Gut dysbiosis, inflammation and type 2 diabetes in mice using synthetic gut microbiota from diabetic humans

The study was aimed to assess impact of high fat diet (HFD) and synthetic human gut microbiota (GM) combined with HFD and chow diet (CD) in inducing type-2 diabetes (T2D) using mice model. To our knowledge, this is the first study using selected human GM transplantation via culture based method coupled dietary modulation in mice for in vivo establishment of inflammation leading to T2D and gut dysbiosis. Twenty bacteria (T2D1-T2D20) from stool samples of confirmed T2D subjects were found to be morphologically different and subjected to purification on different media both aerobically and anerobically, which revealed seven bacteria more common among 20 isolates on the basis of biochemical characterization. On the basis of 16S rRNA gene sequencing, these seven isolates were identified as Bacteroides stercoris (MT152636), Lactobacillus acidophilus (MT152637), Lactobacillus salivarius (MT152638), Ruminococcus bromii (MT152639), Klebsiella aerogenes (MT152640), Bacteroides fragilis (MT152909), Clostridium botulinum (MT152910). The seven isolates were subsequently used as synthetic gut microbiome (GM) for their role in inducing T2D in mice. Inbred strains of albino mice were divided into four groups and were fed with CD, HFD, GM+HFD and GM+CD. Mice receiving HFD and GM+modified diet (CD/HFD) showed highly significant (P<0.05) increase in weight and blood glucose concentration as well as elevated level of inflammatory cytokines (TNF-α, IL-6, and MCP-1) compared to mice receiving CD only. The 16S rRNA gene sequencing of 11 fecal bacteria obtained from three randomly selected animals from each group revealed gut dysbiosis in animals receiving GM. Bacterial strains including Bacteroides gallinarum (MT152630), Ruminococcus bromii (MT152631), Lactobacillus acidophilus (MT152632), Parabacteroides gordonii (MT152633), Prevotella copri (MT152634) and Lactobacillus gasseri (MT152635) were isolated from mice treated with GM+modified diet (HFD/CD) compared to strains Akkermansia muciniphila (MT152625), Bacteriodes sp. (MT152626), Bacteroides faecis (MT152627), Bacteroides vulgatus (MT152628), Lactobacillus plantarum (MT152629) which were isolated from mice receiving CD/HFD. In conclusion, these findings suggest that constitution of GM and diet plays significant role in inflammation leading to onset or/and possibly progression of T2D. .

Molecular detection of Mycoplasma gallisepticum in different poultry breeds of Abbottabad and Rawalpindi, Pakistan

The poultry sector in Pakistan is contributing mainly in bridging gap between demand and supply for protein. Mycoplasma gallisepticum is an emerging bacterium causing serious problems in poultry industry of Pakistan. A cross-sectional study was conducted to evaluate the M. gallisepticum load in poultry populated regions of Pakistan. Total 600 serum and 600 swab samples were collected, 200 from each broiler, layers and breeders poultry in Rawalpindi and Abbottabad districts. Serum samples were analyzed through ELISA for seroprevalence. Swabs were cultured on Frey’s medium followed by PCR and partial mgc2 gene sequencing. Results of seroprevalence of M. gallisepticum showed that layers (75%, n=150) are more positive as compared to breeders (70%, n=140) and broilers (50%, n=100). Typical colonies of the M. gallisepticum were observed in breeder (26.5%), followed by layer (21%) and broilers (9%). A total of 37.1% (n=42) samples were identified positive through PCR out of total 113 cultured based positive samples. A total of six M. gallisepticum isolates of current study showed 98-99 percent similarity with previously reported isolates on the basis of mgc2 gene partial sequencing. The M. gallisepticum was found highly prevalent in different poultry breads. Results of this study would add into basic data and provide a direction for livestock sector to strengthen a control strategy for mycoplasmosis in poultry farms.

Role of GDNF, GFRα1 and GFAP in a Bifidobacterium-Intervention Induced Mouse Model of Intestinal Neuronal Dysplasia

ObjectiveTo investigate the effects of glial cell-derived neurotrophic factor (GDNF), GDNF family receptor alpha 1 (GFRα1), and glial fibrillary acidic protein (GFAP) on colonic motility in a mouse model of intestinal neuronal dysplasia by intervention with Bifidobacterium and to explore the influence of Bifidobacterium on enteric glial cells (EGCs).MethodsWestern blotting and qRT-PCR were employed to detect the expression of GFRα1 and GFAP in colonic tissues of mice with or without Tlx2 mutations, and ELISA was used to detect the expression of GDNF in serum. IHC was used to detect the appearance of the ganglion cells. Subsequently, Tlx2 homozygous mutant (Tlx2−/−) mice were treated with Bifidobacterium. Colonic motility was measured before and after intervention by measuring the glass bead expelling time. The variations in abdominal circumference and GDNF, GFRα1, and GFAP expression were measured. In addition, 16SrRNA gene sequencing was performed to detect the abundance of the intestinal microbiota.ResultsThe mRNA and protein expression of GFRα1 and GFAP was decreased in the colonic tissues of Tlx2−/− mice and GDNF expression was decreased in serum compared with Tlx2+/− and WT mice. After confirming the colonization of Bifidobacterium by 16S rRNA gene sequencing, the expelling time and abdominal distension were ameliorated, and the expression of GFAP, GDNF, and GFRα1 was increased.ConclusionsThe expression of GDNF, GFRα1, and GFAP is associated with colonic motility. The altered expression of EGC-related factors suggested that Bifidobacterium may be involved in the EGC activation process. The amelioration of IND symptoms after intervention with Bifidobacterium prompted the elicitation of adjuvant therapy.

Cytauxzoon europaeus infections in domestic cats in Switzerland and in European wildcats in France: a tale that started more than two decades ago

Background Cytauxzoon spp. infection is believed to be a newly emerging tick-borne disease in felids in Europe, with three species of the haemoparasite having recently been differentiated in wild felids. In Switzerland, rare infections have been documented in domestic cats in the west and northwest of the country, the first of which was in 2014. The aims of the present study were: (i) to characterize a Cytauxzoon spp. hotspot in domestic cats in central Switzerland; (ii) to elucidate the geographic distribution of Cytauxzoon spp. in domestic cats in Switzerland; (iii) to assess suspected high-risk populations, such as stray and anaemic cats; and (iv) to investigate the newly emerging nature of the infection. Cytauxzoon spp. were further differentiated using mitochondrial gene sequencing. Methods The overall study included samples from 13 cats from two households in central Switzerland (study A), 881 cats from all regions of Switzerland (study B), 91 stray cats from a hotspot region in the northwest of Switzerland and 501 anaemic cats from across Switzerland (study C), and 65 Swiss domestic cats sampled in 2003 and 34 European wildcats from eastern France sampled in the period 1995–1996 (study D). The samples were analysed for Cytauxzoon spp. using real-time TaqMan quantitative PCR, and positive samples were subjected to 18S rRNA, cytochrome b (CytB) and cytochrome c oxidase subunit I (COI) gene sequencing. Results In study A, six of 13 cats from two neighbouring households in central Switzerland tested postive for Cytauxzoon spp.; two of the six infected cats died from bacterial infections. In studies B and C, only one of the 881 cats (0.1%; 95% confidence interval [CI]: 0–0.3%) in the countrywide survey and one of the 501 anaemic cats (0.2%; 95% CI: 0–0.6%) tested postive for Cytauxzoon spp. while eight of the 91 stray cats in the northwest of Switzerland tested positive (8.8%; 95% CI: 3.0–14.6%). In study D, Cytauxzoon spp. was detected in one of the 65 domestic cat samples from 2003 (1.5%; 95% CI: 0–4.5%) and in ten of the 34 European wildcat samples from 1995 to 1996 (29%; 95% CI: 14.2–44.7%). The isolates showed ≥ 98.6% sequence identities among the 18S rRNA, CytB and COI genes, respectively, and fell in the subclade Cytauxzoon europaeus based on CytB and COI gene phylogenetic analyses. Conclusions The study challenges the newly emerging nature of Cytauxzoon spp. in central Europe and confirms that isolates from domestic cats in Switzerland and European wild felids belong to the same species. Graphical Abstract

COVID-19-Associated Pulmonary Aspergillosis in a Series of Complete Autopsies from the Brazilian Amazon

Between April and July 2020, and, therefore, prior to the broad recommendation of corticosteroids for severe COVID-19, a total of 50 full autopsies were performed in Manaus. We confirmed two invasive cases of aspergillosis through histopathology and gene sequencing (4%) in our autopsy series. The confirmed invasive aspergillosis incidence seems much lower than expected based on the “probable and possible” definitions, and an individualized approach should be considered for each country scenario. Interestingly, a prolonged length of stay in the intensive care unit was not observed in any of the cases. Timely diagnosis and treatment of fungal infection can reduce mortality rates.

Pharmaceutical interest of in-silico approaches

The virtual environment within the computer using software performed on the computer is known as in-silico studies. These drugs designing software play a vital task in discovering new drugs in the field of pharmaceuticals. These designing programs and software are employed in gene sequencing, molecular modeling, and in assessing the three-dimensional structure of the molecule, which can further be used in drug designing and development. Drug development and discovery is not only a powerful, extensive, and an interdisciplinary system but also a very complex and time-consuming method. This book chapter mainly focused on different types of in-silico approaches along with their pharmaceutical applications in numerous diseases.

Neuroprotective Effects of Probiotic-Supplemented Diet on Cognitive Behavior of 3xTg-AD Mice

Alzheimer’s disease (AD) is recognized as one of the most common types of senile dementia. AD patients first suffer memory loss for recent events (short-term memory impairment). As the disease progresses, they are deprived of self-awareness. This study aims to explore the effects of a probiotic-supplemented diet on the cognitive behaviors and pathological features of mouse models of Alzheimer’s disease (AD). Mice in the control group and the 3xTg-AD group were fed a regular diet and a probiotic-supplemented diet, respectively, for 20 weeks. Behavioral experiments like Morris’s water maze and Y maze were conducted. Then, feces of mice were collected for 16S sRNA gene sequencing for microorganisms. In the end, soluble and insoluble Aβ40 and Aβ42 in the hippocampus and cortex of mice in each group were quantitatively analyzed with a double-antibody Sandwich ELISA. The expression levels of tau protein and gliocyte in the hippocampus and cortex were detected using the Western Blot method. The result of the Morris water maze experiment indicated that, in the place navigation test, the mice in the 3xTg-AD group experienced a significant decline in the learning ability and a longer escape latency and in the space exploration test, the swimming time of mice in the 3xTg-AD group in the target quadrant decreased and after being treated with the probiotic diet, mice in the 3xTg-AD group had improved learning and memory ability. The result of Y maze showed that the probiotic diet can improve the spontaneous alternation accuracy of mice in the 3xTg-AD group. The result of 16s rRNA gene sequencing showed that, compared with mice in the WT group, those in the 3xTg-AD group experienced a change in the intestinal flora. The Western Blot result displayed a decreased expression level of tau (pS202) ( P < 0.05 ) and decreased expression levels of Iba-1 and GFAP ( P < 0.05 ). The result of the ELISA experiment showed decreased levels of soluble and insoluble Aβ40 and Aβ42 in 3xTg-AD mice ( P < 0.05 ). In conclusion, a probiotic diet can prevent and treat AD by improving the intestinal flora of 3xTg-AD.

Investigation on prevalence, risk factors, and genetic diversity of Pediculus humanus capitis among primary school children

Pediculosis is an integumentary disease caused by the ecto-parasite Pediculus humanus capitis, which infests human hair. It is a common public health problem that is most prominent worldwide in elementary school children. The current study aimed to investigate the prevalence, risk factors, and genetic diversity of P. humanus capitis among primary school children in the Erbil province. For this purpose, this study was conducted from October 2019 to December 2019 among 1100 randomly selected elementary school children aged 6-12. Data collection was performed via a regular questionnaire and physical hair examination. For the genetic diversity part, after collecting one louse randomly from each individual, DNA was extracted. The mitochondrial Cox1 gene was then amplified by universal primer and PCR. Gene sequencing was performed by ABI (BioNEER, South Korea). Data analysis was done by Chi-Square and T-test using the SPSS ver. 23. The overall infestation rate was 21.27%, and the rate was significantly higher among females (34.93%) compared to males (7.91%). Some variables had found the prevalence rate to be strongly affected. This included age; the rate was not significant (26.87%) in the age group 8-9 years compared to other age groups. According to hair length, the rate was significantly increased (36.52%) among children with tall hair. In terms of hair type, the incidence of curly-haired children was significantly higher (31.54%); in terms of hair color, there were not significant differences among blonde children (25.90%) and others. According to the results of Cox1 gene sequencing, of 234 infested children to lice, 86 (36.75%) of them were exposed to clade A, 38 (16.24%) were exposed to clade B, clade C has not been seen among any children (0%), 105 students (44.87%) were exposed to clade D, and 5 of them exposed to clade E (2.14%). Eventually, a significantly higher incidence (33.78%) was reported in rural primary school children. The infection rate of human head lice in Erbil province is still high, which is one of the health problems of children in public schools.