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Streptococcus gallolyticus subspecies gallolyticus (Sgg) has a strong clinical association with colorectal cancer (CRC) and actively promotes the development of colon tumors. However, the molecular determinants involved in Sgg pathogenicity in the gut are unknown. Bacterial type VII secretion systems (T7SS) mediate pathogen interactions with their host and are important for virulence in pathogenic mycobacteria and Staphylococcus aureus. Through genome analysis, we identified a locus in Sgg strain TX20005 that encodes a putative type VII secretion system (designated as SggT7SST05). We showed that core genes within the SggT7SST05 locus are expressed in vitro and in the colon of mice. Western blot analysis showed that SggEsxA, a protein predicted to be a T7SS secretion substrate, is detected in the bacterial culture supernatant, indicating that this SggT7SST05 is functional. Deletion of SggT7SST05 (TX20005Δesx) resulted in impaired bacterial adherence to HT29 cells and abolished the ability of Sgg to stimulate HT29 cell proliferation. Analysis of bacterial culture supernatants suggest that SggT7SST05-secreted factors are responsible for the pro-proliferative activity of Sgg, whereas Sgg adherence to host cells requires both SggT7SST05-secreted and bacterial surface-associated factors. In a murine gut colonization model, TX20005Δesx showed significantly reduced colonization compared to the parent strain. Furthermore, in a mouse model of CRC, mice exposed to TX20005 had a significantly higher tumor burden compared to saline-treated mice, whereas those exposed to TX20005Δesx did not. Examination of the Sgg load in the colon in the CRC model suggests that SggT7SST05-mediated activities are directly involved in the promotion of colon tumors. Taken together, these results reveal SggT7SST05 as a previously unrecognized pathogenicity determinant for Sgg colonization of the colon and promotion of colon tumors.

Honey bee (Apis mellifera) queens have a remarkable organ, the spermatheca, which successfully stores sperm for years after a virgin queen mates. This study uniquely characterized and quantified the transcriptomes of the spermathecae from mated and virgin honey bee queens via RNA sequencing to identify differences in mRNA levels based on a queen's mating status. The transcriptome of drone semen was analyzed for comparison. Samples from three individual bees were independently analyzed for mated queen spermathecae and virgin queen spermathecae, and three pools of semen from ten drones each were collected from three separate colonies. In total, the expression of 11,233 genes was identified in mated queen spermathecae, 10,521 in virgin queen spermathecae, and 10,407 in drone semen. Using a cutoff log2 fold-change value of 2.0, we identified 212 differentially expressed genes between mated and virgin spermathecal queen tissues: 129 (1.4% of total) were up-regulated and 83 (0.9% of total) were down-regulated in mated queen spermathecae. Three genes in mated queen spermathecae, three genes in virgin queen spermathecae and four genes in drone semen that were more highly expressed in those tissues from the RNA sequencing data were further validated by real time quantitative PCR. Among others, expression of Kielin/chordin-like and Trehalase mRNAs was highest in the spermathecae of mated queens compared to virgin queen spermathecae and drone semen. Expression of the mRNA encoding Alpha glucosidase 2 was higher in the spermathecae of virgin queens. Finally, expression of Facilitated trehalose transporter 1 mRNA was greatest in drone semen. This is the first characterization of gene expression in the spermathecae of honey bee queens revealing the alterations in mRNA levels within them after mating. Future studies will extend to other reproductive tissues with the purpose of relating levels of specific mRNAs to the functional competence of honey bee queens and the colonies they head.

ABSTRACT Staphylococcus epidermidis is a leading cause of nosocomial infections in patients with a compromised immune system and/or an implanted medical device. Seventy to 90% of S. epidermidis clinical isolates are methicillin resistant and carry the mecA gene, present in a mobile genetic element (MGE) called the staphylococcal cassette chromosome mec (SCCmec) element. Along with the presence of antibiotic and heavy metal resistance genes, MGEs can also contain genes encoding secreted or cell wall-anchored virulence factors. In our earlier studies of S. epidermidis clinical isolates, we discovered S. epidermidis surface protein J (SesJ), a prototype of a recently discovered subfamily of the microbial surface component recognizing adhesive matrix molecule (MSCRAMM) group. MSCRAMMs are major virulence factors of pathogenic Gram-positive bacteria. Here, we report that the sesJ gene is always accompanied by two glycosyltransferase genes, gtfA and gtfB, and is present in two MGEs, called the arginine catabolic mobile element (ACME) and the staphylococcal cassette chromosome (SCC) element. The presence of the sesJ gene was associated with the left-hand direct repeat DR_B or DR_E. When inserted via DR_E, the sesJ gene was encoded in the SCC element. When inserted via DR_B, the sesJ gene was accompanied by the genes for the type 1 restriction modification system and was encoded in the ACME. Additionally, the SCC element and ACME carry different isoforms of the SesJ protein. To date, the genes encoding MSCRAMMs have been seen to be located in the bacterial core genome. Here, we report the presence of an MSCRAMM in an MGE in S. epidermidis clinical isolates. IMPORTANCE S. epidermidis is an opportunistic bacterium that has established itself as a successful nosocomial pathogen. The modern era of novel therapeutics and medical devices has extended the longevity of human life, but at the same time, we also witness the evolution of pathogens to adapt to newly available niches in the host. Increasing antibiotic resistance among pathogens provides an example of such pathogen adaptation. With limited opportunities to modify the core genome, most of the adaptation occurs by acquiring new genes, such as virulence factors and antibiotic resistance determinants present in MGEs. In this study, we describe that the sesJ gene, encoding a recently discovered cell wall-anchored protein in S. epidermidis, is present in both ACME and the SCC element. The presence of virulence factors in MGEs can influence the virulence potential of a specific strain. Therefore, it is critical to study the virulence factors found in MGEs in emerging pathogenic bacteria or strains to understand the mechanisms used by these bacteria to cause infections.

Virus-induced neurological sequelae resulting from infection by Theiler’s murine encephalomyelitis virus (TMEV) are used for studying human conditions ranging from epileptic seizures to demyelinating disease. Mouse strains are typically considered susceptible or resistant to TMEV infection based on viral persistence and extreme phenotypes, such as demyelination. We have identified a broader spectrum of phenotypic outcomes by infecting strains of the genetically diverse Collaborative Cross (CC) mouse resource. We evaluated the chronic-infection gene expression profiles of hippocampi and thoracic spinal cords for 19 CC strains in relation to phenotypic severity and TMEV persistence. Strains were clustered based on similar phenotypic profiles and TMEV levels at 90 days post-infection, and we categorized distinct TMEV response profiles. The three most common profiles included “resistant” and “susceptible,” as before, as well as a “resilient” TMEV response group which experienced both TMEV persistence and mild neurological phenotypes even at 90 days post-infection. Each profile had a distinct gene expression signature, allowing the identification of pathways and networks specific to each TMEV response group. CC founder haplotypes for genes involved in these pathways/networks revealed candidate response-specific alleles. These alleles demonstrated pleiotropy and epigenetic (miRNA) regulation in long-term TMEV infection, with particular relevance for resilient mouse strains.

Abstract Introduction Prenatal alcohol exposure can contribute to fetal alcohol spectrum disorders (FASD), characterized by a myriad of developmental impairments affecting behavior and cognition. Studies show that many of these functional impairments are associated with the hippocampus, a structure exhibiting exquisite vulnerability to developmental alcohol exposure and critically implicated in learning and memory; however, mechanisms underlying alcohol‐induced hippocampal deficits remain poorly understood. By utilizing a high‐throughput RNA‐sequencing (RNA‐seq) approach to address the neurobiological and molecular basis of prenatal alcohol‐induced hippocampal functional deficits, we hypothesized that chronic binge prenatal alcohol exposure alters gene expression and global molecular pathways in the fetal hippocampus. Methods Timed‐pregnant Sprague–Dawley rats were randomly assigned to a pair‐fed control (PF) or binge alcohol (ALC) treatment group on gestational day (GD) 4. ALC dams acclimatized from GDs 5–10 with a daily treatment of 4.5 g/kg alcohol and subsequently received 6 g/kg on GDs 11–20. PF dams received a once daily maltose dextrin gavage on GDs 5–20, isocalorically matching ALC counterparts. On GD 21, bilateral hippocampi were dissected, flash frozen, and stored at −80°C. Total RNA was then isolated from homogenized tissues. Samples were normalized to ~4nM and pooled equally. Sequencing was performed by Illumina NextSeq 500 on a 75 cycle, single‐end sequencing run. Results RNA‐seq identified 13,388 genes, of these, 76 genes showed a significant difference (p

Melanocortin 1 receptor (MC1R), the agouti signaling protein (ASIP), and tyrosinase related protein 1 (TYRP1) are among the major regulators of pigmentation in mammals. Recently, MC1R and ASIP sequence variants were associated with white and black/dark brown coat colors, respectively, in the dromedary. Here we confirmed this association by independent sequencing and mutation discovery of MC1R and ASIP coding regions and by TaqMan genotyping in 188 dromedaries from Saudi Arabia and United States, including 38 black, 53 white, and 97 beige/brown/red animals. We showed that heterozygosity for a missense mutation c.901C > T in MC1R is sufficient for the white coat color suggesting a possible dominant negative effect. Likewise, we confirmed that the majority of black dromedaries were homozygous for a frameshift mutation in ASIP exon 2, except for 4 animals, which were heterozygous. In search for additional mutations underlying the black color, we identified another frameshift mutation in ASIP exon 4 and 6 new variants in MC1R including a significantly associated SNP in 3′UTR. In pursuit of sequence variants that may modify dromedary wild-type color from dark-reddish brown to light beige, we identified 4 SNPs and one insertion in TYRP1 non-coding regions. However, none of these were associated with variations in wild-type colors. Finally, the three genes were cytogenetically mapped in New World (alpaca) and Old World (dromedary and Bactrian camel) camelids. The MC1R was assigned to chr21, ASIP to chr19 and TYRP1 to chr4 in all 3 species confirming extensive conservation of camelid karyotypes. Notably, while the locations of ASIP and TYRP1 were in agreement with human-camelid comparative map, mapping MC1R identified a new evolutionary conserved synteny segment between camelid chromosome 21 and HSA16. The findings contribute to coat color genomics and the development of molecular tests in camelids and toward the chromosome level reference assemblies of camelid genomes.

The rapid divergence of male genitalia is a preeminent evolutionary pattern. This rapid divergence is especially striking in the baculum, a bone that occurs in the penis of many mammalian species. Closely related species often display diverse baculum morphology where no other morphological differences can be discerned. While this fundamental pattern of evolution has been appreciated at the level of gross morphology, nearly nothing is known about the genetic basis of size and shape divergence. Quantifying the genetic basis of baculum size and shape variation has been difficult because these structures generally lack obvious landmarks, so comparing them in three dimensions is not straightforward. Here, we develop a novel morphometric approach to quantify size and shape variation from three-dimensional micro-CT scans taken from 369 bacula, representing 75 distinct strains of the BXD family of mice. We identify two quantitative trait loci (QTL) that explain ∼50% of the variance in baculum size, and a third QTL that explains more than 20% of the variance in shape. Together, our study demonstrates that baculum morphology may diverge relatively easily, with mutations at a few loci of large effect that independently modulate size and shape. Based on a combination of bioinformatic investigations and new data on RNA expression, we prioritized these QTL to 16 candidate genes, which have hypothesized roles in bone morphogenesis and may enable future genetic manipulation of baculum morphology.