Identifying SARS-CoV-2-related coronaviruses in Malayan pangolins.

The ongoing outbreak of viral pneumonia in China and across the world is associated with a new coronavirus, SARS-CoV-21. This outbreak has been tentatively associated with a seafood market in Wuhan, China, where the sale of wild animals may be the source of zoonotic infection2. Although bats are probable reservoir hosts for SARS-CoV-2, the identity of any intermediate host that may have facilitated transfer to humans is unknown. Here we report the identification of SARS-CoV-2-related coronaviruses in Malayan pangolins (Manis javanica) seized in anti-smuggling operations in southern China. Metagenomic sequencing identified pangolin-associated coronaviruses that belong to two sub-lineages of SARS-CoV-2-related coronaviruses, including one that exhibits strong similarity in the receptor-binding domain to SARS-CoV-2. The discovery of multiple lineages of pangolin coronavirus and their similarity to SARS-CoV-2 suggests that pangolins should be considered as possible hosts in the emergence of new coronaviruses and should be removed from wet markets to prevent zoonotic transmission.

Osthole exhibits the remedial potential for polycystic ovary syndrome mice through Nrf2-Foxo1-GSH-NF-κB pathway.

Polycystic ovary syndrome (PCOS) is the primary cause of female infertility with a lack of universal therapeutic regimen. Although osthole exhibits numerous pharmacological activities in treating various diseases, its therapeutic effect on PCOS is undiscovered. The present study found that application of osthole improved the symptoms of PCOS mice through preventing ovarian granulosa cells (GCs) production of more estrogen and alleviating the liberation of pro-inflammatory cytokine interleukin (IL)-1ß, IL-6, and tumor necrosis factor alpha. Meanwhile, osthole enhanced ovarian antioxidant capacity and alleviated intracellular reactive oxygen species (ROS) accumulation with a concurrent attenuation for oxidative stress, while intervention of antioxidant enzymic activity and glutathione (GSH) synthesis neutralized the salvation of osthole on GCs secretory disorder and chronic inflammation. Further analysis revealed that osthole restored the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and forkhead box O 1 (Foxo1) whose repression antagonized the amelioration of osthole on the insufficiency of antioxidant capacity and accumulation of ROS. Moreover, Nrf2 served as an intermedium to mediate the regulation of osthole on Foxo1. Additionally, osthole restricted the phosphorylation of IκBα and nuclear factor kappa B (NF-κB) subunit p65 by DHEA and weakened the transcriptional activity of NF-κB, but this effectiveness was abrogated by the obstruction of Nrf2 and Foxo1, whereas adjunction of GSH renewed the redemptive effect of osthole on NF-κB whose activation caused an invalidation of osthole in rescuing the aberration of GCs secretory function and inflammation response. Collectively, osthole might relieve the symptoms of PCOS mice via Nrf2-Foxo1-GSH-NF-κB pathway.

Melatonin modulates endometrial decidualization via NOTCH1-NRF2-FOXO1-GSH pathway†.

Melatonin is important for oocyte maturation, fertilization, early embryonic development, and embryo implantation, but less knowledge is available regarding its role in decidualization. The present study found that melatonin did not alter the proliferation of human endometrial stromal cells (ESCs), as well as cell cycle progress, but suppressed stromal differentiation after binding to the melatonin receptor 1B (MTNR1B), which was visualized in decidualizing ESCs. Further analysis evidenced that application of melatonin resulted in the diminishment for NOTCH1 and RBPJ expression. Supplementation of recombinant NOTCH1 protein (rNOTCH1) counteracted the impairment of stromal differentiation conferred by melatonin, while the addition of the NOTCH signaling pathway inhibitor DAPT aggravated the differentiation progress. Meanwhile, melatonin might restrain the expression and transcriptional activity of nuclear factor erythroid 2-related factor 2 (NRF2), whose blockage accelerated the fault of stromal differentiation under the context of melatonin, but this restraint was subsequently ameliorated by rNOTCH1. Forkhead box O 1 (FOXO1) was identified as a downstream target of melatonin in decidualization. Repression of NRF2 antagonized the retrieval of rNOTCH1 due to aberrant FOXO1 expression elicited by melatonin. Moreover, melatonin brought about the occurrence of oxidative stress accompanied by an obvious accumulation of intracellular reactive oxygen species and a significant reduction in glutathione (GSH) content, as well as enzymatic activities of glutathione peroxidase and glutathione reductase, whereas supplementation of rNOTCH1 improved the above-mentioned effects. Nevertheless, this improvement was disrupted by the blockage of NRF2 and FOXO1. Furthermore, addition of GSH rescued the defect of stromal differentiation by melatonin. Collectively, melatonin might impair endometrial decidualization by restraining the differentiation of ESCs dependent on NOTCH1-NRF2-FOXO1-GSH pathway after binding to the MTNR1B receptor.

Bmp2 regulates Serpinb6b expression via cAMP/PKA/Wnt4 pathway during uterine decidualization.

Serpinb6b is a novel member of Serpinb family and found in germ and somatic cells of mouse gonads, but its physiological function in uterine decidualization remains unclear. The present study revealed that abundant Serpinb6b was noted in decidual cells, and advanced the proliferation and differentiation of stromal cells, indicating a creative role of Serpinb6b in uterine decidualization. Further analysis found that Serpinb6b modulated the expression of Mmp2 and Mmp9. Meanwhile, Serpinb6b was identified as a target of Bmp2 regulation in stromal differentiation. Treatment with rBmp2 resulted in an accumulation of intracellular cAMP level whose function in this differentiation program was mediated by Serpinb6b. Addition of PKA inhibitor H89 impeded the Bmp2 induction of Serpinb6b, whereas 8-Br-cAMP rescued the defect of Serpinb6b expression elicited by Bmp2 knock-down. Attenuation of Serpinb6b greatly reduced the induction of constitutive Wnt4 activation on stromal cell differentiation. By contrast, overexpression of Serpinb6b prevented this inhibition of differentiation process by Wnt4 siRNA. Moreover, blockage of Wnt4 abrogated the up-regulation of cAMP on Serpinb6b. Collectively, Serpinb6b mediates uterine decidualization via Mmp2/9 in response to Bmp2/cAMP/PKA/Wnt4 pathway.

Hydroxysafflor yellow A exerts beneficial effects by restoring hormone secretion and alleviating oxidative stress in polycystic ovary syndrome mice.

NEW FINDINGS: What is the central question of this study? What are the potential therapeutic roles of ginsenoside Rb1 and hydroxysafflor yellow A (HSYA) in polycystic ovary syndrome (PCOS). What is the main finding and its importance? HSYA restored the oestrous cycles of PCOS mice, reduced follicular cysts in ovaries and rescued abnormal hormone secretion; ginsenoside Rb1 did not ameliorate the main symptoms of PCOS mice. HSYA alleviated oxidative stress along with an enhancement of antioxidant enzyme activity. This highlights a potential role of HSYA in PCOS therapy. ABSTRACT: Polycystic ovary syndrome (PCOS) is the most common endocrine disease resulting in female infertility. Hydroxysafflor yellow A (HSYA) and ginsenoside Rb1 have been shown to have antioxidant properties, but little is known about their impact in PCOS. Here dehydroepiandrosterone was used to induce PCOS in a mouse model that was characterized by an irregular oestrous cycle, cystic follicles and an elevated serum testosterone level. Supplementation of HSYA restored the oestrous cycle of PCOS mice, reduced follicular cysts in PCOS mouse ovaries and brought about a decline in serum testosterone level, while ginsenoside Rb1 did not ameliorate the above symptoms of PCOS mice. After HSYA treatment, there was elevation of serum oestradiol, progesterone, luteinizing hormone and anti-Müllerian hormone levels and a reduction of follicle-stimulating hormone level, but ginsenoside Rb1 only rescued the levels of follicle-stimulating hormone and anti-Müllerian hormone. Further analysis evidenced that HSYA reversed the expression of steroid hormone secretion-related genes Star, Hsd3b1, Cyp11a1 and Cyp19a1. In PCOS mice HSYA weakened the elevation of ovarian malondialdehyde, which is regarded as a biomarker for oxidative stress. Moreover, HSYA improved reduced glutathione content accompanied by a simultaneous increase in reduced to oxidized glutathione ratio, and enhanced the activities of the antioxidant enzymes superoxide dismutase, glutathione peroxidase and catalase. Collectively, HSYA exerted beneficial effects on PCOS mice by restoring hormone secretion and alleviating oxidative stress.

Genistein protects ovarian granulosa cells from oxidative stress via cAMP-PKA signaling.

Genistein is an isoflavone that has estrogen (E2 )-like activity and is beneficial for follicular development, but little is known regarding its function in oxidative stress (OS)-mediated granulosa cell (GC) injury. Here, we found that after exposure to H2 O2 , Genistein weakened the elevated levels of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA), which were regarded as the biomarkers for OS, and rescued glutathione (GSH) content and GSH/GSSG ratio accompanying with a simultaneous increase in cyclic adenosine monophosphate (cAMP) level, whereas addition of protein kinase A (PKA) inhibitor H89 impeded the effects of Genistein on the levels of ROS and MDA. Further analysis evidenced that Genistein enhanced the activities of antioxidant enzymes superoxide dismutase (SOD), GSH-peroxidase (GSH-Px), and catalase (CAT) in H2 O2 -treated GCs, but this enhancement was attenuated by H89. Under OS, Genistein improved cell viability and lessened the apoptotic rate of GCs along with a reduction in the activity of Casp3 and levels of Bax and Bad messenger RNA (mRNA), while H89 reversed the above effects. Moreover, Genistein treatment caused an obvious elevation in mitochondrial membrane potential (MMP) followed by a decline in the levels of intracellular mitochondrial superoxide, but H89 inhibited the regulation of Genistein on MMP and mitochondrial superoxide. Supplementation of Genistein promoted the secretion of E2 and increased the expression of Star and Cyp19a1 mRNA, whereas suppressed the level of progesterone (P4 ) accompanied with a decline in the level of Hsd3b1 mRNA expression. H89 blocked the regulation of Genistein on the secretion of E2 and P4 , and alleviated the ascending of Star and Cyp19a1 elicited by Genistein. Collectively, Genistein protects GCs from OS via cAMP-PKA signaling.

[Diagnosis of the first imported case of Plasmodium ovale infection at Guangdong Port].

The first imported case of Plasmodium ovale infection in Guangdong Province was identified. The patient worked in Myanmar for one week and had a fever when he arrived at Guangzhou Baiyun International Airport. Epidemiological information and blood sample were collected. The detection was conducted by microscopy, right VIEW rapid malaria test (RDTs) and real-time PCR with Plasmodium genus-specific and species-specific primers and probes. The case showed weak positive RDT result, and was confirmed as P. ovale infection by microscopy and real-time PCR. After treatment with artemether, his symptoms improved.

Puerarin Ameliorated PCOS through Preventing Mitochondrial Dysfunction Dependent on the Maintenance of Intracellular Calcium Homeostasis.

Polycystic ovarian syndrome (PCOS) is the major cause of infertility in reproductive women, but no universal drug is feasible. Although puerarin clinically treats cerebrovascular and cardiovascular diseases, its curative effect on PCOS remains elusive. The present study discovered that administration of puerarin restored estrous cycle of PCOS mice and diminished the number of cystic follicles with the concomitant recovery for circulating testosterone, LH and FSH levels, and LH/FSH ratio, indicating the therapeutic role of puerarin in PCOS. KEGG analysis of differential genes between PCOS and control revealed the enrichment in MAPK and calcium signaling pathway. Application of puerarin restricted the phosphorylation of ERK1/2 and JNK, whose activation neutralized the improvement of puerarin on the secretory function and apoptosis of ovarian granulosa cells (GCs). Meanwhile, puerarin alleviated the accumulation of cytosolic Ca2+ through restricting the opening of Ryr and Itpr channels, but this effectiveness was counteracted by the activatory ERK1/2 and JNK. Attenuation of cytosolic Ca2+ counteracted the antagonistic effects of ERK1/2 and JNK activation on puerarin's role in rescuing the calcineurin and Nfatc. Further analysis manifested that Mcu had been authenticated as a direct downstream target of Nfatc to mediate the amelioration of puerarin on mitochondrial Ca2+ uptake. Moreover, puerarin prevented the disorder of ATP content, mitochondrial membrane potential, and mitochondrial permeability transition pore opening through maintaining mitochondrial Ca2+ homeostasis. Collectively, puerarin might ameliorate the symptoms of PCOS mice through preventing mitochondrial dysfunction that is dependent on the maintenance of intracellular Ca2+ homeostasis after inactivation of ERK1/2 and JNK.

TAZ ameliorates the microglia-mediated inflammatory response via the Nrf2-ROS-NF-κB pathway.

Transcriptional co-activator with PDZ-binding motif (TAZ), one of core modules of the Hippo pathway, involves inflammatory cell infiltration in the liver, but little information is available regarding its physiological function in the microglia-mediated inflammatory response. Here we revealed that activation of TAZ prevented microglia production of proinflammatory cytokines, indicating TAZ's importance in anti-inflammation. After translocation into the nucleus, TAZ interacted with transcriptional enhanced associate domain (TEAD) and bound to the promoter of nuclear factor erythroid 2-related factor 2 (Nrf2), whose blockage caused inability of TAZ to improve inflammation, implying that Nrf2 is a direct target of TAZ. Further analysis showed that TAZ induced Nrf2 nuclear translocation to enhance antioxidant capacity with attenuation of oxidative stress and the inflammatory response. Under inflammatory conditions, TAZ impeded mitochondrial dysfunction, as indicated by amelioration of ATP levels, mtDNA copy numbers, and mitochondrial membrane potential with an obvious reduction in mitochondrial superoxide, but this impediment was neutralized by blockage of Nrf2. TAZ hindered opening of the mitochondrial permeability transition pore, restrained release of cytochrome c from mitochondria into the cytosol, and was sufficient to rescue microglia from apoptosis dependent on Nrf2. Nrf2 acted as a downstream target of TAZ to repress NF-κB activation by enhancing antioxidant capacity. Collectively, TAZ might ameliorate the microglia-mediated inflammatory response through the Nrf2-reactive oxygen species (ROS)-nuclear factor κB (NF-κB) pathway.

HB-EGF induces mitochondrial dysfunction via estrogen hypersecretion in granulosa cells dependent on cAMP-PKA-JNK/ERK-Ca2+-FOXO1 pathway.

Polycystic ovarian syndrome (PCOS) is one of the most prevalent endocrinopathies and the leading cause of anovulatory infertility, but its pathogenesis remains elusive. Although HB-EGF is involved in ovarian cancer progression, there is still no clarity about its relevance with PCOS. The present study exhibited that abundant HB-EGF was noted in follicular fluid from PCOS women, where it might induce the granulosa cells (GCs) production of more estrogen via the elevation of CYP19A1 expression after binding to EGFR. Furthermore, HB-EGF transduced intracellular downstream cAMP-PKA signaling to promote the phosphorylation of JNK and ERK whose blockage impeded the induction of HB-EGF on estrogen secretion. Meanwhile, HB-EGF enhanced the accumulation of intracellular Ca2+ whose chelation by BAPTA-AM abrogated the stimulation of HB-EGF on FOXO1 along with an obvious diminishment for estrogen production. cAMP-PKA-JNK/ERK-Ca2+ pathway played an important role in the crosstalk between HB-EGF and FOXO1. Treatment of GCs with HB-EGF resulted in mitochondrial dysfunction as evinced by the reduction of ATP content, mtDNA copy number and mitochondrial membrane potential. Additionally, HB-EGF facilitated the opening of mitochondrial permeability transition pore via targeting BAX and raised the release of cytochrome C from mitochondria into the cytosol to trigger the apoptosis of GCs, but this effectiveness was counteracted by estrogen receptor antagonist. Collectively, HB-EGF might induce mitochondrial dysfunction and GCs apoptosis through advancing estrogen hypersecretion dependent on cAMP-PKA-JNK/ERK-Ca2+-FOXO1 pathway and act as a promising therapeutic target for PCOS.

Yap is essential for uterine decidualization through Rrm2/GSH/ROS pathway in response to Bmp2.

Yap is required for ovarian follicle and early embryo development, but little information is available regarding its physiological significance in decidualization. Here we determine the effects of YAP on decidualization, mitochondrial function, cell apoptosis and DNA damage, and explore its interplay with Bmp2, Rrm2, GSH and ROS. The results exhibited that Yap was abundant in decidual cells and its inactivation impaired the proliferation and differentiation of stromal cells along with the deferral of G1/S phase transition, indicating Yap importance in decidualization. Bmp2 via Alk2 receptor promoted nuclear translocation of Yap where it might interact with Tead and then bind to the promoter of Rrm2 whose activation rescued the faultiness of differentiation program and attenuated oxidative DNA damage caused by Yap impediment. Meanwhile, Yap had an important part in the crosstalk between Bmp2 and Rrm2. Furthermore, inactivation of Yap resulted in an obvious accumulation of intracellular ROS followed by the abnormal GR activity and GSH content dependent on Rrm2. Replenishment of GSH counteracted the regulation of Yap inactivation on stromal differentiation and DNA damage with distinct reduction for intracellular ROS. Additionally, blockage of Yap caused the enhancement of stromal cell apoptosis and brought about mitochondrial dysfunction as indicated by the aberration for ATP level, mtDNA copy number and mitochondrial membrane potential concomitant with the opening of mitochondrial permeability transition pore, but these abnormalities were neutralized by GSH. Administration of mitochondrial antioxidant Mito-TEMPO rescued the fault of stromal differentiation conferred by Yap inactivation. Collectively, Yap was essential for uterine decidualization through Rrm2/GSH/ROS pathway in response to Bmp2.

[Nested PCR for malaria detection and Plasmodium species identification].

According to the sequences of small subunit ribosomal RNA (SSU rRNA) gene of Plasmodium spp., universal and species-specific primers were designed to detect malaria and identify species. 60 blood samples were detected by the established nested PCR method. The results were compared with those of microscopic examination. 40 blood samples were Plasmodium-positive by nested PCR with 22 samples of P. falciparum, 13 of P. vivas, 3 with P. falciparum and P. vivax mixed infection, 1 of P. ovale and 1 of unclassified malaria infection. Altogether, the coincidence between the results of nested PCR and microscopy stood for 76.7% (46/60), including 18 of P. falciparum, 11 of P. vivax and 17 negatives. Further sequence analysis and real-time PCR were performed to detect blood samples with discrepancy, results of which were the same as that of nested PCR. The amplified product of P. ovale was sequenced and showed 100% homology to the corresponding part of P. ovale SSU rRNA gene sequence (GenBank No. DQ845247), which confirmed that the case was imported ovale malaria.

Genistein exhibits therapeutic potential for PCOS mice via the ER-Nrf2-Foxo1-ROS pathway.

Polycystic ovarian syndrome (PCOS) is a complex endocrinopathy in women of reproductive age and the main cause of female infertility, but there is no universal drug for PCOS therapy. As a predominant dietary isoflavone present in soybeans, genistein (GEN) possesses estrogenic and antioxidative properties, but limited information is available regarding its therapeutic potential and underlying molecular mechanism in PCOS. In this study, we found that GEN might restore the estrous cycle of PCOS mice and ameliorate the elevation of circulating T, AMH and LH levels as well as LH/FSH ratios along with reduced cystic follicles, indicating the importance of GEN in PCOS therapy. Meanwhile, GEN improved the ovarian secretion function of PCOS mice and attenuated oxidative damage of the ovary through enhancing its antioxidant capability dependent on ER. Supplementation of GEN improved the defect of the ATP level and mitochondrial membrane potential, indicating the significance of GEN in preventing mitochondrial dysfunction. Further analysis demonstrated that GEN via ER heightened the expression of Nrf2 and Foxo1 whose blockage antagonized the defence of GEN on the secretory and mitochondrial functions of ovarian granulosa cells followed by the limited antioxidant capability and increased intracellular ROS level. Moreover, nuclear translocation and transcriptional activity of Nrf2 presented a notable enhancement after exposure to GEN. Addition of the Nrf2 inhibitor ML385 hampered the GEN induction of Foxo1. Nrf2 might directly bind to the antioxidant response element of the Foxo1 promoter region. Collectively, GEN might exhibit therapeutic potential for PCOS mice via the ER-Nrf2-Foxo1-ROS pathway.

Perianal mucinous adenocarcinoma arising from chronic anorectal fistulae: a review from single institution.

BACKGROUND: Mucinous adenocarcinoma arising from a chronic anorectal fistula is rare, with few reports in the literature. Such lesions can be misdiagnosed for the more common benign perianal abscess or fistula. METHODS: From our retrospective chart review, we identified three patients with chronic perianal fistula-in-ano who were subsequently found to have developed perianal mucinous adenocarcinoma on biopsy. We recorded the symptomatology, subsequent management and further follow-up of each patient. RESULTS: Two of three patients who received irradiation and chemotherapy were still alive during 28 and 24 months of follow-up, respectively without any evidence of distant metastasis. One patient with inguinal lymph node metastases died due to distant metastasis 6 months after diagnosis. CONCLUSIONS: Fistula-associated perianal mucinous adenocarcinoma is an uncommon malignant transformation of chronic fistula-in-ano. MRI can provide important diagnostic information on patient with this suspicious inflammatory condition. Although radical resection of the tumour with abdominoperineal resection remains the surgical treatment of choice. Combined chemoradiotherapy may be appropriate for these patients with promising results.

Clinical, virological and immunological features from patients infected with re-emergent avian-origin human H7N9 influenza disease of varying severity in Guangdong province.

BACKGROUND: The second wave of avian influenza H7N9 virus outbreak in humans spread to the Guangdong province of China by August of 2013 and this virus is now endemic in poultry in this region. METHODS: Five patients with H7N9 virus infection admitted to our hospital during August 2013 to February 2014 were intensively investigated. Viral load in the respiratory tract was determined by quantitative polymerase chain reaction (Q-PCR) and cytokine levels were measured by bead-based flow cytometery. RESULTS: Four patients survived and one died. Viral load in different clinical specimens was correlated with cytokine levels in plasma and broncho-alveolar fluid (BALF), therapeutic modalities used and clinical outcome. Intravenous zanamivir appeared to be better than peramivir as salvage therapy in patients who failed to respond to oseltamivir. Higher and more prolonged viral load was found in the sputum or endotracheal aspirates compared to throat swabs. Upregulation of proinflammatory cytokines IP-10, MCP-1, MIG, MIP-1α/ß, IL-1ß and IL-8 was found in the plasma and BALF samples. The levels of cytokines in the plasma and viral load were correlated with disease severity. Reactivation of herpes simplex virus type 1(HSV-1) was found in three out of five patients (60%). CONCLUSION: Expectorated sputum or endotracheal aspirate specimens are preferable to throat swabs for detecting and monitoring H7N9 virus. Severity of the disease was correlated to the viral load in the respiratory tract as well as the extents of cytokinemia. Reactivation of HSV-1 may contribute to clinical outcome.

Epidemiological and viral genome characteristics of the first human H7N9 influenza infection in Guangdong Province, China.

BACKGROUND: The first H7N9 human case in south of China was confirmed in Guangdong Province on August 2013, outside of the typical influenza season. For investigating the H7N9 virus source and transmission in the local community, we analyze the epidemiology and genome features of the virus isolated from the first human infection detected in Guangdong Province. METHODS: The data including medical records, exposure history and time line of events for the H7N9 patient and close contacts was collected. Variation and genetic signatures of H7N9 virus in Guangdong was analyzed using ClustalW algorithm and comparison with mutations associated with changes in biological characteristics of the virus. RESULTS: The female patient had a history of poultry exposure, and she was transferred from a local primary hospital to an intensive care unit (ICU) upon deterioration. No additional cases were reported. Similar to previous infections with avian influenza A (H7N9) virus, the patient presented with both upper and lower respiratory tract symptoms. Respiratory failure progressed quickly, and the patient recovered 4 weeks after the onset of symptoms. Genome analysis of the virus indicated that the predicted antigen city and internal genes of the virus are similar to previously reported H7N9 viruses. The isolated virus is susceptible to neuraminidase (NA) inhibitors but resistant to adamantine. Although this virus contains some unique mutations that were only detected in avian or environment-origin avian influenza A (H7N9) viruses, it is still quite similar to other human H7N9 isolates. CONCLUSIONS: The epidemiological features and genome of the first H7N9 virus in Guangdong Province are similar to other human H7N9 infections. This virus may have existed in the environment and live poultry locally; therefore, it is important to be alert of the risk of H7N9 re-emergence in China, including emergence outside the typical influenza season.

[Isolation and identification of SARS virus in Guangdong province].

BACKGROUND: To isolate and identify pathogen of atypical pneumonia in Guangdong. METHODS: Pathogens were isolated from variety of samples collected from atypical pneumonia patient by using MDCK cells, and identified with serological and molecular methods. RESULTS: A novel coronavirus was isolated from patients with atypical pneumonia, from which an RNA fragment of 279 nt was amplified by nested RT-PCR. And sequence assay showed that only 39-65 percent of sequence of the virus was homogenous to known coronavirus, but almost 100% homogenous (with one base exception, 12a to t) to SARS-associated coronavirus isolated from patients outside Guangdong, such as in Beijing, Hong Kong, Taiwan, Germany, Italy and so on. Indirect immunofluorescence test showed a specific antigen-antibody reactivity between the coronavirus and convalescent-phase sera of SARS patients. CONCLUSION: The pathogen of the atypical pneumonia in Guangdong province was a novel type of coronavirus, which could be isolated by using MDCK cells.