Effects of compression force on elasticity index and elasticity ratio in ultrasound elastography.

OBJECTIVES: The purpose of this study was to investigate the relationship between compression force and hardness values in ultrasound elastography. METHODS: Ultrasound elastography was performed using an elastography phantom, comprising inclusions with different elasticities and echogenicities. The compression force was set to approximately 100 gw (light force) and approximately 500 gw (heavy force). The elasticity index (EI) of the inclusion was measured. The EI was a relative hardness value of a structure within an elastographic image. Similarly, the EI of the background was measured as a reference. The elasticity ratio (ER) was calculated as the EI of the inclusion divided by the EI of the reference. RESULTS: The hardness of the phantom could be discerned with both the EI and ER, regardless of the compression force. The EI and ER with heavy force tended to be higher than those with light force, but the difference was not significant. A strong correlation was observed between the EI and ER of soft structures, whereas the correlation between the EI and ER of hard structures was weak, and the ER values varied widely. CONCLUSIONS: The EI offers potential as a good indicator for assessing the hardness.

Nationwide surveillance of West Nile virus targeting mosquitoes and dead birds from April 2004 through March 2007 in Japan.

We conducted nationwide West Nile virus (WNV) surveillance targeting mosquitoes and dead birds to reveal whether the virus and its potential vectors are present in Japan. A total of 12 766 mosquitoes and 230 dead birds were collected in April 2004-March 2005 (the 2004-2005 period), 10 755 mosquitoes and 267 dead birds in April 2005-March 2006 (the 2005-2006 period), and 8624 mosquitoes and 245 dead birds in April 2006-March 2007 (the 2006-2007 period). The species of most of the mosquitoes collected over the 3 years were Culex tritaeniorhynchus (47.82%) and Anopheles sinensis (28.49%), and other species included Aedes albopictus (6.75%), the Culex pipiens group (Cx. pipiens pallens and Cx. pipiens molestus: 5.37%), Aedes vexans nipponii (2.54%), Armigeres subalbatus (1.08%), and Aedes japonicus (0.95%). As for the dead birds, most were Passeriformes (456 specimens), which included several crow species, and the other orders included Anseriformes, Columbiformes and Ciconiiformes (78, 66 and 36 specimens, respectively). All the specimens tested negative for WNV RNA by reverse-transcriptase polymerase chain reaction (RT-PCR) in the 2004-2005 period and by real-time RT-PCR in the 2005-2006 and the 2006-2007 periods, respectively. Our surveillance provided no evidence for WNV in Japan as of the end of the surveillance period, but on the other hand, it revealed that several species of potential WNV vectors are distributed widely in Japan, which suggests that WNV in principle could be transmitted by the potential vectors if introduced. Thus, it is essential to take continued precautions against WNV introduction.

Molecular characterization of a troponin I-like protein from the hard tick Haemaphysalis longicornis.

A cDNA expression library prepared from mRNA of Haemaphysalis longicornis (H. longicornis) was screened with a H. longicornis-infested rabbit serum. A cDNA encoding 27/30kDa proteins was cloned and designated P27/30 gene. The predicted amino acid sequence of the P27/30 gene shows a rather high homology (58% amino acid identities and 11% amino acid similarity) with Drosophila melanogaster troponin I clone E2. H. longicornis P27/30 possesses amino acid sequence of actin-binding domains of troponin I at the amino acid residues 128-148, suggesting that H. longicornis P27/30 is a troponin I-like protein. By immunoblot analysis, mouse anti-recombinant P27/30 serum reacted with major constituent protein bands in extracts of adult ticks, and also immunoreacted with muscle, cuticle, gut, and salivary gland in H. longicornis ticks. Moreover, immunohistochemistry using the anti-P27/30 serum showed a strong reactivity in muscle, suggesting that native P27/30 is expressed abundantly in that tissue.

Molecular characterization of a peroxiredoxin from the hard tick Haemaphysalis longicornis.

Antioxidant enzymes in eukaryotes play an important role in protection against the oxygen radicals generated during aerobic metabolism. Here we report the cloning and characterization of a cDNA encoding the antioxidant enzyme peroxiredoxin from the hard tick Haemaphysalis longicornis (HlPrx). HlPrx is 939 bp long and contains a 101 bp non-translated sequence at the 5' end and a polyadenylation singnal followed by a poly(A) tail at the 3' end. HlPrx encodes a full-length protein with a predicted molecular mass of 26 kDa that possesses one cysteine residue at amino acid 49 that is conserved among Prx proteins of various species. GenBanktrade mark analysis showed that the deduced amino acid sequence had significant similarity to mammalian and plant Prxs at the amino acid level. A DNA-nicking assay revealed that Escherichia coli-expressed recombinant HlPrx (rHlPrx) inhibited oxidative-nicking of supercoiled plasmid DNA. Two-dimensional immunoblot analysis with mouse antirHlPrx serum showed reaction with a major constituent protein spot in extracts of adult ticks. In addition, immunoblot analysis showed that rHlPrx was immunoreacted with serum from rabbits repeatedly infested with H. longicornis. Localization analysis using mouse antirHlPrx serum revealed that native HlPrx was highly expressed in the salivary gland of the tick. Moreover, Northern blot analysis showed that the level of HlPrx transcripts was increased during blood sucking. The present results indicate that HlPrx may be an important detoxifying enzyme during the normal life span as well as during blood sucking in ticks.

Analysis of the 16S rRNA gene sequence of Anaplasma centrale and its phylogenetic relatedness to other ehrlichiae.

The nucleotide sequence of the Anaplasma centrale 16S rRNA gene was determined and compared with the sequences of ehrlichial bacteria. The sequence of A. centrale was closely related to Anaplasma marginale by both level-of-similarity (98.08% identical) and distance analysis. A species-specific PCR was developed based upon the alignment data. The PCR can detect A. centrale DNA extracted from 10 infected bovine red blood cells in a reaction mixture. A. centrale DNA was amplified in the reaction, but not other related ehrlichial species.

[A case of a left atrial primary malignant fibrous histiocytoma].

Primary malignant fibrous histiocytoma (MFH) of the heart is very rere. We report a case of a primary intracardiac MFH. A 50-year-old woman underwent an operation for congestive heart failure due to left atrial (LA) tumor. Cardiac echocardiogram showed the tumor attached to the septal wall and posterior wall of LA. We extendedly resected the tumor with septal and partially posterior wall and reconstructed there with pericardial substitute. Pathological diagnosis was MFH. Although she did well postoperatively, cardiac echocardiogram showed a recurrent tumor in the LA at 10 days after operation. She developed respiratory failure, and chest and abdominal CT demonstrated metabolism to the lung, liver and bladder at post operative 47 days. At 77 days after operation, she expired. The prognosis of MFH is poor despite surgery. We recommend extended surgery, if the LA tumor had wide base attached to the septal or posterior wall.

Extended high-frequency ototoxicity induced by the first administration of cisplatin.

The ototoxicity induced by cisplatin results in high-frequency hearing loss. The sound-pressure thresholds at extended high frequencies of 8 to 20 kHz were measured in 12 patients with head and neck cancer before and after the first administration of cisplatin. Ototoxicity was defined statistically by the newly introduced regression-line analysis. This analysis revealed that the threshold of 1 of the 12 patients increased evenly from 8 to 10 kHz, and the damage to the hearing system of this patient was permanent. The reasons for such a low rate of ototoxicity might be the long-term administration of low doses of cisplatin and the addition of fosfomycin.

Phylogenetic relationships of the benign Theileria species in cattle and Asian buffalo based on the major piroplasm surface protein (p33/34) gene sequences.

In order to examine the taxonomic relationship of Theileria sp. of Asian buffalo to the benign Theileria spp. of cattle, we sequenced and compared the major piroplasm protein (p33/34) genes of these parasites. The two consensus sequences determined for the buffalo parasite were of the same length (852 bp) and showed >80% identity with the sequences of the homologous genes (849 bp) in the cattle parasites. Alignment of the inferred aa sequences with those of Theileria sergenti and Theileria buffeli predicted that there is an insertion of a single residue at the N-terminus in the inferred polypeptide of the buffalo parasite. Phylogenetic analyses based on the aa sequences suggested that Theileria sp. of the Asian buffalo should be classified within the benign Theileria parasite group as a separate species from the cattle parasites. Based on this, we propose a rearrangement of the currently used classification for the benign Theileria species in cattle and Asian buffalo.

Molecular phylogenetic studies on Theileria parasites based on small subunit ribosomal RNA gene sequences.

Classification of Theileria parasites of south-east Asian countries is still ambiguous due to the lack of basic studies, especially their molecular genetic information. In this study, we included 6 known species and 14 unclassified Theileria parasite isolates: Theileria annulata, Theileria parva, Theileria taurotragi, Theileria sergenti, Theileria buffeli, Theileria types Sable, Theileria types A, B, B1, B2, C, D, E, F, G, G1, Theileria type Medan (Indonesia), Theileria type Ipoh (Malaysia) and Theileria type Thong Song (Thailand). Small subunit ribosomal RNA (srRNA) nucleotide sequence data were collected by PCR, cloning and dideoxy sequencing. The srRNA nucleotide sequences were aligned and analyzed by distance methods, maximum parsimony algorithms and maximum likelihood methods to construct phylogenetic trees. Bootstrap analysis was used to test the strength of the different phylogenetic reconstructions. The data indicated that all of the tree-building methods gave very similar results. This study identified two groups of Theileria, the pathogenic and benign groups, which are strongly supported by bootstrap analysis. The analysis also indicated that three subgroups (A, B and C) were generated within the benign Theileria group whereas the classification of Theileria type D and Thong Song is questionable. However, more basic information such as life cycle differences, vectors, modes of transmission, virulent and genetic/sexual compatability is essential for clearer taxonomic definition of the benign Theileria parasites.

New Prognostic Factors Associated with Long-term Survival in Node-Negative Breast Cancer Patients.

BACKGROUND: This study was undertaken to determine the absolute and relative value of angiogenesis, proliferating cell nuclear antigen (PCNA) and conventional prognostic factors in predicting relapse-free survival (RFS) and overall survival (OS) rates associated with long-term survival in Japanese patients with node-negative breast cancer. PATIENTS AND METHODS: Two hundred patients with histological node-negative breast cancer were studied. We investigated nine clinicopathological factors, including angiogenesis, PCNA using per-manent-section immunohistochemistry, clinicaltumor size, histological grade (HG), tumor necrosis, lymphatic vessel invasion (LVI), histological extension, histological classification, and infiltrating growth (INF), followed for a median of 10 years (range, 1 to 20). RESULTS: Twenty-one patients (10.5%) had recurrence and 15 patients (7.5%) died of breast cancer. Univariate analysis showed that PCNA, clinical tumor size, HG, angiogenesis, and LVI were significantly predictive of 20-year RFS or OS. Tumor necrosis was significantly predictive of OS, not of RFS. Multi-variate analysis showed that clinical tumor size (P = 0.0003), angiogenesis (P = 0.0003), PCNA (P = 0.0064), and HG (P = 0.0401) were significant independent prognostic factors for RFS. PCNA (P< 0.0001) and clinical tumor size (P = 0.0112) were significant independent prognostic factors for OS, while angiogenesis was a borderline significant factor. CONCLUSION: PCNA and angiogenesis were important new prognostic factors in node-negative breast cancer patients.

DNA polymorphism of srRNA gene among Eimeria tenella strains isolated in Japan.

DNA polymorphism in twelve starains of Eimeria tenella isolated from various places in Japan was examined using 1.l kb small subunits ribosomal RNA amplified by PCR. Genetic variation was evaluated by random amplification of polymorphic DNA (RAPD) analysis. DNA fingerprint patterns were grouped into two, indicating that at least two DNA polymorphisms exist in Japanese E. tenella strains.

Inter-species differentiation of benign Theilerias by genomic fingerprinting with arbitrary primers.

A method was developed to obtain reproducible DNA fingerprints from five distinct purified benign Theileria genomic DNAs by PCR-based amplification. Randomly amplified polymorphic DNA (RAPD) profiles were obtained from 10 randomly designed 12-mers. However, nine of the 10 primers could generate the difference in RAPD-PCR profiles which allowed discrimination of Theileria species. The method has advantage of being simple, fast and sensitive for diagnosis and characterization of the parasites since it does not require prior DNA sequence information to construct species-specific probes or primers. The results are also beneficial for a proper understanding of the epidemiology and designing rational control programmes for Theileriosis in Asian and South-East Asian countries.

Average thresholds in the 8 to 20 kHz range in young adults.

The sound-pressure level thresholds in the extended high-frequency range (8 to 20 kHz) were measured in 25 non-hearing-impaired young adults from 20 to 29 years of age. The result was not unlike that obtained by previous investigators; the thresholds increased gradually as a function of frequency. However, two notable points were found: one that the threshold reached a plateau above 18 kHz, and the other that it decreased slightly at 12 kHz. As the subjects might respond to the low-frequency noise of the stimulus wave, the threshold became a plateau above 18 kHz. An acoustic resonance in the ear canal caused the threshold to decrease at 12 kHz. In clinical studies of extended high-frequency audiometry, the threshold data should be carefully evaluated above 18 kHz and at 12 kHz.

Average thresholds in the 8 to 20 kHz range as a function of age.

The sound-pressure thresholds at the extended high frequencies of 8-20 kHz were measured for 65 normal subjects aged between 10 and 69 years. The results are not unlike those obtained by previous investigators. The thresholds increased gradually as a function of frequency, except around 12 kHz and above 19 kHz, and also as a function of age. To clarify the connections between threshold, frequency and age, we introduced the regression lines for the threshold by analysing two ranges of frequencies (8-10 kHz and 14-19 kHz) and determining their slopes and intercepts. The regression line analysis reveals that the thresholds at 8-10 kHz tend to increase more at higher frequencies as subject age increased above 30 to 39 years, and those at 14-19 kHz increase translationally with increase of age. Our results did not contradict earlier reports on the pathological changes of the inner ear.

A polymorphism observed in the experimentally successful peptide vaccine sequence derived from Theileria sergenti piroplasm major surface antigen (p33).

A polymorphism in the experimentally successful peptide vaccine sequence (EVVWKEKKEVKDLDA, amino acids 134-148) derived from the 33 kDa piroplasm major surface antigen (p33) of Theileria sergenti was examined. The vaccine sequences obtained by PCR amplification and sequencing of the p33 gene from a total of 15 parasite-infected cattle blood samples collected from 4 prefectures through Hokkaido to Kumamoto revealed the two major sequences (Ikeda and Chitose stock types) either of which was identified in all samples. Since the peptide vaccine develops the parasite species- or stock-specific immunity in the animals, an application of the two major peptide sequences as cocktailed vaccine should be evaluated for a practical use of this strategy to controlling T. sergenti infection in Japan.