Anna N. Walker

Bioaccumulation and Metabolic Effects of the Endocrine Disruptor Methoprene in the Lobster, Homarus americanus

Abstract Methoprene is a pesticide that acts as a juvenile hormone agonist. Although developed initially against insects, it has since been shown to have toxic effects on larval and adult crustaceans. Methoprene was one of several pesticides applied to the Western Long Island Sound (WLIS) watershed area during the summer of 1999; the other pesticides were malathion, resmethrin, and sumethrin. These pesticides were applied as part of a county-by-county effort to control the mosquito vector of West Nile Virus. Subsequently, the seasonal lobster catches from the WLIS have decreased dramatically. The lethality of the pesticides to lobsters had been unknown. We studied the effects of methoprene while other investigators studied effects of the other pesticides. We questioned whether methoprene, through its effects on larvae, adults or both, could have contributed to this decline. We found that low levels of methoprene had adverse effects on lobster larvae. It was toxic to stage II larvae at 1 ppb. Stage IV larvae were more resistant, but did exhibit significant increases in molt frequency beginning at exposures of 5 ppb. Juvenile lobsters exhibited variations in tissue susceptibility to methoprene: hepatopancreas appeared to be the most vulnerable, reflected by environmental concentrations of methoprene inhibiting almost all protein synthesis in this organ. Our results indicated that methoprene concentrates in the hepatopancreas, nervous tissue and epidermal cells of the adult lobster. Methoprene altered the synthesis and incorporation of chitoproteins (cuticle proteins) into adult postmolt lobster explant shells. SDS PAGE analyses of adult post–molt shell extracts revealed changes in the synthesis of chitoproteins in the methoprene-treated specimens, suggesting that methoprene affects the normal pathway of lobster cuticle synthesis and the quality of the post-molt shell. Although it is likely that a combination of factors led to the reduced lobster population in WLIS, methoprene may have contributed both by direct toxic effects and by disrupting homeostatic events under endocrine control.

Toxin-antitoxin loci vapBC-1 and vapXD contribute to survival and virulence in nontypeable Haemophilus influenzae

BackgroundNontypeable Haemophilus influenzae (NTHi) is a significant human pathogen responsible for respiratory tract infections and the most common cause of recurrent otitis media. Type II toxin-antitoxin (TA) systems are genetic elements that code for a stable protein toxin and a labile antitoxin that are thought to be involved in metabolic regulation of bacteria by enabling a switch to a dormant state under stress conditions. The contribution to infection persistence of the NTHi TA loci vapBC-1 and vapXD was examined in this study.ResultsDeletions in vapBC-1, vapXD and vapBC-1 vapXD significantly decreased the survival of NTHi co-cultured with primary human respiratory tissue at the air-liquid interface and in the chinchilla model of otitis media. The TA deletions did not affect the growth dynamics of the mutants in rich media, their ultra-structural morphology, or display appreciable synergy during NTHi infections. The toxin and antitoxin proteins of both pairs heterodimerized in vivo. Consistent with our previous findings regarding the VapC-1 toxin, the NTHi VapD toxin also displayed ribonuclease activity.ConclusionsWe conclude that the vapBC-1 and vapXD TA loci enhance NTHi survival and virulence during infection in vitro and in vivo using a mechanism of mRNA cleavage, and that these conserved TA pairs represent new targets for the prophylaxis and therapy of otitis media and other NTHi-caused mucosal diseases.

Cronkhite-Canada Syndrome: Sustained Remission After Corticosteroid Treatment

testinal metaplasia in any of our patients, in contrast to the literature that suggests intestinal metaplasia and atrophy of gastric mucosa when exposed to bile acid (1, 2). Although we could not determine the duration and degree of DGR, this may be a factor in the development of intestinal metaplasia or atrophy. The effect of UDCA on DGR gastritis and dyspeptic symptoms due to DGR is beneficial, and improvement in symptoms after UDCA therapy has been shown to be highly significant compared with placebo (5, 6). Also UDCA is safe and effective for children with DGR (7). We used triple therapy for H. pylori infection and UDCA for DGR, but the eradication of H. pylori infection and symptom relief were achieved in only two of our patients. This low eradication rate may support the synergistic action of DGR and H. pylori infection on gastric mucosa of children. Further controlled studies are needed to clarify the relationship between DGR and H. pylori infection in children.

EFFECTS OF THE PESTICIDE METHOPRENE ON MORPHOGENESIS AND SHELL FORMATION IN THE BLUE CRAB CALLINECTES SAPIDUS

The juvenile hormone analog methoprene causes both cytologic and biochemical alterations in larval and adult stages of the blue crab Callinectes sapidus. This insect growth regulator, used for mosquito control, caused (at a concentration of 10 gM) profound ultrastructural changes in the cuticular epithelial cells of postmolt adult blue crabs studied in vitro; these changes included loss of secretory organelles as well as distention and blebbing of the outer membrane of the nuclear envelope. Biochemically, 10 gM methoprene caused decreased deposition of extracellular cuticular chitin and protein, as well as a remarkable intracellular accumulation of chitoprotein precursors. These findings suggest that methoprene alters exocytosis and deposition of cuticular components. In vivo studies indicated that 5-10 gM methoprene is able to penetrate the embryonic investment coat to localize in lipovitellin. Exposure to methoprene at environmental concentrations (2-10 giM) produced morbidity and mortality in the form of an overall reduction in the number of successful hatching and lethargic behavior exhibited by the surviving zoeae. Methoprene exposure (3.3 gM) was also toxic to megalopae, delaying the molt to the first crab form and resulting in death of 80% of larvae after 10 days. The blue crab Callinectes sapidus Rathbun is a crustacean of both ecologic and economic importance to the coastal zones of the eastern and southern United States. Its life cycle and feeding habits bring it into estuarine environments subjected to pesticide treatments for mosquito control. It is logical to assume that the blue crab might be impacted by compounds active in other arthropods. The pesticide methoprene, a compound frequently applied to wetlands and salt marshes, belongs to the group of pesticides known as insect growth regulators (IGRs), which, in general, exert their toxic effects by disrupting insect development and/or reproduction. We offer in this report evidence that methoprene can penetrate the embryonic cuticle and can produce morbidity and mortality in larval forms of the blue crab. We also present in vitro findings indicating that methoprene may be toxic to the adult blue crab as well by altering the exocytosis of cuticular material.

Transmission of the parasitic dinoflagellate Hematodinium sp. infection in blue crabs Callinectes sapidus by cannibalism.

Infection with the parasitic dinoflagellate Hematodinium sp. can be devastating to blue crab Callinectes sapidus populations. Morbidity and mortality appear to depend on the burden of parasitic organisms. Heavily infected crabs become lethargic and, if not preyed upon, succumb to overwhelming infection. We report on the transmission of Hematodinium sp. into blue crabs that were fed pieces of infected tissues and examined for evidence of infection at time periods from 1 to 48 h and for the general state of their health after 4 d. During the first 16 h after feeding, Hematodinium sp. was found in the gut, followed by large increases in hemolymph hemocytes and the appearance of hemocytic nodules in tissues. By 16 h, the hemocytic nodules appeared poorly circumscribed and disorganized. No nodules were seen in a heavily infected crab after 24 h. By the end of the 48 h after feeding, 73% (11 of 15) of the crabs had shown evidence of infection with Hematodinium sp. Those crabs with infection intensities (Hematodinium sp. as percent of cells in hemolymph) higher than 20% were dead within 4 d.

Cutaneous sclerosing extramedullary hematopoietic tumor in chronic myelogenous leukemia

Background:  Extramedullary hematopoiesis is a well‐documented manifestation of chronic myeloproliferative disorders, most commonly seen in chronic idiopathic myelofibrosis (agnogenic myeloid metaplasia), but rarely in chronic myelogenous leukemia. It typically occurs in the spleen and liver, but has also been described in skin. Microscopically, foci of extramedullary hematopoiesis consist of erythroid and myeloid precursors intermixed with megakaryocytes. The megakaryocytes may elaborate fibrogenic cytokines, which induce proliferation of fibroblasts. The term ‘sclerosing extramedullary hematopoietic tumor’ has been applied to this latter entity and its resemblance to a fibrohistiocytic neoplasm has been noted.

METABOLIC EFFECTS OF ACUTE EXPOSURE TO METHOPRENE IN THE AMERICAN LOBSTER, HOMARUS AMERICANUS

Abstract Methoprene was a constituent of the pesticide cocktail applied to the Western Long Island Sound (WLIS) watershed area during the summer of 1999. Subsequently, the seasonal lobster catches from the WLIS have decreased dramatically. We have been engaged in ongoing studies of the effects of methoprene on larval, juvenile and adult lobsters. Most recently, we found that Stage IV larvae exposed to 50 ppb methoprene experience >90% mortality rate after 3 days. Bioaccumulation studies on adult lobsters showed that methoprene concentrated against the gradient of the surrounding seawater (50 ppb) in hepatopancreas (1.55 ppm), gonad (5.18 ppm), epithelial tissue (6.17 ppm) and, most significantly, the eyestalks (28.83 ppm). Exposure to methoprene altered the expression of the stress proteins and the pattern of ubiquitinylation of cytosolic proteins by Day 1 Stage I larvae and by epithelial tissue of postmolt juvenile lobsters. Postmolt juvenile animals also demonstrated an altered pattern of protein phosphorylation in their epithelial tissues following methoprene exposure, indicating that it may interfere with cell signaling pathways. Increasing concentrations of methoprene were associated with increasing chitoproteins in the microsomal fractions of Day 1 Stage I larvae, suggesting that methoprene may compromise the exocytosis of shell matrix precursors from the epithelial cells. Methoprene did not, however, alter the activity of chitin synthase in these larvae. Although it is likely that a combination of harmful events and exposures led to the reduced lobster population in WLIS, methoprene may have contributed to the decline both by direct toxic effects and by disrupting homeostatic processes.

Small-cell undifferentiated carcinoma of neuroendocrine type originating in the gallbladder

PURPOSE We report a case of small-cell undifferentiated carcinoma with neuroendocrine (SCUCN) of the gallbladder in a 67-year-old man who presented with suspected cholelithiasis. Treatment included a cholecystectomy and a 4-cycle course of etoposide and carboplatin. CONCLUSIONS Small-cell undifferentiated carcinoma with neuroendocrine features of the gallbladder is a rare disease with approximately 30 cases reported in the literature. Clinical characteristics include an association with cholelithiasis, an elderly age distribution, a female preponderance, and a correlation with cigarette smoking. It is known to behave aggressively and carry a grave prognosis, with extensive local invasion and early metastasis being characteristic. Medical and surgical therapies exist and have demonstrated best results when used in combination.

Morphologic effects of in vivo acute exposure to the pesticide methoprene on the hepatopancreas of a non-target organism, Homarus americanus.

Methoprene is a pesticide widely used for mosquito control. It is an endocrine disruptor, acting as an analog of juvenile hormone. While targeting insect larvae, it also impacts non-target animals including crustaceans. Anecdotal reports suggested that methoprene has unintended effects on adult arthropods. Earlier, we documented effects in adult lobsters at the metabolic and gene expression levels. In this study we have documented morphologic corollaries to our prior observations. We examined the light and electron microscopic changes in the hepatopancreas of adult lobsters following in vivo acute exposure to methoprene. Changes by light and electron microscopy levels were evident following exposure to sub-lethal concentrations of methoprene for 24h. Tissue from exposed animals showed the formation of extensive cytoplasmic spaces (vesiculation) with disruption and loss of specific subcellular organelles. The findings provide morphologic correlates to the metabolic and genomic alterations we have observed in previous investigations.

Synthesis of a high-density lipoprotein in the developing blue crab (Callinectes sapidus).

An important lipoprotein in the hemolymph of crustaceans is LpI. It transports lipid to peripheral tissues and also has a role in crustacean immune recognition. We employed a monoclonal antibody specific for the LpI peptide to demonstrate by ELISA, western blot and immunohistochemistry the appearance of LpI during development of Callinectes sapidus, the blue crab. LpI was first found in stage 5 embryos and appeared to be synthesized by lateral basophilic cuboidal cells that demonstrated cytoplasmic immunoreactivity for LpI at their interface with the yolk mass. The embryonic cuboidal cells bore a strong cytologic resemblance to the hepatopancreas cells of later stages (zoea, megalopae, adults), which were also immunoreactive for LpI.