Assessment of Zoonotic Transmission of Giardia and Cryptosporidium between Cattle and Humans in Rural Villages in Bangladesh

Giardia and Cryptosporidium are important causes of diarrhoea in Bangladesh. The high prevalence of both parasites in humans and cattle in rural Bangladesh and the common use of water ponds by village inhabitants and their animals suggest a potential for zoonotic transmission. Direct transmission of Giardia and Cryptosporidium between cattle and their handlers and indirect transmission through water ponds was investigated. Faecal/stool samples were collected from 623 calves and 125 calf handlers in a cross-sectional survey. In two villages, water samples were collected monthly from water ponds and faecal/stool samples were collected monthly from inhabitants and their cattle. Giardia cysts and Cryptosporidium oocysts were detected in water samples and in faecal/stool samples and positive samples were genotyped, to determine their human or animal origin. The prevalence of Giardia and Cryptosporidium in calves was 22% and 5% respectively. In calf handlers, the prevalence of Giardia and Cryptosporidium was 11.2% and 3.2% respectively. Both in the cross-sectional survey and in the longitudinal study in the villages, G. duodenalis assemblage E was most prevalent in calves, while in humans assemblage AII, BIII and BIV were found. In cattle, Cryptosporidium parvum, C. bovis and C. andersoni were identified, but no Cryptosporidium sequences were obtained from humans. Giardia and Cryptosporidium were detected in 14/24 and 12/24 water samples respectively. G. duodenalis assemblage E and BIV (-like), as well as C. andersoni and C. hominis were identified. Although the presence of Giardia and Cryptosporidium in both water ponds suggests that water-borne transmission of Giardia and Cryptosporidium is possible, the genotyping results indicate that there is no significant direct or indirect (water-borne) transmission of Giardia between cattle and people in this area of rural Bangladesh. No conclusions could be drawn for Cryptosporidium, because of the low number of sequences that were obtained from human and water samples.     

HER2+ mCRC patients with exon 20 R784G substitution mutation do not respond to the cetuximab therapy

The human epidermal growth factor 2 (HER2) gene undergoes various mutations that could alter its activity or respond to the antibody therapies. Cetuximab, a known anti-EGFR monoclonal antibody (mAB), is widely administered in metastatic colorectal cancer (mCRC) cases. Here we identified mCRC patients who did not respond to cetuximab (500 mg/m², q2w) after fluoropyrimidine/oxaliplatin regimen failure. Tumor samples were examined with immunohistochemistry for protein distribution, polymerase chain reaction (PCR) sequencing for mutation detection and real-time PCR for mRNA expression pattern analysis between cetuximab sensitive and resistance patients. The conformational differences of normal and mutated protein structures were predicted by bioinformatics analysis. The 5-year survival rates of target groups were estimated using the Kaplan–Meier method. Immunohistochemistry showed that all cases had high level of HER2 protein. No K-Ras or B-Raf mutation was observed among the study population; however, cetuximab resistance patients harbored a somatic mutation R784G at the exon 20 region of HER2 coding sequence. According to bioinformatics analysis, this mutation caused a notable misfold in protein conformation. Meanwhile, survival analysis showed R784G mutated mCRC patients had shortened survival rate compared with the mCRC cases with wild-type HER2. Collectively, these data report a new mechanism of resistance to cetuximab and might be applicable in modifying new therapeutic strategies for HER2 involved cancers.     

In vivo SPECT reporter gene imaging of regulatory T cells

Regulatory T cells (Tregs) were identified several years ago and are key in controlling autoimmune diseases and limiting immune responses to foreign antigens, including alloantigens. In vivo imaging techniques including intravital microscopy as well as whole body imaging using bioluminescence probes have contributed to the understanding of in vivo Treg function, their mechanisms of action and target cells. Imaging of the human sodium/iodide symporter via Single Photon Emission Computed Tomography (SPECT) has been used to image various cell types in vivo. It has several advantages over the aforementioned imaging techniques including high sensitivity, it allows non-invasive whole body studies of viable cell migration and localisation of cells over time and lastly it may offer the possibility to be translated to the clinic. This study addresses whether SPECT/CT imaging can be used to visualise the migratory pattern of Tregs in vivo. Treg lines derived from CD4(+)CD25(+)FoxP3(+) cells were retrovirally transduced with a construct encoding for the human Sodium Iodide Symporter (NIS) and the fluorescent protein mCherry and stimulated with autologous DCs. NIS expressing self-specific Tregs were specifically radiolabelled in vitro with Technetium-99m pertechnetate ((99m)TcO(4)(-)) and exposure of these cells to radioactivity did not affect cell viability, phenotype or function. In addition adoptively transferred Treg-NIS cells were imaged in vivo in C57BL/6 (BL/6) mice by SPECT/CT using (99m)TcO(4)(-). After 24 hours NIS expressing Tregs were observed in the spleen and their localisation was further confirmed by organ biodistribution studies and flow cytometry analysis. The data presented here suggests that SPECT/CT imaging can be utilised in preclinical imaging studies of adoptively transferred Tregs without affecting Treg function and viability thereby allowing longitudinal studies within disease models.     

Retinoic acid and/or progesterone differentiate mouse induced pluripotent stem cells into male germ cells in vitro

Numerous reagents were employed for differentiating induced pluripotent stem cells (iPSCs) into male germ cells; however, the induction procedure was ineffective. The aim of this study was to improve the in vitro differentiation of mice iPSCs (miPSCs) into male germ cells with retinoic acid (RA) and progesterone (P). miPSCs were differentiated to embryoid bodies (EBs) in suspension with RA with or without progesterone for 0, 4, and 7 days. Then, the expression of certain genes at different stages of male germ cell development including Ddx4 (pre meiosis), Stra8 (meiosis), AKAP3 (post meiosis), and Mvh protein was examined in RNA and/or protein levels by real-time polymerase chain reaction or flow cytometry, respectively. The Stra8 gene expression increased in the RA groups on all days. But, expression of this gene declined in RA + P groups. In addition, an increased expression of Ddx4 gene was observed on day 0 in the P group. Also, a significant upregulation was observed in the expression of AKAP3 gene in the RA + P group on days 0 and 4. However, gene expression decreased in P and RA groups on day 7. The expression of Mvh protein significantly increased in the RA group on day 7. The Mvh expression was also enhanced in the P group on day 4, but it decreased on day 7, while this protein upregulated on day 0 and 7 in the RA + P group. The miPSCs have the capacity for in vitro differentiation into male germ cells by RA and/or progesterone. However, the effects of these inducers depend on the type of combination and an effective time.     

Brucellosis Causes Alteration in Trace Elements and Oxidative Stress Factors

Brucellosis is regarded as one of the most common diseases among humans and livestock. In the present study, we aimed to assess the effect of this disease on the level of various cations including copper (Cu), manganese (Mn), zinc (Zn), and magnesium (Mg) as well as oxidative stress status in the serum of people suffering from brucellosis. The present case-study was carried out on 40 patients with brucellosis (case) and 20 healthy people (control). Blood specimens were taken from all the people and the level of essential trace elements and oxidative stress status were measured. The serum level of copper in the case group (165.39 ± 43.19 μg/dl) was significantly higher compared with that in the control group (122.12 ± 28.88 μg/dl). Whereas the serum level of zinc was significantly lower in the case group compared with that in the control group (76.47 ± 28.88 vs. 92.85 ± 23.16 μg/dl). The manganese and magnesium serum levels did not differ significantly between the two groups. Furthermore, total antioxidant capacity level was significantly lower in the case group (122.12 ± 28.22 μmol/ml) than that in the control group (3.08 ± 0.12 μmol/ml) and the level of serum malondialdehyde was significantly higher in the case group (7.20 ± 0.23 mmol/ml) than that in the control group (4.0 ± 0.19 mmol/ml). Brucellosis can cause alteration in the serum level of essential trace elements. Moreover, the present study indicated that brucellosis produces oxidative stress in patients.     

Implementation of fed-batch strategies for vitamin K (menaquinone-7) production by Bacillus subtilis natto in biofilm reactors

Recent studies show the essential health benefits associated with vitamin K, especially menaquinone-7 (MK-7). These benefits include reducing risks of cardiovascular diseases, osteoporosis, and even cancer. However, MK-7 production on an industrial level is only possible through bacterial fermentation and also current static fermentation strategies are not potent enough with difficulties to scale up. Biofilm reactors, however, may be a practical alternative. Biofilm reactors provide a controlled environment for the microorganisms to form mature and robust biofilms that enable them to produce value-added products with enhanced efficiencies. In this study, fed-batch addition of glucose and glycerol were investigated to the base media in biofilm reactors, as carbon source addition seemed crucial in batch fermentations. Results indicated that fed-batch strategies can be significantly effective in glucose-based medium, increasing the end-product concentrations to 28.7 ± 0.3 mg/L of MK-7 which was 2.3 fold higher than the level produced in suspended-cell bioreactors and renders the biofilm reactors as a potential replacement for static fermentation strategies. Moreover, morphological changes of B. subtilis were tracked during the 12-day long fermentation runs and finally, SEM investigations confirmed significant biofilm and extracellular matrices formed on the plastic composite support (PCS) in the biofilm reactors. In conclusion, biofilm reactors especially with fed-batch fermentation regimes seem to be an effective tool for MK-7 production at industrial scales.