Development and bench validation of real-time reverse transcription polymerase chain reaction protocols for rapid detection of the subtypes H6, H9, and H11 of avian influenza viruses in experimental samples.
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Real-time reverse transcription polymerase chain reaction (RRT-PCR) is commonly used for the rapid detection, as well as to determine the subtype, of avian influenza viruses (AIVs). There are 16 known serologically distinct hemagglutinin (HA) subtypes of AIV described. Currently, determination of the subtypes of AIVs by RRT-PCR tests has been limited to the H5 and H7 subtypes. In this study, RRT-PCR assays were developed in simplex formats for rapid detection of AIV subtypes H6, H9, and H11. The primers and probes for RRT-PCR were designed from nucleotide sequences of the HA genes, which were either downloaded from GenBank (for H6 and H9) or sequenced for this study. The specificity and sensitivity of the RRT-PCR assays were determined based on the detection of the virus from a proficiency panel consisting of 15 different HA subtypes of AIVs and from serial dilutions of target viral RNA. The subtype-specific RRT-PCR assays were used to detect the virus in cloacal and oropharyngeal swabs of experimental chickens inoculated with H6, H9, and H11 AIVs, and the test results were compared with validated RRT-PCR assays based on the amplification of AI matrix (MA) gene. A high correlation of the matrix test and the specific H6, H9, and H11 by the RRT-PCR assays was observed; kappa coefficients for the agreement of test results in cloacal and oropharyngeal swabs combined were 0.927, 0.962, and 0.981, respectively. (+info)
Active transport of sodium by the malpighian tubules of the tsetse fly Glossian morsitans.
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Isolated Malpighian tubules of Glossina morsitans are able to transport sodium against its concentration gradient. Their rate of secretion is dependent on the sodium concentration of the bathing medium. Potassium must be present in the bathing solution for rapid secretion to be maintained, but it does not play an active role in fluid secretion. Lithium and ammonium ions are able to substitute partially for sodium, other monovalent cations cannot. Ouabain does not affect rapid secretion by Glossina tubules in vitro. Conclusions drawn from the results are incorporated into a model of Malpighian tubule cell function in this insect. (+info)
Pathogenesis of digestive tract lesions in duck plague.
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White Pekin ducklings were inoculated orally with duck plague virus. Tissues from the digestive tract were collected daily after inoculation and examined by light, electron and fluorescent microscopy. There were necrosis and degeneration of stratified squamous epithelium of the esophagus and cloaca, epithelium of intestinal crypt and esophageal submucosal glands, macrophages in the lamina propria, and submucosal fibrocytes and lymphocytes. Submucosal hemorrhages occurred after degeneration and necrosis of lymphocytes, macrophages, fibrocytes and epithelial cells. Viral antigens were detected in all these cells by use of fluorescein-labeled antibodies. With the electron microscope, nucleocapsids were seen in the nuclei, budding through the inner nuclear membrane; enveloped virions were present in cytoplasmic vacuoles of macrophages, epithelial cells and fibrocytes. In lymphocytes, nucleocapsids were also in the nuclei, but karyorrhexis and cytolysis occurred before viral maturation was completed. (+info)
Role of poultry meat in sporadic Campylobacter infections in Bosnia and Herzegovina: laboratory-based study.
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AIM: To investigate genetic diversity and specificity of Campylobacter jejuni and Campylobacter coli strains isolated from humans, retail poultry meat, and live farm chickens in Zenica-Doboj Canton, Bosnia and Herzegovina, and identify the role of poultry meat in sporadic Campylobacter infections. METHODS: We determined the type of Campylobacter species using standard microbiological methods and multiplex polymerase chain reaction (PCR), and performed pulsed field gel-electrophoresis (PFGE) and restriction fragment length polymorphism (RFLP) typing of the flaA gene to investigate genetic diversity among the isolates. RESULTS: We isolated C jejuni and C coli from 75 (5.2%) of 1453 samples of consecutive outpatients with sporadic diarrhea; from 51 (34.7%) of 147 samples of poultry meat; and from 15 out of 23 farm chicken samples. The proportion of C coli found among human (30.1%), poultry meat (56.9%), and farm chicken isolates (53.3%), was greater than the proportion of C jejuni. Fourteen and 24 PFGE genotypes were identified among 20 C coli and 37 C jejuni isolates, respectively. Identical PFGE genotypes were found in two cases of human and poultry meat isolates and two cases of poultry meat and farm chicken isolates. CONCLUSION: Only a minority of human Campylobacter isolates shared identical PFGE type with poultry meat isolates. Although poultry is the source of a certain number of human infections, there may be other more important sources. Further research is required to identify the environmental reservoir of Campylobacter spp responsible for causing human disease and the reason for the high prevalence of C coli human infections in this region. (+info)
Prenatal diagnosis of persistent cloaca associated with VATER (vertebral defects, anal atresia, tracheo-esophageal fistula, and renal dysplasia).
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The cloaca is a single canal from which the urinary, genital, and intestinal tracts arise around gestational weeks 5-6. Persistent cloaca can result from cystic mass formation within the pelvis, which is commonly association with multiple developmental defects. VATER association, which is a spectrum of anomalies, manifested by vertebral defects, anal atresia, tracheo-esophageal fistula with esophageal atresia, and renal dysplasia, arises from abnormalities in mesodermal differentiation. Recently, both conditions have been proposed to represent a continuous spectrum of anomalies, but the pathophysiology concerning the continuity of the development and the clinical condition are still unclear. Since renal failure becomes a serious problem after birth, timely infant delivery is essential to avoid loss of renal function. We report a patient, in whom the overlap between these two conditions was identified, and renal function was lost from one kidney. A polycystic mass was found in the fetal abdomen at 26 weeks of gestation. By ultrasonography, we detected a polycystic left kidney, a single umbilical artery, a ventricular septal defect, an esophageal atresia, ascites, an anal atresia, and a cystic mass with debris behind the bladder. The left kidney was non-functioning and the right kidney showed signs of hydronephrosis at 30 weeks of gestation. We measured the size and the blood flow of renal artery sequentially, and could deliver the fetus before the function was lost from the right kidney. Our observations will help inform future patients where prompt intervention can help improve renal function and infant health. (+info)
Disease associated with integumentary and cloacal parasites in tadpoles of northern red-legged frog Rana aurora aurora.
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A total of 6830 northern red-legged frog Rana aurora aurora tadpoles were examined under a dissecting microscope for oral disc, integumentary, and cloacal abnormalities in 13 ponds in and near Redwood National Park in northern California. Of these, 163 tadpoles were collected for histopathological investigation, including 115 randomly collected individuals, 38 collected with oral disc abnormalities, and 10 collected due to severe morbidity of unknown etiology. The tadpoles were infected with 8 parasites, including Batrachochytrium dendrobatidis (the amphibian chytrid), trematodes, leeches, and protozoa. Chytridiomycosis was detected at an overall prevalence of 6.4%, but prevalence was higher in tadpoles with oral disc lesions than in those with normal oral discs (43.5% versus 6.1%). Interestingly, infection was associated with some environmental and co-infection risk factors. Individual tadpoles possessed 0 to 5 species of parasites in varying intensities. Apiosoma sp. was the most prevalent (66%) and widespread. Tadpoles infected with B. dendrobatidis had a lower diversity of oral parasites than those uninfected. During the field portion of the study, a large number (approximately 500) of moribund and dead tadpoles was seen occurring at multiple locations within and surrounding Redwood National Park. Ten animals were collected for histological examination and a diverse protozoal infection was discovered, including some known pathogens of fish. This study is the first reporting parasitism and disease in natural populations of northern red-legged frogs. (+info)
Diversity of extended-spectrum beta-lactamases and class C beta-lactamases among cloacal Escherichia coli Isolates in Belgian broiler farms.
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