Transitioning from conventional to cover crop systems with minimum tillage does not alter nutrient loading.

Nutrient loading from conventional row-crop production systems impairs surface waterbodies in the mid-southern United States. This study was conducted to determine whether minimum tillage and winter cover crops can decrease nutrient loading in surface runoff from conventionally tilled row-crop fields. The effects of winter cover crops and minimum tillage on N and P loading from a corn (Zea mays L.)-soybean [Glycine max (L.) Merr.] rotation system were investigated on production fields in northwestern Mississippi using a split-field approach. As measured at the edge of the field, minimum tillage with cover crops had no effect on surface runoff from production fields regarding N or P loading (p > .10 for all nutrient loads), discharge (p > .10), or loss of suspended solids (p > .10). Minimum tillage and cover crops decreased sediment and nutrient concentrations in runoff for total N (p = .05) and total P (p = .09) but had no effect on other nutrients of interest. Although these practices decreased total N concentration by 36% in surface runoff to receiving waters, this reduction was only seen when aboveground cover crop biomass was present (p = .07). Regardless of the time of year, minimum tillage with cover crops decreased total P concentration in surface runoff by 27% (p = .09). These data indicate that it is unlikely that minimum tillage and cover crops will affect N and P loading while transitioning to a conservation production system in the mid-southern United States.

ACOEM OEM Core Competencies: 2021.

Competency is defined as possession of sufficient physical, intellectual, and behavioral qualifications to perform a task or serve in a role which adequately accomplishes a desired outcome. Recognizing the need for defining competencies essential to occupational and environmental medicine (OEM) physicians, the American College of Occupational and Environmental Medicine developed its first set of OEM Competencies in 1998. Later updated in 2008, and again in 2014, the increasing globalization and modernization of the workplace, along with published research on OEM practice, required an update to ensure OEM physicians stay current with the field and practice of OEM. Delineation of core competencies for the profession provides employers, government agencies, health care organizations, and other health practitioners a solid context of the role and expertise of OEM physicians.

A preliminary investigation of wild pig (Sus scrofa) impacts in water quality.

The United States, particularly the southern portion, has recently suffered drastic population expansion of wild pigs causing destruction of prime farmland. An associated concern, which has been understudied, is the potential transfer of nutrients and pathogens to surface water. This study aimed to identify the abiotic and biotic impacts of captive wild pigs on water quality, including nutrients, fecal indicator and pathogenic bacteria, and antimicrobial resistance. Overall, the study demonstrated that wild pigs harbored Salmonella spp., Campylobacter spp., Escherichia coli, and Clostridium perfringens, which were found in water runoff collected directly beneath the hog paddock, often 2 log10 greater than above-paddock levels. However, the impacts to downstream water quality were limited, perhaps because of a relatively large riparian buffer between the paddock and surface water. A higher rate of ammonium concentration changes over time was detected in the runoff water below the paddock; additionally, microbial releases detected in runoff were also time dependent, possibly associated with increasing pig numbers. Antibiotic resistance was generally not associated with the wild pigs. Antibiotic resistance genes were found in upstream as well as downstream surface water, suggesting that nonpoint sources of microbial contamination were present. Interestingly, intI1 levels were greater in below-paddock runoff by nearly 2 log10 . Overall, it appears that wild pigs potentially pose a threat to water quality but only if they have direct access to the water. Pathogen, fecal indicator bacteria, and some nutrient release were significantly associated with wild pigs, but riparian buffers limited water quality impairment.

Using cameras to index waterfowl abundance in winter-flooded rice fields.

Extensive wetland habitat loss across the continental United States has caused post-harvested rice fields to become an important surrogate wetland habitat for migratory waterfowl. Flooded rice fields used by waterfowl have the potential to provide agronomic benefits to soil. Increasing interest in the reciprocal relationship between birds and flooded rice fields has given rise to many studies that aim to quantify bird abundance. However, surveying large flocks of birds in open agricultural fields is challenging because traditional ground and aerial surveys can cause birds to flush or re-allocate spatially, thus biasing counts that are reflected in following management practice recommendations. To avoid this, we used camera surveys and an open-access image manipulation program to estimate 24-h bird use of rice fields. Indices of bird abundance from counts were used to estimate fecal matter input to rice fields. Camera surveys have the potential to limit biases seen in other methods because of their ability to capture bird use over a 24-h period over an entire season and the ability for multiple researchers to survey the same site.•Surveying bird flocks by traditional ground or aerial surveys can bias bird abundance estimates.•Camera surveys of waterfowl in rice fields were used to estimate bird abundance and fecal matter input.•Camera surveys reflect static bird use over 24-h which can lower bias seen in traditional methods.

Investigating the role of organic carbon amendments and microbial denitrification gene abundance in nitrogen removal from experimental agricultural drainage ditches with low-grade weirs.

Low-grade weirs placed within agricultural drainage ditches in the Lower Mississippi Alluvial Valley can be used as a management practice to enhance nitrogen removal. The addition of organic carbon amendments within ditches that contain weirs could further increase nitrogen removal. Through repeated trials, changes in NO 3 - -N concentration between inflow and outflow were variable in the ditch without weirs, while only decreases in concentration were observed in ditches with weirs. Significant differences in NO 3 - -N concentrations were observed between treatments, with greater removal of NO 3 - -N observed in dissolved organic carbon treatments compared to control and particulate organic carbon treatments. At medium- and high-flow rates, respectively, dissolved organic carbon treatments resulted in greater NO 3 - -N concentration decreases of 31.6% and 27.1% compared to 19% and 11.6% in particulate organic carbon treatments and 18.6% and 17.2% in control treatments. Significant effects of weirs and sampling date on nirS, nirK, nosZ, and 16S rRNA gene abundances were observed. Observed increases in NO 3 - -N removal with organic carbon amendments, provides support for continued investigation on improving the efficacy of organic carbon amendments as a best management practice for NO 3 - -N removal in agricultural drainage ditches. PRACTITIONER POINTS: Dissolved organic carbon amendments increased nitrate-nitrogen removal. Only decreases in nitrate-nitrogen concentration were observed in ditches with weirs. Increasing flow rate did not affect nitrate-nitrogen removal. Abundance of denitrification-performing microbes likely did not affect N removal. Lack of anaerobic soil conditions and short residence time reduced nitrate-N removal.

Arsenic Exposure, Assessment, Toxicity, Diagnosis, and Management: Guidance for Occupational and Environmental Physicians.

: Arsenic is ubiquitous in the environment and human exposure can occur from multiple possible routes including diet. Occupational medicine physicians asked to evaluate workers with elevated urine arsenic levels may be unaware that many sources of arsenic exposure are not work related. In this paper, we address arsenic exposure sources and pathways, adverse health effects of arsenic exposure and those subpopulations at increased risk, and the evaluation and treatment of those exposed to elevated arsenic levels.

Effects of Low-Grade Weirs on Soil Microbial Communities in Agricultural Drainage Ditches.

Agricultural fertilizer application throughout the Mississippi River basin has been identified as a major source of N pollution to the Gulf of Mexico. Using best management practices, such as low-grade weirs, has been identified as a potential solution to mitigate nutrient loads in agricultural runoff. This study assessed impacts of weir implementation in four agricultural drainage ditches (three with weirs and one control site) in the Mississippi Delta. Soil samples collected from field locations in spring 2013 were analyzed for denitrifier abundance using genes (16s ribosomal RNA [rRNA] genes, , , and ) via quantitative polymerase chain reaction (qPCR), microbial community profiles via terminal-restriction fragment length polymorphism (T-RFLP) of 16s rRNA genes, soil parameters (C, N, and moisture), and vegetation presence at sample locations. Gene quantification was successful, except for , which was found below detection limits (5000 gene copies g soil). Distance from weirs was negatively correlated with 16S rRNA genes and soil moisture, and soil moisture was positively correlated with 16s rRNA and S gene abundance. Results of empirical Bayesian kriging did not exhibit obvious patterns of microbial diversity in relation to weir proximity. Preliminary assessment of seasonal trends showed genes 16s rRNA and , soil N, and mean T-RF values to be greater in fall than in spring. Results highlight that weirs had no direct impact on microbial diversity or denitrification functional gene abundance. Correlations between microbial measures and environmental parameters suggest that adequate management of N runoff from agricultural landscapes will require ecological engineering beyond weirs to optimize N mitigation.

Haemophilus ducreyi Seeks Alternative Carbon Sources and Adapts to Nutrient Stress and Anaerobiosis during Experimental Infection of Human Volunteers.

Haemophilus ducreyi causes the sexually transmitted disease chancroid in adults and cutaneous ulcers in children. In humans, H. ducreyi resides in an abscess and must adapt to a variety of stresses. Previous studies (D. Gangaiah, M. Labandeira-Rey, X. Zhang, K. R. Fortney, S. Ellinger, B. Zwickl, B. Baker, Y. Liu, D. M. Janowicz, B. P. Katz, C. A. Brautigam, R. S. MunsonJr, E. J. Hansen, and S. M. Spinola, mBio 5:e01081-13, 2014, http://dx.doi.org/10.1128/mBio.01081-13) suggested that H. ducreyi encounters growth conditions in human lesions resembling those found in stationary phase. However, how H. ducreyi transcriptionally responds to stress during human infection is unknown. Here, we determined the H. ducreyi transcriptome in biopsy specimens of human lesions and compared it to the transcriptomes of bacteria grown to mid-log, transition, and stationary phases. Multidimensional scaling showed that the in vivo transcriptome is distinct from those of in vitro growth. Compared to the inoculum (mid-log-phase bacteria), H. ducreyi harvested from pustules differentially expressed ∼93 genes, of which 62 were upregulated. The upregulated genes encode homologs of proteins involved in nutrient transport, alternative carbon pathways (l-ascorbate utilization and metabolism), growth arrest response, heat shock response, DNA recombination, and anaerobiosis. H. ducreyi upregulated few genes (hgbA, flp-tad, and lspB-lspA2) encoding virulence determinants required for human infection. Most genes regulated by CpxRA, RpoE, Hfq, (p)ppGpp, and DksA, which control the expression of virulence determinants and adaptation to a variety of stresses, were not differentially expressed in vivo, suggesting that these systems are cycling on and off during infection. Taken together, these data suggest that the in vivo transcriptome is distinct from those of in vitro growth and that adaptation to nutrient stress and anaerobiosis is crucial for H. ducreyi survival in humans.

Investigation of denitrifying microbial communities within an agricultural drainage system fitted with low-grade weirs.

Enhancing wetland characteristics in agricultural drainage ditches with the use of low-grade weirs, has been identified as a best management practice (BMP) to mitigate nutrient runoff from agriculture landscapes. A major objective of utilizing low-grade weirs as a BMP includes fostering environments suitable for the biogeochemical removal of nitrogen via denitrification. This study examined the spatial resolution of microbial communities involved in denitrification in agricultural drainage systems fitted with low-grade weirs. Appropriate sampling scales of microbial communities were investigated using 16S rRNA and denitrification functional genes nosZ, nirS, and nirK via quantitative polymerase chain reaction (qPCR) and terminal-restriction fragment length polymorphism (T-RFLP) analysis. Genes 16S rRNA, nosZ, and nirS were all successfully detected in soil samples, while nirK was below the detection limit throughout the study. Utilizing a combination of three sampling regimes (management, reach, catchment) was found to be effective in capturing microbial community patterns, as ANOVA results revealed nosZ gene abundance was significantly greater at the management rather than reach scale (p = 0.045; F = 3.311), although, no significant differences were observed in 16S rRNA or nirS between sampling scales (p > 0.05). A Pearson correlation matrix confirmed that 16S rRNA and nosZ gene abundances were positively correlated with soil carbon (C), nitrogen (N), and moisture, while nirS abundance was only positively correlated with soil C and soil moisture. This highlights the potential for wetland-like characteristics to be recovered in agricultural drainage systems, as weir proximity is observed to enhance soil moisture and conditions for N remediation. This study provides the basis for additional investigations of these unique environments in the Mississippi Alluvial Valley and a starting point for adaptive management to enhance agricultural drainage systems for microbial communities towards nutrient remediation goals.

DksA and (p)ppGpp have unique and overlapping contributions to Haemophilus ducreyi pathogenesis in humans.

The (p)ppGpp-mediated stringent response is important for bacterial survival in nutrient limiting conditions. For maximal effect, (p)ppGpp interacts with the cofactor DksA, which stabilizes (p)ppGpp's interaction with RNA polymerase. We previously demonstrated that (p)ppGpp was required for the virulence of Haemophilus ducreyi in humans. Here, we constructed an H. ducreyi dksA mutant and showed it was also partially attenuated for pustule formation in human volunteers. To understand the roles of (p)ppGpp and DksA in gene regulation in H. ducreyi, we defined genes potentially altered by (p)ppGpp and DksA deficiency using transcriptome sequencing (RNA-seq). In bacteria collected at stationary phase, lack of (p)ppGpp and DksA altered expression of 28% and 17% of H. ducreyi open reading frames, respectively, including genes involved in transcription, translation, and metabolism. There was significant overlap in genes differentially expressed in the (p)ppGpp mutant relative to the dksA mutant. Loss of (p)ppGpp or DksA resulted in the dysregulation of several known virulence determinants. Deletion of dksA downregulated lspB and rendered the organism less resistant to phagocytosis and increased its sensitivity to oxidative stress. Both mutants had reduced ability to attach to human foreskin fibroblasts; the defect correlated with reduced expression of the Flp adhesin proteins in the (p)ppGpp mutant but not in the dksA mutant, suggesting that DksA regulates the expression of an unknown cofactor(s) required for Flp-mediated adherence. We conclude that both (p)ppGpp and DksA serve as major regulators of H. ducreyi gene expression in stationary phase and have both overlapping and unique contributions to pathogenesis.

Overlapping and complementary oxidative stress defense mechanisms in nontypeable Haemophilus influenzae.

The Gram-negative commensal bacterium nontypeable Haemophilus influenzae (NTHI) can cause respiratory tract diseases that include otitis media, sinusitis, exacerbations of chronic obstructive pulmonary disease, and bronchitis. During colonization and infection, NTHI withstands oxidative stress generated by reactive oxygen species produced endogenously, by the host, and by other copathogens and flora. These reactive oxygen species include superoxide, hydrogen peroxide (H2O2), and hydroxyl radicals, whose killing is amplified by iron via the Fenton reaction. We previously identified genes that encode proteins with putative roles in protection of the NTHI isolate strain 86-028NP against oxidative stress. These include catalase (HktE), peroxiredoxin/glutaredoxin (PgdX), and a ferritin-like protein (Dps). Strains were generated with mutations in hktE, pgdX, and dps. The hktE mutant and a pgdX hktE double mutant were more sensitive than the parent to killing by H2O2. Conversely, the pgdX mutant was more resistant to H2O2 due to increased catalase activity. Supporting the role of killing via the Fenton reaction, binding of iron by Dps significantly mitigated the effect of H2O2-mediated killing. NTHI thus utilizes several effectors to resist oxidative stress, and regulation of free iron is critical to this protection. These mechanisms will be important for successful colonization and infection by this opportunistic human pathogen.

Haemophilus ducreyi RpoE and CpxRA appear to play distinct yet complementary roles in regulation of envelope-related functions.

Haemophilus ducreyi causes the sexually transmitted disease chancroid and a chronic limb ulceration syndrome in children. In humans, H. ducreyi is found in an abscess and overcomes a hostile environment to establish infection. To sense and respond to membrane stress, bacteria utilize two-component systems (TCSs) and extracytoplasmic function (ECF) sigma factors. We previously showed that activation of CpxRA, the only intact TCS in H. ducreyi, does not regulate homologues of envelope protein folding factors but does downregulate genes encoding envelope-localized proteins, including many virulence determinants. H. ducreyi also harbors a homologue of RpoE, which is the only ECF sigma factor in the organism. To potentially understand how H. ducreyi responds to membrane stress, here we defined RpoE-dependent genes using transcriptome sequencing (RNA-Seq). We identified 180 RpoE-dependent genes, of which 98% were upregulated; a major set of these genes encodes homologues of envelope maintenance and repair factors. We also identified and validated a putative RpoE promoter consensus sequence, which was enriched in the majority of RpoE-dependent targets. Comparison of RpoE-dependent genes to those controlled by CpxR showed that each transcription factor regulated a distinct set of genes. Given that RpoE activated a large number of genes encoding envelope maintenance and repair factors and that CpxRA represses genes encoding envelope-localized proteins, these data suggest that RpoE and CpxRA appear to play distinct yet complementary roles in regulating envelope homeostasis in H. ducreyi.

HrrF is the Fur-regulated small RNA in nontypeable Haemophilus influenzae.

Nontypeable Haemophilus influenzae (NTHi) are Gram-negative commensal bacteria that reside in the nasopharynx. NTHi can also cause multiple upper and lower respiratory tract diseases that include sinusitis, conjunctivitis, bronchitis, and otitis media. In numerous bacterial species the ferric uptake regulator (Fur) acts as a global regulator of iron homeostasis by negatively regulating the expression of iron uptake systems. However in NTHi strain 86-028NP and numerous other bacterial species there are multiple instances where Fur positively affects gene expression. It is known that many instances of positive regulation by Fur occur indirectly through a small RNA intermediate. However, no examples of small RNAs have been described in NTHi. Therefore we used RNA-Seq analysis to analyze the transcriptome of NTHi strain 86-028NPrpsL and an isogenic 86-028NPrpsLΔfur strain to identify Fur-regulated intergenic transcripts. From this analysis we identified HrrF, the first small RNA described in any Haemophilus species. Orthologues of this small RNA exist only among other Pasteurellaceae. Our analysis showed that HrrF is maximally expressed when iron levels are low. Additionally, Fur was shown to bind upstream of the hrrF promoter. RNA-Seq analysis was used to identify targets of HrrF which include genes whose products are involved in molybdate uptake, deoxyribonucleotide synthesis, and amino acid biosynthesis. The stability of HrrF is not dependent on the RNA chaperone Hfq. This study is the first step in an effort to investigate the role small RNAs play in altering gene expression in response to iron limitation in NTHi.

Identifying non-point sources of endocrine active compounds and their biological impacts in freshwater lakes.

Contaminants of emerging concern, particularly endocrine active compounds (EACs), have been identified as a threat to aquatic wildlife. However, little is known about the impact of EACs on lakes through groundwater from onsite wastewater treatment systems (OWTS). This study aims to identify specific contributions of OWTS to Sullivan Lake, Minnesota, USA. Lake hydrology, water chemistry, caged bluegill sunfish (Lepomis macrochirus), and larval fathead minnow (Pimephales promelas) exposures were used to assess whether EACs entered the lake through OWTS inflow and the resultant biological impact on fish. Study areas included two OWTS-influenced near-shore sites with native bluegill spawning habitats and two in-lake control sites without nearby EAC sources. Caged bluegill sunfish were analyzed for plasma vitellogenin concentrations, organosomatic indices, and histological pathologies. Surface and porewater was collected from each site and analyzed for EACs. Porewater was also collected for laboratory exposure of larval fathead minnow, before analysis of predator escape performance and gene expression profiles. Chemical analysis showed EACs present at low concentrations at each study site, whereas discrete variations were reported between sites and between summer and fall samplings. Body condition index and liver vacuolization of sunfish were found to differ among study sites as did gene expression in exposed larval fathead minnows. Interestingly, biological exposure data and water chemistry did not match. Therefore, although results highlight the potential impacts of seepage from OWTS, further investigation of mixture effects and life history factor as well as chemical fate is warranted.

Haemophilus ducreyi Hfq contributes to virulence gene regulation as cells enter stationary phase.

UNLABELLED: To adapt to stresses encountered in stationary phase, Gram-negative bacteria utilize the alternative sigma factor RpoS. However, some species lack RpoS; thus, it is unclear how stationary-phase adaptation is regulated in these organisms. Here we defined the growth-phase-dependent transcriptomes of Haemophilus ducreyi, which lacks an RpoS homolog. Compared to mid-log-phase organisms, cells harvested from the stationary phase upregulated genes encoding several virulence determinants and a homolog of hfq. Insertional inactivation of hfq altered the expression of ~16% of the H. ducreyi genes. Importantly, there were a significant overlap and an inverse correlation in the transcript levels of genes differentially expressed in the hfq inactivation mutant relative to its parent and the genes differentially expressed in stationary phase relative to mid-log phase in the parent. Inactivation of hfq downregulated genes in the flp-tad and lspB-lspA2 operons, which encode several virulence determinants. To comply with FDA guidelines for human inoculation experiments, an unmarked hfq deletion mutant was constructed and was fully attenuated for virulence in humans. Inactivation or deletion of hfq downregulated Flp1 and impaired the ability of H. ducreyi to form microcolonies, downregulated DsrA and rendered H. ducreyi serum susceptible, and downregulated LspB and LspA2, which allow H. ducreyi to resist phagocytosis. We propose that, in the absence of an RpoS homolog, Hfq serves as a major contributor of H. ducreyi stationary-phase and virulence gene regulation. The contribution of Hfq to stationary-phase gene regulation may have broad implications for other organisms that lack an RpoS homolog. IMPORTANCE: Pathogenic bacteria encounter a wide range of stresses in their hosts, including nutrient limitation; the ability to sense and respond to such stresses is crucial for bacterial pathogens to successfully establish an infection. Gram-negative bacteria frequently utilize the alternative sigma factor RpoS to adapt to stresses and stationary phase. However, homologs of RpoS are absent in some bacterial pathogens, including Haemophilus ducreyi, which causes chancroid and facilitates the acquisition and transmission of HIV-1. Here, we provide evidence that, in the absence of an RpoS homolog, Hfq serves as a major contributor of stationary-phase gene regulation and that Hfq is required for H. ducreyi to infect humans. To our knowledge, this is the first study describing Hfq as a major contributor of stationary-phase gene regulation in bacteria and the requirement of Hfq for the virulence of a bacterial pathogen in humans.

Draft Genome Sequence of the Clinical Isolate Acinetobacter nosocomialis Strain M2.

We report the 3.78-Mbp high-quality draft assembly of the genome from a clinical isolate of Acinetobacter nosocomialis called strain M2 (previously known as Acinetobacter baumannii strain M2).

Activation of CpxRA in Haemophilus ducreyi primarily inhibits the expression of its targets, including major virulence determinants.

Haemophilus ducreyi causes chancroid, a genital ulcer disease that facilitates the transmission of human immunodeficiency virus type 1. In humans, H. ducreyi is surrounded by phagocytes and must adapt to a hostile environment to survive. To sense and respond to environmental cues, bacteria frequently use two-component signal transduction (2CST) systems. The only obvious 2CST system in H. ducreyi is CpxRA; CpxR is a response regulator, and CpxA is a sensor kinase. Previous studies by Hansen and coworkers showed that CpxR directly represses the expression of dsrA, the lspB-lspA2 operon, and the flp operon, which are required for virulence in humans. They further showed that CpxA functions predominantly as a phosphatase in vitro to maintain the expression of virulence determinants. Since a cpxA mutant is avirulent while a cpxR mutant is fully virulent in humans, CpxA also likely functions predominantly as a phosphatase in vivo. To better understand the role of H. ducreyi CpxRA in controlling virulence determinants, here we defined genes potentially regulated by CpxRA by using RNA-Seq. Activation of CpxR by deletion of cpxA repressed nearly 70% of its targets, including seven established virulence determinants. Inactivation of CpxR by deletion of cpxR differentially regulated few genes and increased the expression of one virulence determinant. We identified a CpxR binding motif that was enriched in downregulated but not upregulated targets. These data reinforce the hypothesis that CpxA phosphatase activity plays a critical role in controlling H. ducreyi virulence in vivo. Characterization of the downregulated genes may offer new insights into pathogenesis.