Detection of human papillomavirus genotypes, herpes simplex, varicella zoster and cytomegalovirus in breast cancer patients.

BACKGROUND: The role of viruses as a cause of breast cancer (BC) has been significantly investigated in recent years. Human papillomavirus (HPV) has been detected in invasive breast carcinomas, while most studies have only focused on the detection of viral DNA, we aimed to examine the prevalence and genotypes of HPV among Iranian BC patients. We also examined the presence of herpes simplex-1 (HSV-1), herpes simplex-2 (HSV-2), varicella zoster virus (VZV), and cytomegalovirus (CMV) in these samples. METHODS: We collected and analyzed 70 Formalin-Fixed Paraffin-Embedded (FFPE) blocks including 59 BC samples, and 11 benign breast lesions as control from Iranian patients using nested PCR. Real-time PCR utilized as a confirming test to nested PCR findings. Genotyping of HPV positive samples was performed, the samples were also subjected to a multiplex PCR to detect HSV-1, HSV-2, VZV, and CMV in BC. RESULTS: Papillomavirus DNA was present in 7 of 59 BC samples (11.8%); while none was detected in control samples. The most prevalent type was HPV18, followed by HPV 6. All HPV positive patients had high tumor grades (II/ III) with a histologic diagnosis of ductal carcinoma. The patient age range was 33 to 73 years with a median of 51 years. Most of HPV positive patients had low levels of education. HPV16 was not detected. Also, 5 of 59 BC specimens (8.47%), were positive for HSV-1. But none of the samples were positive for HSV-2, VZV, and CMV. CONCLUSIONS: Our results suggest a carcinogenesis role for High-risk HPV (HPV18) in breast tumors. Our findings of HSV-1 and low-risk HPV (HPV6) in BCs may propose a cancer-causing role for them. Further large-scale studies are warranted to assess the significance of our findings.

Bovine leukemia virus detected in the breast tissue and blood of Iranian women.

BACKGROUND: Breast cancer is one of the most common cancers in the world particularly among Iranian women. Bovine leukemia virus (BLV) is an enzootic, exogenous, and oncogenic retrovirus that causes B-cell leukosis in 1-5% of infected cattle. The current study aimed at evaluating the correlation between BLV infection and breast cancer in an Iranian population. MATERIALS AND TECHNIQUES: A total of 400 samples including 200 breast cancer-suspected tissue samples and 200 blood samples of women without breast cancer, were collected from July 2017 to October 2018 from women referred to two general hospitals in Qom Province, Iran. The nested PCR technique was performed to determine the presence of tax and gag gene of BLV in the collected samples. RESULTS: Out of 200 breast cancer-suspected tissue samples, 172 samples were malignant in terms of pathology. Other samples were reported as non-malignant and non-tumor. Based on nested PCR technique, tax and gag genes of BLV were detected in 30% and 8% of breast cancer-suspected tissue samples, respectively. The frequency of BLV in blood samples collected from women without breast cancer was 16.5% (33/200). CONCLUSION: It seems that human breast cancer and BLV infection in cattle could be associated using nested PCR technique.

Detection and identification of mouse mammary tumor virus-like DNA sequences in blood and breast tissues of breast cancer patients.

Mouse mammary tumor virus (MMTV) is a well-known cause of mammary tumors in mice transmitted as endogenous proviruses or exogenously as infectious virions. The hypothesis that a retrovirus homologous to MMTV is involved in human breast cancers has resulted in renewed interest in the etiology of human breast cancer. Therefore, the detection of MMTV-like exogenous sequences in 30-40 % of invasive breast cancer has increased attention towards this hypothesis. To detect the prevalence of MMTV in Pakistani population, 666-bp-long MMTV envelop and 630-bp LTR sequences were amplified from breast cancer patient samples (tissue biopsies and peripheral blood) using mouse with mammary tumor as control. MMTV-like virus env and LTR DNA sequences were detected in 20 and 26 % of breast tumor samples, respectively, from the total of 80 breast cancer patients' blood and tissue samples. No significant association was observed between age, grade of disease, and lymph node involvement with the prevalence of MMTV-like sequences. Our data add to the growing number of studies implicating MMTV-like virus in human breast cancer, but still clear causal association of MMTV to breast cancer remains to be reputable.

Understanding HIV compartments and reservoirs.

The spectrum of HIV-1 cellular reservoirs is highly diversified, and their role varies according to the milieu of the anatomical sites in which the virus replicates. In this light, mechanisms underlying HIV-1 persistence in anatomical compartments may be profoundly different from what is observed in peripheral blood. This scenario is further complicated by sub-optimal drug penetration in tissues allowing persistent and cryptic HIV-1 replication in body districts despite undetectable viremia. On this basis, this review aims at providing recent insights regarding the critical role of HIV-1 cellular reservoirs in different anatomical compartments, and their relationship with the pathogenesis of HIV-1 infection. A comprehensive definition of the complex interplay between the virus and its reservoir is critical in order to set up prophylactic and therapeutic strategies aimed at achieving the maximal virological suppression and hopefully in the near future the cure of HIV-1 infection (either functional or biological).

Limited evidence of human papillomavirus in [corrected] breast tissue using molecular in situ methods.

BACKGROUND: Human papillomavirus (HPV) has been proposed as an etiologic agent of breast cancer based on numerous reports of high-risk (oncogenic) HPV types in malignant breast tissues. However, most of those studies used standard and nested solution polymerase chain reaction (PCR) techniques, both of which are disadvantaged by vulnerability to laboratory contamination from positive control DNA and the inability to localize the signal to a specific cell type. To overcome these drawbacks, the authors of this report explored the use of in situ molecular methods of viral detection to reassess the frequency of HPV in malignant breast tissue. METHODS: In situ hybridization (ISH) was used with probes that were specific for the capsid region of 12 oncogenic HPV types, and in situ PCR (IS-PCR) was used with primers that were specific for the capsid region of HPV-16, which is the most common oncogenic HPV type. These methods were resistant to molecular contamination and allowed identification of the positive cell type. The specimens examined were malignant tissues from patients with 70 breast cancer patients at The University of Texas M. D. Anderson Cancer Center in Houston, Texas. RESULTS: HPV was observed in 4 of 70 specimens (5.7%) using ISH and in 2 of 70 specimens (2.9%) of specimens using IS-PCR. Concordance between the 2 methods was high for negative specimens; both methods yielded negative results in 66 of 70 specimens (94.3%). However, there was no concordance for the few positive specimens, probably because of differences in sensitivity and the targeted HPV types. CONCLUSIONS: Oncogenic (high-risk) HPV types were present in malignant breast epithelium very infrequently and, thus, may be causative agents of only a relatively small proportion of all breast cancers.

Primary human mammary epithelial cells endocytose HIV-1 and facilitate viral infection of CD4+ T lymphocytes.

The contribution of mammary epithelial cells (MEC) to human immunodeficiency virus type 1 (HIV-1) in breast milk remains largely unknown. While breast milk contains CD4(+) cells throughout the breast-feeding period, it is not known whether MEC directly support HIV-1 infection or facilitate infection of CD4(+) cells in the breast compartment. This study evaluated primary human MEC for direct infection with HIV-1 and for indirect transfer of infection to CD4(+) target cells. Primary human MEC were isolated and assessed for expression of HIV-1 receptors. MEC were exposed to CCR5-, CXCR4- and dual-tropic strains of HIV-1 and evaluated for viral reverse transcription and integration and productive viral infection. MEC were also tested for the ability to transfer HIV to CD4(+) target cells and to activate resting CD4(+) T cells. Our results demonstrate that MEC express HIV-1 receptor proteins CD4, CCR5, CXCR4, and galactosyl ceramide (GalCer). While no evidence for direct infection of MEC was found, HIV-1 virions were observed in MEC endosomal compartments. Coculture of HIV-exposed MEC resulted in productive infection of activated CD4(+) T cells. In addition, MEC secretions increased HIV-1 replication and proliferation of infected target cells. Overall, our results indicate that MEC are capable of endosomal uptake of HIV-1 and can facilitate virus infection and replication in CD4(+) target cells. These findings suggest that MEC may serve as a viral reservoir for HIV-1 and may enhance infection of CD4(+) T lymphocytes in vivo.

High prevalence of human papillomaviruses in fresh frozen breast cancer samples.

While the etiology of breast cancer remains enigmatic, some recent reports have examined the role of human papillomavirus (HPV) in breast carcinogenesis. The purpose of this study was to determine the prevalence of HPV in breast cancer tissue using PCR analysis and sequencing. Fifty-four (54) fresh frozen breast cancers samples that were removed from a cohort of breast cancer patients were analyzed. Samples were tested for HPV using comprehensive PCR primers, and in situ hybridization was performed on paraffin embedded tissue sections. Findings were correlated with clinical and pathological characteristics. The HPV DNA prevalence in the breast cancer samples was 50% (27/54) with sequence analysis indicating all cases to be positive for HPV-18 type. While HPV patients were slightly younger, no correlation was noted for menopausal status or family history. HPV positive tumors were smaller with earlier T staging and demonstrated lesser nodal involvement compared to HPV negative cancers. In situ hybridization analyses proved negative. The high proportion of HPV positive breast cancers detected in this series using fresh frozen tissues cannot be dismissed, however the role of HPV in breast carcinogenesis remains unclear and may ultimately be ascertained by monitoring future breast cancer incidence amongst women vaccinated against high risk HPV types.

Risk factor for breast cancer development under exposure to bovine leukemia virus in Colombian women: A case-control study.

Viruses have been implicated in cancer development in both humans and animals. The role of viruses in cancer is typically to initiate cellular transformation through cellular DNA damage, although specific mechanisms remain unknown. Silent and long-term viral infections need to be present, in order to initiate cancer disease. In efforts to establish a causative role of viruses, first is needed to demonstrate the strength and consistency of associations in different populations. The aim of this study was to determine the association of bovine leukemia virus (BLV), a causative agent of leukemia in cattle, with breast cancer and its biomarkers used as prognosis of the severity of the disease (Ki67, HER2, hormonal receptors) in Colombian women. An unmatched, observational case-control study was conducted among women undergoing breast surgery between 2016-2018. Malignant samples (n = 75) were considered as cases and benign samples (n = 83) as controls. Nested-liquid PCR, in-situ PCR and immunohistochemistry were used for viral detection in blood and breast tissues. For the risk assessment, only BLV positive samples from breast tissues were included in the analysis. BLV was higher in cases group (61.3%) compared with controls (48.2%), with a statistically significant association between the virus and breast cancer in the unconditional logistic regression (adjusted-OR = 2.450,95%CI:1.088-5.517, p = 0.031). In this study, BLV was found in both blood and breast tissues of participants and an association between breast cancer and the virus was confirmed in Colombia, as an intermediate risk factor.

Distinct Oncogenic Transcriptomes in Human Mammary Epithelial Cells Infected With Cytomegalovirus.

Human cytomegalovirus is being recognized as a potential oncovirus beside its oncomodulation role. We previously isolated two clinical isolates, HCMV-DB (KT959235) and HCMV-BL (MW980585), which in primary human mammary epithelial cells promoted oncogenic molecular pathways, established anchorage-independent growth in vitro, and produced tumorigenicity in mice models, therefore named high-risk oncogenic strains. In contrast, other clinical HCMV strains such as HCMV-FS, KM, and SC did not trigger such traits, therefore named low-risk oncogenic strains. In this study, we compared high-risk oncogenic HCMV-DB and BL strains (high-risk) with low-risk oncogenic strains HCMV-FS, KM, and SC (low-risk) additionally to the prototypic HCMV-TB40/E, knowing that all strains infect HMECs in vitro. Numerous pro-oncogenic features including enhanced expression of oncogenes, cell survival, proliferation, and epithelial-mesenchymal transition genes were observed with HCMV-BL. In vitro, mammosphere formation was observed only in high-risk strains. HCMV-TB40/E showed an intermediate transcriptome landscape with limited mammosphere formation. Since we observed that Ki67 gene expression allows us to discriminate between high and low-risk HCMV strains in vitro, we further tested its expression in vivo. Among HCMV-positive breast cancer biopsies, we only detected high expression of the Ki67 gene in basal tumors which may correspond to the presence of high-risk HCMV strains within tumors. Altogether, the transcriptome of HMECs infected with HCMV clinical isolates displays an "oncogenic gradient" where high-risk strains specifically induce a prooncogenic environment which might participate in breast cancer development.

Characterization of SARS-CoV-2 RNA, Antibodies, and Neutralizing Capacity in Milk Produced by Women with COVID-19.

Whether mother-to-infant SARS-CoV-2 transmission can occur during breastfeeding and, if so, whether the benefits of breastfeeding outweigh this risk during maternal COVID-19 illness remain important questions. Using RT-qPCR, we did not detect SARS-CoV-2 RNA in any milk sample (n = 37) collected from 18 women following COVID-19 diagnosis. Although we detected evidence of viral RNA on 8 out of 70 breast skin swabs, only one was considered a conclusive positive result. In contrast, 76% of the milk samples collected from women with COVID-19 contained SARS-CoV-2-specific IgA, and 80% had SARS-CoV-2-specific IgG. In addition, 62% of the milk samples were able to neutralize SARS-CoV-2 infectivity in vitro, whereas milk samples collected prior to the COVID-19 pandemic were unable to do so. Taken together, our data do not support mother-to-infant transmission of SARS-CoV-2 via milk. Importantly, milk produced by infected mothers is a beneficial source of anti-SARS-CoV-2 IgA and IgG and neutralizes SARS-CoV-2 activity. These results support recommendations to continue breastfeeding during mild-to-moderate maternal COVID-19 illness.IMPORTANCE Results from prior studies assaying human milk for the presence of SARS-CoV-2, the causative virus of COVID-19, have suggested milk may act as a potential vehicle for mother-to-child transmission. Most previous studies are limited because they followed only a few participants, were cross-sectional, and/or failed to report how milk was collected and/or analyzed. As such, considerable uncertainty remains regarding whether human milk is capable of transmitting SARS-CoV-2 from mother to child. Here, we report that repeated milk samples collected from 18 women following COVID-19 diagnosis did not contain SARS-CoV-2 RNA; however, risk of transmission via breast skin should be further evaluated. Importantly, we found that milk produced by infected mothers is a source of anti-SARS-CoV-2 IgA and IgG and neutralizes SARS-CoV-2 activity. These results support recommendations to continue breastfeeding during mild-to-moderate maternal COVID-19 illness as milk likely provides specific immunologic benefits to infants.

Local replication of simian immunodeficiency virus in the breast milk compartment of chronically-infected, lactating rhesus monkeys.

Breast milk transmission remains a major mode of infant HIV acquisition, yet anatomic and immunologic forces shaping virus quasispecies in milk are not well characterized. In this study, phylogenic analysis of envelope sequences of milk SIV variants revealed groups of nearly identical viruses, indicating local virus production. However, comparison of the patterns and rates of CTL escape of blood and milk virus demonstrated only subtle differences between the compartments. These findings suggest that a substantial fraction of milk viruses are produced by locally-infected cells, but are shaped by cellular immune pressures similar to that in the blood.

Progress in research on the detection of the novel coronavirus in human samples of different groups.

OBJECTIVE: Among the illnesses that may develop from COVID-19, the disease caused by the novel coronavirus (SARS-CoV-2), is pneumonia, a severe acute respiratory infectious disease. SARS-CoV-2 continues to spread worldwide and has caused hundreds of thousands of deaths thus far and has disrupted the world economy. PATIENTS AND METHODS: This review summarized the reported distributions of SARS-CoV-2 in 13 biological samples of the human body, including nose, feces, sperm, tears, breast milk, cerebrospinal fluid, urine, organs, sputum, cell lines, bronchial brush, blood, throat, and bronchoalveolar lavage fluid. Moreover, this review briefly describes the detection of SARS-CoV-2 in human body samples of five other coronaviruses. CONCLUSIONS: This review offers several recommendations for controlling the spread of SARS-CoV-2 control, specifically, sample collection from suspected cases from foreign countries and risk assessment of imported special goods (biological materials).

Female reproductive tract has low concentration of SARS-CoV2 receptors.

There has been significant concern regarding fertility and reproductive outcomes during the SARS-CoV2 pandemic. Recent data suggests a high concentration of SARS-Cov2 receptors, ACE2 or TMPRSS2, in nasal epithelium and cornea, which explains person-to-person transmission. We investigated the prevalence of SARS-CoV2 receptors among reproductive tissues by exploring the single-cell sequencing datasets from uterus, myometrium, ovary, fallopian tube, and breast epithelium. We did not detect significant expression of either ACE2 or TMPRSS2 in the normal human myometrium, uterus, ovaries, fallopian tube, or breast. Furthermore, none of the cell types in the female reproductive organs we investigated, showed the co-expression of ACE2 with proteases, TMPRSS2, Cathepsin B (CTSB), and Cathepsin L (CTSL) known to facilitate the entry of SARS2-CoV2 into the host cell. These results suggest that myometrium, uterus, ovaries, fallopian tube, and breast are unlikely to be susceptible to infection by SARS-CoV2.

Koilocytes indicate a role for human papilloma virus in breast cancer.

BACKGROUND: High-risk human papilloma viruses (HPVs) are candidates as causal viruses in breast cancer. The scientific challenge is to determine whether HPVs are causal and not merely passengers or parasites. Studies of HPV-related koilocytes in breast cancer offer an opportunity to address this crucial issue. Koilocytes are epithelial cells characterised by perinuclear haloes surrounding condensed nuclei and are commonly present in cervical intraepithelial neoplasia. Koilocytosis is accepted as pathognomonic (characteristic of a particular disease) of HPV infection. The aim of this investigation is to determine whether putative koilocytes in normal and malignant breast tissues are because of HPV infection. METHODS: Archival formalin-fixed normal and malignant breast specimens were investigated by histology, in situ PCR with confirmation of the findings by standard PCR and sequencing of the products, plus immunohistochemistry to identify HPV E6 oncoproteins. RESULTS: human papilloma virus-associated koilocytes were present in normal breast skin and lobules and in the breast skin and cancer tissue of patients with ductal carcinoma in situ (DCIS) and invasive ductal carcinomas (IDCs). INTERPRETATION: As koilocytes are known to be the precursors of some HPV-associated cervical cancer, it follows that HPVs may be causally associated with breast cancer.

Low frequency of human papillomavirus DNA in breast cancer tissue.

Human papillomavirus (HPV) is considered the aetiological agent for cervical cancer. Several reports have addressed a relationship with HPV and breast cancer, as different HPVs have been identified. The purpose of this study was to detect HPV DNA in 67 breast cancer patients and 40 non-malignant disease breast tissues by means of Polymerase Chain Reaction with consensus primers. The frequency of HPV in the cases group were 4.4% (3/67) and no positive samples among the reference group were identified. From the 3 positive samples, HPV types 16, 18 and 33 were identified by restriction patterns and direct sequencing. The high diversity among detection in the related studies shows that population genomic heterogeneity plays an important role in the disease. The low frequency detected in the present study suggests that HPV does not play an important role in breast cancer.

Bovine leukemia virus DNA associated with breast cancer in women from South Brazil.

Breast cancer is a neoplastic condition with a high morbidity and mortality amongst women worldwide. Recent data linking bovine leukemia virus (BLV) with breast cancer has been contested already. Our study investigated the presence of BLV genome in healthy (n = 72) and cancerous (n = 72) paraffin-embedded samples of breast tissues from women in south Brazil. BLV DNA was found most frequently (30.5%) in breast cancer tissue than in healthy breast (13.9%) (Odds ratio = 2.73; confidence interval = 1.18-6.29; p = 0.027). In contrast, antibodies to BLV were found in a very small percentage of healthy blood donors. There was no association between BLV DNA and other tumor prognostic biological markers such as hormonal receptors, HER2 oncoprotein, proliferation index, metastasis in sentinels lymph nodes, and tumor grade and size. Our findings suggest that BLV should be considered a potential predisposing factor to breast cancer in women.

Mouse mammary tumor viral env sequences are not present in the human genome but are present in breast tumors and normal breast tissues.

In 1936, John Joseph Bittner identified mouse mammary tumor virus (MMTV), a milk transmitted beta retrovirus, a form of single-stranded positive-sense RNA virus. A retrovirus inserts a copy of its genome into the DNA of a host cell, thus altering the cell's genome. In the current analysis, we searched for MMTV sequences within the human genome. To compare the MMTV genome to the human genome, we used BLAT, the Blast-Like Alignment Tool of the UCSC Genome Browser. BLAT can align a user sequence of 25 bases or more to the genome. 60 MMTV sequences were in the human genome. Of 56 sequences from the MMTV POL gene, 36 POL sequences were from the same part of the gene, beginning at viral nucleotide 4800 but of different lengths. 8 viral sequences began at nucleotide ∼3430 of the POL gene. Four viral sequences were from GAGdUTPase, encoded by the MMTV PRO gene. Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) is an enzyme present in several major retroviral families. In MMTV dUTPase may be essential for viral replication. Since BLAT identified no MMTV envelope (env) sequence in the human genome, the env sequences from breast tumors and normal breast tissue found in other studies may have come from an MMTV infection. However, no one is certain how MMTV could enter human cells, since the cells do not have a cellular receptor for MMTV, as do mouse cells.

Hypothetic association between human papillomavirus infection and breast carcinoma.

High-risk human Papillomaviruses (HPVs) has been proved to be the major cause of cervical cancer. It has been considered that HPV may also cause squamous cell carcinomas of the other sites such as anus, vulva and esophagus. Furthermore, a number of studies have detected HPV DNA in breast carcinoma tissues. This raises the question that whether HPV plays a carcinogenic role in breast carcinomas. On the other hand, human Papillomaviruses do not seem to be able infect normal mammary cells in vitro, nor have HPV infections in human breast glands been observed among patients with AIDS. At present, there is no explanation for these "conflicting observations". In this paper, we propose the hypothesis that mammary epithelial cells that partly lose control in proliferation are more susceptible for persistent HPV infection. The potential role of HPV infection in the carcinogenic steps of breast cancer should be further tested. One possible cost-effective way for further investigation is to conduct a case-control study comparing the prevalence of previous HPV exposure to the breast, such as history of cervical HPV infections and HPV infections in nipples between cases and controls.

HPV infection and breast cancer. Results of a microarray approach.

OBJECTIVES: Human papilloma virus (HPV) has been implicated in several types of epithelial cancer. The role of HPV in breast carcinogenesis has been a matter of debate fueled by conflicting reports in recent years. The aim of this study is to identify the prevalence of breast and cervical HPV infection in cancer patients by using a modern microarray approach. MATERIALS AND METHODS: In the present prospective study, 201 breast cancer patients were included. For each patient a detailed medical history was taken and during the operation, under sterile conditions, samples were collected, from the tumour, the healthy adjacent breast tissue and any positive sentinel lymph nodes. In addition, for each patient a cervical sample was also collected. All samples were analysed for DNA of 24 types of HPV using a microarray technique. RESULTS: Despite the high sensitivity of the technique used, no HPV DNA was identified in any of the breast or lymph node samples. Our analysis showed that patients with HPV positive cervical samples (28 cases) were more likely to have tumors with positive progesterone receptors (p=0.041) and were also more likely to have two or three positive lymph nodes (p=0.002). CONCLUSION: In the present study, a combination of careful sample collection and a very sensitive microarray approach showed no correlation between HPV and breast cancer. However some characteristics of the breast tumors were different among patients with HPV DNA in their cervical samples.