Clinical Risk Factors and Microbiological and Intestinal Characteristics of Carbapenemase-Producing Enterobacteriaceae Colonization and Subsequent Infection.

Gastrointestinal colonization with carbapenem-resistant Enterobacteriaceae (CRE) is always a prerequisite for the development of translocated infections. Here, we sought to screen for fecal carriage of CRE and identify the risk factors for CRE colonization as well as subsequent translocated pneumonia in critically ill patients admitted to the intensive care unit (ICU) of a university hospital in China. We further focused on the intestinal flora composition and fecal metabolic profiles in CRE rectal colonization and translocated infection patients. Animal models of gastrointestinal colonization with a carbapenemase-producing Klebsiella pneumoniae (carbapenem-resistant K. pneumoniae [CRKP]) clinical isolate expressing green fluorescent protein (GFP) were established, and systemic infection was subsequently traced using an in vivo imaging system (IVIS). The intestinal barrier, inflammatory factors, and infiltrating immune cells were further investigated. In this study, we screened 54 patients hospitalized in the ICU with CRE rectal colonization, and 50% of the colonized patients developed CRE-associated pneumonia, in line with the significantly high mortality rate. Upon multivariate analysis, risk factors associated with subsequent pneumonia caused by CRE in patients with fecal colonization included enteral feeding and carbapenem exposure. Furthermore, CRKP colonization and translocated infection influenced the diversity and community composition of the intestinal microbiome. Downregulated propionate and butyrate probably play important and multiangle roles in regulating immune cell infiltration, inflammatory factor expression, and mucus and intestinal epithelial barrier integrity. Although the risk factors and intestinal biomarkers for subsequent infections among CRE-colonized patients were explored, further work is needed to elucidate the complicated mechanisms. IMPORTANCE Carbapenem-resistant Enterobacteriaceae have emerged as a major threat to modern medicine, and the spread of carbapenem-resistant Enterobacteriaceae is a clinical and public health problem. Gastrointestinal colonization by potential pathogens is always a prerequisite for the development of translocated infections, and there is a growing need to assess clinical risk factors and microbiological and intestinal characteristics to prevent the development of clinical infection by carbapenem-resistant Enterobacteriaceae.

Toxoplasma gondii α-amylase deletion mutant is a promising vaccine against acute and chronic toxoplasmosis.

Individuals with inhibited immunity may develop lethal toxoplasmosis; thus, a safe and effective vaccine is urged to be developed. Toxoplasma gondii (T. gondii) α-amylase (α-AMY) is one of the enzymes responsible for starch digestion. In the present study, we first generated a ME49Δα-amy mutant and discovered that loss of α-AMY robustly grew in vitro but contributed to significant virulence attenuation in vivo. Therefore, we established a mouse model to explore the protective immunity of Δα-amy mutant against acute and chronic toxoplasmosis. The results indicated that the survival rates of short-term or long-term immunized mice re-infected with the tachyzoites of multiple T. gondii strains were nearly 100%. ME49Δα-amy not only could provide protective immunity against tachyzoites infection but also could resist the infection of tissue cysts. Furthermore, we detected that ME49Δα-amy vaccination could effectively eliminate the proliferation of parasites in mice and prevent the formation of cysts. The significant increases of Th1-type cytokines, Th2-type cytokines and specific total IgG and IgG subclasses (IgG2a and IgG1) confirmed efficiency of a combination of cellular and humoral immunity against infection. In conclusion, ME49Δα-amy attenuated strain can produce strong immune responses to provide efficient protection against toxoplasmosis, which signifies that ME49Δα-amy mutant may be a potential vaccine candidate.

Comparative transcriptome analysis of normal and CD44-deleted mouse brain under chronic infection with Toxoplasma gondii.

Toxoplasma gondii is a globally-distributed intracellular parasitic protozoon with wide host range. Chronic infection is the most prevalent form of T. gondii infection, which can lead to significant damage. CD44 plays an important role in body's immune response, however, little is known about the function and mechanism of CD44 in T. gondii infection until now. In the present study, total RNA isolated from four groups including C57BL/6 mouse (C57), C57BL/6△CD44 mouse(C57△CD44), C57BL/6 mouse infected with T. gondii (C57-TG) and C57BL/6△CD44 infected with T. gondii (C57△CD44-TG)were subjected to comparative transcriptome analyses using RNA-seq techniques to explore the possible function of CD44 in mouse brain during chronic Toxoplasma infection. The results indicated a total of 35,908, 54,428, 51,473 and 22,387 unigenes were annotated in KOG, Swissprot, GO and KEGG databases by transcriptome analysis, respectively, and all the databases shared 9,833 unigenes. Subsequently, differentially expressed GO terms and enriched KEGG Pathways showed 20,303 unigenes were annotated belonging to three main GO categories (namely biological process, cellular component and molecular function) and six main KEGG categories (cellular processes, environmental information processing, genetic information processing, human diseases, metabolism and organismal systems) between normal C57 and C57△CD44 mice, as well as for C57-TG and C57△CD44-TG mice. For up-regulated genes, Mid1, Ttr and Cd4 were significantly up-regulated in the C57△CD44 mouse compared with the C57 mouse, and Pcp2, Ppp1r17 and Nrk were significantly up-regulated in the C57△CD44-TG mouse compared with the C57-TG mouse. As to down-regulated genes, AC114588.1, Cbln3 and Pmch were significantly down-regulated in the C57△CD44 the mouse compared with the C57 mouse, and down-regulated genes were enriched for immunoglobulins, major histocompatibility complex (MHC) class II antigens, chemokines ligands and interferon (IFN)-inducible GTPase families in the C57△CD44-TG mouse compared with the C57-TG mouse. The present study is the first trial for exploring the function of CD44 in the mouse brain during chronic infection with T. gondii at the transcriptional level, which can provide a basis for the study of the host immune defense mechanism against T. gondii infection.

Toxoplasma gondii ADSL Knockout Provides Excellent Immune Protection against a Variety of Strains.

Toxoplasma gondii is a protozoan parasite, occurring worldwide, endangers human health and causes enormous economic losses to the Ministry of Agriculture. A safe and effective vaccination is needed to handle these problems. In addition, ideal vaccine production is a challenge in the future. In this study, we knocked out the adenylosuccinate lyase (ADSL) gene and found that the gene reduces the growth rate of T. gondii tachyzoites in vitro under standard growth conditions by plaque or replication experiments. Furthermore, mice that were immunized with tachyzoites of the ME49ΔADSL strain induced 100% protection efficacy against challenge with the type 1 strain RH, type 2 strain ME49 and type 3 strain VEG. All mice that were immunized with ME49ΔADSL had a survival rate of 100% when they were reinfected with wild-type strains, either 30 days or 70 days after immunization, and immunization was also protective against homologous infection with 50 T. gondii ME49 tissue cysts. In addition, the level of Toxoplasma-specific IgG was significantly elevated at 30 and 70 days after immunization. ME49ΔADSL induced high levels of Th1 cytokines (interferon gamma (IFN-γ), interleukin (IL)-12) at 4 weeks after immunization and spleen cell cultures from mice vaccinated for 150 days were able to produce robust INF-γ and IL-12 levels in the supernatant. The results of the present study showed that ΔADSL vaccination induced a T. gondii-specific cellular immune response against further infections. These results suggest that the ADSL-deficient vaccine can induce anti-Toxoplasma gondii humoral and cellular immune responses and has 100% immune protection against post-challenge by the type 1 strain RH, type 2 strain ME49 and type 3 strain VEG. It will be used as an excellent candidate for live vaccines and may contribute in a positive meaning to control human toxoplasmosis.

Transcriptomic Profiling Identifies Candidate Genes Involved in the Salt Tolerance of the Xerophyte Pugionium cornutum.

The xerophyte Pugionium cornutum adapts to salt stress by accumulating inorganic ions (e.g., Cl-) for osmotic adjustment and enhancing the activity of antioxidant enzymes, but the associated molecular basis remains unclear. In this study, we first found that P. cornutum could also maintain cell membrane stability due to its prominent ROS-scavenging ability and exhibits efficient carbon assimilation capacity under salt stress. Then, the candidate genes associated with the important physiological traits of the salt tolerance of P. cornutum were identified through transcriptomic analysis. The results showed that after 50 mM NaCl treatment for 6 or 24 h, multiple genes encoding proteins facilitating Cl- accumulation and NO3- homeostasis, as well as the transport of other major inorganic osmoticums, were significantly upregulated in roots and shoots, which should be favorable for enhancing osmotic adjustment capacity and maintaining the uptake and transport of nutrient elements; a large number of genes related to ROS-scavenging pathways were also significantly upregulated, which might be beneficial for mitigating salt-induced oxidative damage to the cells. Meanwhile, many genes encoding components of the photosynthetic electron transport pathway and carbon fixation enzymes were significantly upregulated in shoots, possibly resulting in high carbon assimilation efficiency in P. cornutum. Additionally, numerous salt-inducible transcription factor genes that probably regulate the abovementioned processes were found. This work lays a preliminary foundation for clarifying the molecular mechanism underlying the adaptation of xerophytes to harsh environments.