Microbial determinants of dementia risk in subjects of Mexican descent with type 2 diabetes living in South Texas.

Type 2 diabetes (T2D) is a common forerunner of neurodegeneration and dementia, including Alzheimer's Disease (AD), yet the underlying mechanisms remain unresolved. Individuals of Mexican descent living in South Texas have increased prevalence of comorbid T2D and early onset AD, despite low incidence of the predisposing APOE-e4 variant and an absence of the phenotype among relatives residing in Mexico - suggesting a role for environmental factors in coincident T2D and AD susceptibility. Here, in a small clinical trial, we show dysbiosis of the human gut microbiome could contribute to neuroinflammation and risk for AD in this population. Divergent Gastrointestinal Symptom Rating Scale (GSRS) responses, despite no differences in expressed dietary preferences, provided the first evidence for altered gut microbial ecology among T2D subjects (sT2D) versus population-matched healthy controls (HC). Metataxonomic 16S rRNA sequencing of participant stool revealed a decrease in alpha diversity of sT2D versus HC gut communities and identified BMI as a driver of gut community structure. Linear discriminant analysis effect size (LEfSe) identified a significant decrease in the relative abundance of the short-chain fatty acid-producing taxa Lachnospiraceae, Faecalibacterium, and Alistipes and an increase in pathobionts Escherichia-Shigella, Enterobacter, and Clostridia innocuum among sT2D gut microbiota, as well as differentially abundant gene and metabolic pathways. These results suggest characterization of the gut microbiome of individuals with T2D could identify key actors among "disease state" microbiota which may increase risk for or accelerate the onset of neurodegeneration. Furthermore, they identify candidate microbiome-targeted approaches for prevention and treatment of neuroinflammation in AD.

Monospecies probiotic preparation and administration with downstream analysis of sex-specific effects on gut microbiome composition in mice.

Dysbiosis of the gut microbiome is implicated in the growing burden of non-communicable chronic diseases, including neurodevelopmental disorders, and both preclinical and clinical studies highlight the potential for precision probiotic therapies in their prevention and treatment. Here, we present an optimized protocol for the preparation and administration of Limosilactobacillus reuteri MM4-1A (ATCC-PTA-6475) to adolescent mice. We also describe steps for performing downstream analysis of metataxonomic sequencing data with careful assessment of sex-specific effects on microbiome composition and structure. For complete details on the use and execution of this protocol, please refer to Di Gesù et al.1.

Maternal gut microbiota mediate intergenerational effects of high-fat diet on descendant social behavior.

Dysbiosis of the maternal gut microbiome during pregnancy is associated with adverse neurodevelopmental outcomes. We previously showed that maternal high-fat diet (MHFD) in mice induces gut dysbiosis, social dysfunction, and underlying synaptic plasticity deficits in male offspring (F1). Here, we reason that, if HFD-mediated changes in maternal gut microbiota drive offspring social deficits, then MHFD-induced dysbiosis in F1 female MHFD offspring would likewise impair F2 social behavior. Metataxonomic sequencing reveals reduced microbial richness among female F1 MHFD offspring. Despite recovery of microbial richness among MHFD-descendant F2 mice, they display social dysfunction. Post-weaning Limosilactobacillus reuteri treatment increases the abundance of short-chain fatty acid-producing taxa and rescues MHFD-descendant F2 social deficits. L. reuteri exerts a sexually dimorphic impact on gut microbiota configuration, increasing discriminant taxa between female cohorts. Collectively, these results show multigenerational impacts of HFD-induced dysbiosis in the maternal lineage and highlight the potential of maternal microbiome-targeted interventions for neurodevelopmental disorders.

A study of innate immune kinetics reveals a role for a chloride transporter in a virulent Francisella tularensis type B strain.

Tularemia is a zoonotic disease of global proportions. Francisella tularensis subspecies tularensis (type A) and holarctica (type B) cause disease in healthy humans, with type A infections resulting in higher mortality. Repeated passage of a type B strain in the mid-20th century generated the Live Vaccine Strain (LVS). LVS remains unlicensed, does not protect against high inhalational doses of type A, and its exact mechanisms of attenuation are poorly understood. Recent data suggest that live attenuated vaccines derived from type B may cross-protect against type A. However, there is a dearth of knowledge regarding virulent type B pathogenesis and its capacity to stimulate the host's innate immune response. We therefore sought to increase our understanding of virulent type B in vitro characteristics using strain OR96-0246 as a model. Adding to our knowledge of innate immune kinetics in macrophages following infection with virulent type B, we observed robust replication of strain OR96-0246 in murine and human macrophages, reduced expression of pro-inflammatory cytokine genes from "wild type" type B-infected macrophages compared to LVS, and delayed macrophage cell death suggesting that virulent type B may suppress macrophage activation. One disruption in LVS is in the gene encoding the chloride transporter ClcA. We investigated the role of ClcA in macrophage infection and observed a replication delay in a clcA mutant. Here, we propose its role in acid tolerance. A greater understanding of LVS attenuation may reveal new mechanisms of pathogenesis and inform strategies toward the development of an improved vaccine against tularemia.

Diet-induced dysbiosis of the maternal gut microbiome in early life programming of neurodevelopmental disorders.

The maternal gut microbiome plays a critical role in fetal and early postnatal development, shaping fundamental processes including immune maturation and brain development, among others. Consequently, it also contributes to fetal programming of health and disease. Over the last decade, epidemiological studies and work in preclinical animal models have begun to uncover a link between dysbiosis of the maternal gut microbiome and neurodevelopmental disorders in offspring. Neurodevelopmental disorders are caused by both genetic and environmental factors, and their interactions; however, clinical heterogeneity, phenotypic variability, and comorbidities make identification of underlying mechanisms difficult. Among environmental factors, exposure to maternal obesity in utero confers a significant increase in risk for neurodevelopmental disorders. Obesogenic diets in humans, non-human primates, and rodents induce functional modifications in maternal gut microbiome composition, which animal studies suggest are causally related to adverse mental health outcomes in offspring. Here, we review evidence linking maternal diet-induced gut dysbiosis to neurodevelopmental disorders and discuss how it could affect pre- and early postnatal brain development. We are hopeful that this burgeoning field of research will revolutionize antenatal care by leading to accessible prophylactic strategies, such as prenatal probiotics, to improve mental health outcomes in children affected by maternal diet-induced obesity.

Polynucleotide phosphorylase promotes the stability and function of Hfq-binding sRNAs by degrading target mRNA-derived fragments.

In many Gram-negative and some Gram-positive bacteria, small regulatory RNAs (sRNAs) that bind the RNA chaperone Hfq have a pivotal role in modulating virulence, stress responses, metabolism and biofilm formation. These sRNAs recognize transcripts through base-pairing, and sRNA-mRNA annealing consequently alters the translation and/or stability of transcripts leading to changes in gene expression. We have previously found that the highly conserved 3'-to-5' exoribonuclease polynucleotide phosphorylase (PNPase) has an indispensable role in paradoxically stabilizing Hfq-bound sRNAs and promoting their function in gene regulation in Escherichia coli. Here, we report that PNPase contributes to the degradation of specific short mRNA fragments, the majority of which bind Hfq and are derived from targets of sRNAs. Specifically, we found that these mRNA-derived fragments accumulate in the absence of PNPase or its exoribonuclease activity and interact with PNPase. Additionally, we show that mutations in hfq or in the seed pairing region of some sRNAs eliminated the requirement of PNPase for their stability. Altogether, our results are consistent with a model that PNPase degrades mRNA-derived fragments that could otherwise deplete cells of Hfq-binding sRNAs through pairing-mediated decay.

Polynucleotide phosphorylase: Not merely an RNase but a pivotal post-transcriptional regulator.

Almost 60 years ago, Severo Ochoa was awarded the Nobel Prize in Physiology or Medicine for his discovery of the enzymatic synthesis of RNA by polynucleotide phosphorylase (PNPase). Although this discovery provided an important tool for deciphering the genetic code, subsequent work revealed that the predominant function of PNPase in bacteria and eukaryotes is catalyzing the reverse reaction, i.e., the release of ribonucleotides from RNA. PNPase has a crucial role in RNA metabolism in bacteria and eukaryotes mainly through its roles in processing and degrading RNAs, but additional functions in RNA metabolism have recently been reported for this enzyme. Here, we discuss these established and noncanonical functions for PNPase and the possibility that the major impact of PNPase on cell physiology is through its unorthodox roles.

Poly(A) polymerase is required for RyhB sRNA stability and function in Escherichia coli.

Small regulatory RNAs (sRNAs) are an important class of bacterial post-transcriptional regulators that control numerous physiological processes, including stress responses. In Gram-negative bacteria including Escherichia coli, the RNA chaperone Hfq binds many sRNAs and facilitates pairing to target transcripts, resulting in changes in mRNA transcription, translation, or stability. Here, we report that poly(A) polymerase (PAP I), which promotes RNA degradation by exoribonucleases through the addition of poly(A) tails, has a crucial role in the regulation of gene expression by Hfq-dependent sRNAs. Specifically, we show that deletion of pcnB, encoding PAP I, paradoxically resulted in an increased turnover of certain Hfq-dependent sRNAs, including RyhB. RyhB instability in the pcnB deletion strain was suppressed by mutations in hfq or ryhB that disrupt pairing of RyhB with target RNAs, by mutations in the 3' external transcribed spacer of the glyW-cysT-leuZ transcript (3'ETSLeuZ) involved in pairing with RyhB, or an internal deletion in rne, which encodes the endoribonuclease RNase E. Finally, the reduced stability of RyhB in the pcnB deletion strain resulted in impaired regulation of some of its target mRNAs, specifically sodB and sdhCDAB. Altogether our data support a model where PAP I plays a critical role in ensuring the efficient decay of the 3'ETSLeuZ In the absence of PAP I, the 3'ETSLeuZ transcripts accumulate, bind Hfq, and pair with RyhB, resulting in its depletion via RNase E-mediated decay. This ultimately leads to a defect in RyhB function in a PAP I deficient strain.

Alcohol Consumption and Incident Stroke Among Older Adults.

Objectives: This study examines the relationship between alcohol consumption and incident stroke among older adults and tests whether alcohol consumption contributes to observed race and sex differences in stroke. Method: Data are from a U.S. national cohort of black and white adults aged 45 and older, the REasons for Geographic And Racial Differences in Stroke (REGARDS) study. Current and past drinking levels were reported at baseline (2003-2007). Participants who had never had a stroke were followed for adjudicated stroke events through September 2015 (n = 27,265). We calculated Cox proportional hazard models for stroke, adjusting for demographic, socioeconomic, behavioral, and health characteristics. Results: Participants, mean age 64.7 years, consumed on average 2.2 drinks/week and experienced 1,140 first-time stroke events over median 9.1 years follow-up. Nondrinkers had a 12% higher risk of stroke than current drinkers; the risk of stroke among nondrinkers largely reflected high risks among past drinkers; these differences were explained by socioeconomic characteristics. Among current drinkers, light drinkers had significantly lower stroke risks than moderate drinkers after accounting for demographic, socioeconomic, behavioral, and health characteristics. Implications of alcohol did not differ between blacks and whites but did differ by sex: Especially among women, nondrinkers, and specifically past drinkers, had higher risks; these differences were largely explained by health characteristics and behaviors. Alcohol did not explain race and sex differences in stroke incidence. Discussion: Among older adults, those who used to, but no longer, drink had higher risks of stroke, especially among women; current light drinkers had the lowest risk of stroke.