The significance of standardized fetal ultrasonographic screening in the first trimester / 中华医学超声杂志（电子版）

Objective To evaluate the significance of standardized ultrasound examination of fetal structures in the ifrst trimester (11-13+6 weeks). Methods The ultrasound data of 29 858 fetuses who underwent the ifrst trimester screening were analyzed retrospectively in Beijing Obstetrics and Gynecology Hospital of Capital Medical University from January 2010 to December 2012 and followed up the cases with structural abnormalities and increased nuchal translucency (NT). Results Among 29 858 fetuses, 328 structural defects were detected in 284 fetuses (0.95%) by ultrasound in the first trimester, including fetal hydrops (124 cases), choroid plexus cysts (36 cases), exencephaly (32 cases), gastroschisis (24 cases), exomphalos (22 cases), cardiac defects (19 cases), megacystis (14 cases), spine abnormalities (10 cases), meningoceles/encephaloceles (9 cases), alobar holoprosencephaly (8 cases), hydrocephaly (7 cases), abnormalities of extremities (7 cases), acrania (6 cases), amniotic band syndrome (4 cases), abdominal cyst (3 cases) and conjoined twins (3 cases). Pregnancy was terminated in all cases (115) with exencephaly, gastroschisis, exomphalos, meningoceles/encephaloceles, alobar holoprosencephaly, hydrocephaly,abnormalities of extremities, acrania, amniotic band syndrome and conjoined twins, and the defects were verified by the gross appearance of fetuses. Two cases of megacystis were turned out to be normal during follow-up while 1 case was proved to be aneuploid and 1 other case progressed to multiple abnormalities during the second trimester. Most choroid plexus cysts, except 1 case of aneuploid, were normal during the second and the third trimester follow-ups by ultrasound. Thirteen cases of fetal hydrops were found to be aneuploids and 4 hydrops cases were proved to be complicated with other structural defects during the second and the third trimester. NT was increased in 422 cases (1.41%), among which 122 cases (28.91%) were complicated with structural defects and/or fetal hydrops. Ninty-nine cases with increased NT underwent chromosomal examination and 38 cases were found to be aneuploides. During the follow-up of 298 cases with increased NT who continued pregnancy, 21 structural abnormalities were found, including cardiac defects (14 cases), neural abnormalities (4 cases) and diaphragmatic hernia (3 cases). Conclusion The standardized fetal ultrasound in the first trimester is a effective tool for the screening of aneupolides and major structural abnormalities.

Ultrasonographic characteristics of ovarian yolk sac tumor / 中华医学超声杂志（电子版）

Objective To investigate ultrasonographic features of primary and metastatic ovarian yolk sac tumors.Methods Ultrasonographic features of 19 primary lesions and 33 metastatic lesions in 35 patients were retrospectively analyzed.Results Primary tumors were sized (14.6±3.6)cm in maximum diameter,manifesting as cysti-solid masses.Solid components of primary tumors were mainly hypoechoic or isoechoic(16/19)with rich blood supplies.Thirty-three metastatic lesions were located in pelvoceliac cavity(26/33) and liver parenchyma(7/33),sized (9.4±4.5)cm,(9.2±4.9)cm and (5.6±1.6)cm in maximum diameter respectively.Metastatic lesions in pelvoceliac lesions mainly demonstrated as hypoechoic masses(21/26), however lesions in the liver were mainly hyperechoic(5/7).Anechoic regions could be found in 9/26 of the pelvoceliac lesions.Blood supply was found less rich in metastatic masses than that in primary ones.Elevated serum level of alpha fetoprotein (AFP) was observed in all patients; ranging from 217 to 211 682 μg/L.Conclusions Primary and metastatic lesions of ovarian yolk sac tumor have obvious ultrasonographic characteristics.Combined with serum AFP level,the accuracy of diagnosis could be improved.

Microarray Analysis of Differentially Expressed Genes between Cysts and Trophozoites of Acanthamoeba castellanii

Acanthamoeba infection is difficult to treat because of the resistance property of Acanthamoeba cyst against the host immune system, diverse antibiotics, and therapeutic agents. To identify encystation mediating factors of Acanthamoeba, we compared the transcription profile between cysts and trophozoites using microarray analysis. The DNA chip was composed of 12,544 genes based on expressed sequence tag (EST) from an Acanthamoeba ESTs database (DB) constructed in our laboratory, genetic information of Acanthamoeba from TBest DB, and all of Acanthamoeba related genes registered in the NCBI. Microarray analysis indicated that 701 genes showed higher expression than 2 folds in cysts than in trophozoites, and 859 genes were less expressed in cysts than in trophozoites. The results of real-time PCR analysis of randomly selected 9 genes of which expression was increased during cyst formation were coincided well with the microarray results. Eukaryotic orthologous groups (KOG) analysis showed an increment in T article (signal transduction mechanisms) and O article (posttranslational modification, protein turnover, and chaperones) whereas significant decrement of C article (energy production and conversion) during cyst formation. Especially, cystein proteinases showed high expression changes (282 folds) with significant increases in real-time PCR, suggesting a pivotal role of this proteinase in the cyst formation of Acanthamoeba. The present study provides important clues for the identification and characterization of encystation mediating factors of Acanthamoeba.

Endosymbionts of Acanthamoeba Isolated from Domestic Tap Water in Korea

In a previous study, we reported our discovery of Acanthamoeba contamination in domestic tap water; in that study, we determined that some Acanthamoeba strains harbor endosymbiotic bacteria, via our molecular characterization by mitochondrial DNA restriction fragment length polymorphism (Mt DNA RFLP). Five (29.4%) among 17 Acanthamoeba isolates contained endosymbionts in their cytoplasm, as demonstrated via orcein staining. In order to estimate their pathogenicity, we conducted a genetic characterization of the endosymbionts in Acanthamoeba isolated from domestic tap water via 16S rDNA sequencing. The endosymbionts of Acanthamoeba sp. KA/WP3 and KA/WP4 evidenced the highest level of similarity, at 97% of the recently published 16S rDNA sequence of the bacterium, Candidatus Amoebophilus asiaticus. The endosymbionts of Acanthamoeba sp. KA/WP8 and KA/WP12 shared a 97% sequence similarity with each other, and were also highly similar to Candidatus Odyssella thessalonicensis, a member of the alpha-proteobacteria. The endosymbiont of Acanthamoeba sp. KA/WP9 exhibits a high degree of similarity (85-95%) with genus Methylophilus, which is not yet known to harbor any endosymbionts. This is the first report, to the best of our knowledge, to show that Methylophilus spp. can live in the cytoplasm of Acanthamoeba.

Construction of EST Database for Comparative Gene Studies of Acanthamoeba

The genus Acanthamoeba can cause severe infections such as granulomatous amebic encephalitis and amebic keratitis in humans. However, little genomic information of Acanthamoeba has been reported. Here, we constructed Acanthamoeba expressed sequence tags (EST) database (Acanthamoeba EST DB) derived from our 4 kinds of Acanthamoeba cDNA library. The Acanthamoeba EST DB contains 3,897 EST generated from amebae under various conditions of long term in vitro culture, mouse brain passage, or encystation, and downloaded data of Acanthamoeba from National Center for Biotechnology Information (NCBI) and Taxonomically Broad EST Database (TBestDB). The almost reported cDNA/genomic sequences of Acanthamoeba provide stand alone BLAST system with nucleotide (BLAST NT) and amino acid (BLAST AA) sequence database. In BLAST results, each gene links for the significant information including sequence data, gene orthology annotations, relevant references, and a BlastX result. This is the first attempt for construction of Acanthamoeba database with genes expressed in diverse conditions. These data were integrated into a database (http://www.amoeba.or.kr).

Keratitis by Acanthamoeba triangularis: Report of Cases and Characterization of Isolates

Three Acanthamoeba isolates (KA/E9, KA/E17, and KA/E23) from patients with keratitis were identified as Acanthamoeba triangularis by analysis of their molecular characteristics, a species not previously recognized to be a corneal pathogen. Epidemiologic significance of A. triangularis as a keratopathogen in Korea has been discussed. Morphologic features of Acanthamoeba cysts were examined under a microscope with differential interference contrast (DIC) optics. Mitochondrial DNA (mtDNA) of the ocular isolates KA/E9, KA/E17, and KA/E23 were digested with restriction enzymes, and the restriction patterns were compared with those of reference strains. Complete nuclear 18S and mitochondrial (mt) 16S rDNA sequences were subjected to phylogenetic analysis and species identification. mtDNA RFLP of 3 isolates showed very similar patterns to those of SH621, the type strain of A. triangularis. 16S and 18S rDNA sequence analysis confirmed 3 isolates to be A. triangularis. 18S rDNA sequence differences of the isolates were 1.3% to 1.6% and those of 16S rDNA, 0.4% to 0.9% from A. triangularis SH621. To the best of our knowledge, this is the first report, confirmed by 18S and 16S rDNA sequence analysis, of keratitis caused by A. triangularis of which the type strain was isolated from human feces. Six isolates of A. triangularis had been reported from contaminated contact lens cases in southeastern Korea.

Molecular characterization of bacterial endosymbionts of Acanthamoeba isolates from infected corneas of Korean patients

The endosymbionts of 4 strains of Acanthamoeba (KA/E9, KA/E21, KA/E22, and KA/E23) isolated from the infected corneas of Korean patients were characterized via orcein stain, transmission electron microscopic examination, and 16S rDNA sequence analysis. Double membrane-bound, rod-shaped endosymbionts were distributed randomly throughout both the trophozoites and cysts of each of Acanthamoeba isolates. The endosymbionts of KA/E9, KA/E22, and KA/E23 were surrounded by electron-translucent areas. No lacunae-like structures were observed in the endosymbionts of KA/E21, the bacterial cell walls of which were studded with host ribosomes. Comparative analyses of the 16S rDNA sequences showed that the endosymbionts of KA/E9, KA/E22 and KA/E23 were closely related to Caedibacter caryophilus, whereas the KA/E21 endosymbiont was assigned to the Cytophaga-Flavobacterium-Bacteroides (CFB) phylum. In the 4 strains of Acanthamoeba, the hosts of the endosymbionts were identified as belonging to the Acanthamoeba castellanii complex, which corresponds to the T4 genotype. Acanthamoeba KA/E21 evidenced characteristics almost identical to those of KA/E6, with the exception of the existence of endosymbionts. The discovery of these endosymbionts from Acanthamoeba may prove essential to future studies focusing on interactions between the endosymbionts and the amoebic hosts.

Molecular characterization of Acanthamoeba isolated from amebic keratitis related to orthokeratology lens overnight wear

In an effort to characterize, on the molecular scale, the Acanthamoeba initially isolated from the cornea of an amoebic keratitis patient associated with overnight-wear orthokeratology lens in Korea, we conducted mitochondrial DNA restriction fragment length polymorphism, 18S rDNA sequencing, and drug sensitivity analyses on the isolate (KA/PE1). The patient was treated with polyhexamethylene biguanide, chlorhexidine and oral itraconazole, which resulted in resolution of the patient's ocular inflammation. The majority of the molecular characteristics of the KA/PE1 were determined to be identical, or quite similar, to those of A. castellanii Ma strain, which had been isolated also from amoebic keratitis. The risk of Acanthamoeba keratitis as a potential complication of overnight orthokeratology is briefly discussed.