Detection of Cryptosporidium and Giardia in agricultural and water environments in the Qinghai area of China by IFT and PCR

Qinghai Province in northwest China is strongly influenced by agricultural activities and is an important source of food and drinking water. Here, we present findings regarding the occurrence and molecular epidemiology of Cryptosporidium and Giardia species based on a large-scale investigation of areas of Qinghai Province. The diagnosis and molecular detection of Cryptosporidium oocysts and Giardia cysts was carried out using immunofluorescence microscopy (IFT), whereas nested polymerase chain reaction (PCR) in fecal smears and water samples was used for the detection and molecular characterization of the species. In total, 561 samples (260 water samples and 301 fecal samples from animals) were collected and analyzed. Of the 260 water samples, 66 samples were Cryptosporidium-positive by IFT and 71 samples were positive by nested PCR; in addition, 39 samples were Giardia-positive by IFT and 40 samples were positive by nested PCR. Of the 301 fecal samples from animals, 98 samples were Cryptosporidium-positive by IFT and 61 samples were positive by nested PCR, whereas 52 samples were Giardia-positive by IFT and 31 samples were positive by nested PCR. We showed that the water supplies and animals investigated contained Cryptosporidium and Giardia (oo)cysts. Thus, we recommend that the Chinese Government and Chinese health authorities undertake control measures to protect the food and drinking water sources in Qinghai from these pathogenic protozoa

Cystic fibrosis transmembrane conductance regulator trafficking modulates the barrier function of airway epithelial cell monolayers

The cystic fibrosis transmembrane conductance regulator (CFTR) is an integral membrane glycoprotein which functions as an anion channel and influences diverse cellular processes. We studied its role in the development of epithelial tightness by expressing wild-type (WT-CFTR) or mutant (ΔF508-CFTR) CFTR in human airway epithelial cell monolayers cultured at the air–liquid interface. Green fluorescent protein (GFP)-tagged WT or ΔF508 constructs were expressed in the CF bronchial cell line CFBE41o− using adenoviruses, and the results were compared with those obtained using CFBE41o− lines stably complemented with wild-type or mutant CFTR. As predicted, GFP-WT-CFTR reached the apical membrane whereas GFP-ΔF508-CFTR was only detected intracellularly. Although CFTR expression would be expected to reduce transepithelial resistance (TER), expressing GFP-CFTR significantly increased the TER of CFBE41o− monolayers whilst GFP-ΔF508-CFTR had no effect. Similar results were obtained with cell lines stably overexpressing ΔF508-CFTR or WT-CFTR. Preincubating ΔF508-CFTR monolayers at 29°C reduced mannitol permeability and restored TER, and the effect on TER was reversible during temperature oscillations. Expression of GFP-ΔF508-CFTR or GFP-WT-CFTR in a cell line already containing endogenous WT-CFTR (Calu-3) did not alter TER. The CFTR- and temperature-dependence of TER were not affected by the CFTR inhibitor CFTRinh172 or low-chloride medium; therefore the effect of CFTR on barrier function was unrelated to its ion channel activity. Modulation of TER was blunted but not eliminated by genistein, implying the involvement of tyrosine phosphorylation and other mechanisms. Modulation of CFTR trafficking was correlated with an increase in tight junction depth. The results suggest that CFTR trafficking is required for the normal organisation and function of tight junctions. A reduction in barrier function caused by endoplasmic reticulum retention of ΔF508-CFTR may contribute to fluid hyperabsorption in CF airways