in the frequency of the CCR5Δ32 heterozygous allele between HIV seropositive and seronegative individuals. Therefore, it seems that this allele alone cannot explain the natural resistance to HIV-1 infection and probably several mechanisms are responsible for these processes and it should be further investigated.

Presence of different hepatitis C virus genotypes in plasma and peripheral blood mononuclear cell samples of Iranian patients with HIV infection.

Due to the similar routes of transmission, individuals infected with the human immunodeficiency virus (HIV) may become infected with the hepatitis C virus (HCV) simultaneously. The aim of this study was to investigate the frequency of HCV co-infection in Iranian individuals with HIV infection, and to genotype HCV in plasma and PBMC specimens of these patients. From September 2015 to October 2016, a total of 140 Iranian individuals with HIV infection were enrolled in this cross-sectional study. The RNA from plasma and PBMC specimens was extracted, and genomic HCV-RNA was amplified using RT-nested PCR with primers that target 5'-UTR. The HCV genotyping used the RFLP technique. To confirm HCV genotype, 10 randomly selected HCV-positive samples were also submitted for sequencing. The mean age of patients was 35.7 ± 13.5 years (range: 1-66). Out of 140 patients, 62 (44.3 %) were positive for anti-HCV antibodies; among these, viral genomic RNA was detected in 34 (24.3%) and 39 (27.9%) of the plasma and PBMC specimens, respectively. The HCV genotyping showed a similar pattern of subtypes 1a (44% vs 46.2%), 3a (32.4% vs 33.3%), and 1b (17.6% vs 17.9%) in all sera and PBMC samples. It is noteworthy that the HCV genotypes in plasma and PBMC specimens of 6 HCV co-infected patients were not the same. This study reveals that HIV/HCV co-infection is high in Iranian patients (44.3%), especially in people who have high-risk factors (83.9%). Also, HIV/HCV co-infected individuals may have dissimilar HCV genotypes in their plasma and PBMC specimens.

Detection of HBV genome in the plasma and peripheral blood mononuclear cells of Iranian HBsAg negative patients with HIV infection: occult HBV infection.

The presence of hepatitis B virus (HBV) DNA in the absence of traceable hepatitis B surface antigen (HBsAg) in the plasma specimen of patients is defined as occult HBV infection (OBI). This study aimed to detect HBV-DNA in the plasma and peripheral blood mononuclear cells (PBMCs) of Iranian HBsAg negative patients with human immunodeficiency virus (HIV) infection. This cross-sectional study was conducted on 172 patients with HIV infection from September 2015 to August 2017. The patients were tested for serological parameters (HBsAg, HBcAb, HBeAg and HBeAb) against HBV infection. Moreover, they were tested for HBV viral load (using COBAS TaqMan 48 Kit, Roche, USA) in plasma and the presence of the HBV genome in PBMC specimens using real-time PCR. The mean age of the patients was 35.4 ± 13.4 years. Of the 172 studied patients, 109 (63.4%) were male. In this study, 151 (87.8%) patients were negative for HBsAg, 111 (64.5%) patients were negative for all HBV infection serological markers, 9 (5.2%) patients were only positive for HBsAg and 29 (16.9%) patients were only positive for HBcAb. Moreover, five (3.3%) patients with HBsAg negative had OBI (in the plasma sample of four patients and PBMC specimens of all five patients, HBV-DNA was detected). The present study revealed that 3.3% of the patients with HIV infection had occult HBV infection. Presumably, designing prospective studies to identify this infection in patients with HIV infection is informative and valuable.

Investigation of the effects of a prevention of mother-to-child HIV transmission program among Iranian neonates.

Human immunodeficiency virus (HIV) infection is mostly spreading in developing countries. One of the most important pathways of HIV infection in these nations is the vertical route, from mother to infant. Therefore, this study evaluated the effectiveness of the prevention of mother-to-child transmission (PMTCT) program for HIV among Iranian neonates born to HIV-positive mothers. A total of 54 neonates born to HIV-1 positive mothers, all of whom were in a PMTCT program for HIV, as per the Iranian guidelines, were enrolled in this descriptive cross sectional study from March 2014 to July 2017. After RNA extraction of a plasma specimen, HIV-1 viral load was tested by an Artus HIV-1 RG RT-PCR Kit. Out of 54 evaluated neonates, 32 (59.3%) were male. The mean age of the HIV-infected mothers was 30.1 ± 5.4 (range: 19-47) years, and 36 (66.7%) of the mothers were in the age group 26-34 years. In the present study, it was found that none of the neonates whose mothers had previously entered PMTCT programs had HIV. 15 children were found who were born to HIV-positive mothers who had not entered the PMTCT program. Three of these children were infected with HIV (CRF35_AD), and none of them carried HIV-1 variants with SDRMs. The results of this study indicate that if HIV-positive pregnant women enter the PMTCT program for HIV, they can realistically hope to give birth to a non-infected child.

HIV-1 genetic diversity and transmitted drug resistance frequency among Iranian treatment-naive, sexually infected individuals.

In recent years, the patterns of human immunodeficiency virus 1 (HIV-1) transmission in Iran have been changing gradually from drug injection to unprotected sexual contact. This study sought to investigate the phylogenetic trends and characteristics of transmitted drug resistance (TDR) mutations of HIV-1 in a population that is mainly infected through homo/heterosexual contacts. Sixty newly diagnosed antiretroviral-naive individuals with HIV infection living in Tehran were recruited to this survey, and among them, 42 subjects were established to be infected through sexual intercourse. Following amplification and sequencing of the main part of the HIV-1 pol region, phylogenetic and drug-resistance mutation (DRM) analysis was successfully performed on these 42 patients. Phylogenetic analysis showed that the majority of the subjects were infected with subtype CRF35_AD (88%), followed by subtype B, with 7.1%, and subtype CRF01_AE, with 4.7%. A total of 7.1% of the subjects were found to be infected with HIV-1 variants with surveillance drug-resistant mutations (SDRMs) according to the last world health organisation (WHO) algorithm. All of the identified SDRMs belonged to the non-nucleoside reverse transcriptase inhibitors (NNRTIs) class, including K103 N and V106A, which were found in three patients. Two minor HIV protease-inhibitor-related mutations (L10I and G73S) were detected in two patients, but these mutations are not included in the WHO SDRMs list. The dominance of HIV-1 subtype CRF35_AD was observed among subjects of this study who were infected through sexual contact. The moderate prevalence of SDRMs (7.1%) in this population emphasises the fact that the risk of treatment failure in HIV-infected individuals might increase in the future, and preventive measures should be considered by health authorities.

Evaluation of the Expression Pattern of 4 microRNAs and their Correlation with Cellular/viral Factors in PBMCs of Long Term Non-progressors and HIV Infected Naïve Individuals.

BACKGROUND: Long-term non-progressors (LTNPs) are small subsets of HIV-infected subjects that can control HIV-1 replication for several years without receiving ART. The exact mechanism of HIV-1 suppression has not yet been completely elucidated. Although the modulatory role of microRNAs (miRNAs) in HIV-1 replication has been reported, their importance in LTNPs is unclear. OBJECTIVE: The aim of this cross-sectional study was to assess the expression pattern of miR-27b, -29, -150, and -221, as well as their relationship with CD4+ T-cell count, HIV-1 viral load, and nef gene expression in peripheral blood mononuclear cells (PBMCs) of untreated viremic patients and in LTNPs. METHODS: MiRNAs expression levels were evaluated with real-time PCR assay using RNA isolated from PBMCs of LTNPs, HIV-1 infected naive patients, and healthy people. Moreover, CD4 T-cell count, HIV viral load, and nef gene expression were assessed. RESULTS: The expression level of all miRNAs significantly decreased in the HIV-1 patient group compared to the control group, while the expression pattern of miRNAs in the LNTPs group was similar to that in the healthy subject group. In addition, there were significant correlations between some miRNA expression with viral load, CD4+ T-cell count, and nef gene expression. CONCLUSION: The significant similarity and difference of the miRNA expression pattern between LNTPs and healthy individuals as well as between elite controllers and HIV-infected patients, respectively, showed that these miRNAs could be used as diagnostic biomarkers. Further, positive and negative correlations between miRNAs expression and viral/cellular factors could justify the role of these miRNAs in HIV-1 disease monitoring.

The frequency of HIV-1 infection and surveillance drug-resistant mutations determination among Iranians with high-risk behaviors, during 2014 to 2020.

BACKGROUND AND OBJECTIVES: Human immunodeficiency virus (HIV) has various transmission routes. Instant antiretroviral therapy (ART) is the recommended treatment for HIV infection. Highly active antiretroviral therapy (HAART) significantly decreases the acquired immunodeficiency syndrome (AIDS) and AIDS-related co-morbidities. Notwithstanding the suitability of HAART, the antiretrovirals (ARVs) have adverse effects and antiretroviral drug resistance mutations are reported among those who receive ARVs. In this survey, the abundance of HIV-1 infection in Iranians with high-risk behaviors, and detection of the surveillance drug-resistant mutations (SDRMs) were evaluated. MATERIALS AND METHODS: This cross-sectional study was conducted on 250 individuals with high-risk behaviors from September 2014 to February 2020. HIV-1 Ag/Ab in plasma samples was detected using enzyme immunoassay (EIA) kits. The conserved region of HIV-1 was detected in the plasma samples by real-time polymerase chain reaction (PCR) assay. Furthermore, in individuals with positive HIV-1 RNA, HIV-1 viral load testing was performed. After amplification and sequencing of the HIV-1 protease, reverse transcriptase, and integrase genes, surveillance drug resistance mutation (SDRM) and phylogenetic analysis were determined. RESULTS: Out of the 250 participants with high-risk behaviors, six (2.4%) were infected with HIV-1. According to the phylogenetic analysis, the CRF35_AD (83.3% or 5/6) was the dominant subtype, followed by CRF01_AE (16.7% or 1/6). In this research, in none of the HIV-1 infected patients, SDRM for protease inhibitors (PIs), nucleoside reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), and integrase inhibitors (INs) were observed. Nevertheless, in one of the patients, V179L mutation was detected which is a rare non-polymorphic mutation and is listed as a rilpivirine (RPV) -associated resistance mutation. CONCLUSION: The results of the current survey revealed that 2.4% of people with high-risk behaviors are infected with HIV and the level of drug resistance mutations (DRMs) in these people is very low.

Detection of Toxoplasma gondii bradyzoite genes in the peripheral blood mononuclear cells among patients with toxoplasmic chorioretinitis.

BACKGROUND: Toxoplasmic chorioretinitis may occur as a result of acquired toxoplasmosis or reactivated congenital toxoplasmosis. In this study, Toxoplasma gondii bradyzoite genes along with the B1 gene were evaluated to detect T. gondii DNA in serum and peripheral blood mononuclear cells (PBMCs) of patients with toxoplasmic chorioretinitis. METHODS: Blood samples were collected from 10 patients (7 cases of active chorioretinal lesions and 3 cases of old chorioretinal scars). The genomic DNA was extracted from the patients' serum and PBMCs and a polymerase chain reaction (PCR) assay was performed using bradyzoite genes along with B1. The subjects were also evaluated in terms of the T. gondii antibodies. RESULTS: The PCR results were positive in four of seven patients (57.1%) with active ocular toxoplasmosis lesions. In three patients (42.8%), the PCR results were positive for MAG-1 and SAG-4 and in one patient (14.3%) the PCR results were only positive for the B1 gene. The PCR results were positive only in the PBMCs, whereas they were negative in the serum samples. Two patients with positive PCR results showed high Toxoplasma immunoglobulin G (IgG) antibody titres. However, none of the patients showed positive Toxoplasma IgM antibodies. CONCLUSIONS: The PBMCs are suitable for evaluating toxoplasmic chorioretinitis. The present results showed that PCR with bradyzoite genes is useful in the diagnosis of toxoplasmic chorioretinitis in PBMCs.

Acute and post-acute phase of COVID-19: Analyzing expression patterns of miRNA-29a-3p, 146a-3p, 155-5p, and let-7b-3p in PBMC.

BACKGROUND: When a new pathogen, such as severe acute respiratory syndrome coronavirus 2, appears all novel information can aid in the process of monitoring and in the diagnosis of the coronavirus disease (COVID-19). The aim of the current study is to elucidate the specific miRNA profile which can act as new biomarkers for distinguishing acute COVID-19 disease from the healthy group and those in the post-acute phase of the COVID-19 disease. METHODS: The expression level of selected miRNAs including let-7b-3p, miR-29a-3p, miR-146a-3p and miR-155-5p were evaluated in peripheral blood mononuclear cells (PBMCs) of COVID-19 patients, in both the acute and post-acute COVID-19 phase of the disease and healthy groups, by real-time PCR assays. Specificity and sensitivity of miRNAs was tested by receiver operating characteristic (ROC) analysis in COVID-19 patients. RESULTS: The expression level of all miRNAs in COVID-19 patients was significantly higher than in the healthy group. Therefore, the expression pattern of miR-29a-3p, miR-146a-3p and let-7b-3p in the post-acute COVID-19 phase was significantly different from the acute COVID-19 phase. ROC analyses demonstrated that miR-29a-3p, -155-5p and -146a-3p may serve as the novel biomarker for COVID-19 diagnosis with high specificity and sensitivity. In addition, miR-29a-3p, and -146a-3p can maybe act as novel biomarkers for distinguishing acute from post-acute phase of COVID-19 disease. DISCUSSION: The difference in miRNA expression pattern between COVID-19 patients and those in the healthy group, and between acute COVID-19 with post-acute COVID-19, suggested that cellular miRNAs could be used as promising biomarkers for diagnosis and monitoring of COVID-19.

Evaluation of a PCR assay for diagnosis of toxoplasmosis in serum and peripheral blood mononuclear cell among HIV/AIDS patients.

Cerebral toxoplasmosis is one of the neurological infections with high morbidity and mortality in patients with AIDS, so the accurate method for diagnosis of cerebral toxoplasmosis seems necessary. In this study, nested PCR assay using B1 gene was evaluated in diagnosis of toxoplasmosis in serum and peripheral blood mononuclear cell (PBMC) among HIV/AIDS patients. One hundred eight blood samples from HIV/AIDS patients, including four patients with cerebral toxoplasmosis and 104 HIV/AIDS patients without cerebral toxoplasmosis were evaluated for the Toxoplasma gondii antibodies using Enzyme Linked immunosorbent Assay. DNA of serum and PBMC of these patients were extracted and nested-PCR was carried out. Of 108 participants, 95 cases (88%) were positive for Toxoplasma IgG antibodies and one patient was found positive for Toxoplasma IgM antibody. In general, four patients, including three patients with cerebral toxoplasmosis, who were positive for Toxoplasma IgG antibodies and one patient without cerebral toxoplasmosis who was positive for Toxoplasma IgM antibody were found to be PCR positive. DNA of T. gondii was detected in both serum and PBMC in two cerebral toxoplasmosis patients; however DNA was detected in only PBMC in other cerebral toxoplasmosis patient. All cases with cerebral toxoplasmosis were also diagnosed by clinical and radiological manifestations. The results of this study showed that the numbers of positive samples by PCR in PBMC were higher than serum specimens for diagnosis of toxoplasmosis. If molecular method and immunological assay are complemented with magnetic resonance imaging, the results can be useful for diagnosis of cerebral toxoplasmosis.