[Analysis of the clinical characteristics and species distribution of non-tuberculous mycobacteria in a general hospital].

Objective: To summarize the clinical characteristics and species distribution of nontuberculous mycobacteria(NTM). Methods: We conducted a retrospectively chart review of 453 adult patients with 532 positive cultures with NTM diagnosed and treated in Peking Union Medical College Hospital from January 2014 to December 2018. The information including clinical findings, imaging and etiological data were obtained from clinical examination system. Among these 453 patients, 75 cases met the diagnostic criteria for NTM, with detailed clinical data, culture results and radiological images. Of these patients, 38 were males and 37 were females, with an average age of (50.8±16.2) years. Clinical manifestations, imaging features, NTM species and treatment were analyzed. Results: Among 75 NTM patients, 43 cases (57.3%, 43/75) were infected with Mycobacterium avium Complex (MAC)/intracellular, 18 cases (24.0%, 18/75) with Mycobacterium turtle/abscess, and 7 cases (9.3%, 7/75) with Mycobacterium kansasii. Comorbidities existed in 64.0% patients (48/75). The top two common comorbidities were previous tuberculosis (25.3%, 19/75) infection and clinical bronchiectasis (18.7%, 14/75). After the respiratory disease (82.7%, 62/75), we identified bloodstream as the most frequent manifestation of NTM infection (25.3%, 19/75). MAC was often associated with infection in respiratory system and bloodstream, while rapidly growing mycobacteria (RGM) was more commonly detected in lymph nodes and skin and soft tissues. Cough/expectoration (77.4%, 48/62) was the most common clinical symptom of NTM disease, followed by fever and dyspnea. The frequent imaging findings were nodules (41.9%, 26/62), bronchiectasis (37.1%, 23/62), and fibrous cavities (24.2%, 15/62). In disseminated NTM disease, 94.7% (18/19) patients had underlying diseases and 89.5% (17/19) patients had long-term immunosuppression. Conclusion: The most common NTM species was Mycobacterium avium/intracellular in our series. The predominant infected organ was the respiratory system. Previous tuberculosis history likely increased the risk of infection.

Rapamycin induces human acute promyelocytic leukemia cell HL-60 autophagic apoptosis.

OBJECTIVE: We aimed at investigating the effects of rapamycin on apoptosis and autophagy of human acute promyelocytic leukemia cell line HL-60, and to preliminarily explore the mechanism of extra medullary infiltration of leukemia cells with human acute promyelocytic leukemia cell line HL-60 as the object of study, providing a theoretical basis for the clinical treatment of leukemia. MATERIALS AND METHODS: After HL-60 cells were cultured in vitro, the effect of rapamycin on proliferation ability of HL-60 cells was determined by methyl thiazolyl tetrazolium (MTT) method, the cell apoptosis ratio was detected by flow cytometer, the change of autophagy after HL-60 cells acted by rapamycin was tested by monodansylcadaverine (MDC) fluorescence staining, the mRNA expression of autophagy-related molecule was detected by polymerase chain reaction (PCR), and the expressions of apoptosis-related protein and autophagy-related protein were determined by Western blotting (WB). RESULTS: HL-60 cell proliferation could be significantly inhibited by rapamycin (80 µg/mL-640 µg/mL), which was in a dose-dependent manner. HL-60 cell apoptosis ratio and apoptosis-related protein expression were distinctly improved by rapamycin. Cell autophagy level, mRNA expression of autophagy-related molecule and autophagy-related protein expression were remarkably induced by rapamycin. CONCLUSIONS: Rapamycin can induce HL-60 cell apoptosis, which is produced mainly by inducing cell autophagy.

[A retrospective study of 10 cases of post-transplant bronchiolitis obliterans].

Objective: To analyze the clinical features of bronchiolitis obliterans syndrome (BOS) in patients with allogeneic hematopoietic stem cell transplantation (HSCT). Methods: This retrospective study included patients who underwent allogeneic HSCT from January 1998 to December 2016. The clinical features, radiological manifestations and treatment of clinically proven BOS were reviewed. Results: Of 681 patients who experienced HSCT, 10(1.47%) met the diagnostic criteria. The duration of BOS onset after transplantation was 5-48 months, averaging (18±15) months. Cough and worsening dyspnea were present in most cases, and 9 (90%) of the cases had manifestations of chronic graft versus host disease. Hyperinflation with areas of decreased attenuation and bronchiectasis were present in 7 (70%) cases and air trapping was present during the expiratory phase of imaging. The management of BOS consisted of high dose systemic corticosteroids and immunosuppressive therapy. Patients were treated with bronchodilators if they were symptomatic and during acute exacerbations of respiratory symptoms. In 5 (50%) patients the condition deteriorated and caused death, while in 3 patients the condition was improved. Two patients were lost to follow up. Conclusion: BOS is the most common late noninfectious pulmonary complication following allogeneic HSCT and has a poor prognosis.

Crystal structure of human epidermal growth factor and its dimerization.

Epidermal growth factor (EGF) is a typical growth-stimulating peptide and functions by binding to specific cell-surface receptors and inducing dimerization of the receptors. Little is known about the molecular mechanism of EGF-induced dimerization of EGF receptors. The crystal structure of human EGF has been determined at pH 8.1. There are two human EGF molecules A and B in the asymmetric unit of the crystals, which form a potential dimer. Importantly, a number of residues known to be indispensable for EGF binding to its receptor are involved in the interface between the two EGF molecules, suggesting a crucial role of EGF dimerization in the EGF-induced dimerization of receptors. In addition, the crystal structure of EGF shares the main features of the NMR structure of mouse EGF determined at pH 2.0, but structural comparisons between different models have revealed new detailed features and properties of the EGF structure.

[Efficacy of ablendazole emulsion in treatment of 212 patients with cystic echinococcosis].

OBJECTIVE: To verify the efficacy of abendazole emulsion, a new formulation of abendazole, in treatment of human cystic echinococcosis. METHODS: 212 patients with liver cystic echinococcosis were treated orally with albendazole emulsion at a daily dose of 10 mg/kg or 12.5 mg/kg for 3 to 12 months or over one year. The therapeutic efficacy was mainly evaluated by image feature examined with B ultrasound examination, a short-term efficacy at the completion of treatment and a long-term efficacy followed-up for 1-4 years. RESULTS: In 212 patients treated with albendazole emulsion at a daily dose of 10 mg/kg and 12.5 mg/kg, the average cure rate, improved rate and the rate of no avail were 74.5%, 99.1% and 0.9% respectively after termination of the treatment, and the average long-term rates were 83.1%, 89.3% and 0.6% respectively. Recurrence occurred in 18 patients(10.2%). The results indicated that the best efficacy was seen in patients treated with albendazole 12.5 mg/kg daily for 9 months. Better response was also found when the recurrent patients were re-treated with albendazole. CONCLUSION: The efficacy of albendazole emulsion on patients with liver cystic echinococcosis is much better than that of albendazole tablet or capsule and mebendazole. Meanwhile, the efficacy of albendazole emulsion is stable with less adverse effects. The results suggest that albendazole emulsion could be the drug of choice for treatment of cystic hydatid disease.

Molecular-replacement studies of Trichosanthes kirilowii lectin 1: a structure belonging to the family of type 2 ribosome-inactivating proteins.

Trichosanthes kirilowii lectin 1 (TKL-1) isolated from the tuber of T. kirilowii consists of two chains, each with a molecular weight of about 30 kDa. It has immunological properties which are similar to some ribosome-inactivating proteins (RIPs). TKL-1 was crystallized in space group P2(1)2(1)2(1) and diffraction data were collected to 2.7 A resolution. The molecular-replacement method was applied to solve the structure, using different chains of ricin, abrin-a and trichosanthin as search models. A set of consistent solutions was further verified by R(omit) profile analysis. In addition, the spatial arrangement of the two chains of TKL-1 is identical to that of type 2 RIPs.

Crystallization and preliminary X-ray diffraction studies of human epidermal growth factor.

Human epidermal growth factor (hEGF), a 6.2 kDa protein of 53 amino acids with three internal disulfide bridges, has been crystallized by the hanging-drop method. hEGF crystallizes in space group P3(1)21 (or P3(2)21) using MgCl(2) as precipitant, with unit-cell parameters a = b = 61.4, c = 87.0 A. Another type of crystal, obtained using NaCl as precipitant, belongs to a tetragonal point group and has unit-cell dimensions a = b = 102.5, c = 166.6 A. The trigonal crystals with the smaller unit cell diffract X-rays better and a native data set from a single crystal has been collected to 3.0 A resolution.

Rapid screening with a recombinant antigen (rK39) for diagnosis of visceral leishmaniasis using dipstick.

OBJECTIVE: To evaluate the diagnostic value of the recombinant antigen of 39 amino acid repeats encoded by a kinesin-like gene of Leishmania changasi (rK39) in serodiagnosis of visceral leishmaniasis (VL). METHODS: In Kashi, Xinjiang, 13 VL patients with splenomegaly and bone marrow aspirate culture positive were subjected to dipstick assay. A drop of whole blood or serum from patient was placed at the absorbing pad at the bottom of the dipstick. Flooding of the bottom protein with buffer allows serum proteins to migrate upwards, producing the positive band and Western blot analysis of rK39 subsequently performed with the sera collected. RESULTS: The end-point titers of anti-rK39 antibodies of these sera were determined by ELISA and found to fall within the range of 10(-2) to 10(-4), which were consistent with the intensity of their reaction with rK39 in dipstick assay. The positive sera could also recognize the specific rK39 band as analyzed by Western blot analysis. CONCLUSION: The rK39 dipstick assay is more rapid, specific, sensitive and less invasive than the conventional methods of diagnosis for VL in the areas of low endemicity.

Differential serodiagnosis for cystic and alveolar echinococcosis using fractions of Echinococcus granulosus cyst fluid (antigen B) and E. multilocularis protoscolex (EM18).

Echinococcus granulosus cyst fluid and E. multilocularis protoscolex extract were fractionated by a single step of preparative isoelectric focusing, resulting in an antigen B-rich fraction (8-kD) and an Em18-rich fraction, respectively. The usefulness of both fractions for differential serodiagnosis of cystic (CE) and alveolar (AE) echinococcosis was evaluated by a large-scale immunoblot analysis on a battery of 354 serum samples. These included 66 from AE patients originating from four different endemic areas, 173 from CE patients originating from seven different endemic areas, 71 from patients with other parasitic diseases, 15 from patients with hepatomas, and 29 from healthy individuals. In an immunoblot with the antigen B-rich fraction, 92% (158 of 173) of the CE sera as well as 79% (52 of 66) of the AE sera reacted with the 8-kD subunit. No cross-reactivity occurred with any sera from patients with cysticercosis, other parasitic diseases, or with hepatomas, or from healthy controls. In an immunoblot with the Em18-rich fraction, all but two sera from AE patients (64 of 66, 97%) recognized Em18, and only nine of 34 CE sera from China reacted with it. All other (139) CE sera from six other countries were negative as were all (115) other non-echinococcosis sera. These findings indicate that antigen B (8-kD) is not species-specific for E. granulosus but is genus-specific for Echinococcus, and that the Em18 antigen is a reliable serologic marker for species-specific differentiation of AE from CE.

Crystal structure of carboxypeptidase A complexed with an inactivator in two crystal forms.

Two different crystal forms of carboxypeptidase A (CPA) complexed with an inactivator were obtained by the method of hanging drop vapor diffusion. The inactivator, 2-benzyl-3-iodo-propanoic acid (BIPA), binds covalently to an active site residue Glu270 of CPA. The complexes were crystallized in the space group P2(1) (CPA-I) and P2(1)2(1)2(1) (CPA-II), respectively. The structures of both crystal forms were determined by molecular replacement using the native CPA crystal structure as the search model. The final crystallographic residuals are 0.163 for CPA-I and 0.152 for CPA-II. Except for the modification of Glu270, the inactivator exhibits normal binding mode compared with other ligand complexes of CPA. In the final electron density difference maps (2Fo-Fc, Fo-Fc), the density of the iodo ion could not be found in both crystal forms while the conserved water molecule remains coordinated to Zn2+ as in the native CPA. Comparisons of the complexes of CPA-BIPA with the native CPA and the CPA-D-Phe complex are presented. The mechanism of the inactivation of CPA and its implication for catalytic mechanism were discussed.

Diagnosis of kala-azar by nested PCR based on amplification of the Leishmania mini-exon gene.

To diagnose visceral leishmaniasis (kala-azar), we have developed a nested PCR method based on amplification of the mini-exon gene, which is unique and tandomly repeated in the Leishmania genome. Nested PCR was sufficiently sensitive for the detection of DNA in an amount equivalent to a single Leishmania parasite or less. We examined the usefulness of this PCR method using bone marrow aspirates and buffy coat cells collected from kala-azar patients who had or had not received chemotherapy in northwest China. We obtained PCR positivity for all of the parasitologically positive bone marrow samples from the patients. Some ambiguities with the primary PCR results were eliminated by the subsequent nested PCR. The buffy coat samples from 7 of 12 patients with splenomegaly were positive by the nested PCR, although only 2 of them were positive for parasites by culture. However, buffy coat samples from nine children, whose splenomegaly has been reduced and clinically cured by antimony treatment, were all negative. Thus, this nested PCR method represents a new tool for the diagnosis of kala-azar with patient blood samples instead of bone marrow or spleen aspirates obtained by more invasive procedures.

Short report: An imported case of cystic echinococcosis in Japan diagnosed by imaging and serology with confirmation of Echinococcus granulosus-specific DNA sequences.

We report one case of cystic echinococcosis (CE) in Japan in a native of Nepal. Ultrasonography and computed tomography scan of the liver revealed unique cystic lesions with or without daughter cysts of Echinococcus granulosus. Immunoblot analysis using crude antigens of E. multilocularis and cyst fluid of E. granulosus, without reference to these image analyses, strongly suggested this was a case of CE. We found protoscoleces in surgically removed hepatic lesions and analyzed the mitochondrial cytochrome c oxidase subunit I (COI) gene by the polymerase chain reaction. Based on the similarity in DNA sequences of the COI gene of this Echinococcus spp. with that of previously reported sheep-dog strain (GI), the parasite was considered to be the so-called common sheep strain of E. granulosus.

Mitochondrial genomic markers confirm the presence of the camel strain (G6 genotype) of Echinococcus granulosus in north-western China.

Twenty-eight isolates of E. granulosus, collected from humans at surgery, and a range of intermediate hosts, including sheep, cattle and camels from abattoirs in North and South Xinjiang Uygur Autonomous Region, People's Republic of China, were analysed for DNA sequence variation within regions of the mitochondrial cytochrome c oxidase I (COI) and NADH dehydrogenase subunit I (NDI) genes. The isolates were categorized into 2 distinct and uniform genotypic groupings, based on the sequences obtained, and the data clearly indicated that the camel/dog strain (G6 genotype) of E. granulosus as well as the cosmopolitan, common sheep strain (G1 genotype) occur in north Xinjiang. The presence of the camel strain has thus been confirmed in Xinjiang but it is evident from this and a previous molecular genetic survey of E. granulosus isolates from north-western China that the common sheep strain is the most predominant in the region. From the public health perspective, the majority of infected livestock will act as reservoirs of human infection there. During the course of the study, a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) assay, based on the NDI sequence variation, was developed that allows rapid discrimination of the G1 and G6 genotypes.

[The structural elucidation of new polyoxygenated cyclohexenes from Uvaria grandiflora].

Four new polyoxygenated cyclohexenes, named uvarigranol A(1), B(2), E(3) and F(4), along with the known zeylenol were obtained from the roots of Uvaria grandiflora Roxb (Annonaceae). Based on spectral and X-ray analysis as well as Mosher methodology, their structures and absolute configuration were established.

Epidemiological studies on cystic echinococcosis in China--a review.

In the four decades from 1951 to 1990, the six provinces or autonomous regions (Xinjiang, Gansu, Qinghai, Ningxia, Xizang and Nei Monggol) reported a total of 26,065 surgical cases of hydatid disease, most of which were reported in the recent decade. About one third of the patients was children and adolescents under 15 years old. So far, cystic hydatid infections of local origin have been confirmed in 22 provinces, autonomous regions and municipalities in the whole nation. Findings of X-ray examination and real-time B-mode ultrasonography in agricultural and pastoral areas of Xinjiang, Gansu, Qinghai, Ningxia and Xizang showed that the morbidity rate of hydatidosis in human population varied between 0.5% and 4.5%. The main animal intermediate host in all these regions is sheep, the morbidity of which varied between 3.3% and 90%. The infection rate of adult Echinococcus granulosus in dogs varied between 7% and 71%. The high-risk period for humans contracting hydatid disease is the pre-school age. Direct contact of children with dogs and ingestion of water, vegetables and foods contaminated by worm eggs are the chief mode of transmission for human hydatidosis. The common practices of home slaughter and of feeding dogs on offal containing hydatid cysts facilitate the life cycle of the parasite. On the basis of hydatid control efforts for several years, the Ministry of Public Health promulgated officially the "1992-1995 National Programme for Hydatid Disease Control" in April, 1992, and pilot studies in which the control of hydatidosis is composed of extensive health education, sanitation of slaughtering and management and deworming of dogs are being established. Cystic echinococcosis has wide distribution in China and is a major public health problem in hyperendemic areas, poses a great threat against people's health and influences the development of livestock husbandry. This problem has received great attention from the medical and veterinary departments. In the recent decade great efforts have been made in the epidemiology, parasitology and clinical treatment of echinococcosis, resulting in a better understanding of the disease and procedures effective in bringing about control of the disease.

Effects of benzimidazole compounds on mice infected with secondary cysts of Echinococcus granulosus.

The effects of mebendazole (Meb), albendazole (Alb) and albendazole sulfoxide (AlbSO) on metacestodes of Echinococcus granulosus have been studied. The results show that Meb and Alb exhibit less effect on protoscoleces in vitro, but in vivo the protoscolicidal effect of Meb is higher than that of Alb. Both Meb and Alb are efficacious in the treatment of mice infected with secondary cysts of E. granulosus. In terms of the minimal effective dose required, the occurrence of collapsed and shrunk cysts after treatment, the drug-induced damage on the germinal layer and the relationship between the drug content in the cyst wall and the damage of germinal layer, the effect of Meb on metacestodes of E. granulosus is higher than that of Alb. The results also indicated that lower drug absorption rate and thickening of the adventitia during longer disease course are the two major factors affecting the efficacy of Meb and Alb, hence suggesting that increase of Meb absorption may be expected to raise the therapeutic effect of the drug. Finally, Alb is not only metabolized to AlbSo and AlbSP in vivo, but also metabolized by the cysts exposed to Alb in vitro. Experimental chemotherapy showed that AlbSO is the major effective metabolite of Alb, and its curative dose is only one half of the parent compound.

Separation of Chinese Leishmania isolates into five genotypes by kinetoplast and chromosomal DNA heterogeneity.

Leishmaniasis remains endemic in China, especially in the west and northwest frontier regions in central Asia. Epidemic outbreaks of both visceral and cutaneous forms of the disease have become a serious concern in view of such events occurring in neighboring countries. In the present study, we have begun to characterize available parasites as an initial step in understanding the epidemiology of leishmaniasis in central Asia. Nineteen Leishmania isolates collected since the 1950s from epidemiologically different foci in China were separated into five genotypes (Groups I-V) based on their polymorphisms in both kinetoplast (kDNA) and nuclear (nDNA) DNAs. Both kDNA and nDNA are conserved in Group I, which consists of six isolates, i.e., five cases of human kala-azar and one case of canine leishmaniasis isolated from three distant foci more than 30 years apart. In contrast, both kDNA and nDNA are heterogeneous in Group II, consisting of 10 isolates scattered in the plain area from the eastern coast to the western desert. This group includes five kala-azar cases, one post-kala-azar dermal leishmaniasis case, two sand fly isolates, and two canine isolates. The remaining three groups (III-V), two from great gerbils (Rhombomys opimus) and one from a kala-azar case, differ among themselves and from the aforementioned groups. Groups I, II/III, IV, and V contain isolates that have been recognized epidemiologically or typed isoenzymatically as L. donovani s.l., L. infantum s.l., L. turinica, and L. gerbilli, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

[Effects of mebendazole on glucose, glycogen, lactic acid, and lactate dehydrogenase in Echinococcus granulosus cyst wall].

The glucose, glycogen, and lactic acid contents in Echinococcus granulosus cyst wall were 3570 +/- 623, 1276 +/- 126, and 267 +/- 29 micrograms.g-1, respectively. The lactate dehydrogenase (LDH) activity in the cyst wall was 4307 +/- 297 U/(min.mg protein). When infected mice were treated ig with mebendazole (Meb) 25 or 50 mg.kg-1.d-1 for 7-14 d, the glucose and glycogen contents in the cyst wall decreased markedly. No apparent changes in lactic acid and LDH activity were seen. The results suggested that Meb had no effect on glycolysis of the cyst wall, but it exhibited an inhibitory effect on the transport of exogenous glucose from the host to the cyst wall, which might further promote the decomposition of endogenous glycogen for energy supply and resulted in a decrease of endogenous glycogen content in the cyst wall.