Clinical and laboratory features of PCR-confirmed periocular tuberculosis in China

Experts lack knowledge of periocular tuberculosis（TB） in China. Nested polymerase chain reaction（PCR） shows advantages in diagnosis of extrapulmonary TB. Our study aims to explore the clinical and laboratory features of PCR-confirmed periocular TB. We retrospectively reviewed medical records of presumptive periocular TB and performed nested PCR test to confirm diagnosis. Nine cases were recruited. Clinical symptoms were chronic and insidious. Eight cases achieved favorable visual acuity, while one underwent enucleation due to fungalTB panophthalmitis. Sensitivity of caseous necrosis, acidfast bacilli（AFB） staining and interferon γ release assay（T-SPOT） test are 33.3%, 44.4% and 85.7% respectively. Low lymphocyte percentage（P=0.019） and high monocytelymphocyte ratio（P=0.042） positively correlate with AFB staining. Male gender（P=0.048） and Langhans giant cell（P=0.048） positively correlate with caseous necrosis. To conclude, traditional TB ancillary tests are not as sensitive as nested PCR technique. Several factors facilitate diagnosis including male gender, decreased lymphocytes, and typical Langhans giant cells.

DNA diagnostics for reliable and universal identification of Helicobacter pylori

Reliable diagnostics are a major challenge for the detection and treatment of Helicobacter pylori(H.pylori)infection.Currently at the forefront are non-invasive urea breath test(UBT)and stool antigen test(SAT).Polymerase chain reaction(PCR)is not endorsed due to nonspecific primers and the threat of false-positives.The specificity of DNA amplification can be achieved by nested PCR(NPCR),which involves two rounds of PCR.If the primers are properly designed for the variable regions of the 16S rRNA gene,it is not difficult to develop an NPCR assay for the unambiguous identification of H.pylori.Elaborate NPCR for a 454 bp amplicon was validated on 81 clinical biopsy,stool,and saliva samples,each from the same individuals,and compared with available H.pylori assays,namely histology,rapid urease test,SAT,and 13C-UBT.The assay was much more sensitive than simple PCR,and it was equally sensitive in biopsy samples as the 13CUBT test,which is considered the gold standard.In addition,it is sufficiently specific because sequencing of the PCR products exclusively confirmed the presence of H.pylori-specific DNA.However,due to the threshold and lower abundance,the sensitivity was much lower in amplifications from stool or saliva.Reliable detection in saliva also complicates the ability of H.pylori to survive in the oral cavity aside from and independent of the stomach.The reason for the lower sensitivity in stool is DNA degradation;therefore,a new NPCR assay was developed to obtain a shorter 148 bp 16S rRNA amplicon.The assay was validated on stool samples from 208 gastroenterological patients and compared to SAT results.Surprisingly,this NPCR revealed the presence of H.pylori in twice the number of samples as SAT,indicating that many patients are misdiagnosed,not treated by antibiotics,and their problems are interpreted as chronic.Thus,it is unclear how to properly diagnose H.pylori in practice.In the first approach,SAT or UBT is sufficient.If samples are negative,the 148 bp amplicon NPCR assay should be performed.If problems persist,patients should not be considered negative,but due to threshold H.pylori abundance,they should be periodically tested.The advantage of NPCR over UBT is that it can be used universally,including questionable samples taken from patients with achlorhydria,receiving proton pump inhibitors,antibiotics,bismuth compound,intestinal metaplasia,or gastric ulcer bleeding.

Y Specific Sequence Gene Analysis of Single Fetal Nucleated Erythroblasts from the Peripheral Blood of Pregnant Women

The single cell isolation technique was used to detect fetal nucleated erythroblasts at a single cell level from the peripheral blood of pregnant women in order to investigate the feasibility of this method for noninvasive prenatal diagnosis. Single fetal nucleated erythroblasts were isolated from the peripheral blood samples from 51 pregnant women by micromanipulation techniques after density gradient centrifugation. Nested polymerase chain reaction method was used to amplify the SRY gene. It was found that the concordance rate of amplification results with real fetal sex was 82.61 %. The sensitivity and specificity were 80 % and 87.50 % respectively. It was suggested that it is feasible and promising in non invasive prenatal diagnosis to detect fetal nucleated erythroblasts at a single cell level by using micromanipulation techniques.