Evelyn Santana

Colonization with Extended-Spectrum β-Lactamase-Producing Escherichia coli and Klebsiella Species in Long-Term Care Facility Residents

We describe the prevalence of and risk factors for colonization with extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL-EB) in the long-term care facility (LTCF) setting. Colonization prevalence differed significantly across the 3 LTCFs evaluated in the study, with recent use of levofloxacin and fecal incontinence demonstrating borderline significant associations with ESBL-EB colonization.

Risk factors for the development of gastrointestinal colonization with fluoroquinolone-resistant Escherichia coli in residents of long-term care facilities.

BACKGROUND The objective of this study was to assess risk factors for the development of fluoroquinolone (FQ)-resistant Escherichia coli gastrointestinal tract colonization in long-term care facility (LTCF) residents. METHODS A prospective cohort study was conducted from 2006 to 2008 at 3 LTCFs. Residents initially colonized with FQ-susceptible E. coli were followed by means of serial fecal sampling for new FQ-resistant E. coli colonization for up to 12 months or until discharge or death. A Cox proportional hazards regression model was developed to identify risk factors for new FQ-resistant E. coli colonization, with antibiotic and device exposures modeled as time-varying covariates. RESULTS Fifty-seven (47.5%) of 120 residents became newly colonized with FQ-resistant E. coli, with a median time to colonization of 57 days. Fecal incontinence (hazard ratio [HR], 1.78; 95% confidence interval [CI], 1.04-3.06; P = .04) was significantly associated with FQ-resistant E. coli acquisition. Receipt of amoxicillin-clavulanate (HR, 6.48; 95% CI, 1.43-29.4; P = .02) and the presence of a urinary catheter (HR, 3.81; 95% CI, 1.06-13.8; P = .04) during LTCF stay increased the risk of new FQ-resistant E. coli colonization. CONCLUSIONS Acquisition of FQ-resistant E. coli was common, with nearly half of LTCF residents developing new FQ-resistant E. coli colonization. Further studies are needed on interventions to limit the emergence of FQ-resistant E. coli in LTCFs.

Photoreceptor proliferation and dysregulation of cell cycle genes in early onset inherited retinal degenerations

BackgroundMitotic terminally differentiated photoreceptors (PRs) are observed in early retinal degeneration (erd), an inherited canine retinal disease driven by mutations in the NDR kinase STK38L (NDR2).ResultsWe demonstrate that a similar proliferative response, but of lower magnitude, occurs in two other early onset disease models, X-linked progressive retinal atrophy 2 (xlpra2) and rod cone dysplasia 1 (rcd1). Proliferating cells are rod PRs, and not microglia or Müller cells. Expression of the cell cycle related genes RB1 and E2F1 as well as CDK2,4,6 was up-regulated, but changes were mutation-specific. Changes in cyclin expression differed across all genes, diseases and time points analyzed, although CCNA1 and CCNE1 expression increased with age in the three models suggesting that there is a dysregulation of cell cycle gene expression in all three diseases. Unique to erd, however, are mutation-specific changes in the expression of NDR kinases and Hippo signaling members with increased expression of MOB1 and LATS1 in the newly generated hybrid rod/S-cones.ConclusionsOur data raise the intriguing possibility that terminally differentiated normal PRs are kept from dividing by NDR2-MOB1 interaction. Furthermore, they provide the framework for the selection of candidate genes for further investigation as potential targets of therapy.

A Naturally Occurring Canine Model of Autosomal Recessive Congenital Stationary Night Blindness.

Congenital stationary night blindness (CSNB) is a non-progressive, clinically and genetically heterogeneous disease of impaired night vision. We report a naturally-occurring, stationary, autosomal recessive phenotype in beagle dogs with normal daylight vision but absent night vision. Affected dogs had normal retinas on clinical examination, but showed no detectable rod responses. They had “negative-type” mixed rod and cone responses in full-field ERGs. Their photopic long-flash ERGs had normal OFF-responses associated with severely reduced ON-responses. The phenotype is similar to the Schubert-Bornschein form of complete CSNB in humans. Homozygosity mapping ruled out most known CSNB candidates as well as CACNA2D4 and GNB3. Three remaining genes were excluded based on sequencing the open reading frame and intron-exon boundaries (RHO, NYX), causal to a different form of CSNB (RHO) or X-chromosome (NYX, CACNA1F) location. Among the genes expressed in the photoreceptors and their synaptic terminals, and mGluR6 cascade and modulators, reduced expression of GNAT1, CACNA2D4 and NYX was observed by qRT-PCR in both carrier (n = 2) and affected (n = 2) retinas whereas CACNA1F was down-regulated only in the affecteds. Retinal morphology revealed normal cellular layers and structure, and electron microscopy showed normal rod spherules and synaptic ribbons. No difference from normal was observed by immunohistochemistry (IHC) for antibodies labeling rods, cones and their presynaptic terminals. None of the retinas showed any sign of stress. Selected proteins of mGluR6 cascade and its modulators were examined by IHC and showed that PKCα weakly labeled the rod bipolar somata in the affected, but intensely labeled axonal terminals that appeared thickened and irregular. Dendritic terminals of ON-bipolar cells showed increased Goα labeling. Both PKCα and Goα labeled the more prominent bipolar dendrites that extended into the OPL in affected but not normal retinas. Interestingly, RGS11 showed no labeling in the affected retina. Our results indicate involvement of a yet unknown gene in this canine model of complete CSNB.

Strong upregulation of inflammatory genes accompanies photoreceptor demise in canine models of retinal degeneration

We have analyzed the complex pattern of the inflammatory response in early-onset canine models of human retinitis pigmentosa, rcd1, xlpra2 and erd, as well as late-onset xlpra1, in comparative manner. The time course of immune response genes and proteins expression was examined along the timeline of photoreceptors degeneration. Gene expression analysis of the early-onset models prior to and after the peak of photoreceptors death identified the involvement of multiple immune response genes including those encoding constituents of the NLRP3 inflammasome, its substrates, pro-IL1B, pro-IL18, and common components of IL1B, IL18 and TLR4 pathways. Out of two activated caspase-1 cleavage products, IL1B and IL18, only IL1B was detected in rcd1 and xlpra2 while precursor IL18 remained unprocessed in the same protein extract highlighting prominence of IL1B pathway. An overall immune response was most prominent in rcd1 followed by xlpra2 and least prominent in erd. Noticeably, in rcd1 and xlpra2, but not in erd, early induction of the immune response was accompanied by sustained intraretinal migration and activation of retinal microglia. Lastly, delayed activation of the anti-inflammatory factors in all early-onset models was insufficient to counterbalance rapidly progressing inflammation. In contrast to early-onset models, in late-onset xlpra1 retinas a subset of the pro-inflammatory genes was highly upregulated long before any disease-related structural changes occurred, but was counterbalanced by an adequate anti-inflammatory response. Results point out to upregulated immune response accompanying disease progression in animal models of retinal degeneration, and to potential benefits of early anti-inflammatory therapy.

Epidemiology and characteristics of Escherichia coli sequence type 131 (ST131) from long-term care facility residents colonized intestinally with fluoroquinolone-resistant Escherichia coli

The objective of this study was to evaluate molecular and epidemiologic factors associated with Escherichia coli sequence type 131 (ST131) among long-term care facility (LTCF) residents who acquired gastrointestinal tract colonization with fluoroquinolone-resistant E. coli (FQREC). Colonizing isolates from 37 residents who newly developed FQREC colonization at three LTCFs from 2006 to 2008 were evaluated. Twenty-nine (78%) of 37 total FQREC colonizing isolates were ST131. Most ST131 isolates had a distinctive combination of gyrA and parC replacement mutations. The ST131 and non-ST131 isolates differed significantly for the prevalence of many individual virulence factors but not for the proportion that qualified molecularly as extraintestinal pathogenic E. coli (ExPEC) or aggregate virulence factor scores. E. coli ST131 was highly prevalent among LTCF residents with FQREC colonization. Future studies should determine the risk factors for infection among ST131-colonized residents, and assess the potential for increased transmissibility of ST131 in the long-term care setting.

Efficient recovery of fluoroquinolone-susceptible and fluoroquinolone-resistant Escherichia coli strains from frozen samples.

We assessed the rate of recovery of fluoroquinolone-resistant and fluoroquinolone-susceptible Escherichia coli isolates from culture of frozen perirectal swab samples compared with the results for culture of the same specimen before freezing. Recovery rates for these 2 classes of E. coli were 91% and 83%, respectively. The majority of distinct strains recovered from the initial sample were also recovered from the frozen sample. The strains that were not recovered were typically present only in low numbers in the initial sample. These findings emphasize the utility of frozen surveillance samples.

Ndr kinases regulate retinal interneuron proliferation and homeostasis

Ndr2/Stk38l encodes a protein kinase associated with the Hippo tumor suppressor pathway and is mutated in a naturally-occurring canine early retinal degeneration (erd). To elucidate the retinal functions of Ndr2 and its paralog Ndr1/Stk38, we generated Ndr1 and Ndr2 single knockout mice. Although retinal lamination appeared normal in these mice, Ndr deletion caused a subset of Pax6-positive amacrine cells to proliferate in differentiated retinas, while concurrently decreasing the number of GABAergic, HuD and Pax6-positive amacrine cells. Retinal transcriptome analyses revealed that Ndr2 deletion increased expression of neuronal stress genes and decreased expression of synaptic organization genes. Consistent with the latter, Ndr deletion dramatically reduced levels of Aak1, an Ndr substrate that regulates vesicle trafficking. Our findings indicate that Ndr kinases are important regulators of amacrine and photoreceptor cells and suggest that Ndr kinases inhibit the proliferation of a subset of terminally differentiated cells and modulate interneuron synapse function via Aak1.

Author Correction: Variabilities in retinal function and structure in a canine model of cone-rod dystrophy associated with RPGRIP1 support multigenic etiology

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Variabilities in retinal function and structure in a canine model of cone-rod dystrophy associated with RPGRIP1 support multigenic etiology

Defects in the cilia gene RPGRIP1 cause Leber congenital amaurosis and cone-rod dystrophy in humans. A form of canine cone-rod dystrophy (cord1) was originally associated with a homozygous insertion in RPGRIP1 (RPGRIP1ins/ins) as the primary disease locus while a homozygous deletion in MAP9 (MAP9del/del) was later identified as a modifier associated with the early onset form. However, we find further variability in cone electroretinograms (ERGs) ranging from normal to absent in an extended RPGRIP1ins/ins canine colony, irrespective of the MAP9 genotype. Ophthalmoscopically, cone ERGabsentRPGRIP1ins/ins eyes show discolouration of the tapetal fundus with varying onset and disease progression, while sd-OCT reveals atrophic changes. Despite marked changes in cone ERG and retinal morphology, photopic vision-guided behaviour is comparable between normal and cone ERGabsentRPGRIP1ins/ins littermates. Cone morphology of the dogs lacking cone ERG are truncated with shortened outer and inner segments. Immunohistochemically, cone ERGabsentRPGRIP1ins/ins retinas have extensive L/M-opsin mislocalization, lack CNGB3 labelling in the L/M-cones, and lack GC1 in all cones. Our results indicate that cord1 is a multigenic disease in which mutations in neither RPGRIP1 nor MAP9 alone lead to visual deficits, and additional gene(s) contribute to cone-specific functional and morphologic defects.