One Health and antimicrobial resistance, a United States perspective.

Antimicrobial drugs are a precious resource, responsible for saving millions of lives since their discovery. Unfortunately, some antimicrobials are rapidly losing their effectiveness due to the development and spread of antimicrobial resistance (AMR), a multi-faceted and complex problem affecting humans, animals, plants and the environment. While AMR is a global problem, in this paper, the authors briefly highlight some ongoing efforts in the United States of America aimed at integrating a One Health approach into policies and programmes that address this important health threat.

Les antibiotiques sont des ressources de grande valeur qui ont sauvé des millions de vies depuis leur découverte. Malheureusement, certains agents antimicrobiens perdent rapidement leur efficacité en raison de l'apparition et propagation des résistances à ces agents, phénomène complexe et multidimensionnel qui affecte l'homme, les animaux, les plantes et l'environnement. La résistance aux agents antimicrobiens est un problème mondial ; dans cet article, les auteurs décrivent certaines initiatives actuellement mises en oeuvre aux États-Unis d'Amérique pour intégrer l'approche Une seule santé dans les politiques et les programmes conçus pour lutter contre cette menace sanitaire majeure.

Los fármacos antimicrobianos son un recurso valiosísimo, cuyo uso ha salvado millones de vidas desde que fueron descubiertos. Lamentablemente, algunos de ellos están perdiendo rápidamente eficacia debido a la aparición y propagación de resistencias, lo que plantea un problema tan complejo como poliédrico, que afecta a personas, animales, plantas y ecosistemas. Aunque la dimensión del problema es planetaria, los autores destacan aquí brevemente algunas de las iniciativas en curso en los Estados Unidos de América que tienen por objetivo integrar los planteamientos de Una sola salud en el conjunto de políticas y programas desde los cuales se aborda esta importante amenaza sanitaria.

Development of a (silent) speech recognition system for patients following laryngectomy.

Surgical voice restoration post-laryngectomy has a number of limitations and drawbacks. The present gold standard involves the use of a tracheo-oesophageal fistula (TOF) valve to divert air from the lungs into the throat, which vibrates, and from this, speech can be formed. Not all patients can use these valves and those who do are susceptible to complications associated with valve failure. Thus there is still a place for other voice restoration options. With advances in electronic miniaturization and portable computing power a computing-intensive solution has been investigated. Magnets were placed on the lips, teeth and tongue of a volunteer causing a change in the surrounding magnetic field when the individual mouthed words. These changes were detected by 6 dual axis magnetic sensors, which were incorporated into a pair of special glasses. The resulting signals were compared to training data recorded previously by means of a dynamic time warping algorithm using dynamic programming. When compared to a small vocabulary database, the patterns were found to be recognised with an accuracy of 97% for words and 94% for phonemes. On this basis we plan to develop a speech system for patients who have lost laryngeal function.

The dhfr oribeta-binding protein RIP60 contains 15 zinc fingers: DNA binding and looping by the central three fingers and an associated proline-rich region.

Initiation of DNA replication occurs with high frequency within oribeta, a short region 3' to the Chinese hamster dhfr gene. Homodimers of RIP60 (replication initiation-region protein 60 kDA) purified from nuclear extract bind two ATT-rich sites in oribeta and foster the formation of a twisted 720 bp DNA loop in vitro. Using a one hybrid screen in yeast, we have cloned the cDNA for human RIP60. RIP60 contains 15 C(2)H(2)zinc finger (ZF) DNA binding motifs organized in three clusters, termed hand Z1 (ZFs 1-5), hand Z2 (ZFs 6-8) and hand Z3 (ZFs 9-15). A proline-rich region is located between hands Z2 and Z3. Gel mobility shift and DNase I footprinting experiments show hands Z1 and Z2 independently bind the oribeta RIP60 sites specifically, but with different affinities. Hand Z3 binds DNA, but displays no specificity for RIP60 sites. Ligation enhancement, DNase I footprinting, and atomic force microscopy assays show that hand Z2 and a portion of the associated proline-rich region is sufficient for protein multimerization on DNA and DNA looping in vitro. Polyomavirus origin-dependent plasmid replication assays show RIP60 has weak replication enhancer activity, suggesting that RIP60 does not harbor a transcriptional transactivation domain. Because vertebrate origins of replication have no known consensus sequence, we suggest that sequence-specific DNA binding proteins such as RIP60 may act as accessory factors in origin identification prior to the assembly of pre-initiation complexes.

Comparison of ribosomal proteins from neoplastic and non-neoplastic cells. Resolution by two-dimensional gel electrophoresis.

Functionally active mammalian membrane-free ribosomes were prepared from rat forebrain, a solid tumor (mouse neuroblastoma C-1300) and a human tumor cell-line in suspension culture (HeLa). Proteins were extracted from these ribosomes and were resolved by a newly developed. Two dimensional gel electrophoretic procedure. Although the ribosomal protein patterns from the three preparations were generally similar, there were four obvious differences between the forebrain ribosomes and the ribosomes from the two neoplastic cell types.

Evidence for the synthesis of tubulin on membrane-bound and free ribosomes from rat forebrain.

Evidence is presented that tubulin is synthesized in cell-free systems derived from rat forebrain containing either membrane-bound or free ribosomes. Both systems were supplemented with saturating amounts of messenger RNA (from either membrane-bound of free ribosomes) and soluble enzyme and initiation factors from whole forebrain. Synthesized products were chracterized by two biochemical purification methods for brain tubulin isolation. These methods of tubulin analysis indicated that both cell free systems synthesized the same relative amounts of tubulin.

Altered pituitary growth hormone (GH) regulation in streptozotocin-diabetic rats: a combined defect of hypothalamic somatostatin and GH-releasing factor.

Diabetes mellitus in the rat is associated with loss of pulsatile GH secretion. An interplay between hypothalamic GH-releasing factor (GRF) and inhibitory factor [somatostatin (SRIF)] secretion is thought to account for episodic pituitary GH release. An increase in SRIF tone/action or a decrease in GRF release/response in diabetic rats could account for the suppressed GH levels. Pituitaries from streptozotocin-diabetic rats contained less GH than controls (15.9 +/- 2.5 vs. 29.5 +/- 4.6 micrograms/mg; P less than 0.05) despite normal somatotrope representation, as demonstrated using immunofluorescence studies. Basal GH secretion from monolayer culture of dispersed anterior pituitary (AP) cells from diabetic rats was proportionately decreased (150 +/- 10 vs. 103 +/- 10 ng/10(5) cells; P less than 0.005). GRF (10(-11)-10(-8) M)-induced release of GH from AP cells was decreased in diabetic rats (maximum response to 10(-8) M GRF, 401 +/- 60 vs. 618 +/- 41 ng/10(5) cells; P less than 0.01); however, sensitivity to GRF was unchanged (EC50, 79 +/- 41 vs. 128 +/- 67 pM). By contrast, SRIF (10(-7)-10(-10)-induced inhibition of GRF (10(-8) M)-mediated GH release was impaired in AP cells of diabetic rats compared to that in controls (IC50, 112 +/- 33 vs. 55 +/- 31 pM; P less than 0.05) associated with a decrease in AP plasma membrane SRIF receptor concentration (63.4 +/- 15.6 vs. 160.3 +/- 13.7 fmol/mg protein; P less than 0.05), with no change in affinity. These findings are consistent with chronic exposure to increased hypothalamic SRIF influence. GH synthesis has been shown to be independent of SRIF regulation; however, insulin-like growth factor-I and GRF inhibit and stimulate GH synthesis, respectively. In diabetic rats insulin-like growth factor-I levels were decreased, appropriate to low GH status, in serum (290 +/- 66 vs. 1662 +/- 92 ng/ml; P less than 0.001) and hypothalamus (6.8 +/- 1.0 vs. 13.0 +/- 0.4 pg/mg wet wt; P less than 0.001) and, thus, did not seem to account for the low AP GH content. Hypothalamic GRF content in diabetic rats (1.11 +/- 0.10 ng/hypothalamus) did not differ from that in controls (1.16 +/- 0.17 ng/hypothalamus). GRF mRNA levels, however, were reduced by 80% in diabetic rats compared to controls. Taken together these data support a combined role for decreased hypothalamic GRF and increased SRIF in mediating alterations of GH synthesis and secretion in streptozotocin-induced diabetes.

Distribution of metastases at necropsy in colorectal cancer.

A series of 372 necropsies on patients with colorectal cancer (CRC) was examined retrospectively. Patients were excluded if the primary tumour was not removed, if the operation was palliative or if death occurred within three months of operation. Only 43 cases remained after these exclusions and of these six died of intercurrent disease. The 37 remaining patients were all thought to have undergone a curative resection, following which they died with recurrent tumour. They represent the group of patients who might have benefited from adjuvant chemotherapy. The pattern of metastases and the cause of death was examined in these 37 patients. Local recurrence was found more commonly (25 cases, 68 per cent) than hepatic metastases (21 cases, 57 per cent) or metastases at other sites. This finding indicates that excision of the primary tumour must be as complete as possible and that adjuvant therapies must be directed both locally as well as systemically.

Prospective serial liver ultrasound scanning in resectable colorectal cancer treated with adjuvant razoxane.

Prospective serial ultrasound scanning (U/S) of the liver at 3-monthly intervals has been used to detect the presence of hepatic metastases from successfully resected Dukes' A, B and C grade colorectal carcinomas in a trial of the antimetastatic agent razoxane (ICRF 159). One hundred and twenty-six consecutive patients were randomly allocated to either adjuvant razoxane treatment (61) or to no further treatment (65). Twenty-six patients, 15 in the control group (23 per cent) and 11 in the razoxane treatment group (18 per cent) have developed hepatic metastases. All but three patients in the treated group and four in the controls were detected by U/S. In the control group four patients had definite metastases when metastases were first seen by ultrasound and three more, among the eight who initially had probable metastases, became definite metastases on ultrasound. In the treated group four patients had definite metastases and four more developed them amongst 12 who had probable metastases. The remaining eight of these 12 all regressed completely. This compared with only one complete regression (additionally on 5-FU for lung secondaries) amongst the eight controls who had probable metastases. The median time to development of liver metastases detected by U/S and other means was 59 weeks in the control group and 91 weeks in the razoxane treatment group. Ultrasound has proved to be a safe, sensitive and acceptable serial imaging technique for detecting colorectal liver metastases. Used prospectively, U/S has been valuable in monitoring the natural history of liver metastases and their behaviour during razoxane treatment.

Cyclopia and congenital cytomegalovirus infection.

We describe the first documented association of congenital cytomegalovirus (CMV) infection and cyclopia. A previous report has suggested that any infant with congenital ocular defects should be investigated for CMV infection [Frenkel et al, 1980]. Our case underlines this suggestion and questions a teratogenic role for CMV in cyclopiaholoprosencephaly. More documented cases may help clarify the relationship of transplacental CMV infection to the holoprosencephaly developmental field defect, including cyclopia.

In vitro studies of the biosynthesis of brain tubulin on membranes.

Membrane elements in brain tissue contain relatively large amounts of alpha- and beta-tubulin (FIGURES 2 and 3). We have investigated the subcellular sites of tubulin biosynthesis in order to determine the origin of this membrane-associated tubulin. Free and membrane-bound polysomes from rat forebrain were separated by differential centrifugation, and the products of translation from these polysome populations were analyzed by 2DGE (FIGURES 4 and 6). Alpha- and beta-tubulin subunits were synthesized by the free polysome population (FIGURES 4 and 5A and B). The membrane-bound polysome fraction synthesized a protein with similar (but not identical) characteristics to alpha-tubulin (denoted as "MB" in FIGURE 6), including isoelectric point, molecular weight, peptide map, and copurification with microtubules after aggregation-disaggregation. Tubulin subunits synthesized in vitro by free polysomes could associate posttranslationally with a microsome fraction (FIGURE 7A). The association of the tubulin translation products with membranes was not disrupted by high salt; the associated tubulin, however, was susceptible to proteolytic digestion, with the exception of one of the beta-tubulin subunits (FIGURE 7B). There was an identical protease-resistant beta-tubulin subunit among the native proteins of the smooth microsome fractions. Our data is consistent with the conclusion that at least one beta subunit of membrane-associated tubulin is synthesized by free polysomes and becomes posttranslationally added to membrane structures. It is unlikely that a cotranslational mechanism is responsible, in which there is a signal-mediated insertion of a growing polypeptide chain to membrane. Our results, however, are consistent with a "membrane trigger" mechanism proposed by Wickner in which the membrane lipid bilayer triggers the folding of a polypeptide into a configuration that allows integral membrane insertion. The association of tubulin with membranes may also be secondary to the interaction of hydrophobic elements. The amino acid sequence of beta tubulin is known to contain several hydrophobic domains. Tubulin can be incorporated into phospholipid vesicles and various subcellular membrane elements. In our studies, in vitro synthesized tubulin from free polysome was found to be purified by hydrophobic affinity chromatography with ethane-sepharose (FIGURE 8). Thus, the hydrophobic characteristics of newly synthesized tubulin could be partially responsible for the posttranslational association of tubulin subunit with membranes. Native tubulin in a soluble fraction of CNS tissue was not purified by hydrophobic affinity chromatography.(ABSTRACT TRUNCATED AT 400 WORDS)

Properties and function of brain carbonic anhydrase.

This chapter has described the characterization and biogenesis of soluble and membrane-bound CA in the central nervous system. The two forms of the enzyme appear to be quite similar in their molecular characteristics, however the data strongly indicate that they are synthesized on separate polysomal populations; the membrane-bound form resulting from synthesis on the RER. Our preliminary data suggest that the partitioning of mRNA for CA on the different polysomes results from the interaction of partial nascent chains with a specific receptor on the RER. We feel a function of membrane-associated synthesis is for the targeting of CA to sites in the cell where there are enzymes that can rapidly utilize the protons and bicarbonate produced by CA catalytic activity for ion exchange reactions. We have also presented arguments that CA may function as a bicarbonate source in the control of metabolism specifically in the acceleration of fatty acid synthesis in the oligodendrocyte.

Adjuvant oral razoxane (ICRF-159) in resectable colorectal cancer.

One hundred and seventy six patients (81 controls, 95 receiving treatment) have entered a prospective randomized trial of long-term oral adjuvant razoxane (ICRF-159) following removal of a colorectal cancer. The median follow-up is 34 months. The treated patients in Dukes' groups B and C have a significantly longer disease-free interval than the control patients (P = 0.01 'as randomized' and P = 0.004 'as treated'). The differences in survival for Dukes' groups B and C are not significant, although follow-up is short. In Dukes' groups B and C, however, 24 of 56 of the patients in the control group have died (43%), as against only 17 of 64 in the treatment group (27%). The treatment produces very few side-effects, is well tolerated by patients, and is taken orally.

Sununit structure of synaptosomal tubulin.

The subunit structure of rat brain synaptosomal tubulin was examined by high resolution two-dimensional gel fractionation. Whole brain cytoplasmic tubulin consists of two groups of alpha subunits (alpha1 and alpha2), and a minimum of two beta subunits (beta1 and beta2). Both alpha subunits consist of a major relatively acidic form and minor relatively basic forms. In contrast, tubulin purified from synaptoplasm contains an additional subunit, alpha3, which has the same isoelectric point but slightly faster electrophoretic mobility than alpha1 and alpha2. All synaptosomal alpha subunits are the relatively acidic forms and the minor basic forms are absent. The synaptosomal beta subunits have electrophoretic properties similar to the corresponding cytoplasmic forms. The alpha3 synaptosomal tubulin subunit has affinity for colchicine, has a tryptic peptide map similar to whole brain cytoplasmic alpha tubulin, and can be purified by a standard tubulin purification method.

The subunit composition of cerebellar tubulin: evidence for multiple beta tubulin messenger RNAs.

Cytoplasmic proteins were isolated from adult rat forebrain and cerebellum and analyzed by two-dimensional gel electrophoresis under conditions which the major subunits of tubulin were separated. Forebrain cytoplasmic tubulin consisted of two groups of alpha subunits (alpha 1 and alpha 2) and a minimum of two beta subunits (beta 1 and beta 2). However, the rat cerebellar cytoplasmic proteins contained greatly decreased amounts of the beta 1 tubulin subunit relative to the analysis of forebrain proteins. Messenger RNA (mRNA) was purified from cerebellum and forebrain and translated in wheat germ homogenate. Analysis of the translation products of cerebellar mRNA indicated only a trace amount of the beta 1 subunit, whereas the expected amount of beta 1 was found among the translation products of forebrain mRNA. This data is consistent with the conclusion that the beta 1 and beta 2 subunits of tubulin are synthesized from different mRNAs. A decrease in beta 1 mRNA relative to other tubulin mRNAs may be one of the factors responsible for the low steady state amounts of beta 1 tubulin in the cerebellum.

A controlled prospective trial of adjuvant razoxane in resectable colorectal cancer.

Following resection of their tumour, 162 patients with colorectal cancer entered a prospective randomized controlled clinical trial of adjuvant oral razoxane. Thirty-one patients were Duke's group A; 49 group B; 61 group C; and 17 group D; an additional four patients were randomized in error. The adjuvant group received the usual clinical care and 125 mg razoxane twice daily for 5 consecutive days (monday-friday) every week indefinitely. Control patients received the same clinical care as the adjuvant group, but no razoxane. At 3 years, 134 patients (84%) are evaluable. The recurrence rate in the first 6 months was 20% and 28% respectively in the Duke's B and C controls compared with 4% and 9% in the corresponding razoxane treated patients. Most recurrences occurred within the first 6 months from randomization. When all patients as randomized are included in the analysis of survival, irrespective of whether they were cured by surgery (Duke's A), had advanced cancer (Duke's D), or took no razoxane when randomized to take it, then as might be expected any differences there may be between the razoxane-treated and control patients with minimal residual disease (Duke's B and C) are so distorted that the p value of the difference in survival was 0.93. If however only patients with Duke's group B or C are taken (49 controls and 47 treated), log-rank analysis reveals a difference in the cancer mortality curves (p = 0.07). If patients who had been randomized to take razoxane, but who had not taken it at any time (and therefore received the same treatment as controls) are analysed with the controls, the difference between the two groups increases further, with p less than 0.05. The razoxane-treated patients experienced no significant toxicity apart from a readily reversible mild leukopenia in 52% while gastrointestinal symptoms necessitated stopping the drug in only four patients. These four all took the drug for less than 4 weeks. Because there was no toxicity to subtract from any benefit razoxane adjuvant treatment produced and the quality of life was not impaired, the therapeutic benefit of surgery wsa increased to the extent that razoxane increased survival of patients with Duke's B and C tumours.

Serum unconjugated bile acids: qualitative and quantitative profiles in ileal resection and bacterial overgrowth.

Qualitative and quantitative profiles of unconjugated bile acids in the serum obtained over a 24-h period from three patients with ileal resections and one with a bacterial overgrowth are described. Unconjugated serum bile acids were determined using the high sensitivity and resolution of capillary column gas liquid chromatography after their rapid extraction and isolation using reverse phase octadecylsilane bonded silica cartridges and the lipophilic gel Lipidex 1000. Unconjugated serum bile acid concentrations were elevated throughout the day in both ileum resected patients and in conditions involving bacterial overgrowth when compared to healthy subjects. Total conjugated cholic acid concentrations were expectedly low in both intestinal disorders and were without the postprandial increases generally observed in healthy subjects. Qualitative gas chromatographic profiles of serum unconjugated bile acids in bacterial overgrowth distinctly revealed a predominance of deoxycholic acid and other secondary bile acids in all samples, while, in conditions of an impaired enterohepatic circulation, deoxycholic acid was absent or present in only trace amounts. The potential significance of measuring serum unconjugated bile acids in intestinal disorders is discussed.

Avoiding inappropriate surgery for secondary rectal cancer.

AIMS: Secondary rectal carcinoma occurs by invasion of the rectum by local primaries or by metastatic spread from a distant primary. The principle management of primary rectal carcinoma is surgery, but this is not usually the case for secondary carcinoma. This study investigates how these two may be differentiated and inappropriate surgery, in particular inappropriate abdominoperineal excision, can be avoided. METHODS: There were six patients with secondary carcinoma of the rectum. The primary tumours were: prostate (three), endometrium (two), breast (one). RESULTS: All the patients presented with lower gastrointestinal symptoms and four had a palpable mass on rectal examination. The diagnosis was made on histology and immunohistochemistry. Treatment was with endocrine therapy, chemotherapy, radiotherapy, and surgery. Three patients had palliative surgical procedures, and one had a curative anterior resection. The median survival was 7.5 months. CONCLUSIONS: This study has found that the presentation of primary and secondary rectal carcinoma is similar, and the method for distinguishing between the two is histology and immunohistochemistry. Staining for prostate-specific antigen was 100% accurate in the diagnosis of secondary rectal carcinoma arising from a prostate primary. The treatment of secondary rectal carcinoma is with systemic therapy and surgery is usually palliative, and therefore abdominoperineal excision should be avoided.

The aetiology and surgery of carcinoma of the anus, rectum and sigmoid colon in Crohn's disease. Negative correlation with human papillomavirus type 16 (HPV 16).

Anal and recto-sigmoid Crohn's disease may rarely be complicated by the development of local malignancy. Eight patients with this association were identified from the records of St. Mark's Hospital between 1947 and 1988 and two from The London Hospital. The aetiology of the malignancies and their surgical management have been examined. Eight patients had adenocarcinoma of the anus, rectum or sigmoid colon and two had squamous cell carcinoma of the anus. Synchronous dysplasia, adenomas and carcinomas were found in four of the eight patients with adenocarcinoma and a progression to malignancy which is analogous to that in ulcerative colitis is proposed for these cases via a 'dysplasia-carcinoma' or 'adenoma-carcinoma' sequence. In the four other patients with adenocarcinoma, the tumour arose within an area of Crohn's disease or in association with a chronic Crohn's fistula. In these four cases no dysplasia was found in the specimens at that time and long-standing infection is the only aetiological factor identified. Evidence for infection with human papillomavirus (HPV 16) was sought by DNA hybridisation of archival material but none was found in material from the Crohn's disease, fistulae or adeno- or squamous cell carcinomas. Anal and rectosigmoid Crohn's disease disguised the presence of malignancy, and diagnosis was usually delayed. All patients underwent excisional surgery and eight of the ten had all of the large bowel removed either as a single procedure or in stages. Only three of the eight cases of adenocarcinoma had early malignancies (Dukes' stage A) and the remaining five cases had locally advanced disease (Dukes' stage B).

Detailed faecal bile acid profile: a diagnostic test for colorectal cancer?

Detailed profiles of bile acids in faeces were evaluated as a diagnostic test for colorectal cancer in rats. Twenty-seven bile acid peaks were measured using improved methods of extraction and separation followed by the sensitive and specific techniques of capillary column gas liquid chromatography and mass spectrometry. Colorectal cancer was induced in experimental animals (female Sprague-Dawley rats, n = 20) by subcutaneous injection of dimethylhydrazine (DMH) and faecal unconjugated bile acids compared with those in the control group (n = 20). The amount of total faecal unconjugated bile acids was lower in the animals administered DMH (255 mg/day vs 334 mg/day: (P = 0.04), and the excretion of seven individual bile acids was reduced when compared with those in the control group (P less than 0.01). In order to use the faecal bile acid profiles as a diagnostic test, linear discriminant analysis was performed. A discriminant score was derived which was applied to each profile, to determine to which group (control or DMH) each animal belonged retrospectively. All analyses were performed blind, and 90% of the animals were correctly assigned. In man, as in rats, the bile acid profile of faces is equally complex and the bile acid profile may be useful as a diagnostic test.

Invasive carcinoma in juvenile polyposis--report of a case in an African patient.

Since the 1980s it has been recognized that the polyps of juvenile polyposis have malignant potential. Since the introduction of colonoscopy this syndrome has been redefined and the hamartoma-adenoma-carcinoma sequence recognized. We believe this case is the first report of an invasive carcinoma associated with juvenile polyposis to be described in an African patient.