Placental differences between severe fetal growth restriction and hypertensive disorders of pregnancy requiring early preterm delivery: morphometric analysis of the villous tree supported by artificial intelligence.

BACKGROUND: The great obstetrical syndromes of fetal growth restriction and hypertensive disorders of pregnancy can occur individually or be interrelated. Placental pathologic findings often overlap between these conditions, regardless of whether 1 or both diagnoses are present. Quantification of placental villous structures in each of these settings may identify distinct differences in developmental pathways. OBJECTIVE: This study aimed to determine how the quantity and surface area of placental villi and vessels differ between severe, early-onset fetal growth restriction with absent or reversed umbilical artery Doppler indices and hypertensive disorders of pregnancy or the 2 conditions combined among subjects with disease severity that warrant early preterm delivery. We hypothesized that the trajectories of placental morphogenesis diverge after a common initiating insult of deep defective placentation. Specifically, we postulated that only villi are affected in pregnancy-related hypertension, whereas both villous and vascular structures are proportionally diminished in severe fetal growth restriction with no additional effect when hypertension is concomitantly present. STUDY DESIGN: In this retrospective cohort study, paraffin-embedded placental tissue was obtained from 4 groups, namely (1) patients with severe fetal growth restriction with absent or reversed umbilical artery end-diastolic velocities and hypertensive disorders of pregnancy, (2) patients with severe fetal growth restriction with absent or reversed umbilical artery Doppler indices and no hypertension, (3) gestational age-matched, appropriately grown pregnancies with hypertensive disease, and (4) gestational age-matched, appropriately grown pregnancies without hypertension. Dual immunohistochemistry for cytokeratin-7 (trophoblast) and CD34 (endothelial cells) was performed, followed by artificial intelligence-driven morphometric analyses. The number of villi, total villous area, number of fetoplacental vessels, and total vascular area across villi within a uniform region of interest were quantified. Quantitative analyses of placental structures were modeled using linear regression. RESULTS: Placentas from pregnancies complicated by hypertensive disorders of pregnancy exhibited significantly fewer stem villi (-282 stem villi; 95% confidence interval, -467 to -98; P<.01), a smaller stem villous area (-4.3 mm2; 95% confidence interval, -7.3 to -1.2; P<.01), and fewer stem villous vessels (-4967 stem villous vessels; 95% confidence interval, -8501 to -1433; P<.01) with no difference in the total vascular area. In contrast, placental abnormalities in cases with severe growth restriction were limited to terminal villi with global decreases in the number of villi (-873 terminal villi; 95% confidence interval, -1501 to -246; P<.01), the villous area (-1.5 mm2; 95% confidence interval, -2.7 to -0.4; P<.01), the number of blood vessels (-5165 terminal villous vessels; 95% confidence interval, -8201 to -2128; P<.01), and the vascular area (-0.6 mm2; 95% confidence interval, -1.1 to -0.1; P=.02). The combination of hypertension and growth restriction had no additional effect beyond the individual impact of each state. CONCLUSION: Pregnancies complicated by hypertensive disorders of pregnancy exhibited defects in the stem villi only, whereas placental abnormalities in severely growth restricted pregnancies with absent or reversed umbilical artery end-diastolic velocities were limited to the terminal villi. There were no significant statistical interactions in the combination of growth restriction and hypertension, suggesting that distinct pathophysiological pathways downstream of the initial insult of defective placentation are involved in each entity and do not synergize to lead to more severe pathologic consequences. Delineating mechanisms that underly the divergence in placental development after a common inciting event of defective deep placentation may shed light on new targets for prevention or treatment.

Missed Antimicrobial Stewardship Opportunities for Hospitalized Patients with Urinary Tract Infections Receiving Ceftriaxone.

Background: Ceftriaxone is a commonly utilized antibiotic for the treatment of urinary tract infections (UTI) despite the limited literature supporting its use. Opportunities for antimicrobial stewardship (ASP), including IV-to-PO conversions and de-escalation of therapy, are often missed in the hospital setting. Objective: The study reported here describes the utilization of ceftriaxone in patients admitted to the hospital and treated for UTIs in a large health system, focusing on opportunities for IV-to-PO conversion of antibiotic therapy. Methods: This was a multi-center, retrospective, descriptive study conducted in a large health system. Patients admitted from January 2019 to July 2019 were included for analysis if they were 18 years or older at admission, diagnosed with acute cystitis, acute pyelonephritis, or unspecified UTI, and received two or more doses of ceftriaxone. The primary outcome was to evaluate the percentage of patients who were eligible for conversion from IV ceftriaxone to oral antibiotics while admitted to the hospital based on the defined criteria for automatic pharmacist conversion in the health system. Percentage of urine cultures with susceptibility to cefazolin, the duration of antibiotic therapy in the hospital, and an evaluation of oral antibiotics prescribed at discharge were also recorded. Results: A total of 300 patients were included; 88% met the pre-specified criteria for IV-to-PO conversion, but only 12% were converted from IV-to-PO antibiotics during hospitalization. Approximately 65% of patients remained on IV ceftriaxone until discharge, at which time they were converted to a PO antibiotic, most commonly fluoroquinolones followed by third-generation cephalosporins. Conclusion: Patients admitted to the hospital and receiving treatment with ceftriaxone for UTI were infrequently converted to oral therapy prior to discharge despite meeting criteria for automatic pharmacist IV-to-PO conversion. Findings highlight opportunities to contribute to antimicrobial stewardship initiatives across the health system and the importance of tracking and reporting results to frontline providers.

Evaluation of Pseudomonas aeruginosa pathogenesis and therapeutics in military-relevant animal infection models.

Modern combat-related injuries are often associated with acute polytrauma. As a consequence of severe combat-related injuries, a dysregulated immune response results in serious infectious complications. The gram-negative bacterium Pseudomonas aeruginosa is an opportunistic pathogen that often causes life-threatening bloodstream, lung, bone, urinary tract, and wound infections following combat-related injuries. The rise in the number of multidrug-resistant P. aeruginosa strains has elevated its importance to civilian clinicians and military medicine. Development of novel therapeutics and treatment options for P. aeruginosa infections is urgently needed. During the process of drug discovery and therapeutic testing, in vivo testing in animal models is a critical step in the bench-to-bedside approach, and required for Food and Drug Administration approval. Here, we review current and past literature with a focus on combat injury-relevant animal models often used to understand infection development, the interplay between P. aeruginosa and the host, and evaluation of novel treatments. Specifically, this review focuses on the following animal infection models: wound, burn, bone, lung, urinary tract, foreign body, and sepsis.

Synergistic Killing and Re-Sensitization of Pseudomonas aeruginosa to Antibiotics by Phage-Antibiotic Combination Treatment.

Multidrug-resistant (MDR) Pseudomonas aeruginosa infections pose a serious health threat. Bacteriophage-antibiotic combination therapy is a promising candidate for combating these infections. A 5-phage P. aeruginosa cocktail, PAM2H, was tested in combination with antibiotics (ceftazidime, ciprofloxacin, gentamicin, meropenem) to determine if PAM2H enhances antibiotic activity. Combination treatment in vitro resulted in a significant increase in susceptibility of MDR strains to antibiotics. Treatment with ceftazidime (CAZ), meropenem, gentamicin, or ciprofloxacin in the presence of the phage increased the number of P. aeruginosa strains susceptible to these antibiotics by 63%, 56%, 31%, and 81%, respectively. Additionally, in a mouse dorsal wound model, seven of eight mice treated with a combination of CAZ and PAM2H for three days had no detectable bacteria remaining in their wounds on day 4, while all mice treated with CAZ or PAM2H alone had ~107 colony forming units (CFU) remaining in their wounds. P. aeruginosa recovered from mouse wounds post-treatment showed decreased virulence in a wax worm model, and DNA sequencing indicated that the combination treatment prevented mutations in genes encoding known phage receptors. Treatment with PAM2H in combination with antibiotics resulted in the re-sensitization of P. aeruginosa to antibiotics in vitro and a synergistic reduction in bacterial burden in vivo.

Complete Genome Sequences of 10 Phages Lytic against Multidrug-Resistant Pseudomonas aeruginosa.

We report the genome sequences of 10 Pseudomonas aeruginosa phages studied for their potential for formulation of a therapeutic cocktail; they represent the families Myoviridae, Podoviridae, and Siphoviridae Genome sizes ranged from 43,299 to 88,728 nucleotides, with G+C contents of 52.1% to 62.2%. The genomes contained 68 to 168 coding sequences.

Impact of Frequent Administration of Bacteriophage on Therapeutic Efficacy in an A. baumannii Mouse Wound Infection Model.

The spread of multidrug antibiotic resistance (MDR) is a widely recognized crisis in the treatment of bacterial infections, including those occurring in military communities. Recently, the World Health Organization published its first ever list of antibiotic-resistant "priority pathogens" - a catalog of 12 families of bacteria that pose the greatest threat to human health with A. baumannii listed in the "Priority 1: Critical" category of pathogens. With the increasing prevalence of antibiotic resistance and limited development of new classes of antibiotics, alternative antimicrobial therapies are needed, with lytic bacteriophage (phage) specifically targeted against each of the high priority bacterial infections as a potential approach currently in development toward regulatory approval for clinical use. Balb/c mice were prophylactically administered PBS or phage selected against A. baumannii strain AB5075. After 3 weeks, mice were anesthetized, wounded (dorsal), and challenged topically with AB5075. Following infection, mice were subsequently treated with PBS or phage for three consecutive days, and evaluated for 3 weeks to assess the safety and efficacy of the phage treatment relative to the control. We assessed mortality, bacterial burden, time to wound closure, systemic and local cytokine profiles, alterations in host cellular immunity, and finally presence of neutralizing antibodies to the phage mixture. In our study, we found that prophylactic phage administration led to a significant reduction in monocyte-related cytokines in serum compared to mice given PBS. However, we detected no significant changes to circulating blood populations or immune cell populations of secondary lymphoid organs compared to PBS-treated mice. Following prophylactic phage administration, we detected a marked increase in total immunoglobulins in serum, particularly IgG2a and IgG2b. Furthermore, we determined that these antibodies were able to specifically target phage and effectively neutralize their ability to lyse their respective target. In regards to their therapeutic efficacy, administration of phage treatment effectively decreased wound size of mice infected with AB5075 without adverse effects. In conclusion, our data demonstrate that phage can serve as a safe and effective novel therapeutic agent against A. baumannii without adverse reactions to the host and pre-exposure to phage does not seem to adversely affect therapeutic efficacy. This study is an important proof of concept to support the efforts to develop phage as a novel therapeutic product for treatment of complex bacterial wound infections.

Characterization of Acinetobacter baumannii Copper Resistance Reveals a Role in Virulence.

Acinetobacter baumannii is often highly drug-resistant and causes severe infections in compromised patients. These infections can be life threatening due to limited treatment options. Copper is inherently antimicrobial and increasing evidence indicates that copper containing formulations may serve as non-traditional therapeutics against multidrug-resistant bacteria. We previously reported that A. baumannii is sensitive to high concentrations of copper. To understand A. baumannii copper resistance at the molecular level, herein we identified putative copper resistance components and characterized 21 strains bearing mutations in these genes. Eight of the strains displayed a copper sensitive phenotype (pcoA, pcoB, copB, copA/cueO, copR/cusR, copS/cusS, copC, copD); the putative functions of these proteins include copper transport, oxidation, sequestration, and regulation. Importantly, many of these mutant strains still showed increased sensitivity to copper while in a biofilm. Inductively coupled plasma mass spectrometry revealed that many of these strains had defects in copper mobilization, as the mutant strains accumulated more intracellular copper than the wild-type strain. Given the crucial antimicrobial role of copper-mediated killing employed by the immune system, virulence of these mutant strains was investigated in Galleria mellonella; many of the mutant strains were attenuated. Finally, the cusR and copD strains were also investigated in the murine pneumonia model; both were found to be important for full virulence. Thus, copper possesses antimicrobial activity against multidrug-resistant A. baumannii, and copper sensitivity is further increased when copper homeostasis mechanisms are interrupted. Importantly, these proteins are crucial for full virulence of A. baumannii and may represent novel drug targets.

How Postbaccalaureate Career Changer and Traditional Medical Students Differ Academically.

OBJECTIVE: This retrospective descriptive study compared the academic performance of postbaccalaureate career changer students with that of traditional students during the classroom-based, science-dominated early years of medical school. Earlier studies documented the eventual success of nontraditional medical students, although we found little information specific to the medical school performance of career changers. Our objective was to determine whether postbaccalaureate career changer medical students perform differently from traditionally prepared medical students in the science-dominated early years of medical school classroom education. METHODS: This study analyzed the admission data and academic performance of medical students at the University of Central Florida College of Medicine across 8 years (N = 630). Differences in performance were assessed using examination grades from the first 2 years of medical school, and US Medical Licensing Examination (USMLE) Step 1 and Step 2 scores. RESULTS: Statistically significant differences were found between traditional and career changer students for all science modules in year 1, and 4 of the 5 modules in year 2. Traditional students performed better on USMLE Step 1. Significant differences between the groups disappeared by USMLE Step 2. CONCLUSIONS: Career changer medical students show a small, persistent academic lag in the first 2 years of medical school and on USMLE Step 1 scores. By USMLE Step 2 the difference disappears. Similar undergraduate grade point averages and Medical College Admission Test scores suggest that science exposure, not ability may explain these differences. An unexpected finding is the number of career changer students is not increasing proportional to the proliferation of postbaccalaureate programs in the United States. This study may benefit student advisors and residency directors, and, it is hoped, provide reassurance to career changer students.

Precarious sleep? Nonstandard work, gender, and sleep disturbance in 31 European countries.

Despite the advent of precarious work, little is known about how this form of employment can generate disparities in sleep outcomes. We extend existing work by providing a theoretical framework linking different measures of work precarity to sleep problems. We argue that the association between objective precarious working conditions and sleep disturbance is channeled through and mediated by subjective work precarity. We further argue that gender moderates the relationship between objective and subjective work precarity. We test this theoretical framework using the 2010 European Working Conditions Survey. Our results indicate that objective precarious working conditions undermine sleep by promoting the subjective experience of insecurity. Furthermore, the indirect effect of objective precarious work on sleep disturbance through subjective employment insecurity varies by gender: compared to women in similar working conditions, men report higher levels of subjective precarity. This research makes important contributions to the literatures on the health consequences of nonstandard work and social determinants of well-being.

Crop Rotational Effects on Yield Formation in Current Sugar Beet Production - Results From a Farm Survey and Field Trials.

In Europe, the framework for sugar beet (Beta vulgaris L.) production was subject to considerable changes and for the future it is expected that sugar beet cultivation might concentrate around the sugar factories for economic reasons. Based on data from a national sugar beet farmers' survey and multi-year crop rotation trials, the effects of cropping interval (number of years in between two subsequent sugar beet crops) and of preceding crops on sugar yield were elucidated under current Central European management conditions. The dominating sugar beet cropping interval was ≥4 years in the farm survey with pronounced differences between regions. However, the cropping intervals 2, 3, and ≥4 years did not affect the sugar yield. Therefore, significant differences in sugar yield between regions were assumed to be caused by multiple interactions between year, site, and farmers' skills. Throughout Germany, the dominating preceding crops in sugar beet cultivation were winter wheat (Triticum aestivum L.) and winter barley (Hordeum vulgare L.). In the field trials, the sugar yield was 5% higher after pea (Pisum sativum L.) compared to maize (Zea mays L.) as preceding crop, while differences between the preceding crops pea and winter wheat, and wheat and maize were not significant. Repeated measurements of canopy development and leaf color during the growing season revealed a higher N-availability after pea as preceding crop. However, decreased growth after maize was not completely compensated for by high N-fertilizer doses. Overall, the causes for the differences in sugar yield between the preceding crops remained open. The results do not support concerns about substantial yield losses in sugar beet production due to a reduction in the cropping interval from 3 to 2 years. Nevertheless, short rotations with maize and sugar beet might increase the risk of Rhizoctonia solani crown and root rot infestation. Leguminous crops such as pea offer the potential for higher sugar beet yield with lower N-fertilizer doses.

Update on the use of direct oral anticoagulants for the prevention and treatment of thromboembolism.

Following publication of our review article 4 years ago, there has been an uptake in the use of nonvitamin K oral antagonists, also known as direct oral anticoagulants. The most recent Xa inhibitors to receive approval are edoxaban, which has been approved for use in both atrial fibrillation and venous thromboembolism prevention and betrixaban, which has been approved in the USA for extended thromboprophylaxis in the medically ill population. Additional analyses of certain types of atrial fibrillation patients have now become available. Ongoing prescriber vigilance is recommended as additional information continues to emerge with this class of medications. The purpose of this paper is to provide an update on the use of the direct oral anticoagulant agents for the prevention and treatment of thromboembolism.

1,2,4-Triazolidine-3-thiones as Narrow Spectrum Antibiotics against Multidrug-Resistant Acinetobacter baumannii.

With only two new classes of antibiotics developed in the last 40 years, novel antibiotics are desperately needed to combat the growing problem of multidrug-resistant and extensively drug resistant bacteria, particularly Gram-negative bacteria. Described in this letter is the synthesis and antibiotic activity of 1,2,4-triazolidine-3-thiones as narrow spectrum antibiotics. Optimization of the 1,2,4-triazolidine-3-thione scaffold identified a small molecule with potent antibiotic activity against multiple strains of multidrug-resistant and extensively drug-resistant Acinetobacter baumannii. This small molecule also shows single dose, in vivo activity in a Galleria mellonella infection model with A. baumannii and represents a promising start in the development of a class of drugs that can target this bacterial pathogen.

1,2,4-Triazolidine-3-thiones Have Specific Activity against Acinetobacter baumannii among Common Nosocomial Pathogens.

Acinetobacter baumannii are Gram-negative bacilli that pose a constant threat to susceptible patients because of increased resistance to multiple antibiotics and persistence in the hospital environment. After genome analysis, we discovered that A. baumannii harbors genes that share homology to an enzymatic pathway that elongates long-chain fatty acids (LCFA) in fungi. Previously, 1,2,4-triazolidine-3-thiones (T-3-Ts) were shown to inhibit hyphae production in fungi, and this same LCFA elongation pathway was implicated as the possible target. Therefore, we investigated if T-3-Ts also have activity against multidrug-resistant A. baumannii. Surprisingly, all of the clinical isolates of A. baumannii that were tested have susceptibility to ECC145 and ECC188 with MIC90 values of 8.0 µg/mL. In contrast, reference strains and clinical isolates of other common nosocomial bacteria that lack the LCFA pathway also lacked susceptibility. Time-kill experiments revealed that both ECC145 and ECC188 have a bacteriostatic effect against A. baumannii. Mass spectrometry analysis suggested that exposure to T-3-Ts resulted in less LCFA production. Supplementation of media with either 0.02% w/v oleic or linoleic acid abrogated the bacteriostatic effect of the compounds, which again implicated LCFA elongation as the target. Our results suggest these molecules could be a promising start to further exploit what appears to be an important aspect of A. baumannii membrane function and integrity.

Personalized Therapeutic Cocktail of Wild Environmental Phages Rescues Mice from Acinetobacter baumannii Wound Infections.

Multidrug-resistant bacterial pathogens are an increasing threat to public health, and lytic bacteriophages have reemerged as a potential therapeutic option. In this work, we isolated and assembled a five-member cocktail of wild phages against Acinetobacter baumannii and demonstrated therapeutic efficacy in a mouse full-thickness dorsal infected wound model. The cocktail lowers the bioburden in the wound, prevents the spread of infection and necrosis to surrounding tissue, and decreases infection-associated morbidity. Interestingly, this effective cocktail is composed of four phages that do not kill the parent strain of the infection and one phage that simply delays bacterial growth in vitro via a strong but incomplete selection event. The cocktail here appears to function in a combinatorial manner, as one constituent phage targets capsulated A. baumannii bacteria and selects for loss of receptor, shifting the population to an uncapsulated state that is then sensitized to the remaining four phages in the cocktail. Additionally, capsule is a known virulence factor for A. baumannii, and we demonstrated that the emergent uncapsulated bacteria are avirulent in a Galleria mellonella model. These results highlight the importance of anticipating population changes during phage therapy and designing intelligent cocktails to control emergent strains, as well as the benefits of using phages that target virulence factors. Because of the efficacy of this cocktail isolated from a limited environmental pool, we have established a pipeline for developing new phage therapeutics against additional clinically relevant multidrug-resistant pathogens by using environmental phages sourced from around the globe.

Employment Transitions, Child Care Conflict, and the Mental Health of Low-Income Urban Women With Children.

OBJECTIVE: Although studies suggest that employment promotes mental health, it is unclear whether this pattern extends to low-income urban women with children who are disproportionately employed in unstable jobs and often unable to obtain child care. In this paper, we consider whether becoming employed reduces symptoms of psychological distress among low-income women with children. We also assess whether having trouble securing adequate child care offsets these benefits. STUDY DESIGN: We use longitudinal data from the Welfare, Children, and Families project, a probability sample of low-income women with children living in Boston, Chicago, and San Antonio, to test whether becoming employed reduces symptoms of psychological distress over time and whether having trouble securing child care moderates this association. RESULTS: We find that employment is associated with lower levels of distress among women who have no trouble with child care and higher levels of distress among women who struggle with child care. CONCLUSION: Taken together, our results suggest that valuing the benefits of paid work over unpaid work is an oversimplification and that the emphasis on placing poor women with children into paid work could be misguided. Policies that focus on moving low-income women off of government assistance and into paid work could be more effective if greater resources were devoted to increasing access to quality child care.

Environmental impacts of different crop rotations in terms of soil compaction.

Avoiding soil compaction caused by agricultural management is a key aim of sustainable land management, and the soil compaction risk should be considered when assessing the environmental impacts of land use systems. Therefore this project compares different crop rotations in terms of soil structure and the soil compaction risk. It is based on a field trial in Germany, in which the crop rotations (i) silage maize (SM) monoculture, (ii) catch crop mustard (Mu)_sugar beet (SB)-winter wheat (WW)-WW, (iii) Mu_SM-WW-WW and (iv) SB-WW-Mu_SM are established since 2010. Based on the cultivation dates, the operation specific soil compaction risks and the soil compaction risk of the entire crop rotations are modelled at two soil depths (20 and 35 cm). To this end, based on assumptions of the equipment currently used in practice by a model farm, two scenarios are modelled (100 and 50% hopper load for SB and WW harvest). In addition, after one complete rotation, in 2013 and in 2014, the physical soil parameters saturated hydraulic conductivity (kS) and air capacity (AC) were determined at soil depths 2-8, 12-18, 22-28 and 32-38 cm in order to quantify the soil structure. At both soil depths, the modelled soil compaction risks for the crop rotations including SB (Mu_SB-WW-WW, SB-WW-Mu_SM) are higher (20 cm: medium to very high risks; 35 cm: no to medium risks) than for those without SB (SM monoculture, Mu_SM-WW-WW; 20 cm: medium risks; 35 cm: no to low risks). This increased soil compaction risk is largely influenced by the SB harvest in years where soil water content is high. Halving the hopper load and adjusting the tyre inflation pressure reduces the soil compaction risk for the crop rotation as a whole. Under these conditions, there are no to low soil compaction risks for all variants in the subsoil (soil depth 35 cm). Soil structure is mainly influenced in the topsoil (2-8 cm) related to the cultivation of Mu as a catch crop and WW as a preceding crop. Concerning kS, Mu_SB-WW-WW (240 cm d(-1)) and Mu_SM-WW-WW (196 cm d(-1)) displayed significantly higher values than the SM monoculture (67 cm d(-1)), indicating better structural stability and infiltration capacity. At other soil depths, and for the parameter AC, there are no systematic differences in soil structure between the variants. Under the circumstances described, all crop rotations investigated are not associated with environmental impacts caused by soil compaction.

Evaluation of Parameters for High Efficiency Transformation of Acinetobacter baumannii.

Acinetobacter baumannii is an emerging, nosocomial pathogen that is poorly characterized due to a paucity of genetic tools and methods. While whole genome sequence data from several epidemic and environmental strains have recently become available, the functional characterization of genes is significantly lagging. Efficient transformation is one of the first steps to develop molecular tools that can be used to address these shortcomings. Here we report parameters allowing high efficiency transformation of A. baumannii. Using a multi-factorial experimental design we found that growth phase, voltage, and resistance all significantly contribute to transformation efficiency. The highest efficiency (4.3 × 10(8) Transformants/µg DNA) was obtained at the stationary growth phase of the bacterium (OD 6.0) using 25 ng of plasmid DNA under 100 Ohms resistance and 1.7 kV/cm voltage. The optimized electroporation parameters reported here provide a useful tool for genetic manipulation of A. baumannii.

Laboratory Maintenance of Acinetobacter baumannii.

Acinetobacter baumannii has recently drawn great interest in the microbiology research community due to the increase in clinical antibiotic resistance of this organism, and persistence of this bacterial species in the hospital environment. This unit outlines protocols for the growth and maintenance of A. baumannii in the laboratory.

Genetic Manipulation of Acinetobacter baumannii.

Acinetobacter baumannii is a Gram-negative nosocomial pathogen of clinical importance. A lack of genetic tools has hindered the research of this organism in the past; however, recently, various methods have been designed, modified, and optimized to facilitate the genetic manipulation of A. baumannii. This unit describes some of the recent genetic advances and new recombinant tools developed for this pathogen, including standard transformation and conjugation techniques specifically developed for the bacteria. As the need to understand the basic biology of A. baumannii increases with the prospect of developing new therapeutics, the use of the basic genetic methods herein can provide the critical first step to identify genes required for infection.