The ß-encapsulation cage of rearrangement hotspot (Rhs) effectors is required for type VI secretion.

Bacteria deploy rearrangement hotspot (Rhs) proteins as toxic effectors against both prokaryotic and eukaryotic target cells. Rhs proteins are characterized by YD-peptide repeats, which fold into a large ß-cage structure that encapsulates the C-terminal toxin domain. Here, we show that Rhs effectors are essential for type VI secretion system (T6SS) activity in Enterobacter cloacae (ECL). ECL rhs- mutants do not kill Escherichia coli target bacteria and are defective for T6SS-dependent export of hemolysin-coregulated protein (Hcp). The RhsA and RhsB effectors of ECL both contain Pro-Ala-Ala-Arg (PAAR) repeat domains, which bind the ß-spike of trimeric valine-glycine repeat protein G (VgrG) and are important for T6SS activity in other bacteria. Truncated RhsA that retains the PAAR domain is capable of forming higher-order, thermostable complexes with VgrG, yet these assemblies fail to restore secretion activity to ∆rhsA ∆rhsB mutants. Full T6SS-1 activity requires Rhs that contains N-terminal transmembrane helices, the PAAR domain, and an intact ß-cage. Although ∆rhsA ∆rhsB mutants do not kill target bacteria, time-lapse microscopy reveals that they assemble and fire T6SS contractile sheaths at ∼6% of the frequency of rhs+ cells. Therefore, Rhs proteins are not strictly required for T6SS assembly, although they greatly increase secretion efficiency. We propose that PAAR and the ß-cage provide distinct structures that promote secretion. PAAR is clearly sufficient to stabilize trimeric VgrG, but efficient assembly of T6SS-1 also depends on an intact ß-cage. Together, these domains enforce a quality control checkpoint to ensure that VgrG is loaded with toxic cargo before assembling the secretion apparatus.

Clinical Scores Predict Acute and Chronic Complications in Pediatric Osteomyelitis: An External Validation.

BACKGROUND: Pediatric acute hematogenous osteomyelitis (AHO) outcomes are highly dependent on the disease severity. Recently, the A-SCORE and C-SCORE, were proposed as predictors of an acute complicated course and chronic morbidity, respectively. The purpose of this study was to externally validate the A-SCORE and C-SCORE at a single institution. METHODS: This IRB-approved retrospective chart review included AHO patients admitted at a tertiary referral hospital between October 1, 2015 and December 31, 2019. The inclusion criteria were ages 0 to 18 and clinical response to treatment. The exclusion criteria were immunocompromised status or penetrating inoculation. RESULTS: The A-SCORE demonstrated an area under the receiver operator curve (ROC area) of >86% with regards to all acute complications. It also demonstrated sensitivities >85% and specificities >92% at the cut-off of 4 (Youden index) for all acute complications. The C-SCORE demonstrated an ROC area of 100% with regards to chronic osteomyelitis. It also demonstrated sensitivities >70% and specificities >93% for the chronic morbidity variables seen in our population at the cut-off of 3 (Youden index.). CONCLUSIONS: These novel composite clinical scores, in combination with clinical judgment, could help guide early care decisions. The A-SCORE and C-SCORE are useful risk stratification tools in the management of pediatric AHO and in predicting acute complicated courses or chronic sequelae of AHO, respectively. These scoring systems, if integrated into standardized pediatric AHO guidelines, can allow clinicians to stratify the AHO population and guide clinical decision making. LEVEL OF EVIDENCE: Level III (prognostic study, retrospective chart review).

Diversity and Inclusion in an Orthopaedic Surgical Society: A Longitudinal Study.

BACKGROUND: Diversity and inclusion are critical to providing the best possible health care. Previous studies have shown that diversity among physicians increases cultural competency, which in turn enhances the quality of care provided and increases minoritized patients' participation in decisions regarding their health care. However, physician diversity in both race and sex is lacking in orthopaedic surgery. This study seeks to determine the sex and racial diversity in the membership and leadership of the Pediatric Orthopaedic Society of North America (POSNA). METHODS: POSNA membership and leadership were reviewed for the years 2010, 2015, and 2020. This data was gathered from membership directories and committee reference books. All North American Active Members' race/ethnicity and sex were recorded for each year. The categories for race/ethnicity are Caucasian, East/South/Middle Eastern Asian American (Asian), Hispanic/Latin/South American (HLSA), and African American. RESULTS: From 2010 to 2020, Active Members of POSNA increased from 608 to 818, and the percentage of female (14.6% to 23.7%), Asian (7.4% to 11.2%), HLSA (2.5% to 2.9%), and African American membership (1.6% to 1.8%) increased. Male (85.4% to 76.3%) and Caucasian (88.5% to 84.0%) membership decreased. From 2010 to 2020, male leadership decreased on both the Board of Directors and Committee Chairs (89.5% to 81.8% and 86.4% to 64.7%, respectively), as did Caucasians (94.7% to 81.8% and 90.9% to 88.2%, respectively). The number of Asian members holding positions on both the Board of Directors and Committee Chairs increased (0% to 18.2% and 4.5% to 11.8%, respectively) as did the number of females (10.5% to 18.2% and 13.6% to 35.3%, respectively). HLSA and African American members were proportionally represented in leadership for the years 2010 and 2015. CONCLUSIONS: Membership in POSNA has increased between 2010 to 2020 for every diversity category examined and POSNA membership exhibits significantly more diversity than the orthopaedic specialty as a whole. Leadership as a whole is more diverse in 2020 than it was in 2010. LEVEL OF EVIDENCE: Level II-retrospective.

Microbes associated with fresh produce: Sources, types and methods to reduce spoilage and contamination.

Global food security remains one of the most important challenges that needs to be addressed to ensure the increasing demand for food of the fast growing human population is satisfied. Fruits and vegetables comprise an essential component of a healthy balanced diet as they are the major source of both macro- and micronutrients. They are particularly important for communities in developing countries whose nutrition often relies solely on a plant-based diet. Recent advances in agriculture and food processing technologies have facilitated production of fresh, nutritious and safe food for consumers. However, despite the development of sophisticated chemical and physical methods of food and equipment disinfection, fresh-cut produce and fruit juice industry still faces significant economic losses due to microbial spoilage. Furthermore, fresh produce remains an important source of pathogens that have been causing outbreaks of human illness worldwide. This chapter characterizes common spoilage and human pathogenic microorganisms associated with fresh-cut produce and fruit juice products, and discusses the methods and technology that have been developed and utilized over the years to combat them. Substantial attention is given to highlight advantages and disadvantages of using these methods to reduce microbial spoilage and their efficacy to eliminate human pathogenic microbes associated with consumption of fresh-cut produce and fruit juice products.

Perineal repair of a full-thickness rectal prolapse in a wild Sumatran orangutan (Pongo abelii).

A Delorme's procedure perineal surgical repair was performed in a wild adult male Sumatran orangutan (Pongo abelii) with a chronic persistent rectal prolapse that had been unsuccessfully treated by 6 previous surgeries. The rectal prolapse did not recur, and the orangutan was successfully released to the wild, 6 weeks later.

Sources and contamination routes of microbial pathogens to fresh produce during field cultivation: A review.

Foodborne illness resulting from the consumption of contaminated fresh produce is a common phenomenon and has severe effects on human health together with severe economic and social impacts. The implications of foodborne diseases associated with fresh produce have urged research into the numerous ways and mechanisms through which pathogens may gain access to produce, thereby compromising microbiological safety. This review provides a background on the various sources and pathways through which pathogenic bacteria contaminate fresh produce; the survival and proliferation of pathogens on fresh produce while growing and potential methods to reduce microbial contamination before harvest. Some of the established bacterial contamination sources include contaminated manure, irrigation water, soil, livestock/ wildlife, and numerous factors influence the incidence, fate, transport, survival and proliferation of pathogens in the wide variety of sources where they are found. Once pathogenic bacteria have been introduced into the growing environment, they can colonize and persist on fresh produce using a variety of mechanisms. Overall, microbiological hazards are significant; therefore, ways to reduce sources of contamination and a deeper understanding of pathogen survival and growth on fresh produce in the field are required to reduce risk to human health and the associated economic consequences.

Reconstructing the demographic history of orang-utans using Approximate Bayesian Computation.

Investigating how different evolutionary forces have shaped patterns of DNA variation within and among species requires detailed knowledge of their demographic history. Orang-utans, whose distribution is currently restricted to the South-East Asian islands of Borneo (Pongo pygmaeus) and Sumatra (Pongo abelii), have likely experienced a complex demographic history, influenced by recurrent changes in climate and sea levels, volcanic activities and anthropogenic pressures. Using the most extensive sample set of wild orang-utans to date, we employed an Approximate Bayesian Computation (ABC) approach to test the fit of 12 different demographic scenarios to the observed patterns of variation in autosomal, X-chromosomal, mitochondrial and Y-chromosomal markers. In the best-fitting model, Sumatran orang-utans exhibit a deep split of populations north and south of Lake Toba, probably caused by multiple eruptions of the Toba volcano. In addition, we found signals for a strong decline in all Sumatran populations ~24 ka, probably associated with hunting by human colonizers. In contrast, Bornean orang-utans experienced a severe bottleneck ~135 ka, followed by a population expansion and substructuring starting ~82 ka, which we link to an expansion from a glacial refugium. We showed that orang-utans went through drastic changes in population size and connectedness, caused by recurrent contraction and expansion of rainforest habitat during Pleistocene glaciations and probably hunting by early humans. Our findings emphasize the fact that important aspects of the evolutionary past of species with complex demographic histories might remain obscured when applying overly simplified models.

Generation of SNP datasets for orangutan population genomics using improved reduced-representation sequencing and direct comparisons of SNP calling algorithms.

BACKGROUND: High-throughput sequencing has opened up exciting possibilities in population and conservation genetics by enabling the assessment of genetic variation at genome-wide scales. One approach to reduce genome complexity, i.e. investigating only parts of the genome, is reduced-representation library (RRL) sequencing. Like similar approaches, RRL sequencing reduces ascertainment bias due to simultaneous discovery and genotyping of single-nucleotide polymorphisms (SNPs) and does not require reference genomes. Yet, generating such datasets remains challenging due to laboratory and bioinformatical issues. In the laboratory, current protocols require improvements with regards to sequencing homologous fragments to reduce the number of missing genotypes. From the bioinformatical perspective, the reliance of most studies on a single SNP caller disregards the possibility that different algorithms may produce disparate SNP datasets. RESULTS: We present an improved RRL (iRRL) protocol that maximizes the generation of homologous DNA sequences, thus achieving improved genotyping-by-sequencing efficiency. Our modifications facilitate generation of single-sample libraries, enabling individual genotype assignments instead of pooled-sample analysis. We sequenced ~1% of the orangutan genome with 41-fold median coverage in 31 wild-born individuals from two populations. SNPs and genotypes were called using three different algorithms. We obtained substantially different SNP datasets depending on the SNP caller. Genotype validations revealed that the Unified Genotyper of the Genome Analysis Toolkit and SAMtools performed significantly better than a caller from CLC Genomics Workbench (CLC). Of all conflicting genotype calls, CLC was only correct in 17% of the cases. Furthermore, conflicting genotypes between two algorithms showed a systematic bias in that one caller almost exclusively assigned heterozygotes, while the other one almost exclusively assigned homozygotes. CONCLUSIONS: Our enhanced iRRL approach greatly facilitates genotyping-by-sequencing and thus direct estimates of allele frequencies. Our direct comparison of three commonly used SNP callers emphasizes the need to question the accuracy of SNP and genotype calling, as we obtained considerably different SNP datasets depending on caller algorithms, sequencing depths and filtering criteria. These differences affected scans for signatures of natural selection, but will also exert undue influences on demographic inferences. This study presents the first effort to generate a population genomic dataset for wild-born orangutans with known population provenance.

Laminoplasty in Motion: Evolving Techniques and Complications.

Cervical laminoplasty is an increasingly popular surgical option for the treatment of cervical spondylotic myelopathy (CSM) and ossification of the posterior longitudinal ligament (OPLL). Over the past few decades, there have been substantial developments in both surgical technique and hardware options. As the field of cervical surgery rapidly evolves, there is a timely need to reassess the evolving complications associated with newer techniques. This review aims to synthesize the available literature on cervical laminoplasty and associated mechanical complications pertaining to different laminoplasty hinge fixation options.

Marked population structure and recent migration in the critically endangered Sumatran orangutan (Pongo abelii).

A multitude of factors influence how natural populations are genetically structured, including dispersal barriers, inhomogeneous habitats, and social organization. Such population subdivision is of special concern in endangered species, as it may lead to reduced adaptive potential and inbreeding in local subpopulations, thus increasing the risk of future extinctions. With only 6600 animals left in the wild, Sumatran orangutans (Pongo abelii) are among the most endangered, but also most enigmatic, great ape species. In order to infer the fine-scale population structure and connectivity of Sumatran orangutans, we analyzed the most comprehensive set of samples to date, including mitochondrial hyper-variable region I haplotypes for 123 individuals and genotypes of 27 autosomal microsatellite markers for 109 individuals. For both mitochondrial and autosomal markers, we found a pronounced population structure, caused by major rivers, mountain ridges, and the Toba caldera. We found that genetic diversity and corresponding long-term effective population size estimates vary strongly among sampling regions for mitochondrial DNA, but show remarkable similarity for autosomal markers, hinting at male-driven long-distance gene flow. In support of this, we identified several individuals that were most likely sired by males originating from other genetic clusters. Our results highlight the effect of natural barriers in shaping the genetic structure of great ape populations, but also point toward important dispersal corridors on northern Sumatra that allow for genetic exchange.

Accuracy of Multimodality Fetal Imaging (US, MRI, and CT) for Congenital Musculoskeletal Anomalies.

BACKGROUND: Ultrasonography (US) is the first-line diagnostic tool used to assess fetal musculoskeletal (MSK) anomalies. Associated anomalies in other organ systems may benefit from evaluation via Magnetic Resonance Imaging (MRI). In this study, we compared the diagnostic accuracy of US and MRI to diagnose fetal MSK (primary objective) and non-MSK anomalies (secondary objective). We describe additional findings by low-dose computerized tomography (CT) in two cases incompletely characterized via US and MRI. MATERIALS AND METHODS: This was an IRB-approved retrospective study of consecutive patients with suspected fetal MSK anomalies examined between December 2015 and June 2020. We compared individual MSK and non-MSK anomalies identified via US, MRI, and CT with postnatal outcomes. Sensitivity and specificity for US and MRI were calculated and compared. RESULTS: A total of 31 patients with 112 MSK and 43 non-MSK anomalies were included. The sensitivity of MRI and US for MSK anomalies was not significantly different (76.6% vs. 61.3%, p = 0.3). Low-dose CT identified eight additional skeletal anomalies. MRI diagnosed a higher number of non-MSK anomalies compared to US (81.4% vs. 37.2%, p < 0.05). CONCLUSIONS: Fetal MRI and US have comparable sensitivity for MSK anomalies. In selected cases, low-dose CT may provide additional information. Fetal MRI detected a larger number of non-MSK anomalies in other organ systems compared to US. Multimodality imaging combining all the information provided by MRI, US, and CT, if necessary, ultimately achieved a sensitivity of 89.2% (95% CI: 83.4% to 95.0%) for the diagnosis of musculoskeletal anomalies and 81.4% for additional anomalies in other organs and systems.

C1-C2 Facet Joint Penetration by C2 Pedicle Screws: Influence of Local Anatomy, Bone Mineral Density, and Screw Length.

BACKGROUND: A challenge of C2 pedicle screw placement is to avoid penetration into the C1-C2 facet joint, as this may alter normal biomechanics and accelerate joint degeneration. Our objective was to clarify how local anatomy and surgical technique may relate to C2 pedicle screw penetration into the C1-C2 facet joint. METHODS: C2 pedicle screws were inserted using a fluoroscopically assisted freehand technique. Independent fellowship-trained spine surgeons blindly reviewed intraoperative fluoroscopic and postoperative computed tomography (CT) images for evidence of facet joint penetration (FJP). C2 pedicle morphometry, the sagittal angle of the facet joint, axial and sagittal pedicle screw angles, and screw length were measured on the relevant CT images. RESULTS: A total of 34 patients fulfilled the study criteria, and a total of 68 C2 pedicle screws were placed. Eight screws (16%) penetrated the C1-C2 facet joint. The mean sagittal angle of the C1-C2 facet joint was significantly lower in the FJP group compared with the non-FJP group. The mean sagittal angle of the screws was significantly higher in the FJP group compared with the non-FJP group. The mean screw length was significantly greater for screws causing FJP compared with the non-FJP group. The mean axial screw angle was significantly lower in the FJP group compared with the non-FJP group. Pedicle width, length, height, and transverse angle were not significantly associated with FJP. Independent reviewers were able to identify FJP on intraoperative fluoroscopic imaging in 2 out of 8 cases. CONCLUSION: Lower sagittal angle of the facet joint, higher sagittal angle of the pedicle screw, and screw length >24 mm are associated with higher risk of C1-C2 FJP. When placing C2 pedicle screws under these conditions, caution should be taken to avoid FJP. CLINICAL RELEVANCE: Several anatomical and technical factors may increase the risk of C1-C2 FJP during placement of C2 pedicle screws using a fluoroscopically assisted freehand technique, underscoring the importance of preoperative planning and limiting screw length.

Sex-biased dispersal and volcanic activities shaped phylogeographic patterns of extant Orangutans (genus: Pongo).

The Southeast Asian Sunda archipelago harbors a rich biodiversity with a substantial proportion of endemic species. The evolutionary history of these species has been drastically influenced by environmental forces, such as fluctuating sea levels, climatic changes, and severe volcanic activities. Orangutans (genus: Pongo), the only Asian great apes, are well suited to study the relative impact of these forces due to their well-documented behavioral ecology, strict habitat requirements, and exceptionally slow life history. We investigated the phylogeographic patterns and evolutionary history of orangutans in the light of the complex geological and climatic history of the Sunda archipelago. Our study is based on the most extensive genetic sampling to date, covering the entire range of extant orangutan populations. Using data from three mitochondrial DNA (mtDNA) genes from 112 wild orangutans, we show that Sumatran orangutans, Pongo abelii, are paraphyletic with respect to Bornean orangutans (P. pygmaeus), the only other currently recognized species within this genus. The deepest split in the mtDNA phylogeny of orangutans occurs across the Toba caldera in northern Sumatra and, not as expected, between both islands. Until the recent past, the Toba region has experienced extensive volcanic activity, which has shaped the current phylogeographic patterns. Like their Bornean counterparts, Sumatran orangutans exhibit a strong, yet previously undocumented structuring into four geographical clusters. However, with 3.50 Ma, the Sumatran haplotypes have a much older coalescence than their Bornean counterparts (178 kya). In sharp contrast to the mtDNA data, 18 Y-chromosomal polymorphisms show a much more recent coalescence within Sumatra compared with Borneo. Moreover, the deep geographic structure evident in mtDNA is not reflected in the male population history, strongly suggesting male-biased dispersal. We conclude that volcanic activities have played an important role in the evolutionary history of orangutans and potentially of many other forest-dwelling Sundaland species. Furthermore, we demonstrate that a strong sex bias in dispersal can lead to conflicting patterns in uniparentally inherited markers even at a genus-wide scale, highlighting the need for a combined usage of maternally and paternally inherited marker systems in phylogenetic studies.

Microsatellites for microbiologists.

Microsatellites are repeating sequences of 2-6base pairs of DNA. Currently, they are used as molecular markers in many organisms, specifically in genetic studies analyzing kinship and population structure. In addition, they can be used to study gene duplication and/or deletion. Although they are used in studies on microorganisms including fungi, bacteria, protists, and archaea, it appears that these genetic markers are not being utilized to their full microbiological potential. Microsatellites have many advantages over other genetic markers currently in use as they are in general species specific, and therefore, cross-contamination by nontarget organisms is rare. Furthermore, microsatellites are suitable for use with fast and cheap DNA extraction methods, with ancient DNA or DNA from hair and fecal samples used in noninvasive sampling, making them widely available as a genetic marker. Microsatellites have already proven to be a useful tool for evolutionary studies of pathogenic microorganisms such as Candida albicans and Helicobacter pylori, and the onset of new sequencing techniques (such as 454, PACBIO, and mini-ion sequencing) means the ability to detect such markers will become less time consuming and cheaper, thus further expanding their potential to answer important microbial ecology questions.

Review: metal-based nanoparticles; size, function, and areas for advancement in applied microbiology.

Nanoparticles (NPs) are attracting increased attention in commerce and applied microbiology due to their antimicrobial activity, high electrical conductivity, and optical properties. For example, silver NPs have broad spectrum antimicrobial properties against a wide range of bacteria and fungi, making them ideal for minimizing biofouling. By controlling the size, shape, surface, and agglomeration state of the NPs, specific ion release profiles can be developed for any given application. Currently, NPs are formed in a wide variety of different shapes and sizes including spheres, plates, and wires. This review looks at both commercially and naturally produced NPs with a focus on silver NPs and addresses how these are formed. Furthermore, potential areas for improving these techniques will be highlighted, focusing on advancing shape and structure formation using modern applications. Finally, the review evaluates the feasibility of bioengineering microorganisms to synthesize particles of defined shape and size, by examining genes associated with NP production.

Perceived Physician Empathy in Pediatric Orthopedics: A Cross-Sectional Study.

Empathy is the cornerstone of the patient-physician relationship and is consistently ranked by patients as one of the most important factors in the quality of their care. In this paper we examine the degree to which perceived physician empathy is associated with the characteristics of the caregiver (parent or legal guardian) and physician in pediatric orthopedic surgery. This was a cross-sectional survey study of 200 English-speaking caregivers of pediatric patients at a large children's hospital. The Consultation and Relational Empathy (CARE) Measure was used to measure perceived physician empathy. Only if the caregiver felt carefully listened to by the physician (p-value < 0.001), and if the physician showed respect for what the caregiver had to say (p-value = 0.007) were statistically significant and positively associated with perceived physician empathy. The most significant determinant of perceived physician empathy is whether the caregiver felt listened to during the encounter. Other factors such as caregiver demographics, health literacy, self-rated mental health, wait time, and time spent with the physician do not significantly affect perceived physician empathy.

Ethnic and Sex Diversity in Academic Orthopaedic Surgery: A Cross-sectional Study.

INTRODUCTION: Although the diversity in orthopaedic residency programs has been studied, the diversity within academic orthopaedics has not. METHODS: The board of specialty societies, five leading journals and the National Institutes of Health RePORTER tool, and three accreditation organizations were explored. RESULTS: The board of directors comprised 220 (72%) Caucasians, 36 (12%) Asians, 4 (1%) Hispanic/Latinos, 29 (9%) African Americans, and 18 (6%) Other individuals; 250 (81%) were men, and 57 (19%) were women. The editorial boards comprised 288 (77%) Caucasians, 62 (16%) Asians, 14 (4%) Hispanic/Latinos, 8 (2%) African Americans, and 4 (1%) Other individuals; 341 (91%) were men, and 35 (9%) were women. The National Institutes of Health grant recipients comprised 117 (64%) Caucasians, 58 (32%) Asians, 4 (2%) Hispanic/Latinos, and 3 (2%) African Americans; 128 (70%) were men, and 54 (30%) were women. On average, Caucasians, Asians, Hispanic/Latinos, and African Americans received $776,543, $439,600, $420,182, and $494,049, respectively. On average, men and women received $759,426 and $419,518, respectively. The accreditation boards comprised 45 (82%) Caucasians, 6 (11%) Asians, and 4 (7%) African Americans; 45 (82%) were men, and 10 (18%) were women. CONCLUSIONS: Academic orthopaedic surgery does not resemble the United States. Residency, fellowship committees, and professional organization boards need to become aware of these disparities.

Postoperative Urinary Retention after Pediatric Orthopedic Surgery.

Purpose: This study aims to describe the incidence of postoperative urinary retention among pediatric patients undergoing orthopedic surgery and identify risk factors. Methods: The Pediatric Health Information System was used to identify children aged 1−18 years who underwent orthopedic surgery. Collected from each patient's record were demographic information, principal procedure during hospitalization, the presence of neurologic/neuromuscular conditions and other complex chronic medical conditions, the total postoperative length of stay, and the presence of postoperative urinary retention. Results: The overall incidence of postoperative urinary retention was 0.38%. Children with complex chronic neuromuscular conditions (OR 11.54 (95% CI 9.60−13.88), p = < 0.001) and complex chronic non-neuromuscular medical conditions (OR 5.07 (95% CI 4.11−6.25), p ≤ 0.001) had a substantially increased incidence of urinary retention. Surgeries on the spine (OR 3.98 (95% CI 3.28−4.82, p ≤ 0.001) and femur/hip (OR 3.63 (95% CI 3.03−4.36), p ≤ 0.001) were also associated with an increased incidence. Conclusions: Children with complex chronic neuromuscular conditions have a substantially increased risk of experiencing postoperative urinary retention. Complex chronic non-neuromuscular medical conditions and surgeries to the spine, hip, and femur also carry a notably increased risk.

Quinol-cytochrome c oxidoreductase and cytochrome c4 mediate electron transfer during selenate respiration in Thauera selenatis.

Selenate reductase (SER) from Thauera selenatis is a periplasmic enzyme that has been classified as a type II molybdoenzyme. The enzyme comprises three subunits SerABC, where SerC is an unusual b-heme cytochrome. In the present work the spectropotentiometric characterization of the SerC component and the identification of redox partners to SER are reported. The mid-point redox potential of the b-heme was determined by optical titration (E(m) + 234 +/- 10 mV). A profile of periplasmic c-type cytochromes expressed in T. selenatis under selenate respiring conditions was undertaken. Two c-type cytochromes were purified ( approximately 24 and approximately 6 kDa), and the 24-kDa protein (cytc-Ts4) was shown to donate electrons to SerABC in vitro. Protein sequence of cytc-Ts4 was obtained by N-terminal sequencing and liquid chromatography-tandem mass spectrometry analysis, and based upon sequence similarities, was assigned as a member of cytochrome c(4) family. Redox potentiometry, combined with UV-visible spectroscopy, showed that cytc-Ts4 is a diheme cytochrome with a redox potential of +282 +/- 10 mV, and both hemes are predicted to have His-Met ligation. To identify the membrane-bound electron donors to cytc-Ts4, growth of T. selenatis in the presence of respiratory inhibitors was monitored. The specific quinol-cytochrome c oxidoreductase (QCR) inhibitors myxothiazol and antimycin A partially inhibited selenate respiration, demonstrating that some electron flux is via the QCR. Electron transfer via a QCR and a diheme cytochrome c(4) is a novel route for a member of the DMSO reductase family of molybdoenzymes.