Cholinergic septohippocampal projections from the medial septal area to the hippocampus are proposed to have important roles in cognition by modulating properties of the hippocampal network. However, the precise spatial and temporal profile of acetylcholine release in the hippocampus remains unclear making it difficult to define specific roles for cholinergic transmission in hippocampal dependent behaviors. This is partly due to a lack of tools enabling specific intervention in, and recording of, cholinergic transmission. Here, we review the organization of septohippocampal cholinergic projections and hippocampal acetylcholine receptors as well as the role of cholinergic transmission in modulating cellular excitability, synaptic plasticity, and rhythmic network oscillations. We point to a number of open questions that remain unanswered and discuss the potential for recently developed techniques to provide a radical reappraisal of the function of cholinergic inputs to the hippocampus. Neuromodulation is a feature of complex nervous systems that is proposed to play an essential role in behavior allowing anatomically defined neural circuitry to be re-purposed, thereby enabling rapid adaptation in response to external stimuli Katz, . NeuromoThe neuromodulator acetylcholine has been extensively studied due to its prominent role in attention, learning, and synaptic plasticity  or depression (LTD)  and nicotinic (nAChRs) acetylcholine receptors, causing the subsequent modulation of cellular excitability and synaptic transmission. These two types of receptors are differentially expressed across the hippocampus Table  and fulfNicotinic acetylcholine receptors are ionotropic pentameric receptors made up of heteromeric or homomeric assemblies of α2−α10 and β2−β4 subunits. In the hippocampus, the nAChR subtypes predominantly expressed are α7, α4β2, and α3β4  or Gi/o proteins (M2 and M4). Differences in G-protein coupling preferences lie within an amino acid sequence divergence in the third intracellular loop between the M1/M3/M5 sequences compared with the M2/M4 sequences  cause inhibition of voltage-gated Ca2+ channels, a decrease in cAMP-mediated signaling and inhibition of neurotransmitter release at cholinergic, GABAergic and glutamatergic terminals  potentiate NMDA currents , causing membrane depolarization and increasing input resistance    restricts neurotransmission to classic synapses or junctional complexes. Following axon terminal depolarization, neurotransmitter release and binding to nicotinic and muscarinic acetylcholine receptors, acetylcholine is rapidly hydrolyzed by AChE to yield choline and acetate. Choline is then transported back into the terminal by high-affinity choline transporters to be resynthesized into acetylcholine by the enzyme choline acetyltransferase (ChAT).Conversely, volume transmission is characterized by multiple extracellular pathways through which signals can diffuse in a three-dimensional fashion and activate extrasynaptic receptors. In this case, presynaptic cholinergic terminals do not make synaptic contacts and therefore do not face a defined postsynaptic density  are particularly prominent in the hippocampus. Two types of hippocampal theta oscillations have been defined—type 1 (7–12 Hz) is associated with voluntary movement and exploratory behavior, whereas type 2 (4–6 Hz) is present during immobility, in particular REM sleep, and occurs spontaneously during urethane anesthesia  cholinergic, fast- and burst-firing (~10–18 Hz) GABAergic and fast- and cluster-firing (~8–14 Hz) glutamatergic neurons  channels facilitates the induction of LTP (Markram and Segal, 2 receptors (Shimoshige et al., 1 receptor-dependent inhibition of voltage-activated Kv7 potassium channels (Tsubokawa and Ross, 1 receptors and NMDA receptors regulate dendritic voltage-gated Kv4.2 potassium channels (Losonczy et al., 2 receptors coupled with an increase in pyramidal cell excitability by α7 receptors both facilitate the induction of LTP (Ji et al., As well as directly causing short- and long-term synaptic plasticity, cholinergic receptor activation modulates the induction of synaptic plasticity (Shimoshige et al., In vitro and in vivo studies have supported the idea that theta and gamma oscillations provide a mechanism for bringing together in time glutamatergic inputs to pyramidal cell dendrites and dendritic invasion of fast spikes, the key elements for the induction of synaptic plasticity. Furthermore, inhibition from different classes of interneurons, creating gamma oscillations within each theta cycle and the modulated efficacy of excitatory inputs at different theta phases can selectively influence the timing of pyramidal cell firing (Lengyel et al., Questions:Most studies examining the role of acetylcholine in synaptic transmission and plasticity use exogenous agonist application. Which forms of synaptic transmission and plasticity modulation are engaged by physiological acetylcholine release? What mechanisms are important? Is the physiological timing of cholinergic input important for modulating synaptic plasticity in the hippocampus?Currently, there is considerable focus on dissecting the neuronal substrates of behavior by linking specific cell types and populations to their activity patterns and extending this knowledge to probe the neuronal substrates of behavior. This approach is proving critical for understanding how neuronal circuits contribute to nervous system function.Although much is known about the effects of acetylcholine in the hippocampus, it is difficult to predict, from the detail of individual receptors and synaptic pathways, what the overall effect will be. To understand this we need better tools to measure cholinergic neuron firing rates and the consequent release of acetylcholine, in addition to specific interventions to disrupt or activate cholinergic input to the hippocampus. The diffuse nature of this cholinergic pathway makes the isolation and experimental stimulation/silencing of well-defined groups of cholinergic fibers difficult to achieve. Recently developed tools could circumvent many of these technical problems and lead to new insights into the role of acetylcholine in modulating network activity, synaptic plasticity and behavioral output.In this context, bioengineering and genetic tools, combined with transgenic animals, have become a powerful resource for the anatomical and functional deconstruction of neuronal networks. Cell type-specific expression of protein markers and light-gated ion channels allow the structural dynamics and electrical activity of genetically defined neurons to be manipulated and analysed on the millisecond timescale (Luo et al., in vivo patterns of acetylcholine release will be critical for identifying and dissecting the physiological effects of cholinergic neuromodulation in the hippocampus. Two complementary approaches could reveal the temporal specificity of cholinergic neuron firing and subsequent acetylcholine release. The first is accurate measurement of the release kinetics and extracellular concentrations of acetylcholine/choline both in vivo and in vitro at high temporal resolution. This has been performed in discrete areas of the cortex during different behavioral states but information is not yet available for the hippocampus (Parikh et al., Although optogenetic tools will allow more precise simulation of acetylcholine release, the patterns of release will not be optimal unless there is a better understanding of the physiological patterns of cholinergic cell firing and fluctuations in acetylcholine concentrations in the extracellular space. The ability to mimic The combination of these new approaches may provoke a fundamental rethink of the functions of cholinergic inputs to the hippocampus and reveal how specific receptors mediate those functions, therefore presenting an opportunity to establish causal connections between the activity of the septohippocampal cholinergic system, hippocampal network function, learning and memory.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Porcine torque teno sus virus (TTSuV) is a small icosahedral and non-enveloped virus which contains a single-stranded (ssDNA), circular and negative DNA genome and infects mainly vertebrates and is currently classified into the 'floating' genus Anellovirus of Circoviridae with two species. Viral DNA of both porcine TTSuV species has a high prevalence in both healthy and diseased pigs worldwide and multiple infections of TTSuV with distinct genotypes or subtypes of the same species has been documented in the United States, Europe and Asia. However, there exists no information about histopathological lesions caused by infection with porcine TTSuV2.7genomic copies viral loads of porcine TTSuV2 that had positive result for torque teno sus virus type 2 and negative result for torque teno sus virus type 1 and porcine pseudorabies virus type 2 were used to inoculate specific pathogen-free piglets by intramuscular route and humanely killed at 3,7,10,14,17,21 and 24 days post inoculation (dpi), the control pigs were injected intramuscularly with 1 ml of sterile DMEM and humanely killed the end of the study for histopathological examination routinely processed, respectively.Porcine liver tissue homogenate with 1 ml of 6.91 × 10All porcine TTSuV2 inoculated piglets were clinic asymptomatic but developed myocardial fibroklasts and endocardium, interstitial pneumonia, membranous glomerular nephropathy, and modest inflammatory cells infiltration in portal areas in the liver, foci of hemorrhage in some pancreas islet, a tiny amount red blood cells in venule of muscularis mucosae and outer longitudinal muscle, rarely red blood cells in the microvasculation and infiltration of inflammatory cells (lymphocytes and eosinophils) of tonsil and hilar lymph nodes, infiltration of inflammatory lymphocytes and necrosis or degeneration and focal gliosis of lymphocytes in the paracortical zone after inoculation with porcine TTSuV2-containing tissue homogenate.Analysis of these presentations revealed that porcine TTSuV2 was readily transmitted to TTSuV-negative swine and that infection was associated with characteristic pathologic changes in specific pathogen-free piglets inoculated with porcine TTSuV2. Those results indicated no markedly histopathological changes happened in those parenchymatous organs, especially the digestive system and immune system when the specific pathogen-free pigs were infected with porcine TTSuV2, hence, to some extent, it was not remarkable pathological agent for domestic pigs at least. So, porcine TTSuV2 could be an unrecognized pathogenic viral infectious etiology of swine. This study indicated a directly related description of lesions responsible for TTSuV2 infection in swine. In 1997, torque teno sus virus (TTSuV) was discovered in Japan in a patient with post-transfusion hepatitis of unknown aetiology -8. In 20Recently, TTSuVs have attracted markedly interest within the research community  and porcTo date, much attention has been paid to TTSuV infection in other vertebrates ,27,28, eNo tissue culture system for TTSuVs propagation has been identified . AlthougControl animals displayed no clinical manifestations throughout the study. No obvious gross lesions were observed between the control group and the infected animals on necropsy examination. In totally, specific microscopical lesions were found in the tissues from the twenty-one pigs infection with porcine torque teno sus virus 2 throughout the study, and there existed subtle pathologic changes in the tissues containing myocardial fibroklasts and endocardium, interstitial pneumonia, membranous glomerular nephropathy, and modest inflammatory cells infiltration in portal areas in the liver, foci of hemorrhage in some pancreas islet, a tiny amount red blood cells in venule of muscularis mucosae and outer longitudinal muscle, rarely red blood cells in the microvasculation and infiltration of inflammatory cells (lymphocytes and eosinophils) of tonsil and hilar lymph nodes, infiltration of inflammatory lymphocytes and necrosis or degeneration and focal gliosis of lymphocytes in the paracortical zone after inoculation with the porcine TTSuV2-containing tissue homogenate; these changes were not detected in uninoculated control pigs or pigs injected with tissue homogenate devoid of porcine TTSuV2 genome. Firstly, hyperemia and congestion were found in the myocardial fibroklasts and endocardium seen in Figure The lung lesions contained of a mild interstitial pneumonia characterized by a slight thickening of the alveolar septa by mononuclear cells and congestion, and bronchial epithelial cells happened defluvium and inflammation infiltration in pulmonary alveolus seen in Figure Figure Figure The micrographic pathological changes of the immune system showed in Figure In the experimental study, porcine TTSuV2 genome were identified in the serum of a conventional pig used as the source material of porcine TTSuV2 for inoculation into gnotobiotic porcine. TTSuV has been elucidated that it was readily transmitted to young gnotobiotic swine that had negative results for TTSuV genome , serial The present study evaluated the histopathological lesions of inoculation infection of the porcine Circoviridae family DNA virus torque tero sus virus (TTSuV) in main parenchmatous organs containing heart, lung, kidney, digestive system and immune system tonsil, spleen and lymph nodes on account of the viral tropism to lymphoid cells ,49. The To date, it has been found that it is common for an individual to coinfect with different genogroups of TTSuV  and the Viral persistence and sporadic infection may be a risk factor for dissemination of PCV2 or TTSuV to negative sows, potentially resulting in reproductive failures. This irregular pattern of infection also paid attention to the fact that boars which tested negative on the first collection may be positive in subsequent ejaculates. A periodic monitoring for PCV2 or TTSuV2 must be established in boar community. PCV2 and TTSuV2 presence in semen samples of younger animals may indicate a recent infection or may even be related to management or measures of stress. Morphology and sperm motility analysis did not indicate significant diversities between PCV2 positive and negative boars. All semen samples tested presented motility superior to 80%, which could allow their use for processing, dilution and artificial insemination. No manifestation of reproductive failure was associated with this co-infection. These findings raised the problem of the importance of these viral infections in the pathology of reproductive failures . AlthougMoreover, in both species, genotype 2 is more prevalent than genotype 1. In domestic pig, TTSuV2 infections have been shown to be more common in pigs affected by postweaning multisystemic wasting syndrome (PMWS), a porcine circovirus type 2 (PCV2) disease, than non-PMWS affected pigs ,49. Due Up to now, to our knowledge, the only available method for detection of TTSuV is focused on nucleic acid detection, such as nested polymerase chain reaction (nPCR) , quantitAnalysis of these presentations revealed that porcine TTSuV2 was readily transmitted to TTSuV-negative swine and that infection was associated with characteristic pathologic changes in specific pathogen-free piglets inoculated with porcine TTSuV2. Those results indicated no markedly histopathological changes happened in those parenchymatous organs, especially the digestive system and immune system when the specific pathogen-free pigs were infected with porcine TTSuV2, hence, to some extent, it was not remarkable pathological agent for domestic pigs at least. So, porcine TTSuV2 could be an unrecognized pathogenic viral infectious etiology of swine. This study indicated a directly related description of lesions responsible for TTSuV2 infection in swine.Date-mated specific pathogen-free pregnant primiparous sows fetched in Canada were purchased from a new constructed pigfarm located in Chongqing and transported to National Animal Experimental Teaching Demonstration Center of Sichuan Agricultural University for caesarian section of specific pathogen-free piglets litters using methods described elsewhere . Twenty-7genomic copies/ml) is determined by SYBR green-based real-time quantitative PCR.Porcine torque teno sus virus 2 was derived from 20% (w/v) liver tissue homogenate which was extracted twice with chloroform so as to remove infectivity of any extraneous enveloped viruses  and was 7genomic copies viral loads of porcine TTSuV2 liver tissue homogenate and humanely killed at 3,7,10,14,17,21 and 24 days post inoculation (dpi). The three remaining pigs, which were used as control pigs, were injected intramuscularly with 1 ml of sterile Dulbecco's modified Eagle's medium , supplemented with 10% (v/v) fetal calf serum , 100 IU/ml of streptomycin and penicillin, respectively, and humanely killed the end of the study.Twenty-four 28-day-old specific pathogen-free piglets were randomly divided two groups, housed separately in the experimental animal houses at National Animal Experimental Teaching Demonstration Center of Sichuan Agricultural University, twenty-one piglets used as experimental group which were inoculated by intramuscular route with 1 ml of 6.91 × 10All pigs were subjected to gross necropsy examination and main tissues and organs samples for microscopical examination were collected from each pig at the same time of necropsy. These samples contained portions of heart, liver, spleen, lung, kidney, hilar lymph nodes, mesenteric lymph nodes, inguinal lymph nodes, pancreas, tonsil and duodenum for histopathological examination. All tissues were fixed in 10% neutral buffered formalin for 24-48 hours and routinely processed and embedded in paraffin wax. Sections were cut into slices with 4 ~ 5 micrometer thick and stained with hematoxylin and eosin (HE). Those slices were detected under a light microscope for conventional morphological evaluation.The authors declare that they have no competing interests.MM, LZ carried out most of the experiments and drafted the manuscript. ZWX and WZG critically revised the manuscript and the experimental design. YW, XP, LZ, YFW and SL helped with the study. All of the authors read and approved the final version of the manuscript.


Exposure of mothers to chronic stressors during pregnancy or the postpartum period often leads to the development of depression, anxiety, or other related mood disorders. The adverse effects of mood disorders are often mediated through maternal behavior and recent work has identified arginine vasopressin (AVP) as a key neuropeptide hormone in the expression of maternal behavior in both rats and humans. Using an established rodent model that elicits behavioral and physiological responses similar to human mood disorders, this study tested the effectiveness of chronic AVP infusion as a novel treatment for the adverse effects of exposure to chronic social stress during lactation in rats. During early (day 3) and mid (day 10) lactation, AVP treatment significantly decreased the latency to initiate nursing and time spent retrieving pups, and increased pup grooming and total maternal care (sum of pup grooming and nursing). AVP treatment was also effective in decreasing maternal aggression and the average duration of aggressive bouts on day 3 of lactation. Central AVP may be an effective target for the development of treatments for enhancing maternal behavior in individuals exposed to chronic social stress. Depression, chronic anxiety, and other mood disorders can have negative consequences on mothers as well as their offspring, and the occurrence of these disorders during lactation has been associated with impaired child growth and development ,2. WhileWhile numerous animal models of stress-induced depression and anxiety disrupt maternal behavior, most involve significant physiological components and may not replicate the effects of chronic psychosocial stressors in humans. In rodents for example, chronic restraint stress or exposure to wet bedding decreases maternal aggression  and incrThe various roles of neuropeptide hormones in the regulation and expression of maternal behavior in both rats and humans have been studied for decades, and several have been linked to the etiology of stress related mood disorders ,25. ArgiThe objective of this investigation was to test the effectiveness of AVP as a novel treatment for stress induced disorders that affect maternal behavior. It was postulated that chronic central AVP infusion would alter maternal behavior and growth patterns in dams and their pups exposed to the CSS rodent model for postpartum depression and anxiety. More specifically, it was hypothesized that AVP treatment would decrease aggression towards a novel male intruder, increase maternal care towards offspring, and prevent attenuated growth in both the dam and pups.p-values < 0.02, repeated measures ANOVA). However, the primary focus of the current study is to investigate the effects of AVP on maternal behavior; therefore, the effects of lactation day (time) will not be discussed in detail. During maternal care testing on day 10, the time required to retrieve all 8 pups was decreased in AVP dams . There were no differences between the saline and AVP groups in any measure of behavior during the maternal care tests on lactation days 3 or 17 . However, the primary focus of the current study is to investigate the effects of AVP on maternal behavior; therefore, the effects of lactation day (time) will not be discussed in detail. During the maternal aggression tests on lactation day 3, latency to initiate nursing was decreased in AVP dams . Locomotor activity duration was decreased in the AVP group on day 3 . There were no differences in behavior between the saline and AVP groups during maternal aggression tests on lactation days 10 or 17 , day 10  or day 17 . Similarly there were no differences in mean body weight between the pups of saline and AVP treated dams on day 3 , day 10  or day 17 . Absolute weight gain (in grams) and percent weight gain (relative to day 3) were similar for both dams and pups of saline and AVP groups on day 10, but not day 17 , the percent weight increase was larger for saline than AVP pups . The exposure of saline control dams to a novel male intruder resulted in behavioral and growth responses consistent with stressed dams in the other studies ,22, thusTo date, AVP research has focused primarily on male-typical social behaviors, including aggression and pair-bond formation, with less attention devoted to the effects of neuropeptides on maternal social behavior . The obsAVP treatment significantly affected maternal care behaviors during maternal care testing  and when dams were faced with an acute stressor during maternal aggression testing . During maternal care testing on day 10 of lactation, AVP dams significantly decreased the amount of time spent retrieving pups back to the nest  and increased pup grooming. Similarly, during the maternal aggression testing on lactation day 3, AVP treated dams initiated nursing more quickly (decreased nursing latency) than saline dams, and AVP dams exhibited a significant increase in total maternal care behavior. These results are consistent with previous work investigating the neuroendocrine and behavioral parameters of rats selectively bred for high anxiety-related behavior (HAB). While it has been shown that the responsiveness of the hypothalamo-pituitary-adrenal (HPA) axis is attenuated during the natural progression of pregnancy and lactation ,65, HAB The significant effects of chronic icv AVP on maternal aggression behavior support the hypothesis that AVP administration decreases maternal aggression towards a novel male intruder. While maternal aggression is typically highest during early lactation ,45,46, iThe absence of AVP effects on any behavioral measure during maternal care testing on lactation day 3 may suggest that AVP has no acute effect on maternal care. However, the positive effects of AVP on both maternal care and aggression during the maternal aggression tests on day 3 suggest otherwise. The consistent effects of AVP on maternal care both during typical dam-pup interactions as well as during the exposure to a social stressor indicate that the treatment was effective after 24 h (day 3) and one week (day 10). It is likely that the lack of effect on maternal care during aggression testing on day 10 was due to the fact that the maternal care test immediately preceded the maternal aggression test. The lack of significant treatment effects on lactation day 17 may be due to a down-regulation of central endogenous AVP production in response to the continuous infusion, resulting in a dampened effect of the exogenous treatment. Conversely, the lack of significant effects during late lactation may be due to a ceiling effect , where AThe hypothesis that AVP treatment would increase growth in dams and/or pups following CSS exposure was not supported by the growth profiles. There was no difference in body weight or weight gain between control and AVP animals on either day 3 or 10 of lactation. Interestingly, while there was no difference in absolute weight gain between pups of saline and AVP treated dams on day 17, the percent weight increase from day 3 to day 17 was less in AVP litters compared with controls. Similar effects are seen in humans, where infants of mothers suffering from postnatal depression suffered poorer growth than infants of non-depressed mothers , presumaad libitum throughout the study. Rats were mated by placing 2 females with 1 male for 8 days. Following mating, females were housed communally (3 per cage) until one day prior to parturition when each female was housed individually. Normal maternal behavior  was observed in all animals on lactation day 1, litters were culled to 8 pups , and dams were randomly assigned to saline control, low dose AVP, or high dose AVP groups. On lactation day 2, rats were anesthetized with isoflurane and implanted with chronic guide cannulae directed into the right lateral ventricle (placements were confirmed with India ink). ALZET  osmotic pumps  were primed and connected to the chronic guide cannulae and implanted subcutaneously in the upper back region. The pumps contained one of two doses of AVP , or saline vehicle. While the initial group sizes for control, low dose AVP and high dose AVP were 16, 12 and 14, respectively, intruder males attacked and killed the pups of 4 dams during the study. As a result, these dams were removed from subsequent maternal behavior testing . Animals in this study were maintained in accordance with the guidelines of the Committee of the Care and Use of Laboratory Animals Resources, National Research Council, and the research protocol was approved by the Tufts University Institutional Animal Care and Use Committee. Female Sprague-Dawley rats (175–200 g) were obtained from Charles River Laboratories  and maintained in temperature (21–25 °C) and light  controlled rooms. Food (Purina rat chow) and water were provided vs. non-stressed dams), but rather to test the effectiveness of AVP vs. saline control in ameliorating the negative behavioral effects of chronic stress.The CSS protocol used in the current study was closely based on the 2011 study by Nephew and Bridges . On lacti.e., maternal care test). The latency to retrieve all pups back to the nest, the latency to initiate nursing, as well as the frequencies and durations of pup retrieval, pup grooming, self grooming, nursing, total maternal care (sum of pup grooming and nursing), nesting and general locomotor activity were all scored. Immediately following the maternal care test, a novel intruder male was introduced to the home cage (with both dam and pups present as per the CSS paradigm described above) and maternal aggression was recorded for a subsequent 30 min . During this maternal aggression test, the latency to initiate nursing, the latency to initiate aggression, as well as the frequencies and durations of pup retrieval, pup grooming, self grooming, nursing, nesting, attacking , biting, kicking with hindlimbs, pinning of intruder to the cage floor, and general locomotor activity were scored. In addition, total maternal care, total aggression , and mean aggressive bout duration  were also calculated. All behavior analyses were conducted using ODlog behavioral analysis software (Macropod Inc.) by an observer that was blind to the treatment of each dam. The ODlog software records continuous data in 5 s bins and generates frequency and duration summaries for all behavior measures over the 30-min observation periods. Maternal behavior, which consists of maternal care of the pups and maternal aggression towards a novel intruder, was assessed in all dams during two 30-min tests on days 3, 10 and 17 of lactation. Behavior testing was conducted at randomly selected times between 9:00 a.m. and 12:00 p.m. to avoid habituation effects on behavior. A digital video camera (Panasonic PV-GS180) was used to record maternal behaviors without human interference. Prior to maternal behavior testing, the pups were removed from the home cage for 30 min. The pups were then re-introduced and maternal care was recorded for 30 min  were also compared using one-tailed t-tests. All results are presented as the mean ± SEM, and the level of statistical significance was p ≤ 0.05.Initial statistical testing of behavioral data was conducted using 2-way repeated measures ANOVAs with lactation day  as the repeated factor (VassarStats). Given that both maternal care and aggression have been previously shown to change over the course of lactation ,46,48, tIn summary, the data support the hypotheses that chronic infusion of AVP promotes maternal care and suppresses maternal aggression during exposure to social stress during early and mid lactation. While the current results from maternal females are not consistent with most male studies, they are consistent with several studies of AVP and maternal behavior that served as the basis for the present study. The most significant implication of the gender differences in the effects of central AVP is in the development of novel treatments for depression and anxiety. AVP antagonism (V1a and V1b receptors) is an active area of preclinical and clinical research on depression and anxiety disorders , but rec


To the Editor: Since the beginning of May 2011, a large outbreak of infections associated with Shiga toxin (Stx)–producing Escherichia coli (STEC) O104:H4 has occurred in Germany , another group of diarrheagenic strains found frequently in developing countries  from a case-patient with HUS in 2009 in Italy had the same combination of virulence factors as the strain from Germany: Stx2 production and enteroaggregative adhesion genetic markers. This strain, which had not been typed when it was isolated, showed positive PCR results for O104  on August 5, 2009, after 5 days of bloody diarrhea, vomiting, and abdominal pain. Diagnosis of HUS was based on the presence of hemolytic anemia, thrombocytopenia, and anuria. Neurologic symptoms  occurred during hospitalization; magnetic resonance imaging showed signal abnormalities in the lenticular nuclei.Because of severe cardiac impairment with ejection fraction reduction and troponin increase, inotropic support and mechanical ventilation were temporarily needed. After improvement of clinical conditions, the patient was discharged, but she was readmitted a few days later because of headache, vomiting, confusion, dysarthria, hypertension, and visual impairment. Ischemic lesions were found by magnetic resonance imaging at fundus oculi. Neurologic status improved the next day, but the visual deficit persisted. Hemodialysis was needed for 2 months. Long-term sequelae of the disease were stage IV chronic kidney disease, hypertension, and severe visual impairment.Informed consent and an epidemiologic interview were obtained from the patient’s parents. The household, including her mother and 2 siblings (4 and 5 years of age), had traveled for 1 week to a resort in Tunisia; they had returned 3 weeks before the onset of the prodromal symptoms of HUS. Four days after their return, the youngest sister was hospitalized for 3 days because of bloody diarrhea, but no laboratory diagnosis was established. The mother reported having had watery diarrhea and abdominal pain on August 2. The patient history did not show any other usual risk factor for STEC infection, such as consumption of unpasteurized milk or dairy products, undercooked meat, or raw sprouts or direct exposure to ruminants or their manure. This finding suggests that the infection was probably acquired through person-to-person transmission.,This case report confirms that strains of STEC O104 strictly related to the epidemic strain in Germany had already caused sporadic infections in Europe (,The history of this patient supports the hypothesis that ruminants would not have had a specific role in the transmission of STEC O104:H4, as already suggested by the epidemiologic features of the recent outbreaks (,The clinical course of our patient closely resembles those of persons who had HUS associated with the German outbreak (


The conserved brain design that primates inherited from early mammals differs from the variable adult brain size and species-specific brain dominances observed across mammals. This variability relies on the emergence of specialized cerebral cortical regions and sub-compartments, triggering an increase in brain size, areal interconnectivity and histological complexity that ultimately lies on the activation of developmental programs. Structural placental features are not well correlated with brain enlargement; however, several endocrine pathways could be tuned with the activation of neuronal progenitors in the proliferative neocortical compartments. In this article, we reviewed some mechanisms of eutherians maternal–fetal unit interactions associated with brain development and evolution. We propose a hypothesis of brain evolution where proliferative compartments in primates become activated by “non-classical” endocrine placental signals participating in different steps of corticogenesis. Changes in the inner placental structure, along with placenta endocrine stimuli over the cortical proliferative activity would allow mammalian brain enlargement with a concomitant shorter gestation span, as an evolutionary strategy to escape from parent-offspring conflict. During fication . In addiThe mammalian lineage arises from synapsids, a mammal-like reptile ancestor, diverged from other tetrapods about 300 million years ago (Mya) in the Carboniferous . The basal diversification of eutherians has been historically considered an unstable node . Using 18 homologous genes segments, Afrotheria was originally positioned as the most basal mammalian clade, with Xenarthra as the second, and Euarchontoglires and Laurasiatheria as the third branches of the mammalian tree . Using genomic sequences, the phylogenetic relationship of mammals has been re-informed . A recent publication renewed this debate resolving this basal relation as a split between Xenarthra and Epitheria .Three mammalian subclasses became extinct in different evolutionary moments. Triconodonta, the earliest lineage diverged in mammalian phylogeny, disappeared at the end of the Cretaceous period (extended from ~145 to ~66 Mya) leading to a lack of basal-related in living mammals. The second was the Multituberculata, and the latest extinguished was the infraclass of Theria called Palaeoryctoides. A recent analysis suggests that the first modern placental orders (eutherians) emerged around 2–3 million years later than the Cretaceous–Paleogene (K–Pg) extinction event occurred 66 Mya . It was ble node  and the informed  and “coninformed , positioUnderstanding the organization of the phylogenetic tree of mammals allows the visualization of the evolutionary history of different traits and the definition of the presumed ancestral structure of the placenta . The acqFigure 2). This extensive maternal–fetal communication also implies some disadvantages like a major risk of maternal bleeding after delivery and a greater chance of fetal cells transfer to the maternal system , variations in the interhemal barrier mainly characterized by different degrees of hypertrophy of maternal endothelium and presence of cytotrophoblast and/or syncytial trophoblast are commonly observed . Three ml system .Figure 2) is necessary for fetal brain growth. However, this relation is not supported once the analyzes are refined . Accordingly, invasiveness of the placenta is not a requirement to develop large brains , or the addition of preimplantation stages lacking placenta formation would hamper these comparisons between marsupial and eutherian mammals . RecentlSvet1, Cux2, and Tbr2 genes. In the SVZ, cells undergo one to three more cell divisions and then migrate to build up the superficial layers of the neocortex. Neurons generated in successively later moments are incorporated into progressively more superficial layers, generating the inside-out neurogenetic gradient that is characteristic of the neocortex.Most neurons in the neocortex derive from multipotent neural stem cells in the proliferative epithelium of the VZ lining the ventricular surface of the telencephalic wall. In the VZ, radial glial cells will generate lower- and upper-layer neurons according to distinct fate potentials , the CuxFigure 3). We believe that several traits of cortical neurodevelopment would account for the high mean EQ of Euarchontoglires and especially of large-brained primates  of the neocortex depending mainly on the proliferation of cortical intermediate progenitors  and gliaprimates . Primateprimates , and an primates . Togetheprimates .In some way, placenta resembles the function of several endocrine systems; thus it is positioned as the main endocrine organ throughout intrauterine development. Placenta produces and releases a number of signaling substances including cytokines, neuropeptides, neurosteroids, and amines . Some ofFigure 4). Instead, the exogenous source of 5-HT produced by the placenta is required to maintain normal levels of forebrain 5-HT during early stages of brain development  pathway. This neuroactive factor has been associated with proliferative activity, migration, and differentiation processes during neocortical development . Originaelopment  and woulelopment . For an elopment . Moreoveelopment , insteadelopment . 5-HT stelopment . Intereselopment  and cocaelopment , both inelopment .During pregnancy in mammals, placenta produces a large amount of steroids, which are crucial for the survival, development, and health of the developing embryo. Progesterone and estrogens  are the main hormones during pregnancy. Progesterone is fundamental to modulate the maternal immune response allowing the maternal tolerance of the fetal “semi-allograft.” On the other hand, estrogens are needed to promote the placental growth and angiogenesis and influence fetal growth and metabolism. These steroids are in extremely high concentrations in maternal circulation. However, placental tissue can metabolize them to inactive forms in order to avoid fetal exposure. Many of these metabolites can act as neurosteroids in adult brain, however, their contribution to fetal brain development remains under debate. For example, allopregnanolone, a progesterone metabolite modulates the activity of GABAergic and glutamatergic neurons in fetal brain and may mitigate the brain injury provoked by asphyxia in the hippocampal region, since allopregnanolone can control the proliferation and apoptotic patterns in cerebellum and hippocampus , 2009.Similar to sex steroids, glucocorticoids levels are lower in fetal than in maternal circulation. This difference is attributable to the high expression of 11β hydroxysteroid dehydrogenase type 2 (11β-HSD2) in both the placenta and fetus. In the placenta, 11β-HSD2 catalyzes the rapid inactivation of cortisol and corticosterone to inert 11 keto-products, and then it serves as a “glucocorticoid barrier,” modulating the transfer of glucocorticoids to the fetus. In the placenta, 11β-HSD2 is highly expressed in the syncytiotrophoblast of humans and in the labyrinthine zone of rodents . InteresIn general, normal concentrations of glucocorticoids are essential in the development of many organs, including central nervous system. However, some conditions associated with elevated levels of glucocorticoids, as stress or reduced capacity of placenta to metabolize it, can leads to detrimental effects on brain development and long-term behavioral effects, since glucocorticoid receptors are highly expressed in some brain areas like hippocampus . PrimateFigure 5A) and developing neocortical brain compartments . At this point of our on-going studies, we become interested in the published list of human placental differentially expressed genes . The top one gene module is enriched in signal molecules .High-throughput analysis of active transcripts is a powerful tool to explore possible molecular interactions across organs and species. Brain and placenta share some remarkable transcriptional features as the expression of imprinted genes and the expression of transcriptional pathways of immune interactions. Transcriptomes from placenta , a network analysis database focused on protein interactions .We compared these human placental signal-enriched transcripts, and those transcripts from human developing cortex  is expressed predominantly in the developing cortex participating in dendrite branching and synapses formation (CSPG4 proteoglycan (chondroitin sulfate proteoglycan 4) and with the transmembrane protein SDC4 (syndecan 4) placenta–brain coexpressed genes. Interestingly, transcriptomic analyzes have suggested that cell adhesion and cell–extracellular matrix interactions promote the proliferation and self-renewal of neural progenitors in the developing human neocortex , expressed in the developing cortex, encodes for galectin, a known regulator of neural stem cell proliferation , CSH2 (chorionic somatomammotropin hormone 2), GH2 (growth hormone 2), and VPREB1  placental genes. PTK7 (protein tyrosine kinase 7) expressed in the developing cortex, interacts with the peptide hormone CCK (cholecystokinin) from placenta and encodes a transmembrane receptor in the brain that controls a variety of developmental and physiological processes, including cell polarity, cell migration, invasion, and antagonize Wnt signaling  and CCK could be related to the fetal brain development in humans. Moreover, according to comparative placental transcriptomic analysis , these nGABBR1 [gamma-aminobutyric acid (GABA) B receptor 1] is expressed in the developing cortex and interacts with other GABA receptor in the placenta . GABBR1 has been associated with anxiety (EDN (endothelin 3) is an endothelium-derived vasoactive peptide expressed in the placenta, involved in a variety of biological functions and interacting with EDNRA (endothelin receptor type A), which is expressed in the developing cortex and upregulated after hypoxic preconditioning in the immature brain . During pregnancy in some species,including human, CG is secreted by the trophoblast and maintains the progesterone secretion from the corpus luteum at the beginning of gestation. In humans, the production of CG declines around 6–8 weeks of pregnancy. Interestingly, receptors for luteinizing hormone (LH)/human chorionic gonadotropin (hCG) have been identified in multiple areas of the brain, including the cortex. Also, rat brain expresses these receptors and neurotropic effects of LH and hCG have been demonstrated in fetal rat brain . Thus, the active interaction between the placenta and the proliferative cortical compartments would amplify the number of neural progenitors as a strategy to increase the total number of neurons in the mature brain. If this proposal is correct, it not only allows to obtain a bigger brain, but also it would permit to reduce the length of the pregnancy required to generate bigger brains, supporting a developmental strategy to escape from fetal-maternal parental conflict. Interestingly, such interactions would conduce to fetal programming changes representing plausible adaptive advantages to environmental demands before and after birth, or maladaptation when there is a mismatch between the programming and the environment requirements  would be associated with developmental long distance interactions through molecular signals displayed by the placenta. This hypothesis is complementary to those relied on the intrinsic control of neocortical development. In addition to the currently described 5-HT developmental interactions between the placenta and the cerebral cortical proliferative compartments, our transcriptomic analysis indicates new candidates for promoting neocortical expansion (e.g., irements . For exairements . Severalirements . We are The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Despite some problems related to accuracy and applicability of malaria rapid diagnostic tests (RDTs), they are currently the best option in areas with limited laboratory services for improving case management through parasitological diagnosis and reducing over-treatment. This study was conducted in areas with declining malaria burden to assess; 1) the accuracy of RDTs when used at different community settings, 2) the impact of using RDTs on anti-malarial dispensing by community-owned resource persons (CORPs) and 3) adherence of CORPs to treatment guidelines by providing treatment based on RDT results.Data were obtained from: 1) a longitudinal study of passive case detection of fevers using CORPs in six villages in Korogwe; and 2) cross-sectional surveys (CSS) in six villages of Korogwe and Muheza districts, north-eastern, Tanzania. Performance of RDTs was compared with microscopy as a gold standard, and factors affecting their accuracy were explored using a multivariate logistic regression model.2 = 367.7, p < 0.001), while the specificity was significantly higher  when compared to the longitudinal study. As determinants of sensitivity of RDTs in both studies, parasite density of < 200 asexual parasites/μl was significantly associated with high risk of false negative RDTs , while the risk of false negative test was significantly lower among cases with fever (axillary temperature ≥37.5°C) . The risk of false positive RDT (as a determinant of specificity) was significantly higher in cases with fever compared to afebrile cases . Using RDTs reduced anti-malarials dispensing from 98.9% to 32.1% in cases aged ≥5 years.Overall sensitivity and specificity of RDTs in the longitudinal study  were 88.6% and 88.2%, respectively. In the CSS, the sensitivity was significantly lower (63.4%; χAlthough RDTs had low sensitivity and specificity, which varied widely depending on fever and parasite density, using RDTs reduced over-treatment with anti-malarials significantly. Thus, with declining malaria prevalence, RDTs will potentially identify majority of febrile cases with parasites and lead to improved management of malaria and non-malaria fevers. Prompt and correct diagnosis is important for effective management of malaria cases in order to reduce morbidity and mortality caused by delayed or poor management of patients particularly among under-fives and pregnant women. Malaria diagnosis has for a long time, and particularly at community level, depended on clinical diagnosis (based on patients' history and symptoms), which has low specificity leading to over-diagnosis and over-treatment . Thus, rMicroscopic examination of Giemsa-stained blood smears confirms the presence of malaria parasites and remains the gold standard for malaria diagnosis despite its technical challenges and demand for trained personnel. However, most peripheral health facilities in malaria endemic countries lack the capacity to carry out parasitological diagnosis of malaria by microscopy -6. In TaPlasmodium falciparum histidine rich protein 2 (PfHRP-2), Plasmodium lactate dehydrogenase (pLDH) or aldolase  were neither treated with ALu nor referred to the nearest health facility by CORPs. Using a cut-off of parasite density of 5,000 asexual parasites/μl, only 36(0.2%) cases [including 5(0.1%) under-fives and 31(0.2%) aged ≥5 years] were not treated with anti-malarial drugs and thus put at risk of developing a severe disease in case they did not return to CORPs or go to find treatment elsewhere.The WHO has recently recommended that treatment with anti-malarial drugs should be given to patients with malaria parasites confirmed by laboratory tests or RDTs in areas lacking the capacity for detection of malaria parasite by microscopy . The Tanet al. [et al (unpublished data). The low sensitivity observed in the CSS might mainly be due to large number of cases with low level parasitaemia, which was below the detection limits of RDTs giving rise to high rate of false negative results. The low sensitivity of RDTs observed in the current studies was similar to what has been reported in other field studies conducted in South-eastern Tanzania [The findings of this study showed that both RDT brands (Paracheck and ParaHIT) had low sensitivity but relatively high specificity depending on fever status, parasite density, malaria parasite prevalence and transmission intensity. The overall sensitivity of RDTs was significantly higher in the longitudinal study compared to CSS and decreased with declining malaria parasite prevalence in both studies corresponding to the declining burden of malaria in the study areas as recently shown by Mmbando et al.  and IsheTanzania ,32 and oTanzania -35. HoweTanzania ,36 and aTanzania and UgandTanzania -39, and Tanzania ,40,41).Despite the differences in parasite density in the two studies  which increased with decreasing parasite prevalence, the sensitivity of RDTs was higher in febrile cases, and this was similar to what has been reported in studies conducted elsewhere in Africa ,37,42. IDespite significantly higher specificity in the CSS compared to the longitudinal study , the specificity observed in both studies was higher than what has been recently reported with similar RDTs (based on HRP-2) in Malawi . False pBefore introduction of RDTs, almost all cases attended by CORPs were treated with SP based on presenting symptoms leading to a significant level of over-treatment, since only 20.1% of all cases treated with SP during this period had malaria parasites confirmed by microscopy. However, this approach was cost-effective due to low cost of SP and was also safe since most of the febrile cases (regardless of parasite infection status) were promptly treated with anti-malarials thus reducing the risks associated with severe malaria, which often occurs when a patient is left without treatment. Despite the observed low accuracy of RDTs, which was lower than the sensitivity and specificity of ≥95% recommended by WHO , deploymAdherence to treatment guidelines by CORPs as stipulated in the SOPs was high (> 95%) in cases of all age groups. The level of adherence to treatment guidelines shown in this study was higher than what was reported by previous studies conducted in Zanzibar  and BurkThe current study utilized two brands of HRP-2 based RDTs (Paracheck and ParaHIT) which showed variable results regarding their accuracy, whereby both brands had similar sensitivity in the longitudinal study while Paracheck showed higher sensitivity in the CSS. In both studies, ParaHIT had higher specificity compared to Paracheck. Although these differences could be a true reflection of their performance under field conditions in an area where malaria has remarkably declined , this stin-vivo efficacy study of anti-malarials and in-vitro monitoring of drug resistance (which were stopped due to lack of enough samples). Unlike in health facilities and community studies where febrile patients are commonly attended and RDTs are intended to detect patients with parasites, the tests cannot be similarly used in CSS due to low number of patients with fever, low parasite density and therefore low sensitivity as shown by the findings of the CSS. However, these findings provide important information which will potentially guide future applicability of RDTs when fully introduced in many endemic countries for malaria diagnosis in the health facilities and communities; emphasizing that the tests have more relevance for case management than for disease surveillance.Furthermore, the performance of RDTs was compared to expert microscopy whose quality cannot be attained under health facility settings as recently shown by studies conducted in Tanzania  and otheet al. personal communication). Based on the setting of the longitudinal study where medical services were made available in the community through CORPs, cases not given anti-malarials had an opportunity to get treatment whenever they felt unwell, thus making this strategy safe and rational for testing the implementation of malaria case management based on RDT results.In the longitudinal study, some cases with malaria parasites (confirmed by microscopy) were neither treated with anti-malarials  nor given a referral to be attended at nearby health facilities as stipulated in the guidelines given to CORPs. While it would intuitively be important to know what happened to such cases, these issues are beyond the scope of this study since only passive case detection of fever episodes was performed, and follow-up of cases after treatment or those with negative RDT results who were not given anti-malarials was not done. However, other studies conducted in northern Tanzania showed that only 1% of children with negative RDT results developed parasitaemia during the 28 days of follow-up and only 1 out of 816 children developed a severe malaria condition demanding hospitalisation (Reyburn This study showed that RDTs used in the two community studies had varying accuracy (low sensitivity but relatively higher specificity) indicating that diagnosis of malaria using these HRP-2 based RDTs particularly in this era of declining burden of malaria remains a problem. Patients presenting in the health facilities from these communities for diagnosis of malaria are likely to have low parasite density which the current tests may fail to detect due to low sensitivity. High specificity of the RDTs indicates that most of the patients without malaria were correctly detected. However, introduction of RDTs in the longitudinal study reduced the number of cases without malaria who were treated with antimalarial drugs during the pre-RDTs as compared to the post-RDT period. Therefore, deployment of RDTs, coupled with supportive supervision can potentially reduce over-treatment and provide an opportunity for improved malaria diagnosis, and proper management of both malaria and non-malaria fevers. However, continued search and eventually introducing other alternative and sensitive malaria diagnostic methods should be explored.The authors declare that they have no competing interests.DSI, JPA, MA, TGT, IBC and MML conceived of and designed the study, DSI and PM conducted the field work and supervised the laboratory analyses. JPA and MML supervised field and laboratory work in Korogwe and Muheza districts. DSI, FF and BPM participated in data management and analysis, and DSI wrote the initial draft of the manuscript. All authors contributed during writing, read and approved the manuscript.


Approximately 90% of non-medullary thyroid malignancies originate from the follicular cell and are classified as papillary or follicular (well-differentiated) thyroid carcinomas, showing an overall favourable prognosis. However, recurrence or persistence of the disease occurs in some cases associated with the presence of loco-regional or distant metastatic lesions that generally become resistant to radioiodine therapy, while glucose uptake and metabolism are increased. Recent advances in the field of tumor progression have shown that CTC (circulating tumour cells) are metabolic and genetically heterogeneous. There is now special interest in unravelling the mechanisms that allow the reminiscence of dormant tumour lesions that might be related to late disease progression and increased risk of recurrence. AMPK (AMP-activated protein kinase) is activated by the depletion in cellular energy levels and allows adaptive changes in cell metabolism that are fundamental for cell survival in a stressful environment; nevertheless, the activation of this kinase also decreases cell proliferation rate and induces tumour cell apoptosis. In the thyroid field, AMPK emerged as a novel important intracellular pathway, since it regulates both iodide and glucose uptakes in normal thyroid cells. Furthermore, it has recently been demonstrated that the AMPK pathway is highly activated in papillary thyroid carcinomas, although the clinical significance of these findings remains elusive. Herein we review the current knowledge about the role of AMPK activation in thyroid physiology and pathophysiology, with special focus on thyroid cancer. Recurrence or persistence of differentiated thyroid cancer can be associated with resistance to radioiodine therapy, which is associated with higher glucose uptake and metabolism by the remnant lesions. The possible role of AMPK activation in this phenomenon is discussed. Palpable and non-palpable thyroid nodules are common endocrine tumours that can be detected in as much as 70% of a population if sensitive image-assisted examination methods are used . IndepenAlthough nodular lesions are frequent in the overall population, only 5–10% of thyroid nodules correspond to malignant lesions, and thyroid carcinoma is a rare disease that accounts for just 1% of all human cancers, although it is the most common endocrine malignancy . ApproxiFour types of genetic alterations comprise the majority of known mutations in DTC: BRAF and RAS (renin–angiotensin system) point mutations, and RET/PTC rearrangements for papillary thyroid cancers, and PAX8/PPARγ rearrangement in the follicular subtype . The inc131I-radioiodide and 18F-FDG-PET (18F-fluoro-deoxiglucose positron emission tomography) are very important criteria for assessment of recurrent or persistent disease. Indeed, the increase of 18F-FDG uptake associated with a decrease in iodide uptake indicates poor prognostics [+/I−-symporter), which is a thyrocyte differentiation marker that is lost in the course of thyroid carcinogenesis [The adequate intervention for DTC vary depending on the disease stage, rather than the different causative mutations. In the past 10 years, risk stratification systems were reported in order to try to better establish the adequate intervention for each patient ,14. In tgnostics . These iogenesis ,17. On tAlthough, iodide uptake through NIS is involved in both diagnostic and treatment of thyroid cancer, the mechanism underlying NIS expression and subcellular localization in thyroid cells has not been completely elucidated. Also, the molecular mechanisms involved in the metabolic shift that leads to higher glucose uptake by tumour cells are poorly defined. Recently, we described that the energy sensor AMPK (AMP-activated protein kinase) plays an important physiological role in the thyroid gland by regulating both iodide and glucose uptakes ,19. In fIn this review, we focused on the current knowledge about NIS regulation by AMPK in thyroid cells and the possible involvement of AMPK signalling pathway in thyroid cancer cell biology.de novo lipid biosynthesis. In turn, manolyl-CoA is a potent inhibitor of carnitine palmitoyl transferase-1, responsible for the translocation of long-chain fat acid into mitochondrial matrix. The reduction of malonyl-CoA induced by AMPK activation is believed to favour fat acid translocation into the mitochondria and β-oxidation to increase ATP production [AMPK is a metabolic stress-sensing cytosolic enzyme, composed of an α-catalytic and two regulatory (β and γ) subunits . Stresseoduction –25.18F-FDG-PET is accompanied by decreased radioiodide uptake ability in DTC [Our group was the first to describe the expression and function of AMPK in the thyroid gland, and its ability to regulate iodide and glucose uptakes in thyroid cells ,19. Indey in DTC ,15. Howey in DTC . Thus, tMore recently, Abdulrahman et al.  confirmeThe evidence that the tumour suppressor LKB1 is the major upstream kinase responsible for AMP activation linked for the first time AMPK activity and cancer development . Since LSeveral studies demonstrated that activated AMPK causes cell-cycle arrest associated with stabilization of p53 and decreased protein and lipid synthesis and ribosomal RNA ,28. IndeAltogether, these previously described AMPK effects are mainly anti-proliferative and anti-tumourigenic. However, different studies demonstrated that AMPK activation mediated by the reduction of ATP/AMP ratio promotes cellular survival under stressful metabolic conditions that are characteristic of tumour microenvironment ,25,31. IRecently, the participation of ROS (reactive oxygen species)/AMPK activation as fundamental for the Warburg effect in cancer cells has been confirmed . In factRecent advances in the field of tumour progression have shown that CTC (circulating tumour cells) are metabolic and genetically heterogeneous . Based oThe strikingly findings that type 2 diabetic patients have increased incidence of thyroid nodules and thyroid volume  are undeSubsequent reports described that metformin, a drug used for the management of type 2 diabetes, might present antineoplastic effects in various human cancers ,41. TakiStudies on various cancer cell lines and animal models demonstrated that metformin inhibits tumour growth via the activation of AMPK and the inhibition of mTOR/S6K (S6 kinase) signalling pathway ,45,46. IThe AMPK pathway has not been extensively studied in thyroid cancer and until recently the expression and function of AMPK in this type of cancer was not evaluated so far. In a recent report, AMPK expression was first described in the human thyroid gland and both its expression and activity were shown to be strongly up-regulated in PTC . Using aUsing thyroid cancer cell line expressing either wild-type BRAF or V600E-mutant BRAF, Choi et al.  observedThe mTOR pathway is also up-regulated in DTC , and mTOPtenthy−/− mice had a strong decreased in AMPK activity (AMPK phophorylation on Thr172), mainly through PI3K and PKA-mediated AMPK phosphorylation at Ser485. In order to reactivated AMPK, they treated the mutants with AICAR for 4 weeks. They observed a restoration in the expression of TCA enzymes and a reversion in the glycolytic switch induced by constitutive PI3K activation. Also, AMPK reactivation led to a slower growth rate in mutant glands compared with untreated mutants, which was confirmed by a drastically reduced thyroid BrdUrd (bromodeoxyuridine) incorporation in AICAR treated, when compared with that of untreated mice. These findings are in agreement with the tumour suppressor role of AMPK, but are not fully concordant with the finding of increased AMPK signalling pathway that was detected in human thyroid cancer samples.Recently, Antico Arciuch et al.  describeMetformin treatment inhibited growth and induced apoptosis in anaplastic thyroid cancer cell lines . FurtherThe new findings of higher AMPK activation in human papillary thyroid cancer in relation to benign lesions and the possibility that this pathway modulate cell growth, apoptosis and survival raises several questions that need to be answered in order to better define whether AMPK could be a novel target in thyroid cancer patients. The differentiated papillary thyroid cancer is indolent and most of these tumours do not present an aggressive behaviour. It is intriguing that this same tumour subtype seems to express high levels of both mTOR  and AMPKAMPK functions as a tumour suppressor gene in several cancer subtypes, and accordingly its activation in tumour cells lead to cell death. As shown, AMPK activation in normal cells or its loss of function in tumours can also be the molecular pathway implicated in the higher glucose uptake that occurs during the tumour metabolic shift known as the Warburg effect. Thus, it is of great importance to better understand the pleiotropic actions of the different AMPK isoforms, their levels of activation and their perspectives in cancer cell biology. Since AMPK pathway is overexpressed in primary differentiated tumours of the human thyroid, it is now intriguing to know what are the consequences for tumour cells of its further activation or rather silencing. Also, the study of AMPK pathway in advanced and the undifferentiated anaplastic thyroid cancer can bring further insight into the role of AMPK in the different stages of cancer progression.


In recent years an increasing number of neurodegenerative diseases has been linked to the misfolding of a specific protein and its subsequent accumulation into aggregated species, often toxic to the cell. Of all the factors that affect the behavior of these proteins, disulfide bonds are likely to be important, being very conserved in protein sequences and being the enzymes devoted to their formation among the most conserved machineries in mammals. Their crucial role in the folding and in the function of a big fraction of the human proteome is well established. The role of disulfide bonding in preventing and managing protein misfolding and aggregation is currently under investigation. New insights into their involvement in neurodegenerative diseases, their effect on the process of protein misfolding and aggregation, and into the role of the cellular machineries devoted to disulfide bond formation in neurodegenerative diseases are emerging. These studies mark a step forward in the comprehension of the biological base of neurodegenerative disorders and highlight the numerous questions that still remain open. Neurodegenerative misfolding diseases (NMD) are a group of diseases involving the misfolding of one or two proteins and their accumulation into aggregated species toxic to neurons, leading to a wide range of neurological symptoms , whose role has been revealed as central in many neurodegenerative disorders, being upregulated in many NMD and in some cases interacting directly with misfolded and aggregated proteins [The fate of a protein depends on two major factors, its sequence and its cellular environment. From the sequence perspective, many studies have identified several features of the amino acid ructures , 3. Afteructures . From thproteins , 6. Disulfide bonds are present in 15% of the human proteome and they are enriched in secreted proteins (65%), due to the need of greater protein stability in the absence of the accurate quality control systems present inside the cell. Interestingly they are present in 55% of the proteins involved in pathologic amyloid formation , suggestThis distribution of disulfide bonds presence in proteins involved in misfolding diseases  shows thThe questions about protein aggregation, disulfide bonds, and neurotoxicity are numerous, and the proposed answers are often controversial. Here we focus on two simple aspects of disulfide bonds and neurodegenerative diseases: the role of disulfide bonds in the stabilization of proteins involved in aggregation and neurodegeneration and the involvement of PDIs in neurodegenerative misfolding diseases. β-propensity, hydrophobicity, and net charge [Proteins have evolved several structural and sequence-based strategies to avoid misfolding and potentially toxic aggregation such as stabilizing protein folding, controlling aggregation by gatekeepers residues, limiting t charge .β-lactoglobulin, and lysozyme) [Many studies on the effects of disulfide bonds on protein misfolding and aggregation revealed that (i) some disulfide bonds are positioned to prevent the population of aggregation prone conformation of some proteins  , 7, 10, ysozyme) , (iii) cysozyme) , and  to a highly β-sheet, protease-resistant, scrapie conformation (PrPSc). The only PrP disulfide bond (Cys179–Cys214) stabilizes the overall fold of the protein by connecting the two long C-terminal helices [C to PrPSc conversion in vivo, and whether intermolecular disulfide bonds play a role in stabilizing PrPSc aggregates [Prion-related disorders are neurodegenerative diseases characterized by the conversion of the prion protein PrP from its normal cellular structure  is the most common degenerative disease of the motor neuron system. The cause of ALS is unknown although 5–10% of cases are familial with a Mendelian pattern of inheritance . About 2Intermolecular disulfide bonds have been identified in SOD1 aggregates in animal models of familial ALS , 25. ExpDespite the importance of disulfide bonds in SOD1 and ALS, it is not known how the intramolecular disulfide bond is formed in the reducing environment of the cytosol and how the thiol-disulfide status of SOD1 changes in the course of the disease.in vitro, there are lines of evidence that intramolecular disulfide bonds retard Tau aggregation in vitro [Tauopathies are a group of neurodegenerative diseases, including AD, Pick disease, and corticobasal degeneration, characterized by the accumulation of insoluble Tau fibrils. Tau exists as six different isoforms that result from alternative splicing of mRNA , and thein vitro . The prein vitro .β, where disulfide bonds have been engineered to allow the homogeneous population and stabilization of specific aggregated species  and the subsequent characterization of their biochemical, structural, and biological properties [In many cases a disulfide bond has been added to proteins that misfold and aggregate in neurodegenerative disorders. The most studied case is Aoperties . In eukaryotic organisms enzyme-catalyzed disulfide bonds are formed in specialized cell compartments, such as the endoplasmic reticulum (ER) and the intermembrane space of the mitochondria .2O2 [The ER not only provides an environment suitable for disulfide formation, being very oxidizing with a low GSH : GSSH ratio, but it contains also many proteins dedicated to protein folding and correct disulfide bonding. Already during their translocation into the ER lumen, proteins are oxidized by protein disulfide isomerases (PDIs). The PDIs are maintained in an oxidized state by the flavoprotein Ero1, which transfers electrons directly onto oxygen generating H2O2 . PDI was the first enzyme characterized to catalyze thiol-disulfide exchange reactions in the ER, but in recent years the number of PDI family members has been growing very quickly . In humaPDIs are a class of proteins activated upon ER stress . Since eCreutzfeldt-Jacob (CJD) disease was the first human brain disease shown to be associated with an upregulation of PDIs, both by a proteomic study of sporadic CJD brain tissue  and by tMoreover, PDI can physically interact with misfolded prion , 39, andin vitro [Many studies report a link between PDI and the pathogenesis of ALS. PDI and ERp57 were identified as the two main proteins upregulated in the spinal cord of a symptomatic ALS transgenic mice , and in in vitro .Besides physically interacting with SOD1 , moreoveDespite its demonstrated protective role, increased levels of PDI in ALS were not beneficial in patients. As previously shown for Parkinson's and Alzheimer's brain tissues , posttraα-synuclein transgenic mice [in vivo, however, is still not clear.Increased expression of PDIs family members has been observed in many studies on PD. PDI and ERp57 were found to be upregulated in two gene profile studies of PD cell culture models , 53. PDInic mice . The funα-synuclein in cell-free in vitro systems [PDI decreases the aggregation of the Parkinson's disease-associated synphilin-1 protein in neuroblastoma cells . It also systems .β [β peptide.PDI levels are increased in the brain of AD patients, where PDI colocalizes with neurofibrillary tangles in cells containing neurofibrillary tangles , 58. ERpβ , suggestProtein sequences have used different strategies to evolve and reach their functional folding in an efficient way, minimizing the risks of misfolding and (toxic) aggregation . DisulfiIntramolecular disulfide bonds stabilize the monomeric folding of prion, SOD1, and Tau retarding their aggregation. Intermolecular disulfide bonds, instead, have a more controversial role in pathogenesis. In many cases their presence is still under debate, although their importance in determining the biophysical and biological features of the formed aggregates and as a possible target for treatment.the upregulation of PDIs in animal models of NMD and in postmortem tissues from NMD patients can be part of the general UPR response to the ER stress driven by extensive protein misfolding in the cell; the colocalization of PDIs with protein aggregates suggests a chaperone activity of PDIs aimed at limiting protein aggregation itself. If this chaperone activity acts in stabilizing monomers or also in disrupting aggregates has not been addressed so far;the physical interaction of PDIs with the monomeric forms of several aggregating proteins indicates a role in modulation of the aggregation of specific disease-associated proteins by a direct association.Different potential roles of PDIs in neurodegenerative diseases are suggested by experimental findings: α1-antitrypsin [PDIs appear as a cellular strategy to avoid protein aggregation, but several questions remain unanswered. Which are the mechanisms of PDIs actions in affecting protein aggregation? And what about their effect on protein aggregates? Are different PDIs members specific for different substrates? Interestingly, some observations indeed indicate that some disease and protein specificity of PDI protection is likely to exist. PDI action, in fact, is not specific for amyloid-like aggregates: PDI overexpression inhibits the formation of Ig aggregation in a model of Russell Bodies formation  but doesitrypsin .Given the urgent need for novel therapies to treat neurodegenerative misfolding diseases, a solid understanding of disulfide bonding role in protein misfolding and aggregation could lead to the identification of new potential therapeutic targets.


The accumulation of Na+ in soil through irrigation for sustainable agricultural crop production, particularly in arid land, and by changes in environmental and climate conditions leads to the buildup of toxic level of salts in the soil. Since the latter half of the twentieth century, extensive molecular research has identified several classes of Na+ transporters that play major roles in the alleviation of ionic stress by excluding toxic Na+ from the cytosol or preventing Na+ transport to the photosynthetic organs, and also in osmotic stress by modulating intra/extracellular osmotic balance. In this review, we summarize the current knowledge of three major Na+ transporters, namely NHX, SOS1, and HKT transporters, including recently revealed characteristics of these transporters.Since sodium, Na, is a non-essential element for the plant growth, the molecular mechanism of Na Many studies have demonstrated significant involvements of these Na+ transporters in Na+ sequestration in plant vacuoles, Na+ extrusion from cells, and Na+ circulation for the alleviation of sodium stress under saline conditions. Moreover, some of these Na+ transporters have been shown to improve salinity tolerance of crop plants when their encoding gene is overexpressed . Most of the plant species sequenced to date contain both types of NHXs . Interaction with AtCaM15 decreases the Na+ transport activity of AtNHX1, while keeping K+ transport activity almost unchanged. Given that the plant vacuole usually keeps a high Ca2+ concentration and an acidic luminal pH, AtNHX1 would favor a K+/H+ antiport mode under normal physiological conditions due to the binding with AtCaM15. However, under salinity stress, which often causes vacuolar alkalization  transporters are the best-characterized class of transporters attributed with this function . The SOS1 gene has been identified in several plants including Arabidopsis  that is categorized into the CPA1 family along with NHXs, although it is more closely related to bacterial NhaP antiporters , AtNHX8 seemed to prefer Li+ over Na+ as its substrate  exhibited increased Na+ content in roots and leaves as compared to wild type under salt stress, while stems of silenced plants accumulated less Na+ than the wild type  mutant exhibited increased Na+ accumulation in aerial organs and decreased it in roots  were responsible for the expression in roots (AtHKT1 expression (+ in the shoot (AtHKT1-overexpressor reduced salt content in the shoot, whereas AtHKT1 expression driven by the 35S promoter in whole plant resulted in salt hypersensitivity (AtHKT1. It is believed that this region is a potential small RNA target site and the methylation in this region may control the expression of AtHKT1 at a low level (Some transcriptional regulatory elements have been identified in the in roots . The secpression . Reductihe shoot . In accositivity . The putow level .+-transporters, may provide further insight into the development of efficient strategies for the engineering of “super salt-tolerant” crop plants and therefore progress in this research field is highly anticipated.Almost two decades of extensive molecular studies have clearly established the involvement of NHX, SOS1, and HKT transporters in plant salt tolerance. The genes encoding some of these transporters have since been successfully utilized as genetic tools for enhancing salt tolerance of model and crop plants. Recent advancements have also provided evidence on the involvement of these transporters in other vital cellular processes. This indicates the possibility that further enhancement of salt tolerance by overexpression of any of the above-mentioned genes might reach its limit due to excessive perturbation of related cellular and physiological processes. A much broader and profound understanding of the molecular properties, regulations, and cellular/physiological functions of NaThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Obesity and its associated complications, which can lead to the development of metabolic syndrome, are a worldwide major public health concern especially in developed countries where they have a very high prevalence. RIP140 is a nuclear coregulator with a pivotal role in controlling lipid and glucose metabolism. Genetically manipulated mice devoid of RIP140 are lean with increased oxygen consumption and are resistant to high-fat diet-induced obesity and hepatic steatosis with improved insulin sensitivity. Moreover, white adipocytes with targeted disruption of RIP140 express genes characteristic of brown fat including CIDEA and UCP1 while skeletal muscles show a shift in fibre type composition enriched in more oxidative fibres. Thus, RIP140 is a potential therapeutic target in metabolic disorders. In this article we will review the role of RIP140 in tissues relevant to the appearance and progression of the metabolic syndrome and discuss how the manipulation of RIP140 levels or activity might represent a therapeutic approach to combat obesity and associated metabolic disorders. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease. ► RIP140 regulates energy expenditure and storage in adipose tissue. ► RIP140 modulates muscle fiber type. ► RIP140 affects inflammatory responses. ► RIP140 is a promising target for treatment of metabolic disorders. However, they play largely opposing roles in mammalian physiology. WAT serves as the prime reservoir for storing excess caloric energy in the form of TAG. It is also an important source of endocrine hormones involved in metabolic regulation such as leptin, adiponectin, and resistin RIP140 is most highly expressed in WAT where it regulates the expression of many genes involved in catabolic pathways (energy consuming), especially those involved in lipid and glucose metabolism. Mice devoid of the coregulator express higher levels of genes that regulate fatty acid oxidation and proteins involved in energy dissipation, highlighting its role as a corepressor and its role controlling the balance between energy consumption and energy expenditure Recent studies by Li Na Wei and her colleagues indicate that RIP140 may not only be a transcriptional coregulator but may also function in the cell cytoplasm. They have found that cytoplasmic RIP140 inhibits glucose metabolism by reducing insulin-stimulated glucose transporter 4 (GLUT4) trafficking and glucose uptake Very few studies have been carried out in humans. A decrease in RIP140 mRNA in biopsies of visceral WAT depots from obese patients has been reported with a strong correlation between body mass index and RIP140 mRNA levels RIP140 is highly expressed in BAT albeit to a lesser extent than in WAT. Interest in the study of BAT physiology has been renewed by recent demonstration of considerable amounts of active tissue in many adult humans The amount of BAT and the expression of UCP1 correlate with increased basal metabolic rate and protection from obesity in both mice and humans. Genetic ablation of BAT induces obesity in mice 3Lipotoxicity and inflammation are intimately involved in the evolution of a number of metabolic diseases. Lipid overload of macrophages in vascular endothelium leads to phenotypic transformation into foam cells, secretion of pro-inflammatory cytokines, and apoptosis, contributing to atherosclerosis In contrast to the corepressor activities in adipose tissue and muscle, RIP140 can function as a coactivator in macrophages . CompariIt is perhaps surprising that the expression of metabolic genes is essentially unaltered in macrophages in the absence of RIP140 in contrast to the situation in skeletal muscle and adipocytes Communications between adipocytes and macrophages are important in triggering inflammation in adipose tissue and it is increasingly recognized that both cell types contribute to the development of obesity-induced insulin resistance. Although the infiltrated macrophages are the main source of TNFα, adipocytes are responsible for a sizable fraction of the IL6 concentration in the circulation of obese patients 44.1In both mouse and human, skeletal muscle contains distinct types of fibres, characterized by differential expression of specific myosin heavy chains. This difference in fibre composition accounts for different types of contractile function. Type I fibres (slow twitch fibres) are rich in mitochondria and show high levels of oxidative phosphorylation relaying mainly on fatty acid oxidation. These are the fibers resistant to fatigue and predominant in red muscles, such as the soleous. Type II (fast twitch fibers) contain a low number of mitochondria and relay more on anaerobic respiration via glycolysis or glycogenolysis that enable them to undergo rapid and short contractile bursts. In comparison to type I fibers, type II are more prone to fatigue. These types of fibers are predominant in muscles like the gastrocnemius and extensor digitorum longus (EDL) Skeletal muscles play an important role in whole-body metabolism as they account for the 70% of the total insulin-stimulated glucose uptake. Alterations in its function or fibre composition can have a big impact on the whole-body metabolism. For example, in insulin-resistant states, such as obesity and type II diabetes, insulin-stimulated glucose uptake is markedly impaired In skeletal muscle, RIP140 is expressed in a fiber type specific manner. RIP140 mRNA levels seem to be higher in glycolytic muscles, like gastrocnemius and EDL, rather than in oxidative muscles, like soleus and diaphragm 4.2Recent evidence suggests that impaired myocardial mitochondrial biogenesis, fatty acid metabolism, and antioxidant defence mechanisms lead to diminished cardiac substrate flexibility, decreased cardiac energetic efficiency, and diastolic dysfunction. These mitochondrial abnormalities can predispose to a metabolic cardiomyopathy and have been observed in insulin-resistant animal models and persons that are at higher risk of developing type 2 diabetes mellitus, hypertension, and cardiovascular disease RIP140 mRNA expression is high in cardiac muscle 5The hepatic manifestation of the Metabolic Syndrome is non-alcoholic fatty liver disease (NAFLD) and it is associated with insulin resistance although the cause and consequence relationship is debated 6Lipotoxic events are the underlying cause of cellular damage in the metabolic syndrome and therefore tackling the oversupply of lipid to affected organs is important. Reducing caloric intake and increasing exercise to favour a negative energy balance should lead to clearance of ectopic lipid deposition and improve an array of metabolic parameters Excess fatty acids reaching non-adipose tissue can be activated to fatty acyl-CoA (FACoA) that undergo a series of esterification reaction to form lysophosphatidic acid (LPA), phosphatidic acid (PA), diacylglycerol (DAG) and finally TAG. Alternatively, FACoA can be channelled down the sphingosine synthesis pathway, where ceramide is an intermediate, or it can undergo β-oxidation to feed the mitochondrial electron transport chain The pathways underlying the metabolic syndrome are interlinked in a complex network and an effective approach to treatment may need to target multiple points. For example, TDZ treatment, which activates PPARγ to reduce inflammation and promote adipogenesis and enhances adiponectin secretion to increase lipid oxidation What pharmacological approaches could be used to modulate the function of RIP140? Three possibilities are outlined: (a) regulation of RIP140 expression, (b) regulation of RIP140 activity and (c) regulation of NR-dependent recruitment. In the first instance, several NR ligands have been shown to control RIP140 expression. In breast cancer cells estrogens In its role as a transcriptional regulator, RIP140 lacks any intrinsic enzymatic activities but is thought to function as a scaffold protein, providing a platform for the recruitment of enzymes such as C-terminal binding protein (CtBP) A third method of regulating RIP140 function involves taking advantage of its recruitment by NR. Partial agonists or antagonists may selectively prevent interaction with RIP140 while not interfering with other activities of the receptor. Since the metabolic repressor action of RIP140 is supported by NR-dependent recruitment, the discovery of such compounds would prove valuable for limiting the dampening effect on catabolism in adipose tissue and skeletal muscle. Notably, MRL24 has been recently identified as a PPARγ minimal agonist that inhibits Cdk5-mediated receptor phosphorylation to prevent obesity-associated gene dysregulation in fat cells 7Overall this article summarizes the regulatory effects of RIP140 in tissues with an important role in the development and progression of the metabolic syndrome including white and brown adipose tissue, inflammatory cells, skeletal and cardiac muscles and liver. We have outlined signalling pathways that are subject to regulation by RIP140 and contribute to the development of the metabolic syndrome and speculated about the possibility of targeting RIP140 to ameliorate associated metabolic disorders.In our opinion, therapeutic strategies targeting RIP140 have to be considered with caution, considering the broad spectrum of actions of RIP140 as well as its recently described role as a coactivator. Given that RIP140 is likely to be a difficult metabolic target to finely tune it would be necessary to screen for compounds that modify the recruitment of RIP140 to specific nuclear receptors at a subset of genes, or alternatively, compounds that can alter its binding to other components of the transcriptional machinery, such as histone modifying enzymes.


Mus spp.) by screening 764 small mammals in Ghana. The Natal multimammate mouse , the known Lassa virus reservoir, was the dominant indoor rodent species in 4 of 10 sites, and accounted for 27% of all captured rodents. No rodent captured indoors tested positive for an arenavirus.Two arenaviruses were detected in pygmy mice ( Lassa fever is an arenavirus infection transmitted to humans from rodents in a limited geographic region of western Africa. Nosocomial outbreaks have been recorded in Sierra Leone, Liberia, Guinea, and Nigeria: the countries best known to report Lassa fever (LASV has a bisegmented genome: the nucleoprotein (NP) and glycoprotein (GP) genes are on the small RNA segment, and the polymerase (L) and matrix protein (Z) genes are on the large RNA segment. LASV circulates in rodent populations even when infections in humans are not occurring, providing a source for subsequent outbreaks among humans.Mastomys natalensis to be the only rodent reservoir for LASV  of captured species, and was outnumbered by another commensal rodent, Praomys daltoni, in 6 of 10 sites  with QIAshredder columns and purified with on-column RNase-free DNase set (QIAGEN). The samples were tested for the presence of arenavirus GP gene RNA by using Power SYBR Green RNA-to-C 1-Step PCR Kit   and Natorduori (site 1), respectively, were found to be positive for arenavirus RNA. For these 2 positive specimens, additional PCRs were performed by using primers OWS1+, OWS1000-, OWS2165A+, OWS2165B+, OWS2840A-, OWS2840B-, OWS2770+, OWS3400A-, and OWS3400- to acquire longer fragments of GP and NP genes  but with low branch support (20% bootstrap support) in the GP gene tree  were identified as Mastomys natalensis, compared with 54% (601/1123) in Guinea and 80% (82/103) in Mali (P. daltoni, a species not known to harbor LASV, outnumbered M. natalensis in Ghana, and was the predominant rodent species found indoors in 6 of our 10 sites.This study aimed to provide data on the risk for Lassa fever in Ghana, a country situated between well-known Lassa fever–endemic regions, but with little known disease itself. Our findings support the idea that Lassa virus is not widely prevalent in Ghana, and the Mus baoulei and Mus mattheyi. Jirandogo, the sequence found in Mus baoulei, is phylogenetically close to LASV clade viruses. However, the maximum amino acid difference of 18% in NP between Jirandogo and LASV exceeds the 12% cutoff criteria, and therefore places it outside the LASV clade (Sequences of arenaviruses were detected in 2 species of pygmy mice, Mus species in Africa are sometimes found indoors where exposure to humans is more likely, but most are found outdoors, including those from which we collected the 2 positive samples tested in our study. Infrequent cases in humans would be expected from arenavirus carriage in outdoor species, such as Mus.Arenaviruses other than LASV have recently been reported to cause human disease in Africa, but it is not known whether the viral sequences we found in Ghana are from viruses pathogenic to humans. Mastomys natalensis represented 15% of all rodents captured, and showed no evidence of arenavirus infection by PCR testing. The cases in humans were reported on the basis of the results of PCR tests, and further sequencing will be necessary to confirm the finding.It is notable that possible cases of Lassa fever have recently been reported in Ghana by the Ministry of Health from the high risk area near site 5 of our study (Mastomys natalensis species tested from Ghana, although 2 were detected in other rodent species from sites with high predicted risk. More information about geographic and temporal fluctuations in Mastomys natalensis rodent populations, the frequency of virus host-switching among rodents and the degree of arenavirus circulation is needed to better understand the implications of our findings for the risk for disease outbreaks from LASV or other arenaviruses in Ghana.Although arenaviruses have species–specific reservoirs, recent work suggests host switching may be more common than previously believed (